Resolvin D1 alleviates apoptosis triggered by endoplasmic reticulum stress in IPEC-J2 cells.

BACKGROUND: Resolvin D1 (RvD1), a specialized pro-resolving lipid mediator (SPM), is derived from docosahexaenoic acid (DHA). It plays a key role in actively resolving inflammatory responses, which further reduces small intestinal damage. However, its regulation of the apoptosis triggered by endoplasmic reticulum (ER) stress in intestinal epithelial cells is still poorly understood. The intestinal porcine epithelial cells (IPEC-J2) were stimulated with tunicamycin to screen an optimal stimulation time and concentration to establish an ER stress model. Meanwhile, RvD1 (0, 1, 10, 20, and 50 nM) cytotoxicity and its impact on cell viability and the effective concentration for reducing ER stress and apoptosis were determined. Finally, the effects of RvD1 on ER stress and associated apoptosis were furtherly explored by flow cytometry analysis, AO/EB staining, RT-qPCR, and western blotting. RESULTS: The ER stress model of IPEC-J2 cells was successfully built by stimulating the cells with 1 µg/mL tunicamycin for 9 h. Certainly, the increased apoptosis and cell viability inhibition also appeared under the ER stress condition. RvD1 had no cytotoxicity, and its concentration of 1 nM significantly decreased cell viability inhibition (p= 0.0154) and the total apoptosis rate of the cells from 14.13 to 10.00% (p= 0.0000). RvD1 at the concentration of 1 nM also significantly reduced the expression of glucose-regulated protein 78 (GRP-78, an ER stress marker gene) (p= 0.0000) and pro-apoptotic gene Caspase-3 (p= 0.0368) and promoted the expression of B cell lymphoma 2 (Bcl-2, an anti-apoptotic gene)(p= 0.0008). CONCLUSIONS: Collectively, the results shed light on the potential of RvD1 for alleviating apoptosis triggered by ER stress, which may indicate an essential role of RvD1 in maintaining intestinal health and homeostasis.

Probiotic Potential and Safety Assessment of Lactiplantibacillus plantarum cqf-43 and Whole-Genome Sequence Analysis.

This study reports the whole-genome sequence of Lactiplantibacillus plantarum cqf-43 isolated from healthy sow feces. Based on genomic analysis, we performed a comprehensive safety assessment of strain cqf-43, using both in vitro and in vivo experiments, and explored its probiotic potential. The total genome length measures 3,169,201 bp, boasting a GC content of 44.59%. Through phylogenetic analyses, leveraging both 16S rRNA gene and whole-genome sequences, we confidently categorize strain cqf-43 as a member of Lactiplantibacillus. Genome annotation using Prokka unveiled a total of 3141 genes, encompassing 2990 protein-coding sequences, 71 tRNAs, 16 rRNAs, and 1 tmRNA. Functional annotations derived from COG and KEGG databases highlighted a significant abundance of genes related to metabolism, with a notable emphasis on carbohydrate utilization. The genome also revealed the presence of prophage regions and CRISPR-Cas regions while lacking virulence and toxin genes. Screening for antibiotic resistance genes via the CARD database yielded no detectable transferable resistance genes, effectively eliminating the potential for harmful gene transfer. It is worth highlighting that the virulence factors identified via the VFDB database primarily contribute to bolstering pathogen resilience in hostile environments. This characteristic is particularly advantageous for probiotics. Furthermore, the genome is devoid of menacing genes such as hemolysin, gelatinase, and biogenic amine-producing genes. Our investigation also unveiled the presence of three unannotated secondary metabolite biosynthetic gene clusters, as detected by the online tool antiSMASH, suggesting a great deal of unknown potential for this strain. Rigorous in vitro experiments confirmed tolerance of strain cqf-43 in the intestinal environment, its antimicrobial efficacy, sensitivity to antibiotics, absence of hemolysis and gelatinase activity, and its inability to produce biogenic amines. In addition, a 28-day oral toxicity test showed that the strain cqf-43 did not pose a health hazard in mice, further establishing it as a safe strain.

Growth performance, bile acid profile, fecal microbiome and serum metabolomics of growing-finishing pigs fed diets with bile acids supplementation.

