Effects of cytochalasin D on relaxation process of skinned taenia cecum and carotid artery from guinea pig.

Actin linked regulatory mechanisms are known to contribute contraction/relaxation in smooth muscle. In order to clarify whether modulation of polymerization/depolymerization of actin filaments affects relaxation process, we examined the effects of cytochalasin D on relaxation process by Ca2+ removal after Ca2+-induced contraction of ß-escin skinned (cell membrane permeabilized) taenia cecum and carotid artery preparations from guinea pigs. Cytochalasin D, an inhibitor of actin polymerization, significantly suppressed the force during relaxation both in skinned taenia cecum and carotid artery. The data fitting analysis of the relaxation processes indicates that cytochalasin D accelerates slow (latch-like) bridge dissociation. Cytochalasin D seems to directly disrupts actin filament organization or its length, resulting in modulation of actin filament structure that prevents myosin binding.

A type 4 resistant potato starch alters the cecal microbiome and gene expression in mice fed a western diet based on NHANES data.

Four major types of resistant starch (RS1-4) are present in foods, all of which can alter the microbiome and are fermented in the cecum and colon to produce short-chain fatty acids (SCFAs). Type 4 RSs are chemically modified starches, not normally found in foods, but have become a popular food additive as their addition increases fiber content. Multiple studies, in humans and rodents, have explored how different RS4 affect post-prandial glucose metabolism, but fewer studies have examined the effects of RS4 consumption on the microbiome. In addition, many RS studies conducted in rodents use high-fat diets that do not approximate what is typically consumed by humans. To address this, mice were fed a Total Western Diet (TWD), based on National Health and Nutrition Examination Survey (NHANES) data that mimics the macro and micronutrient composition of a typical American diet, for six weeks, and then supplemented with 0, 2, 5, or 10% of the RS4, Versafibe 1490™ (VF), a phosphorylated and cross-linked potato starch, for an additional three weeks. The cecal contents were analyzed for SCFA content and microbiota composition. Butyrate production was increased while branched chain SCFA production decreased. The alpha-diversity of the microbiome decreased in mice fed the TWD with 10% VF 1490 added while the beta-diversity plot showed that the 5% and 10% VF groups were distinct from mice fed the TWD. Similarly, the largest changes in relative abundance of various genera were greatest in mice fed the 10% VF diet. To examine the effect of VF consumption on tissue gene expression, cecal and distal colon tissue mRNA abundance were analyzed by RNASeq. Gene expression changes were more prevalent in the cecum than the colon and in mice fed the 10% VF diet, but the number of changes was substantially lower than we previously observed in mice fed the TWD supplemented with native potato starch (RPS). These results provide additional evidence that the structure of the RS is a major factor determining its effects on the microbiome and gene expression in the cecum and colon.

M1 cholinergic signaling in the brain modulates cytokine levels and splenic cell sub-phenotypes following cecal ligation and puncture.

