The subject of inflammation seems to be appearing with greater frequency in the media and even as the feature subject in health magazines (e.g. in 2004 the July Life Extension and the August Alive) and newsletters devoted to health issues. Inflammation "The Secret Killer" was the cover story of the February 23, 2004 issue of Time magazine. If the title or abstract key word "inflammation," is used in a MEDLINE (PubMed) search of the medical literature, it brings up almost 16,000 citations for just 2002-2003. Chronic inflammation appears to be associated with many health issues where the connection is neither obvious nor even generally appreciated. This review will explore a number of aspects of this subject. First we will examine why inflammation can be viewed as a double-edged sword by looking at the inflammatory component of the immune response.

INFLAMMATION AND THE IMMUNE RESPONSE

This review will be concerned with chronic inflammation and the associated health issues (1-3). Its counterpart, acute inflammation, is presumably well known to readers, and is characterized by its relatively short duration which is frequently accompanied by pain, fever, swelling, etc. For example, the occurrence of a cut will immediately result in the body marshalling forces to deal with bacterial and other foreign matter introduced by the injury. This immune reaction is accompanied by inflammatory processes of great biochemical complexity. Within a short period the injury may exhibit swelling, pain and redness. There follows a complex series of repair processes which eventually lead to healing, perhaps with the formation of some scare tissue. The entire episode is characterized by its short duration and by the success of the body in dealing with the problems of infectious agents, foreign material, injured tissue disposal, and repair of the damaged area. Thus inflammation is the body's immediate response to injury or infection, and the normal end result is the elimination of invading pathogens or toxins and the repair of damaged tissue. This is critical to our everyday survival.

The acute inflammatory response is normally subject to a series of complicated control mechanisms which turn off the generation of dangerous chemicals secreted by cells of the immune system when their task is completed. Failure of this control mechanism can lead to uncontrolled inflammation and serious disease. Also, if the body is unable to successfully deal with the cause of the acute response, for example due to a severely impaired immune system or an infectious agent that overwhelms the immune system, the problem can escalate to one of critical or even fatal proportions. An example is the Systemic Inflammatory Response Syndrome (SIRS) seen in the critical care setting.

Chronic inflammation, on the other hand, may develop in several different ways, depending on the circumstances (2). For example, if the cause of an acute inflammatory episode is not completely resolved due to the inability to completely eliminate the agent responsible, a low-level immune/inflammatory reaction will remain. It is also possible that there is no acute phase due to the stimuli responsible having low toxicity and thus being incapable of initiating the acute inflammatory response but still able to maintain a low-level immune/inflammatory reaction. Normally, the term chronic is used to describe an inflammatory process that persists for more than a few days or weeks. Chronic inflammation has been described as "frustrated repair," repair that is thwarted because of the presence of an irritant that cannot be eliminated, such as a persistent antigen that continues to trigger a low-level immune response. The continuous immune response with its associated inflammation can be extremely damaging.

Causes of chronic inflammation include:

• Persistent infectious agents, especially those of low toxicity that are not eliminated by the normal immune/inflammatory response.
• Remnants of dead organisms such as bacteria which remain after the bacteria have been killed by the normal immune reaction.
• Foreign material for which the body has no mechanism for complete removal, such as silica dust, talcum powder, splinters, etc.
• Metabolic products that accumulate in abnormal amounts, deposit in inappropriate locations and become a source of irritation and inflammation. A classic example is the accumulation of uric acid crystals in joints to yield the painful disorder gout.
• Psychological stress which can have much more widespread inflammatory effects than is generally appreciated. This type of stress is now known to stimulate the production of a variety of pro-inflammatory substances.
• Non-self tissue such as in organ transplants or the failure of the immune system to recognize "self" as in autoimmune disease.
• Toxins in food, air, water or tobacco smoke.
• Obesity and overeating in general.
• Hyperglycemia and diabetes.

The immune system is clearly and directly involved in chronic inflammation and inflammatory lesions are frequently characterized by the presence of cells involved in the immune response. Chronic inflammation may be asymptomatic, may "simmer" for years, release of a whole host of potentially toxic substances and may continue unnoticed until a resultant disease state becomes symptomatic and recognizable. Some of these toxic substances are capable of causing DNA or RNA damage and can, for example, initiate cancer (4-6). These substances may also create a pro-cancerous environment which promotes cancer cells or tumors to grow, establish blood supply, and even metastasize (7). The list of toxic substances associated with inflammation is long, and the damage and potential problems manifold. On the other hand, chronic inflammation can even at a fairly early stage result in painful symptoms and potentially life-altering problems such as are seen in rheumatoid arthritis. In fact, rheumatoid arthritis is the classic example of a failure in the body's struggle and confusion regarding self versus non-self.

In this review we will examine in some detail the connection between chronic inflammation and a number of diseases and health problems and review the current ideas as to what preventive action may be appropriate. Hopefully it will become clear that the current and intense interest in this subject in both lay and professional circles is completely justified.

THE OMEGA-3 AND OMEGA-6 FATTY ACIDS AND INFLAMMATION

While the evolution of the recognition of the importance of the omega-3 (n-3) and omega-6 (n-6) polyunsaturated fatty acids (PUFAs) is long and complicated, what is clear is that the health benefits of eating large amounts of fish as seen in certain native and other populations, the frequent association of n-3 consumption with decreased risk of certain diseases, and a growing understanding of the key role of these fatty acids in the complex biochemistry and microbiology of the immune/inflammation process has been in a large part responsible for the current intense interest both in the laboratory and clinically (1). Enthusiasm for the n-3 fats has also spread to the general public, judging by the popularity of such books as the Omega RX Zone, the Omega Diet and other similar diet books. The recommendation to eat more fish is now coming from mainstream medicine (8). Thus the obvious question—why are the n-3 fatty acids so important, what is the evidence, and what mechanisms are at work? The answer mainly involves inflammation.

There are four general types of fats which are termed saturated (SF), monounsaturated fatty acids (MUFA), polyunsaturated fatty (PUFA), and trans-fats (TF). While the last type does occur in nature, it is in very small amounts and has become a significant part of human fat intake only very recently in the form of stick or tub margarine or extended shelf-life oils produced through partial hydrogenation of PUFAs. Today there is almost universal agreement that TFs should be strictly avoided (see the IHN research report "Dietary Fat and Heart Disease"). In connection with inflammation, attention is focused mostly on certain n-3 and n-6 PUFAs.

The parent dietary n-6 fatty acid is linoleic acid (LA) which is found mainly in corn, safflower, sunflower, soybean, and peanut oil, whereas the parent fatty acid for the n-3 family is alpha-linolenic acid (ALNA), which is present in flax seeds and flax seed, soybean and rapeseed oils, walnuts, and some green vegetables. The importance of LA and ALNA derives mostly from the fatty acids that the body makes from these two starting materials, of which the most important are arachidonic acid (AA) from LA and eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) from ALNA. They are frequently called long-chain PUFAs. While EPA and DHA are produced in humans with very low efficiency from ALNA (9), they can also be acquired from eating fish or consuming fish oil. The quantities of EPA and DHA available in normal servings of cold water oily fish (salmon and herring for example) can only be duplicated by consuming quite large amounts of flax seed, its oil or other dietary sources of ALNA. This is an important consideration when EPA and DHA are being used for preventive or therapeutic reasons.

The LA to AA chemistry is complicated (10), but an important aspect is that EPA inhibits and insulin promotes the last enzyme controlled step leading to AA. Also, since some DHA is retro-converted into EPA increasing the intake of EPA and DHA decreases the production of AA from LA, and while this is not reflected in the dietary n- 6:n-3 ratio, it is reflected in the AA/EPA ratio as seen in cellular phospholipid membranes. Also, inhibiting this last step allows the accumulation of an intermediate that is involved in the production of anti-inflammatory substances. This may in part explain the much stronger health benefits seen from increasing the intake of EPA and DHA rather than in decreasing the intake of LA. However, as will be discussed below, different diseases respond differently with cancer apparently somewhat more sensitive to the intake of LA and the n-6:n-3 ratio than is coronary heart disease (CHD).

EPA and AA are incorporated into the cell wall of most cells, including cells involved in immunity and inflammation. When large amounts of AA are available, this can result in abnormally low concentrations of cellular EPA. Increasing the intake of ALNA or especially EPA from fish, fish oil or supplements can alter the cell wall balance between these two fatty acids. This is important in part because the immune response involves the release of both AA and EPA from cells of the immune system. These fatty acids are converted to powerful cell signaling and inflammation mediating chemicals called eicosanoids, with EPA leading to the n-3 family and AA leading to the n-6 family of these very bioactive and highly important compounds. High levels of EPA can also decrease the production of n-6 eicosanoids by virtue of inhibitory reactions. The frequently quoted notion that eicosanoids produced from EPA are anti-inflammatory and those from AA are inflammatory is however, an oversimplification. Both AA and EPA are involved in the synthesis of a large number of eicosanoids and in the two families there are both pro- and anti-inflammatory products (1,11). Balance is thus the key, but nevertheless it appears that labeling AA, its parent LA, and the resultant eicosanoids as pro-inflammatory is a useful generalization, especially since AA generates eicosanoids that are among the most powerful inflammatory agents, while those from EPA are in general much less so. The fact that insulin is a promoter of the last step from LA to AA focuses attention on the inflammatory aspects of insulin resistance, the Metabolic Syndrome and the dangers of high-glycemic load diets, as will be discussed later.

The advent of agriculture, the large scale production of vegetable oils high in n-6 PUFAs used for baking and cooking, and the emphasis on n-6 rich grain feeds for domestic livestock has in the last half-century resulted in a significant increase in human consumption of n-6 fatty acids as compared to pre-agricultural times (12). While this was happening there was a decrease in fish consumption, a major source of n-3 fatty acids. Also, due to feeding practices, the n-6:n-3 ratio in eggs rose dramatically from about 1-4:1 to 20:1 found in typical supermarket eggs. Also the n-3 content of some farmed salmon decreased along with the decreased supermarket availability of wild salmon (one of the best sources of n-3 fatty acids). The result is that the n-6:n-3 ratio in the diet of the Western world is now between 16:1 and 20:1. Estimates of this ratio in our hunter- gatherer ancestors and present-day hunter-gatherer societies is 1:1 to 4:1 (12).

These rapid and profound changes in diet are not something to which humans can easily adapt. The spontaneous mutation rate of nuclear DNA is about 0.5% per million years, and thus over the 10,000 years during which the transition occurred from the hunter-gatherer way of life to the cultivation of grains and then to highly industrialized food production, our genes and thus our metabolism have remained essentially unchanged and today we live in a nutritional environment that is vastly different from that for which our genetic constitution was selected (12). There has not been time to adapt. Also, the change in the balance of n-3 to n-6 fatty acids is implicated in diseases which become symptomatic and potentially fatal either late in life or after the reproductive years, eliminating a powerful mechanism for adaptation. This modern nutritional environment appears intrinsically much more pro-inflammatory than the diet optimum for our genetic make-up. Thus, the frequently seen recommendation of the merits associated with eating a diet similar to Paleolithic or Stone Age man. It was high in fiber, antioxidants and other micronutrients, had a low n-6:n-3 ratio, and of necessity included mostly low glycemic index vegetables, nuts and fruits and no cultivated grains at all. Wild vegetables were good sources of n-3 fatty acids. Wild animal fat is estimated to contain about 4% EPA, whereas modern domesticated beef contains small or undetectable amounts of even ALNA (12). The Paleolithic diet also contained fish when available.

There is also a very important relationship between eicosanoids and what are termed cytokines (and leukotrienes), the so-called-hormones of the immune system. These are soluble polypeptide (small protein- like) products of the cells of the immune system. They help regulate the response to injury and infection. Some of the members of these two classes of biochemicals are involved whenever the immune system is activated. Cytokines facilitate the intercellular communication and help orchestrate the immune response, stimulate the action of various immune cells and are involved in a wide variety of reactions including those producing inflammation. They are also thought to promote atherosclerosis. In the context of this review, the most important are Tumor Necrosis Factor Alpha (TNF) and the Interleukins 1, 6 and 8 (IL-1, IL-6 and IL-8), all of which are implicated in inflammation. A very important function of both EPA and DHA is that they can inhibit the production of these inflammatory cytokines. While some of the evidence is from cell culture or animal studies, there are also human studies which back up this view. It is still not clear whether this anti-inflammatory action is through antagonism of eicosanoid production or via an eicosanoid independent path, or both (1). Uncontrolled inflammation can produce very destructive levels of TNF, IL-1 and IL-6, and chronic over production is implicated in endotoxic shock, the acute respiratory distress syndrome, rheumatoid arthritis and irritable bowel syndrome (1). There seems no doubt that EPA and DHA play a role in anti-inflammatory processes that in part involves the modulation of the effects of inflammatory cytokines and leukotrienes.

