Modulation of GABA by sodium butyrate ameliorates hypothalamic inflammation in experimental model of PCOS.

Polycystic ovarian syndrome (PCOS) is a known endocrine disorder that has affected many women of childbearing age, and is accompanied by various neurodegenerative conditions. Hence, this study investigates the impact of butyrate in reversing hypothalamic-related disorder, possibly through γ aminobutyric acid (GABA) in a rat model of PCOS. Eight-week-old female Wistar rats were allotted into four groups (n = 5), which include control, butyrate, letrozole, and letrozole + butyrate groups. PCOS was induced by administering 1 mg/kg of letrozole (oral gavage) for 21 days. After confirmation of PCOS, 200 mg/kg of butyrate (oral gavage) was administered for 6 weeks. Rats with PCOS were characterized by elevated levels of plasma insulin and testosterone. Increases in plasma and hypothalamic triglyceride levels, inflammatory biomarker (SDF-1), apoptotic marker (caspase-6), and decreased plasma GnRH were observed. Additionally, a decrease in hypothalamic GABA was revealed. Nevertheless, the administration of butyrate attenuated these alterations. The present study suggests that butyrate ameliorates hypothalamic inflammation in an experimental model of PCOS, a beneficial effect that is accompanied by enhanced GABA production.

Butyrate Ameliorates Intestinal Epithelial Barrier Injury Via Enhancing Foxp3+ Regulatory T-Cell Function in Severe Acute Pancreatitis Model.

BACKGROUND: This study aimed to examine the effect of sodium butyrate on severe acute pancreatitis-related gut barrier injury in a rat model and explore its mechanism. METHODS: Male rats randomly fell into 3 groups, that is, the control, the severe acute pancreatitis group, and the severe acute pancreatitis+butyrate group. Rats in the control group received sham operation, while rats in the severe acute pancreatitis group and severe acute pancreatitis+butyrate group received severe acute pancreatitis induction by intraductal infusion of 4% sodium taurocholate. After that, rats in the severe acute pancreatitis+butyrate group were fed with sodium butyrate solution with free access. Intestinal barrier injury was measured based on the expression of tight junction proteins by reverse transcription polymerase chain reaction, Western blotting assay as well as immunohistochemical staining. The variation of Treg cells was measured by reverse transcription polymerase chain reaction, Western blotting assay, immunohistochemical staining, and flow cytometry analysis. RESULTS: Compared to rats in the control, rats in the severe acute pancreatitis group showed significantly higher pathohistological scores (P < .001) in the intestine, as well as decreased expression of occludin and ZO-1. While, rats in the severe acute pancreatitis+butyrate group showed mitigated histologic lesions (P < .05) and increased expressions of occludin and ZO-1. In addition, rats in the severe acute pancreatitis group showed the obvious reduction in the expressions of Foxp3 and GPR109a and the decreased percentage of Treg cells in the intestine (P < .001) compared to rats in the control. However, rats in the severe acute pancreatitis+butyrate group showed markedly increased expressions of Foxp3 and GPR109a and the upregulated percentage of Treg cells (P < .01). CONCLUSION: Butyrate could significantly mitigate the intestinal injury induced by severe acute pancreatitis, probably by inducing the differentiation of Treg cells.

Physiological activity of E. coli engineered to produce butyric acid.

Faecalibacterium prausnitzii (F. prausnitzii) is one of the most abundant bacteria in the human intestine, with its anti-inflammatory effects establishing it as a major effector in human intestinal health. However, its extreme sensitivity to oxygen makes its cultivation and physiological study difficult. F. prausnitzii produces butyric acid, which is beneficial to human gut health. Butyric acid is a short-chain fatty acid (SCFA) produced by the fermentation of carbohydrates, such as dietary fibre in the large bowel. The genes encoding butyryl-CoA dehydrogenase (BCD) and butyryl-CoA:acetate CoA transferase (BUT) in F. prausnitzii were cloned and expressed in E. coli to determine the effect of butyric acid production on intestinal health using DSS-induced colitis model mice. The results from the E. coli Nissle 1917 strain, expressing BCD, BUT, or both, showed that BCD was essential, while BUT was dispensable for producing butyric acid. The effects of different carbon sources, such as glucose, N-acetylglucosamine (NAG), N-acetylgalactosamine (NAGA), and inulin, were compared with results showing that the optimal carbon sources for butyric acid production were NAG, a major component of mucin in the human intestine, and glucose. Furthermore, the anti-inflammatory effects of butyric acid production were tested by administering these strains to DSS-induced colitis model mice. The oral administration of the E. coli Nissle 1917 strain, carrying the expression vector for BCD and BUT (EcN-BCD-BUT), was found to prevent DSS-induced damage. Introduction of the BCD expression vector into E. coli Nissle 1917 led to increased butyric acid production, which improved the strain's health-beneficial effects.

