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Would you like a side of metabolic endotoxemia with that order?

“Chronic low-grade inflammation is now considered to be a critical pathological factor underlying many modern chronic diseases, including diabetes, cardiovascular disease, cancer, and neurodegenerative diseases, and is associated with aging. Chronic low-grade inflammation is characterized by elevated circulating levels of inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α), interleukin (IL)-1, and IL-6. A primary cause of chronic low-grade inflammation is metabolic endotoxemia, which presents as a gradual increase in plasma endotoxins, particularly lipopolysaccharides (LPS). The binding of LPS with Toll-like receptor-4 (TLR4) and subsequent activation of NLRP3 inflammasome leads to the increased expression of inflammatory cytokines. Understanding the factors that modulate the development of metabolic endotoxemia is important for the management of chronic disease. Metabolic endotoxemia can often result from gut dysbiosis, such as an increase in LPS-producing bacteria (e.g. Enterobactericeae) and/or a decrease in LPS-suppressing bacteria (those which can lower the numbers of LPS-producing bacteria, such as Bifidobacterium), as well as intestinal barrier dysfunction. Diet is known to be an important modulating factor of metabolic endotoxemia and chronic low-grade inflammation; for example, a high-saturated fat diet can increase LPS levels and induce endotoxemia. Clearly, any factor that can alter the gut microbiota could play a role in regulating metabolic endotoxemia and chronic low-grade inflammation. Identifying the dietary components that can optimize the gut microbiota will be crucial for the prevention and treatment of chronic disease.

It is well recognized that long-chain omega-6 (n-6) and omega-3 (n-3) polyunsaturated fatty acids (PUFA) play important and opposing roles in the modulation of inflammation. Generally, n-6 PUFA promote inflammation, whereas n-3 PUFA have anti-inflammatory properties. Since n-6 and n-3 long-chain PUFA compete for the same enzymes for their synthesis and metabolism, their ratio in body tissues determines the profile of lipid mediators involved in the inflammatory response. Over the past decades, the n-6/n-3 PUFA ratio has undergone a dramatic shift from the human evolutionary ratio of ~1:1 to the modern dietary ratio ranging from 10:1 to 50:1, due to the increased intake of foods rich in n-6 PUFA and deficient in n-3 PUFA. This shift in the n-6/n-3 PUFA ratio is thought to contribute to today’s prevalence of chronic disease.” – A host-microbiome interaction mediates the opposing effects of omega-6 and omega-3 fatty acids on metabolic endotoxemia (1)

This is a very good summation of the role endotoxemia plays in chronic disease, including those diseases classified under the rubric of metabolic syndrome. My only quibble is with the implication that because saturated fats increase lipopolysaccharide (LPS) translocation via chylomicron formation when absorbed, they are somehow uniquely deleterious to our health, as if evolution forgot to provide us with a biological mechanism (lipoproteins) to neutralize this effect when consuming a type of fat that we’ve been consistently eating for approximately 200,000 years. I’ve explained why this hypothesis has no basis in fact here and here, and will leave it at that for now.

What goes conveniently unmentioned in this opening statement, however, is that this imbalance between omega-6 and omega-3 intake is a direct result of the public, especially in the English speaking world, being repeatedly told by those in authority that saturated fats are a dietary villain best avoided, and that “heart healthy” omega-6 polyunsaturated fats (PUFAs) should be eaten in their place.

Is it any wonder then that diseases like type 2 diabetes and non-alcoholic fatty liver disease have exploded in the interim? Yet last time I checked, not one of these authorities has suffered any career or legal ramifications for promoting the consumption of these gut destroying, pro-inflammatory and toxic fats.

The focus of the study whose introductory paragraphs I just quoted is precisely about how omega-6 fats negatively impact gut flora, and by extension, markers of metabolic endotoxemia in lab mice fed these fatty acids in amounts representative of typical human intake.