The production of eicosanoids from either AA or EPA involves two classes of enzymes, the cyclooxgenases (COX) and the lipooxygenases (LOX). The former are now recognized to comprise at least two forms called COX-1 and COX-2. COX-2 has gone from being a term heard only in laboratories to almost a household word due to the advent and very heavy promotion of the pharmaceutical COX-2 inhibitors such as Vioxx and Celebrex. The anti-inflammatory action of the COX inhibitors is due to their disruption of the production of inflammatory eicosanoids. Aspirin and other anti-inflammatory over-the-counter drugs such as ibuprofen are also COX inhibitors, but inhibit both COX-1 and COX-2, and are called non-specific. COX-1 inhibition is thought to interfere with the natural protective mechanisms of the gut mucosa from the adverse effects of stomach acid. This was part of the rationale for the introduction of the COX-2 class of inhibitors, which are promoted as being more "stomach friendly." This is still being debated, as is the significance of other side effects including the possible increased risk of heart attacks associated with COX-2 inhibitor drugs. In fact, Vioxx was recently pulled from the worldwide market by Merck after a study of its anticancer action uncovered excess heart problems (13). Nevertheless, the point is that by inhibiting the enzyme that is required to synthesize inflammatory eicosanoids from AA and perhaps EPA, there is a clinically observable decrease in inflammation and pain which has made both the over-the-counter and prescription COX inhibitors hugely popular for everything from headaches to osteoarthritis and rheumatoid arthritis. If an inhibitor is used which stops all eicosanoid production from both AA and EPA there can be a dramatic reduction in pain and inflammation, but the use of such a drug for more than a few weeks can have serious if not devastating side effects, as seen with the corticosteroid class of drugs (e.g. cortisone).

The n-3 PUFAs can also influence the ability of some immune cells to bind to various surfaces, a process that is thought to be part of the atherosclerotic process. Also, there is growing evidence that these PUFAs also are modifiers of inflammatory gene expression (1). Thus the anti-inflammatory effect of n-3 PUFAs appears to operate through several different mechanisms. These mechanisms provide support to the therapeutic and preventive use of these PUFAs by providing a biological basis for the anti-inflammatory actions observed clinically (1).

The simplified notion that n-6 PUFAs are pro-inflammatory whereas the n-3s are anti-inflammatory and also that n-6 PUFAs inhibit the anti-inflammatory action of the n-3s has resulted in the belief that it should be beneficial to reduce n-6 and increase n-3 consumption. A related question concerns the optimum ratio of dietary n-6 to n-3 fatty acids (FAs). A very recent prospective study of over 50,000 US men and women (from the famous Health Professionals Follow-up Study data base) examined these questions by measuring the effects of both the n-3 and n-6 FAs on inflammatory markers C-reactive protein (CRP), IL-6 and a serum marker for TNF activity. They found that the n-6 FAs do not inhibit the anti-inflammatory action of the n-3 FAs, and that the combination of both types of FAs is associated with the lowest levels of inflammation. The authors suggest that the inhibition of inflammatory cytokines may be one of the mechanisms for the observed beneficial effects of these FAs on chronic-inflammatory related diseases (14). Consumption of ALNA was found to have no effect, probably because of the low conversion to EPA and DHA. They also confirmed the anti-inflammatory effects of long-chain n-3 FA intake, consistent with many other studies (15,16). The authors point out that there are no data from human studies that support the detrimental effect of dietary n-6 FAs (17), whereas several studies have found beneficial effects (18). However, they are ignoring a possible connection with high n-6 intake and cancer, as will be discussed in Part II. Thus they conclude that while n-6 FAs may raise pro-inflammatory cytokine levels, the combination of n-3 and n-6 FAs may decrease the formation of these pro-inflammatory substances. The important point is that a significant decrease in the consumption of n-6 FAs may not be as beneficial as increasing the intake of the n-3s. Again, balance is the name of the game, and health problems appear to be primarily associated with low to very low n-3 levels.

Thus the optimum ratio of dietary n-6 to n-3 FAs for either healthy individuals or those with various inflammation related disorders is far from clear. There is in fact considerable evidence indicating that n-6 FA consumption is beneficial, especially in the context of LDL cholesterol levels, insulin resistance and the threshold for ventricular fibrillation (18). As will be discussed in Part III, both n-3 and n-6 FAs are associated with lower risk of CHD, but the biological pathways are thought to be different (18). Hu et al (18) in fact take the position that, considering the strong protective effect of n-6 FAs against CHD observed in epidemiologic studies, the recommendation frequently seen to reduce n-6 FA consumption does not seem, in their opinion, justifiable. However, their position is based only on heart disease considerations. This is in fact a complex question because both AA, EPA and DHA lead directly to a large number of immune and inflammation related biochemicals (19), and in addition are involved in triggering the production of an additional array of substances of importance in connection with both inflammation and immunity. It may be some time before an evidence-based answer is available regarding the optimum ratio of these very important and essential fatty acids in the context of the major inflammation-related diseases.

This rather detailed discussion of the mechanism of action of the n-3 and n-6 fatty acids may seem excessive, but it is important for the discussion to follow of these fatty acids in connection with a variety of diseases. As will be seen, LA, ALNA, AA, EPA and DHA are principal players in the inflammation game, and an understanding of their interrelationships is very important.

ADVANCED GLYCATION ENDPRODUCTS AND INFLAMMATION

There is one class of pro-inflammatory agent not derived from the usual sources of chronic inflammation that is thought to play an important role in disease and especially diabetes and its related vascular complications (20- 23). This family of molecules is called Advanced Glycation Endproducts (AGE). They result from the reaction (the Maillard Reaction) of sugar (typically glucose or fructose) and protein molecules. The process of glycation can render proteins dysfunctional and has been associated with the "natural" aging process. AGEs can ultimately give rise to what is called cross-linking where molecules are joined through chemical bonds to make bigger molecules (proteins are already "macro" molecules), which for example can result in undesirable stiffening and hardening of tissue. But AGEs are also able to initiate the synthesis of pro-inflammatory substances from various immune cells and, because of the persistence of AGEs, they can be triggers for chronic inflammation (20,23).

AGEs are not only made in vivo via the Maillard reaction, but also occur in foods cooked for long times at high temperatures. The hallmark is browning such as seen on bread crust or the surface of pretzels, or the color of well-done meat. Diet is in fact considered a major environmental source of AGEs (21). A recent human dietary study which limited foods high in AGEs (no cooked foods or roasted foods, no bakery products, no coffee, etc) dramatically reduced a urinary marker for AGEs in a small group of subjects (24). While the body has mechanisms for the degradation and elimination of AGEs, these processes can become impaired and lead to accumulation (20).

Both cell culture (23), animal (25) and human (21,26) studies support the view that AGEs are pro-inflammatory. In a recent study on human subjects, two diets differing by six-fold in AGE content were examined for their ability to generate inflammatory markers. The high-AGE diet produced significantly elevated serum levels of both AGEs and the inflammatory markers (TNF) and vascular adhesion molecule-1 whereas the low AGE diet reduced the levels of these markers by 30-50% (20). Similar results on a group of human subjects were obtained by Vlassara et al (26). These results are consistent with animal studies (25). In fact, it appears to be a general property of AGEs that they can influence serum levels of inflammatory mediators such as TNF, IL-1, IL-6 and vascular cellular adhesion molecule-1 (21).

A key question regards the connection between hyperglycemia (high serum glucose levels) and AGE mediated inflammation from endogenous AGE formation. There is evidence that this connection indeed exists. For example, hyperglycemia is related to the modification of the ocular lens by AGEs, leading to cataracts common both to diabetes and aging. Also, intracellular AGEs are found to be significantly elevated in diabetics, many of whom have poor glucose control and hyperglycemia. The recently reported positive association between glycosylated hemoglobin (a measure of long term serum glucose levels) and cardiovascular disease in diabetics (27-29) was explained in part by the postulated enhanced formation of AGEs and thus inflammation in the presence of hyperglycemia (27,30). These studies are consistent with a recent study from Harvard where the dietary glycemic load was significantly and positively associated with serum C-reactive protein (CRP) in healthy middle-aged women, independent of conventional risk factors for ischemic heart disease. While the authors offer several possible explanations, one was that hyperglycemia could lead to AGEs which might stimulate the liver to release acute phase reactants such as CRP (31). These and other studies are the basis of the hypothesis that AGEs mediate low-grade and potentially chronic inflammation.

ACCESSING INFLAMMATION STATUS

It would be nice if there was a simple, reliable blood test that would indicate the presence or absence of chronic inflammation. If chronic inflammation was found, steps could be taken to ascertain the cause and the test could be used to follow the success of treatment. The measurement of high-sensitivity C-reactive protein (CRP) appears to meet some of these criteria, but the laboratory results can be misleading (see also the recent IHN research review on CRP and heart disease). Another test, called the Omega-3 Essential Fatty Acid Profile has recently become commercially available and appears to hold great promise for identifying chronic inflammation.

CRP is a so-called acute phase reactant produced by the liver in response to IL-6. Levels can reach 1000-fold of normal in the presence of acute infection, and it is frequently elevated in autoimmune diseases, trauma, infection, diabetes and malignancy. Historically, some physicians used CRP to monitor success of the treatment of acute infections. The old "low-sensitivity" CRP assay had a lower limit of detection of about 7 mg/L which, while providing evidence of serious inflammation, failed to discriminate among "normal" individuals, most (about 80%) of whom have serum CRP between 0.1 and 3.8 mg/L (32,33). What is significant is that disease related risks increase considerably within these "normal" limits. In the last decade there has been an explosion of research on the correlation of CRP levels and the risk of cardiovascular disease, Alzheimer's disease, the risk of vascular complications in diabetes, and autoimmune diseases (34). Serum levels of CRP have also been used in examining the connection between cancer and other diseases and inflammation (34,35).

Since CRP is a general marker for inflammation, it naturally can be elevated by conditions that are temporary such as an infection or injury. Thus an elevated value must be confirmed by additional measurements to answer the question of the presence of a chronic inflammatory condition. In other words, a very low value is reassuring, whereas an elevated value merely calls for more tests to eliminate the possibility of temporary elevation. When an elevated value persists, it can be argued that action is indicated to identify the source, and this may present a diagnostic challenge if the patient is asymptomatic. Periodontal (gum) disease is a good example of an inflammatory condition that elevates CRP but is easily overlooked (36). Not only is CRP a marker for inflammation, but there is now evidence that it can also take part in inflammatory processes, thus adding to the danger of high levels (37,38). In Part II and Part III of this review the interplay of CRP, inflammation and the risk of disease or disease progression will be discussed for various conditions.

It does not appear that CRP is, as yet, normally included in the blood tests done during routine physical exams since the test may not be covered by insurance and even be omitted when patients presenting with symptoms of heart disease are evaluated. Those most actively involved in CRP research suggest that it is time for physicians to consider the potential of CRP in the routine assessment of the risk of CVD (32,33). However, organizations like the American Heart Association are still reluctant to recommend the use of CRP testing for general screening (39).

There are other markers of inflammation that merit mention. Fibrinogen is, like CRP, an acute phase inflammation marker and is sometimes measured along with CRP to get a better picture of the overall risk of cardiovascular disease. IL-6 is also an important inflammation marker, but its serum levels are more variable and its measurement is generally associated with inflammation research rather than routine patient assessment. Since IL-6 triggers CRP release, to some extent CRP is a surrogate marker for this cytokine.

The hypothesis that the fatty acid AA is inflammatory and EPA is anti-inflammatory, while an oversimplification, has given rise to a new blood marker for measuring inflammation status. Barry Sears in his new book The Anti-inflammation Zone (10) calls it the "Silent Inflammation Profile." The marker is simply the ratio of AA to EPA, generally measured in the phospholipid fraction of blood fats. A related blood marker is the sum of the concentrations of EPA and DHA, frequently expressed as a percentage of total blood phospholipids. Sears presents arguments based on a small number of studies that the AA/EPA ratio should be in the range of 1-3 with both lower and higher values outside this range representing dangerous ground. For example, the Japanese, who have the lowest rates of heart disease in the world, have values of this ratio in the "good" range (40). In the famous Lyon Diet Heart Study where a Mediterranean type diet enriched with ALNA was compared to the usual European diet, the dramatic decrease in secondary adverse events in heart patients (79%) was accompanied by a drop in the serum AA/EPA from 9.1 to 6.1 but, interestingly, little change in serum cholesterol levels between the intervention and control groups. But there is some evidence based on a 1999 study (41) of Greenlanders with high n-3 PUFA intake, that the risk of hemorrhagic stroke increases when the ratio reached 0.5. However, in which the healthy controls in this study had a ratio of 0.82. Greenlanders have a well known low incidence of ischemic heart disease, but high incidence of cerebrovascular disease (41). Sears stratifies the serum AA/EPA ratio in terms of risk for inflammation related disease as: very high, above 15; high, 9-15; moderate, 3-8; low, 1- 3; and moderate again for less than 1.

An inverse correlation between disease risk and the AA/EPA serum ratio has been observed in studies of clinical depression (42,43), Alzheimer's disease, dementia and cognitive impairment (44), and with regard to the clinical outcome of patients with newly diagnosed multiple sclerosis (45). In all of these studies the ratio was directly measured. Typical value for "normal individuals" was about 6. That it is not lower is probably simply an indication of the high n-6 and low n-3 PUFA consumption that typifies the Western diet. Sears would no doubt point to these "normal" values as further evidence for the widespread presence of chronic "silent" inflammation in individuals who appear healthy. In fact, based on his own experience, he claims that the average AA/EPA ratio in Americans is 12, but then this average includes the obese, those with Metabolic Syndrome, atherosclerosis, diabetes, non-fish eaters, etc., etc., so this is not surprising, but should still be considered alarming.

The AA/EPA ratio is easily altered with supplements, either based on fish oil or purified EPA and DHA. Two studies using 4 capsules a day of a prescription preparation (Omacor) of mixed EPA and DHA containing a total of 1.88g EPA and 1.47g DHA found decreases in the AA/EPA ratio of 20 to 7 and 11.1 to 2.1 (46,47).