The present experiment was conducted to determine the effect of bile acids (BAs) supplementation on growth performance, BAs profile, fecal microbiome, and serum metabolomics in growing-finishing pigs. A total of 60 pigs [Duroc × (Landrace × Yorkshire)] with an average body weight of 27.0 ±â€…1.5 kg were selected and allotted into one of 2 groups (castrated male to female ratio = 1:1), with 10 replicates per treatment and 3 pigs per replicate. The 2 treatments were the control group (control) and a porcine bile extract-supplemented group dosed at 0.5 g/kg feed (BA). After a 16-wk treatment, growth performance, BAs profiles in serum and feces, and fecal microbial composition were determined. An untargeted metabolomics approach using gas chromatography with a time-of-flight mass spectrometer was conducted to identify the metabolic pathways and associated metabolites in the serum of pigs. We found that BAs supplementation had no effect on the growth performance of the growing-finishing pig. However, it tended to increase the gain-to-feed ratio for the whole period (P = 0.07). BAs supplementation resulted in elevated serum concentrations of secondary bile acids, including hyodeoxycholic acid (HDCA), glycoursodeoxycholic acid, and tauro-hyodeoxycholic acid, as well as fecal concentration of HDCA (P < 0.05). Fecal microbiota analysis revealed no differences in alpha and beta diversity indices or the relative abundance of operational taxonomic units (OTUs) at both phylum and genus levels between groups. Metabolic pathway analysis revealed that the differential metabolites between control and BA groups are mainly involved in purine metabolism, ether lipid metabolism, glycerophospholipid metabolism, and amino sugar and nucleotide sugar metabolism, as well as primary bile acid biosynthesis. Our findings indicate that BAs supplementation tended to improve the feed efficiency, and significantly altered the BA profile in the serum and feces of growing-finished pigs, regardless of any changes in the gut microbial composition. The altered metabolic pathways could potentially play a vital role in improving the feed efficiency of growing-finished pigs with BAs supplementation.

Bile acids (BAs), known to exhibit a key role in emulsification and absorption of dietary fat in the intestinal lumen, have also become appreciated as important regulators of intestinal function, lipid and energy metabolism in humans and animals. This study investigated the effect of BAs supplementation on growth performance, BAs profile, fecal microbiome, and serum metabolomics in growing-finishing pigs. The results showed that BAs supplementation had few effects on pig growth performance and fecal microbiota, but modified serum and fecal BAs profile and serum metabolomics profile. The altered metabolic pathways could potentially play a vital role in improving the feed efficiency of growing-finished pigs with BAs supplementation.

MDMA targets miR-124/MEKK3 via MALAT1 to promote Parkinson's disease progression.

BACKGROUND: Parkinson's disease (PD) is a well-known neurodegenerative disease that is usually caused by the progressive loss of dopamine neurons and the formation of Lewy vesicles. 3,4-Methylenedioxymethamphetamine (MDMA) has been reported to cause damage to human substantia nigra neurons and an increased risk of PD, but the exact molecular mechanisms need further investigation. METHODS: MPTP- and MPP+-induced PD cells and animal models were treated with Nissl staining to assess neuronal damage in the substantia nigra (SN) area; immunohistochemistry to detect TH expression in the SN; TUNEL staining to detect apoptosis in the SN area; Western blotting to detect the inflammatory factors NF-κB, TNF-α, IL-6 and mitogen-activated protein kinase kinase kinase 3 (MEKK3); Griess assay for NO; RT‒qPCR for metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) and miR-124 expression; Cell proliferation was assessed by CCK-8. Dual luciferase reporter genes were used to verify targeting relationships. RESULTS: MDMA promoted MALAT1 expression, and knockdown of MALAT1 alleviated the MDMA-induced inhibition of SH-SY5Y cell proliferation, inflammation, NO release, SN neuronal injury, and TH expression inhibition. Both inhibition of miR-124 and overexpression of MEKK3 reversed the neuroprotective effects exhibited by knockdown of MALAT1. CONCLUSION: MDMA promotes MALAT1 expression and inhibits the targeted downregulation of MEKK3 by miR-124, resulting in upregulation of the expression of MEKK3 and finally jointly promoting PD progression.

Mesenchymal stem cell exosomes rich in miR-23b-3p affect the Wnt signaling pathway and promote neuronal autophagy to alleviate PD symptoms.

