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Angiotensin II (AngII) regulates pigment synthesis by tyrosinase in melanocytes. To evaluate the association between AngII and coat color formation, we detected the expression distribution of AngII in white and black sheep skins by LC-ESI-MS/MS, western blot, quantitative real-time-PCR (qPCR) and distribution of AngII by immunohistochemistry.Liquid chromatography-electrospray ionization tandem MS (LC-ESI-MS/MS) results showed that AngII was found in white and black skin tissues of sheep. Western blot results verified the LC-ESI-MS/MS results and suggested that AngII was expressed at significantly higher levels in black sheep skins compared with the white sheep skins. Quantitative real time PCR (qRT-PCR) results also revealed that the expression level of AngII mRNA was higher in black sheep skins than that in white sheep skins. Immunohistochemical analysis further demonstrated that AngII protein was localized in the hair bulb and outer root sheath of hair follicle in sheep. In summary, protein and transcripts exhibited the same expression pattern in white and black sheep skins. Furthermore, the expressions of AngII in the hair bulb and outer root sheath of black sheep were stronger than those in white sheep. These results suggested that AngII functions in sheep coat color regulation and offer a novel insight for further investigation on the role of AngII in the coat color formation in sheep.

Bovine endometrial stromal cells (bESCs) are exposed to a complex environment of bacteria and viruses due to the rupture of epithelial cells after delivery. Inflammatory responses are elicited by the activation of host pattern recognition receptors through pathogen-related molecules such as lipopolysaccharides (LPS) on the cell membrane. Forsythoside A (FTA) is a major active constituent of Forsythia suspensa (Thunb.) Vahl. is a flowering plant widely employed as a traditional Chinese herbal medicine to treat various inflammatory diseases such as nephritis, eye swelling, scabies, ulcers, and mastitis; however, the molecular mechanisms underlying its therapeutic effects on bovine endometritis are still unclear. The aim of this study was to explore the role of miRNA and the mechanisms underlying the protective activity of FTA on the inflammation of bovine endometrial stromal cells induced by LPS. Based on previous research, we isolated and cultured bESCs in vitro and categorized them into LPS and LPS+FTA groups with three replicates. Upon reaching 80% confluence, the bESCs were treated with 0.5 μg/mL of LPS or 0.5 μg/mL of LPS + 100 μg/mL of FTA. We, then, performed high-throughput sequencing (RNA-Seq) to investigate the effects of FTA on LPS-stimulated primary bESCs and their underlying mechanisms. We identified 167 miRNAs differentially expressed in the LPS groups; 72 miRNAs were up-regulated, and 95 were down-regulated. Gene ontology enrichment analysis revealed that differentially expressed microRNA (DEGs) were most enriched during the cellular metabolic process; they were mostly located intracellularly and participated in protein, enzyme, and ion binding. Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed that the DEGs were most enriched in the mitogen-activated protein kinase, tumor necrosis factor, and Interleukin-17 signaling pathways. These results reveal the complex molecular mechanism involved in the FTA and provide a basis for future studies of bovine endometritis treatment with traditional Chinese medicine monomer.

After parturition, bovine uterine stromal cells are often exposed to complex bacterial and viral stimuli owing to epithelial cell rupture, resulting in an inflammatory response. In this study, we used an in vitro model to study the response of bovine endometrial stromal cells to inflammatory mediators and the associated regulated microRNAs in response to lipopolysaccharide. Lipopolysaccharide (LPS) is a bacterial wall component in gram-negative bacteria that causes inflammation upon immune recognition, which is used to create in vitro inflammation models. Thus, we used high-throughput RNA sequencing to identify miRNAs that may have an anti-inflammatory role in the LPS-induced inflammatory response. Two groups of bovine uterine cells were treated with phosphate buffer saline (PBS) and LPS, respectively. Compared with the control (PBS) group, the LPS-treated group had 219 differentially expressed miRNAs, of which 113 were upregulated, and 106 were downregulated. Gene ontology enrichment analysis revealed that the target genes of differentially expressed miRNAs were significantly enriched in several activities, such as transferase activity, small molecule binding, and protein binding. Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated that the target genes of differential miRNAs were significantly enriched in fluid shear stress and atherosclerosis, MAPK signaling pathway, TNF signaling pathway. By analyzing differentially expressed miRNAs, we found that miR-200c, miR-1247-3p, and let-7b are directly related to the inflammatory response. For instance, miR-200c target genes (MAP3K1, MAP4K3, MAPKAPK5, MAP3K8, MAP3K5) and let-7b target genes (CASP3, IL13, MAPK8, CXCL10) were significantly enriched in the MAPK and IL-17 signaling pathways, respectively. In summary, our research provides insight into the molecular mechanism underlying LPS-induced inflammation in vitro, which may unveil new targets for the treatment of endometritis.

