Tear Film miRNAs and Their Association With Human Dry Eye Disease.

PURPOSE: miRNAs can regulate inflammatory pathways. The purpose of this work was to determine if inflammatory-related tear film miRNAs are associated with extracellular vesicles (EVs) in human non-Sjögren's Syndrome dry eye disease (DED) participants. METHODS: Five DED and 5 non-DED human participants were recruited. Tears samples were collected by washing the ocular surface of both eyes with phosphate buffered saline, pooling samples from the right and left eyes, and purifying EVs from the samples with a polyethylene glycol (PEG) 8000 precipitation procedure. Samples were directly analyzed via ELISA or transmission electron microscopy (TEM), or RNA was isolated first from the EVs and evaluated with RNA-Seq. RESULTS: EVs were identified in the tear film of both groups using TEM and ELISA. Following EV purification and RNA isolation, RNA-Seq determined that there were 126 EV miRNAs differentially expressed between the two groups when comparing their RNA cargoes. Ingenuity Pathways Analysis found 9 upregulated miRNAs that were associated with inflammation (miR-127-5p, miR-1273h-3p, miR-1288-5p, miR-130b-5p, miR-139-3p, miR-1910-5p, miR-203b-5p, miR-22-5p, and miR-4632-3p; all p < 0.049; fold regulation range = 1.43-1.67). CONCLUSION: This study determined that EVs are present in the tear film and that tear EVs contain miRNAs that may be associated with DED inflammatory pathways.

Adenovirus infection activates akt1 and induces cell proliferation in pancreatic islets1.

BACKGROUND: : Accumulated evidence has shown that insulin producing beta-cells in pancreatic islets have the limited potential to regenerate. Adenoviruses have been widely employed to deliver genes of interest into pancreatic islets. This study was aimed at investigating whether adenovirus infection has any impact on the potential of beta-cell proliferation. METHODS: : Human adenovirus type 5 (Ad5) encoding rat insulin promoter driven reporter genes were used to infect freshly isolated pancreatic islets. Western blotting assays were performed to evaluate the expression and activation of key molecules involved in cell survival and proliferation following Ad5 infection. Immunofluorescence staining was employed to identify proliferating cells after culturing the infected and control islets in the presence of BrdU, an analog of thymidine that can be incorporated into the genome of proliferating cells. RESULTS: : Ad5 infection of the islets resulted in expression and activation of Akt1, a key molecule in the PI3 kinase signaling pathway. Accordingly, a higher frequency of islet cell proliferation was detected in Ad5-infected islets than in control islets. DISCUSSION: : These data suggest adenovirus infection can activate beta-cell survival and proliferation machinery, in particular operating through the PI3K/Akt signaling pathway. This information has significant ramification for the use of adenovirus as a gene delivery vehicle for pancreatic islet cells.