The EPA + DHA level also appears to be attracting attention (46,48,49). In a recent paper, Rupp et al (46) review both this sum and the AA/EPA ratio. Included is a sensational graph of two studies (50,51) of the effect of EPA and DHA on the risk of sudden cardiac death, where over the range for the sum of 3.5% to about 7% (% by weight of EPA + DHA in the total phospholipids fraction of whole blood) the risk goes from 1.0 (reference) to about 0.1! Harris et al (48) have recently proposed that the EPA + DHA expressed as a percentage of total fatty acids in the red blood cells (RBC), which they term The Omega-3 Index, be used as a risk factor of significant clinical utility for death from CHD. They show that this index correlates very well with the EPA + DHA levels expressed as a percentage of the total plasma phospholipids, a measure used in some studies. Furthermore, they show that the Index is inversely associated with risk for CHD mortality based on a review of five studies. They suggest that a target for reducing risk is an Omega-3 Index value of 8%, whereas levels less than 4% represent the least cardio-protection. Further support for the utility of the Omega-3 Index comes from a study showing that the RBC based EPA + DHA sum was highly correlated with cardiac tissue EPA + DHA (49) The required oral supplementation required to produce levels above the 8% level in the sum is about 500 mg/d of a mixture of EPA and DHA in roughly the proportions found in fish oil. One g/d was found to yield a value of the Omega-3 Index of 10% and 2 g/d a value of 12%. These latter two values would be considered, on the basis of what is now known, to be highly protective. Note that the numbers produced from RBC analysis are higher than those from phospholipids analysis, i.e. the above-mentioned correlation is linear but not 1:1.

The above discussion of these markers would be of only academic interest if they were merely research curiosities, but in fact a test is now commercially available for n-3 status based on these two indicators plus two more. It is called the Omega-3 Essential Fatty Acid Profile and is now available in a number of countries. In Canada the assay has been licensed to MDS Diagnostic Services and can be ordered by any physician. For more details, see www.nutrasource.ca. The developers of the assay, Nutrasource Diagnostics, who are associated with the University of Guelph in Ontario, Canada, quote optimum reference ranges of 1.5 to 3.0 for AA/EPA and greater than 4.6% for the EPA + DHA score. The 4.6% EPA + DHA score, which is a percentage based on total serum phospholipids, is equivalent to a value of 8% for the RBC based Omega-3 Index (see (48), Figure 3). Since according to Harris et al (48), the Omega-3 Index cutoff value of 8% is a lower limit and thus values higher than this appear desirable. RBC based Omega-3 Index values of 10 and 12% are equivalent to EPA + DHA scores (Nutrasource Diagnostics) of about 7 and 9%. Three of the five studies examined to establish the cutoff had the Omega-3 Index above 8% (8.3, 8.9 and 9.5%).

If the n-3 PUFAs are anti-inflammatory, one would expect that supplementation or high levels of fish consumption would lower the biomarkers of inflammation. In a recent study by Lopez-Garcia et al (52) it was found that increased consumption of n-3 PUFAs was associated with decreased levels of six markers indicating lower levels of inflammation and endothelial activation. When comparing the first with fifth quintiles of intake for the sum of n-3 FAs, it was found that mean values of CRP decreased 1.7 to 1.2 mg/L, a decrease that was statistically significant. The total range of n-3 intake was from 0.54 to 3.33 g/day. In a study (53) where ALNA was used as the source of n-3 fatty acids, CRP decreased in a statistically significant manner from 1.24 to 0.93 mg/L. This intervention involved a diet where the n-6 to n-3 ratio for the PUFAs was 1.3:1. In an intervention study (54) involving a Mediterranean diet vs. a "prudent" diet, intake of n-3 fatty acids was increased on average from 0.6 g/d to 1.5 g/day whereas in the control diet it remained essentially the same. CRP decreased from 2.7 to 1.8 mg/L in the intervention group and an insignificant 2.9 to 2.8 mg/L in the control group. The study (55) by Madsen et al found no effect of dietary n-3 PUFAs on CRP, but most of the 40 patients divided between the low- and high-dose interventions had very low levels of CRP at baseline. A decrease in CRP was seen in most of the individuals with initial CPR > 2 mg/L, but the total number of subjects in this category was quite small.

In Parts II and III, the connection between inflammation and various diseases, including cancer, cardiovascular disease, diabetes, Alzheimer's disease, etc., will be discussed. The role of n-3 and n-6 essential fatty acids will be underscored by this discussion, as will the significance of elevated CRP. In addition, the role of non-steroidal anti-inflammatory drugs will be examined, both in connection with implicating the potential role of inflammation in the etiology of some diseases, but also as a possible preventive measure under some circumstances. Finally, the question of anti-inflammatory diets will be addressed.

INFLAMMATION AND CANCER

The suspected connection between inflammation and cancer can be traced back to 1863 when Rudolf Virchow suggested cancer originated at sites of chronic inflammation. Recently there has been renewed interest in the Virchow hypothesis (5). What then in general terms is the evidence for the connection?

• Anti-inflammatory drugs, both prescription and over-the-counter, have been observed to influence the incidence and/or progression of some cancers (56-58). The evidence is strong enough to justify a number of ongoing trials of the use of COX-2 inhibitors as preventive drugs (59,60). The recent downfall of Merck's Vioxx occurred because of adverse cardiac events that occurred in a study, not of pain reduction, but of the drug's effectiveness in preventing the recurrence of neoplastic large bowel polyps in subjects with a genetic predisposition for colorectal cancer (61,62).

• A relationship observed in some but not all studies of the n-3 and n-6 PUFAs (polyunsaturated fatty acids) and the incidence and/or progression of some cancers has strengthened the hypothesis that there is an association with inflammation (63-67). Studies range from the incidence of cancer as a function of fish consumption (68,69) to studies that relate the long-chain fatty acid (EPA and DHA) concentrations in breast tissue to the incidence of breast cancer in the case-control setting (70,71). This is one of the most extensively investigated associations.

• Cell culture and animal studies tend to confirm the association of inflammation and cancer (5).

• Plausible mechanisms derived from both microbiologic and biochemical studies provide a basis for the hypothesis, although such studies are far from definitive and the understanding of the total picture will require a more complete understanding of the etiology of cancer than exists at present (4-7,72-74).

• It has been observed in numerous studies over many years that chronic inflammatory conditions such as asbestosis, silicosis, bronchitis, cystitis, pancreatitis, etc., carry enhanced risk of developing cancer (5).

N-3 FATTY ACIDS AND CANCER
Animal and cell-culture studies as well as some epidemiologic studies suggest that n-3 fatty acids, especially EPA (eicosapentaenoic acid) and DHA (docosahexaenoic acid), inhibit carcinogenesis and tumor growth. The anti-carcinogenesis properties of these substances are not yet fully understood but a number of mechanisms have been proposed (63). These include:

1. altered immune response to cancer cells and modulation of inflammation, cell proliferation, cell death, metastasis and angiogenesis (development of new tumor blood supply) all due to suppression of eicosanoids from AA (arachidonic acid);
2. influence on metabolism, cell growth and differentiation that are due to gene related effects. A number of complex mechanisms have been suggested, some of which are not directly related to the inflammatory or anti-inflammatory actions of n-3 and n-6 derived eicosanoids and related cytokines;
3. reduced estrogen stimulated cell growth due to decreased estrogen production tied to eicosanoid levels;
4. changes in the production of free radicals and active oxygen species;
5. mechanisms involving membrane fluidity and insulin sensitivity changes due to cell wall composition of n- 3 or n-6 PUFAs;
6. formation of oxidation products (lipid per-oxidation) that inhibit the growth of tumors and tumor cells— oxidation products produced in particular from EPA or DHA. Antioxidants are found to interfere with this cytotoxic process in cell cultures, and are implicated in epidemiologic and clinical studies.

Since carcinogenesis, tumor growth and metastasis are only partially understood, trying to fathom possible mechanisms for the action of n-3 and n-6 PUFAs in this context is like trying to understand and apply work in progress. After all, at least 41 eicosanoids derive from AA and EPA via COX and LOX enzyme paths, and AA, EPA and DHA have a number of other biological actions independent of eicosanoid production. Given this, it is perhaps more productive to focus on the human epidemiologic and clinical results, knowing that there are probably plausible biological mechanisms even if they are not fully understood at this point.

However, epidemiologic studies also have their problems. When the intake of fish, EPA and DHA is studied in relation to human cancers, only about one-third to one-half of the studies reports a statistically significant reduction in risk for various cancer types. While some of the remaining studies find inverse associations that are not significant, others observed no effect. Larsson et al (63) offer several possible explanations:

1. the intake of long-chain n-3 fatty acids was too low to produce risk reduction;
2. population variation in intake which limits statistical power and as well, incorrect estimates of intake;
3. the critical period for maximum effect may in some cases be in childhood or early adulthood. Information obtained in middle or old age when cancer is diagnosed may miss this aspect;
4. studies that look at fish intake rarely take into account the large variation in the total fat content from species to species as well as the variation in the EPA and DHA content;
5. looking at ALNA (alpha-linolenic acid) intake can be deceiving since the human biochemical conversion to EPA and DHA is both low and variable;
6. many studies fail to take into account the n-6 intake either as LA (linoleic acid) or as AA (arachidonic acid), and since there are significant interactions with EPA biochemistry, as pointed out above, this can seriously confuse the issue, especially in populations where the intake of n-6 PUFAs is very high and n-3 intake low (e.g. the typical Western diet);
7. if it is indeed true that an important mechanism for risk reduction involves the toxic action of oxidized long-chain fatty acids on cancer cells with the concomitant interaction with antioxidants, then one might expect that in countries like the US and Canada, where antioxidant supplements are popular, studies might find lower levels of protection from n-3 PUFAs.

NON-STEROIDAL ANTI-INFLAMMATORY DRUGS AND CANCER
There is a logic trap here: (a) NSAIDs reduce inflammation; (b) NSAIDs reduce the incidence of disease X; (c) therefore the etiology of disease X involves inflammation. The problem is that NSAIDs probably do many other things besides inhibiting the production of eicosanoids and cytokines involved in inflammation, and until all these other activities are discovered and ruled out as the reason for the anti-cancer action, the conclusion is attractive but tentative.

Epidemiologic studies suggest that the regular use of NSAIDs reduces the risk of several types of cancer, suggesting that these drugs may suppress tumor initiation or growth (75). NSAIDs are inhibitors of the COX enzymes and thus can suppress the production of a variety of eicosanoids that arise from EPA and AA (at least 17 follow COX dependent pathways). Some of these eicosanoids are thought to be involved in both the initiation of cancer and the promotion of tumor growth through pro-inflammatory or other actions. Thus inhibiting the enzymes involved in their production provides a potential mechanism for the observed anti-cancer activity. In addition, NSAIDs are implicated as anti-angiogenesis agents and this is also a commonly proposed mechanism for their observed anti-cancer activity. However, as pointed out above, not only are many details of the initiation and development of cancer still to be elaborated, inhibiting the COX enzymes involves potentially the inhibition of the production of a number of eicosanoids, and at present there is a lack of detailed knowledge of how each COX-dependent eicosanoid is or is not involved in carcinogenesis, tumor growth or metastasis. Mechanisms that are complete and satisfying must of necessity await more research. In addition, the sheer complexity of the eicosanoid system suggests caution in the use of inhibitors. In fact, one of the mechanisms suggested for the excess CV (cardiovascular) risks found with Vioxx involved disturbing the balance in the production of eicosanoids involved in promoting and inhibiting thrombosis (76).

NSAIDs are implicated in the treatment or prevention in the following sites: Colorectal (77-80); Breast (60,81- 84); Prostate (85-88); Esophagus and Stomach (89-91); Brain (92); and Lung (93). The connection between inflammation and cancer in some of these sites will be discussed below.

The downside associated with the notion of using NSAIDs for cancer prevention is that they can cause adverse gastrointestinal events including serious and even fatal bleeding. This in fact is a leading cause for hospitalization among the elderly who take these drugs for pain related to arthritis and other disorders, and NSAIDs are thought to be responsible for the largest number of deaths attributable to any class of therapeutic agent (60). Nevertheless, over 80 million aspirin tablets are consumed each day in the US (94). For those who must take this class of drug for pain, the possibility of cancer prevention is, however, an added benefit.

INFLAMMATION AND BREAST CANCER

POLYUNSATURATED FATS AND BREAST CANCER
Epidemiologic studies have provided a somewhat confusing and inconsistent picture of the relationship between n-3 and n-6 PUFAs intake and breast cancer (BC). Cohort and case-control studies frequently differ, and the studies themselves use different methods and analysis. Also many studies do not allow a meaningful examination of the influence of age, menopausal status or the influence of total n-6 intake and the n-6 to n-3 ratio. Many studies of BC and fish intake fail to take into account the type of fish or the amounts of n-3 PUFAs consumed. It appears that the n-6 dietary intake is also a confounding factor, but is rarely considered, even though there is a large variation. All in all, this turns out to be a difficult area from the point of view of definitive studies and clear indications. Thus meta-analyses appear to offer only limited insight. There are two types of study of interest, the case-control and cohort designs. In the case-control study, diagnosed cases are matched to one or more controls, and the intake of PUFAs is estimated from food frequency questionnaires (FFQ) or blood markers. FFQ based results can be subject to recall bias since they are carried out post-diagnosis for the cases. For blood markers such as EPA and DHA, the content of these FAs (fatty acids) in red blood cells (RBC), total serum fat, serum phospholipids, or fat tissue are used. Only the latter looks back for a significant but unknown time. The RBC method gives a picture of fatty acid consumption essentially at the time of sampling, when the patient already has significant disease. Thus it can be argued that the tissue fat (adipose tissue) approach is more appropriate when markers are used. Cohort studies on the other hand enroll large numbers of subjects, follow their dietary and lifestyle habits over a number of years by interview and FFQ, and observe, in this case, the frequency of breast cancer. Only this type of study has the potential to provide information regarding long-term nutrient intake and associated risk. The following recent studies are of interest:

• In a review (68) that summarized studies up to about the year 2002 of the effect of fish or fish oil consumption on the risk of BC, of 21 case-control studies only 2 showed evidence of protection and one had a significant trend (using the yardstick that for significance, the upper 95% confidence limit must not include or exceed 1.0). Of 5 studies using fat tissue as a source of markers, only one study found a high n-3:n-6 ratio protective and one found DHA significantly lower in cases than controls.

