Administration Of Short-chain Fatty Acids Research Articles

Developing insulin-like peptide 5-based antagonists for the G protein-coupled receptor, RXFP4

Human insulin-like peptide 5 (INSL5) is a gut hormone produced by colonic L-cells, and its biological functions are mediated by Relaxin Family Peptide Receptor 4 (RXFP4). Our preliminary data indicated that RXFP4 agonists are potential drug leads for the treatment of constipation. More recently, we designed and developed a novel RXFP4 antagonist, A13-nR that was shown to block agonist-induced activity in cells and animal models. We showed that A13-nR was able to block agonist-induced increases in colon motility in mice of both genders that express the receptor, RXFP4. Our data also showed that colorectal propulsion induced by intracolonic administration of short-chain fatty acids was antagonized by A13-nR. Therefore, A13-nR is an important research tool and potential drug lead for the treatment of colon motility disorders, such as bacterial diarrhea. However, A13-nR acted as a partial agonist at high concentrations in vitro and demonstrated modest antagonist potency (∼35 nM). Consequently, the primary objective of this study is to pinpoint novel modifications to A13-nR that eliminate partial agonist effects while preserving or augmenting antagonist potency. In this work, we detail the creation of a series of A13-nR-modified analogues, among which analogues 3, 4, and 6 demonstrated significantly improved RXFP4 affinity (∼3 nM) with reduced partial agonist activity, enhanced antagonist potency (∼10 nM) and maximum agonist inhibition (∼80 %) when compared with A13-nR. These compounds have potential as candidates for further preclinical evaluations, marking a significant stride toward innovative therapeutics for colon motility disorders.

SCFAs Supplementation Rescues Anxiety- and Depression-like Phenotypes Generated by Fecal Engraftment of Treatment-Resistant Depression Rats.

Alteration of gut microbiota and microbial metabolites such as short-chain fatty acids (SCFAs) coexisted with stress-generated brain disorders, including depression. Herein, we investigated the effect of SCFAs in a treatment-resistant depression (TRD) model of rat. Rats were exposed to chronic-unpredictable mild stress (CUMS) and repeated adrenocorticotropic hormone (ACTH) injections to generate a TRD-like phenotype. The cecal contents of these animals were engrafted into healthy-recipient rats and allowed to colonize for 4 weeks (TRD-FMT group). Blood, brain, colon, fecal, and cecal samples were collected for molecular studies. Rats exposed to CUMS + ACTH showed TRD-like phenotypes in sucrose-preference (SPT), forced swim (FST), and elevated plus maze (EPM) tests. The TRD-FMT group also exhibited anxiety- and depression-like behaviors. Administration of SCFAs (acetate, propionate, and butyrate at 67.5, 25, and 40 mM, respectively) for 7 days exerted robust antidepressant and antianxiety effects by restoring the levels of SCFAs in plasma and fecal samples, and proinflammatory cytokines (TNF-α and IL-6), serotonin, GABA, norepinephrine, and dopamine in the hippocampus and/or frontal cortex of TRD and TRD-FMT animals. SCFAs treatment elevated the expression of free-fatty acid receptors 2/3, BDNF, doublecortin, and zonula-occludens, and reduced the elevated plasma levels of kynurenine and quinolinic acid and increased mucus-producing goblet cells in TRD and TRD-FMT animals. In 16S sequencing results, decreased microbial diversity in TRD rats corresponds with differences in the genus of Faecalibacterium, Anaerostipes, Allobaculum, Blautia, Peptococcus, Rombustia, Ruminococcaceae_UCG-014, Ruminococcaceae_UCG-002, Solobacterium, Subdolibacterium, and Eubacterium ventriosum. SCFAs may impart beneficial effects via modulation of tryptophan metabolism, inflammation, neurotransmitters, and microbiota-gut-brain axis in TRD rats.

