Mitochondrial remodeling and energy metabolism adaptations in colonic crypts during spontaneous epithelial repair after colitis induction in mice.

Mucosal healing has emerged as a therapeutic goal to achieve lasting clinical remission in ulcerative colitis. Intestinal repair in response to inflammation presumably requires higher energy supplies for the restoration of intestinal barrier and physiological functions. However, epithelial energy metabolism during intestinal mucosal healing has been little studied, whereas inflammation-induced alterations have been reported in the main energy production site, the mitochondria. The aim of the present work was to assess the involvement of mitochondrial activity and the events influencing their function during spontaneous epithelial repair after colitis induction in mouse colonic crypts. The results obtained show adaptations of colonocyte metabolism during colitis to ensure maximal ATP production for supporting energetic demand by both oxidative phosphorylation and glycolysis in a context of decreased mitochondrial biogenesis and through mitochondrial function restoration during colon epithelial repair. In parallel, colitis-induced mitochondrial ROS production in colonic epithelial cells was rapidly associated with transient expression of GSH-related enzymes. Mitochondrial respiration in colonic crypts was markedly increased during both inflammatory and recovery phases despite decreased expression of several mitochondrial respiratory chain complex subunits after colitis induction. Rapid induction of mitochondrial fusion was associated with mitochondrial function restoration. Finally, in contrast with the kinetics expression of genes involved in mitochondrial oxidative metabolism and in glycolysis, the expression of glutaminase was markedly reduced in the colonic crypts both during colitis and repair phases. Overall, our data suggest that the epithelial repair after colitis induction is characterized by a rapid and transient increased capacity for mitochondrial ATP production in a context of apparent restoration of mitochondrial biogenesis and metabolic reorientation of energy production. The potential implication of energy production adaptations within colonic crypts to sustain mucosal healing in a context of altered fuel supply is discussed.

Regulation of enteroendocrine cell respiration by the microbial metabolite hydrogen sulfide.

Endocrine functions of the gut are supported by a scattered population of cells, the enteroendocrine cells (EECs). EECs sense their environment to secrete hormones in a regulated manner. Distal EECs are in contact with various microbial compounds including hydrogen sulfide (H2S) which modulate cell respiration with potential consequences on EEC physiology. However, the effect of H2S on gut hormone secretion remains discussed and the importance of the modulation of cell metabolism on EEC functions remains to be deciphered. The aim of this project was to characterize the metabolic response of EECs to H2S and the consequences on GLP-1 secretion. We used cell line models of EECs to assess their capacity to metabolize H2S at low concentration and the associated modulation of cell respiration. We confirmed that like what is observed in colonocytes, colonic EEC model, NCI-h716 cell line rapidly metabolizes H2S at low concentrations, resulting in transient increased respiration. Higher concentrations of H2S inhibited this respiration, with the concentration threshold for inhibition depending on cell density. However, increased or inhibited oxidative respiration had little effect on acute GLP-1 secretion. Overall, we present here a first study showing the EEC capacity to detoxify low concentrations of H2S and used this model to acutely address the importance of cell respiration on secretory activity.

Obesogenic diet leads to luminal overproduction of the complex IV inhibitor H2 S and mitochondrial dysfunction in mouse colonocytes.

