Analysis of deep sequencing Exosome-microRNA expression profile from Chicken Type Ⅱ Pneumocytes derived reveals potential role of gga-miRNA-451 in inflammation

Background: Exosomes are nanosized extracellular vesicles secreted by multiple cells in the body, including those located in the respiratory tract and lungs. They are emerging as important inflammatory mediators and can release their contents, especially microRNAs (miRNAs), to both neighboring and distal cells. Mycoplasma gallisepticum (MG) can target host cell and cause chronic respiratory disease (CRD) in chickens. Although exosomal miRNAs have been demonstrated to produce an important effect on microbial pathogenesis and inflammatory response as crucial regulatory noncoding RNAs, the mechanism by which exosomal miRNAs regulate MG-induced inflammation remains to be elucidated. Methods: the expression of exosome-microRNA derived from MG-infected chicken type Ⅱ pneumocytes (CP-Ⅱ) was screened, and the target genes and function of differentially expressed miRNAs (DEGs) were predicted. To verify the inflammatory functions of exosomal gga-miR-451 via targeting YWHAZ, Western blot, ELISA, and RT-qPCR were used in this study. Results: A total of 722 miRNAs were identified from the two exosomal small RNA (sRNA) libraries, and 279 novel miRNAs were discovered; 30 miRNAs (9 up-regulated and 21 down-regulated) were significantly changed (P＜0.05). Function annotation analysis of DEGs showed that the target miRNAs were significantly enriched in treatment group, such as cell cycle, Toll-like receptor signaling pathway and MAPK signaling pathway, etc. The results have also confirmed that gga-miR-451-absent exosomes derived from MG-infected CP-Ⅱ cells increased inflammatory cytokine production in DF-1 (chicken embryo fibroblast) cells, and Wild Type CP-Ⅱ cells-derived-exosomes displayed protective effects. Conclusion: our work suggests that exosomes from MG-infected CP-Ⅱ cells alter the dynamics of the DF-1 cells, and may contribute to pathology of the MG infection via exosomal gga-miR-451 targeting YWHAZ involving in inflammation. This could potentially be used as a biomarker for diagnostics and treatment.