A8 LUMINAL SHORT CHAIN FATTY ACIDS INCREASE ACTIVITY IN COLONIC MYENTERIC PLEXUS NEURONS IN MICE

Abstract Background The enteric nervous system (ENS) plays a critical role in the control of gastrointestinal functions. Short chain fatty acids (SCFAs) are the products of non-digestible dietary fiber fermentation by gut microbial metabolism and are a primary energy source for enterocytes. However, the impact of SCFAs on the ENS is not well understood. Aims We aimed to test whether SCFAs affect enteric neuronal function using a novel intact preparation of the colon. Methods Intact segments of the colon from mice expressing a genetically encoded fluorescent calcium (Ca2+) reporter specifically in all enteric neurons (Wnt-1-GCaMP6 mice) or in intrinsic primary afferent neurons (IPANs, Calb1-GCaMP6 mice) were bathed and luminally perfused with Krebs solution. Contractility was inhibited by nicardipine (3 µM) and scopolamine (1 µM). The duration and intensity of neuronal intracellular Ca2+ fluorescence responses to SCFA were visualized by live-cell confocal microscopy and were analyzed using Imaris™ software. The SCFAs were luminally perfused at a constant rate of 10 mL/h for 3 min. NaCl was modified in Krebs solution to correct for the osmolarity of SCFAs. Results In the myenteric plexus of Wnt-1-GCaMP6 mice, SCFAs (acetate, 60 mM; propionate, 20 mM or butyrate, 20 mM) caused a rapid and sustained increase in Ca2+ fluorescence (26.8±5.9, 25.8±1.7, and 27.4±5.9 min, respectively). The peak Ca2+ responses were 92.5%± 28.3%, 122.6%±21.5% and 111.7%± 23.0%, respectively. The combination of these three SCFAs increased Ca2+ fluorescence for 20.5±6.3 min and potentiated levels of Ca2+ fluorescence to 125.1%±21.6% above baseline. The percentages of neurons responding to SCFAs were 85.7%, 94.4%, 97.7% and 93.3 % for acetate, propionate, butyrate and a combination of SCFAs, respectively. In Calb1-GCaMP6 mice, IPANs showed a similar pattern of Ca2+ responses and a proportion of neurons that responded to the SCFAs. Following acetate, propionate, butyrate and combination of SCFAs administration, fluorescence was increased for 13.5±1.2, 20.6±2.7, 24.9±2.2 and 46.1±4.6 min, by a magnitude of 404.3%±153.7%, 129.0%±0.4%, 73.9%±18.8% and 717%±267.0% above baseline and the percentages of neuronal response were 100.0%, 100.0%, 100.0% and 87.5 %, respectively. Conclusions We show that SCFAs activate neurons of the myenteric plexus including IPANs. These studies suggest that some of the actions of SCFAs are mediated by the ENS, but further studies are needed to identify the underlying mechanisms of action of SCFAs in the myenteric plexus. Funding Agencies CAG, CIHRTRIANGLE Canada

A gutsy performance: the potential for supplementation of short-chain fatty acids to benefit athletic health, exercise performance, and recovery.

The gut microbiome is known to play an important role in the day-to-day physiology and health of the human host. It is, therefore, not surprising that there is interest surrounding the gut microbiome and its potential to benefit athletic health and performance. This has, in part, been driven by the consideration that gut bacterial by-products (i.e. metabolic waste) could be harnessed by the host and utilised for a beneficial outcome. The concept of harnessing bacterial metabolites as beneficial health modulators has developed the theory of leveraging short-chain fatty acids (SCFAs) as novel supplements for enhancing athletic performance. This review discusses the current literature investigating SCFA administration in cellular, animal, and human models, with the aim of linking the demonstrated physiological/biochemical mechanisms to potential exercise/athletic benefit. In addition, practical implications and factors relating to SCFA-supplementation in athletic populations are considered. The literature demonstrates a tangible rationale that SCFAs can have a positive impact on human physiology to afford benefits to the athletic population. These advantages include the capacity to improve respiratory immunity to combat elevated levels/severity of upper respiratory tract infections often reported in athletes; the blunting of pro-inflammatory and pro-fibrotic pathways to aid in exercise recovery; and the role of SCFAs as usable energy sources and metabolism modulators to fuel exercise and improve performance and/or endurance capacity. However, there is currently minimal research completed in human participants and thus further investigations into the direct benefit of SCFAs in exercise performance and/or recovery-based studies are required.

Fecal microbiota transplantation and short-chain fatty acids improve learning and memory in fluorosis mice by BDNF-PI3K/AKT pathway

Fluoride, an environmental toxicant, not only arouses intestinal microbiota dysbiosis, but also causes neuronal apoptosis and a decline in learning and memory ability. The purpose of this study was to explore whether fecal microbiota transplantation (FMT) from healthy mice and bacteria-derived metabolites short-chain fatty acids (SCFAs) supplement protect against fluoride-induced learning and memory impairment. Results showed that FMT reversed the elevated percentage of working memory errors (WME) and reference memory errors (RME) in fluorosis mice during the eight-arm maze test. Nissl and TUNEL staining presented that fluoride led to a decreased proportion of Nissl bodies area in the hippocampal CA3 region and an increased apoptotic ratio of nerve cells in CA1, CA3 and DG areas, whereas FMT alleviated those pathological damages. Moreover, the expressions of mRNA in hippocampal BDNF, PDK1, AKT, Bcl-2, and Bcl-xL were downregulated in mice exposed to fluoride, but the levels of PI3K, Bax, Bak, and Caspase-7 mRNA were upregulated. NaF treatment had an increase in PI3K and Caspase-3 protein levels and reduced the expressions of these four proteins, including BDNF, p-PI3K, AKT and p-AKT. By contrast, FMT enhanced the expression of BDNF and thus activated the PI3K/AKT pathway. Besides, the 16S rRNA sequencing revealed that fluoride caused a reduction in certain SCFA producers in the colon as evidenced by a decline in Erysipelatoclostridiaceae, and a downward trend in Akkermansia, Blautia and Alistipes. However, the disordered gut microbiome was restored via frequent FMT. Of note, SCFAs administration also increased BDNF levels and regulated its downstream pathways, which contributed to cell survival and learning and memory function recovery. In conclusion, FMT and SCFAs may activate the BDNF-PI3K/AKT pathway to play an anti-apoptotic role and ultimately improve learning and memory deficits in fluorosis mice.