Background: Endometritis adversely affects the ability of cattle to reproduce, and significantly reduces milk production. Consequently, it has great influence on the economic value of dairy cows. The endometrium is mainly composed of epithelial and stromal cells and they produce the first immune response to invading pathogens. Epithelial cells are the first cellular barrier through which bacteria enter the uterine endometrium. However, most of the epithelial cells are disrupted and stromal cells are exposed to an inflammatory environment when endometritis occurs, especially postpartum. A loss of the protective epithelium allows bacteria or toxins to access the underlying stromal cells. The activation of Toll-like receptor（TLRs）on stromal cells induces increased production of cytokines. Understanding the genome-wide characterization of the bovine endometritis will be beneficial for prevention and treatment of endometritis. In this study, whole-transcriptomic gene changes in bovine stromal cells treated with LPS were compared with those treated with PBS (control group) and were analyzed by RNA sequencing (RNA-seq). This was done in a cell culture model in vitro.Results: Compared with the control group, a total of 366 differentially expressed genes (DEGs) were identified in LPS-induced group (234 upregulated and 132 downregulated genes), with an adjusted P-value＜0.05 by DESeq. Gene ontology (GO) enrichment analysis revealed DEGs were most enriched in lymphocyte activation, interleukin-1 receptor binding, regulation of cell activation, and lymphocyte-activated interleukin-12 production. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed DEGs were most enriched in TNF signaling pathway, Toll-like receptor signaling pathway, cytokine-cytokine receptor interaction, nucleotide-binding oligomerization domain-like (NOD-like) receptor signaling pathway, NF-κB signaling pathway, and chemokine signaling pathway.Conclusion: The results of this study unraveled bovine endometrial stromal cells affected with LPS transcriptome profile alterations，which may have a significant effect on the treatment inflammation by comprehending molecular mechanisms and authenticating unique genes related to endometritis.

Background Medulloblastoma (MB) is the most common intracranial malignant tumor in children. The genes and pathways involved in the pathogenesis of MB are relatively unknown. We aimed to identify potential biomarkers and small-molecule drugs for MB. Methods Gene expression profile data sets were obtained from the Gene Expression Omnibus (GEO) database and the differentially expressed genes (DEGs) were identified using the Limma package in R. Functional annotation, and cell signaling pathway analysis of DEGs was carried out using DAVID and Kobas. A protein-protein interaction network was generated using STRING. Potential small-molecule drugs were identified using CMap. Result We identified 104 DEGs (29 upregulated; 75 downregulated). Gene ontology analysis showed enrichment in the mitotic cell cycle, cell cycle, spindle, and DNA binding. Cell signaling pathway analysis identified cell cycle, HIF-1 signaling pathway, and phospholipase D signaling pathway as key pathways. SYN1, CNTN2, FAIM2, MT3, and SH3GL2 were the prominent hub genes and their expression level were verified by RT-qPCR. Vorinostat, resveratrol, trichostatin A, pyrvinium, and prochlorperazine were identified as potential drugs for MB. The five hub genes may be targets for diagnosis and treatment of MB, and the small-molecule compounds are promising drugs for effective treatment of MB. Conclusion In this study we obtained five hub genes of MB, SYN1, CNTN2, FAIM2, MT3, and SH3GL2 were confirmed as hub genes. Meanwhile, Vorinostat, resveratrol, trichostatin A, pyrvinium, and prochlorperazine were identified as potential drugs for MB.

OBJECTIVEIt has been reported that microRNA-195 (miR-195) protects against chronic brain injury induced by chronic brain hypoperfusion. However, neither the expression profile of miR-195 nor its potential role during acute ischemic stroke has been investigated. In this study, the authors’ aim was to verify the mechanism of miR-195 in acute ischemic stroke.METHODSThe plasma levels of miR-195 expression were assessed using real-time PCR in 96 patients with acute ischemic stroke, and the correlation with the National Institutes of Health Stroke Scale score was evaluated. In addition, cerebral infarct volume, neurological score, and levels of miR-195 and CX3CL1/CX3CR1 mRNA and protein expression were assessed in mice subjected to middle cerebral artery occlusion (MCAO) with or without intra-cerebroventricular infusion of lentiviral vector. The inflammatory cytokines tumor necrosis factor–α (TNFα), interleukin (IL)–1β, and IL-6 of mouse brains after MCAO and BV2 cells treated with oxygen-glucose deprivation were measured using enzyme-linked immunosorbent assay, and apoptotic proteins were examined by Western blotting. Direct targeting of CX3CL1/CX3CR1 by miR-195 was determined by immunoblotting and dual luciferase assay.RESULTSIn ischemic stroke patients, miR-195 was significantly downregulated and expression levels of miR-195 in these patients negatively correlated with the National Institutes of Health Stroke Scale score. In mice after MCAO, miR-195 overexpression decreased infarct volume, alleviated neurological deficits, and most importantly, suppressed an inflammatory response. Meanwhile, miR-195 suppressed the expression of the inflammatory cytokines TNFα, IL-1β, and IL-6 in vitro and in vivo. The authors further discovered that both CX3CL1 and CX3CR1 are direct targets of miR-195, but miR-195 exerts neuroprotective roles mainly through inhibiting CX3CR1-mediated neuroinflammation and subsequent neuronal cell apoptosis.CONCLUSIONSTaken together, these findings suggest that miR-195 promotes neuronal cell survival against chronic cerebral ischemic damage by inhibiting CX3CR1-mediated neuroinflammation. This indicates that miR-195 may represent a novel target that regulates neuroinflammation and brain injury, thus offering a new treatment strategy for cerebral ischemic disorders.