Obesity is characterized by systemic low-grade inflammation associated with disturbances of intestinal homeostasis and microbiota dysbiosis. Mitochondrial metabolism sustains epithelial homeostasis by providing energy to colonic epithelial cells (CEC) but can be altered by dietary modulations of the luminal environment. Our study aimed at evaluating whether the consumption of an obesogenic diet alters the mitochondrial function of CEC in mice. Mice were fed for 22 weeks with a 58% kcal fat diet (diet-induced obesity [DIO] group) or a 10% kcal fat diet (control diet, CTRL). Colonic crypts were isolated to assess mitochondrial function while colonic content was collected to characterize microbiota and metabolites. DIO mice developed obesity, intestinal hyperpermeability, and increased endotoxemia. Analysis of isolated colonic crypt bioenergetics revealed a mitochondrial dysfunction marked by decreased basal and maximal respirations and lower respiration linked to ATP production in DIO mice. Yet, CEC gene expression of mitochondrial respiration chain complexes and mitochondrial dynamics were not altered in DIO mice. In parallel, DIO mice displayed increased colonic bile acid concentrations, associated with higher abundance of Desulfovibrionaceae. Sulfide concentration was markedly increased in the colon content of DIO mice. Hence, chronic treatment of CTRL mouse colon organoids with sodium sulfide provoked mitochondrial dysfunction similar to that observed in vivo in DIO mice while acute exposure of isolated mitochondria from CEC of CTRL mice to sodium sulfide diminished complex IV activity. Our study provides new insights into colon mitochondrial dysfunction in obesity by revealing that increased sulfide production by DIO-induced dysbiosis impairs complex IV activity in mouse CEC.

Fate of undigested proteins in the pig large intestine: What impact on the colon epithelium?

Apart from its obvious agronomic interest in feeding billions of people worldwide, the porcine species represents an irreplaceable experimental model for intestinal physiologists and nutritionists. In this review, we give an overview on the fate of proteins that are not fully digested in the pig small intestine, and thus are transferred into the large intestine. In the large intestine, dietary and endogenous proteins are converted to peptides and amino acids (AA) by the action of bacterial proteases and peptidases. AA, which cannot, except in the neonatal period, be absorbed to any significant level by the colonocytes, are used by the intestinal microbes for protein synthesis and for the production of numerous metabolites. Of note, the production of the AA-derived metabolites greatly depends on the amount of undigested polysaccharides in the pig's diet. The effects of these AA-derived bacterial metabolites on the pig colonic epithelium have not yet been largely studied. However, the available data, performed on colonic mucosa, isolated colonic crypts and colonocytes, indicate that some of them, like ammonia, butyrate, acetate, hydrogen sulfide (H2S), and p-cresol are active either directly or indirectly on energy metabolism in colonic epithelial cells. Further studies in that area will certainly gain from the utilization of the pig colonic organoid model, which allows for disposal of functional epithelial unities. Such studies will contribute to a better understanding of the potential causal links between diet-induced changes in the luminal concentrations of these AA-derived bacterial metabolites and effects on the colon epithelial barrier function and water/electrolyte absorption.

Production of Indole and Indole-Related Compounds by the Intestinal Microbiota and Consequences for the Host: The Good, the Bad, and the Ugly.

The intestinal microbiota metabolic activity towards the available substrates generates myriad bacterial metabolites that may accumulate in the luminal fluid. Among them, indole and indole-related compounds are produced by specific bacterial species from tryptophan. Although indole-related compounds are, first, involved in intestinal microbial community communication, these molecules are also active on the intestinal mucosa, exerting generally beneficial effects in different experimental situations. After absorption, indole is partly metabolized in the liver into the co-metabolite indoxyl sulfate. Although some anti-inflammatory actions of indole on liver cells have been shown, indoxyl sulfate is a well-known uremic toxin that aggravates chronic kidney disease, through deleterious effects on kidney cells. Indoxyl sulfate is also known to provoke endothelial dysfunction. Regarding the central nervous system, emerging research indicates that indole at excessive concentrations displays a negative impact on emotional behavior. The indole-derived co-metabolite isatin appears, in pre-clinical studies, to accumulate in the brain, modulating brain function either positively or negatively, depending on the doses used. Oxindole, a bacterial metabolite that enters the brain, has shown deleterious effects on the central nervous system in experimental studies. Lastly, recent studies performed with indoxyl sulfate report either beneficial or deleterious effects depending once again on the dose used, with missing information on the physiological concentrations that are reaching the central nervous system. Any intervention aiming at modulating indole and indole-related compound concentrations in the biological fluids should crucially take into account the dual effects of these compounds according to the host tissues considered.