This study aims to elucidate the role of miR-23b-3p in mesenchymal stem cell exosomes in regulating the Wnt signaling pathway to promote autophagy of neurons and alleviate Parkinson's disease (PD) symptoms. We generated rat and cellular PD models with 6-OHDA, treated them with mesenchymal stem cell exosomes rich in miR-23b-3p and determined the expression of α-syn and Wnt/ß-catenin pathway and autophagy-related genes. In the plasma of PD patients, the levels of miR-23b-3p and the Wnt/ß-catenin pathway-related genes ß-catenin and DAT were low, while α-syn expression was high. In the PD cell model, miR-23b-3p was downregulated, the Wnt pathway was inhibited, α-syn was upregulated, neuron autophagy was inhibited, and the revitalization of the Wnt/ß-catenin pathway could promote the autophagy of neurons. Coculture of miR-23b-3p-enriched exosomes with MN9D cells confirmed that miR-23b-3p-enriched exosomes could promote autophagy in MN9D cells in a PD cell model. Moreover, animal experiments confirmed the results of the cell experiments. Therefore, miR-23b-3p-enriched mesenchymal stem cell exosomes promote neuronal autophagy by regulating the Wnt signaling pathway, thus alleviating PD progression and providing an important basis for the clinical treatment of PD.

BDNF alleviates Parkinson's disease by promoting STAT3 phosphorylation and regulating neuronal autophagy.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by the gradual death of dopaminergic neurons. Brain-derived neurotrophic factor (BDNF) and its receptors are widely distributed throughout the central nervous system, which can promote the survival and growth of neurons and protect neurons. This study revealed that BDNF promotes STAT3 phosphorylation and regulates autophagy in neurons. The PD mouse model was established by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Moreover, SH-SY5Y cells were treated with 1-methyl-4-phenyl-pyridinium (MPP+) to establish a PD cell model. The level of BDNF was low in PD model mice and SH-SY5Y cells treated with MPP+. BDNF enhanced the levels of p-TrkB, P-STAT3, PINK1, and DJ-1. BDNF promoted autophagy, inhibited the level of p-α-syn (Ser129) and enhanced cell proliferation. The autophagy inhibitor 3-Methyladenine (3-methyladenine, 3-MA) reversed the protective effects of BDNF on neurons. BiFC assay results showed that there was a direct physical interaction between BDNF and STAT3, and coimmunoprecipitation experiments indicated an interaction between STAT3 and PI3K. The PI3K agonist Recilisib activated the PI3K/AKT/mTOR pathway, promoted autophagy, and alleviated neuronal cell damage. BDNF alleviates PD pathology by promoting STAT3 phosphorylation and regulating neuronal autophagy in SH-SY5Y cells and cultured primary neurons. Finally, BDNF has neuroprotective effects on PD model mice.

Dietary Supplementation of Lactobacillus johnsonii RS-7 Improved Antioxidant and Immune Function of Weaned Piglets.

We investigated the effects of dietary supplementation of lactic acid bacteria on the immune and antioxidant performance of weaned pigs. A total of 128 Duroc × Landrace × Yorkshire piglets weaned on day 28 with an average body weight of 8.95 ± 1.15 kg were selected and randomly divided into four treatment groups according to body weight and sex for a 28-day study. The four dietary treatments were basal diet (CON), and CON with 0.05% (LJ0.05), 0.1% (LJ0.1), and 0.2% (LJ0.2) Lactobacillus johnsonii RS-7, respectively. The lowest feed-to-gain ratio (F:G) was found when LJ0.1 was added to the diet. The addition of compound lactic acid bacteria to the diet increased the concentrations of TP, ALB, IgA, and IgM on day 14 and IgG, IgA, and IgM on day 28 (p < 0.05) in the blood, with trait values greater for pigs fed LJ0.1 than CON pigs (p < 0.05). Concentrations of antioxidants (CAT, T-AOC, MDA, T-SOD, and GSH) in serum, intestinal mucosa, spleen, liver, and pancreas improved. In summary, dietary supplementation of Lactobacillus johnsonii RS-7 improved the antioxidant and immune function of weaned piglets.

MALAT1 Mediates α-Synuclein Expression through miR-23b-3p to Induce Autophagic Impairment and the Inflammatory Response in Microglia to Promote Apoptosis in Dopaminergic Neuronal Cells.

