Structure-dependent and receptor-independent increase in osmotic fragility of rat erythrocytes by short-chain fatty acids

We examined short-chain fatty acids (SCFAs) with 1 (C1) to 5 (C5) carbon atoms for osmotic fragility (OF) in isolated red blood cells (RBCs) in rats. The RBCs were used as prototypical plasma membrane model. The dense packed RBC was incubated in a phosphate-NaCl buffer solution containing each SCFA at 0 to 100 mM. The RBC suspensions were transferred into the OF test tubes containing NaCl from 0.2 to 0.9%. The hemoglobin concentration was determined and the EC50 in hemolysis was calculated. The OF in RBCs was dose-dependently increased by exposure to SCFAs, except for C1, with an increasing number of carbon atoms. Branched-chain fatty acids (isomers of C4 and C5) have a smaller effect on OF than straight-chain fatty acids (C4 and C5). The SCFA-induced increases in OF were not affected by pretreatment of RBCs with trypsin. The response of the RBC membrane to SCFAs depends on their concentration, carbon chain length and chain structure (straight or branched). The SCFAs probably disturb the lipid bilayer of the RBC membrane and result in a decrease in osmotic resistance. The plasma membrane in rat RBCs could respond to the structure of the SCFAs in detail by using the OF as an indicator.     

Suppressive effect of short-chain fatty acids on production of proinflammatory mediators by neutrophils

Short chain fatty acids (SCFAs) are fermentation products of anaerobic bacteria. More than just being an important energy source for intestinal epithelial cells, these compounds are modulators of leukocyte function and potential targets for the development of new drugs. The aim of this study was to evaluate the effects of SCFAs (acetate, propionate and butyrate) on production of nitric oxide (NO) and proinflammatory cytokines [tumor necrosis factor α (TNF-α) and cytokine-induced neutrophil chemoattractant-2 (CINC-2αβ)] by rat neutrophils. The involvement of nuclear factor κB (NF-κB) and histone deacetylase (HDAC) was examined. The effect of butyrate was also investigated in vivo after oral administration of tributyrin (a pro-drug of butyrate). Propionate and butyrate diminished TNF-α, CINC-2αβ and NO production by LPS-stimulated neutrophils. We also observed that these fatty acids inhibit HDAC activity and NF-κB activation, which might be involved in the attenuation of the LPS response. Products of cyclooxygenase and 5-lipoxygenase are not involved in the effects of SCFAs as indicated by the results obtained with the inhibitors of these enzymes. The recruitment of neutrophils to the peritonium after intraperitoneal administration of a glycogen solution (1%) and the ex vivo production of cytokines and NO by neutrophils were attenuated in rats that previously received tributyrin. These results argue that this triglyceride may be effective in the treatment of inflammatory conditions.     

Influence of short-chain fatty acids and osmolality on mucin release in the rat colon

Summary The influence of short-chain fatty acids (SCFA) and osmolality on mucin release in the rat colon was studied histochemically by determining number of stained mucin-containing cells. SCFA did not significantly influence the number of cells staining for mucin. Hypertonic solutions (360 mosm/l) did not affect mucin release in the proximal colon, but stimulated mucin release in the distal colon. Solutions of lower osmolality (300 or 250 mosm/l) caused a considerable release of mucin from goblet cells as well as vacuolated cells in both the proximal and the distal colon; the lower the osmolality, the more mucin was released. The mucosa of the distal colon was conspicuously affected by solutions of lower osmolality. The influence of osmolality on mucin release was entirely local.Supported by a grant from the Deutsche Forschungsgemeinschaft (En 65/9)The authors wish to thank Prof. H. Höller and G. Rechkemmer for critical advice and Miss G. Becker for technical assistanceA preliminary portion of this study was presented at the 3rd Meeting of the European Intestinal Transport Group, Southampton, 21.–23. April, 1980     

Unidirectional fluxes of short-chain fatty acids across segments of the large intestine in pig,sheep and pony compared with guinea pig