Dual effects of the tryptophan-derived bacterial metabolite indole on colonic epithelial cell metabolism and physiology: comparison with its co-metabolite indoxyl sulfate.

Indole, which is produced by the intestinal microbiota from L-tryptophan, is recovered at millimolar concentrations in the human feces. Indoxyl sulfate (IS), the main indole co-metabolite, can be synthesized by the host tissues. Although indole has been shown to restore intestinal barrier function in experimental colitis, little is known on the effects of indole and IS on colonic epithelial cell metabolism and physiology. In this study, we compared the effects of indole and IS on the human colonic epithelial HT-29 Glc-/+ and Caco-2 cell lines, exposed to these compounds for 1-48 h. Indole, but not IS, was cytotoxic at 5 mM, altering markedly colonocyte proliferation. Both molecules, used up to 2.5 mM, induced a transient oxidative stress in colonocytes, that was detected after 1 h, but not after 48 h exposure. This was associated with the induction after 24 h of the expression of glutathione reductase, heme oxygenase, and cytochrome P450 (CYP)1B1. Indole and IS used at 2.5 mM impaired colonocyte respiration by diminishing mitochondrial oxygen consumption and maximal respiratory capacity. Indole, but not IS, displayed a slight genotoxic effect on colonocytes. Indole, but not IS, increased transepithelial resistance in colonocyte monolayers. Indole and IS used at 2.5 mM, induced a secretion of the pro-inflammatory interleukin-8 after 3 h incubation, and an increase of tumor necrosis factor-α secretion after 48 h. Although our results suggest beneficial effect of indole on epithelial integrity, overall they indicate that indole and IS share adverse effects on colonocyte respiration and production of reactive oxygen species, in association with the induction of enzymes of the antioxidant defense system. This latter process can be viewed as an adaptive response toward oxidative stress. Both compounds increased the production of inflammatory cytokines from colonocytes. However, only indole, but not IS, affected DNA integrity in colonocytes. Since colonocytes little convert indole to IS, the deleterious effects of indole on colonocytes appear to be unrelated to its conversion to IS.

Production of hydrogen sulfide by the intestinal microbiota and epithelial cells and consequences for the colonic and rectal mucosa.

Among bacterial metabolites, hydrogen sulfide (H2S) has received increasing attention. The epithelial cells of the large intestine are exposed to two sources of H2S. The main one is the luminal source that results from specific bacteria metabolic activity toward sulfur-containing substrates. The other source in colonocytes is from the intracellular production mainly through cystathionine ß-synthase (CBS) activity. H2S is oxidized by the mitochondrial sulfide oxidation unit, resulting in ATP synthesis, and, thus, establishing this compound as the first mineral energy substrate in colonocytes. However, when the intracellular H2S concentration exceeds the colonocyte capacity for its oxidation, it inhibits the mitochondrial respiratory chain, thus affecting energy metabolism. Higher luminal H2S concentration affects the integrity of the mucus layer and displays proinflammatory effects. However, a low/minimal amount of endogenous H2S exerts an anti-inflammatory effect on the colon mucosa, pointing out the ambivalent effect of H2S depending on its intracellular concentration. Regarding colorectal carcinogenesis, forced CBS expression in late adenoma-like colonocytes increased their proliferative activity, bioenergetics capacity, and tumorigenicity; whereas, genetic ablation of CBS in mice resulted in a reduced number of mutagen-induced aberrant crypt foci. Activation of endogenous H2S production and low H2S extracellular concentration enhance cancerous colorectal cell proliferation. Higher exogenous H2S concentrations markedly reduce mitochondrial ATP synthesis and proliferative capacity in cancerous cells and enhance glycolysis but do not affect their ATP cell content or viability. Thus, it appears that, notably through an effect on colonocyte energy metabolism, endogenous and microbiota-derived H2S are involved in the host intestinal physiology and physiopathology.

Sulfur-Containing Amino Acids and Lipid Metabolism.