The HDAC Inhibitor Butyrate Impairs ß Cell Function and Activates the Disallowed Gene Hexokinase I.

Histone deacetylase (HDAC) inhibitors such as butyrate have been reported to reduce diabetes risk and protect insulin-secreting pancreatic ß cells in animal models. However, studies on insulin-secreting cells in vitro have found that butyrate treatment resulted in impaired or inappropriate insulin secretion. Our study explores the effects of butyrate on insulin secretion by BRIN BD-11 rat pancreatic ß cells and examined effects on the expression of genes implicated in ß cell function. Robust HDAC inhibition with 5 mM butyrate or trichostatin A for 24 h in ß cells decreased basal insulin secretion and content, as well as insulin secretion in response to acute stimulation. Treatment with butyrate also increased expression of the disallowed gene hexokinase I, possibly explaining the impairment to insulin secretion, and of TXNIP, which may increase oxidative stress and ß cell apoptosis. In contrast to robust HDAC inhibition (>70% after 24 h), low-dose and acute high-dose treatment with butyrate enhanced nutrient-stimulated insulin secretion. In conclusion, although protective effects of HDAC inhibition have been observed in vivo, potent HDAC inhibition impairs ß cell function in vitro. The chronic low dose and acute high dose butyrate treatments may be more reflective of in vivo effects.

High doses of butyrate induce a reversible body temperature drop through transient proton leak in mitochondria of brain neurons.

AIMS: Sodium butyrate (SB) is a major product of gut microbiota with signaling activity in the human body. It has become a dietary supplement in the treatment of intestinal disorders. However, the toxic effect of overdosed SB and treatment strategy remain unknown. The two issues are addressed in current study. MATERIALS AND METHODS: SB (0.3-2.5 g/kg) was administrated through a single peritoneal injection in mice. The core body temperature and mitochondrial function in the brown adipose tissue and brain were monitored. Pharmacodynamics, targeted metabolomics, electron microscope, oxygen consumption rate and gene knockdown were employed to dissect the mechanism for the toxic effect. KEY FINDINGS: The temperature was reduced by SB (1.2-2.5 g/kg) in a dose-dependent manner in mice for 2-4 h. In the brain, the effect was associated with SB elevation and neurotransmitter reduction. Metabolites changes were seen in the glycolysis, TCA cycle and pentose phosphate pathways. Adenine nucleotide translocase (ANT) was activated by butyrate for proton transportation leading to a transient potential collapse through proton leak. The SB activity was attenuated by ANT inhibition from gene knockdown or pharmacological blocker. ROS was elevated by SB for the increased ANT activity in proton leak in Neuro-2a. SIGNIFICANCE: Excessive SB generated an immediate and reversible toxic effect for inhibition of body temperature through transient mitochondrial dysfunction in the brain. The mechanism was quick activation of ANT proteins for potential collapse in mitochondria. ROS may be a factor in the ANT activation by SB.

Sodium butyrate induces genotoxic stress in function of photoperiod variations and differentially modulates the expression of genes involved in chromatin modification and DNA repair in Petunia hybrida seedlings.