This study used two sets of mice: your run-of-the-mill C57BL/6 male WT lab mice and a relatively new rodent by the name of fat-1 transgenic mice. And what, pray tell, are fat-1 transgenic mice? Well, they’re genetically modified rodents (created by a modern day Dr. Frankenmouse, or was it Dr. Frankencat?) that can endogenously convert omega-6 PUFAs from their diet to omega-3 PUFAs. (2) This is obviously something that mammals, including ourselves, are incapable of doing naturally.

So in the graphs I’m about to show you, keep in mind that those bars labeled WT are representative of the conventional mice fed large quantities of omega-6 PUFAs, in this particular case corn oil. Cont stands for C57BL/6 male mice fed a standard control diet and Fat-1 represents the transgenic mice I just mentioned:The top three graphs show markers for metabolic endotoxemia. In the conventional mice (WT) fed corn oil chow, serum lipopolysaccharide (LPS), lipopolysaccharide binding protein (LBP) and sCD14, a marker for immune activation, were all elevated. Conversely, the fat-1 mice who are able to metabolically convert omega 6 to omega 3 showed markers that were slightly worse, the same or somewhat better than controls.

In the second row of graphs, we see the effect of omega-6 feeding on cytokine production. To refresh your memory, cytokines are signaling proteins that coordinate immune responses.

Three pro-inflammatory cytokines—tumor necrosis factor-alpha (TNF-a), interleukin 1 beta (IL-1ß) and interleukin 6 (interleukin 6)—were significantly elevated in the conventional mice fed corn oil in contrast to both control and fat-1 mice.

Interleukin 10, an anti-inflammatory cytokine, was elevated in both the conventional and transgenic mice. Interleukin 10 is always elevated whenever inflammation is present, and serves as somewhat of a brake on inflammatory processes much like cortisol. The very high rate noted in the fat-1 mice speaks to the immune suppressing actions of omega-3 fatty acids. I’ll have more to say about this towards the end of the post.

Row three shows body weight gain, fasting blood glucose, fasting insulin levels and homeostatic model assessment (HOMA-IR), which is a test to quantify insulin resistance and pancreatic beta cell function. Not surprisingly, the corn oil fed mice fared far worse than the other two groups.

A second leg of this research experiment involved taking half the conventional mice and half of the fat-1 mice and nuking their gut flora with broad spectrum antibiotics, after which they were all housed together and fed the high corn oil chow. Interestingly enough, the conventional mice showed similarly low levels of metabolic endotoxemia as their fat-1 littermates. And the reason for this was because they were continuously “dining” on the gut contents of the transgenic fat-1 roommates by—oh dear, how do I put this delicately—eating their poop (i.e. engaging in coprophagia), which is what mice do when housed together. Lovely.

This behavior, however, is not at all recommended for my dear readers. I must admit, however, to dispensing such advice to certain people in my life when they’ve crossed me. My sincerest heartfelt apologies to anyone so offended.

Returning to the topic at hand, these researchers then proceeded to study the gut flora composition that was protecting the fat-1 mice from the god-awful “heart healthy” corn oil. They discovered that:

“Using both bacterial culture and quantitative PCR to identify and quantify the bacteria in stool, we found that the fat-1 mice exhibited significantly lowered or undetectable quantities of LPS-producing and/or pro-inflammatory bacterial groups, including the phylum Proteobacteria and its members (Enterobacteriaceae, Escherichia coli, gamma- and delta-proteobacteria), Prevotella, Fusobacterium, Clostridium cluster XI, and Segmented Filamentous Bacteria (SFB), compared to their WT littermates…In contrast, the levels of LPS-suppressing and/or anti-inflammatory bacterial groups, such as Bifidobacterium, Akkermansia muciniphila, Lactobacillus (primarily L. gasseri), Clostridium clusters IV and XIVa, and Enterococcus faecium, were markedly higher in the fat-1 mice compared to their WT littermates…Notably, the bacterial overgrowth in the WT group was mainly due to the increased proportion of LPS-producing members of the phylum Proteobacteria…Intestinal permeability was also lower in the fat-1 mice than in the WT mice…As anticipated, the overall stool microbiota profile of the co-housed WT mice was similar to those of the fat-1 groups, particularly the abundance of stool bacteria favoring or suppressing metabolic endotoxemia…These findings indicate that the tissue n-6/n-3 PUFA ratio is a determinant of the gut microbiota profile: a high tissue n-6/n-3 PUFA ratio can increase the proportions of LPS-producing and/or pro-inflammatory bacteria and decrease those of LPS-suppressing and/or anti-inflammatory bacteria, while reducing the tissue n-6/n-3 PUFA ratio has the opposite effect.”

