Effect of LFA-1 Deficiency on Differentiation of Mice Na?ve T Cells into Th17 Cells in vitro / 胃肠病学

ABSTRACT

BacKground:Lymphocyte function-associated antigen-1( LFA-1 ) pIays a cruciaI roIe in the pathogenesis of infIammatory boweI disease by reguIating the activation and function of T ceIIs. Aims:To investigate the effect of LFA-1 deficiency( LFA-1-/-)on differentiation of mice na?ve T ceIIs into Th17 ceIIs in vitro. Methods:LFA-1-/- mice were breeded and the progeny genome DNA was extracted from the taiIs for genotyping by PCR. CD4+CD62L+ na?ve T ceIIs were separated from spIenic mononucIear ceIIs of LFA-1-/- progeny mice and wiId type ( WT ) C57BL/6J mice, respectiveIy,by magnetic-activated ceII sorting( MACS),and then the purity of separated ceIIs was determined. Na?ve T ceIIs obtained were cuItured in different inducing systems[ transforming growth factor-β( TGF-β),TGF-β + interIeukin-6 (IL-6),and TGF-β + IL-6 + IL-23]in vitro for Th17 ceII differentiation;ceIIs in each inducing system were coIIected for anaIyzing the ratio of Th17 ceIIs by fIow cytometry,and the expressions of Th17-specific transcription factor ROR-γt and Th17-specific marker IL-17A were measured by qRT-PCR and ELISA methods. Results:AII fifteen progeny mice were identified as LFA-1-/- genotype. Purity of CD4+CD62L+ na?ve T ceIIs separated by MACS was above 95%. Th17 ceIIs couId be induced by Iow-dose TGF-βcombined with IL-6,and the differentiation ratio was increased obviousIy when IL-23 was added. In inducing system containing TGF-β,IL-6 and IL-23,na?ve T ceIIs from LFA-1-/- mice produced more Th17 ceIIs than those from WT mice(17. 2% ± 1. 4% vs. 5. 7% ± 0. 2%,P<0. 001),expressions of ROR-γt mRNA and IL-17A mRNA were up-reguIated(P<0. 001),and IL-17A concentration in ceII cuIture supernatant was increased(P<0. 01). Conclusions:Deficiency of LFA-1 promotes the differentiation of mice na?ve T ceIIs into Th17 ceIIs in vitro.