BACKGROUND: The contribution of the central nervous system to sepsis pathobiology is incompletely understood. In previous studies, administration of endotoxin to mice decreased activity of the vagus anti-inflammatory reflex. Treatment with the centrally-acting M1 muscarinic acetylcholine (ACh) receptor (M1AChR) attenuated this endotoxin-mediated change. We hypothesize that decreased M1AChR-mediated activity contributes to inflammation following cecal ligation and puncture (CLP), a mouse model of sepsis. METHODS: In male C57Bl/6 mice, we quantified basal forebrain cholinergic activity (immunostaining), hippocampal neuronal activity, serum cytokine/chemokine levels (ELISA) and splenic cell subtypes (flow cytometry) at baseline, following CLP and following CLP in mice also treated with the M1AChR agonist xanomeline. RESULTS: At 48 h. post-CLP, activity in basal forebrain cells expressing choline acetyltransferase (ChAT) was half of that observed at baseline. Lower activity was also noted in the hippocampus, which contains projections from ChAT-expressing basal forebrain neurons. Serum levels of TNFα, IL-1ß, MIP-1α, IL-6, KC and G-CSF were higher post-CLP than at baseline. Post-CLP numbers of splenic macrophages and inflammatory monocytes, TNFα+ and ILß+ neutrophils and ILß+ monocytes were higher than baseline while numbers of central Dendritic Cells (cDCs), CD4+ and CD8+ T cells were lower. When, following CLP, mice were treated with xanomeline activity in basal forebrain ChAT-expressing neurons and in the hippocampus was significantly higher than in untreated animals. Post-CLP serum concentrations of TNFα, IL-1ß, and MIP-1α, but not of IL-6, KC and G-CSF, were significantly lower in xanomeline-treated mice than in untreated mice. Post-CLP numbers of splenic neutrophils, macrophages, inflammatory monocytes and TNFα+ neutrophils also were lower in xanomeline-treated mice than in untreated animals. Percentages of IL-1ß+ neutrophils, IL-1ß+ monocytes, cDCs, CD4+ T cells and CD8+ T cells were similar in xanomeline-treated and untreated post-CLP mice. CONCLUSION: Our findings indicate that M1AChR-mediated responses modulate CLP-induced alterations in serum levels of some, but not all, cytokines/chemokines and affected splenic immune response phenotypes.

Evaluating the effects of tannic acid on rabbit growth performance, digestibility, antioxidant status, intestinal morphology and caecal fermentation and microbiota.

The objective of this study was to assess the effects of dietary supplementation with tannic acid (TA) on the growth performance, digestibility, antioxidant status, intestinal morphology and the caecal fermentation and microbiota in rabbits. A total number of 120 Ira rabbits (30 days of age) were randomly allotted to four dietary treatment groups: TA 0 (control), TA 0.75, TA 1.5 and TA 3, administered basal diets with 0, 0.75, 1.5 and 3 g TA/kg of feed for 28 days. Compared to the control group, dietary 3 g TA/kg inclusion decreased the average daily feed intake (p < 0.05). No significant differences were found in the digestibility among the groups (p > 0.05). Serum total antioxidant capacity was significantly higher in the 3 g/kg TA group than in the other groups (p < 0.05). There was a significant increase in the concentration of propionic acid and butyric acid in the 3 g/kg TA group. The addition of TA had no effect on villus height and crypt depth of small intestine (p > 0.05). The 16S rRNA high-throughput sequencing results showed that at the phylum level, dietary 3 g/kg TA increased the abundance of Bacteroidetes in the caecum of rabbits (p < 0.05). Based on the results, dietary TA is effective in antioxidant capacity of rabbits, improving caecal fermentation and optimizing the caecal microflora. However, the appropriate dosage supplementation of TA in rabbits needs further research.

Rabbits (Oryctolagus cuniculus) increase caecal calcium absorption at increasing dietary calcium levels.

Hindgut fermenting herbivores from different vertebrate taxa, including tortoises, and among mammals some afrotheria, perissodactyla incl. equids, several rodents as well as lagomorphs absorb more calcium (Ca) from the digesta than they require, and excrete the surplus via urine. Both proximate and ultimate causes are elusive. It was suggested that this mechanism might ensure phosphorus availability for the hindgut microbiome by removing potentially complex-building Ca from the digesta. Here we use Ussing chamber experiments to show that rabbits (Oryctolagus cuniculus) maintained on four different diets (six animals/diet) increase active Ca absorption at increasing Ca levels. This contradicts the common assumption that at higher dietary levels, where passive uptake should be more prevalent, active transport can relax and hence supports the deliberate removal hypothesis. In the rabbits, this absorption was distinctively higher in the caecum than in the duodenum, which is unexpected in mammals. Additional quantification of the presence of two proteins involved in active Ca absorption (calbindin-D9K CB; vitamin D receptor, VDR) showed higher presence with higher dietary Ca. However, their detailed distribution across the intestinal tract and the diet groups suggests that other factors not investigated in this study must play major roles in Ca absorption in rabbits. Investigating strategies of herbivores to mitigate potential negative effects of Ca in the digesta on microbial activity and growth might represent a promising area of future research.

