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Insights into specific and nonspecific butyrate-producing pathways during the in vitro fecal fermentation of butyrylated starch.

Butyrylated starch is a special type-4 resistant starch with butyrate-carrying attribute. In this study, the unique butyrate-producing capability of butyrylated starch was deeply investigated by focusing on its specific and nonspecific butyrate-producing pathways, respectively, using specially designed substrates as controls. In vitro fermentation studies revealed that butyrylated and isobutyrylated starches generated high levels of butyrate and isobutyrate, respectively, highlighting the role of butyryl group metabolism in the specificity of butyrate production. Carboxylesterase assays have demonstrated that butyryl group metabolism is primarily facilitated by carbohydrate esterases expressed in the gut microbiota. Combined with 16S rRNA sequencing and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, it was found that butyrylated starch fermentation did not significantly enhance traditional butyrate synthesis pathways but modified the balance between the butyryl-CoA:acetyl-CoA transferase and butyrate kinase pathways by altering the gut microbiota composition, specifically by upregulating the relative abundance of indicator species such as Bacteroides, the Lachnospiraceae_NK4A136_group, and Parabacteroides. These insights offer theoretical guidance for designing butyrylated starch structures and regulating intestinal health.

Starch

[A new butyric acid-producing bacteroides species: B. splanchnicus n. sp. (author's transl)].

Three butyric acid-producing saccharolytic Bacteroides cultures (1651/6, BM 158, and IPP 3751) were described by WERNER and REICHERTZ in 1971 (Zbl.Bakt.Hyg., I. Abt. Orig. A 217,206-216). Since then, 6 strains closely resembling 1651/6 were isolated from stool specimens and surgically removed appendices. In the present communication, strains 1651/6, S2/34, S3/38, S4/28, S6/6, A5/2 are described as members of a new species, Bacteroides splanchnicus n.sp. The strains were morphologically very similar (Gram negative non-sporing non-motile rods, 1-2.5 mu in length and 0.7 mu in width) and fermented glucose, fructose, galactose, mannose, lactose, and arabinose (pH values of 4.6-5.4, moderate gas formation). Negative reactions (pH values of 5.8-7.2) were observed with 20 other carbohydrates. The strains were positive in the glutamic acid decarboxylase test and formed indole and H2S. In peptone-yeast extract broth and peptone-yeast extract-glucose broth acetic, propionic, isobutyric, butyric, and isovaleric acids were produced. Washed cells of strains 1651/6 and S4/28 incubated anaerobically in sterile solutions of single amino acids produced butyrate from lysine only. Abundant butyric acid was also produced from glucose. The in vitro activity of 15 antibiotics on 5 strains was studied by broth dilution tests. Uniformly, the strains showed resistance to aminoglycosides and polymyxins (MIC values, 60-500 mug/ml) and susceptibility to tetracyclines, lincomycin, clindamycin, rifampicin, and erythromycin (MIC values, 0.05-0.5 mug/ml). Chloramphenicol, penicillins, and cephalosporins showed bacteriostatic activity at concentrations of 5-40 mug/ml. The serological behaviour of 5 strains was studied in cross-agglutination and gel-diffusion experiments. Cross-reactivity was pronounced in gel-diffusion tests using rabbit antisera and autoclaved extracts and extracts prepared by repeated deep-freezing and thawing of whole cell suspensions as antigens. However, antisera against the B. splanchnicus strains did not react with antigens of B. fragilis 6869, B. thetaiotaomicron AS 126, B. vulgatus AM 45 a, B. distasonis AII 104 and butyric acid-producing strains BM 158 and IPP 3751 in agglutination and gel-diffusion experiments, and there were no cross-reactions between B. splanchnicus antigens and antisera against B. fragilis 6869 and the other aforementioned strains. The new species has been confirmed by members of the ICSB Taxonomic Subcommittee for Gram negative anaerobic rods. Strains 1651/6 and S4/28 have been deposited in The National Collection of Type Cultures, London (NCTC numbers 10825 and 10826). Strain NCTC 10825 (= 1651/6) is the type of the species, B. splanchnicus.

Acetates

Effect of sodium butyrate on mammalian cells in culture: a review.

Sodium butyrate produces reversible changes in morphology, growth rate, and enzyme activities of several mammalian cell types in culture. Some of these changes are similar to those produced by agents which increase the intracellular level of adenosine 3',5'-cyclic monophosphate (cAMP) or by analogs of cAMP. Sodium butyrate increases the intracellular level of cAMP by about two fold in neuroblastoma cells; therefore, some of the effects of sodium butyrate on these cells may in part be mediated by cAMP. Sodium butyrate appears to have properties of a good chemotherapeutic agent for neuroblastoma tumors because the treatment of neuroblastoma cells in culture causes cell death and "differentiation"; however, it is either innocuous or produces reversible morphological and biochemical alterations in other cell types.