Systemic antibiotics cause deterioration of emphysema associated with exaggerated inflammation and autophagy

The interaction between the microbial environment and the host is important for immune homeostasis. Recent research suggests that microbiota dysbiosis can be involved in respiratory diseases. Emphysema is a chronic inflammatory disease, but it is unclear whether dysbiosis caused by antibiotics can affect disease progression. Here, we tried to elucidate the effect of systemic antibiotics on smoking-exposed emphysema models. In this study, the antibiotic mixture caused more alveolar destruction and airspace expansion in the smoking group than in the smoking only or control groups. This emphysema aggravation as a result of antibiotic exposure was associated with increased levels of inflammatory cells, IL-6, IFNγ and protein concentrations in bronchoalveolar lavage fluid. Proteomics analysis indicated that autophagy could be involved in antibiotic-associated emphysema aggravation, and increased protein levels of LC3B, atg3, and atg7 were identified by Western blotting. In microbiome and metabolome analyses, the composition of the gut microbiota was different with smoking and antibiotic exposure, and the levels of short-chain fatty acids (SCFAs), including acetate and propionate, were reduced by antibiotic exposure. SCFA administration restored emphysema development with reduced inflammatory cells, IL-6, and IFNγ and decreased LC3B, atg3, and atg7 levels. In conclusion, antibiotics can aggravate emphysema, and inflammation and autophagy may be associated with this aggravation. This study provides important insight into the systemic impact of microbial dysbiosis and the therapeutic potential of utilizing the gut microbiota in emphysema.

Gut microbiota regulates blood-cerebrospinal fluid barrier function and Aβ pathology.

Accumulating evidence indicates that gut microbiota dysbiosis is associated with increased blood-brain barrier (BBB) permeability and contributes to Alzheimer's disease (AD) pathogenesis. In contrast, the influence of gut microbiota on the blood-cerebrospinal fluid (CSF) barrier has not yet been studied. Here, we report that mice lacking gut microbiota display increased blood-CSF barrier permeability associated with disorganized tight junctions (TJs), which can be rescued by recolonization with gut microbiota or supplementation with short-chain fatty acids (SCFAs). Our data reveal that gut microbiota is important not only for the establishment but also for the maintenance of a tight barrier. Also, we report that the vagus nerve plays an important role in this process and that SCFAs can independently tighten the barrier. Administration of SCFAs in AppNL-G-F mice improved the subcellular localization of TJs at the blood-CSF barrier, reduced the β-amyloid (Aβ) burden, and affected microglial phenotype. Altogether, our results suggest that modulating the microbiota and administering SCFAs might have therapeutic potential in AD via blood-CSF barrier tightening and maintaining microglial activity and Aβ clearance.

Butyrate regulates neutrophil homeostasis and impairs early antimicrobial activity in the lung

Short-chain fatty acids (SCFAs) are metabolites that are produced after microbial fermentation of dietary fiber and impact cell metabolism and anti-inflammatory pathways both locally in the gut and systemically. In preclinical models, administration of SCFAs, such as butyrate, ameliorates a range of inflammatory disease models including allergic airway inflammation, atopic dermatitis, and influenza infection. Here we report the effect of butyrate on a bacteria-induced acute neutrophil-driven immune response in the airways. Butyrate impacted discrete aspects of hematopoiesis in the bone marrow resulting in the accumulation of immature neutrophils. During Pseudomonas aeruginosa infection, butyrate treatment led to the enhanced mobilization of neutrophils to the lungs as a result of increased CXCL2 expression by lung macrophages. Despite this increase in granulocyte numbers and their enhanced phagocytic capacity, neutrophils failed to control early bacterial growth. Butyrate reduced the expression of nicotinamide adenine dinucleotide phosphate, oxidase complex components required for reactive oxygen species production, and reduced secondary granule enzymes, culminating in impaired bactericidal activity. These data reveal that SCFAs tune neutrophil maturation and effector function in the bone marrow under homeostatic conditions, potentially to mitigate against excessive granulocyte-driven immunopathology, but their consequently restricted bactericidal capacity impairs early control of Pseudomonas infection.