So it must have been the beneficial bacteria ingested when eating the stool of fat-1 mice (eww) that was protective of the conventional mice when they were housed together, right?

Well no, that wasn’t the case. What they found instead was that it was the intake of intestinal alkaline phosphatase (IAP), found in the feces of fat-1 mice (double eww), that was the decisive factor in keeping our ordinary furry blokes free from an overgrowth of gram-negative gut pathogens when consuming a high omega-6 chow.

To prove their point, an IAP inhibitor by the name of L-phenylalanine was fed to the fat-1 transgenic mice along with the same omega-6 rich PUFA diet for eight weeks. This caused the fat-1 mice to exhibit a significant increase in LPS-producing bacteria and endotoxemia, just like their conventional counterparts when eating the same diet.

Finally, these researchers wanted to discover if supplementing with omega-3 fish oil would correct dysbiosis in conventional mice fed omega-6 corn oil since weaning. And indeed, the mice taking fish oil showed decreased levels of LPS producing pathogens and increased levels of beneficial bacteria.

So let’s sum up what this study found:

  1. Omega-6 vegetable oils (in this case corn oil) increased pathogenic bacteria in the guts of conventional C57BL/6 male mice and induced metabolic endotoxemia.
  2. Housing these dysbiotic mice with genetically modified fat-1 mice normalized their gut flora because they ate the feces of fat-1 mice.
  3. This normalization was not due to the bacteria contained in the fat-1 stool, but rather to the presence of intestinal alkaline phosphatase (IAP).
  4. Confirmation of this was arrived at by inhibiting IAP in fat-1 mice by the administration of L-phenylalanine resulting in the same level of gut dysbiosis and metabolic endotoxemia experienced by the non-GMO rodents eating large amounts of omega-6 chow.

As I wrote in my post on intestinal alkaline phosphatase:

“IAP has some very important functions. First, it’s involved in regulating secretion of bicarbonate in the duodenum.

Bicarbonate helps to neutralize the very acidic semi-digested food (chyme) entering the small intestine after passing through the stomach. Failure to raise pH here can lead to acidified chyme injuring cells lining this part of the digestive tract. That can increase inflammation and intestinal permeability.

But IAP’s most important role is detoxifying lipopolysaccharides (LPSs) derived from the cell wall components of gram-negative gut bacteria. It is therefore an important defense against endotoxemia.”

This study once again emphasizes how important IAP is to maintaining gut health. But there is a major concern I have with the conclusion arrived at by these researchers:

“From a translational standpoint, we utilized n-3 PUFA supplementation in this study to confirm that the elevated tissue n-3 PUFA status and the beneficial effects observed in the fat-1 mice could also be achieved through dietary intervention. Our discovery that elevating tissue n-3 PUFA status and lowering the n-6/n-3 PUFA ratio can improve the gut microbiota profile and suppress chronic low-grade inflammation provides two major implications for today’s health problems. On one hand [sic], this study highlights the excess of n-6 PUFA and deficiency of n-3 PUFA in the Western diet as a cause of modern health epidemics. On the other hand, this study points to a new strategy for the prevention and treatment of chronic diseases by reducing the tissue n-6/n-3 PUFA ratio through n-3 PUFA supplementation and reducing n-6 PUFA intake. Furthermore, the mediating role of IAP in the pathway explored in this study supports the use of exogenous IAP as a potential therapy for inflammatory diseases. Finally, given that gut dysbiosis and metabolic endotoxemia are often linked to many clinical problems, the capability of n-3 PUFA to reverse these conditions indicates the potential of n-3 PUFA supplementation as a therapeutic means for treating such cases, and its efficacy could be evaluated through the measurement of LPS and related factors as biomarkers.”