3',5'-Cyclic-AMP Phosphodiesterases

Enzymology of butyrate formation by Butyrivibrio fibrisolvens.

Butyrivibrio fibrisolvens is a major butyrate-forming species in the bovine and ovine rumen. The enzymology of butyrate formation from pyruvate was investigated in cell-free extracts of B. fibrisolvens D1. Pyruvate owas oxidized to acetylcoenzyme A (CoA) in the presence of CoA.SH and benzyl viologen or flavin nucleotides. The bacterium uses thiolase, beta-hydroxybutyryl-CoA dehydrogenase, crotonase, and crotonyl-CoA reductase to form butyryl-CoA from acetyl-CoA. Reduction of acetoacetyl-CoA to beta-hydroxybutyryl-CoA was faster with NADH than with NADPH. Crotonyl-CoA was reduced to butyryl-CoA by NADH, but not by NADPH, only in the presence of flavin nucleotides. Reduction of flavin nucleotides by NADH was much slower than the flavin-dependent reduction of crotonyl-CoA. This indicates that flavoproteins rather than free flavin participated in the reduction of crotonyl-CoA. Butyryl-CoA was converted to butyrate by phosphate butyryl transferase and butyrate kinase.

3-Hydroxyacyl CoA Dehydrogenases

The actions of cyclic AMP, its butyryl derivatives and Na butyrate on the proliferation of malignant trophoblast cells in vitro.

Cyclic AMP, and its derivatives N6-monobutyryl cyclic AMP and dibutyryl cyclic AMP, have been found to inhibit the proliferation of trophoblast cells of the BeWo cell line in vitro. Sodium butyrate (1 mM), a possible degradation product of the butyrate derivatives, also inhibited cell proliferation, giving similar growth rates to equimolar dibutyryl cyclic AMP. The inhibition by butyrate was however, not sufficient to account for the action of 1 mM N6-monobutyryl cycli AMP, which, like cyclic AMP, completely inhibited cell proliferation. The potency, specificity and toxicity of the substances were compared. The results suggest different modes of action for cyclic AMP and dibutyryl cyclic AMP.

Bucladesine

Regulation of growth and morphological modulation of HeLa65 cells in monolayer culture by dibutyryl cyclic AMP, butyrate and their analogs.

N6-O2'-Dibutyryl adenosine-3',5' monophosphate (DBcAMP) markedly altered the morphology of HeLa cells by increasing average cell size with an increase in total cell protein and RNA. Such effects were not caused by adenosine 3',5' monophosphate (cAMP) or related nucleosides and nucleotides. Butyrate, an enzyme catalyzed hydrolysis product of DBcAMP, induced a jagged spindle shape in HeLa cells within 8 hours and then caused them to enlarge and resemble those grown with DBcAMP. These effects were specific for butyrate (C4) and pentanoate (C5) and were not observed with isomers, substituted analogs, or other fatty acid derivatives. These morphological effects were prevented by blocking protein synthesis or by altering the cytoskeleton with Colcemide or cytochalasin B.

Adenosine Triphosphate

Contact allergy to hydrocortisone 17-butyrate.

Two female patients with stasis dermatitis developed allergic contact dermatitis to hydrocortisone 17-butyrate cream. Patch tests with hydrocortisone 17-butyrate were positive, but not with the vehicle. One patient was also allergic to two other commerical corticosteroids, but patch tests revealed positive reactions only to ingredients of the vehicles. Attention is drawn to the frequency of contact allergy to corticosteroids in patients with stasis dermatitis.

Dermatitis, Contact

Comparison of desoximetasone and hydrocortisone butyrate in psoriasis.

Thirty psoriatics were treated for 2 weeks on a double-blind controlled basis with desoximetasone (0.25%) and with hydrocortisone butyrate (0.1%). It was a randomised left-right comparative trial. Thirteen out of 27 patients preferred desoximetasone, 3 patients preferred hydrocortisone butyrate. There was also a significantly better effect of desoximetasone as judged by the observer after the second week of treatment.

Administration, Topical

Intestinal organoid screen reveals that Bacillus velezensis PGM541 promotes epithelial proliferation via its metabolite butyric acid.