• A large pooled cohort study (351,821 subjects, 732 cases) was reported (95) in 2001. No association was found with any PUFA and BC risk. Analysis was based on substituting 5% of energy from carbohydrates with various types of fat. These results are consistent with the large Nurses' Health Study (96).

• In 2004, Saadatian-Elahi (97) published a meta-analysis of studies that used biomarkers. Case-control and cohort studies were considered separately. The cohort studies did not include those in the above mentioned study (95). In the case-control studies, only high levels of ALNA (alpha-linolenic acid) showed protection against BC, whereas in the cohort studies, relative risks of 0.58 for total n-3 PUFAs, 0.66 for DHA and 0.91 for EPA were statistically significant. Studies using RBC and serum phospholipids assays were combined and studies using tissue fat were not included.

• In 2003, Chajes and Bougnoux (98) published a review that emphasized the n-3:n-6 ratio in relation to risk of BC. Nine studies were examined. Of the four that used blood biomarkers, no significant results were found. Of the five that used fat tissue, one found significant protection from BC only in postmenopausal women, and two found significant protection associated with a high n-3:n-6 ratio and in addition one of these also found significant protection from total n-3 PUFAs. The authors conclude that the overall picture supports the idea that the protective effect of n-3 PUFAs depends on the background levels of n-6 PUFAs. What is significant may be the ratio and not the absolute amounts either consumed or present in tissue. This is in keeping with the criticism of many studies that no account was taken of the n-6 intake or levels which now appear to be a possible and significant confounding factor.

• A recent cohort study (99), the Singapore Chinese Health Study, bears mentioning because of the attention to the interplay of n-3 and n-6 PUFAs. The study, based on a FFQ and follow-up approach, enrolled over 35,000 subjects age 45-74 between 1993 and 1998. As of the end of 2000, 314 cases of BC were recorded. Overall there was no association with risk and n-6 PUFAs, but in the subgroup of subjects who consumed low levels of marine n-3 PUFAs, a statistically significant increase in risk was observed to be associated with n-6 intake (RR = 1.87). Also, high levels of dietary n-3 fatty acids from fish/shellfish were significantly associated with reduced risk. Relative to the lowest quartile of intake, individuals in the higher three quartiles had a 26% reduction in risk. Stronger associations were found with post- rather than pre-menopausal subjects.

• The above study was followed by a recent examination (100) of the hypothesis mentioned above of a direct role for peroxidation products from marine n-3 fatty acids and BC protection. Because a glutathione transferase enzyme (GST) is thought to be the principal catalyst for the elimination of these oxidation products, the authors examined a sub-group of women genetically predisposed to low GST activity (the null genotypes) since on the basis of the hypothesis they would have the greatest protection as compared to other genotypes. This was borne out by the study of the cases drawn from the Singapore Chinese Health Study (99). Women with the high activity GST genotype exhibited no protective effect from n-3 consumption, whereas those with the null genotypes showed a statistically significant reduction in BC risk with an impressive odds ratio of 0.34. Thus in studies of n-3 PUFAs on BC risk, the "responders" may be diluted by the "non-responders", and as well, this study adds credibility to the observation that antioxidants interfere with some of the beneficial actions of EPA and DHA (101,102). Also implied is the presence of non-inflammatory, non-eicosanoid related actions for EPA and DHA, in this case, the cytotoxic action of their oxidation products, and that in populations where taking antioxidants is common the results of studies may be significantly impacted. This might also explain the inconsistent results that characterize attempts to determine the effectiveness of the n-3 PUFA in this context.

The question of the interaction of antioxidants and the n-3 PUFAs has been addressed in two very recent case- control studies involving a fairly homogeneous population of French Canadians in Montreal, Canada. The first (103) determined dietary intake with a FFQ. No overall association was found between specific fatty acids and BC risk. However, postmenopausal women with low vitamin E intake exhibited a statistically significant, dose- dependent relationship between AA status and BC risk, with an odds ratio of 0.41. That is, the lower the vitamin E status or intake, the lower the risk. In a second study (104) of the same population, the intake of specific carotenoids and fatty acids was examined in connection with BC risk. In postmenopausal women, total carotenoids were positively associated with BC risk in those with high AA intake and inversely associated with those with a high DHA intake. Das (102) has suggested that the long-chain fatty acids not only induce cell death by enhancing lipid peroxidation, but also act by suppressing or enhancing other cellular processes that act at the gene/oncogene level to produce cytotoxic action on tumor cells. In view of the proposed negative effects of lipid peroxidation in the context of CHD (coronary heart disease), and the evidence, albeit controversial and inconsistent, regarding the effect of antioxidants on the risk of CHD, this hypothesis that lipid peroxidation products are in fact cytotoxic to cancer cells is an odd twist of fate. More studies have been suggested (101)!

Thus while a very large number of studies related to this subject are in the medical literature, it is clear that a definite evidence-based conclusion must await more carefully conducted studies. Nevertheless, the hypothesis is attractive and is supported by the observed low incidence of BC in populations that consume a lot of fish and as well, low cancer rates are seen in those who consume the traditional diet of Crete which contained about 30 times the amount of fish that is common in the present Western diet (12). In view of the benefits of high n-3 PUFA consumption to be discussed below in connection with other diseases, additional reasons for increased n- 3 consumption do not appear necessary and the possibility of protection from BC is merely an added bonus.

NSAIDs AND BREAST CANCER
Cell culture and animal studies have consistently shown that NSAIDs inhibit mammary carcinogenesis, and there are now a number of epidemiologic investigations that have provided evidence for the breast cancer risk reduction associated with the use of aspirin, other non-specific NSAIDs (e.g. ibuprofen) and specific COX-2 inhibitors such as Celebrex (60,84). A recent meta-analysis of six cohort and eight case-control studies found that the use of NSAIDs was associated with a significant 22% decrease in breast cancer risk (105). Results from the prospective Woman's Health Initiative Study of non-specific NSAIDs yielded similar results (84). Regular use of a NSAID for 5-9 years produced a 21% reduction in breast cancer risk and for > 10 years use the reduction was 28%, and there was a statistically significant inverse trend with duration of use. Long-term ibuprofen use resulted in a risk reduction of nearly 50% which was larger than the relative risk reduction of 0.7, with aspirin. In a case-control study (81,94) reported in May 2004, a similar result was obtained for aspirin use which was frequency and dose dependent. In this study, ibuprofen was less effective compared to aspirin. The case- control design of this study made it possible to examine the connection between aspirin's protective action and the estrogen or progesterone receptor status. It was found that the aspirin associated risk reduction was seen only among women with hormone receptor-positive tumors (81). Finally, low dose aspirin (<100mg) such as is used for heart attack prevention was found in one study to be ineffective in breast cancer prevention (84).

Recent studies have confirmed the earlier observation that the COX-2 enzyme is over-expressed in invasive human breast cancer and this impacts survival (106). A number of studies are now in progress using COX-2 inhibitors, and in particular Celebrex, either alone or in conjunction with an estrogen inhibitor, in an attempt to both prevent and treat breast cancer (82,83). Preliminary results are encouraging. The connection with the COX-2 enzyme is thought to be related, in part, to its role in the generation of the prostaglandin PGE-2. Increased levels of this eicosanoid can influence the normal process of cell death (apoptosis), cell invasion, immune function and tumor-related angiogenesis (82,83,107). PGE-2 can also influence the concentration of an enzyme involved in estrogen production. Since estrogen stimulates PGE-2 production, positive feed-back can result and a vicious cycle leading to an estrogen promoted increase in tumor cell proliferation (83). Non-specific NSAIDs like aspirin and ibuprofen also impact the production of the prostaglandin PGE-2 which incidentally, as discussed above, is derived from AA.

INFLAMMATION AND PROSTATE CANCER

Cell culture and animal studies and a few epidemiologic studies have provided the basis for the hypothesis that prostate cancer (PC) has an inflammatory component. However, the evidence is far from compelling. There is weak evidence that acute or chronic bacterial prostatitis may be associated with PC (108), and while there is sufficient evidence indicating that chronic inflammation may be a legitimate target for chemoprevention, it appears clear that more studies are required to establish inflammation's role in PC (109,110).

POLYUNSATURATED FATS AND PROSTATE CANCER
The subject of dietary n-6 and n-3 PUFAs and prostate cancer (PC) risk with emphasis on epidemiologic and experimental evidence has recently been reviewed by Pierre Astrog (111). A large number of studies are critically examined. What emerges is that while there are some indications favoring n-3 PUFAs from fish, in fact there is little epidemiologic support in general for the long chain n-3 PUFAs such as EPA and DHA in this context. A puzzling finding from some studies is that there appears to be an increased risk of PC for men having higher intake or higher blood levels of ALNA. However, other studies fail to find this association. The conversion efficiency of ALNA to EPA and DHA is very low, and ALNA may act in some fashion unrelated to eicosanoid chemistry. Also, since supplemental EPA and DHA were not considered, the only significant source may have been seafood, and the levels consumed may not have been large enough to yield a protective effect. The odd result with ALNA also appears specific to PC, since such a positive association has not been found for breast or colon cancer, and in fact, ALNA appears to lower the risk of BC. Little or no evidence was found linking either LA acid or AA to the risk of PC.

A very recently published observational study (112) by Leitzmann et al based on over 47,000 men with no cancer followed for 14 years found results similar to those summarized by Astrog. In this study it was also found that ALNA may increase the risk of advanced prostate cancer, but EPA and DHA may reduce the risk of total and advanced PC. This paper also contains a good review of past studies, which highlights the inconsistent picture that emerges. In addition, a new meta-analysis (113) also found increased risk of prostate cancer with high intake or high blood levels of ALNA whereas a beneficial effect was found for heart disease. Leitzmann et al comment on this apparent dilemma. It can be concluded that more studies are needed to resolve the inconsistencies and contradictions that characterize the literature regarding PUFAs and PC, but EPA and DHA do emerge as potential protective agents. Men might want to consider limiting the intake of ALNA from, for example, flax seeds or flax oil.

NSAIDs AND PROSTATE CANCER
In the last 10 years there have been 4 case-control studies (85,114-116) and one follow-up study (86) that examined the question of the effect of NSAIDs on PC risk. Four found inverse relations, mostly statistically significant, and one gave a puzzling positive association. None of the studies examined the COX-2 inhibitors and aspirin was the most commonly studied NSAID. These studies provide limited guidance because of the inconsistent reporting of duration of use, dose data, age and NSAID type stratification. In addition, as Michael Barry points out in an editorial (87), there is considerable potential for serious confounding, especially since there was no correction for vitamin E and selenium intake, a popular anti-PC supplement strategy and a combination currently in a large clinical trial (the SELECT trial) because of findings of strong preventive effects (63% reduction in incidence with selenium, 32% with vitamin E) in trials conducted for other purposes (117,118).

The COX-2 level of expression in vivo appears to be higher in neoplastic as compared to benign prostate glands, although there are conflicting reports on this issue (88). COX-2 is also associated with increased levels of PGE-2 which may mediate cancer cell proliferation. However, there appear to be no clinical or epidemiologic studies that are relevant, and at present there appear to be only two ongoing studies dealing with prevention. One is using Celebrex (88). Merck had started a study with Vioxx, but it is unlikely that that study will continue, given the fact that the drug is now withdrawn from markets worldwide due to excess cardiovascular events that occurred in another cancer prevention trial.

For men undecided about taking aspirin to reduce their risk of fatal and nonfatal heart attack, the above studies might tip their personal risk-benefit analysis in favor of taking aspirin. However, at this point there is no information relating to the effect of aspirin or other NSAIDs use on PC mortality, but some relating just to the reduced risk of PC. The question of dose remains unresolved.

INFLAMMATION AND COLORECTAL CANCER

The development of colorectal cancer (CRC) in the majority of cases is thought to proceed first with the formation of polyps (a small growth projecting from the mucosa of the large intestine) which then develop into larger, tumor like growths termed adenomas, which represent the precancerous state and can go on to develop into full blown invasive and metastatic cancer. Hereditary predisposition to polyp formation, termed Familial Adenomatous Polyposis (FAP) is present in 1-2% of patients diagnosed with colon carcinoma. Cell culture, epidemiologic and animal studies have provided evidence for a connection between inflammation and CRC. Of interest is the potential preventive action of NSAIDs and in particular aspirin and the specific COX-2 inhibitors.