Microbiota-derived SCFAs Promote Chronic Neurogenesis and Anti-inflammatory Gene Expression after Traumatic Brain Injury (P1-6.008)

<h3>Objective:</h3> To evaluate whether short-chain fatty acids (SCFAs) will benefit the long-term recovery of traumatic brain injury (TBI) in mice. <h3>Background:</h3> SCFAs, including acetate and butyrate, are the main metabolites generated by gut microbiota consuming dietary fiber in the colon. SCFAs have pleotropic effects throughout the body including influences on immunity and neurogenesis. We hypothesized that SCFAs could influence recovery from TBI. <h3>Design/Methods:</h3> Adult C57BL6/J mice (n=4/group) were randomized to standard high-amylose maize starch (HAMS) chow or acetylated- and butyrylated-HAMS (HAMS-AB). Fermentation of HAMS-AB by gut bacteria releases acetate and butyrate in the intestinal lumen. The controlled cortical impact (CCI) model of TBI (6m/s, 2.2mm depth, 50msec dwell) was performed. Sham mice received anesthesia and skin incision without craniotomy. At 6 months post-injury, mice were sacrificed and single-cell mRNA from pericontusional tissue sequenced following the 10X Genomics protocol. After pre-processing by <i>CellRanger</i> and SCT normalization by <i>Seurat</i>, cells were annotated based on canonical markers. Cell subtype gene modules were clustered by weighted gene co-expression network analysis (WGCNA) and enriched at gene-ontology (GO) pathways. The GO pathway and regulon activity were evaluated by <i>AUCell</i> and <i>SCENIC</i> respectively. <h3>Results:</h3> In a total of 116,134 cells, consisting of 12 cell types, from 3 groups (41,509 from sham_control, 36,409 from injury_control, 38,216 from injury_SCFA,) passed the quality control and doublet filtering. Both the cell composition and transcriptome were different in each group. SCFAs decreased gene expression associated with neuronal activity and increased the immature neuron frequency. During immature neuron differentiation, SCFAs downregulated neurodegenerative disease-related genes expression in injured mice. In addition, SCFAs decreased chronic microglia/macrophage activation and promoted an M2-like phenotype. Lastly, SCFAs attenuated T cell activation and cytotoxic-related pathways. <h3>Conclusions:</h3> SCFAs impact gene expression in multiple cell types at 6 months after TBI. The pathways affected suggest a beneficial role for gut-derived SCFA administration for recovery after TBI. <b>Disclosure:</b> Dr. Xiong has nothing to disclose. Mrs. Nelson has nothing to disclose. Mr. Sneiderman has nothing to disclose. Ms. Feldman has nothing to disclose. Patrick M. Kochanek, MD, MCCM has received personal compensation in the range of $10,000-$49,999 for serving as an Expert Witness for Johns Hopkins Health System. The institution of Patrick M. Kochanek, MD, MCCM has received research support from Chuck Noll Foundation. The institution of Patrick M. Kochanek, MD, MCCM has received research support from NIH. Patrick M. Kochanek, MD, MCCM has received publishing royalties from a publication relating to health care. Dr. Rajasundaram has nothing to disclose. Dr. Jha has received personal compensation in the range of $5,000-$9,999 for serving as a Consultant for Biogen. An immediate family member of Dr. Jha has received personal compensation in the range of $5,000-$9,999 for serving as an Expert Witness for Legal fees. The institution of Dr. Jha has received research support from NIH/NINDS, Chuck Noll Foundation, University of Pittsburgh, Barrow Neurological Foundation. The institution of Dr. Kohanbash has received research support from NIH. Dr. Simon has nothing to disclose.

Short-chain fatty acid-releasing nano-prodrugs for attenuating growth and metastasis of melanoma.