BACKGROUND: Probiotics have been widely used for the regulation of intestinal health. Current screening methods for probiotics typically rely on animal or two-dimensional cell models. In this study, we employed intestinal organoids to identify a candidate probiotic strain. Furthermore, we investigated the potential mechanisms through which this strain and its active metabolites exert their effects, thereby evaluating the efficacy of this screening approach. RESULTS: Firstly, candidate probiotic strain PGM541 was identified from a porcine-derived Bacillus library by assessing organoid viability. Subsequently, to validate the organoid screening reliability, the potential mechanism of strain PGM541 on the intestinal epithelium was investigated; it was found to exhibit probiotic functions by regulating cell proliferation in both in vitro organoid and in vivo piglet models. Furthermore, organoid screening combined with metabolomic analysis identified butyric acid (BA) as the key bioactive metabolite responsible for driving epithelial proliferation. Whole-genome and transcriptomic analyses revealed the biosynthetic pathway of BA in strain PGM541. Importantly, BA receptor blockade experiments directly confirmed that BA enhances epithelial proliferation via interaction with the FFAR2 receptor, thereby validating its functional activity. Additionally, strain PGM541 exhibited protective effects against dextran sulfate sodium (DSS)-induced colitis, further validating the effectiveness of the intestinal organoid platform for probiotic screening. CONCLUSIONS: The probiotic strain PGM541, which was screened using intestinal organoids, promotes intestinal epithelial cell proliferation via its metabolite BA activating the FFAR2 receptor. These findings demonstrate that the intestinal organoid model serves as an effective platform for both preliminary probiotic screening and mechanistic investigation. Video Abstract.

Animals

Regulation of the induction of alkaline phosphatase in choriocarcinoma cells by sodium butyrate.

Alkaline phosphatase is induced in human choriocarcinoma cells by short-chain fatty acids, especially sodium butyrate. This fatty acid increases the phosphatase activity immediately and in a nearly linear fashion. Only phosphatase with an alkaline pH optimum is induced. Both the induced alkaline phosphatase and the basal enzyme are precipitated by antiserum against term-placental alkaline phosphatase, but the choriocarcinoma phosphatase is less stable to heating than is the term-placental enzyme. The induction of alkaline phosphatase activity requires cellular synthesis of protein, RNA and DNA. The regulation of induction probably occurs at the transcriptional level.

Alkaline Phosphatase

Simple butyrate esterase stain for monocytes.

The esterases used to identify monocytes are best demonstrated using alpha-naphthyl butyrate as substrate. However, the reagents commonly used for this stain are time-consuming to prepare and are unstable. This report describes a quick, easy, and reproducible staining method using stable reagents which are readily available commercially but which may also be prepared in the laboratory.

Butyrates

Utlization of D-3-hydroxy[3-14C]butyrate for lipogenesis in vivo in lactating rat mammary gland.

Incorporation of D-3-hydroxy[3-14C]butyrate into lipid in vivo suggests that lactating mammary gland is a major site of ketone-body utilization. The incorporation decreases in short-term insulin deficiency (2h) and on starvation (24h), but increases again on refeeding (2h). The activity of cytosolic acetoacetyl-CoA synthetase parallels the changes in nutritional state, but is not affected by short-term insulin deficiency.

Acetyl-CoA C-Acetyltransferase

Comparative study of triamcinolone acetonide and hydrocortisone 17-butyrate in rosacea with special regard to the rebound phenomenon.

The clinical efficacy and the rebound phenomenon were studied in a left-right double-blind trial comparing triamcinolone acetonide (TA) and hydrocortisone 17-butyrate (HC 17-B, Locoid). The trial comprised 19 patients with rosacea-like dermatitis of whom 7 did not receive treatment and 12 were pretreated with betamethasone valerate (BMV). Tetracyclince was given all the time as additional treatment. Clinically there was no significant difference between TA and HC 17-B. No rebound phenomenon was observed. If corticosteroids are to be used at all in rosacea or resoacea-like dermatitis, preference is given to HC 17-B.

Administration, Topical

[Effects of estradiol-7 alpha-butyric acid on hypothalamus cells].

When Rat uterus was incubated at 37 degrees with estradiol-7 alpha-butyric acid (OII-7 alpha-bu), no interference was observed with the intracellular estradiol receptors. In addition, OII-7 alpha-bu did not display estrogenic effect such as in vivo inhibition of LH secretion in Rat and in vitro increased activity of the enzyme ornithine-decarboxylase in the chick oviduct. Contrary to these negative findings, we have observed preoptic and septal cells in the guinea pig where micro-iontophoresis of OII-7 alpha-bu triggers changes of the electric activity within a second. We submit therefore, that this latter response is due to an interaction between the acid estrogen and the neuron membrane.

Animals

Effect of n-dipropylacetate and L-glutamine on gamma amino-n-butyric acid levels in brains of rats intoxicated with ethanol.

The concentrations of gamma-amino-n-butyric acid in brains of rats intoxicated with half intraperitoneal LD50 ethanol were determined at various times and found increased at early times (10 min) and decreased later (1 to 3 hours). The concentration tended to be restored to normal values upon administration of n-dipropylacetate (400 mg/kg) or L-glutamine (340 mg/kg). The treatment with both substances showed a positive interaction with an overall positive effect greater than the sum of each.

Alcoholic Intoxication