Unidirectional fluxes of short-chain fatty acids across pig, sheep and pony caecum, proximal and distal colon were studied under short-circuit current conditions in Ussing chambers. Findings are compared with results from guinea pig. Marked species differences are apparent; highest mucosal-to-serosal fluxes of acetate, propionate and butyrate were seen in guinea pig, lower values in pig and smallest fluxes in sheep and pony. Segmental differences between caecum, proximal and distal colon exist mainly in guinea pig and are less developed in pig, sheep and pony. Inhibition of Na⁺/H⁺ exchange by amiloride added to the mucosal solution decreased the mucosal-to-serosal fluxes of short-chain fatty acids clearly in guinea pig caecum and proximal colon, and very little in distal colon. This effect was somewhat less pronounced in pig caecum and distal colon, in caecum and distal colon of sheep and caecum of the pony. In pig, sheep and pony proximal colon and pony distal colon no significant inhibition was observed. Inhibition of the K⁺-H⁺ ATPase by addition of ouabain to the mucosal solution diminished mucosal-to-serosal fluxes of short-chain fatty acids in the guinea pig distal colon extensively. No comparable inhibition was seen in any of the other segments in the animals studied.     

Acetylation of the Entamoeba histone H4 N-terminal domain is influenced by short-chain fatty acids that enter trophozoites in a pH-dependent manner

Treatment of higher eukaryotic cells with short-chain fatty acids (SCFA) such as butyrate causes decreased levels of histone deacetylase (HDAC) activity and hyperacetylation of histones, and thereby affects gene expression, cell growth and differentiation. Entamoeba parasites encounter high levels of SCFA in the host colon, and in vitro these compounds allow trophozoite stage parasites to multiply but prevent their differentiation into infectious cysts. The Entamoeba invadens IP-1 histone H4 protein has an unusual number of lysines in its N-terminus, and these become hyperacetylated in trophozoites exposed to the HDAC inhibitors trichostatin A (TSA) or HC-toxin, but not in trophozoites exposed to butyrate. We have now found that several other commonly studied isolates of Entamoeba parasites also have an extended set of histone H4 acetylation sites that become hyperacetylated in response to TSA, but hypoacetylated in response to butyrate, suggesting an unusual sensitivity of this parasite's histone modifying enzymes to SCFA. Butyrate was found to enter trophozoites in a pH-dependent manner consistent with diffusive entry of the un-ionised form of the fatty acid into the amoebae. Transit of the Entamoeba organism through areas of the host intestine with distinct pH and SCFA concentrations would therefore result in very different levels of SCFA within the parasite. Entamoeba appears to have acquired unique alterations of its histone acetylation mechanism that may allow for its growth in the presence of varying amounts of the bacterial fermentation products.     

Oxidation of short-chain fatty acids by sulfate-reducing bacteria in freshwater and in marine sediments

Colony counts of acetate-, propionate- and l-lactate-oxidizing sulfate-reducing bacteria in marine sediments were made. The vertical distribution of these organisms were equal for the three types considered. The highest numbers were found just beneath the border of aerobic and anaerobic layers.Anaerobic mineralization of acetate, propionate and l-lactate was studied in the presence and in the absence of sulfate. In freshwater and in marine sediments, acetate and propionate were oxidized completely with concomitant reduction of sulfate. l-Lactate was always fermented. Lactate-oxidizing, sulfate-reducing bacteria, belonging to the species Desulfovibrio desulfuricans, and lactate-fermenting bacteria were found in approximately equal amounts in the sediments. Acetate-oxidizing, sulfate-reducing bacteria could only be isolated from marine sediments, they belonged to the genus Desulfobacter and oxidized only acetate and ethanol by sulfate reduction. Propionate-oxidizing, sulfate-reducing bacteria belonged to the genus Desulfobulbus. They were isolated from freshwater as well as from marine sediments and showed a relatively large range of usable substrates: hydrogen, formate, propionate, l-lactate and ethanol were oxidized with concomitant sulfate reduction. l-Lactate and pyruvate could be fermented by most of the isolated strains.     

Effect of organic loading rate on VFA production, organic matter removal and microbial activity of a two-stage thermophilic anaerobic membrane bioreactor

This study focused on the VFA (volatile fatty acid) profile variation with organic loading rate (OLR) of a two stage thermophilic anaerobic membrane bioreactor (TAnMBR). The two stage TAnMBR treating high strength molasses-based synthetic wastewater was operated under a side-stream partial sedimentation mode at 55 °C. Reactor performances were studied at different OLR ranging from 5 to 12 kg COD m⁻³ d⁻¹. Operational performance of TAnMBR was monitored by assessing biological activity, organic removal efficiency, and VFA. The major intermediate products of anaerobic digestion were identified as acetate, propionate, iso-butyrate, n-butyrate and valerate. Among them acetate and n-butyrate were identified as the most abundant components. Increase of OLR changes the predominant VFA type from acetic acid to n-butyric acid and the total VFA concentration was increased with increased OLR. Moreover, increased OLR increased organic removal efficiency up to second loading rate and dropped in third loading rate while biological activity was increased continuously.     