MAIN CONCLUSION: Sodium butyrate applied to Petunia hybrida seeds under a long-day photoperiod has a negative impact (reduced seedling length, decreased production of photosynthetic pigments, and accumulation of DNA damage) on early seedling development, whereas its administration under dark/light conditions (complete dark conditions for 5 days followed by exposure to long-day photoperiod for 5 days) bypasses some of the adverse effects. Genotoxic stress impairs plant development. To circumvent DNA damage, plants activate DNA repair pathways in concert with chromatin dynamics. These are essential during seed germination and seedling establishment, and may be influenced by photoperiod variations. To assess this interplay, an experimental design was developed in Petunia hybrida, a relevant horticultural crop and model species. Seeds were treated with different doses of sodium butyrate (NaB, 1 mM and 5 mM) as a stress agent applied under different light/dark conditions throughout a time period of 10 days. Phenotypic (germination percentage and speed, seedling length, and photosynthetic pigments) and molecular (DNA damage and gene expression profiles) analyses were performed to monitor the response to the imposed conditions. Seed germination was not affected by the treatments. Seedling development was hampered by increasing NaB concentrations applied under a long-day photoperiod (L) as reflected by the decreased seedling length accompanied by increased DNA damage. When seedlings were grown under dark conditions for 5 days and then exposed to long-day photoperiod for the remaining 5 days (D/L), the damaging effects of NaB were circumvented. NaB exposure under L conditions resulted in enhanced expression of HAT/HDAC (HISTONE ACETYLTRANSFERASES/HISTONE DEACTEYLASES) genes along with repression of genes involved in DNA repair. Differently, under D/L conditions, the expression of DNA repair genes was increased by NaB treatment and this was associated with lower levels of DNA damage. The observed DNA damage and gene expression profiles suggest the involvement of chromatin modification- and DNA repair-associated pathways in response to NaB and dark/light exposure during seedling development.

Oral butyrate does not affect innate immunity and islet autoimmunity in individuals with longstanding type 1 diabetes: a randomised controlled trial.

AIMS/HYPOTHESIS: The pathophysiology of type 1 diabetes has been linked to altered gut microbiota and more specifically to a shortage of intestinal production of the short-chain fatty acid (SCFA) butyrate, which may play key roles in maintaining intestinal epithelial integrity and in human and gut microbial metabolism. Butyrate supplementation can protect against autoimmune diabetes in mouse models. We thus set out to study the effect of oral butyrate vs placebo on glucose regulation and immune variables in human participants with longstanding type 1 diabetes. METHODS: We administered a daily oral dose of 4 g sodium butyrate or placebo for 1 month to 30 individuals with longstanding type 1 diabetes, without comorbidity or medication use, in a randomised (1:1), controlled, double-blind crossover trial, with a washout period of 1 month in between. Participants were randomly allocated to the 'oral sodium butyrate capsules first' or 'oral placebo capsules first' study arm in blocks of five. The clinical investigator received blinded medication from the clinical trial pharmacy. All participants, people doing measurements or examinations, or people assessing the outcomes were blinded to group assignment. The primary outcome was a change in the innate immune phenotype (monocyte subsets and in vitro cytokine production). Secondary outcomes were changes in blood markers of islet autoimmunity (cell counts, lymphocyte stimulation indices and CD8 quantum dot assays), glucose and lipid metabolism, beta cell function (by mixed-meal test), gut microbiota and faecal SCFA. The data was collected at the Amsterdam University Medical Centers. RESULTS: All 30 participants were analysed. Faecal butyrate and propionate levels were significantly affected by oral butyrate supplementation and butyrate treatment was safe. However, this modulation of intestinal SCFAs did not result in any significant changes in adaptive or innate immunity, or in any of the other outcome variables. In our discussion, we elaborate on this important discrepancy with previous animal work. CONCLUSIONS/INTERPRETATION: Oral butyrate supplementation does not significantly affect innate or adaptive immunity in humans with longstanding type 1 diabetes. TRIAL REGISTRATION: Netherlands Trial Register: NL4832 (www.trialregister.nl). DATA AVAILABILITY: Raw sequencing data are available in the European Nucleotide Archive repository (https://www.ebi.ac.uk/ena/browse) under study PRJEB30292. FUNDING: The study was funded by a Le Ducq consortium grant, a CVON grant, a personal ZONMW-VIDI grant and a Dutch Heart Foundation grant.

Sensory irritation of vapours of formic, acetic, propionic and butyric acid.