Single-cell profiling of the pig cecum at various developmental stages.

The gastrointestinal tract is essential for food digestion, nutrient absorption, waste elimination, and microbial defense. Single-cell transcriptome profiling of the intestinal tract has greatly enriched our understanding of cellular diversity, functional heterogeneity, and their importance in intestinal tract development and disease. Although such profiling has been extensively conducted in humans and mice, the single-cell gene expression landscape of the pig cecum remains unexplored. Here, single-cell RNA sequencing was performed on 45 572 cells obtained from seven cecal samples in pigs at four different developmental stages (days (D) 30, 42, 150, and 730). Analysis revealed 12 major cell types and 38 subtypes, as well as their distinctive genes, transcription factors, and regulons, many of which were conserved in humans. An increase in the relative proportions of CD8 + T and Granzyme A (low expression) natural killer T cells (GZMA low NKT) cells and a decrease in the relative proportions of epithelial stem cells, Tregs, RHEX + T cells, and plasmacytoid dendritic cells (pDCs) were noted across the developmental stages. Moreover, the post-weaning period exhibited an up-regulation in mitochondrial genes, COX2 and ND2, as well as genes involved in immune activation in multiple cell types. Cell-cell crosstalk analysis indicated that IBP6 + fibroblasts were the main signal senders at D30, whereas IBP6 - fibroblasts assumed this role at the other stages. NKT cells established interactions with epithelial cells and IBP6 + fibroblasts in the D730 cecum through mediation of GZMA-F2RL1/F2RL2 pairs. This study provides valuable insights into cellular heterogeneity and function in the pig cecum at different development stages.

The feasibility of partial replacement of berseem hay by spent mushroom (Pleurotus osteratus) substrate in rabbit diets on growth performance, digestibility, caecum fermentation, and economic efficiency.

Nowadays, agricultural by-product disposal is a major concern. The mushroom by-products could be used as an alternative feed source in rabbit diets. Therefore, partial replacement of berseem hay (BH) with spent mushroom substrate (SMS) was conducted in four experimental groups as follows: 0, 20, 40, and 60% of SMS. Forty weaned New Zealand White (NZW) rabbits at 6th weeks of age with an initial body weight of 520.25 ± 70.01 g were fed the experimental diet for 8 weeks. The results showed that ash content and cell wall constituents in SMS were higher than in BH, while the other nutrient compounds in SMS were lower than in BH. Dietary SMS at a level of 60% increased the final body weight (p = 0.05) and feed conversion ratio (p ≤ 0.05). However, average daily gain and total feed intake were not affected by treatments. The replacement of SMS at levels of 40 and 60% significantly improved nutrient digestibility and total digestible nutrient value. SMS 60% decreased (p = 0.040) faecal N and improved (p = 0.006) retained nitrogen. The SMS replacement increased caecum length (p = 0.001), and full and empty caecum weight (p = 0.001 and 0.021, respectively) compared to the control. The SMS inclusion caused a decrease (p = 0.021 and 0.007) in the pH and NH3-N concentrations, respectively. Total VFA, acetic acid, butyric acid, and propionic acid proportions increased with the dietary inclusion of SMS in a level-dependent manner. Using SMS as a replacement for BH in growing rabbit diets reduced the total feed cost, and consequently improved net revenue, economic efficiency, and relative economic efficiency.

Addition of soluble fiber to standard purified diets is important for gut morphology in mice.

