Metabolism Of Short-chain Fatty Acids Research Articles

Colonic microbial metabolite and flux analysis

The colon is one of the body’s metabolically most active organs. While insight in colonic microbial diversity increases, little is known of the metabolic function of the microbes, the way diet affects metabolic fluxes, and how produced metabolites affect human health. Therefore, as a first step in detailed metabolomic characterization of colonic carbohydrate metabolism, we studied the conversion by human gut bacteria of the model substrate glucose. An in vitro model of the colon was inoculated with a standardised human fecal microbiota. D-[U-13C]glucose was added at t=0, and lumen as well as dialysate samples were taken at various time points. GC-MS, enzymatic methods and 1- and 2-dimensional NMR techniques at 500 MHz were used to identify microbial metabolites and to analyze their 13C contents. SCFA (short-chain fatty acids), formate and lactate were determined as the principal glucose degradation products. Carbon balancing revealed a 95% recovery of the 13C glucose carbon. Several minor components, including succinate, aspartate and alanine, were identified using 2D NMR experiments. Quantitative insight in bacterial metabolic routes was obtained from analysis of 13C-13C coupling patterns in 2D HSQC spectra that resulted from incorporation of intact 13C glucose backbone fragments in SCFA. In conclusion, this stable isotope study underlines the importance of colonic bacterial SCFA metabolism.

A novel class of CoA-transferase involved in short-chain fatty acid metabolism in butyrate-producing human colonic bacteria

Bacterial butyryl-CoA CoA-transferase activity plays a key role in butyrate formation in the human colon, but the enzyme and corresponding gene responsible for this activity have not previously been identified. A novel CoA-transferase gene is described from the colonic bacterium Roseburia sp. A2-183, with similarity to acetyl-CoA hydrolase as well as 4-hydroxybutyrate CoA-transferase sequences. The gene product, overexpressed in an Escherichia coli lysate, showed activity with butyryl-CoA and to a lesser degree propionyl-CoA in the presence of acetate. Butyrate, propionate, isobutyrate and valerate competed with acetate as the co-substrate. Despite the sequence similarity to 4-hydroxybutyrate CoA-transferases, 4-hydroxybutyrate did not compete with acetate as the co-substrate. Thus the CoA-transferase preferentially uses butyryl-CoA as substrate. Similar genes were identified in other butyrate-producing human gut bacteria from clostridial clusters IV and XIVa, while other candidate CoA-transferases for butyrate formation could not be detected in Roseburia sp. A2-183. This suggests strongly that the newly identified group of CoA-transferases described here plays a key role in butyrate formation in the human colon.

Crystallographic Trapping of the Glutamyl-CoA Thioester Intermediate of Family I CoA Transferases

Coenzyme A transferases are involved in a broad range of biochemical processes in both prokaryotes and eukaryotes, and exhibit a diverse range of substrate specificities. The YdiF protein from Escherichia coli O157:H7 is an acyl-CoA transferase of unknown physiological function, and belongs to a large sequence family of CoA transferases, present in bacteria to humans, which utilize oxoacids as acceptors. In vitro measurements showed that YdiF displays enzymatic activity with short-chain acyl-CoAs. The crystal structures of YdiF and its complex with CoA, the first co-crystal structure for any Family I CoA transferase, have been determined and refined at 1.9 and 2.0 A resolution, respectively. YdiF is organized into tetramers, with each monomer having an open alpha/beta structure characteristic of Family I CoA transferases. Co-crystallization of YdiF with a variety of CoA thioesters in the absence of acceptor carboxylic acid resulted in trapping a covalent gamma-glutamyl-CoA thioester intermediate. The CoA binds within a well defined pocket at the N- and C-terminal domain interface, but makes contact only with the C-terminal domain. The structure of the YdiF complex provides a basis for understanding the different catalytic steps in the reaction of Family I CoA transferases.