The metabolism of methionine and cysteine in the body tissues determines the concentrations of several metabolites with various biologic activities, including homocysteine, hydrogen sulfide (H2S), taurine, and glutathione. Hyperhomocysteinemia, which is correlated with lower HDL cholesterol in blood in volunteers and animal models, has been associated with an increased risk for cardiovascular diseases. In humans, the relation between methionine intake and hyperhomocysteinemia is dependent on vitamin status (vitamins B-6 and B-12 and folic acid) and on the supply of other amino acids. However, lowering homocysteinemia by itself is not sufficient for decreasing the risk of cardiovascular disease progression. Other compounds related to methionine metabolism have recently been identified as being involved in the risk of atherosclerosis and steatohepatitis. Indeed, the metabolism of sulfur amino acids has an impact on phosphatidylcholine (PC) metabolism, and anomalies in PC synthesis due to global hypomethylation have been associated with disturbances of lipid metabolism. In addition, impairment of H2S synthesis from cysteine favors atherosclerosis and steatosis in animal models. The effects of taurine on lipid metabolism appear heterogeneous depending on the populations of volunteers studied. A decrease in the concentration of intracellular glutathione, a tripeptide involved in redox homeostasis, is implicated in the etiology of cardiovascular diseases and steatosis. Last, supplementation with betaine, a compound that allows remethylation of homocysteine to methionine, decreases basal and methionine-stimulated homocysteinemia; however, it adversely increases plasma total and LDL cholesterol. The study of these metabolites may help determine the range of optimal and safe intakes of methionine and cysteine in dietary proteins and supplements. The amino acid requirement for protein synthesis in different situations and for optimal production of intracellular compounds involved in the regulation of lipid metabolism also needs to be considered for dietary attenuation of atherosclerosis and steatosis risk.

Hypertonic external medium represses cellular respiration and promotes Warburg/Crabtree effect.

Hyperosmotic conditions are associated to several pathological states. In this article, we evaluate the consequence of hyperosmotic medium on cellular energy metabolism. We demonstrate that exposure of cells to hyperosmotic conditions immediately reduces the mitochondrial oxidative phosphorylation rate. This causes an increase in glycolysis, which represses further respiration. This is known as the Warburg or Crabtree effect. In addition to aerobic glycolysis, we observed two other cellular responses that would help to preserve cellular ATP level and viability: A reduction in the cellular ATP turnover rate and a partial mitochondrial uncoupling which is expected to enhance ATP production by Krebs cycle. The latter is likely to constitute another metabolic adaptation to compensate for deficient oxidative phosphorylation that, importantly, is not dependent on glucose.

Protective Effect of an Avocado Peel Polyphenolic Extract Rich in Proanthocyanidins on the Alterations of Colonic Homeostasis Induced by a High-Protein Diet.

Avocado peel, a byproduct from the avocado pulp industry, is a promising source of polyphenolic compounds. We evaluated the effect of a proanthocyanidin-rich avocado peel polyphenol extract (AvPPE) on the composition and metabolic activity of human fecal microbiota cultured for 24 h in a bioreactor in the presence of high protein (HP) amounts and the effect of the resulting culture supernatants (CSs) on HT-29Glc-/+ and Caco-2 cells. AvPPE decreased the HP-induced production of ammonia, H2S, propionate, and isovalerate and increased that of indole and butyrate. Microbiota composition was marginally affected by HP, whileAvPPE increased the microorganisms/abundance of phylum Actinobacteria, families Coriobacteriaceae and Ruminococcaceae, and genus Faecalibacterium. AvPPE failed to prevent the HP-induced decrease of HT-29Glc-/+ cell viability and energy efficiency but prevented the HP-induced alterations of barrier function in Caco-2 cells. Additionally, the genotoxic effect of the CSs upon HT-29Glc-/+ was attenuated by AvPPE. Therefore, AvPPE may be considered as a promising product for improving colonic homeostasis.