Purified diets (PD) increase standardization and repeatability in rodent studies but lead to differences in the phenotype of animals compared to grain-based "chow" diets. PD contain less fiber and are often devoid of soluble fiber, which can impact gut health. Thus, the aim of the present study was to modify the PD AIN93G by addition of soluble fiber, to promote more natural gut development as seen with chow diets. One hundred twenty male C57BL/6J mice were fed over 12 weeks either a chow diet, AIN93G or one of three modified AIN93G with increased fiber content and different ratios of soluble fiber to cellulose. Gut health was assessed through histological and immunohistochemical parameters and gut barrier gene expression. Gut microbiota composition was analyzed and its activity characterized through short chain fatty acid (SCFA) quantification. Feeding AIN93G led to tissue atrophy, a less diverse microbiota and a lower production of SCFA compared to chow diet. The addition of soluble fiber mitigated these effects, leading to intermediate colon and caecum crypt lengths and microbiota composition compared to both control diets. In conclusion, the addition of soluble fibers in PDs seems essential for gut morphology as well as a diverse and functional gut microbiome.

Apple consumption affects cecal health by regulating 12S-hydroxy-5Z,8Z,10E,14Z-eicosatetraenoic acid (12(S)-HETE) levels through modifying the microbiota in rats.

Apples are rich in many nutrients and functional components. However, the mechanism of the effect of fresh apple consumption on rats remains unclear. In the present study, fresh apples (10 g kg-1) were added to the diet of Wistar rats, and changes in the microbiota and metabolite content of the cecum were analyzed after 28 days of feeding, and changes in the 12S-hydroxy-5Z,8Z,10E,14Z-eicosatetraenoic acid (12(S)-HETE) content and indicators related to inflammation, oxidative stress, and apoptosis were detected. Subsequently, a fecal microbiota transplantation (FMT) protocol was designed and carried out to verify the relationship between the microbiota and 12(S)-HETE, the cecal structure, and inflammatory factors. The results show that apple consumption significantly reduced the serum levels of alanine aminotransferase (ALT) and immunoglobulin G (IgG), altered the cecal histomorphology, and significantly upregulated the gene expression of claudin-1 and zonula occludens-1 (ZO-1), which encode tight junction proteins. Apple consumption also changed the structure of the cecal microbiota, increasing the abundance of some species (such as Shuttleworthia) and decreasing the abundance of others (such as Alphaproteobacteria). Metabolomic screening identified 64 significantly different metabolites. The FMT results showed that apple consumption reduced 12(S)-HETE metabolite levels in the cecal contents, improved the intestinal structure, and reduced the levels of proinflammatory factor expression by altering the cecal microbiota. In conclusion, this study provides further insight into the effects of apples on animals using rats as experimental animals. It provides basic data for future exploration of the mechanisms of the effect of apple consumption on humans.

C. difficile intoxicates neurons and pericytes to drive neurogenic inflammation.

Clostridioides difficile infection (CDI) is a major cause of healthcare-associated gastrointestinal infections1,2. The exaggerated colonic inflammation caused by C. difficile toxins such as toxin B (TcdB) damages tissues and promotes C. difficile colonization3-6, but how TcdB causes inflammation is unclear. Here we report that TcdB induces neurogenic inflammation by targeting gut-innervating afferent neurons and pericytes through receptors, including the Frizzled receptors (FZD1, FZD2 and FZD7) in neurons and chondroitin sulfate proteoglycan 4 (CSPG4) in pericytes. TcdB stimulates the secretion of the neuropeptides substance P (SP) and calcitonin gene-related peptide (CGRP) from neurons and pro-inflammatory cytokines from pericytes. Targeted delivery of the TcdB enzymatic domain, through fusion with a detoxified diphtheria toxin, into peptidergic sensory neurons that express exogeneous diphtheria toxin receptor (an approach we term toxogenetics) is sufficient to induce neurogenic inflammation and recapitulates major colonic histopathology associated with CDI. Conversely, mice lacking SP, CGRP or the SP receptor (neurokinin 1 receptor) show reduced pathology in both models of caecal TcdB injection and CDI. Blocking SP or CGRP signalling reduces tissue damage and C. difficile burden in mice infected with a standard C. difficile strain or with hypervirulent strains expressing the TcdB2 variant. Thus, targeting neurogenic inflammation provides a host-oriented therapeutic approach for treating CDI.