Role of short-chain fatty acids in colonic HCO3secretion

Luminal isobutyrate, a relatively poor metabolized short-chain fatty acid (SCFA), induces HCO(3) secretion via a Cl-independent, DIDS-insensitive, carrier-mediated process as well as inhibiting both Cl-dependent and cAMP-induced HCO(3) secretion. The mechanism(s) responsible for these processes have not been well characterized. HCO(3) secretion was measured in isolated colonic mucosa mounted in Lucite chambers using pH stat technique and during microperfusion of isolated colonic crypts. (14)C-labeled butyrate, (14)C-labeled isobutyrate, and (36)Cl uptake were also determined by apical membrane vesicles (AMV) isolated from surface and/or crypt cells. Butyrate stimulation of Cl-independent, DIDS-insensitive 5-nitro-3-(3-phenylpropyl-amino)benzoic acid-insensitive HCO(3) secretion is greater than that by isobutyrate, suggesting that both SCFA transport and metabolism are critical for HCO(3) secretion. Both lumen and serosal 25 mM butyrate inhibit cAMP-induced HCO(3) secretion to a comparable degree (98 vs. 90%). In contrast, Cl-dependent HCO(3) secretion is downregulated by lumen 25 mM butyrate considerably more than by serosal butyrate (98 vs. 37%). Butyrate did not induce HCO(3) secretion in isolated microperfused crypts, whereas an outward-directed HCO(3) gradient-driven induced (14)C-butyrate uptake by surface but not crypt cell AMV. Both (36)Cl/HCO(3) exchange and potential-dependent (36)Cl movement in AMV were inhibited by 96-98% by 20 mM butyrate. We conclude that 1) SCFA-dependent HCO(3) secretion is the result of SCFA transport across the apical membrane via a SCFA/HCO(3) exchange more than intracellular SCFA metabolism; 2) SCFA-dependent HCO(3) secretion is most likely a result of an apical membrane SCFA/HCO(3) exchange in surface epithelial cells; 3) SCFA downregulates Cl-dependent and cAMP-induced HCO(3) secretion secondary to SCFA inhibition of apical membrane Cl/HCO(3) exchange and anion channel activity, respectively.

Rumen Development, Intestinal Growth and Hepatic Metabolism In The Pre- and Postweaning Ruminant

Abstract Both the role of intestinal development and the process of transitioning calves from their neonatal reliance on nutrients supplied from milk to nutrients supplied from grain are of substantial economic importance to the producer. Improvements to the calf nutritional regime can decrease mortality and disease susceptibility, increase postweaning rate of gain, ultimately enhancing the rate of herd genetic improvement (due to increased capacity for voluntary culling). Current feeding practices including daily encouragement to eat grain diets result in weaning from milk by 3 to 4 wk of age. Although the mechanisms are not yet completely understood, development of a viable fermentation within the rumen is required to initiate the maturation of the rumen epithelia. This transition results in tremendous metabolic ramifications to calf growth rate, as tissues must convert from reliance on glucose supplied from milk to the metabolism of short-chain fatty acids as primary energy substrates. This maturation is the result of differential expression of numerous genes regulating both physical and metabolic characteristics of the tissue. While the most dramatic physical changes occurring during development are associated with the rumen epithelium, changes in intestinal mass and metabolism are also realized in response to dietary changes. Amino acid use by the intestinal tissues is high and may affect amino acid availability in support of growth. Moreover, because the metabolic and protein synthetic activities of the digestive tract are high, accounting for up to 30% of both whole animal energy use and whole animal protein synthesis, understanding their regulation is vital to the continued improvement in calf management. Specific nutrient-gene interactions have been identified across the digestive tract and serve to increase visceral organ mass and directly change nutrient metabolism by the epithelia. Similarly, humoral factors have been identified as having regulatory function over gastrointestinal tissue mass and metabolism. Continuing efforts to better understand the factors affecting intestinal development will improve weaning strategies and foster better postweanling calf growth performance.

The role of bacterial antizyme: From an inhibitory protein to AtoC transcriptional regulator

This review considers the role of bacterial antizyme in the regulation of polyamine biosynthesis and gives new perspectives on the involvement of antizyme in other significant cellular mechanisms. Antizyme is a protein molecule induced by the end product of the enzymic reaction that it inhibits, in a non-competitive manner. The bacterial ornithine decarboxylase is regulated by nucleotides, phosphorylation and antizyme. The inhibition of ornithine decarboxylase by antizyme can be relieved to different degrees by DNA or by a variety of synthetic nucleic acid polymers, attributed to a specific interaction between nucleic acid and antizyme. Recently, this interplay between bacterial antizyme and nucleic acid was determined by discerning an additional function to antizyme that proved to be the atoC gene product, encoding the response regulator of the bacterial two-component system AtoS-AtoC. The gene located just upstream of atoC encodes the sensor kinase, named AtoS, that modulates AtoC activity. AtoC regulates expression of atoDAEB operon which is involved in short-chain fatty acid metabolism. Antizyme is thus referred to as AtoC, functioning both as a post-translational and transcriptional regulator. Also, the AtoS-AtoC signal transduction system in E. coli has a positive regulatory role on poly-(R)-3-hydroxybutyrate biosynthesis. The properties and gene structural similarities of antizymes from different organisms were compared. It was revealed that conserved domains are present mostly in the C-domain of all antizymes. BLAST analysis of the E. coli antizyme protein (AtoC) showed similarities around 69–58% among proteobacteria, g-proteobacteria, enterobacteria and the thermophilic bacterium Thermus thermophilus. A working hypothesis is proposed for the metabolic role of antizyme (AtoC) describing the significant biological implications of this protein molecule. Whether antizymes exist to other enzymes in different tissues, meeting the criteria discussed in the text remains to be elucidated.