Use of Gifu Anaerobic Medium for culturing 32 dominant species of human gut microbes and its evaluation based on short-chain fatty acids fermentation profiles

Recently, a “human gut microbial gene catalogue,” which ranks the dominance of microbe genus/species in human fecal samples, was published. Most of the bacteria ranked in the catalog are currently publicly available; however, the growth media recommended by the distributors vary among species, hampering physiological comparisons among the bacteria. To address this problem, we evaluated Gifu anaerobic medium (GAM) as a standard medium. Forty-four publicly available species of the top 56 species listed in the “human gut microbial gene catalogue” were cultured in GAM, and out of these, 32 (72%) were successfully cultured. Short-chain fatty acids from the bacterial culture supernatants were then quantified, and bacterial metabolic pathways were predicted based on in silico genomic sequence analysis. Our system provides a useful platform for assessing growth properties and analyzing metabolites of dominant human gut bacteria grown in GAM and supplemented with compounds of interest.     

The effect of volatile fatty acids on the inactivation of <Emphasis Type="Italic">Clostridium perfringens</Emphasis> in anaerobic digestion

The application of sludge digestion systems to remove pathogens has been employed to generate biosolids suitable for reuse in agriculture. Traditionally, temperature is considered the principal agent responsible for pathogen reduction in anaerobic digestion. However, other substances such as volatile fatty acids may also have an antimicrobial effect. The objective of this study was to assess the impact of fatty acid mixtures on the inactivation of C. perfringens over a range of digestion temperatures. An equimolar mixture of acetic acid, propionic acid and butyric acid was applied to digester effluent for a period of 24 h at temperatures of 35 °C, 42 °C, 49 °C and 55 °C. C. perfringens inactivation in digester effluents, when dosed with volatile organic acids, was found to depend on pH, acid concentration and temperature. Temperatures above 55 °C appeared to increase the inhibitory effects of the organic acids at higher concentrations. An interaction between temperature and pH on survival of C. perfringens was observed. The results suggest that high concentrations of organic acids at a pH value of 4.5–5.5 during thermophilic digestion substantially reduce concentrations of C. perfringens in municipal sludge.     

Effect of short-chain fatty acids on the ethylene pathway in embryonic axes of Cicer arietinum during germination

The effect of short-chain saturated fatty acids (C₅–C₁₀) on the biosynthesis of ethylene in embryonic axes of chick-pea ( Cicer arietinum L.) seeds was investigated. The emergence of radicle and fresh weight of embryonic axes diminished with increasing number of carbons. The inhibition of germination caused by lower concentrations (1 m M ) of fatty acids (C₅–C₁₀) was partially reversed by exogenous 1-aminocyclopropane-1-carboxylic acid (ACC), whereas exogenous ethylene was able to overcome the inhibitory effect provoked by all concentrations (1–5 m M ) of applied fatty acids (C₅–C₁₀). Ethylene production rates, and enzyme activities of ACC synthase and ACC oxidase decreased concomitantly with the molecular mass and increasing concentration of fatty acids. The inhibitory effect of these acids on ethylene production seems to result not only from a decreased ACC synthesis, but also from an enhancement of 1-malonylamino)cyclopropane-1-carboxylic acid (MACC) synthesis.     