In mice, inhalation of formic, acetic, propionic and butyric acid caused a rapid decrease in the respiratory rate, which decreased to a stable level during the remaining part of the 30 min exposure period; this was due to sensory irritation. The concentration decreasing the respiratory rate (RD) by 50% (RD50) was 438, 308, 386 and 285 ppm, respectively, which allowed an adequate prediction of the Threshold Limit Values. In mice inhaling through a tracheal cannula, bypassing the trigeminal nerves, caused a slower decrease in respiratory rate due to pulmonary irritation. In the low concentration range, the pulmonary irritation response was less pronounced than the sensory irritation response. As the response in the normal (non-cannulated) mice was not influenced by pulmonary irritation, sensory irritation is the key effect, presumably due to the scrubbing effect of the upper airways, preventing access to the lungs. The activated receptors were in a non-lipophilic (hydrophilic) environment, from where the receptors may be activated by means of liberated protons. At the RD0, formic acid may, at least partly, activates ASIC, TRPV1 and TRPA1 receptors, whereas acetic, propionic and butyric acid may activate ASIC and TRPA1 receptors, based on the estimated pH in the mucus layer.

Oral sodium butyrate impacts brain metabolism and hippocampal neurogenesis, with limited effects on gut anatomy and function in pigs.

Butyrate can improve gut functions, whereas histone deacetylase inhibitors might alleviate neurocognitive alterations. Our aim was to assess whether oral butyrate could modulate brain metabolism and plasticity and if this would relate to gut function. Sixteen pigs were subjected to sodium butyrate (SB) supplementation via beverage water or water only [control (C)]. All pigs had blood sampled after 2 and 3 wk of treatment, and were subjected to a brain positron emission tomography after 3 wk. Animals were euthanized after 4 wk to sample pancreas, intestine, and brain for gut physiology and anatomy measurements, as well as hippocampal histology, Ki67, and doublecortin (DCX) immunohistochemistry. SB compared with C treatment triggered basal brain glucose metabolism changes in the nucleus accumbens and hippocampus ( P = 0.003), increased hippocampal granular cell layer volume ( P = 0.006), and neurogenesis (Ki67: P = 0.026; DCX: P = 0.029). After 2 wk of treatment, plasma levels of glucose, insulin, lactate, glucagon-like peptide 1, and peptide tyrosine tyrosine remained unchanged. After 3 wk, plasma levels of lactate were lower in SB compared with C animals ( P = 0.028), with no difference for glucose and insulin. Butyrate intake impacted very little gut anatomy and function. These results demonstrate that oral SB impacted brain functions with little effects on the gut.-Val-Laillet, D., Guérin, S., Coquery, N., Nogret, I., Formal, M., Romé, V., Le Normand, L., Meurice, P., Randuineau, G., Guilloteau, P., Malbert, C.-H., Parnet, P., Lallès, J.-P., Segain, J.-P. Oral sodium butyrate impacts brain metabolism and hippocampal neurogenesis, with limited effects on gut anatomy and function in pigs.

Differentiation Affects the Release of Exosomes from Colon Cancer Cells and Their Ability to Modulate the Behavior of Recipient Cells.

Exosomes are involved in intercellular communication. We previously reported that sodium butyrate-induced differentiation of HT29 colon cancer cells is associated with a reduced CD133 expression. Herein, we analyzed the role of exosomes in the differentiation of HT29 cells. Exosomes were prepared using ultracentrifugation. Gene expression levels were evaluated by real-time PCR. The cell proliferation rate was assessed by MTT assay and with the electric cell-substrate impedance sensing system, whereas cell motility was assessed using the scratch test and confocal microscopy. Sodium butyrate-induced differentiation of HT29 and Caco-2 cells increased the levels of released exosomes and their expression of CD133. Cell differentiation and the decrease of cellular CD133 expression levels were prevented by blocking multivesicular body maturation. Exosomes released by HT29 differentiating cells carried increased levels of miRNAs, induced an increased proliferation and motility of both colon cancer cells and normal fibroblasts, increased the colony-forming efficiency of cancer cells, and reduced the sodium butyrate-induced differentiation of HT29 cells. Such effects were associated with an increased phosphorylation level of both Src and extracellular signal regulated kinase proteins and with an increased expression of epithelial-to-mesenchymal transition-related genes. Release of exosomes is affected by differentiation of colon cancer cells; exosomes might be used by differentiating cells to get rid of components that are no longer necessary but might continue to exert their effects on recipient cells.