Background: Parkinson's disease (PD) is a very common neurodegenerative disease that adversely affects the physical and mental health of many patients, but there is currently no effective treatment. Objective: To this end, this study focused on investigating the potential mechanisms leading to dopaminergic neuronal apoptosis in PD. Methods: Rotenone induces damage in dopaminergic neuronal MN9D cells. Apoptosis was detected by flow cytometry, and the expression of apoptosis-related proteins was detected by western blot. RT-qPCR was used to detect the expression of MALAT1 and miR-23b-3p. The expression of α-synuclein was detected by ELISA. A dual luciferase gene reporter assay was used to determine the targeted regulatory relationship between MALAT1 and miR-23b-3p and miR-23b-3p and α-synuclein. MN9D supernatant was cocultured with BV-2 cells, or BV-2 cells were treated with exogenous α-synuclein and then treated with an autophagy inhibitor (3-MA) and autophagy activator (RAPA). The expression of α-synuclein in BV-2 cells was detected by immunofluorescence. The expression of MIP-1α, a marker of microglial activation, was detected by ELISA. The nuclear translocation of NF-κB p65 was detected by immunofluorescence. The expression of proinflammatory cytokines was detected by ELISA. Western blotting was used to detect the expression of autophagy-related proteins. Apoptosis of MN9D cells was detected after coculture of BV-2 supernatant with MN9D. Results: The expression of MALAT1 and α-synuclein was upregulated, while the expression of miR-23b-3p was downregulated in damaged MN9D cells, resulting in cell apoptosis. MALAT1 can negatively regulate the expression of miR-23b-3p, while miR-23b-3p negatively regulates the expression of α-synuclein. α-synuclein can enter BV-2 cells through cell phagocytosis. Coculture of BV-2 cells with α-synuclein or with MN9D supernatant overexpressing MALAT1 resulted in a decrease in the autophagy level of BV-2 cells and an inflammatory reaction. However, miR-23b-3p mimics and knockdown of α-synuclein reversed the effect of MALAT1 on autophagy and the inflammatory response of BV-2 cells. In addition, after coculture of BV-2 cells with α-synuclein, the level of autophagy further decreased when 3-MA was added, while the opposite result occurred when RAPA was added. After coculture of α-synuclein-treated BV-2 cell supernatant with MN9D cells, autophagy-impaired BV-2 promoted the apoptosis of MN9D cells, and 3-MA aggravated the autophagy disorder of BV-2 and further promoted the apoptosis of MN9D cells, while RAPA reversed the autophagy disorder of BV-2 and alleviated the apoptosis of MN9D cells. Conclusion: MALAT1 can promote α-synuclein expression by regulating miR-23b-3p, thereby inducing microglial autophagy disorder and an inflammatory response leading to apoptosis of dopaminergic neurons. This newly discovered molecular mechanism may provide a potential target for the treatment of PD.

Alteration of gut microbiome and metabolome by Clostridium butyricum can repair the intestinal dysbiosis caused by antibiotics in mice.

This study evaluated the repair effects of Clostridium butyricum (CBX 2021) on the antibiotic (ABX)-induced intestinal dysbiosis in mice by the multi-omics method. Results showed that ABX eliminated more than 90% of cecal bacteria and also exerted adverse effects on the intestinal structure and overall health in mice after 10 days of the treatment. Of interest, supplementing CBX 2021 in the mice for the next 10 days colonized more butyrate-producing bacteria and accelerated butyrate production compared with the mice by natural recovery. The reconstruction of intestinal microbiota efficiently promoted the improvement of the damaged gut morphology and physical barrier in the mice. In addition, CBX 2021 significantly reduced the content of disease-related metabolites and meanwhile promoted carbohydrate digestion and absorption in mice followed the microbiome alternation. In conclusion, CBX 2021 can repair the intestinal ecology of mice damaged by the antibiotics through reconstructing gut microbiota and optimizing metabolic functions.

Soybean oligosaccharides combined with probiotics reduce faecal odour compound content by improving intestinal microbiota in pigs.

As a potential prebiotic, soybean oligosaccharides (SBOS) can improve animal health by modulating gut microbiota. The aim of this study was to investigate the different effects of supplementing SBOS and supplementing SBOS plus probiotic on the growth and health of pigs. Three groups of growing pigs (n = 12) were fed with basal diet (Control), basal diet + 0.5% SBOS (SBOS), or basal diet +0.5% SBOS + 0.1% compound probiotics (SOP) for 42 days. Results showed that SBOS and SOP treatments had positive effects on the pigs in the experiment, and the latter was more effective. Compared with the control pigs, the average daily gain of SBOS group and SOP group slightly increased, SOP significantly increased the serum levels of growth hormone and thyroid hormone T3. Importantly, serum concentrations of immunoglobulin (IgA, IgG and IgM), total antioxidant capacity and superoxide dismutase in both treatments were increased significantly, SOP group most. Moreover, the faecal odour compounds of pigs, especially skatole, were significantly reduced by the treatments. Additionally, SOP significantly increased the diversity and richness of the faecal microbiota, both the treatments increased genera of norank_f_Muribaculaceae and Ruminococcaceae but reduced Lactobacillus. Correlation analysis indicated that Lactobacillus was significantly positively correlated with odour compounds, while Ruminococcaceae was the opposite. Conclusively, synbiotics combined with SBOS and probiotics had stronger promotion effects on the growth and health of pigs.