Hence the recommendation given by many health practitioners, both conventional and alternative, to use omega-3 supplements as part of a standard treatment protocol for correcting gut dysbiosis. However, there is no proof offered in this paper that increasing omega-3 intake in conventional mice or humans increases expression of IAP in the gut. On the contrary, the evidence points in the opposite direction. As I wrote in my post on IAP:

“Unfortunately for the public, omega-3 PUFAs also inhibit IAP expression. As I wrote in my post Ulcerative Colitis and Dietary PUFAs, omega-3s in conjunction with omega-6s have been shown to lower IAP expression in mice even when adjusted to levels that are typical for humans who take omega-3 supplements”

So the fact that genetically modified mice high in omega-3 tissue concentration from endogenous production of this fatty acid can produce IAP in large amounts is no reason to expect the same from exogenous omega-3 sources.

But there is another reason to be leery of this recommendation. Dousing your intestines with copious quantities of highly reactive and immune suppressing omega-3s carries very real risks.

This study didn’t track longevity rates or death from infection in mice fed omega-3s. Other studies have, however.

For example, one study that assessed caloric restriction on life span found that while all mice placed on calorie-restricted diets experienced increases in longevity, mice fed a fish oil diet saw the least benefit, while those fed a lard diet rich in monounsaturated and saturated fatty acids lived the longest. (3)

Courtesy: The Influence of Dietary Fat Source on Life Span in Calorie Restricted Mice

Here we see a chart plotting the length of days extended by a 40% calorie-restricted diet (CR) against controls fed a 5% restricted diet. Fish oil fed CR mice are representd by the upside down purple triangles, soybean omega-6 fed mice by the green triangles and lard fed mice by the yellow squares.

As expected, all three calorie-restricted groups extended their time on our dear Earth. However, of the three groups, the omega-3 fed mice had the shortest extension of the bunch. As these researchers put it:

“The results of the present study indicate that dietary lipid composition influences life span in CR mice. In particular, life span was increased in CR mice consuming a diet with a high proportion of monounsaturated and saturated fatty acids compared to CR mice consuming diets with high proportions of n-3 or n-6 PUFAs.”

There were a number of reasons given for why this might be the case,  but one in particular grabbed my attention:

“Thus, differences in life span between the CR lard mice and the other CR groups were likely due to delay in onset of disease rather than preventing the occurrence of specific disease conditions.”

But why would this be the case? Well permit me to reprint part of a study abstract that I’ve already blogged about concerning supplementing with omega-3 PUFAs to counter a high omega-6 diet:

“Addition of ω-3 PUFA on a high ω-6 PUFA diet, reversed inflammatory-inducing microbial blooms and enriched beneficial microbes like Lactobacillus and Bifidobacteria, reduced immune cell infiltration and impaired cytokine/chemokine induction during infection. (4)

I’m sure you’re thinking “Well isn’t that great news Ray? I mean this jives with the results of the study you’re writing about, so what’s the big fricking deal?”

Well my little non-GMO human critters, this is the big deal:

“While, ω-3 PUFA supplementation protected against severe colitis, these mice suffered greater mortality associated with sepsis-related serum factors such as LPS binding protein, IL-15 and TNF-α. These mice also demonstrated decreased expression of intestinal alkaline phosphatase and an inability to dephosphorylate LPS. Thus, the colonic microbiota is altered differentially through varying PUFA composition, conferring altered susceptibility to colitis. Overall, ω-6 PUFA enriches pro-inflammatory microbes and augments colitis; but prevents infection-induced systemic inflammation. In contrast, ω-3 PUFA supplementation reverses the effects of the ω-6 PUFA diet but impairs infection-induced responses resulting in sepsis. We conclude that as an anti-inflammatory agent, ω-3 PUFA supplementation during infection may prove detrimental when host inflammatory responses are critical for survival.”

