Thursday, April 25, 2013

The interplay among the human intestinal microbial landscape, obesity, metabolism, and nutritional status: an overview


INTRODUCTION 

I’ve been paying much more attention to the human intestinal microbial landscape because of the red meat-carnitine study, and more recently, this one.  There is a growing body of compelling evidence linking the types of bacteria that colonize our intestines, and therefore the types of toxins we are exposed to, and our risk of diseases that include obesity. 

I’ve written about this before, namely with respect to the gram-negative bacterial toxin, lipopolysaccharide (LPS), as it relates to physical attractiveness, as well as the apparent beneficial metabolic effects of sterilizing the intestines of all microbial life.

I don’t want to dwell on this matter, but I do want to try to delve further into the intricacies of the topic and the evidence in which many of the suppositions are based, and towards the end, shed light on some pathways that open up possibilities for intervention.


ENDOTOXIN: A BACTERIAL TOXIN

Certain intestinal bacteria (gram-negative) have an outer cell wall component called LPS.  Unlike other bacterial toxins, LPS is not secreted, but is instead released by bacteria upon their destruction or death.

LPS is predominantly taken into the body from the intestines, via newly made chylomicrons, subsequently reaching the bloodstream, whereupon coming into contact with pathogen-recognizing receptor complexes on the surface of immune cells, activate inflammatory pathways. (Obese people have higher circulating levels of LPS than lean people do.)

Intestinal bacteria produce LPS continuously throughout the course of a day, and there is a diurnal variation in blood LPS levels, with peaks following meals, especially high-fat meals.  Chronically feeding rodents high-fat diets leads to changes in their intestinal flora, increasing the proportion of gram-negative bacteria to gram-positive bacteria, and this is accompanied by a modest rise in blood LPS levels and low-grade inflammation.

When present in very high amounts (as seen in septic shock, perforated ulcers, or peritonitis), LPS creates widespread blood clotting, stimulates the secretion of proinflammatory cytokines by macrophages, and activates the complement system to generate chemoattractants and adhesion molecules.  In short, LPS initiates an immune response, and large doses of LPS can result in life-threatening illnesses.

Many of the effects elicited by LPS are executed through its interaction with a receptor complex found, for all intents and purposes, on the surface of immune cells, namely monocytes and macrophages.  Upon this interaction, the monocytes and macrophages begin to upregulate their expression of NF-κB, which, in turn, activates a host of related genes that promote inflammation and aging, notably skin aging.1

NF-κB, for instance, induces the gene expression of COX-2, and, via TNF-α, the inducible nitric oxide synthase (iNOS), which generates a reactive gas called nitric oxide (NO).  In the pancreas, NO suppresses insulin secretion; in the muscles, NO reduces insulin sensitivity; and in the fat cells, NO, by way of phosphorylation, activates HSL.  As a result, free fatty acids (FFAs) are released into the bloodstream in droves.2

TNF-α and FFAs act on the same receptor complex that LPS does, so we can immediately conceive of a vicious cycle, whereby the inflammatory state initiated by LPS becomes amplified.  Blocking the receptors for LPS, deleting the gene for the LPS receptor,3 or eradicating intestinal bacteria all together with antibiotics, protect against diet-induced anything,4 as we saw again with red meat and eggs, and, most interestingly (to me at least), allow for animals to eat more food without getting fatter.5


FREE FATTY ACIDS AND ADIPOSE TISSUE

A vicious cycle is initiated upon the exposure to LPS that leads to a persistent elevation in blood FFAs. (This is a topic for a future post, so I will keep this discussion as brief as possible.) This probably occurs through the development of inflammation in the adipose tissue, with the activation of adipose tissue-resident macrophages and thereafter, the recruitment of additional macrophages via the enhanced expression of the chemoattractant protein called MCP-1.  Inflammation, through mechanisms that are less clear, begets FFA mobilization, and elevated levels of FFAs, in turn, beget inflammation in the adipose tissue, and so on.

Inflammation and oxidative stress are initiated in the adipose tissue by LPS, as well as by FFAs, and we at least know that these factors are not acting in isolation, but rather in a feed-forward mechanism of sorts.

Different fatty acids also have been shown to have differential effects in the body upon mobilization.6 Saturated fats, as a rule, are the greatest promoters of inflammation, which should be expected as they bear the closest resemblance to the LPS molecule itself. (The lipid portion of LPS is composed of medium-chain saturated fatty acids, like those found in abundance in coconut oil.) 

An analogous process occurs in the adipose tissue in obesity, in which large fat cells become inflamed via the infiltration of adipose tissue by macrophages, and as a result of adipocytes themselves producing inflammatory proteins that in turn, also leads to the development of insulin resistance in the body.  Like in the case of LPS, this is probably executed through NO, as deletion of iNOS prevented the impairment of insulin signaling by a high-fat diet.7


EICOSANOID PATHWAYS, ASPIRIN, AND OTHER NSAID

Arachidonic acid (AA), upon liberation by phospholipases, is metabolized by cyclooxygenases (COX).  