Glucose alleviates ammonia-induced inhibition of short-chain fatty acid metabolism in rat colonic epithelial cells.

Ammonia decreased metabolism by rat colonic epithelial cells of butyrate and acetate to CO2 and ketones but increased oxidation of glucose and glutamine. Ammonia decreased cellular concentrations of oxaloacetate for all substrates evaluated. The extent to which butyrate carbon was oxidized to CO2 after entering the tricarboxylic acid (TCA) cycle was not significantly influenced by ammonia, suggesting there was no major shift toward efflux of carbon from the TCA cycle. Ammonia reduced entry of butyrate carbon into the TCA cycle, and the proportion of CoA esterified with acetate and butyrate correlated positively with the production of CO2 and ketone bodies. Also, ammonia reduced oxidation of propionate but had no effect on oxidation of 3-hydroxybutyrate. Inclusion of glucose, lactate, or glutamine with butyrate and acetate counteracted the ability of ammonia to decrease their oxidation. In rat colonocytes, it appears that ammonia suppresses short-chain fatty acid (SCFA) oxidation by inhibiting a step before or during their activation. This inhibition is alleviated by glucose and other energy-generating compounds. These results suggest that ammonia may only affect SCFA metabolism in vivo when glucose availability is compromised.

Production rates and metabolism of short-chain fatty acids in the colon and whole body using stable isotopes.

Short-chain fatty acids (SCFA; mainly acetate, propionate and butyrate) are largely produced in non-ruminants during the colonic bacterial fermentation of non-digestible carbohydrates. These intestinal exogenous SCFA pass in part through the splanchnic bed and reach the peripheral bloodstream, mixing with the endogenous circulating SCFA. The whole-body turnover of SCFA is thus composed of an endogenous peripheral turnover and an exogenous production that depends on dietary intake of non-digestible carbohydrates. In the present work methods were developed for determining the SCFA turnover in animals and in human subjects using stable isotopes. The original studies performed to determine endogenous and exogenous metabolism of SCFA in animals and in human subjects are summarised. Using intravenous infusion of 13C-labelled SCFA the whole-body turnover of acetate, propionate and butyrate was assessed in rats in a fasted state. The endogenous turnover of acetate and its oxidation were determined in healthy human subjects in the post-absorptive state, using intravenous infusion of [1-13C]acetate. Intragastric tracer infusions were performed to evaluate the splanchnic first-pass retention of acetate in adults. Finally, an original model was developed in healthy human subjects using intravenous infusion of [1-13C]acetate to determine in vivo the true colonic acetate production after ingestion of a non-digestible disaccharide. These present studies using stable isotopes provide the basis for a novel strategy to evaluate in vivo, in human subjects, the production of SCFA in the large intestine.

Transport of ketone bodies and lactate in the sheep ruminal epithelium by monocarboxylate transporter 1.

Due to intensive intracellular metabolism of short-chain fatty acids, ruminal epithelial cells generate large amounts of D-beta-hydroxybutyric acid, acetoacetic acid, and lactic acid. These acids have to be extruded from the cytosol to avoid disturbances of intracellular pH (pH(i)). To evaluate acid extrusion, pH(i) was studied in cultured ruminal epithelial cells of sheep using the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. Extracellular addition of D-beta-hydroxybutyrate, acetoacetate, or lactate (20 mM) resulted in intracellular acidification. Vice versa, removing extracellular D-beta-hydroxybutyrate, acetoacetate, or lactate after preincubation with the respective monocarboxylate induced an increase of pH(i). Initial rate of pH(i) decrease as well as of pH(i) recovery was strongly inhibited by pCMBS (400 microM) and phloretin (20 microM). Both cultured cells and intact ruminal epithelium were tested for the possible presence of proton-linked monocarboxylate transporter (MCT) on both the mRNA and protein levels. With the use of RT-PCR, mRNA encoding for MCT1 isoform was demonstrated in cultured ruminal epithelial cells and the ruminal epithelium. Immunostaining with MCT1 antibodies intensively labeled cultured ruminal epithelial cells and cells located in the stratum basale of the ruminal epithelium. In conclusion, our data indicate that MCT1 is expressed in the stratum basale of the ruminal epithelium and may function as a main mechanism for removing ketone bodies and lactate together with H+ from the cytosol into the blood.