Impaired cecal motility and secretion alongside expansion of gut-associated lymphoid tissue in the Nlgn3R451C mouse model of autism.

Individuals with Autism Spectrum Disorder (ASD; autism) commonly present with gastrointestinal (GI) illness in addition to core diagnostic behavioural traits. The appendix, or cecum in mice, is important for GI homeostasis via its function as a key site for fermentation and a microbial reservoir. Even so, the role of the appendix and cecum in autism-associated GI symptoms remains uninvestigated. Here, we studied mice with an autism-associated missense mutation in the post-synaptic protein neuroligin-3 (Nlgn3R451C), which impacts brain and enteric neuronal activity. We assessed for changes in cecal motility using a tri-cannulation video-imaging approach in ex vivo preparations from wild-type and Nlgn3R451C mice. We investigated cecal permeability and neurally-evoked secretion in wild-type and Nlgn3R451C tissues using an Ussing chamber set-up. The number of cecal patches in fresh tissue samples were assessed and key immune populations including gut macrophages and dendritic cells were visualised using immunofluorescence. Nlgn3R451C mice displayed accelerated cecal motor complexes and reduced cecal weight in comparison to wildtype littermates. Nlgn3R451C mice also demonstrated reduced neurally-evoked cecal secretion in response to the nicotinic acetylcholine receptor agonist 1,1-dimethyl-4-phenylpiperazinium (DMPP), but permeability was unchanged. We observed an increase in the number of cecal patches in Nlgn3R451C mice, however the cellular morphologies of key immune populations studied were not significantly altered. We show that the R451C nervous system mutation leads to cecal dysmotility, impaired secretion, and neuro-immune alterations. Together, these results suggest that the R451C mutation disrupts the gut-brain axis with GI dysfunction in autism.

Extremely hair follicle density is associated with a significantly different cecal microbiota in rex rabbits.

It has become increasingly clear that gut microbiota and skin are interconnected since the discovery of the 'gut-brain-skin' axis. Hair follicles (HFs) are skin microorganisms, but few studies have investigated their relationship to gut microbiota. Hence, we hypothesize that HFs have a close relationship with the gut, similarly to what was reported for the skin. Using rex rabbits as an animal model, one hundred healthy half-sibling rex rabbits were selected for the experiment, and 16 s rRNA gene sequencing was performed on the cecal microbiota of nine rabbits with the extremely high (HS) and low (LS) hair density (n = 9 per group) to determine differences between the composition and function of these communities. In comparison with the LS group, several alpha diversity index values were significantly lower in the HS group, although the higher variation in species composition in the HS group. Additionally, species diversity and abundance differed significantly in the cecum microbiota of HS and LS rabbits. Further, primary and secondary HF density was significantly correlated with the families Muribaculaceae and Bacteroidaceae, and genera Blautia, Bacteroides and Desulfovibrio. In particular, Muribaculaceae, Bacteroidaceae, Blautia and Bacteroides may support the development of HFs. Moreover, the expression of WNT4, WNT10a, WNT10b, CTNNB1 (ß-catenin) and LEF1 in the skin was significantly higher in the HS group compared with the LS group. Altogether, the results of this study suggest that the extremely high density of HF in rabbits is associated with a significantly different microbiota diversity and community structure, and the Wnt/ß-catenin signalling pathway was activated in the HS group. Thus, key bacteria may promote the development of HF.

Consumption of 1-Kestose Upregulates MicroRNA-200 and -192/215 Families in Lamina Propria Leukocytes of the Murine Large Intestine.