Without a doubt, the COX pathway is crucial for generating an immune response to LPS, but because linoleic acid (LA) is present in such higher amounts compared to AA, and because LA is not a substrate for the COX, the products of the lipoxygenases (LOX), which can act on both LA and AA, are probably of greater physiological relevance, at least in mammals.  No one, I think, would dispute the statement that the LOX pathway, namely the 5-LOX pathway, is highly involved in the LPS-induced metabolic derangements.8

Aspirin is an interesting drug, and I’ve been thinking about it more of late, and this recent blog post spurred the writing of the one you’re now reading.  Aspirin irreversibly inhibits COX-1 and modifies (via acetylation) COX-2. (Is there a COX-3?9)

COX-1 is said to be “constitutively active,” meaning it is active, to varying extents depending on the tissue, all the time, and is involved in general “housekeeping” functions, like maintaining he stomach’s mucus lining.  COX-1 also generates thromboxane A2, a prostanoid that promotes platelet activation and aggregation, thereby regulating blood clotting.

COX-2, on the other hand, is normally undetectable in most cells, being induced in response to inflammatory stimuli.  Bear in mind that both COX-1 and COX-2 carry out the same basic reaction.  The main difference lies in their tissue distribution.

Selective COX-2 inhibitors have not fared so well in clinical trials, as they’ve been associated with an increased risk of heart attacks and strokes, largely attributed to their ability to inhibit the vascular production of PGI2 that synergizes with 13-HODE—derived from LA—to inhibit the aggregation of platelets and to relax the blood vessels.

However, opinions are divided as to whether this applies to low-doses of aspirin as well.  Ordinarily speaking, such deductions would not be unreasonable, if only we were uninformed of aspirin’s 15-LOX stimulating effect.10

15-LOX not only acts on AA, but it also acts on LA, generating 15-HETE and 13-HODE, respectively.  15-HETE inhibits the production of superoxide radicals, suppresses white blood cell (PMNL) migration across cytokine-activated endothelia, relaxes vascular smooth muscle cells,11,12 and can be further metabolized to the anti-inflammatory lipoxins (and the inflammatory leukotriene B4, which the presence of mead acid [ETA] inhibits the generation of).  15-LOX appears to preferentially oxidize LA to 13-HODE over AA to 15-HETE, irrespective of the levels of AA and LA present in tissues.

There are a couple of observations that point to the possible therapeutic potential of low-dose aspirin therapy.

1.      In colorectal cancers, the expression of 15-LOX is downregulated.
2.      In advanced stages of atherosclerosis, the expression of 15-LOX is upregulated.

The upregulation of 15-LOX is critical for the induction of apoptosis by NSAIDs.  This would, I suppose, play a major role in aspirin’s purported chemoprotective effects, as would the slow conversion of 13-HODE to 13-KODE (ketooctadecadienoic acid), which has been shown to have tumor-suppressant properties. (To be clear, I’m merely speculating here.)

Recall from my lipid peroxidation series (parts 1, 2, and 3) that there is a dramatic accumulation of HODEs that accompanies atherosclerosis.  However, I’m open to the possibility that this increase could represent a protective response that serves to limit the progression of atherosclerosis. 

But at the same time, I’m not ruling out the possibility that these LA, and to a much lesser extent AA, derived 15-LOX products are merely correcting the damages initiated by other lipid peroxidation processes, especially those that go on nonenzymatically.  Another LA derived HODE, for instance, synergizes with 2,4-decadienal (a PUFA oxidation product derived from both LA and AA) to stimulate the release of IL-1β by macrophages.  IL-1β, in turn, promotes white blood cell adhesion to the endothelia, inflammation, and vascular smooth muscle cell proliferation (all bad).

Aspirin is useful in that it inhibits COX, stimulates 15-LOX, and inactivates NF-κB, providing multilayers of protection with minimal risk of side effects, and because of this I think it could serve to curtail the damages caused by both obesity and LPS when used in low doses.


CONCLUSION

The processes outlined above I think illustrate clearly the interplay, and overlap, among the immune system, metabolism, and our nutritional status.  Now that I have a chance to read other blogs, I see this motif about “gut bacteria” recurring, and how all our problems would be solved if we would just put more focus there.  And according to the general consensus, the liberal use of probiotics is precisely what everyone ought to be doing. (Of course, fermented foods are superior because they are more “natural.”)

I do think the animal studies are interesting, and fun to read, but I have my doubts as to how effective “probiotics” really are, and whether simply adopting a normal diet and losing excess body fat, if you have it, would not correct the metabolic derangements that are affecting people like an epidemic. 

In the meantime, low doses of aspirin, eating smaller and balanced meals, spread out equally throughout the day, and if you suspect dysbiosis, getting tested for it and treated properly by a medical doctor I think affords adequate protection against the things discussed herein.  A stimulant laxative I think could help to reduce the intestinal bacterial load, and therefore the exposure to bacterial toxins like LPS.


REFERENCES

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3.       Davis, J. E., Gabler, N. K., Walker-Daniels, J. & Spurlock, M. E. Tlr-4 deficiency selectively protects against obesity induced by diets high in saturated fat. Obesity (Silver Spring, Md.) 16, 1248–55 (2008).
4.       Suganami, T. et al. Attenuation of obesity-induced adipose tissue inflammation in C3H/HeJ mice carrying a Toll-like receptor 4 mutation. Biochemical and biophysical research communications 354, 45–9 (2007).
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