Thousands of long noncoding RNAs (lncRNAs) have been identified in mouse, rat, and human testes, some of which play important roles in testis development and spermatogenesis. However, systematic analysis of lncRNAs expressed in postnatal pig testes has not been reported. Thus, in this study, we present the expression and characterization of lncRNAs in immature (30-day-old [D30]) and mature (180-day-old [D180]) pig testes. A total of 90 440 168 (85.75%) and 97 001 700 (95.35%) 150-base-pair paired-end clean reads were generated in D30 and D180 cDNA libraries, respectively, using the Illumina HiSeq 4000 platform; 36 727 transcripts were assembled in those two libraries, 777 lncRNA transcripts from 752 lncRNA gene loci were identified using the highly stringent pipeline, and 101 of those lncRNA transcripts were significantly differentially expressed. Those lncRNAs shared some characteristics with other mammals, including fewer exons, shorter length and exon length, and lower expression level compared with those of protein-coding genes; 402 protein-coding genes (<10 kb) were found as nearest neighbors of 294 out of 752 lncRNA genes, and gene ontology enrichment showed that they were enriched in transcription- and development-related processes. Fifteen differentially expressed and 10 novel lncRNAs were randomly selected and validated by quantitative polymerase chain reaction (PCR) and reverse transcriptase PCR. In addition, one of the 10 novel lncRNAs was further confirmed using RACE clone technology. This study provides a catalog of porcine testes lncRNAs for further understanding their regulation roles in pig testis development and spermatogenesis.

The aim of this study was to investigate whether supplementation with chitosan (COS) could reduce diarrhea and to explore how COS alleviates intestinal inflammation in weaned pigs. Thirty pigs (Duroc×Landrace×Yorkshire, initial BW of 5.65±0.27) weaned at age 21 d were challenged with enterotoxigenic Escherichia coli during a preliminary trial period, and then divided into three treatment groups. Pigs in individual pens were fed a corn-soybean meal diet, that contained either 0 (control), 50 mg/kg chlortetracycline, or 300 mg/kg COS for 21 days. The post-weaning diarrhea frequency, calprotectin levels and TLR4 protein expression were decreased (P<0.05) in both the COS and chlortetracycline groups compared with control. Simultaneously, supplemental COS and chlortetracycline had no effect on the mRNA expression of TNF-α in the jejunal mucosa, or on the concentrations of IL-1β, IL-6 and TNF-α in serum. However, COS supplementation improved (P<0.05) the mRNA expression of IL-1β and IL-6 in the jejunal mucosa. The results indicate that supplementation with COS at 300 mg/kg was effective for alleviating intestinal inflammation and enhancing the cell-mediated immune response. As feed additives, chitosan and chlortetracycline may influence different mechanisms for alleviating inflammation in piglets.

A comprehensive and systematic understanding of the roles of lncRNAs in the postnatal development of the pig testis has still not been achieved. In the present study, we obtained more than one billion clean reads and identified 15,528 lncRNA transcripts; these transcripts included 5032 known and 10,496 novel porcine lncRNA transcripts and corresponded to 10,041 lncRNA genes. Pairwise comparisons identified 449 known and 324 novel lncRNAs that showed differential expression patterns. GO and KEGG pathway enrichment analyses revealed that the targeted genes were involved in metabolic pathways regulating testis development and spermatogenesis, such as the TGF-beta pathway, the PI3K-Akt pathway, the Wnt/β-catenin pathway, and the AMPK pathway. Using this information, we predicted some lncRNAs and coding gene pairs were predicted that may function in testis development and spermatogenesis; these are listed in detail. This study has provided the most comprehensive catalog to date of lncRNAs in the postnatal pig testis and will aid our understanding of their functional roles in testis development and spermatogenesis.

To understand the complex physiological process underlying pig testis development and spermatogenesis, this study aims to characterize the change in miRNA and mRNA profiles at four developmental stages of embryonic and postnatal testes, including 60 dpc (days post coitus, E60), 90 dpc (E90), 30-day-old (D30) and 180-day-old (D180). A total of 304 mature, 50 novel miRNAs, and 8343 differentiallyexpressed genes were identified. 93 (48 up and 45 down), 104 (49 up and 55 down), 122 (49 up and 73 down) differentially-expressed miRNAs, as well as 1007 (646 up and 361 down), 1929 (911 up and 1018 down), 7420 (3998 up and 3422 down) differentially-expressed genes were identified in E90 vs. E60, D30 vs. E90 and D180 vs. D30, respectively. Integrating analysis of miRNA and mRNA expression profiles predicted more than 50 000 miRNA-mRNA interaction sites. GO and KEGG pathway analysis of the predicted target genes illustrated the likely roles of differentially expressed miRNAs in testis development and spermatogenesis. For example, PI3K-Akt signaling pathway and Hippo signaling pathway related development, and carbon metabolism, fatty acid metabolism, protein digestion and absorption, were involved in metabolite synthesis. These integrated high-throughput expression data show that miRNA is a critical factor in porcine testis development, providing a useful resource to understand global genome expression change in porcine testis development and spermatogenesis.

Accumulating reports have demonstrated that microRNAs (miRNAs) participate in regulating the complex processes of animal testis development and spermatogenesis; yet, the mechanisms by which miRNAs regulate spermatogenesis are poorly understood. miR-26a was identified as a miRNA that is differentially expressed among different pig testicular tissue developmental stages in our previous study. In this study, p21 activated kinase 2 (PAK2) gene was determined as one target gene of miR-26a by luciferase reporter assay, and miR-26a repressed the PAK2 mRNA abundance in porcine Sertoli cells. The Cell Counting Kit-8 (CCK8) assay, 5-Ethynyl-2'-deoxyuridine (EdU) assay and annexin V-FITC/PI staining assay results showed that miR-26a overexpression inhibited proliferation and promoted apoptosis in porcine Sertoli cells. These phenomena were similar to the siRNA-mediated knockdown of the PAK2 gene. Taken together, our results demonstrate that miR-26a inhibits proliferation and promotes apoptosis in porcine Sertoli cells by targeting the PAK2 gene, which may be a regulator of porcine spermatogenesis.