Hyperosmolar environment and intestinal epithelial cells: impact on mitochondrial oxygen consumption, proliferation, and barrier function in vitro.

The aim of the present study was to elucidate the in vitro short-term (2-h) and longer-term (24-h) effects of hyperosmolar media (500 and 680 mOsm/L) on intestinal epithelial cells using the human colonocyte Caco-2 cell line model. We found that a hyperosmolar environment slowed down cell proliferation compared to normal osmolarity (336 mOsm/L) without inducing cell detachment or necrosis. This was associated with a transient reduction of cell mitochondrial oxygen consumption, increase in proton leak, and decrease in intracellular ATP content. The barrier function of Caco-2 monolayers was also transiently affected since increased paracellular apical-to-basal permeability and modified electrolyte permeability were measured, allowing partial equilibration of the trans-epithelial osmotic difference. In addition, hyperosmotic stress induced secretion of the pro-inflammatory cytokine IL-8. By measuring expression of genes involved in energy metabolism, tight junction forming, electrolyte permeability and intracellular signaling, different response patterns to hyperosmotic stress occurred depending on its intensity and duration. These data highlight the potential impact of increased luminal osmolarity on the intestinal epithelium renewal and barrier function and point out some cellular adaptive capacities towards luminal hyperosmolar environment.

Mucosal healing progression after acute colitis in mice.

BACKGROUND: Mucosal healing has become a therapeutic goal to achieve stable remission in patients with inflammatory bowel diseases. To achieve this objective, overlapping actions of complex cellular processes, such as migration, proliferation, and differentiation, are required. These events are longitudinally and tightly controlled by numerous factors including a wide range of distinct regulatory proteins. However, the sequence of events associated with colon mucosal repair after colitis and the evolution of the luminal content characteristics during this process have been little studied. AIM: To document the evolution of colon mucosal characteristics during mucosal healing using a mouse model with chemically-induced colitis. METHODS: C57BL/6 male mice were given 3.5% dextran sodium sulfate (DSS) in drinking water for 5 d. They were euthanized 2 (day 7), 5 (day 10), 8 (day 13), and 23 (day 28) d after DSS removal. The colonic luminal environment and epithelial repair processes during the inflammatory flare and colitis resolution were analyzed with reference to a non-DSS treated control group, euthanized at day 0. Epithelial repair events were assessed histo-morphologically in combination with functional permeability tests, expression of key inflammatory and repairing factors, and evaluation of colon mucosa-adherent microbiota composition by 16S rRNA sequencing. RESULTS: The maximal intensity of colitis was concomitant with maximal alterations of intestinal barrier function and histological damage associated with goblet cell depletion in colon mucosa. It was recorded 2 d after termination of the DSS-treatment, followed by a progressive return to values similar to those of control mice. Although signs of colitis were severe (inflammatory cell infiltrate, crypt disarray, increased permeability) and associated with colonic luminal alterations (hyperosmolarity, dysbiosis, decrease in short-chain fatty acid content), epithelial healing processes were launched early during the inflammatory flare with increased gene expression of certain key epithelial repair modulators, including transforming growth factor-ß, interleukin (Il)-15, Il-22, Il-33, and serum amyloid A. Whereas signs of inflammation progressively diminished, luminal colonic environment alterations and microscopic abnormalities of colon mucosa persisted long after colitis induction. CONCLUSION: This study shows that colon repair can be initiated in the context of inflamed mucosa associated with alterations of the luminal environment and highlights the longitudinal involvement of key modulators.

Effect of a proanthocyanidin-rich polyphenol extract from avocado on the production of amino acid-derived bacterial metabolites and the microbiota composition in rats fed a high-protein diet.