Low-molecular-weight (LMW) short-chain fatty acids (SCFAs), such as propionic and butyric acids, have been reported to possess anti-neoplastic effects; however, rapid renal clearance and high dose-based side effects limit their clinical translation. Hence, in this study, we have designed a new self-assembling nano-prodrugs that can effectively supply SCFAs: endogenous enzyme-metabolizable block copolymer poly(ethylene glycol)block-poly(vinyl ester) possessing several units of SCFAs conjugated as side chains via ester linkages. These amphiphilic polymers spontaneously self-assemble into nanostructures under aqueous conditions to form orally administrable nano-prodrugs (butyric acid: NanoBA and propionic acid: NanoPA). Herein, we show the therapeutic efficacy of SCFA nanoparticles (NanoSCFA) in a mouse model of metastasis (melanoma). Ad libitum intake of our NanoSCFA markedly demonstrated a decrease in the metastatic tumor nodules in the lungs compared with the effect observed after LMW SCFA administration with no discernible toxicity to the GI tract. In contrast, LMW SCFAs, even at a lower concentration than that of the NanoSCFA, facilitated villus atrophy. Taken together, our work suggests that the use of NanoSCFA as a therapeutic intervention for metastatic cancer is preferable over typical LMW SCFAs. STATEMENT OF SIGNIFICANCE: Low-molecular-weight (LMW) short-chain fatty acids (SCFAs) have shown versatile therapeutic effects on various diseases, including anti-tumorigenesis effects. However, their clinical translation is limited due to their poor pharmacokinetic profile and adverse effects. To overcome these limitations, we have developed new amphiphilic block copolymer-based SCFA-prodrugs, which self-assemble into nanoparticles in aqueous media (NanoSCFA). SCFAs are covalently conjugated to the hydrophobic polymer segment via ester linkage, which can be enzymatically metabolized after oral administration. In the present study, we confirmed that ad libitum intake of NanoSCFAs retarded the growth and metastatic potential of B16-F10 tumors compared to the LMW SCFAs with negligible discernible toxicity, reflecting NanoSCFA as a preferable therapeutic intervention to LMW SCFA counterparts.

Fecal microbiota transplantation and short-chain fatty acids protected against cognitive dysfunction in a rat model of chronic cerebral hypoperfusion.

Clear roles and mechanisms in explaining gut microbial dysbiosis and microbial metabolites short-chain fatty acids (SCFAs) alterations in chronic cerebral ischemic pathogenesis have yet to be explored. In this study, we investigated chronic cerebral hypoperfusion (CCH)-induced gut microbiota and metabolic profiles of SCFAs as well as the effects and mechanisms of fecal microbiota transplantation (FMT) and SCFAs treatment on CCH-induced hippocampal neuronal injury. Bilateral common carotid artery occlusion (BCCAo) was used to establish the CCH model. Gut microbiota and SCFAs profiles in feces and hippocampus were evaluated by 16S ribosomal RNA sequencing and gas chromatography-mass spectrometry. RNA sequencing analysis was performed in hippocampal tissues. The potential molecular pathways and differential genes were verified through western blot, immunoprecipitation, immunofluorescence, and ELISA. Cognitive function was assessed via the Morris water maze test. Ultrastructures of mitochondria and synapses were tested through a transmission electron microscope. Chronic cerebral hypoperfusion induced decreased fecal acetic and propionic acid and reduced hippocampal acetic acid, which were reversed after FMT and SCFAs administration by changing fecal microbial community structure and compositions. Furthermore, in the hippocampus, FMT and SCFAs replenishment exerted anti-neuroinflammatory effects through inhibiting microglial and astrocytic activation as well as switching microglial phenotype from M1 toward M2. Moreover, FMT and SCFAs treatment alleviated neuronal loss and microglia-mediated synaptic loss and maintained the normal process of synaptic vesicle fusion and release, resulting in the improvement of synaptic plasticity. In addition, FMT and SCFAs supplement prevented oxidative phosphorylation dysfunction via mitochondrial metabolic reprogramming. The above effects of FMT and SCFAs treatment led to the inhibition of CCH-induced cognitive impairment. Our findings highlight FMT and SCFAs replenishment would be the feasible gut microbiota-based strategy to mitigate chronic cerebral ischemia-induced neuronal injury.

Antibiotic Knockdown of Gut Bacteria Sex-Dependently Enhances Intravenous Fentanyl Self-Administration in Adult Sprague Dawley Rats.

Communication between the brain and gut bacteria impacts drug- and addiction-related behaviors. To investigate the role of gut microbiota on fentanyl reinforcement and reward, we depleted gut bacteria in adult Sprague Dawley male and female rats using an oral, nonabsorbable antibiotic cocktail and allowed rats to intravenously self-administer fentanyl on an escalating schedule of reinforcement. We found that antibiotic treatment enhanced fentanyl self-administration in males, but not females, at the lowest schedule of reinforcement (i.e., fixed ratio 1). Both males and females treated with antibiotics self-administered greater amounts of fentanyl at higher schedules of reinforcement. We then replete microbial metabolites via short-chain fatty acid administration to evaluate a potential mechanism in gut-brain communication and found that restoring metabolites decreases fentanyl self-administration back to controls at higher fixed ratio schedules of reinforcement. Our findings highlight an important relationship between the knockdown and rescue of gut bacterial metabolites and fentanyl self-administration in adult rats, which provides support for a significant relationship between the gut microbiome and opioid use. Further work in this field may lead to effective, targeted treatment interventions in opioid-related disorders.