Whole Wheat and Triticale Flours with Differing Viscosities Stimulate Cecal Fermentations and Lower Plasma and Hepatic Lipids in Rats

Whole flours from oat, rye or barley effectively modify digestive fermentation and lipid metabolism, whereas the effectiveness of whole wheat flour has not been established. To address this question, cecal digestion, short-chain fatty acid (SCFA) metabolism and cholesterol metabolism were investigated in four groups of rats fed the following semipurified diets differing in their carbohydrate source: a control diet (purified wheat starch) and three whole cereal flour diets [Valoris wheat (Wv), Soissons wheat (Ws), or Carnac triticale (Tc)]. Wv is particularly viscous and rich in arabinoxylans, and Tc is richer in hemicellulose than wheat. Compared with controls, rats fed the whole-flour diets had enlarged ceca and a moderate acidification of the bulk pH ( approximately 6.4). In these rats, the cecal SCFA pool size was enhanced (P < 0.05), and the SCFA molar ratio reflected propionic/butyric acid-rich fermentations, especially in those fed TC: The portal SCFA concentrations reflected the rise of the acetic and propionic acid pools in the cecum, whereas portal butyric acid remained relatively low, probably reflecting extensive metabolism by the cecal wall. The fecal excretion of total steroids (bile acids + sterols) was markedly enhanced by all of the whole-flour diets, with Wv (+78%) > Tc (+64%) > Ws (+47%). In parallel, there was a significant plasma cholesterol-lowering effect for rats fed Wv (-27%) and Tc (-32%) and a plasma triglyceride-lowering effect (approximately -40%) in all rats fed whole-flour diets (P < 0.05). This effect was observed mainly for triglyceride-rich lipoprotein-cholesterol, whereas HDL cholesterol was unaffected. These results indicate that whole wheat flours can strikingly affect cecal SCFA, especially butyrate, and are effective plasma cholesterol-lowering agents.

In vivo absorption of medium-chain fatty acids by the rat colon exceeds that of short-chain fatty acids

Short-chain fatty acids (SCFAs) are main fuels of the colonic epithelium, and are avidly absorbed by the colon of animal and man. The current knowledge on colonic metabolism and absorption of medium-chain fatty acids (MCFAs) is limited. In some clinical situations, colonic absorption of high-energy substances could compensate for reduced absorptive capacity because of a shortened or malfunctioning small bowel. We evaluated and compared colonic absorption and metabolism of MCFAs (octanoate, decanoate, and dodecanoate), SCFAs (acetate and butyrate), and long-chain fatty acids (LCFAs) (oleate). Rats were surgically operated on to cannulate a 7-cm segment of proximal colon, isolate the vasculature, and cannulate the right colic vein draining this segment. The lumen was perfused with (14)C-labeled substrates for 100 minutes. Right colic vein blood was analyzed for total (14)C, (14)CO(2), and metabolites by scintillation counting and high-performance liquid chromatography. The transport from the colonic lumen to mesenteric blood of substrate carbon from MCFAs exceeded by 2-13-fold that of SCFAs and LCFAs. The CO(2) production from the oxidation of MCFAs was as high as or higher than that from SCFAs. CO(2) produced from the LCFA, oleate, was lower than from SCFAs or MCFAs. In addition to CO(2), ketone bodies were major metabolites of SCFAs and MCFAs. Ketogenesis from butyrate and the MCFAs was significantly higher than from acetate and oleate. A substantial proportion (50%-90%) of all substrates was absorbed without being metabolized. The colonic epithelium serves to absorb and partially metabolize MCFAs. For patients with a compromised small-bowel function, colonic absorption of MCFAs could represent an important way of receiving calories.