The consumption of high-protein diets (HPDs) increases the flux of undigested proteins moving to the colon. These proteins are hydrolyzed by bacterial proteases and peptidases, releasing amino acids, which in turn are metabolized by the intestinal microbiota (IM) for protein synthesis and production of various metabolites that can exert positive or deleterious effects, depending on their concentrations, at the colonic or systemic level. On the other hand, proanthocyanidins are polymers of flavan-3-ols which cannot be absorbed at the intestinal level, accumulating in the colon where they are fermented by the IM producing metabolites that appear beneficial for colonocytes and also at the peripheral level. This study evaluated the effect of an avocado peel polyphenol extract (AvPPE) rich in proanthocyanidins on the production of cecal bacterial metabolites and microbiota composition in rats fed a HPD. Compared with the normal-protein (NP) group, HPD did not markedly affect the body weight gain of the animals, but increased the kidney weight. Additionally, the HPD induced a higher cecal concentration of ammonia (NH4+/NH3), hydrogen sulfide (H2S) and branched-chain fatty acids (BCFAs). The supplementation with AvPPE attenuated the production of H2S and increased the production of indole. On the other hand, the HPD affected the composition of the cecal microbiota, increasing the relative abundance of the genera Bacteroides and Lactobacillus, while decreasing Prevotella. The AvPPE counteracted the increase induced by the HPD on the genus Lactobacillus, and increased the relative abundance of [Prevotella]. Our results contribute towards explaining the health-promoting effects of proanthocyanidin-rich dietary foodstuffs including fruits and vegetables.

In vitro impact of amino acid-derived bacterial metabolites on colonocyte mitochondrial activity, oxidative stress response and DNA integrity.

BACKGROUND: 4-hydroxyphenylacetic acid (HO-PAA) is produced by intestinal microbiota from L-tyrosine. High concentrations in human fecal water have been associated with cytotoxicity, urging us to test HO-PAA's effects on human colonocytes. We compared these effects with those of phenylacetic acid (PAA), phenol and acetaldehyde, also issued from amino acids fermentation. METHODS: HT-29 Glc-/+ human colonocytes were exposed for 24 h to metabolites at concentrations between 350 and 1000 µM for HO-PAA and PAA, 250-1500 µM for phenol and 25-500 µM for acetaldehyde. We evaluated metabolites'cytotoxicity with 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide and DNA quantification assays, reactive oxygen species (ROS) production with H2DCF-DA, and DNA damage with the comet assay. We measured cell oxygen consumption and mitochondrial complexes activity by polarography. RESULTS: Although HO-PAA displayed no cytotoxic effect on colonocytes, it decreased mitochondrial complex I activity and oxygen consumption. This was paralleled by an increase in ROS production and DNA alteration. Cells pretreatment with N-acetylcysteine, a ROS scavenger, decreased genotoxic effects of HO-PAA, indicating implication of oxidative stress in HO-PAA's genotoxicity. PAA and phenol did not reproduce these effects, but were cytotoxic towards colonocytes. Last, acetaldehyde displayed no effect in terms of cytotoxicity and mitochondrial metabolic activity, but increased DNA damage. CONCLUSIONS: Several bacterial metabolites produced from amino acids displayed deleterious effects on human colonocytes, in terms of genotoxicity (HO-PAA and acetaldehyde) or cytotoxicity (PAA and phenol). GENERAL SIGNIFICANCE: This study helps understanding the consequences of intestinal microbiota's metabolic activity on the host since amino acids fermentation can lead to the formation of compounds toxic towards colonic epithelial cells.

Dietary Protein Intake Level Modulates Mucosal Healing and Mucosa-Adherent Microbiota in Mouse Model of Colitis.