NSAIDs AND THE PREVENTION OF COLORECTAL CANCER
There appear to have been only three randomized clinical trials of aspirin for the prevention of CRC (119), and these all involved studies of adenoma recurrence rather than primary prevention. One study (120) found a reduction with 300 mg/d use but not with 160 mg/d, but the results failed to achieve statistical significance because of the low number of patients in both aspirin groups. The second study (121) randomized 635 patients to receive either 325 mg/d (the standard strength aspirin dose per pill) or a placebo. A statistically significant relative risk (RR) of 0.65 for any recurrent adenoma was found, and as well, aspirin delayed the appearance of the first adenoma. The third trial (122) randomly assigned 1121 patients to receive either 81 mg/d (the typical dose used for prevention of heart attacks), 325 mg/d or a placebo. Only the low dose of aspirin produced a significant reduction in the relative risk of recurrence of one or more adenomas (RR = 0.81). For advanced neoplasmas the low dose yielded a significant RR of 0.59. The authors suggest that the failure to achieve significant reduction with the higher dose, a result that conflicts with some studies, was a matter of chance since the low and high doses of aspirin have both been found to reduce colorectal prostaglandin levels to a similar extent. They also suggest that various eicosanoids have anti-carcinogenic effects and excessive suppression could have deleterious effects. It is unlikely that there will be large-scale clinical trials addressing the question of primary prevention of CRC with aspirin because of the long follow-up and large sample required, especially for primary prevention. Nevertheless, the above results seem highly suggestive that aspirin intake is associated with a protective effect, especially since they involved randomized, controlled clinical trials which were in contrast to the observational studies to be discussed below.

In a meta-analysis published in 2004 (119) which included results of randomized clinical trials up to late 2003 that used NSAIDs for the prevention of adenomas and CRC, three studies using aspirin that met the criteria for inclusion yielded a relative risk of 0.77 for recurrent sporadic colorectal adenomas. In three trials involving patients with FAP, use of the NSAID yielded the result that users had an 11.9 to 44% reduction in the number of colorectal adenomas compared to the control group that had 4.5 to 10%. These later studies were short-term and indicated the support of NSAIDs (silindac or Celebrex) in the above clinical setting.

Garcia Rodriguez and Huerta-Alverez (78) have presented a pooled analysis of cohort and case-control studies of the effect of aspirin on the incidence of CRC published from 1985 to 1999. For fourteen studies they found a highly significant RR of 0.71 (9 showed a significant RR, one a non-significant null result, and 3 each a non- significant but still inverse relationships with RRs in the range of 0.38 to 0.85). What appears to be the most recent observational study was published in February 2004 (123). It is from Harvard and is based on the data obtained in the famous Nurses' Health Study. The objective was to examine the dose-duration aspect of aspirin use in the primary prevention of colorectal adenoma. Over 27,000 women participated. It was found that women who regularly used aspirin (> 2 standard tablets/week) had a significant adjusted RR of 0.75 compared with non-regular users. When non-users were used for comparison, the RR for 0.5-1.5 standard tablets per week was 0.8, while 2-5 tablets per week gave a RR of 0.74, 6-14 tablets a RR of 0.72, and > 14 tablets a week a RR of 0.65. These results were all statistically significant and the trend with frequency of use had a probability of occurring by chance of only 1/1000. Similar dose-response relationships were found for both short term (<5years) and long term (>5 years) use. The authors point out that similar results were obtained for men in the Health Professionals Follow-up Study published in 1994 (124). Thus while there are some inconsistencies among the above discussed studies, the same general picture emerges. The greatest effect in primary prevention of CRC or colorectal adenomas may be associated with relatively high doses of aspirin (>2 tablets— 325 mg—per day), but the difference between 1-2 a day vs. >2 was not great in the Harvard study.

The downside associated with daily aspirin consumption, especially at high doses, is the risk of hemorrhagic stroke and serious gastrointestinal (GI) side effects (bleeding). In a meta analysis (125) published in 2002 which had as its goal to establish clinical guidelines for the use of aspirin in primary prevention of cardiovascular events, it was found that the rate of hemorrhagic strokes due to aspirin was 0-2 per 1000 patients per 5 years, whereas for major GI bleeding events, the prediction was 2-4 per 1000 patients per 5 years. The aspirin dose ranged from 75 to 500 mg/d but dose was considered an issue only in the context of stroke, where the authors quote a cut-off of 175mg/d below which the risk becomes statistically insignificant. In this context, Ladabaum et al (126) argue that aspirin chemoprophylaxis is not a reasonable substitute for colorectal cancer screening (e.g. sigmoidoscopy every 5 years and fecal occult blood testing every year or a colonoscopy every 10 years).

The effects of COX-2 inhibitors on animal models of CRC have demonstrated effective inhibition of tumor growth in a number of studies (127), and this has been a primary driving force for human studies. In 2000 Steinbach et al (128) published a study restricted to patients with FAP. After 6 months of twice daily treatment with 400 mg of Celebrex, a significant reduction in the number of colorectal polyps was observed. This appears to be the only published clinical study involved in the FDA approval of Celebrex for this indication, and in fact the results are presented in a dramatic figure in the Physician's Desk Reference in the section on Celebrex. There appears to be only two other studies published to date for the use of COX-2 inhibitors in this context. Rahme et al (129) in a case-control study examined the effect of both Celebrex and Vioxx on the occurrence or recurrence of colorectal neoplasia in average to high risk patients. Three months exposure to COX-2 inhibitors conferred a significant protective effect against carcinomas and colorectal adenomas. The non-selective NSAIDs (e.g. aspirin) were more effective than Vioxx which was more effective than Celebrex. The authors point out that this is the first demonstration that specific COX-2 inhibitors offer protection against these lesions in the general population.

The above results, along with animal and cell culture studies have prompted a surge in interest and clinical studies using Celebrex for primary and secondary prevention of colorectal adenomas and CRC which are now ongoing. As is now well known, it was a study using Vioxx for this application that finally solidified the case for excess adverse CV events and its withdrawal from the worldwide market (see the Dec-Jan 05 IHN for a perspective on this rather sensational event). It is known that COX-2 is highly expressed (present at abnormal concentrations) in CRC tumor cells. Potential mechanisms for the action of COX-2 inhibitors on the various stages of CRC include suppression of cell proliferation and a more favorable ratio of cell death (apoptosis) to proliferation, i.e. a cytotoxic effect (130). COX-2 inhibitors may also interfere with angiogenesis and stimulate immune surveillance (131). COX-2 is known to stimulate angiogenesis partly via the production of certain eicosanoids. However, it should be clear that this application of COX-2 inhibitors is only in its initial stages, and aside from the approved use in FAP, their application in primary prevention of CRC or the precursor to colon growths is speculative and routine use appears hardly advisable considering the potential for adverse GI events. Also, questions are just now being raised about Celebrex and the risk of adverse CV events. FAP obviously presents a difficult risk-benefit problem. However, for individuals taking Celebrex for arthritis or other pain producing problems, the potential for protection against this common cancer should be comforting.

N-3 AND N-6 PUFAs AND COLORECTAL CANCER
There is remarkably little evidence from human studies on the effect of n-3 and n-6 PUFAs on the primary prevention of CRC. Roynette et al (132) in a review published in 2004 present as evidence for a protective affect of n-3 PUFAs the following:

• The lower incidence of CRC in Greenland Eskimo populations eating their traditional diet compared to populations in the West (>10 g/d of long-chain PUFAs EPA and DHA compared to < 0.25 g/d).
• Japanese migrants to the US who adopt the American diet have increased CRC incidence compared to their counterparts in Japan.
• Data from 24 European countries indicates that a high n-6 to n-3 ratio of PUFAs increases the risk for CRC.

It is well known that such arguments have significant weaknesses and can be subject to serious confounding. Observational studies are limited. In a study that combined results from case-control studies in Switzerland and Italy, Tavani et al (133) found a relative risk (RR) of 0.7 for CRC when the first and fifth quintile of n-3 PUFAs intakes were compared. However, Kobayashi et al (134) found no effect of fish consumption on the etiology of CRC and Lin et al (135) failed to find significant associations with fish or either n-3 or n-6 PUFAs, although the n- 3 fats were not stratified. They also found red meat significantly protective, a result at variance with some studies. In a case-control study that included stratification according to the presence of a COX-2 promoting genotype, Koh et al (136) found a statistically increased risk of CRC for the combination of this gene type and high intake of dietary n-6 PUFAs, which they hypothesize is due to prostaglandins playing an important role in colorectal carcinogenesis by enhancing cell proliferation and growth, and by promoting angiogenesis and inhibiting apoptosis. Aside from cell culture studies, this is essentially where the subject of the role of the PUFAs in the prevention of CRC stands at the moment, and the case for intervention through increased n-3 consumption is weak and to a large extent theoretical.

INFLAMMATION AND LUNG CANCER

It is universally accepted that there is a connection between smoking and lung cancer, but in addition, the inhalation of particulate matter such as nickel oxide (from auto exhaust), crystalline silica, some wood dusts, asbestos and refractory ceramic fibers (from insulation) are all thought to be carcinogenic. Particle accumulation in the lung creates a milieu where inflammatory cell influx and the release of oxidants play a role to generate a pro-mutagenic environment that can lead to malignant lung disease (137,138). Previous lung disease is also implicated (139,140). Thus the indication that inflammation may play an important etiological role. Also, chronic bronchitis, emphysema and pulmonary tuberculosis have been positively related with lung cancer incidence, although the evidence appears strongest for emphysema. These associations remain after correcting for smoking.

A connection with inflammation prompts the question of the role of PUFAs and NSAIDs. As regards the former, there appears to be almost no significant literature bearing on prevention or progression. NSAIDs, however, have received some attention. For aspirin, two recent case-control studies (141,142) found a statistical reduction in risk associated with more than occasional use. On the other hand, a large prospective study published in 2003 (143) found that regular aspirin use was not associated with reduced lung cancer risk in men. This paper also reviewed older studies and points out that when viewed together, epidemiologic studies have provided in fact a rather mixed picture. However, results of this study conflict with that of Ratnasinghe et al (144) published a year later who found an RR of 0.6, among male aspirin users.

A number of studies have indicated a close relationship between the COX-2 enzyme and lung carcinogenesis and progression (145). Thus there is considerable interest in the use of specific COX-2 inhibitors both for primary prevention and for adjuvant use with radiation and/or conventional chemotherapy (145,146). While studies are now underway, little has been reported even of a preliminary nature. Progress will present challenges since lung cancer is highly heterogeneous requiring subgroup identification, and in addition, smoking, diet and environmental exposure to particulates can confound the results.

INFLAMMATION AND OTHER CANCERS

Very preliminary research has been done on the role of inflammation in the etiology of pancreatic, esophageal, gastric, ovary and brain cancer (89,147,148). At this point, the results have been mostly hypothesis generating and much research has to be done before definite conclusions and preventive recommendations can be made based on scientific evidence.

INFLAMMATION AND RHEUMATOID ARTHRITIS

Rheumatoid arthritis (RA) is a classic example of an inflammatory auto-immune disease, and its most obvious manifestation is joint pain and dysfunction. The Japanese, who have a diet rich in n-3 PUFAs from fish have a lower rate of RA than is observed in Western countries. This is in spite of the fact that a genetic predisposition toward RA is more common in Japanese than in most other populations, leading to a prediction of a higher prevalence of RA, whereas the opposite is seen.

According to a just published pair of papers from the Mayo Clinic, CHD (coronary heart disease) and congestive heart failure (CHF) not only occur much more frequently in RA patients than in the general population, but also do not present as expected (149,150). The authors caution that in RA patients the absence of traditional cardiovascular risk factors does not rule out the presence of CV disease. Infarctions are also frequently accompanied by minimal or no symptoms and escape detection until sudden death occurs. One of the pathological mechanisms implicated is a lack of balance between pro- and anti-inflammatory cytokines. The research also revealed that RA patients have twice the risk of developing CHF. The authors regard this as evidence for the hypothesis that systemic inflammation may promote the development of CHF(149).

James et al (151) have recently reviewed the use of n-3 PUFAs for both prevention and therapy. As regards the former, in a case-control study of women with RA, fish consumption was lower in the cases than in the healthy controls. Being in the top 10% for n-3 intake (>1.6 g/d) was associated with a 60% decrease in the probability of having blood markers indicating RA. As James et al point out, there is a need for controlled intervention studies in the context of prevention. As regards treatment, the authors of this review examined 13 double-blind randomized controlled trials that examined the therapeutic effect of fish oil on established RA. In 12 of the 13 studies, there was improvement in outcome measures, the most common being improvement in tender joint count. The use of fish oil supplementation also is observed to decrease the use of NSAIDs. The question of dose response is still open, but in the studies James et al discuss, there was more rapid clinical improvement with 5.9 g/d of EPA + DHA vs. 2.9 g/d. This is consistent with a recent study that found 1.6 g/d of EPA + DHA to be ineffective (152).

The use of NSAIDs in the treatment of RA is standard practice, and this was the principal use of the specific COX-2 inhibitors. Now that Vioxx is withdrawn and the safety of Celebrex is being questioned, there may be more interest in "natural" approaches to pain relief. In this connection, it is worth mentioning that the stomach friendliness of Celebrex has also been called into question and there are those who consider it no better in this respect than non-specific NSAIDs (153,154).

Rennie et al (155) have recently reviewed the subject of nutritional management of RA and concludes that there is evidence to back up the recommendation that patients should consume a diet rich in the long-chain n-3 PUFAs and as well, antioxidants. The authors also point out that supplementation with EPA and DHA consistently demonstrates an improvement in symptoms and a decrease in NSAID use. There is also clinical evidence that a diet low in AA (arachidonic acid) ameliorates the clinical signs of inflammation in RA patients and augments the beneficial effects of fish oil supplementation (156). The subject of anti-inflammatory diets will be discussed later in this review.

