Relationship between HO-1/CO signaling pathway and mitochondrial fission in LPS-challenged alveolar type Ⅱ epithelial cells of rats / 中华麻醉学杂志

ABSTRACT

Objective:To evaluate the relationship between heme oxygenase-1/carbon monoxide (HO-1/CO) signaling pathway and mitochondrial fission in lipopolysaccharide (LPS)-challenged alveolar type Ⅱ epithelial cells (AECⅡ) of rats.Methods:The cultured AECⅡ were subcultured and inoculated in 96-well plates (about 2 × 10 5 cells/ml) and divided into 7 groups ( n=12 each) using a random number table method: blank control group (group C), LPS group (group L), carbon monoxide release molecule-2 (CORM -2) + LPS group (group CL), HO-1 inducer hemin+ LPS group (group HL), HO-1 inhibitor ZnPP-Ⅸ+ LPS group (group ZL), CORM-2+ ZnPP-Ⅸ+ LPS group (group CZL) and hemin+ ZnPP-Ⅸ+ LPS group (group HZL). In group C, the cells were continuously incubated and received no treatment.Cells were incubated with LPS 10 μg/ml in group LPS. In CL, HL and ZL groups, cells were incubated with 100 μmol/L CORM-2 for 1 h, with 20 μmol/L hemin for 1 h and with 10 μmol/L ZnPP- Ⅸ for 0.5 h, respectively, and then the cells were incubated with 10 μg/ml LPS.In group CZL, the cells were incubated with 100 μmol/L CORM-2 for 1 h, then with 10 μmol/L ZnPP-Ⅸ for 0.5 h, and finally with 10 μg/ml LPS.In group HZL, the cells were incubated with 20 μmol/L hemin for 1 h, then with 10 μmol/L ZnPP-Ⅸ for 0.5 h, and finally with 10 μg/ml LPS.Cell viability was evaluated by methyl thiazolyl tetrazolium assay at 48 h of culture or incubation with LPS.The expression of HO-1, dynamin-related protein 1 (Drp1) and mitochondrial fission protein 1 (Fis1) was determined using Western blot. Results:Compared with group C, the cell viability was significantly decreased, and the expression of HO-1, Drp1 and Fis1 was up-regulated in other six groups ( P<0.05). Compared with group L, the cell viability was significantly increased, the expression of HO-1 was up-regulated, and the expression of Drp1 and Fis1 was down-regulated in CL and HL groups, the cell viability was decreased, HO-1 expression was down-regulated and the expression of Drp1 and Fis1 was up-regulated in group ZL ( P<0.05), and no significant change was found in the parameters mentioned above in CZL and HZL groups ( P>0.05). Compared with CL and HL groups, the cell viability was significantly decreased, HO-1 expression was down-regulated, and the expression of Drp1 and Fis1 was up-regulated in group ZL ( P<0.05). Conclusion:HO-1/CO signaling pathway can inhibit mitochondrial fission, which exerts endogenous protection when LPS induces injury to AEC Ⅱ in rats.