miR-425-5p Regulates Proliferation of Bovine Mammary Epithelial Cells by Targeting TOB2.

In recent years, rising temperatures have caused heat stress (HS), which has had a significant impact on livestock production and growth, presenting considerable challenges to the agricultural industry. Research has shown that miR-425-5p regulates cellular proliferation in organisms. However, the specific role of miR-425-5p in bovine mammary epithelial cells (BMECs) remains to be determined. The aim of this study was to investigate the potential of miR-425-5p in alleviating the HS-induced proliferation stagnation in BMECs. The results showed that the expression of miR-425-5p significantly decreased when BMEC were exposed to HS. However, the overexpression of miR-425-5p effectively alleviated the inhibitory effect of HS on BMEC proliferation. Furthermore, RNA sequencing analysis revealed 753 differentially expressed genes (DEGs), comprising 361 upregulated and 392 downregulated genes. Some of these genes were associated with proliferation and thermogenesis through enrichment analyses. Further experimentation revealed that TOB2, which acts as a target gene of miR-425-5p, is involved in the regulatory mechanism of BMEC proliferation. In summary, this study suggests that miR-425-5p can promote the proliferation of BMECs by regulating TOB2. The miR-425-5p/TOB2 axis may represent a potential pathway through which miR-425-5p ameliorates the proliferation stagnation of BMECs induced by HS.

miR-196a Promotes Proliferation of Mammary Epithelial Cells by Targeting CDKN1B.

Heat stress (HS) has become one of the key challenges faced by the dairy industry due to global warming. Studies have reported that miR-196a may exert a role in the organism's response to HS, enhancing cell proliferation and mitigating cellular stress. However, its specific role in bovine mammary epithelial cells (BMECs) remains to be elucidated. In this study, we aimed to investigate whether miR-196a could protect BMECs against proliferation arrest induced by HS and explore its potential underlying mechanism. In this research, we developed an HS model for BMECs and observed a significant suppression of cell proliferation as well as a significant decrease in miR-196a expression when BMECs were exposed to HS. Importantly, when miR-196a was overexpressed, it alleviated the inhibitory effect of HS on cell proliferation. We conducted RNA-seq and identified 105 differentially expressed genes (DEGs). Some of these DEGs were associated with pathways related to thermogenesis and proliferation. Through RT-qPCR, Western blotting, and dual-luciferase reporter assays, we identified CDKN1B as a target gene of miR-196a. In summary, our findings highlight that miR-196a may promote BMEC proliferation by inhibiting CDKN1B and suggest that the miR-196a/CDKN1B axis may be a potential pathway by which miR-196a alleviates heat-stress-induced proliferation arrest in BMECs.

miR-383-5p Regulates Preadipocyte Proliferation and Differentiation by Targeting RAD51AP1.

Obesity has become a major health problem worldwide, and increasing evidence supports the importance of microRNAs (miRNAs) in its pathogenesis. Recently, we found that miR-383-5p_1 is highly expressed in the perirenal fat of high-fat-fed rabbits, but it is not yet known whether miR-383-5p is involved in lipid metabolism. Here, we used transcriptome sequencing technology to screen 1642 known differentially expressed genes between miR-383-5p mimic groups and miR-383-5p negative control groups. Gene Ontology Resource (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were enriched in the pathway related to lipid metabolism, and glycine biosynthesis, the NOD receptor signal pathway and nonalcoholic fatty liver were significantly enriched. Afterwards, our research results indicated that miR-383-5p can promote the proliferation and differentiation of rabbit preadipocytes, and there is a direct targeting relationship with RAD51AP1. Mechanistically, miR-383-5p directly interacts with the lipid metabolism and participates in adipogenesis and lipid accumulation by targeting RAD51AP1. In conclusion, our data highlight a physiological role for miRNA in lipid metabolism and suggest the miR-383-5p/RAD51AP1 axis may represent a potential mechanism for controlling lipid accumulation in obesity.

Negative effects of heat stress on ovarian tissue in female rabbit.

Numerous studies have highlighted the role of miRNA in the deformation and necrosis of cells of ovarian tissue caused by heat stress (HS), which ultimately affects ovarian function. Although the role of small RNAs has been investigated in alterations in ovarian tissue functioning in response to HS, the expression profile of ovarian miRNA has been explored to a lesser extent. In this study, female rabbits were subject to HS treatment by using electrical heater. The current work demonstrated that HS could significantly change physiological performance of female rabbits including body weight, rectal temperature and relative ovary weight, and significantly reduce serum IL-2, IL-8, CAT, and GSH-Px concentrations by enzyme-linked immunosorbent assay (ELISA) technique. As a result, an increase in apoptosis in ovarian cells, as well as unhealthy follicles, were observed by Hematoxylin-eosin (HE) and TUNEL staining. Additionally, small RNA-seq revealed changes in the miRNA expression profile of rabbit ovaries under HS. Five hundred fourteen miRNAs were obtained including known miRNAs 442 and novel miRNAs 72. Among these miRNAs, 23 miRNAs were significantly expressed under HS. Eleven differentially expressed miRNAs (DE miRNAs) and 9 their predicted targets were confirmed by qPCR, which were expected miRNA-mRNA negative regulation pattern. Among the DE miRNAs and targets, miR-141-39 may target COQ6, miR-449a-5p and miR-34c-5p may control RFC5 and RTN2 together, miR-449a-5p may target ACADVL, miR-34c-5p potentially targets Bcl-2 and miR-196b-5p potentially regulates CASK and HOXB6. Thus, the current work suggested the negative effects of HS on the ovarian tissue of female rabbits, and in conclusion these changes could be caused by decreased serum IL-2, IL-8, CAT and GSH-Px levels, increased ovarian apoptosis, and changed the expression of miRNAs.

