Effects of lipopolysaccharide on the expression of tribbles homolog 3 in acute lung injury / 安徽医科大学学报

ABSTRACT

Objective To examine the expression of tribbles homologous 3 (TRB3) on lipopolysaccharide (LPS) induced acute lung injury (ALI) and its relationship with p38-mitogen-activated protein kinase (MAPK) pathway.Methods Rats received a intravenous injection by LPS (5 ml/kg) as models of ALI and a intravenous injection by NS (5 ml/kg) as the control.In rat lung tissue the expression of TRB3 protein was examined using immunohistochemical staining, the expression of TRB3 mRNA was determined by reverse transcript polymerase chain reaction (RT-PCR).Cultured rat pulmonary microvascular cells (PMVEC) were randomly divided into dose-dependent, time-dependent and intervention groups in vitro.In dose-dependent group, PMVEC were stimulated by various concentrations of LPS (0, 2, 4, 10 μg/ml) for 4 h, and in time-dependent group PMVEC were challenged by 10 μg/ml LPS for different time (0, 4, 8, 12 h).In intervention group, PMVEC grown in normal medium or medium with 10 μg/ml LPS for 4 h were pretreated using p38-MAPK inhibitor (10 μmol/L SB203580) for 2 h.Western blot was used to examine expression of TRB3, p-p38 and p38-MAPK.Results Immunohistochemical staining showed that TRB3 protein distributed in rat alveolar walls and glandular epithelium.Increased TRB3 mRNA expression using RT-PCR were found in lung tissue of rats injected by LPS when compared to those in NS group (t=15.524, P<0.01). Increased TRB3 mRNA expression using RT-PCR had also been found in PMVEC stimulated by LPS when compared to those in NS group (t=7.549, P<0.01). In PMVEC, LPS significantly increased the expression of TRB3 protein in a dose-dependent manner (0, 2, 4, 10 μg/ml) after stimulation for 4 h (F=12.619, P<0.001).At indicated time-points after PMVEC were challenged by 10 μg/ml LPS, the expression of TRB3 protein raised at 4 h , then decreased gradually at 8 h , but still was higher than 0 h group , there were significant difference (F=11.273, P<0.001). LPS significantly increased the expression of p-p38 protein after stimulation for 4 h when compared to the control group, LPS also increased the expression of TRB3 protein after stimulation for 4 h when compared to the control group (t=49.121,15.113,P<0.001). SB203580 decreased the protein levels of p-p38 in response to LPS, SB203580 also decreased the protein levels of TRB3 in response to LPS(t=7.040,11.900,P<0.05,0.001).SB203580 alone had no effect on the expression of p-p38 and TRB3,when compared to the control group,there was not statistically significant.Conclusion Levels of TRB3 protein increase in LPS-induced acute lung injury, and is regulated by p38-MAPK pathway.