By comparing germ-free mice and specific pathogen-free mice, we recently demonstrated that the presence of gut commensals upregulates microRNA-200 family members in lamina propria leukocytes (LPL) of the murine large intestine. The present study tested whether the consumption of 1-kestose (KES), an indigestible oligosaccharide that alters gut microbiota composition, influences the microRNA expression in the LPL. Supplementation of KES (4%) in drinking water for 2 wk increased the levels of miR-182-5p, -205-5p, -290a-5p, miR-200 family members (miR-141-3p, -200a-3p, -200b-3p, -200c-3p, and -429-3p) as well as miR-192/215 family members (miR-192-5p, -194-5p, and -215-5p) as determined by microarray analysis in large intestinal LPL of C57BL/6 mice. Quantitative reverse transcription-PCR further confirmed the increase in miR-192-5p, -194-5p, -200a-3p, -200b-3p, -200c-3p, -205-5p, and 215-5p. KES consumption significantly increased Bifidobacterium pseudolongum in the cecal contents. In a separate experiment, intragastric administration of B. pseudolongum (109 CFU/d) for 7 d increased the levels of miR-182-5p, -194-5p, and -200a-3p and tended to increase the levels of miR-200b-3p, -215-5p, and -429-3p. These results suggest that dietary KES influences miRNA expression in the large intestinal LPL, which may be associated with the increased population of B. pseudolongum.

Comparison of the effect of quercetin and daidzein on production performance, anti-oxidation, hormones, and cecal microflora in laying hens during the late laying period.

This study aims to compare the effect of quercetin and daidzein on production performance, anti-oxidation, hormones, and cecal microflora in laying hens during the late laying period. A total of 360 53-week-old healthy Hyline brown laying hens were randomly divided into 3 groups (control, 0.05% quercetin, and 0.003% daidzein). Diets were fed for 10 wk, afterwards 1 bird per replicate (6 replicates) were euthanized for sampling blood, liver and cecal digesta. Compared with the control, quercetin significantly increased laying rate and decreased feed-to-egg weight ratio from wk 1 to 4, wk 5 to 10, and wk 1 to 10 (P < 0.05). Quercetin significantly increased the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) and decreased catalase (CAT) activity and malondialdehyde (MDA) content in serum and liver (P < 0.05) and increased content of total antioxidant capacity (T-AOC) in liver (P < 0.05). Quercetin increased content of estradiol (E2), luteinizing hormone (LH), follicle-stimulating hormone (FSH), growth hormone (GH), insulin-like growth factor 1 (IGF-1), triiodothyronine (T3) and thyroxine (T4) in serum (P < 0.05). Quercetin significantly decreased the relative abundance of Bacteroidaceae and Bacteroides (P < 0.01) and significantly increased the relative abundance of Lactobacillaceae and Lactobacillus (P < 0.05) at family and genus levels in cecum. Daidzein did not significantly influence production performance from wk 1 to 10. Daidzein significantly increased SOD activity and decreased CAT activity and MDA content in serum and liver (P < 0.05), and increased T-AOC content in liver (P < 0.05). Daidzein increased content of FSH, IGF-1, T3 in serum (P < 0.05). Daidzein increased the relative abundance of Rikenellaceae RC9 gut group at genus level in cecum (P < 0.05). Quercetin increased economic efficiency by 137.59% and 8.77%, respectively, compared with daidzein and control. In conclusion, quercetin improved production performance through enhancing antioxidant state, hormone levels, and regulating cecal microflora in laying hens during the late laying period. Quercetin was more effective than daidzein in improving economic efficiency.

An in vitro model for caecal proteolytic fermentation potential of ingredients in broilers.