Airway-delivered short-chain fatty acid acetate boosts antiviral immunity during rhinovirus infection

Microbiota are recognized to play a major role in regulation of immunity through release of immunomodulatory metabolites such as short-chain fatty acids (SCFAs). Rhinoviruses (RVs) induce upper respiratory tract illnesses and precipitate exacerbations of asthma and chronic obstructive pulmonary disease through poorly understood mechanisms. Local interactions between SCFAs and antiviral immune responses in the respiratory tract have not been previously investigated. We sought to investigate whether pulmonary metabolite manipulation through lung-delivered administration of SCFAs can modulate antiviral immunity to RV infection. We studied the effects of intranasal administration of the SCFAs acetate, butyrate, and propionate on basal expression of antiviral signatures, and of acetate in a mouse model of RV infection and in RV-infected lung epithelial cell lines. We additionally assessed the effects of acetate, butyrate, and propionate on RV infection in differentiated human primary bronchial epithelial cells. Intranasal acetate administration induced basal upregulation of IFN-β, an effect not observed with other SCFAs. Butyrate induced RIG-I expression. Intranasal acetate treatment of mice increased interferon-stimulated gene and IFN-λ expression during RV infection and reduced lung virus loads at 8 hours postinfection. Acetate ameliorated virus-induced proinflammatory responses with attenuated pulmonary mucin and IL-6 expression observed at day 4 and 6 postinfection. This interferon-enhancing effect of acetate was confirmed in human bronchial and alveolar epithelial cell lines. In differentiated primary bronchial epithelial cells, butyrate treatment better modulated IFN-β and IFN-λ gene expression during RV infection. SCFAs augment antiviral immunity and reduce virus load and proinflammatory responses during RV infection.

Oligofructose restores postprandial short-chain fatty acid levels during high-fat feeding.

Obesity is associated with consumption of a Western diet low in dietary fiber, while prebiotics reduce body weight. Fiber induces short-chain fatty acid (SCFA) production, and SCFA administration is beneficial to host metabolic homeostasis. However, the role of endogenous SCFA signaling in the development of obesity is contentious. Therefore, the primary objective of this study is to evaluate the postprandial time course of SCFA production and uptake in healthy (chow-fed), Western diet-fed (high-fat diet [HFD]) obese, and oligofructose-treated HFD-fed (HFD + OFS) rats. Male Sprague-Dawley rats were maintained on chow or HFD for 5 weeks, with or without supplementation of 10% OFS for 3 weeks. SCFAs were measured in the ileum, cecum, colon, portal vein, and vena cava at 0, 2, 4, 6, and 8 hours postprandially. Postprandial cecal and portal vein SCFAs were decreased in obese rats compared with lean chow controls, whereas no differences were observed in fasting SCFA concentrations. OFS supplementation increased SCFA levels in the cecum and portal vein during obesity. Butyrate levels were positively associated with portal glucagon-like peptide 1 and adiposity and with Roseburia relative abundance. The current study demonstrates that obesity is associated with reduced SCFA production, and that OFS supplementation increases SCFA levels. Additionally, postprandial butyrate production appears to be beneficial to host energy homeostasis.

Short Chain Fatty Acids Protect the Cognitive Function of Sepsis Associated Encephalopathy Mice via GPR43.

ObjectiveThis study aims to analyze the changes of fecal short chain fatty acids (SCFAs) content and gut microbiota composition in sepsis associated encephalopathy (SAE) mice, further evaluating the effect of SCFAs on cognitive function and the underlying mechanism in SAE mice.MethodsA total of 55 male adult C57BL/6 mice (2–3 months of age, 20–25 g) were divided into four groups randomly: sham group (n = 10), cecal ligation and puncture group (CLP group, n = 15), CLP+SCFAs group (n = 15), and CLP+SCFAs+GLPG0974 group (n = 15). Seven days after surgery, fecal samples were collected for microbiota composition and SCFA analysis from 6 mice in each group randomly. Behavioral test was applied to assess cognitive impairment at the same time. After that, mice were sacrificed and brain tissue was harvested for inflammatory cytokines analysis.ResultsThe levels of acetic acid (.57 ± 0.09 vs 2.00 ± 0.24, p < 0.001) and propionic acid (.32 ± 0.06 vs .66 ± 0.12, p = 0.002) were significantly decreased in the CLP group compared with the sham group. The administration of SCFAs significantly increased the levels of acetic acid (1.51 ± 0.12 vs. 0.57 ± 0.09, p < 0.001) and propionic acid (0.54 ± 0.03 vs. 0.32 ± 0.06, p = 0.033) in CLP+SCFAs group compared with CLP group. Relative abundance of SCFAs-producing bacteria, including Allobaculum (0.16 ± 0.14 vs. 15.21 ± 8.12, p = 0.037), Bacteroides (1.82 ± 0.38 vs. 15.21 ± 5.95, p = 0.002) and Bifidobacterium (0.16 ± 0.06 vs. 2.24 ± 0.48, p = 0.002), significantly decreased in the CLP group compared with the sham group. The behavioral tests suggested that cognitive function was impaired in SAE mice, which could be alleviated by SCFAs pretreatment. ELISA tests indicated that the levels of IL-1β, IL-6, and TNF-α were elevated in SAE mice and SCFAs could lower them. However, the GPR43 antagonist, GLPG0974, could reverse the cognitive protective effect and anti-neuroinflammation effect of SCFAs.ConclusionOur study suggested that in SAE, the levels of acetate and propionate decreased significantly, accompanied by gut microbiota dysbiosis, particularly a decrease in SCFAs-producing bacteria. GPR43 was essential for the anti-neuroinflammation and cognitive protective effect of SCFAs in SAE.

