The Effect Of Antioxidative Stress On The MAPK And STAT Signalling In LPS-induced Mice Liver And Kupffer Cells

Objective: To investagete the effects of antioxidant N-acetylcysteine (NAC) on MAPK and STAT signalling pathway of liver and kupffer cell in LPS-induced liver injury.Methods: The experiment (Exp) 1. Male 54 mice were divided into three groups. (1) Control (n=6): 0.9 % Sodium Chloride 0.2 ml, i.p.. (2) LPS group (n=24): (GalN 520 mg+LPS 120μg)/kg i.p.. (3) NAC+LPS group (n=24): NAC (150 mg/kg) was administered in lavage for 3 days. At 3rd day, GalN/LPS (520 mg+120μg/kg) was injected after 1h of NAC administration.The blood was gathered from eye vein and liver was excised with carbrital anaesthesia after LPS or 0.9 % Sodium Chloride injected at 0.5h, 1h, 2h and 6h for ALT, GSH and MDA assaies for each groups. Exp 2. The animal and groups were same as Exp 1. (1) Control (n=6): 0.9 % Sodium Chloride 0.2 ml, i.p.. (2) LPS group (n=24): LPS 5 mg/kg i.p.. (3)NAC+LPS group(n=24): NAC (150 mg/kg) was administered as Exp 1. LPS (5 mg/kg i.p.) was injected after 1h of NAC administration. The livers were excised in anaesthetic state by carbrital after LPS or 0.9 % Sodium Chloride injected at 0.5h, 1h, 2h and 6h and the protein extracted from livers was assayed for the phosphorylation level of MEK1/2, ERK1/2, p38MAPK, STAT1, STAT3 and expression of HSP70 by western blotting. TNF-αin liver was assayed by radioimmunoassay. Exp 3. KCs was isolated from 40 mice in asepsis and cultured for 24 h.Afterwards, KCs was divided into 3 groups. (1) Control: KCs and supernatant of DMEM mediam was collected directly without any treatment. (2)LPS group: LPS (100 ng/ml) was added in DMEM and stimulated KCs for 0.5h and 1h respectively, then KCs and mediam were collected. (3) NAC+LPS group: After KCs was cultured with NAC (10 mmol/L) in DMEM madiam for 2h, LPS was added in DMEM mediam to stimulate KCs for 0.5 h or 1 h. Afterwards, KCs and mediam ware collected. KCs protein and TNF-αin supernatant was assayed as same as Exp 2.Results:GalN/LPS-induced acute liver injury was attenuated significantly by NAC pretreatment. The plasma ALT activity and liver MDA were decreased remarkably by NAC preteatment and the GSH of liver was increased significantly. The phosphorylation of MEK1/2, ERK1/2 and p38MAPK in KCs and liver were enhanced significantly campared with control after LPS stimulation. p-MEK1/2 and p-ERK1/2 was the highest level at 0.5h than others and came back basal level at 2h after LPS sitmulation. p-p38MAPK was being maintained at a high level after 0.5h. Meanwhile, TNF-αproduction in liver and in isolated KCs was increased observably too. The phosphorylation of STAT1 amd STAT3 was appeared at 0.5h and up to top...