N-3 FATTY ACIDS AND ALLERGIC DISEASE

The large increase in allergic disease in Westernized countries in the past few decades has prompted the hypothesis that one of the aggravating factors might be the huge change in the n-6:n-3 ratio of PUFAs in the modern diet. However, the use of supplemental n-3 PUFAs has had only modest success in impacting either the treatment or prevention of allergies and in particular asthma. One area of interest involves the impact of the n-3 PUFAs during pregnancy on allergies in infants. Large studies are necessary at this point before conclusions can be drawn (157).

INFLAMMATION AND CARDIOVASCULAR DISEASE

While over the last decade the focus in atherosclerosis has been on blood lipids and cholesterol lowering, it is well known that something like half of all adverse cardiovascular (CV) events occur in individuals with a normal blood lipid (cholesterol) profile, many of whom appear perfectly healthy. This has prompted the search for novel risk factors unrelated directly to blood lipids. Considerable attention has been directed at the role and interpretation of the serum markers of inflammation interconnection with CVD, especially the interleukins and C- reactive protein (CRP), although only CRP is currently attractive as a marker for widespread clinical use since the high-sensitivity assay is now standardized and widely available at a low cost. In prospective studies, an elevated level of CRP is associated with increased risk of CV events even in apparently healthy individuals. It also has prognostic value in patients with a prior history of CV disease. The combination of high total cholesterol, low HDL and a high CRP is associated with greatly enhanced risk of heart attack or stroke (see the IHN Research Report on CRP and Heart Disease—published February 2003). But there still remains the question of whether these markers have a causal relation to coronary heart disease (CHD) or are merely markers of the underlying disease process, or both (158). While there is no move toward a general recommendation for measuring CRP levels along with blood lipids for routine risk assessment, it appears generally accepted that such measurements have a place in therapeutic decision making in patients who are at borderline risk based on traditional risk factors (158). One argument for CRP testing is that tests for this marker and for LDL may be each detecting different high-risk groups. CRP measurements have been found to add information of prognostic value at all levels of serum LDL, at all levels of the Metabolic Syndrome, and at all levels of the Framingham Risk Score (160).

An interesting comparison of CRP versus other risk markers of CVD was carried out in the Women's Health Study (160). Four inflammatory markers plus homocysteine, lipoprotein (a), serum amyloid and LDL were compared. In a long-term follow-up, CRP provided the strongest predictive power. In the context of primary prevention of CVD, CRP may be a stronger predictor than even LDL. Patients with elevated levels of both LDL and CRP had 8 times the CV risk as compared to those with low levels of both markers. It is interesting that 46% of the adverse events occurred in patients with LDL < 130 mg/dL, one of the currently accepted goal for primary prevention (160).

STATIN DRUGS AND INFLAMMATION
It has been suspected for a number of years that the statin drugs have beneficial effects over and above their ability to reduce cholesterol levels. There have now been a number of studies (158,160) where even short term statin treatment has resulted in significant decrease in CRP levels, and this reduction was in an LDL independent manner, suggesting both anti-inflammatory as well as lipid lowering effects. In one study, statin treatment was effective in preventing acute coronary events in a primary prevention cohort with elevated CRP, regardless of the baseline levels of LDL or the total cholesterol:HDL ratio.

Two studies published in 2005 relate to the statin-inflammation question. In one (161) intravascular ultrasonography was used to assess the progression atherosclerosis in patients with documented CHD. Moderate (40 mg/d of pravastatin) or intensive (80 mg/d of atorvastatin) interventions were compared. Reduced rate of progression associated with intensive treatment as compared to moderate treatment was found to be significantly related to greater reductions in the levels of both atherogenic lipoproteins and CRP. In the second study (162), which involved the assessment of risk of recurrent MI or death from CHD among patients with acute coronary syndromes, patients who had low CRP levels after statin therapy had better clinical outcomes than those with high CRP, regardless of the level of LDL cholesterol achieved.

Mechanisms for the anti-inflammatory action of the statin drugs have been suggested (160), but much research remains to be done before the multiple actions of these drugs are fully understood. Willerson and Ridker (160) assert that statins are the most effective agents available today for the reduction of vascular inflammation. However, because of the side effects occasionally associated with this class of drug, some of which are very serious, many would not consider this an ideal solution to the problem of inflammation reduction. In this connection, it is interesting that William R. Davis, a cardiologist who has been very active in promoting the so- called calcium scan for measuring and monitoring the plaque load in patients presenting with heart disease symptoms, comments in his new book Track Your Plaque (163) that while the drug companies claim that the incidence of statin caused muscle aches and pain is of the order of 2-3% (not the full-blown muscle inflammation and damage known as rhabdomyolysis), he and his colleagues have found in their own practices that the number is more like 30%. He also finds that 100 mg/day of coenzyme Q-10 (CoQ-10) generally reduces or eliminates the problem within several days. Statin drugs deplete cellular CoQ-10 (164), a side effect that was recognized early in the history of this class of drug. Merck has two patents on the combination of a statin drug with CoQ-10, but such formulations have for some reason never been marketed, nor, it would appear, is there a widespread appreciation of this side effect. As regards rhabdomyolysis, a very recent study (165) found after an analysis of the data bases of 11 health maintenance organizations, that the number of patients needed to treat with the popular statins (Lipitor, Pravachol and Zocor) to generate one case of this muscle disease per year was 22,727, which seems consistent with the claim of low risk. However, the number for the recently withdrawn statin cerivastatin (Baycol) was a shocking 1873.

NSAIDs AND CARDIOVASCULAR EVENTS
Today, when one thinks about anti-inflammatory drugs, both the prescription and over-the-counter COX inhibitors immediately come to mind, including ibuprofen, naproxen, aspirin, the heavily advertised Vioxx and Celebrex, etc. After the withdrawal of Vioxx, the remaining COX-2 inhibitors are coming under increasing scrutiny regarding adverse CV events and would not appear to be candidates for primary or secondary prevention of CHD, although the use of Mobicox in connection with acute coronary syndrome may be significant (166). This leaves the non-specific NSAIDs and since aspirin is unique in this class because of its so-called anti- platelet activity, it is the only NSAID that appears to merit consideration. Aspirin irreversibly modifies COX-1 and COX-2, but the affinity for the former is 50-100 times greater than for COX-2. The COX-1 inhibition blocks eicosanoid production from arachidonic acid (AA) which significantly inhibits platelet thromboxane biosynthesis with the resultant anti-platelet activity. Thromboxane is a potent prothrombotic and vasoconstrictive agent. One dose of 325 mg of aspirin achieves full inhibition in about 3 hours which lasts for several days, but with doses of 40-80 mg/d, a cumulative effect is seen after 4 days and low doses can then maintain this inhibition (167). A meta-analysis of five randomized primary prevention trials by Eidelman et al (168) found a statistically significant 32% reduction in the risk of first heart attack and a 15% reduction in the risk of all important vascular events, but no significant effects on non-fatal stroke or vascular death. The authors regard these results as supporting the position of the American Heart Association and the US Preventive Services Task Force recommendations that individuals with a 10-year (Framingham) risk of a first coronary event that is 10% or greater would find the benefit of aspirin outweighed the risks. In high-risk patients and for secondary prevention, aspirin has a long established therapeutic role (169). Aspirin may have anti-inflammatory action over and above COX inhibition (169).

POLYUNSATURATED FATS AND CARDIOVASCULAR EVENTS
As discussed in detail above, the n-3 PUFAs are in many respects anti-inflammatory, and thus the obvious question, if atherosclerosis and its various clinical manifestations involve inflammation, then can the n-3 PUFAs play a role in primary or secondary prevention? This has been a very active area of epidemiologic and clinical research. In general, high intakes of EPA and DHA combined (2-4g/d) can lower serum triglyceride levels and appear to have mild antihypertensive action as well. The cardio-protective mechanism at intakes below 2-4 g/d appears to be related mainly but not entirely to reduced susceptibility to lethal arrhythmias (170). These benefits may only indirectly involve anti-inflammatory action.

Two large and very long-term prospective studies, one involving over 45,000 men with a 14 year follow-up, the other over 84,000 women with a 16 year follow-up, found inverse associations of CHD and n-3 intake. In the men's study (171) published in 2005, over 45,722 men free of known CV disease were followed for over 14 years. Intake of n-3 and n-6 PUFAs were established by frequent food frequency questionnaires. It was found that plant-based n-3 PUFAs reduced CHD risk when seafood based n-3 consumption is low. Little influence was found from n-6 intake on the benefits of n-3 PUFAs from plant sources or seafood. Men with an intake of EPA + DHA of > 250 mg/d had a reduced risk of sudden death from MI whether n-6 PUFAs intake was high or low.

A study published in 2002 (51) supports the importance of EPA and DHA in the context of sudden cardiac death. Ninety-four men who had sudden death as the first evidence of CVD were matched according to age and smoking habits with 184 men who acted as controls. Blood from cases was frequently collected by paramedics at the scene. As compared with men whose blood levels of EPA and DHA were in the lowest quartile, the relative risk of sudden death was only 0.19 for those in the highest quartile with a significant trend from quartile to quartile. A similar case-control study with almost identical results was reported in 2000 by Siscovick et al (50). They determined the fatty acid composition of the red blood cell membranes and used the sum of EPA and DHA to generate quartiles. These two studies were the basis of the earlier discussion of the utility of the sum EPA + DHA as a marker of inflammation and n-3 status in connection with CVD. The strong connection between n-3 PUFA intake and the risk of sudden cardiac death is generally attributed to an anti-arrhythmia effect.

In the women's study (172), fish consumption 1-3 times a month was associated with a relative risk (RR) of 0.7 for CHD, whereas the RR was 0.71 for once per week, 0.6 for 2-4 times per week, and 0.66 for more than 5 times per week. Across the quintiles of total n-3 fatty acid intake, the RRs were 1.0 (reference), 0.93, 0.78, 0.68, and 0.67. For fish and n-3 fatty acid intake, the inverse association was stronger for CHD deaths as compared to nonfatal MI. For example, the RR for fish consumption 5 times a week was 0.55 for CHD death, 0.73 for nonfatal MI. For diabetic women, a higher consumption of fish and EPA plus DHA was associated with lower CHD incidence and total mortality (173). Differences between men and women are noteworthy. The overall picture of the relationship between fish consumption and CHD is presented in two meta-analysis studies (174,175) published in 2004. Both provide convincing evidence of benefit.

There have been a number of randomized controlled studies of the use of n-3 PUFAs in the prevention of recurrence of adverse cardiac events subsequent to a heart attack. Bucher et al (176) identified 11 studies up to 1999 that met the authors' criteria for acceptability for a meta-analysis. For patients on n-3 enriched diets as compared to placebo, the RR was 0.8 for a non-fatal MI. In 5 trials, sudden death was associated with an RR of 0.7 whereas overall mortality had a RR of 0.8. Dietary and supplement sources were equivalent. This analysis includes the Lyon Heart Study (a secondary prevention clinical trial), where a Mediterranean diet enriched with alpha-linolenic acid vs. a normal European diet produced RRs (relative risks) of 0.3 for both fatal and nonfatal MIs, 0.1 for sudden cardiac death and 0.6 for overall mortality (177). The fact that these rather sensational results occurred in the absence of significant changes in blood lipid profiles strengthened the view of those who consider cholesterol to be only part of the heart disease problem. Also included is the GISSI-Prevenzione trial (178), an intervention trial involving a cohort of individuals who had recently survived a heart attack, which used 1000 mg/d of EPA plus DHA or a placebo. A 20% reduction in mortality was observed. Much of the benefit was attributable to a 53% reduction in cardiac death, a result that emerged in an analysis of the first four months of intervention (179).

Two studies just published relate to the role of n-3 and n-6 PUFAs in heart disease and stroke. In one, researchers (180) found consumption of tuna or other broiled or baked fish was associated with decreased risk (27-30%) of ischemic stroke, but fried fish was associated with increased risk, presumably because of bad fats in the frying oil. In the other study (181), dietary PUFAs and in particular linoleic acid (LA) were found to have a significant cardioprotective benefit. The finding for LA is interesting in connection with the emerging view that the n-6 PUFAs are not as dangerous as first thought and may be beneficial as long as balanced with n-3s.

Adverse atherosclerosis related events generally involve plaque rupture, and thin fibrous caps on plaques rather than thick fibrous caps are thought to increase the risk of rupture. In a recently published study, Thies et al (182) carried out a randomized controlled trial of the association of n-3 PUFAs on the stability of atherosclerotic plaques. Subjects were awaiting carotid endarterectomy (surgical removal of carotid artery plaque). They were randomly assigned to fish oil (1.6 g EPA + DHA per day) or sunflower oil (n-6 fatty acid, and concentrations of EPA, DHA and LA were measured in carotid plaques. As well, the plaque morphology was assessed and the presence of macrophages measured. The time interval of intervention ran between 7 and 190 days. It was found that plaques readily incorporated n-3 PUFAs from fish oil supplementation, and that this induced changes that enhanced plaque stability, whereas n-6 PUFAs did not affect carotid fatty acid composition or stability. The authors comment that the rapid incorporation of n-3 PUFAs suggests that atherosclerotic plaques are fairly dynamic with some degree of lipid turnover even at an advanced stage of atherosclerosis. The most interesting result was that more plaques were seen with well-formed fibrous caps in the fish oil group, even though the intervention period was short, and as well, fewer macrophages were found in the plaques of the fish oil group. Macrophages are thought to make a major contribution to plaque inflammation and instability. Both of these observations relate to a lower risk of rupture in the fish oil group, the importance of inflammation in plaque instability, and the anti-inflammatory role on n-3 PUFAs in atherosclerosis and associated diseases.