Intracerebroventricular Delivery of Gut-derived Microbial Metabolites in Freely Moving Mice.

The impact of gut microbiota and their metabolites on host physiology and behavior has been extensively investigated in this decade. Numerous studies have revealed that gut microbiota-derived metabolites modulate brain-mediated physiological functions through intricate gut-brain pathways in the host. Short-chain fatty acids (SCFAs) are the major bacteria-derived metabolites produced during dietary fiber fermentation by the gut microbiome. Secreted SCFAs from the gut can act at multiple sites in the periphery, affecting the immune, endocrine, and neural responses due to the vast distribution of SCFAs receptors. Therefore, it is challenging to differentiate the central and peripheral effects of SCFAs through oral and intraperitoneal administration of SCFAs. This paper presents a video-based method to interrogate the functional role of SCFAs in the brain via a guide cannula in freely moving mice. The amount and type of SCFAs in the brain can be adjusted by controlling the infusion volume and rate. This method can provide scientists with a way to appreciate the role of gut-derived metabolites in the brain.

Oral short-chain fatty acids administration regulates innate anxiety in adult microbiome-depleted mice

Anxiety is characterized by feelings of tension and worry even in the absence of threatening stimulus. Pathological condition of anxiety elicits defensive behavior and aversive reaction ultimately impacting individuals and society. The gut microbiota has been shown to contribute to the modulation of anxiety-like behavior in rodents through the gut-brain axis. Several studies observed that germ-free (GF) and the broad spectrum of antibiotic cocktail (ABX)-treated rodents display lowered anxiety-like behavior. We speculate that gut microbial short-chain fatty acids (SCFA) modulate the innate anxiety response. Herein, we administered SCFA in the drinking water in adult mice treated with ABX to deplete the microbiota and tested their anxiety-like behavior. To further augment the innate fear response, we enhanced the aversive stimulus of the anxiety-like behavior tests. Strikingly, we found that the anxiety-like behavior in ABX mice was not altered when enhanced aversive stimulus, while control and ABX mice supplemented with SCFA displayed increased anxiety-like behavior. Vagus nerve serves as a promising signaling pathway in the gut-brain axis. We determined the role of vagus nerve by subdiaphragmatic vagotomy (SDV) in ABX mice supplemented with SCFA. We found that the restored anxiety-like behavior in ABX mice by SCFA was unaffected by SDV. These findings suggest that gut microbiota can regulate anxiety-like behavior through their fermentation products SCFA.This article is part of the Special Issue on ‘Microbiome & the Brain: Mechanisms & Maladies’.

Short-chain fatty acid administration via water acidifier improves feed efficiency and modulates fecal microbiota in weaned piglets.

This study examined the effect of a water acidifier containing free and buffered short-chain fatty acids (SCFA-WA) on growth performance and microbiota of weaned piglets. In total, 192 male piglets, approximately 4 wk of age, were allocated to 24 pens (12 per treatment) with 8 piglets per pen. The piglets received either regular drinking water (negative control) or drinking water with the acidifier supplied at 2 L/1,000 L. Body weight and feed intake were measured weekly on pen level. During the first 2 wk, daily visual assessment and scoring of the feces was conducted. Fecal samples of three piglets per pen were collected on days 14 and 42 for high-throughput sequencing analysis of the microbiota. Piglets offered SCFA-WA had significantly improved feed efficiency in the third week (P = 0.025) and over the whole study period (days 0 to 42, P = 0.042) compared with piglets in the negative control group, with a strong tendency observed during the first feeding phase (days 0 to 21, P = 0.055). Furthermore, the water acidifier group had a higher water intake than piglets provided with control water during the second feeding phase (days 21 to 42, P = 0.028) and over the whole study period (days 0 to 42, P = 0.043). There was no significant difference in body weight, average daily gain, or average daily feed intake (days 0 to 21, 21 to 42, 0 to 42). Furthermore, there was no overall significant difference in fecal scoring between the treatments. In terms of the fecal microbiota response, piglets offered the water acidifier showed a significantly higher relative abundance (RA) of genus Clostridium sensu stricto 1 and a lower RA of genus Streptococcus compared to the control. Furthermore, the redundancy analysis showed a positive association between improved feed efficiency and daily weight gain and RA of Butyricicoccus and Faecalibacterium. In conclusion, consumption of the water acidifier containing free and buffered SCFA modulated the microbiota and improved feed efficiency in piglets.