Fermentation of protein in the caeca of chickens may lead to the production of potentially detrimental metabolites, which can reduce gut health. A poor precaecal digestion is expected to increase protein fermentation (PF), as more proteins are likely to enter the caeca. It is unknown if the undigested protein that enters the caeca differs in fermentability depending on their ingredient source. In order to predict which feed ingredients increase the risk of PF, an in vitro procedure was developed, which simulates the gastric and enteric digestion, subsequent caecal fermentation. After digestion, amino acids and peptides smaller than 3.5 kD in the soluble fraction were removed by means of dialysis. These amino acids and peptides are assumed to be hydrolysed and absorbed in the small intestine of poultry and therefore not used in the fermentation assay. The remaining soluble and fine digesta fractions were inoculated with caecal microbes. In chicken, the soluble and fine fractions enter the caeca, to be fermented, while insoluble and coarse fractions bypass them. The inoculum was made N-free to ensure bacteria would require the N from the digesta fractions for their growth and activity. The gas production (GP) from the inoculum, therefore, reflected the ability of bacteria to use N from substrates and was an indirect measure for PF. The Maximum GP rate of ingredients averaged 21.3 ± 0.9 ml/h (mean ± SEM) and was in some cases more rapid than the positive control (urea, maximum GP rate = 16.5 ml/h). Only small differences in GP kinetics were found between protein ingredients. Branched-chain fatty acids and ammonia concentrations in the fermentation fluid after 24 hours showed no differences between ingredients. Results indicate that solubilised undigested proteins larger than 3.5 kD are rapidly fermented independent of its source when an equal amount of N is present.

The effect of claudin-15 deletion on cationic selectivity and transport in paracellular pathways of the cecum and large intestine.

The large intestine plays a pivotal role in water and electrolyte balance. Paracellular transport may play a role in ion transport mechanisms in the cecum and large intestine; however, these molecular mechanisms and their physiological roles have not been fully studied. Claudin-15 forms a cation channel in tight junctions in the small intestine, but its role in the cecum and large intestine has not been investigated. This study aimed to explore the physiological role of claudin-15 in the cecum and large intestine using claudin-15 (Cldn15) KO mice. Electrical conductance, short-circuit current, Na+ flux, and dilution potential were assessed in isolated tissue preparations mounted in Ussing chambers. The induced short-circuit current of short-chain fatty acids, which are fermentative products in the intestinal tract, was also measured. Compared to wild type mice, the electrical conductance and paracellular Na+ flux was decreased in the cecum, but not the middle large intestine, while in both the cecum and the middle large intestine, paracellular Na+ permeability was decreased in Cldn15 KO mice. These results suggest that claudin-15 is responsible for Na+ permeability in the tight junctions of the cecum and large intestine and decreased Na+ permeability in the cecum may cause impaired absorption function.

Microorganisms Involved in Hydrogen Sink in the Gastrointestinal Tract of Chickens.

Hydrogen sink is a beneficial process, which has never been properly examined in chickens. Therefore, the aim of this study was to assess the quantity and quality of microbiota involved in hydrogen uptake with the use of real-time PCR and metagenome sequencing. Analyses were carried out in 50 free-range chickens, 50 commercial broilers, and 54 experimental chickens isolated from external factors. The median values of acetogens, methanogens, sulfate-reducing bacteria (SRB), and [NiFe]-hydrogenase utilizers measured in the cecum were approx. 7.6, 0, 0, and 3.2 log10/gram of wet weight, respectively. For the excreta samples, these values were 5.9, 4.8, 4, and 3 log10/gram of wet weight, respectively. Our results showed that the acetogens were dominant over the other tested groups of hydrogen consumers. The quantities of methanogens, SRB, and the [NiFe]-hydrogenase utilizers were dependent on the overall rearing conditions, being the result of diet, environment, agrotechnical measures, and other factors combined. By sequencing of the 16S rRNA gene, archaea of the genus Methanomassiliicoccus (Candidatus Methanomassiliicoccus) were discovered in chickens for the first time. This study provides some indication that in chickens, acetogenesis may be the main metabolic pathway responsible for hydrogen sink.

Altered Lung Heat Shock Protein-70 Expression and Severity of Sepsis-Induced Acute Lung Injury in a Chronic Kidney Disease Rat Model.