Mucosal healing after an inflammatory flare is associated with lasting clinical remission. The aim of the present work was to evaluate the impact of the amount of dietary protein on epithelial repair after an acute inflammatory episode. C57BL/6 DSS-treated mice received isocaloric diets with different levels of dietary protein: 14% (P14), 30% (P30) and 53% (P53) for 3 (day 10), 6 (day 13) and 21 (day 28) days after the time of colitis maximal intensity. While the P53 diet worsened the DSS- induced inflammation both in intensity and duration, the P30 diet, when compared to the P14 diet, showed a beneficial effect during the epithelial repair process by accelerating inflammation resolution, reducing colonic permeability and increasing epithelial repair together with epithelial hyperproliferation. Dietary protein intake also impacted mucosa-adherent microbiota composition after inflammation since P30 fed mice showed increased colonization of butyrate-producing genera throughout the resolution phase. This study revealed that in our colitis model, the amount of protein in the diet modulated mucosal healing, with beneficial effects of a moderately high-protein diet, while very high-protein diet displayed deleterious effects on this process.

High-protein diets for weight management: Interactions with the intestinal microbiota and consequences for gut health. A position paper by the my new gut study group.

BACKGROUND & AIMS: This review examines to what extent high-protein diets (HPD), which may favor body weight loss and improve metabolic outcomes in overweight and obese individuals, may also impact the gut environment, shaping the microbiota and the host-microbe (co)metabolic pathways and products, possibly affecting large intestine mucosa homeostasis. METHODS: PubMed-referenced publications were analyzed with an emphasis on dietary intervention studies involving human volunteers in order to clarify the beneficial vs. deleterious effects of HPD in terms of both metabolic and gut-related health parameters; taking into account the interactions with the gut microbiota. RESULTS: HPD generally decrease body weight and improve blood metabolic parameters, but also modify the fecal and urinary contents in various bacterial metabolites and co-metabolites. The effects of HPD on the intestinal microbiota composition appear rather heterogeneous depending on the type of dietary intervention. Recently, HPD consumption was shown to modify the expression of genes playing key roles in homeostatic processes in the rectal mucosa, without evidence of intestinal inflammation. Importantly, the effects of HPD on the gut were dependent on the protein source (i.e. from plant or animal sources), a result which should be considered for further investigations. CONCLUSION: Although HPD appear to be efficient for weight loss, the effects of HPD on microbiota-derived metabolites and gene expression in the gut raise new questions on the impact of HPD on the large intestine mucosa homeostasis leading the authors to recommend some caution regarding the utilization of HPD, notably in a recurrent and/or long-term ways.

Proanthocyanidin-containing polyphenol extracts from fruits prevent the inhibitory effect of hydrogen sulfide on human colonocyte oxygen consumption.

Hydrogen sulfide (H2S), a metabolic end product synthesized by the microbiota from L-cysteine, has been shown to act at low micromolar concentration as a mineral oxidative substrate in colonocytes while acting as an inhibitor of oxygen consumption at higher luminal concentrations (65 µM and above). From the previous works showing that polyphenols can bind volatile sulfur compounds, we hypothesized that different dietary proanthocyanidin-containing polyphenol (PACs) plant extracts might modulate the inhibitory effect of H2S on colonocyte respiration. Using the model of human HT-29 Glc-/+ cell colonocytes, we show here that pre-incubation of 65 µM of the H2S donor NaHS with the different polyphenol extracts markedly reduced the inhibitory effect of NaHS on colonocyte oxygen consumption. Our studies on HT-29 Glc-/+ cell respiration performed in the absence or the presence of PACs reveal rapid binding of H2S with the sulfide-oxidizing unit and slower binding of H2S to the cytochrome c oxidase (complex IV of the respiratory chain). Despite acute inhibition of colonocyte respiration, no measurable effect of NaHS on paracellular permeability was recorded after 24 h treatment using the Caco-2 colonocyte monolayer model. The results are discussed in the context of the binding of excessive bacterial metabolites by unabsorbed dietary compounds and of the capacity of colonocytes to adapt to changing luminal environment.

Quantity and source of dietary protein influence metabolite production by gut microbiota and rectal mucosa gene expression: a randomized, parallel, double-blind trial in overweight humans.