High doses of the histone deacetylase inhibitor sodium butyrate trigger a stress-like response.

The hypothalamic-pituitary-adrenal (HPA) axis is activated by a wide range of stimuli, including drugs. Here we report that in male rats, a dose of sodium butyrate (NaBu) that is typically used to inhibit histone deacetylation (1200 mg/kg) increased the peripheral levels of HPA hormones and glucose. In a further experiment, we compared the effects of two different doses of NaBu (200 and 1200 mg/kg) and equimolar saline solutions on peripheral neuroendocrine markers and brain c-Fos expression to demonstrate a specific stress-like effect of NaBu that is not related to hypertonicity and to localise putatively involved brain areas. Only the high dose of NaBu increased the plasma levels of stress markers. The equimolar (hypertonic) saline solution also activated the HPA axis and the c-Fos expression in the paraventricular nucleus of the hypothalamus (PVN), a key area for the control of the HPA axis, but the effects were of a lower magnitude than those of NaBu. Regarding other brain areas, group differences in c-Fos expression were not observed in the medial prefrontal cortex or the medial amygdala, but they were observed in the central amygdala and the lateral ventral septum. However, only the latter area of the NaBu group showed enhanced c-Fos expression that was significantly higher than that after hypertonic saline. The present data indicate that high doses of NaBu appear to act as a pharmacological stressor, and this fact should be taken into account when using this drug to study the role of epigenetic processes in learning and emotional behaviour.

Efeito dos ácidos lático e butírico, isolados e associados, sobre o desempenho e morfometria intestinal em frangos de corte / Effect of lactic and butiric acids , isolated and associated, on performance and intestinal morfometric in broilers

A avicultura nacional tem sofrido constantes desafios, tendo como objetivo ampliar o mercado consumidor do frango de corte ao redor do mundo. Cada mercado apresenta exigências diferentes, como no caso do mercado europeu que estabelece a exclusão dos antibióticos como promotores de crescimento na alimentação dos frangos. Em função deste fato, realizou-se este estudo, com o objetivo de avaliar o uso dos ácidos lático e butírico, isolados e associados, como aditivos em dietas de frangos de corte, em relação ao promotor de crescimento usualmente utilizado nas dietas dos mesmos. Foram avaliados o desempenho e a morfometria intestinal das aves. Utilizaram-se 1400 pintinhos machos da linhagem comercial Ross, divididos em cinco tratamentos (controle - sem aditivo; com adição de ácido butírico; com adição de ácido lático; com adição de ácido butírico+ ácido láctico; com adição do antibiótico avilamicina, promotor de crescimento tradicionalmente utilizado na criação de frangos de corte. Os resultados de desempenho indicaram que a interação dos ácidos foi significativa na fase inicial, entretanto não ocorreu um efeito aditivo dos ácidos, sendo o uso do ácido butírico mais recomendável durante essa fase. Já na fase de crescimento, a interação foi significativa com um efeito aditivo, recomendando seu uso nas rações de crescimento. Os resultados obtidos nas analises de morfometria intestinal não foram conclusivos. Em termos gerais, requerem-se mais estudos quanto ao uso dos ácidos orgânicos como promotores de crescimento.