Effects of short chain fatty acids on colonic Na + absorption and enzyme activity

Short chain fatty acids (SCFA) stimulate colonic Na + absorption and inhibit cAMP and cGMP-mediated Cl − secretion. It is uncertain whether SCFA have equivalent effects on absorption and whether SCFA inhibition of Cl − secretion involves effects on mucosal enzymes. Unidirectional Na + fluxes were measured across stripped colonic segments in the Ussing chamber. Enzyme activity was measured in cell fractions of scraped colonic mucosa. Mucosal 50 mM acetate, propionate, butyrate and poorly metabolized isobutyrate stimulated proximal colon Na + absorption equally (300%). Neither 2-bromo-octanoate, an inhibitor of β-oxidation, nor carbonic anhydrase inhibition affected this stimulation. All SCFA except acetate stimulated distal colon Na + absorption 200%. Only one SCFA affected proximal colon cGMP phosphodiesterase (PDE) (18% inhibition by 50 mM butyrate). All SCFA at 50 mM stimulated distal colon cAMP PDE (24–43%) and decreased forskolin-stimulated mucosal cAMP content. None of the SCFA affected forskolin-stimulated adenylyl cyclase in distal colon or ST a-stimulated guanylyl cyclase in proximal colon. Na +–K +-ATPase in distal colon was inhibited 23–51% by the SCFA at 50 mM. We conclude that all SCFA (except acetate in distal colon) stimulate colonic Na + absorption equally, and the mechanism does not involve mucosal SCFA metabolism or carbonic anhydrase. SCFA inhibition of cAMP-mediated secretion may involve SCFA stimulation of PDE and inhibition of Na +–K +-ATPase.

Portal recovery of short-chain fatty acids infused into the temporarily-isolated and washed reticulo-rumen of sheep

The present study was undertaken to study the metabolism of short-chain fatty acids (SCFA) by the reticulo-ruminal epithelium and the portal-drained viscera (PDV) under in vivo conditions with no interference from the metabolism of the rumen microbes. The technique of temporary isolation of the reticulo-rumen was applied to wethers implanted with catheters in a mesenteric artery, the hepatic portal vein and the right ruminal vein. Portal blood flow was measured by downstream dilution of p-aminohippuric acid; the PDV uptake of arterial acetate, as well as the whole-body irreversible loss rate (ILR) of acetate, was estimated by [2-(13)C]acetate infusion into the right ruminal vein. The sheep were maintained with a bicarbonate-buffered solution of SCFA in the reticulo-rumen along with continuous intraruminal infusion of SCFA for 4 h. The portal appearance of SCFA of non-reticulo-ruminal origin was estimated before and after the infusion protocol. Of the acetate absorbed by the sheep, 89 (SE 5), 109 (SE 7) and 101 (SE 7)% was recovered as portal net appearance of acetate, portal net appearance of acetate corrected for PDV uptake of arterial acetate and increase in the ILR of acetate respectively. Of the propionate, isobutyrate, butyrate, isovalerate and valerate absorbed by the sheep, 95 (SE 7), 102 (SE 9), 23 (SE 3), 48 (SE 5) and 32 (SE 4)% respectively was recovered as portal net appearance. In contrast to current concepts, the present study showed that the reticulo-ruminal epithelium metabolizes none (or only a small proportion) of the acetate and propionate absorbed from the rumen. This observation could lead to the more efficient use of results obtained with multi-catheterized animals to quantify the net metabolite output of the rumen microbes.

Ruminal transport and metabolism of short-chain fatty acids (SCFA) in vitro: effect of SCFA chain length and pH

The unidirectional transport and metabolism of 14C-labeled acetate, propionate and butyrate across the isolated bovine rumen epithelium was measured in vitro by the Ussing chamber technique. There was a significant, but relatively small, net secretion of acetate and propionate, and a large and significant net absorption of butyrate. The results demonstrate that the mucosal–serosal (MS) pathway for short-chain fatty acids (SCFA) is different from the serosal–mucosal (SM) pathway, and that butyrate is treated differently from acetate and propionate by the epithelium. The results support that the main route for epithelial SCFA transport is transcellular. The correlation between SCFA lipophility and the flux rate was positive but weak at both pH 7.3 and 6.0. Decreasing pH increased all SCFA fluxes significantly, but not proportionally to the increase of protonized SCFA in the bathing solution. There was a significant and apparently non-competitive interaction between the transport of acetate, propionate and butyrate. It seems that mediated transport mechanisms must be involved in epithelial SCFA transport in the bovine rumen, but the data do not exclude that passive diffusion could account for a significant part of the flux. The metabolism of SCFA in the Ussing chamber system was considerable, and there was a clear preference for excretion of CO 2 from this metabolism to the mucosal side, while side preference for non-CO 2 metabolite excretion was not studied. Of the propionate and butyrate transported in the MS direction, 78 and 95% was metabolised, while only 37 and 38% was metabolised in the SM direction (acetate metabolism could not be measured). There was, however, no simple relation between the degree of metabolism and the transport rate or the transport asymmetry of the SCFA.

