Effect Of Opening Of Mitochondrial ATP-sensitive K~+ Channel On The Expression Of HIF-1a And Cell Proliferation In Pulmonary Arterial Smooth Muscle Cells Of Rats

Objective:The objective of this paper was to investigate the effect of mitochondrial ATP-sensitive K+ channel on the expression of HIF-1a and cell proliferation in pulmonary arterial smooth muscle cells (PASMCs) of rats.Methods:Cultured PASMCs were divided into six groups, as follows: (1) normoxia group: cultured under normoxia; (2) normoxia + diazoxide group: cultured in normoxia with diazoxide, an opener of MitoKATP; (3) normoxia +5-HDgroup: cultured in normoxia with 5-hydroxydecanoate (5-HD), an antagonist of MitoKATP; (4) hypoxia group: cultured under hypoxia(37℃,5%O2,5%CO2,90%N2) for 24 hours; (5) hypoxia+ diazoxide group, cultured under hypoxia(37℃,5%O2,5%CO2,90%N2) with diazoxide for 24 hours; (6) hypoxia + 5-HD group, cultured under hypoxia(37℃,5%O2,5%CO2,90%N2) with 5-HD for 24 hours. The relative changes in mitochondrial potential were tested with Rhodamine fluorescence (R-123) technique. Immunohistochemical method was used to trace the expression of HIF-1a. The proliferation of PASMCs was examined by MTT colorimetric assay.Results: The results were as following: The intensity of R-123 fluorescence in normoxia+diazoxide group was significantly increased as compared with normoxia group (P<0.05),and the intensity of R-123 fluorescence in hypoxia+diazoxide group was also significantly increased as compared with hypoxia group (P<0.05). 24-hour hypoxia or 24-hour hypoxia + diazoxide markedly increase the intensity of R-123 fluorescence in PASMC as compared with normoxia group (P<0.05),and the change was more prominant in hypoxia + diazoxide group than that of hypoxia group (P<0.05). There was no significant difference in the intensity of R-123 fluorescence between normoxia group and normoxia + 5-HD group(P>0.05). However, 5-HD weakened the effect of 24-hour hypoxia on the intensity of R-123 fluorescence. The intensity of R-123 fluorescence in hypoxia + 5-HD group was significantly decreased as compared with hypoxia group (P<0.05). After exposed to hypoxia or hypoxia + diazoxide for 24 hours, the expression of HIF-1a and the proliferation of PASMCs were significantly increased as compared with normoxia or normoxia + diazoxide group (P<0.05). 24-hour hypoxia or 24-hour hypoxia + diazoxide markedly increased the expression of HIF-1a and the proliferation of PASMCs as compared with normoxia group (P<0.05), the change was more significant in hypoxia + diazoxide group than that of hypoxia group (P<0.05).There were no significant difference in the expression of HIF-1a and the proliferation of PASMCs between normoxia group and normoxia + 5-HD group(P>0.05). Since 5-HD could weaken the effect of 24-hour hypoxia, the expression of HIF-1a and the proliferation of PASMCs in hypoxia + 5-HD group were significantly decreased as compared with hypoxia group (P<0.05).Conclusion:All these results suggest that the opening of MitoKATP followed by a depolarization ofΔΨ_m might contribute to the increase of the expression of HIF-1a and the proliferation of PASMCs. This might be a mechanism of the development of hypoxic pulmonary hypertension. The opening of mitochondrial ATP-sensitive K~+ channel and the depolarization of mitochondrial membrane might play an important role in the expression of HIF-1a and the proliferation of PASMCs.