Dynamics of transcriptome and chromatin accessibility revealed sequential regulation of potential transcription factors during the brown adipose tissue whitening in rabbits.

Brown adipose tissue (BAT) represents a valuable target for treating obesity in humans. BAT losses of thermogenic capacity and gains a "white adipose tissue-like (WAT-like)" phenotype (BAT whitening) under thermoneutral environments, which could lead to potential low therapy responsiveness in BAT-based obesity treatments. However, the epigenetic mechanisms of BAT whitening remain largely unknown. In this study, BATs were collected from rabbits at day0 (D0), D15, D85, and 2 years (Y2). RNA-sequencing (RNA-seq) and the assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) were performed to investigate transcriptome and chromatin accessibility of BATs at the four whitening stages, respectively. Our data showed that many genes and chromatin accessible regions (refer to as "peaks") were identified as significantly changed during BAT whitening in rabbits. The BAT-selective genes downregulated while WAT-selective genes upregulated from D0 to Y2, and the de novo lipogenesis-related genes reached the highest expression levels at D85. Both the highly expressed genes and accessible regions in Y2 were significantly enriched in immune response-related signal pathways. Analysis of different relationships between peaks and their nearby genes found an increased proportion of the synchronous changes between chromatin accessibility and gene expression during BAT whitening. The synergistic changes between the chromatin accessibility of promoter and the gene expression were found in the key adipose genes. The upregulated genes which contained increased peaks were significantly enriched in the PI3K-Akt signaling pathway, steroid biosynthesis, TGF-beta signaling pathway, osteoclast differentiation, and dilated cardiomyopathy. Moreover, the footprinting analysis suggested that sequential regulation of potential transcription factors (TFs) mediated the loss of thermogenic phenotype and the gain of a WAT-like phenotype of BAT. In conclusion, our study provided the transcriptional and epigenetic frameworks for understanding BAT whitening in rabbits for the first time and might facilitate potential insights into BAT-based obesity treatments.

Combined analysis of differentially expressed lncRNAs and miRNAs in liver tissues of high-fat fed rabbits by transcriptome sequencing.

High-fat diet could lead to a series of metabolic diseases, including obesity, and its mechanism is not clear. In this study, the rabbit individuals were fed with high-fat diet, the liver tissues were collected, high-throughput sequencing technology was used to reveal the expression of lncRNA and miRNA difference, and the molecular regulation mechanism of lncRNA-miRNA. A total of 24,615 DE lncRNAs and 52 DE miRNAs were identified, including 15 novel discovered DE miRNAs (5 upregulated and 10 downregulated). Furthermore, five miRNAs and three mRNAs were verified by qRT-PCR, and the results showed that the expression of the DE miRNAs and DE lncRNAs in the two groups was consistent with our sequencing results. GO and KEGG analyzed 7,57,139 target genes respectively, enriching the pathways related to lipid metabolism, including mucin O-glycan biosynthesis pathway, insulin resistance and glucagon signaling pathway. Moreover, 65 targeting relationships were obtained. Among them, LOC103348122/miR-450a-5p, LOC103350359/miR-450a-3p and LOC103350429/miR-148a-5p were proposed the first time. Significantly, LOC103348122/miR-450a-5p and LOC103350429/miR-148a-5p were related to lipid metabolism in the liver. This study is of great significance to the CeRNA regulatory network related to lipid metabolism in the liver of rabbits, and provides a basis for understanding hepatic steatosis in rabbits.

The Multi-Omics Analysis Revealed Microbiological Regulation of Rabbit Colon with Diarrhea Fed an Antibiotic-Free Diet.

Diarrhea symptoms appeared after antibiotics were banned from animal feed based on the law of the Chinese government in 2020. The colon and its contents were collected and analyzed from diarrheal and healthy rabbits using three omics analyses. The result of the microbial genomic analysis showed that the abundance of Bacteroidetes and Proteobacteria increased significantly (p-value < 0.01). Transcriptomes analysis showed that differentially expressed genes (DEGs) are abundant in the IL-17 signaling pathway and are highly expressed in the pro-inflammatory pathway. The metabolome analysis investigated differential metabolites (DMs) that were mainly enriched in tryptophan metabolism and bile secretion, which were closely related to the absorption and immune function of the colon. The results of correlation analysis showed that Bacteroidetes was positively correlated with 4-Morpholinobenzoic acid, and 4-Morpholinobenzoic acid could aggravate inflammation through its influence on the bile secretion pathway. The enriched DMs L-Tryptophan in the tryptophan metabolism pathway will lead to the functional disorder of inhibiting inflammation by affecting the protein digestion and absorption pathway. Thus, the colonic epithelial cells were damaged, affecting the function of the colon and leading to diarrhea in rabbits. Therefore, the study provided an idea for feed development and a theoretical basis for maintaining intestinal tract fitness in rabbits.