Enhanced heat shock protein-70 (HSP-70) expression in the lungs is associated with attenuated acute lung injury (ALI) in a sepsis model. Chronic kidney disease (CKD) significantly contributes to the poor prognosis of patients with sepsis. This study examined the relationship between sepsis-induced ALI severity and altered lung HSP-70 expression in CKD. Experimental rats underwent a sham operation (control group) or 5/6 nephrectomy (CKD group). Sepsis was induced with cecal ligation and puncture (CLP). Laboratory tests and lung harvest were performed in the control group (without CLP and after 3, 12, 24, and 72 h of CLP) and in the CKD group (without CLP and after 72 h of CLP). ALI was the most severe after 12 h of sepsis. The mean lung injury score at 72 h after sepsis was significantly higher in the CKD group than in the control group (4.38 versus 3.30, p < 0.01). Nonetheless, enhanced lung HSP-70 expression was not observed in the CKD group. This study shows that altered lung HSP-70 expression is associated with the worsening of sepsis-induced ALI in patients with CKD. Enhancing lung HSP-70 is a novel treatment target for patients with CKD and sepsis-induced ALI.

Aggressive alternative splicing events discovered in cecum ligation and puncture induced lung injury.

AIMS: Acute lung injury (ALI) induced by sepsis and its complications have high morbidity and mortality rates globally. The objective of this study was to enhance our understanding of the underlying mechanism of ALI by identifying potential splicing events that are regulated in this condition. MATERIALS AND METHODS: The CLP mouse model was utilized for mRNA sequencing, and the expression and splicing data were analyzed. Verification of the changes in expression and splicing induced by CLP was conducted using qPCR and RT-PCR. RESULTS: Our results showed that splicing-related genes were regulated, suggesting that splicing regulation may be a key mechanism in ALI. We also found that more than 2900 genes displayed alternative splicing in the lungs of mice with sepsis. Using RT-PCR, we verified that TLR4 and other genes had differential splicing isoforms in the lungs of mice with sepsis. We confirmed the presence of TLR4-s in the lungs of mice with sepsis using RNA-fluorescence in situ hybridization. CONCLUSION: Our results suggest that sepsis-induced ALI can significantly alter splicing in the lungs of mice. The list of DASGs and splicing factors is valuable for further study in the search for new treatment approaches for sepsis-induced ALI.

Downregulation of the gene expression of Cyp2c29 and Cyp3a11 by cecal ligation and puncture-induced sepsis is associated with interleukin-6.

Sepsis is a pathological condition that affects the metabolism of administered drugs, leading to changes in the duration and intensity of their intended efficacies. Proinflammatory cytokines downregulate the expression of cytochrome P450s (P450s). The effects of P450 expression under inflammatory conditions have been studied using prophlogistic substances such as lipopolysaccharide; however, few studies have focused on clinical models of sepsis. Here, we show that cecal ligation and puncture (CLP), an approach for the study of human polymicrobial sepsis, leads to the expression of interleukin-1ß (IL-1ß), IL-6, and tumor necrosis factor α (TNFα) at 24 h after the CLP operation. Following CLP, IL-6-/- mice exhibited markedly lower survival than WT mice. In addition, CLP led to the significant downregulation of Cyp2c29 and Cyp3a11 gene expression in IL-1α-/-/ß-/- (IL-1-/-) and TNFα-/- mice as well as in WT mice. In contrast, CLP elicited no significant effect on Cyp3a11 expression in IL-6-/- mice. Although CLP reduced the Cyp2c29 expression level in IL-6-/- mice, the expression of Cyp2c29 was lower in CLP-operated WT mice than in CLP-operated IL-6-/- mice. The reduction in the respective P450 protein levels and activities due to CLP-induced sepsis, reflected in the mRNA expression levels, was abolished by IL-6 depletion. Thus, CLP-induced sepsis downregulates P450 gene expression, particularly Cyp2c expression, and this effect is associated with IL-6 without affecting resistance to CLP-induced sepsis. These findings demonstrate the usefulness of CLP for studying the regulation of P450s and highlight IL-6 as a potential indicator of drug-metabolizing capacity under septic conditions.