Regulation of the Expression of SARS-CoV-2 Receptor Angiotensin-Converting Enzyme 2 in Nasal Mucosa.

Coronavirus disease 2019 (COVID-19) has caused a global pandemic. Higher expression of the virus receptor angiotensin-converting enzyme 2 (ACE2) in the nasal mucosa may be associated with high transmissibility and asymptomatic infection. In COVID-19, the elucidation of the determinants of ACE2 expression at nasal tissue level is crucial. The development of strategies to downregulate ACE2 expression in nasal epithelial cells might reduce transmission and be useful as a novel therapeutic approach. To verify ACE2 expression in the nasal mucosa of patients with seasonal allergic rhinitis induced by Japanese cedar pollen (SAR-JCP) and chronic rhinosinusitis with nasal polyp (CRSwNP) and to examine the effects of short-chain fatty acids (SCFAs) on ACE2 expression in airway epithelial cells. We assessed ACE2 expression in the nasal mucosa of control subjects, patients with SAR-JCP, and those with CRSwNP using real-time polymerase chain reaction. We also quantified ACE2 gene expression in cultured airway epithelial cells. Although ACE2 expression was greatly increased in a few patients with SAR-JCP during the Japanese cedar pollen season, mean levels were not significantly increased. ACE2 mRNA expression was significantly decreased in nasal polyp tissue from patients with chronic rhinosinusitis compared with the expression in that from control subjects. SCFAs generated by gastrointestinal microbiota significantly reduced resting ACE2 expression in cultured airway epithelial cells. SCFAs also significantly suppressed the dsRNA-dependent upregulation of ACE2 expression in airway epithelial cells. Inflammatory endotype affects ACE2 expression in the nasal mucosa and influences susceptibility to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. In particular, type 2 inflammation could downregulate ACE2 expression in the nasal mucosa and reduces susceptibility to SARS-CoV-2 in patients with CRSwNP. Although in vivo experiments are required, administration of SCFAs to the nasal cavity might be worthy of consideration as a preventative or therapeutic strategy for the early-stage COVID-19.

Gut Microbial Metabolite Short-Chain Fatt Acids Partially Reverse Surgery and Anesthesia-Induced Behavior Deficits in C57BL/6J Mice.

Accumulating evidence has demonstrated that damages of gut microbiota are strongly associated with central nervous system (CNS) diseases, such as perioperative neurocognitive disorders (PND). The present study investigated the role of gut microbial metabolite short-chain fatty acids (SCFAs) in surgery-induced cognitive deficits and neuroinflammation in the hippocampus. Adult male C57BL/6J mice received either SCFA mixture or saline orally for 4 weeks, and then partial hepatectomy was performed. The fecal supernatant of surgical mice was transplanted to normal mice for 3 weeks. The Morris water maze (MWM) and open-field tests were used to evaluate behavioral performance on postoperative or post-transplantation days 3 and 7. In the MWM test, pretreatment with exogenous SCFAs partially reversed surgery-induced impairments in crossing times and the time spent in the target quadrant on postoperative day 3 (p < 0.05, p < 0.05, respectively). In the open-field test, compared with the surgical mice, exogenous SCFA administration prior to surgery partially improved the locomotor activity (p < 0.05) and anxiety-like behavior (p < 0.05) on postoperative day 3. Surgical trauma and anesthesia enhanced ionized calcium-binding adapter molecule 1 (Iba-1) expression (p < 0.001), increased the levels of interleukin (IL)-1β (p < 0.001) and IL-6 (p < 0.001), and inhibited SCFA production (p < 0.001) on postoperative day 3. The expression of the brain-derived neurotrophic factor (BDNF) was also decreased (p < 0.001). Overall, surgical trauma and anesthesia exacerbated cognitive impairment, enhanced neuroinflammatory responses, and inhibited SCFA production. Pretreatment with SCFAs attenuated these effects partially by reversing microglial overactivation, inhibiting neuroinflammatory responses, and enhancing BDNF expression.