Background: Although high-protein diets (HPDs) are frequently consumed for body-weight control, little is known about the consequences for gut microbiota composition and metabolic activity and for large intestine mucosal homeostasis. Moreover, the effects of HPDs according to the source of protein need to be considered in this context.Objective: The objective of this study was to evaluate the effects of the quantity and source of dietary protein on microbiota composition, bacterial metabolite production, and consequences for the large intestinal mucosa in humans.Design: A randomized, double-blind, parallel-design trial was conducted in 38 overweight individuals who received a 3-wk isocaloric supplementation with casein, soy protein, or maltodextrin as a control. Fecal and rectal biopsy-associated microbiota composition was analyzed by 16S ribosomal DNA sequencing. Fecal, urinary, and plasma metabolomes were assessed by 1H-nuclear magnetic resonance. Mucosal transcriptome in rectal biopsies was determined with the use of microarrays.Results: HPDs did not alter the microbiota composition, but induced a shift in bacterial metabolism toward amino acid degradation with different metabolite profiles according to the protein source. Correlation analysis identified new potential bacterial taxa involved in amino acid degradation. Fecal water cytotoxicity was not modified by HPDs, but was associated with a specific microbiota and bacterial metabolite profile. Casein and soy protein HPDs did not induce inflammation, but differentially modified the expression of genes playing key roles in homeostatic processes in rectal mucosa, such as cell cycle or cell death.Conclusions: This human intervention study shows that the quantity and source of dietary proteins act as regulators of gut microbiota metabolite production and host gene expression in the rectal mucosa, raising new questions on the impact of HPDs on the large intestine mucosa homeostasis. This trial was registered at clinicaltrials.gov as NCT02351297.

Epithelial response to a high-protein diet in rat colon.

BACKGROUND: High-protein diets (HPD) alter the large intestine microbiota composition in association with a metabolic shift towards protein degradation. Some amino acid-derived metabolites produced by the colon bacteria are beneficial for the mucosa while others are deleterious at high concentrations. The aim of the present work was to define the colonic epithelial response to an HPD. Transcriptome profiling was performed on colonocytes of rats fed an HPD or an isocaloric normal-protein diet (NPD) for 2 weeks. RESULTS: The HPD downregulated the expression of genes notably implicated in pathways related to cellular metabolism, NF-κB signaling, DNA repair, glutathione metabolism and cellular adhesion in colonocytes. In contrast, the HPD upregulated the expression of genes related to cell proliferation and chemical barrier function. These changes at the mRNA level in colonocytes were not associated with detrimental effects of the HPD on DNA integrity (comet assay), epithelium renewal (quantification of proliferation and apoptosis markers by immunohistochemistry and western blot) and colonic barrier integrity (Ussing chamber experiments). CONCLUSION: The modifications of the luminal environment after an HPD were associated with maintenance of the colonic homeostasis that might be the result of adaptive processes in the epithelium related to the observed transcriptional regulations.

Changes in the Luminal Environment of the Colonic Epithelial Cells and Physiopathological Consequences.

Evidence, mostly from experimental models, has accumulated, indicating that modifications of bacterial metabolite concentrations in the large intestine luminal content, notably after changes in the dietary composition, may have important beneficial or deleterious consequences for the colonic epithelial cell metabolism and physiology in terms of mitochondrial energy metabolism, reactive oxygen species production, gene expression, DNA integrity, proliferation, and viability. Recent data suggest that for some bacterial metabolites, like hydrogen sulfide and butyrate, the extent of their oxidation in colonocytes affects their capacity to modulate gene expression in these cells. Modifications of the luminal bacterial metabolite concentrations may, in addition, affect the colonic pH and osmolarity, which are known to affect colonocyte biology per se. Although the colonic epithelium appears able to face, up to some extent, changes in its luminal environment, notably by developing a metabolic adaptive response, some of these modifications may likely affect the homeostatic process of colonic epithelium renewal and the epithelial barrier function. The contribution of major changes in the colonocyte luminal environment in pathological processes, like mucosal inflammation, preneoplasia, and neoplasia, although suggested by several studies, remains to be precisely evaluated, particularly in a long-term perspective.