ABSTRACT
Purpose: Thymic stromal lymphopoietin (TSLP) is a pro-allergic cytokine that initiates allergic inflammatory reaction between epithelial and dendritic cells (DCs). miR-19b was reported to suppress TSLP expression. The present study aimed to examine miR-19b expression, regulation, and function in allergic conjunctivitis (AC). Methods: A murine model of experimental AC was induced in BALB/c mice by short ragweed pollen. The serum, eye balls, conjunctiva, and cervical lymph nodes (CLN) were used for the study. Gene expression was determined by RT-PCR, whereas protein production and activation were evaluated by immunostaining, ELISA, and Western blotting. Results: In the murine AC model, miR-19b was aberrantly downregulated, whereas the levels of TSLP and p-STAT3, as well as the number of CD11c+ pSTAT3+ DCs were increased. Moreover, Th2 inflammatory cytokine expression was significantly increased. These severe phenotypes could be counteracted by either applying exogenous miR-19b mimic microRNAs or the JAK/STAT inhibitor CYT387. Moreover, overexpression of miR-19b repressed p-STAT3 expression and the number of CD11c+ cells in AC eye and CLN tissues. Conclusions: These findings suggested that miR-19b reduced ocular surface inflammation by inhibiting Stat3 signaling via TSLP downregulation in a murine AC model. Moreover, the present study further demonstrated the clinical potential of applying miR-19b and anti-JAK/STAT therapies in the treatment of AC.
Subject(s)
Conjunctivitis, Allergic/genetics , Janus Kinases/physiology , MicroRNAs/genetics , STAT Transcription Factors/physiology , Animals , Antigens, Plant , CD11 Antigens/metabolism , Cervical Vertebrae , Conjunctiva/metabolism , Conjunctivitis, Allergic/immunology , Conjunctivitis, Allergic/metabolism , Cornea/metabolism , Cytokines/biosynthesis , Disease Models, Animal , Down-Regulation , Female , Janus Kinases/antagonists & inhibitors , Lymph Nodes/metabolism , Mice, Inbred BALB C , MicroRNAs/biosynthesis , Phenotype , Plant Extracts , STAT Transcription Factors/antagonists & inhibitors , STAT3 Transcription Factor/metabolism , Signal Transduction , Thymic Stromal LymphopoietinABSTRACT
BACKGROUND: Pulmonary arterial smooth muscle cell (PASMC) proliferation in response to hypoxia plays an important role in the vascular remodelling that occurs in hypoxic pulmonary hypertension. MicroRNAs (miRs) are emerging as important regulators in the progression of pulmonary hypertension. In this study, we investigated whether the expression of miR-17-5p is modulated by hypoxia and is involved in the hypoxia-induced proliferation of PASMCs. METHODS: Human PASMCs were cultured under hypoxic conditions. miR-17-5p expression was determined by real-time RT-PCR. A BrdU incorporation assay and time-lapse recording were utilized to determine cell proliferation and migration. RESULTS: PASMC proliferation was increased by moderate hypoxia (3% oxygen) but was reduced by severe hypoxia (0.1% oxygen) after 48 h. Moderate hypoxia induced miR-17-5p expression. Overexpression of miR-17-5p by transfection with miR-17-5p enhanced cell proliferation and migration in normoxia, whereas knockdown of miR-17-5p with anti-miR-17-5p inhibitors significantly reduced cell proliferation and migration. The expression of miR-17-5p target genes, specifically phosphatase and tensin homologue (PTEN) and cyclin-dependent kinase inhibitor 1 (p21WAF1/Cip1, p21), was reduced under moderate hypoxia in PASMCs. Under normoxia, overexpression of miR-17-5p in PASMCs reduced the expression of PTEN and p21. CONCLUSION: Our data indicate that miR-17-5p might play a significant role in hypoxia-induced pulmonary vascular smooth muscle cell proliferation by regulating multiple gene targets, including PTEN and p21, and that miR-17-5p could be a novel therapeutic target for the management of hypoxia-induced PH.