Effect of redox mediator, AQDS, on the decolourisation of a reactive azo dye containing triazine group in a thermophilic anaerobic EGSB reactor

The feasibility of thermophilic (55 °C) anaerobic treatment applied to colour removal of a triazine contained reactive azo dye was investigated in two 0.53 l expanded granular sludge blanket (EGSB) reactors in parallel at a hydraulic retention time (HRT) of 10 h. Generally, this group of azo dyes shows the lowest decolourisation rates during mesophilic anaerobic treatment. The impact of the redox mediator addition on colour removal rates was also evaluated. Reactive Red 2 (RR2) and anthraquinone-2,6-disulfonate (AQDS) were selected as model compounds for azo dye and redox mediator, respectively. The reactors achieved excellent colour removal efficiencies with a high stability, even when high loading rates of RR2 were applied (2.7 g RR2 l⁻¹ per day). Although AQDS addition at catalytic concentrations improved the decolourisation rates, the impact of AQDS on colour removal was less apparent than expected. Results show that the AQDS-free reactor R2 achieved excellent colour removal rates with efficiencies around 91%, compared with the efficiencies around 95% for the AQDS-supplied reactor R1. Batch experiments confirmed that the decolourisation rates were co-substrate dependent, in which the volatile fatty acids (VFA) mixture was the least efficient co-substrate. The highest decolourisation rate was achieved in the presence of either hydrogen or formate, although the presence of glucose had a significant impact on the colour removal rates.     

Syntrophus aciditrophicus sp. nov., a new anaerobic bacterium that degrades fatty acids and benzoate in syntrophic association with hydrogen-using microorganisms

Strain SB^T is a new, strictly anaerobic, gram-negative, nonmotile, non-sporeforming, rod-shaped bacterium that degrades benzoate and certain fatty acids in syntrophic association with hydrogen/formate-using microorganisms. Strain SB^T produced approximately 3 mol of acetate and 0.6 mol of methane per mol of benzoate in coculture with Methanospirillum hungatei strain JF1. Saturated fatty acids, some unsaturated fatty acids, and methyl esters of butyrate and hexanoate also supported growth of strain SB^T in coculture with Desulfovibrio strain G11. Strain SB^T grew in pure culture with crotonate, producing acetate, butyrate, caproate, and hydrogen. The molar growth yield was 17 ± 1 g cell dry mass per mol of crotonate. Strain SB^T did not grow with fumarate, iron(III), polysulfide, or oxyanions of sulfur or nitrogen as electron acceptors with benzoate as the electron donor. The DNA base composition of strain SB^T was 43.1 mol% G+C. Analysis of the 16 S rRNA gene sequence placed strain SB^T in the δ-subdivision of the Proteobacteria, with sulfate-reducing bacteria. Strain SB^T was most closely related to members of the genus Syntrophus. The clear phenotypic and genotypic differences between strain SB^T and the two described species in the genus Syntrophus justify the formation of a new species, Syntrophus aciditrophicus. Received: 2 June 1998 / Accepted: 16 November 1998     

Evaluation of intestinal microbiota,short-chain fatty acids,and immunoglobulin a in diversion colitis

It is reported that an increase in aerobic bacteria, a lack of short-chain fatty acids (SCFAs), and immune disorders in the diverted colon are major causes of diversion colitis. However, the precise pathogenesis of this condition remains unclear. The aim of the present study was to examine the microbiota, intestinal SCFAs, and immunoglobulin A (IgA) in the diverted colon. Eight patients underwent operative procedures for colostomies. We assessed the diverted colon using endoscopy and obtained intestinal samples from the diverted colon and oral colon in these patients. We analyzed the microbiota and SCFAs of the intestinal samples. The bacterial communities were investigated using a 16S rRNA gene sequencing method. The microbiota demonstrated a change in the proportion of some species, especially Lactobacillus, which significantly decreased in the diverted colon at the genus level. We also showed that intestinal SCFA values were significantly decreased in the diverted colon. Furthermore, intestinal IgA levels were significantly increased in the diverted colon. This study was the first to show that intestinal SCFAs were significantly decreased and intestinal IgA was significantly increased in the diverted colon. Our data suggest that SCFAs affect the microbiota and may play an immunological role in diversion colitis.     