Naringin reduces fat deposition by promoting the expression of lipolysis and ß-oxidation related genes.

AIMS: Naringin, a flavonoid present in citrus fruits, has been known for the capacity to reduce lipid synthesis and anti-inflammatory. In this study, we investigated whether naringin increases lipolysis and fatty acid ß-oxidation to change fat deposition. METHODS: In in vivo experiment, obese adult mice (20-weeks-old, n = 18) were divided into control group fed with normal diet and naringin-treated group fed with naringin-supplemented diet (5 g/kg) for 60 days, respectively. In in vitro experiment, differentiated 3T3-L1 adipocytes were treated for four days with or without naringin (100 µg/mL). RESULTS: Supplementing naringin significantly reduced the body weight, abdominal fat weight, blood total cholesterol content of mice, but did not affect food intake. In addition, naringin decreased levels of pro-inflammatory factors in adipose tissue including interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and monocyte chemotactic protein 1 (MCP-1). Naringin increased the expression of AMP-activated protein kinase (AMPK), a key factor in cellular energy metabolism, and raised the ratio of p-AMPK/AMPK in mouse liver tissue. The protein expression of hormone-sensitive lipase (HSL), phospho-HSL563 (p-HSL563), p-HSL563/HSL, and adipocyte triglyceride lipase (ATGL) was significantly increased in the adipose tissue of naringin-treated mice. Furthermore, naringin enhanced the expression of fatty acid ß-oxidation genes, including carnitine palmitoyl transferase 1 (CPT1), uncoupling protein 2 (UCP2), and acyl-coenzyme A oxidase 1 (AOX1) in mouse adipose tissue. In in vitro experiment, similar findings were observed in differentiated 3T3-L1 adipocytes with naringin treatment. The treatment remarkably reduced intracellular lipid content, increased the number of mitochondria and promoted the gene expression of HSL, ATGL, CPT1, AOX1, and UCP2 and the phosphorylation of HSL protein. CONCLUSION: Naringin reduced body fat in obese mice and lipid content in differentiated 3T3-L1 adipocytes, which was associated with enhanced AMPK activation and upregulation of the expression of the lipolytic genes HSL, ATGL, and ß-oxidation genes CPT1, AOX1, and UCP2.

Preparation of gastrodin-modified dendrimer-entrapped gold nanoparticles as a drug delivery system for cerebral ischemia-reperfusion injury.

OBJECTIVE: This study sought to evaluate the feasibility of multifunctional gastrodin (GAS)-containing nano-drug carrier system against cerebral ischemia-reperfusion injury (CIRI). METHODS: The drug-loaded nanocomposite (Au-G5.NHAc-PS/GAS) with certain encapsulation efficiency (EE) was prepared by physical adsorption method using different proportions of GAS and drug-carrying system (Au-G5.NHAc-PS). High-performance liquid chromatography was used to determine the drug loading and EE. Cultured rat astrocytes and hypothalamic neurons were assigned into four groups: PBS, Au-G5.NHAc-PS, Au-G5.NHAc-PS/GAS, and GAS. CCK-8 assay, flow cytometry, and quantitative real-time PCR were performed to examine the cell viability, apoptosis, and the expression of tumor necrosis factor-α (TNF-α), IL-1ß, and IL-6 in the astrocytes and hypothalamic neurons, respectively. Cellular uptake of GAS and Au-G5.NHAc-PS/GAS was analyzed by using Hoechst 33342 staining. The animal model with focal cerebral ischemia was generated by middle cerebral artery occlusion (MCAO) in healthy male Sprague Dawley (SD) rats, and pathological changes of brain tissue and major organs in the rats were identified by hematoxylin and eosin (HE) staining. Apoptosis in rat astrocytes and hypothalamic neurons was detected by TUNEL staining and flow cytometry. RESULTS: Au-G5.NHAc-PS had a spherical shape with a uniform size of 157.3 nm. Among the nanoparticles, Au-G5.NHAc-PS/GAS with an EE of 70.3% displayed the best release delay effect. Moreover, we observed that in vitro cytotoxicity and cellular uptake of Au-G5.NHAc-PS/GAS were higher than those of GAS, whereas the expression of TNF-α, IL-1ß, and IL-6 was significantly downregulated in Au-G5.NHAc-PS/GAS group as compared to G5.NHAc-PS group. Notably, HE staining revealed that although Au-G5.NHAc-PS/GAS had no toxic and side effects on the main organs of rats, it alleviated the damage of brain tissue in the MCAO rats. Besides, Au-G5.NHAc/GAS markedly reduced MCAO-induced apoptosis. CONCLUSION: Au-G5.NHAc-PS showed favorable surface morphology, sustained drug release ability, no measurable toxicity, and good biocompatibility, indicating that GAS exerts anti-inflammatory and antiapoptotic effects on CIRI.