Metabolism of short-chain fatty acids by rat colonic mucosa in vivo.

To determine the influence of substrate concentration and substrate interactions on short-chain fatty acid metabolism in vivo, a surgical procedure was established. Rats were surgically operated to cannulate a 5-cm segment of proximal colon, isolate the vasculature, and cannulate the right colic vein draining this segment. Thus metabolism was restricted to the defined colonic segment. The appearance of total (14)C and (14)CO(2) in the mesenteric blood stabilized after 30 min of perfusion. Increasing luminal concentrations of butyrate from 2 to 40 mmol/l resulted in linear increases in total (14)C, but (14)CO(2) production from [(14)C]butyrate increased as a function of concentration only up to 10 mmol/l and was stable at higher butyrate concentrations. In addition to CO(2), 3-hydroxybutyrate and lactate were major metabolites of acetate and butyrate in vivo. The presence of a mixture of alternative substrates in the lumen had no influence on the metabolism of butyrate to CO(2) but significantly reduced the metabolism of acetate to CO(2). When compared with young (4 mo old) animals, transport of butyrate was significantly lower for aged (48 mo old) animals, as evidenced by the rate of appearance in blood of total (14)C (P = 0.04) and (14)C in butyrate (P = 0.03), but metabolism was similar, since differences were not significant for (14)C in the major metabolites 3-hydroxybutyrate (P = 0.06) and CO(2) (P = 0.17). These results show that important aspects of short-chain fatty acid transport and metabolism are not predicted from data using isolated colonocytes but require study using an in vivo model.

Physiological Effects of Resistant Starches on Fecal Bulk, Short Chain Fatty Acids, Blood Lipids and Glycemic Index

Objective: To assess the effects on fecal bulking, fecal short chain fatty acid (SCFA) production, blood lipids and glycemic indices of two different forms of resistant starch (RS2 and RS3) from a high-amylose cornstarch.Methods: Twenty-four healthy subjects (12 men; 12 women) consumed four supplements taken for 2 weeks in random order separated by 2-week washout periods. The supplements were a low-fiber (control) and supplements providing an additional 30 g dietary fiber as wheat bran (high-fiber control) or the equivalent amount of resistant starch analyzed gravimetrically as dietary fiber from RS2 or RS3. Four-day fecal collections and 12-hour breath gas collections were obtained at the end of each period. Fasting blood was taken at the beginning and end of each period. Glycemic indices of supplements were also assessed.Results: The wheat bran supplement increased fecal bulk 96±14 g/day compared with the low-fiber control (p<0.001) with the mean for both resistant starches also being greater (22±8 g/day) than the low-fiber control (p=0.013). On the resistant starch phases, the mean fecal butyrate:SCFA ratio, which has been suggested to have positive implications for colonic health, was significantly above the low-fiber control by 31±14% (p=0.035). Resistant starches did not alter serum lipids, urea or breath H2 or CH4. No significant differences in glycemic index were seen between the RS and control supplements.Conclusion: The potential physiological benefits of the resistant starches studied appear to relate to colonic health in terms of effects on fecal bulk and SCFA metabolism.

In vivo butyrate metabolism and colonic permeability in extensive ulcerative colitis

