Lipopolysaccharide up-regulates IP-10 expression through the activation of NF-κB in rat peritoneal mesothelial cells

Abstract

Objective To observe the expression of interferon induced protein (IP) -10 and the role of nuclear factor-kappaB (NF-κB) signaling pathway in rat peritoneal mesothelial cells (RPMCs) under the action of lipopolysaccharide (LPS). Methods RPMCs were harvested from Sprague-Dawley rat peritoneal cavity and cultured under defined in vitro conditions. The cells were exposed respectively to different concentrations of LPS (0, 10, 100, 1 000, 10 000 ng/ml) for 3 h or treated with LPS (100 ng/ml) for different time points (0, 1, 3, 6, 12, 24, 48 h). For observing the effect of LPS on the expression of p-p65 and p65, the RPMCs were treated with LPS (100 ng/ml) for different time points (0, 15, 30, 60, 120 min). For observing the effect of BAY11-7085 on the expression of IP-10 mRNA, the RPMCs were treated by LPS or pretreated with BAY11-7085 (5 μmol/L) for 2 h, then treated with LPS for another 3 h, respectively. Expression of IP-10 mRNA was examined by reverse transcription-polymerase chain reaction (RT-PCR). Expression of NF-κB and p-NF-κB protein was detected by Western blot. The secretion of IP-10 was determined by enzyme-linked immunosorbent assay (ELISA). Results Compared with the control group, stimulation of RPMCs with 10 ng/ml LPS resulted in a significant increase in the expression of IP-10 mRNA (P<0.05). 1 000 ng/ml LPS has the strongest effect on IP-10 expression compared with that of 10 ng/ml and 100 ng/ml LPS. Treatment with 100 ng/ml LPS resulted in time-dependent increase in the gene level of IP-10, with the peak at 3 h. However, after that time point, the gene level of them was gradually attenuated. Following treatment with LPS (100 ng/ml), the level of p-NF-κB began to increase at 15 min, gradually reached the peak at 1 hour, and then decreased. But the level of which at 2 h is still significant higher than that of medium control. 5 μmol/L BAY11-7085 significantly decreased the up-regulation of IP-10 induced by LPS. Conclusions LPS enhanced the expression of IP-10 on RPMCs in a concentration-dependent and a time-dependent manner. LPS induced expression of IP-10 depended on the NF-κB signal transduction pathway. Key words: Lipopolysaccharides/PD; Peritoneum/CY; Epithelium/DE; Chemokine CXCL10/ME; NF-kappa B; Rats