The effects of gut microbiota colonizing on the porcine hypothalamus revealed by whole transcriptome analysis.

The roles of the microbe-gut-brain axis in metabolic homeostasis, development, and health are well-known. The hypothalamus integrates the higher nerve center system and functions to regulate energy balance, feeding, biological rhythms and mood. However, how the hypothalamus is affected by gut microbes in mammals is unclear. This study demonstrated differences in hypothalamic gene expression between the germ-free (GF) pigs and pigs colonized with gut microbiota (CG) by whole-transcriptome analysis. A total of 938 mRNAs, 385 lncRNAs and 42 miRNAs were identified to be differentially expressed between the two groups of pigs. An mRNA-miRNA-lncRNA competing endogenous RNA network was constructed, and miR-22-3p, miR-24-3p, miR-136-3p, miR-143-3p, and miR-545-3p located in the net hub. Gene function and pathway enrichment analysis showed the altered mRNAs were mainly related to developmental regulation, mitochondrial function, the nervous system, cell signaling and neurodegenerative diseases. Notably, the remarkable upregulation of multiple genes in oxidative phosphorylation enhanced the GF pigs' hypothalamic energy expenditure. Additionally, the reduction in ATP content and the increase in carnitine palmitoyl transterase-1 (CPT1) protein level also confirmed this fact. Furthermore, the hypothalamic cell apoptosis rate in the CG piglets was significantly higher than that in the GF piglets. This may be due to the elevated concentrations of pro-inflammatory factors produced by gut bacteria. The obtained results collectively suggest that the colonization of gut microbes has a significant impact on hypothalamic function and health.

Time-Series Clustering of lncRNA-mRNA Expression during the Adipogenic Transdifferentiation of Porcine Skeletal Muscle Satellite Cells.

Skeletal muscle satellite cells (SMSCs), which are multifunctional muscle-derived stem cells, can differentiate into adipocytes. Long-chain non-coding RNA (lncRNA) has diverse biological functions, including the regulation of gene expression, chromosome silencing, and nuclear transport. However, the regulatory roles and mechanism of lncRNA during adipogenic transdifferentiation in muscle cells have not been thoroughly investigated. Here, porcine SMSCs were isolated, cultured, and induced for adipogenic differentiation. The expressions of lncRNA and mRNA at different time points during transdifferentiation were analysed using RNA-seq analysis. In total, 1005 lncRNAs and 7671 mRNAs showed significant changes in expression at differential differentiation stages. Time-series expression analysis showed that the differentially expressed (DE) lncRNAs and mRNAs were clustered into 5 and 11 different profiles with different changes, respectively. GO, KEGG, and REACTOME enrichment analyses revealed that DE mRNAs with increased expressions during the trans-differentiation were mainly enriched in the pathways for lipid metabolism and fat cell differentiation. The genes with decreased expressions were mainly enriched in the regulation of cell cycle and genetic information processing. In addition, 1883 DE mRNAs were regulated by 193 DE lncRNAs, and these genes were related to the controlling in cell cycle mainly. Notably, three genes in the fatty acid binding protein (FABP) family significantly and continuously increased during trans-differentiation, and 15, 13, and 11 lncRNAs may target FABP3, FABP4, and FABP5 genes by cis- or trans-regulation, respectively. In conclusion, these studies identify a set of new potential regulator for adipogenesis and cell fate and help us in better understanding the molecular mechanisms of trans-differentiation.

Chronic vagus nerve stimulation (VNS) altered IL-6, IL-1ß, CXCL-1 and IL-13 levels in the hippocampus of rats with LiCl-pilocarpine-induced epilepsy.