Background & Aims: Impaired short-chain fatty acid metabolism by the colonocyte has been suggested as a pathogenic factor in ulcerative colitis (UC). The aim of this study was to measure in vivo butyrate metabolism in UC and to correlate butyrate oxidation with colonic permeability. Methods: Butyrate oxidation was measured by means of a 14CO 2-breath test after rectal instillation of 14C-butyrate. 51Cr-ethylenediaminetetraacetic acid (EDTA) was added to the enema, and the urinary % dose excretion of 51Cr-EDTA after 6 hours was a measure for permeability. Results: Patients with active extensive UC showed a significantly lower butyrate oxidation and increased colonic permeability in comparison to healthy controls. Butyrate oxidation correlated significantly negative with clinical activity. Oxidation of butyrate was not decreased in most patients with inactive extensive UC. In 3 patients with inactive disease and decreased oxidation, a relapse occurred within a few weeks after the test, whereas all patients with normal oxidation maintained their remission for at least 3 months. A significantly negative correlation existed between butyrate oxidation and colonic permeability. Conclusions: Patients with active extensive UC have a decreased colonic butyrate oxidation. However, the fact that remission is associated with normal oxidation suggests that UC mucosa is not intrinsically altered in butyrate oxidation, making this unlikely to be a primary defect in UC. GASTROENTEROLOGY 1998;115:584-590

In vitro Studies on Transport and Metabolism of Short-Chain Fatty Acids in Pig Hindgut

An analytical method based on alkaline freeze drying, ultracentrifugation, and quantitative gas chromatography was established to differentiate between mucosal uptake, tissue accumulation, and serosal release of SCFA in pig hindgut. It was shown that serosal release of SCFA was substantially lower than mucosal uptake and tissue accumulation, indicating substantial degradation and/or metabolism during transepithelial movement.

Histological changes in the colonic mucosa following irrigation with short-chain fatty acids

Short-chain fatty acids (SCFAs) derived from bacterial fermentation of complex carbohydrates are preferred luminal nutrients of the colonic mucosa. Starvation of colonocytes through lack or impaired metabolism of luminal SCFAs may be a cofactor in the pathogenesis of ulcerative colitis. A detailed histological evaluation of colonic biopsy specimens was performed in patients with active distal ulcerative colitis who were treated with rectal enemas containing a mixture of SCFAs, n-butyrate alone or saline placebo. Together with light microscopic parameters of mucosal inflammation, the pattern of crypt cell proliferation (proliferating cell nuclear antigen) and the mucosal activity of factor XIII were assessed. Butyrate reduced the density of polymorphonuclear leucocytes in the lamina propria (4 weeks: P = 0.063; 8 weeks: P = 0.091); other inflammatory parameters remained unchanged. Both butyrate and the SCFA mixture reduced significantly the number of proliferating cells in the upper 40% of crypts. Tissue factor XIII activity in active ulcerative colitis was significantly lower than in mucosa from normal colons; however, it was not affected by SCFA or butyrate irrigation. SCFAs and butyrate have a more marked effect on crypt cell proliferation than on parameters of inflammation in patients with active ulcerative colitis.

Non-ionic diffusion and carrier-mediated transport drive extracellullar pH regulation of mouse colonic crypts.

1. Extracellular pH (pHo) regulation within mouse colonic crypt lumens is stimulated by transepithelial gradients of short-chain fatty acids (SCFAs). Current work assesses underlying mechanisms contributing to pHo regulation. 2. Crypt luminal alkalinization was saturable by apical SCFA (substrate concentration activating half-maximal transport (KT) of isobutyrate = 45 mM). However, saturation was consistent with either carrier-mediated SCFA flux or non-ionic diffusion, because the non-ionized form was titrated by luminal alkalinization. Direct acidification of apical perfusates increased the magnitude of SCFA-induced luminal alkalinization, roughly in the same proportion to the increased concentration of non-ionized SCFA in the crypt lumen. 3. Transepithelial gradients of an alternative weak acid (CO2) produce pHo changes similar to SCFA. In contrast, a weak base (NH3) changes pHo with reverse dependence on the orientation of the transepithelial gradient compared with SCFA. Results implicate non-ionic diffusion in pHo regulation, and suggest that pHo changes may underly SCFA-stimulated bicarbonate secretion and ammonium absorption. 4. SCFA metabolism plays a minor role in extracellular pH regulation. An avidly metabolized SCFA (N-butyrate) augments crypt luminal alkalinization only slightly (0.08 pH units) versus a poorly metabolized SCFA (isobutyrate). 5. Apical addition of 1 mM 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) partially inhibits luminal alkalinization caused by apical SCFA. DIDS has no effect on luminal alkalinization caused by transepithelial CO2 gradients. Probenecid (1 mM), alpha-cyano-4-hydroxycinnamic acid (4 mM) or basolateral DIDS (1 mM) do not affect pHo regulation. Results suggest that DIDS-sensitive, SCFA-dependent transport in the colonocyte apical membrane contributes to pHo regulation.