INFLAMMATION AND ALZHEIMER'S DISEASE

While Alzheimer's disease is a heterogeneous disorder arising from multiple etiologies, there is now considerable evidence linking inflammatory processes to the pathology in vulnerable regions of the AD brain (183-186). Inflammatory activation of microglia (small non-neuronal cells) is consistently found in senile plaques in AD. Amyloid beta peptides, found in neuritic plaques, are also thought to be centers for inflammatory activation (186). In a large follow-up study of well functioning individuals aged 70-79, high baseline levels of interleukin-6 (IL-6) and CRP were predictive of poorer cognitive performance over two years of follow-up (187). In the 25-year follow-up in the Honolulu-Asia Aging Study, comparison of men in the lowest vs. highest quartiles of CRP found a significant 3-fold increase in risk for all dementias combined and for both AD and vascular dementia (188). A recent meta-analysis of the use of NSAIDs found a protective role against AD for long term use (189). While this does not prove that inflammation is a causative factor, the hypothesis is attractive. Much additional evidence based on animal and cell culture studies and human autopsy data could be quoted for the role of inflammation in AD (see (183,190) and the Research Report on AD in IHN, July/Aug and Oct 2003). It has been argued now for some time that there is a connection between AD and CVD and atherosclerosis. Casserly and Topol (191) recently reinforced this view, suggesting that AD and atherosclerosis are in fact independent but convergent disease processes with one of the links being inflammation. Another way of stating this is that if one accepts that AD is at least in part a vascular disease sharing many risk factors with CVD, then if inflammation plays a role in CVD it might also be expected to be involved in the initiation and progression of AD.

ALZHEIMER'S DISEASE AND NSAIDs
Long-term epidemiologic studies (observational) of traditional NSAIDs with the object of studying primary prevention have shown considerable benefit, but only with two or more years of use. The importance of long- term use is illustrated in the meta-analysis quoted above (189). In this study, the pooled relative risk (RR) of AD among users of NSAIDs was 0.72. When stratified by length of use, the RR was 0.95 for < 1 month, 0.83 for < 24 months, and 0.27 for > 24 months. Only the last result was statistically significant if one requires the upper 95% confidence limit to not include the null value of 1.00. Pooled RR for aspirin use was 0.87 but was not statistically significant. Bas et al (192) obtained a similar non-significant benefit for aspirin in this context. Thus it appears that only long-term use of NSAIDs (traditional, non-specific) offers protection against the development of AD, with in fact dramatic reduction in risk for periods exceeding two years. This leads to the hypothesis that it is only the early pathology that is being targeted and that lengthy exposure to the action of these drugs is necessary (189;192;193). The failure of clinical trials, which all involved patients with early symptomatic AD, to yield positive results is consistent with this view. As van Gool et al (193) point out, classical NSAIDs inhibit both COX-1 and COX-2 enzymes, but also have other actions that are independent of COX activity. Thus it is dangerous to take these results as proof that the observed benefits are only COX inhibitor related. In fact, some NSAIDs are capable of inhibiting the formation of amyloid-beta, thought to be the main player in plaque formation, by a non-inflammatory mechanism (193).

Very recently there have been media reports associated with an unpublished 1999 clinical trial by Pfizer of Celebrex in connection with AD. No benefits were found, but an increased risk of adverse CV events was observed. Dr. Eric Topol, cardiologist at the Cleveland Clinic, has recently examined the data and concludes that the results related to CV events were statistically inconclusive (Feb.2, 2005, www.theheart.org, online newsletter).

ALZHEIMER'S DISEASE AND N-3 PUFAs
Seven studies confirm that there is an inverse relationship between either intake or serum markers of n-3 PUFAs and the risk or progression of AD, vascular dementia or cognitive impairment. In a prospective study, Heude et al (194) used the fatty acid content of red blood cells as a marker. In a four year follow-up, n-6 PUFAs were associated with an increased risk of cognitive decline, whereas the higher the n-3 content, the lower the risk. In the Rotterdam Study (195) which was also prospective and used a food frequency questionnaire, in a 2 year follow-up fish consumption was inversely related to the occurrence of dementia. The relative risk of 0.4 for dementia and 0.3 for AD were both significant and impressively low. In a case-control study, Tuly et al (196) found lower DHA levels in AD patients than in controls. Otsuka et al (197) compared cases of AD and vascular dementia with controls. There was a positive association with n-6 PUFAs in males. In females, low fish consumption was associated with increased risk and cases presented an absolute deficiency in n-3 PUFAs. Conquer et al (44) in a case-control study using plasma levels found the AD cases had lower total n-3 PUFAs and a higher n-6:n-3 ratio, and the total n-6 PUFA levels were higher in the cases vs. controls. In a follow-up study over 2-3 years, Kalmijn et al (198) (the Zutphen Study) found that a high intake of linoleic acid (LA) was associated with cognitive impairment, while fish consumption was inversely related to the risk of cognitive impairment. Finally, Yehuda et al (199) in an intervention study found feeding a fatty acid formulation that was 4:1 in the n-6:n-3 ratio (compared with approximately 20:1in the typical Western diet) yielded clinical benefits in AD patients. Thus the case for the benefit of n-3 PUFAs, as found in a number of studies published between 1996 and 2003, might be described as fairly strong even though some of the results were merely suggestive rather than statistically significant.

There is considerable evidence that inflammation associated compounds, including various cytokines and prostaglandins, compounds that are known to promote and sustain pro-inflammatory conditions, are present in the brain tissue of AD patients (200). Thus the inflammatory and anti-inflammatory characteristics of the n-3 and n-6 families, as discussed above, are consistent with the various studies described. However, the n-3 PUFAs may also have non anti-inflammatory actions also, including lowering the risk of thrombosis, reducing blood pressure, reducing triglycerides and improving glycemic control (194). The obvious preventive action that might reduce the risk or retard progression of AD, other dementias and cognitive decline—eat fatty fish several times a week or take EPA/DHA or fish oil supplements, or both. It is probably wise not to depend on alpha-linolenic acid (ALNA) from, for example, flax seeds, due to the low conversion rate to the long-chain n-3 PUFAs.

INFLAMMATION, DIABETES AND THE METABOLIC SYNDROME

Type-2 diabetes is characterized by progressive hyperglycemia, insulin resistance and ultimately pancreatic beta-cell failure. The hypothesis that type-2 diabetes at least in part is an inflammatory disease was first advanced in 1993. Two recent reviews (201,202) nicely summarize the current status of the evidence that this hypothesis has merit. There are four prospective studies that found the risk of developing diabetes was positively associated with plasma levels of markers for inflammation including IL-6 and CRP. One study (203,204) developed an overall inflammation score based on serum markers to which they added the total leukocyte count and the plasma fibrinogen level. When comparing the highest with the lowest quintiles based on this score, an increased risk of type-2 diabetes of 3.7 was found in white non-smokers. Thus smoking appears in this context to be protective in spite of the fact that it is in general inflammatory. Failure to correct for this may have seriously confounded some studies. It is thought that nicotine inhibits the release of inflammatory cytokines from fat tissue. Other evidence quoted in these reviews includes:

1. there is a correlation between fasting insulin concentrations and CRP;
2. human fat tissue expresses tumour necrosis factor (TNF), and the concentration of this cytokine is positively associated with the body mass index (BMI) and is elevated in obese patients. In fact, there is now considerable evidence to show that obesity is a state of chronic inflammation as indicated by increased plasma concentrations of CRP, IL-6 and other markers;
3. the pro-inflammatory effects of overeating in normal subjects are similar to those found in the obese in a fasting state;
4. insulin resistance promotes inflammation;
5. individuals with the Metabolic Syndrome have elevated levels of CRP and at high risk of developing type- 2 diabetes. Roberts and Evans (201) in fact suggest that a CRP cut-off of >3 mg/L be used to enhance prognostic power in individuals diagnosed with the Metabolic Syndrome. Incidentally, the diagnosis of the Metabolic Syndrome (any three of the following—hypertension, low HDL and high triglycerides, impaired fasting glucose, abdominal fat with a poor waist to hip measurement and obesity) is easier than diagnosing insulin resistance directly, and thus combining CRP measurements with the presence of the Metabolic Syndrome for diabetes risk assessment is attractive;
6. levels of glycosylated hemoglobin (hemoglobin A1C, a measure of long-term average blood glucose levels) above 9% are significantly associated with elevated CRP levels, suggesting that inflammation may also be related to poor glycemic control;
7. it is well known that type-2 diabetes puts one at high risk for CVD;
8. inflammatory eicosanoids and cytokines are implicated in pancreatic beta-cell failure.

The overall picture as summed up by Roberts and Evans (201) presents the view that the Metabolic Syndrome, type-2 diabetes and CVD are manifestations of a "common soil" patho-physiology with insulin resistance and an inflammatory condition as central features.

A case-control study (205) published in March 2004 supports the above conclusions. Subjects for this study were from the Nurses' Health Study. A positive association was found with diabetes risk for three inflammatory markers, IL-6, CRP and a surrogate marker for TNF. The strongest association was with CRP with an increased risk of 4.36 for diabetes when extreme quintiles were compared. The association of CRP with diabetes was comparable or stronger than the association of CRP with CHD. Thus elevated CRP may help identify high-risk populations for both type-2 diabetes and CVD. Large differences in the risk prediction by CRP were found between aspirin users and non-users. For example, for those in the highest CRP quintile, an odds ratio (similar to the risk ratio) of 9 was found for non users vs. 3 for users of aspirin. The authors suggest that the inflammatory role of CRP may be mitigated by aspirin use. The authors point out that the biological mechanisms through which CRP increases the risk of diabetes are not well understood, but CPR may have an indirect influence on insulin resistance and insulin secretion. Since the production of CRP is regulated by the inflammatory cytokines TNF and especially IL-6, the association between CRP and the risk of diabetes may also reflect the detrimental effects of these cytokines on insulin resistance. Several mechanisms have been discussed (205).

Aside from the inclusion of stratification for aspirin use in the above described study, there appear to have been limited recent human studies of the preventive potential of NSAIDs in the context of type-2 diabetes. The evidence presented by Helmersson et al (206) that COX mediated inflammation may be involved in type-2 diabetes should inspire studies that address the potential role of these inhibitors in more detail, but the recent association of the COX-2 inhibitors with adverse CV events may discourage clinical trials aimed at primary prevention. With regard to PUFAs, one recent large study addresses this question. Based on a 14 year follow- up associated with the Nurses' Health Study, Salmeron et al (207) found that intakes of saturated or mono- saturated fat were not significantly associated with the risk of diabetes, but for a 5% increase in energy from PUFAs, the relative risk (RR) of type-2 diabetes was 0.63 and for a 2% increase in energy from trans fatty acids the RR was 1.39 The RR for the highest quintile of n-3 marine PUFA intake (EPA and DHA) was 0.8 and the trend from the first quintile to the fifth was significant. The ratio of n-6 to n-3 PUFA intake was, however, not significantly associated with the risk of type-2 diabetes. This study was considered by the authors to be superior to earlier studies that found no effect of fat or specific types of fat. They argued that these earlier studies were too small and not adjusted simultaneously for other types of fat.

INFLAMMATION, A RESPONSE TO PSYCHOLOGICAL STRESS

This is an important aspect of inflammation since psychological stress is unfortunately a significant part of everyday life for many individuals. Stress induced inflammation is part of a more general subject, i.e. the mind- disease connection (208). Some may not even be aware of subtle, chronic stress, especially if it is associated with the workplace or a toxic home environment. Stress initiates a multitude of biochemical responses and causes changes in levels of cellular and circulating chemicals, some but not all of which are inflammatory. There appears little doubt that a significant result of psychological stress is a response that mimics the inflammatory response initiated by infection, injury, etc. This is the basis for the relationship between stress and various diseases which have an inflammatory component as part of their etiology (209).

Stress can also be described in terms of so-called negative emotions—depression, anxiety and the combination of hostility and anger. Related aspects include low self-esteem, low socioeconomic status, and impaired interpersonal relationships. A work environment with extreme competition, demanding deadlines, insecurity and perhaps harassment provides a good example of an emotionally toxic environment. Toxic home environments are well known to those who practice family therapy. It is sometimes useful to think of mental stress as acute, episodic or chronic. All three are thought to be dangerous. There are a number of biomarkers that are used to make the association between mental stress and inflammation, including CRP, IL-6, TNF or its surrogate markers the soluble TNF receptors, and a number of so-called stress hormones including cortisol. Of these, IL-6 has turned out to be one of the most important, and its disregulation is thought to be a key aspect of the link between mental or psychological stress and related disease states. These stress related disease states are now generally recognized (209-211) to comprise atherosclerosis, CHD, CVD, type-2 diabetes, Alzheimer's disease, and the various aspects of the Metabolic Syndrome. The evidence that inflammation is an integral part of the response to stress and has a significant impact on the initiation and/or progression of the above listed diseases has been discussed extensively in the literature and recently reviewed by Paul H. Black, one of the active researchers in this area (209). There is a very large literature base associated with the mechanisms whereby psychological factors interact with the immune/inflammatory systems and as well there are a number of epidemiologic studies that reinforce the connection. Space limitations preclude a detailed discussion. With regard to the connection between stress, inflammation and CVD, the reader is referred to a recent comprehensive review by Black and Garbutt (212).

