Previous studies suggested that elevated level of TGF-β2 in AH of POAG maybe central to the pathologic behavior of the disease[5, 7, 8]. However, until now, a comprehensive investigation of the effects of TGF-β2 has not been presented. MicroRNAs housed within exosomes have emerged in recent years as critical modulators to synchronize AH production and drainage for ultimately IOP homeostasis . Our study was designed to test whether a famous model of glaucomatous TM dysfunction, TGF-β2 treatment, can produce pathological changes in exosomal cargo and function. Furthermore, we want to reveal possible physiological or pathological processes and pathways involving pathogenesis of POAG regulated by the identified exosomal miRNAs.
According to the results of microarray analysis, 23 miRNAs were revealed to be up-regulated in the model group compared with the control group. Hsa-miR-6087, hsa-miR-663a and hsa-miR-6821-5p were the top three up-regulated exosomal miRNAs, whereas hsa-miR-6800-3p, hsa-miR-6716-3p, and hsa-let-7i-5p were revealed to be markedly down-regulated. Hsa-miR-6087 has been previously reported to act as an important mediators of cancer[44, 45],intermediate monocytes and regulation of cell differentiation. Has-miR-663a is located in the chromosome 20q11.1 and has been reported to be closely related with the biological behavior of cell differentiation, inflammation, autoimmune diseases, and cancer[48–50]. Let-7 was discovered early as a member of the large class of non-protein-coding RNAs. Dysregulated expression of the let-7i-5p is associated with various diseases such as cancer, and cardiovascular. These exosomal miRNAs of AH may play a crucial role in the cell-cell communication for pathological processes of TM from POAG patients.
Previous study showed that significant up-regulation of mir-518d and mir-143, and significant down-regulation of mir-660 expression were observed in AH from POAG patients . For this phenomenon, we infer the miRNAs in AH are not parallel to the miRNAs of the exosomes. To date, the specific source of miRNAs in AH is unknown. Variable intraocular cells can secrete exosomes into the AH, and numerous molecules, particularly miRNAs, have been shown to change in response to ocular disease or injury. Recent study shows that non-pigmented ciliary epithelium cell (NPCE)-derived exosomes could affect canonical Wnt signaling in TM cells, a pathway involved in IOP regulation. Thus, we suspect that the changes of exosomal miRNAs mediate cell-cell communication and play a major role in the crosstalk between TM cells and the macro−/microenvironment.
The miRNA-miRNA interaction network showed that during the occurrence and development of diseases. Many miRNA molecules are involved in the process. The term interaction means one miRNA may interact and regulate the stability or expression of the other one or the others. Each miRNA regulates a broad spectrum of target genes and affects many signaling pathways. Therefore, disruption of a single miRNA expression may only limit an inhibitory effect, whereas joint interference of multiple miRNAs may be effective. Thus, determining how to silence multiple abnormally expressed miRNAs simultaneously in order to enhance the efficacy of disease treatments is important. For the resaon, we used DIANA-TarBase to predicte the potential targets of 26 exosomal miRNAs. Unlike miRDB and TargetScan, DIANA-TarBase was the first database aiming to catalog published experimentally validated interactions between miRNAs and genes. The GO analysis results revealed that the exosomal miRNAs were involved in biological processes, cellular components and molecular functions, including antigen processing and presentation, spliceosomal complex and insulin-like growth factor I binding. These miRNAs are not specific to a single gene. These miRNAs alter the expression of a number of genes instead of a single gene, and these genes may be involved in a single cellular process like extracellular matrix remodeling.
KEGG pathway enrichment analysis also revealed that the p13k-akt signaling pathway and MAPK signaling pathways may be involved in the process. We then constructed the co-expression network of selective signaling pathways and exosomal miRNAs. Overall, these studies indicate that exosomes and their cargoes can be used as biomarkers for glaucoma, potentially serving as an early diagnostic marker, and may also contribute directly to the progression of the disease.TGF-β2 cause not only gene and protein expression changes in the HTM cells, but also can alter exosomal miRNAs expression. These miRNAs enlisted in the current study represent only a small proportion of genes in exosomes. This condition may be correlated with the specific condition of in vitro culture of the cells.