An increasing number of observations have indicated that the activation of inflammatory processes is involved in the pathogenesis of epilepsy. As an effective adjunctive therapy for medically intractable seizures, vagus nerve stimulation (VNS) is thought to interact with the inflammatory process to play an antiepileptic role. In this study, we examined the levels of multiple cytokine in focal brain tissue and peripheral blood to determine whether the antiepileptic effect of chronic VNS is related to the expression of cytokines. We observed that the frequency and duration of seizures significantly decreased in epileptic rats after two weeks of chronic VNS treatment. Pathological staining showed that the number of neural cells in the hippocampus was higher in the Epi + VNS group than in the Epi group, indicating that chronic VNS had a significant neuroprotective effect on epileptic rats. After comparing the expression of 9 cytokines, we found that the levels of the proinflammatory cytokines IL-6, IL-1ß and CXCL-1 in the hippocampus were significantly increased in the Epi group, while these cytokines were significantly decreased in the Epi + VNS group. Moreover, the level of the anti-inflammatory cytokine IL-13 was found to be reduced in Epi rats, while its levels were increased after VNS treatment. However, these changes in cytokine expression were not found in the hypothalamus or peripheral blood. These results suggest that the antiepileptic mechanism of VNS may work by inhibiting the activation of inflammatory processes in the epileptogenic focus.

Indole-3-Propionic Acid, a Functional Metabolite of Clostridium sporogenes, Promotes Muscle Tissue Development and Reduces Muscle Cell Inflammation.

Clostridium sporogenes (C. sporogenes), as a potential probiotic, metabolizes tryptophan and produces an anti-inflammatory metabolite, indole-3-propionic acid (IPA). Herein, we studied the effects of C. sporogenes and its bioactive metabolite, IPA, on skeletal muscle development and chronic inflammation in mice. In the in vivo study, the muscle tissues and serum samples of mice with C. sporogenes supplementation were used to analyze the effects of C. sporogenes on muscle metabolism; the IPA content was determined by metabonomics and ELISA. In an in vitro study, C2C12 cells were exposed to lipopolysaccharide (LPS) alone or LPS + IPA to verify the effect of IPA on muscle cell inflammation by transcriptome, and the involved mechanism was revealed by different functional assays. We observed that C. sporogenes colonization significantly increased the body weight and muscle weight gain, as well as the myogenic regulatory factors' (MRFs) expression. In addition, C. sporogenes significantly improved host IPA content and decreased pro-inflammatory cytokine levels in the muscle tissue of mice. Subsequently, we confirmed that IPA promoted C2C12 cells' proliferation by activating MRF signaling. IPA also effectively protected against LPS-induced C2C12 cells inflammation by activating Pregnane X Receptor and restoring the inhibited miR-26a-2-3p expression. miR-26a-2-3p serves as a novel muscle inflammation regulatory factor that could directly bind to the 3'-UTR of IL-1ß, a key initiator factor in inflammation. The results suggested that C. sporogenes with its functional metabolite IPA not only helps muscle growth development, but also protects against inflammation, partly by the IPA/ miR-26a-2-3p /IL-1ß cascade.

Electroacupuncture promotes the recovery of rats with spinal cord injury by suppressing the Notch signaling pathway via the H19/EZH2 axis.

BACKGROUND: Spinal cord injury (SCI) is a life-changing event with an extremely poor prognosis. In our preliminary studies, electroacupuncture (EA) was found to promote the repair of SCI, which was closely related to the Notch signaling pathway. Therefore, in the present study, we hypothesized that EA protects against SCI by inhibiting the Notch signaling pathway and sought to investigate the underlying molecular mechanisms. METHODS: Rat and cell models of SCI were established. The expression of long non-coding RNA H19 was measured by real-time quantitative polymerase chain reaction. The expression levels of EZH2, Notch1, Notch3, Notch4, Hes1, and PS1 protein were measured by western blot. Cell apoptosis and viability were analyzed using flow cytometry and Cell Counting Kit-8 assays, respectively. The expressions of glial fibrillary acidic protein (GFAP) and nestin were detected by immunofluorescence staining. RESULTS: The expressions of H19, EZH2, and GFAP were significantly increased after SCI but were inhibited by EA; in contrast, nestin expression was significantly decreased by SCI but was restored by EA. Moreover, oxygen-glucose deprivation (OGD) treatment elevated the expression of H19, EZH2, and Notch-related factors as well as apoptosis in PC-12 cells, while suppressing cell viability. Suppressing H19 alleviated the effects of OGD on cell viability and apoptosis, and inhibited the expression of EZH2 and Notch-related factors expression; these effects were reversed by EZH2 overexpression. Finally, EA promoted the recovery of SCI rats and neural stem cell (NSC) proliferation by inhibiting the Notch signaling pathway, which was reversed by H19 overexpression. CONCLUSIONS: Our results demonstrated that EA promotes the recovery of SCI rats and increases the proliferation and differentiation of NSCs by suppressing the Notch signaling pathway via modulating the H19/EZH2 axis.