OBESITY AND INFLAMMATION

It now appears generally accepted that obesity is associated with a state of chronic inflammation, as indicated by increased plasma concentrations of CRP, IL-6 and other markers (202). Adiposity (the so-called apple shape caused by a large accumulation of abdominal fat) is thought to induce a pro-inflammatory milieu due to the secretion of IL-6, TNF and other pro-inflammatory compounds, and this in turn results in insulin resistance, Metabolic Syndrome, impaired glucose tolerance and finally type-2 diabetes. In addition, a pro-inflammatory milieu enhances endothelial dysfunction and positively influences the progression of atherosclerosis (213). One measure used to establish the presence of the Metabolic Syndrome is in fact the presence of an apple shape. Weight loss in overweight or obese populations results in a decrease in markers such as IL-6 and CRP (214).

Direct evidence of the obesity-inflammation-CVD link is provided by a recent study by Engstrom et al (215) where five inflammation sensitive plasma proteins (ISPs) were measured in over 6000 men who were then followed for about 18 years to ascertain the risk of fatal and non-fatal heart attack and stroke. High levels of inflammation as measured by the ISPs were associated with increased risk in all categories of body mass index (BMI). The age adjusted relative risks for obese men (BMI>30) were 2.1, 2.4, 3.7 and 4.5 (!!) for those with 0, 1, 2, and 3 or more ISPs with serum levels in the top quartile. Individuals with a BMI<25 and no elevated ISPs were used as a reference, i.e. low inflammatory status and neither obese or overweight. Obesity, especially as reflected in poor waist to hip ratio (apple shape), is often accompanied by insulin resistance, a precursor to type- 2 diabetes (214). Large congregations of apple shaped individuals can usually be observed at "all-you-can-eat" buffets.

The influence of n-3 PUFAs on obesity-related insulin resistance was examined in a recent study by Browning (216). Instead of using CRP or the inflammation markers used by Engstrom et al, serum sialic acid was selected as being superior for this study. Premenopausal non-diabetic subjects with a BMI range of 24 to 44 were grouped according to inflammatory status based on this marker. The group with the higher inflammatory status was found to have higher BMI and evidence of greater insulin resistance. The effect of supplementation with 1.3 g/d and DHA 2.9 g/d was compared with a placebo. The n-3 group showed a decrease in insulin resistance, but the large and significant change as compared to the placebo group was in the subgroup with high levels of inflammation.

INFLAMMATION AND DIET

The balance between n-6 and n-3 PUFAs is both a dietary and supplement issue, and because of the low efficiency in vivo for the conversion of alpha-linolenic acid to EPA and DHA, the dietary aspect mainly involves fish consumption, and in particular oily fish such as salmon. The broader issue concerns diet in general. Are some diets pro-inflammatory, others anti-inflammatory? Some recent studies relate to this question.

• Esposito et al (217) randomized 180 patients with Metabolic Syndrome to either a Mediterranean–type diet (increased daily consumption of whole grains, fruits, vegetables, nuts, and olive oil) or a control diet (50-60% of energy from carbohydrates, 15% from proteins and <30% from fat). Serum markers of inflammation (CRP, IL- 6, 7 and 18) were significantly reduced in the intervention group, and as well, endothelial function score improved and insulin resistance decreased significantly. At 2 years of follow-up, only 40/90 patients in the intervention group still had features of the Metabolic Syndrome vs. 78/90 in the control group. In the same 2004 issue of JAMA, Knoops et al (218) reported a study of 10-year mortality associated with the Mediterranean diet. Four factors were associated with low risk, a Mediterranean diet, being physically active, moderate alcohol use and non-smoking. The combination of all four factors reduced the all cause mortality rate to 0.35, and the four factors were each associated with reduced risk, not only for CVD but also for cancer.
• Using a dietary pattern approach (see the IHN research review, The Diet Zoo) to examine the question of diet and markers for inflammation and endothelial dysfunction, Lopez-Garcia et al (219) compared a prudent diet (higher intakes of fruit, vegetables, legumes, fish poultry and whole grains) and a Western diet (higher intakes of processed and red meat, sweets, desserts, french fries, and refined grains). The prudent diet was associated with lower serum CRP levels and low endothelial dysfunction whereas the Western diet showed the reverse.
• A study (220) of the relation between a diet with a high glycemic load and serum CRP found a strong relationship with a median CRP concentration for the lowest quintile of dietary glycemic load of 1.9 mg/L vs. 3.7 mg/L (well into the danger zone) for the highest quintile, independent of conventional risk factors for ischemic heart disease. Thus diets that result in high post-meal blood glucose levels appear to be inflammatory.
• The Lyon Heart Study discussed above found a Mediterranean diet with enhanced alpha-linolenic acid yielded a very large reduction in adverse CHD events when compared to a normal European diet. This was a secondary prevention trial (177).

Other studies could be quoted, some not as strongly supportive as the above, but the picture would remain essentially unchanged. The general dietary principles to be deduced from the above are very similar to those promoted by Sears in his new book on inflammation (10), as well as those suggested by Challem in The Inflammation Syndrome (3). Thus if there is such a thing an anti-inflammation diet, then an evidence based answer would be that it appears to resemble the classical Mediterranean diet, which can also be characterized as a low glycemic load diet. However, on the basis of the frequency with which fish consumption enters into considerations of disease risk, as seen throughout the above review, the anti-inflammatory diet should contain a liberal amount of this natural source of EPA and DHA. Anyone wanting to play it safe, however, would probably want to take supplemental EPA + DHA or fish oil, with the amount adjusted to account for fish consumption. Dislike of fish or fear of mercury, dioxins and PCBs cause some to elect taking ultra-refined EPA/DHA concentrates or a fish oil that is low in contaminants (see www.ifosprogram.com for detailed information on the contamination levels in some fish oil products). According to Barry Sears (10), high levels of contamination are not uncommon in fish oil so it is buyer beware. Also, there appears to be no regulation regarding the use of the terms 'pharmaceutical grade" or "toxin free."

Which fish are safe is a matter of what standard one wishes to apply. The FDA's so-called Action Level is 1.0 part per million (ppm) of mercury but 30 years ago they were enforcing 0.5 ppm and seizing millions of cans of tuna. The FDA website lists mercury levels in various commercial fish and shellfish (http://vm.cfsan.fda.gov/~frf/sea-mehg.html).

CONCLUSIONS

One conclusion that could be drawn from the many studies quoted above is that in developed countries where both agriculture and the food industry are highly industrialized, diets highly "Westernized," and where food and beverage choices are profoundly influenced by advertising, large segments of the population appear to suffer from a chronic deficiency of the n-3 PUFAs and in particular EPA and DHA. This is in part, and perhaps in large part, responsible for, or at least related to, chronic but silent inflammation. This then may play a critical role in many disease states that appear to characterize our present level of "civilization." One is reminded of the often- quoted observation that at the beginning of the last century coronary heart disease was almost unknown.

Chronic inflammation is of course not simple. It can be caused or aggravated by psychological stress, low intake of n-3 PUFAs, chronic infection, autoimmune disease, toxins or irritants in food, air and water, trans-fats, smoking, obesity and overeating, hyperglycemia, etc. The evidence concerning the connection between chronic inflammation and a variety of diseases including the major degenerative diseases is of growing significance. If Barry Sears and others are correct about their interpretation of the serum AA/EPA ratio, then silent, chronic inflammation is present in the North American population to an alarming but almost totally unrecognized extent. This is consistent with the incidence of the major killer diseases such as heart disease, diabetes and cancer, which has dramatically increased over the last century while at the same time the typical diet has become, so it would appear, much more pro-inflammatory. More research seems urgently needed regarding the implications of the AA/EPA ratio which may indeed turn out to be a key indicator of overall health status and future health prospects. It is encouraging that this blood test has at least gained the status of easy accessibility and can be ordered by any physician.

The connection between obesity and inflammation on the one hand, and inflammation and type-2 diabetes on the other, carries the strong message that chronic inflammation merits serious consideration as one of the driving forces of the current diabetes epidemic, especially since the age of onset is decreasing rapidly and "adult onset diabetes" must now be modified to include "teenage onset." The suggestion that type-2 diabetes will in the next few decades bring health care to the brink of bankruptcy and push insurance rates to unaffordable levels appears to be more than just scare mongering. While what might be called the inflammation hypothesis is still quite young and underdeveloped, and much research needs to be done, the existing evidence should cause it to be taken very seriously indeed. The reduction in chronic inflammation could and perhaps should be considered a major goal of preventive medicine and public health.

It is probably overly optimistic to think that progress will be easy. Obesity has proved to be difficult to overcome in practice with diet and exercise, and smoking is actually on the increase in certain segments of the population. All-you-can-eat buffets are popular. The Western style diet, which appears to be pro-inflammatory, is well entrenched. A dislike for fish is quite common. Mental stress permeates many workplaces and as well many home environments and is exacerbated by poverty, low social status and low self-esteem. Knowledge among the general public of the essential aspects of the Mediterranean diet or any other diet thought to be anti- inflammatory is probably not common, nor is what constitutes an adequate intake of n-3 PUFAs and how to get it common knowledge. Most physicians probably are unaware that assessment of the AA/EPA ratio and the EPA + DHA sum is now readily available from commercial labs, nor is the potential information from these two blood markers widely appreciated or even known among general practitioners who could easily order the Omega-3 Essential Fatty Acid Profile along with the standard blood lipid profile. Most people have probably never heard of advanced glycation end (AGE) products, but a possible way to reduce inflammatory levels is through the avoidance of high-AGE foods.

Natural anti-inflammation strategies are limited but potentially significant. Aside from diet, the centerpiece of course consists of EPA and DHA, either in the form of refined fatty acids or from fish or fish oil. Sears takes the position that 3 g/d of fish oil is appropriate for "healthy" individuals. William R. Davis, the cardiologist mentioned above, recommends a minimum of 1.5 g/d of EPA and DHA combined (about 5 g/d of fish oil) for individuals without heart disease, which is similar to Sears' recommendation. Both suggest larger daily intake if serious disease is present. Sears recommends increasing fish oil or EPA + DHA consumption if the AA/EPA ratio is greater than 3. The cardiologist Dr. Stephen Sinatra comments (www.drsinatra.com) "I'm so impressed with the research on fish oil supplements that I include it in my core heart-health program." It is worth repeating that alpha-linolenic acid (ALNA), for example from flax seed or flax seed oil, is not a very good source of EPA and DHA since the efficiency of conversion can be very low. They are however frequently touted as great omega-3 foods. There is some evidence, as mentioned above, that large amounts of ALNA might be harmful for men. While the conventional wisdom is for lowering the intake of the n-6 PUFAs such as LA and AA, as mentioned above there is growing evidence to indicate that either LA or the long-chain fatty acid AA to which it is converted are not as dangerous as originally thought, and that the best action to correct a poor AA/EPA or high dietary n-6:n-3 ratio may be to increase the levels of EPA and DHA. It now appears that the main undesirable aspect of either ratio being high may be due to low n-3 intake rather than high n-6 intake.

In the presence of pain, the natural reaction is to turn to over-the-counter or prescription anti-inflammatory drugs, and there are a number of non-specific NSAIDs from which to choose and as well physicians still have several options for specific COX-2 inhibitors. But there is potentially a high price to pay in upper gastrointestinal (GI) damage, ulcers, perforation, bleeding etc. While not common, such side effects are also not that rare. Aspirin doubles the risk of GI bleeding even at doses as low as 75 mg/d, and for anyone taking aspirin who has a history of bleeding from ulcers, recurrent bleeding will occur in 15% of cases within a year (221). While Celebrex is promoted as being more stomach friendly than non-specific NSAIDs, the research backing this up has been called into serious question (153,154). No one appears to have questioned the gastric advantages of Vioxx, but this drug has been pulled from the shelves, and the whole COX-2 class of drugs is now under suspicion.

In connection with the use of substitutes for the non-specific NSAIDs (not including aspirin), a recent large case- control study (222) found that sudden cessation of use of NSAIDs put patients at enhanced risk of a heart attack, with those having RA and lupus at particular risk, almost 4 times compared to controls. Patients who had used NSAIDs for a long period were also at increased risk. The risk disappeared several weeks after the discontinuation of the drug. The authors suggest an inflammatory rebound effect, but the mechanism is in fact unknown. Individuals switching from NSAIDs to natural anti-inflammatory substances such as the long-chain n-3 PUFAs, turmeric or other supplements should be aware of this potential risk and discuss with their physician a program for termination. Finally, Barry Sears, in his new book (10), makes a strong argument for using fish oil or purified EPA + DHA, on occasion in high doses, to treat pain, and he presents case histories to back up his claim of effectiveness. A comprehensive discussion of alternatives to NSAIDs can be found on Dr. Stephen Sinatra's website.

If an action plan can be derived from the information in this review, it would involve a Mediterranean type diet typical of Crete in the 60s with lots of fish and a glass or two of wine a day, supplementation with EPA and DHA or fish oil to achieve an AA/EPA ratio below 3 and an EPA + DHA sum of > 4% of total serum phospholipids, and the avoidance of stress, environmental toxins and irritants. Persistent CRP levels above 2 to 2.5 mg/L would merit investigation. Obesity or being overweight would be regarded as a highly dangerous state of affairs urgently requiring attention. An action plan similar to this is outlined in great detail in Barry Sears' new book, The Anti-Inflammation Zone (10).

Some would say that this is reading too much into the fatty acid studies where there are inconsistencies and contradictions. Perspective is perhaps gained from the following comment by Walter Willett and Meir Stampfer of Harvard in connection with randomized prevention trials (223): "In general, clearly positive results would be compelling, but negative results would be difficult to interpret." Considering the intense clinical and epidemiologic research interest in EPA and DHA and what is already known, the problem of inconsistent studies may be eventually sorted out, and these two fatty acids could in fact become the heroes of 21st century nutritional supplementation. Only time will tell.

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