During the last years, the national poultry keeping has suffered constant challenges, having as objective to diversify the consuming market of the broiler to around of the world. However, each market presents different requirements, as in the case of the European market that establishes the exclusion of antibiotics as promotional of growth in the feeding of the broilers. Aiming at this problematic one, this study was become fullfilled, with the objective to analyze the results of the association of the butyric and lactic acids as additives in the ration of broilers, in comparison to the results gotten for the usually used promoters of growth in the diets of broilers. They had been evaluated: the animal performance, humoral immunology and intestinal morfometric of the birds. 1400 male chickens of the commercial ancestry Ross had been used, dividing them in five groups with different treatments to that if it relates to the additive use. Being a group it has controlled absent of additive, a group with butyric acid, a group with lactic acid, the fourth group with the association of butyric and lactic acids, and the fifth group with avilamicina (antibiotic) as promotional of traditionally used growth in the production of broilers. The performance results had indicated that the interaction of acid ones was significant in the initial phase, mean while did not occur an additive effect of acid ones, being the use of the butyric acid separately more recommendable during this phase. Already in the growth phase, the interaction was significant with an additive effect, recommending its use in the rations of growth. In accordance with the average headings of antibodies gotten in the study, the interaction was significant in the third sampling and showed a synergic effect of acid ones, being the combination of acids in question a powerful modulator of the humoral immunity. Meanwhile, the results gotten in the analyses of intestinal morfometric had not been conclusive. In general terms, one requires more organic acid studies that confirm the use of as the promotional ones of growth.

Effects of hydrochloric, acetic, butyric, and propionic acids on pathogenesis of ulcers in the nonglandular portion of the stomach of horses.

OBJECTIVE: To identify the pathogenesis of gastric ulcers by comparing injury to the nonglandular gastric mucosa of horses caused by hydrochloric acid (HCl) or volatile fatty acids (VFAs). SAMPLE POPULATION: Gastric tissues from 30 horses. PROCEDURE: Nonglandular gastric mucosa was studied by use of Ussing chambers. Short-circuit current (Isc) and potential difference were measured and electrical resistance calculated for tissues after addition of HCl and VFAs to normal Ringer's solution (NRS). Tissues were examined histologically. RESULTS: Mucosa exposed to HCl in NRS (pH, 1.5) had a significant decrease in Isc, compared with Isc for mucosa exposed to NRS at pH 4.0 or 7.0. Also, exposure to 60mM acetic, propionic, and butyric acids (pH, 4.0 or 1.5) caused an immediate significant decrease in Isc. Recovery of sodium transport was detected only in samples exposed to acetic acid at pH 4.0. Recovery of sodium transport was not seen in other mucosal samples exposed to VFAs at pH < or = 4.0. CONCLUSIONS AND CLINICAL RELEVANCE: Acetic, butyric, and propionic acids and, to a lesser extent, HCl caused decreases in mucosal barrier function of the nonglandular portion of the equine stomach. Because of their lipid solubility at pH < or = 4.0, undissociated VFAs penetrate cells in the nonglandular gastric mucosa, which causes acidification of cellular contents, inhibition of sodium transport, and cellular swelling. Results indicate that HCl alone or in combination with VFAs at gastric pH < or = 4.0 may be important in the pathogenesis of gastric ulcers in the nonglandular portion of the stomach of horses.

Induction of experimental ulcerative colitis by Fusobacterium varium isolated from colonic mucosa of patients with ulcerative colitis.

BACKGROUND: Bacteria are implicated in certain forms of model chronic colitis but the identity and role of bacteria in human ulcerative colitis (UC) are uncertain. AIMS: To isolate pathogenic bacteria from inflamed mucosa of patients with UC, to examine whether the bacteria have a toxin to Vero cells, and to determine whether the toxin induces UC-like lesions in animals. METHODS: Bacteria were isolated from UC patients and supernatants from cultures were filtered and tested for cytotoxicity to Vero cells. Bacterial cells producing the cytotoxic supernatants were examined by polymerase chain reaction for verotoxin genes. Culture supernatants of cytotoxic strains were examined by high performance liquid chromatography for organic acid concentrations. Mice were given enemas containing organic acid at the mean concentration in the supernatants of cytotoxic strains to ascertain whether colonic lesions appear in UC. RESULTS: Only supernatants from cultures of Fusobacterium varium killed Vero cells. Bacterial cells lacked verotoxin genes. Bacterial culture supernatants contained high concentrations of n-butyric acid and the mean concentration (32 mmol/l) was cytotoxic to Vero cells. Twenty four hours after mice were given enemas containing either butyric acid or F varium culture supernatants, colonic ulcers with crypt abscesses, inflammatory cell infiltration, and apoptotic changes were observed. CONCLUSIONS: Butyric acid in culture supernatants from cultures of F varium caused UC-like lesions in mice. This study indicates that F varium may be one of the elusive pathogenic factors in UC.