Remember how the fat-1 mice over-expressed the anti-inflammatory cytokine interleukin 10? Well, since the mammalian immune system utilizes inflammation to destroy invading bacterial, viral and fungal pathogens, too much of a good thing can suppress the ability to fight off an infection.

But bacterial, fungal or viral fighting immune cells aren’t the only cells suppressed via omega-3 intake. Let me quote at length what I wrote many moons ago about the effect these fatty acids have on the immune system:

“A major concern in using omega 3s to treat gut dysbiosis is the increased risk of unbalancing the immune system away from a Th1 response and towards Th2 dominant activation. One of the common outcomes of doing so is to increase allergic reactions to food, pollen and other environmental inhalants. [Could this be the reason so many complain of histamine reactions to food?] Many of these allergic immune reactions also underlie various skin diseases like eczema…As mentioned, omega 3s also suppress Th17 cells and their cytokine, interleukin 17 (IL-17). In my post on Crohn’s disease, I wrote that IL-17 is usually elevated in this disorder, likely in response to a yeast overgrowth in the intestinal tract.

While omega 3s do decrease the inflammatory response to this type of infection, mainly by inducing death in the immune cells that come to attack and destroy the overgrowth, they would also make it that much harder to eradicate it. If fungal infection is really at the heart of Crohn’s disease, then a strategy of supplementing with omega 3 PUFAs may prove counterproductive. This would also hold true for those trying to overcome a Candida albicans overgrowth.

There is another subset of immune cells that is consistently suppressed by these fatty acids: natural killer cells (NK). NK cells are unique in that like helper T cells they can generate lots of cytokines to destroy infected cells.

However, they also have another very important function. They are surveillance cells responsible for the early detection and destruction of tumor cells.

The ability of omega 3s to depress natural killer cell activity is a cause for concern. A study done in humans found that a moderate amount of eicosapentaenoic acid (EPA), an omega 3 found in fish oil, decreased natural killer cell activity in healthy adults over the age of 55. Whether this translates to greater cancer risk is yet to be determined.”

Kind of a damned-if-you-do, damned-if-you-don’t dilemma don’t you think? Yes, supplementing with omega-3s will improve markers of dysbiosis by reducing inflammation in the gut and altering the intestinal environment to favor beneficial gut flora. However, it does so at the risk of immune suppression and increased risk of systemic infection.

So ask yourself this: would you rather have gut dysbiosis and suffer from chronic metabolic endotoxemia, or have no dysbiosis but increase the possibility of contracting, and possibly dying from, an infection?

So are omega-3s beneficial in the treatment of gut dysbiosis?

It depends on how you ingest them and the quantities you take in. By all means, get your omega-3s from whole foods like seafood and flaxseed. Seafood, in particular, comes bundled with other nutrients like selenium and vitamins, which as antioxidants help neutralize the highly reactive chemical nature of unsaturated omega-3s while benefiting from their anti-inflammatory properties.

But if you can’t stomach seafood or flaxseeds and would rather get your omega-3s from a bottle or pill, please be cognizant of their powerful immune suppressing effects. A little goes a long way, and more is definitely not better. Remember, our physiological need for both omega-3 and omega-6 is low.

The key to correcting a screwed up omega-6/omega-3 imbalance is to reduce your intake of foods containing omega-6 fats, not by swallowing bucketloads of immune suppressing omega-3s in the blind hope of compensating for a bad diet. That means avoiding the vast majority of fried foods (overwhelmingly cooked in oxidized vegetable oils) like doughnuts, French fries, onion rings, fried chicken, battered and fried anything, etc., along with many processed foods containing omega-6 PUFAs like mayonnaise made with soybean oil, many salad dressings, potato chips cooked in vegetable oil and other processed packaged foods (read those ingredient labels folks).

Omega-6s are ubiquitous in the food supply because they are cheap to produce and help keep the cost of food low, while simultaneously preserving the profit margins of the food and restaurant industry. This isn’t going to change until consumers educate themselves about the gut destroying properties of these fats and demand more healthful alternatives like monounsaturated fats, and yes, unjustly vilified saturated fat.

 

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