MicroRNA-489-3p promotes adipogenesis by targeting the Postn gene in 3T3-L1 preadipocytes.

AIMS: Accumulating evidence indicates that a number of microRNAs (miRNAs) serve as essential regulators during adipogenesis and adipolysis in humans and animals and play critical roles in the development of fat tissue. In this study, we aimed to determine the functional role and underlying regulatory mechanism of microRNA-489-3p (miR-489) in adipocytes. MATERIALS AND METHODS: The expression patterns of miR-489 in mice were measured by qRT-PCR. Overexpression and knockdown of miR-489 by mimic and inhibitor transfections in 3T3-L1 preadipocytes revealed the regulatory effect of miR-489 on cellular proliferation and differentiation and energy turnover. Furthermore, RNA-seq, bioinformatics prediction, and dual luciferase reporter assays were used to identify the direct target of miR-489. KEY FINDINGS: The results showed that miR-489 was highly expressed in the visceral fat tissue of adult mice, and obese mice exhibited higher levels of miR-489 than normal mice. Overexpression of miR-489 suppressed proliferation but promoted adipogenic differentiation and lipid accumulation in the cells. Mitochondrial oxidation also fluctuated in the cells due to the high expression of miR-489. Notably, knockdown of miR-489 did not have a strong opposing effect on the cells. Periostin (Postn) was identified as a direct target gene for miR-489, and silencing the Postn gene similarly stimulated adipogenesis and differentiation of adipocytes. SIGNIFICANCE: miR-489 provides a strong driving force for adipogenesis metabolism and adipocyte differentiation by targeting the Postn gene. This result may contribute to the treatment of obesity.

The intestinal microbiota contributes to the growth and physiological state of muscle tissue in piglets.

Although the importance of the intestinal microbiota in host growth and health is well known, the relationship between microbiota colonization and muscle development is unclear. In this study, the direct causal effects of the colonization of gut microorganisms on the muscle tissue of piglets were investigated. The body weight and lean mass of germ-free (GF) piglets were approximately 40% lower than those of normal piglets. The deletion of the intestinal microbiota led to weakened muscle function and a reduction in myogenic regulatory proteins, such as MyoG and MyoD, in GF piglets. In addition, the blinded IGF1/AKT/mTOR pathway in GF piglets caused muscle atrophy and autophagy, which were characterized by the high expression of Murf-1 and KLF15. Gut microbiota introduced to GF piglets via fecal microbiota transplantation not only colonized the gut but also partially restored muscle growth and development. Furthermore, the proportion of slow-twitch muscle fibers was lower in the muscle of GF piglets, which was caused by the reduced short-chain fatty acid content in the circulation and impaired mitochondrial function in muscle. Collectively, these findings suggest that the growth, development and function of skeletal muscle in animals are mediated by the intestinal microbiota.

Introduction of Colonic and Fecal Microbiota From an Adult Pig Differently Affects the Growth, Gut Health, Intestinal Microbiota and Blood Metabolome of Newborn Piglets.

Microbiota transplantation is a rapid and effective method for changing and reshaping the intestinal microbiota and metabolic profile in humans and animals. This study compared the different influences of the introduction of fecal microbes and colonic microbes from a fat, adult pig in newborn pigs. Both colonic microbiota transplantation (CMT) and fecal microbiota transplantation (FMT) promoted growth and improved gut functions in suckling pigs up to weaning. FMT was more beneficial for body weight gain and body fat deposition in piglets, while CMT was more beneficial for intestinal health and mucosal immunity. 16S rDNA sequence analysis indicated that both CMT and FMT significantly increased the abundances of beneficial or functional bacteria, such as Lactobacillus and Prevotella_2 genera, in the piglets, and reduced the abundances of harmful bacteria, such as Escherichia-Shigella. Blood metabolome analysis showed that transplantation, especially FMT, enhanced lipid metabolism in piglets. In addition, while CMT also changed amino acid metabolism and increased anti-inflammatory metabolites such as 3-indoleacetic acid and 3-indolepropionic acid in piglets, FMT did not. Of note, FMT damaged the intestinal barrier of piglets to a certain extent and increased the levels of inflammatory factors in the blood that are potentially harmful to the health of pigs. Taken together, these results suggested that intestinal and fecal microbiota transplantations elicited similar but different physiological effects on young animals, so the application of microbiota transplantation in animal production requires the careful selection and evaluation of source bacteria.