Small hairpin RNA silencing Tim-3 affects peripheral blood Th1 and Th17 cells differentiation in asthmatic mice / 中国当代儿科杂志

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effects of down-regulating Tim-3 gene in the peripheral blood mononuclear cells (PBMCs) of an asthmatic mouse model by short hairpin RNA (shRNA) and to explore the effect of Tim-3 on Th1 and Th17 cell differentiation.</p><p><b>METHODS</b>An asthmatic murine model was established by ovalbumin sensitization and challenge. PBMCs were isolated from asthmatic mice and transfected by shRNA targeting Tim-3 gene. The mRNA and protein expressions of Tim-3 were detected by quantitative PCR and Western blot. Flow cytometry analysis was performed to determine the levels of Th1 and Th17, and ELISA was performed to determine concentrations of IFN-γ, IL-4 and IL-17 in the supernatant.</p><p><b>RESULTS</b>Tim-3 mRNA expression in PBMCs was significantly increased in asthmatic mice. The mRNA and protein expression of Tim-3 decreased significantly in the shRNA group. Compared with the negative groups, Th1 cell levels increased and Th17 cell levels decreased significantly in the asthmatic groups after Tim-3 shRNA interference. In the Tim-3 shRNA interference groups concentrations of IFN-γ increased significantly while IL-17 decreased significantly.</p><p><b>CONCLUSIONS</b>Specific Tim-3 shRNA effectively silences the expression of Tim-3 and change in Tim-3 expression could affect T cell differentiation.</p>