Trichinella spiralis: the influence of short chain fatty acids on the proliferation of lymphocytes, the goblet cell count and apoptosis in the mouse intestine

This study was carried out to determine the influence of short chain fatty acids (SCFA) on spleen and mesenteric lymph node lymphocyte proliferation, goblet cells and apoptosis in the mouse small intestine during invasion by Trichinella spiralis. BALB/c mice were infected with 250 larvae of T. spiralis. An SCFA water solution containing acetic, propionic and butyric acids (30:15:20 mM) was administered orally starting 5 days before infection and ending 20 days post infection (dpi). Fragments of the jejunum were collected by dissection 7 and 10 dpi, and were examined for apoptotic cells in the lamina propria of the intestinal mucosa, and for goblet cells. The proliferation index of the cultured spleen and mesenteric lymph node lymphocytes with MTT test was also determined. The orally administered SCFA solution decreased the proliferation of mesenteric lymph node lymphocytes in the mice infected with T. spiralis at both examination times, but did not influence the proliferative activity of the spleen cells. Seven dpi, both in the spleen and mesenteric lymph nodes, the highest proliferation index of concanavalin A (Con A)-stimulated lymphocytes was found in the group of uninfected animals receiving SCFA animals. This tendency could still be seen 10 dpi in the mesenteric lymph nodes but not in the spleen, where the proliferation index in this group had significantly decreased. In vitro studies revealed, that butyric and propionic acids added to the cell cultures suppressed the proliferation of Con A-stimulated mesenteric lymph nodes and spleen lymphocytes taken from uninfected and T. spiralis-infected mice. Acetic acid stimulated proliferation of splenocytes taken from uninfected mice but did not affect lymphocyte proliferation in mesenteric lymph nodes from uninfected or infected mice. Orally administered SCFA increased the number of goblet cells found in the epithelium of the jejunum 7 dpi, but this number had decreased 10 dpi. The number of apoptotic cells in the lamina propria of the intestinal mucosa of animals infected with the T. spiralis and receiving SCFA was also lower, particularly 10 dpi. The above results show that SCFA can participate in the immune response during the course of trichinellosis in mice.     

Characterization of a fatty acid binding protein from the swimming crab Portunus trituberculatus and its effects on the composition of fatty acids in different tissues

Fatty acid binding proteins (FABPs) belonging to the lipocalin protein family, have an affinity for fatty acids (FAs) and participate in both the transport of FAs and lipid metabolism. In this study, a 909-bp full-length pt-FABP cDNA containing a 411-bp open reading frame (ORF) was obtained from the swimming crab Portunus trituberculatus. Quantitative real-time PCR revealed that during larval development, pt-FABP expression increased gradually from the zoea to the megalopa, and decreased in the juvenile crab. pt-FABP mRNA was detected at the highest level in the testis, followed by the muscles, with the lowest level in the hepatopancreas. The composition of fatty acids in three different tissues from crabs of different sex was detected by gas chromatography–mass spectrometry (GC-MS). The results indicated that the FA profile was in accordance with the expression of pt-FABP, which suggested that pt-FABP participates in the unsaturated fatty acid transportation and storage system of P. trituberculatus.     

The effect of organic loading rate and retention time on hydrogen production from a methanogenic CSTR

The possibility of shifting a methanogenic process for hydrogen production by changing the process parameters viz., organic loading rate (OLR) and hydraulic retention time (HRT) was evaluated. At first, two parallel semi-continuously fed continuously stirred tank reactors (CSTR) were operated as methanogenic reactors (M1 and M2) for 78 days. Results showed that a methane yield of 198-218 L/kg volatile solids fed (VS(fed)) was obtained when fed with grass silage at an OLR of 2 kgVS/m3/d and HRT of 30 days. After 78 days of operation, hydrogen production was induced in M2 by increasing the OLR from 2 to 10 kgVS/m3/d and shortening the HRT from 30 to 6 days. The highest H? yield of 42 L/kgVS(fed) was obtained with a maximum H? content of 24%. The present results thus demonstrate that methanogenic process can be shifted towards hydrogen production by increasing the OLR and decreasing HRT.     

Activation of UCPs gene expression in skeletal muscle can be independent on both circulating fatty acids and food intake. Involvement of ROS in a model of mouse cancer cachexia

Implantation of a fast growing tumour to mice (Lewis lung carcinoma) resulted in a clear cachectic state characterized by a profound muscle wasting. This was accompanied by a significant increase in both UCP2 and UCP3 gene expression in skeletal muscle and heart. Interestingly, this increase in gene expression was not linked to a rise in circulating fatty acids or in a decrease in food intake, as previously reported in other pathophysiological states. These results question the concept that hyperlipaemia is the only factor controlling UCP gene expression in different pathophysiological conditions. In addition, the present work suggests that UCPs might participate in a counter-regulatory mechanism to lower the production of ROS.