Role of activated protein C in lipopolysaccharide induced microglia activation

Abstract

Objective To investigate the role of active protein C (APC) in lipopolysaccharide (LPS) induced microglia activation. Methods Microglia from one day old Sprague-Dawley newborn rat was collected, purified and identified by primary culture and immunofluorescence staining, and then was randomly divided into four groups including LPS group (1.0 μg/ml LPS plus 10 μl phosphate buffered saline 12 h later), LPS+ APC group (1.0 μg/ml LPS plus 0.1 μg/ml APC 12 h later), APC group (10 μl phosphate buffered saline plus 0.1 μg/ml APC 12 h later) and control group (10 μl phosphate buffered saline at each time point). The morphology of micaroglia in all groups was observed under microscope, and the expression of tumor necrosis factor-α (TNF-α) and protease-activated receptor-1 (PAR-1) were determined by immunofluorescence staining. One-way analysis of variance and LSD test were applied for statistical analysis. Results Primary culture microglia was successful and the purity was no less than 99%. In LPS group, the microglia morphology was activated, and the expression of TNF-α was increased significantly than the control group (2.11±0.35 vs 1.38±0.28, LSD test, P=0.002). In LPS+APC group, the microglia morphological change was reversed, and the expression of TNF-α had no significant difference with the control group (1.35±0.36 vs 1.38±0.28, LSD test, P>0.05). The expression of PAR-1 in LPS+APC group was higher comparing with that in the control group (4.60±0.84 vs 2.64±0.41, LSD test, P=0.008) and the LPS group (2.44±0.86, LSD test, P=0.002). The expression of PAR-1 in APC group and LPS group had no obvious difference with control group (2.62±0.69, 2.44±0.86 vs 2.64±0.41, LSD test, both P>0.05). Conclusions By increasing the level of PAR-1 in microglia, active protein C could inhibit the activation of miciroglia and the expression of TNF-α induced by lipopolysaccharide, therefore, protecting the brain tissues from inflammation-induced damage. Key words: Microglia; Protein C; Tumor necrosis factor-alpha; Lipopolysaccharides