[Effect of potassium channel on the proliferation, apoptosis and related-gene expression in human bronchial smooth muscle cells]

Abstract

Objective: To investigate the effects of three potassium channels, voltage-dependent K(+) channel (K(V)), calcium-activated K(+) channel (K(Ca)) and ATP-sensitive K(+) channel (K(ATP)), on the proliferation, apoptosis and related-gene expression in human bronchial smooth muscle cells (HBSMCs).

Methods: Normal human bronchial tissues were obtained from 5 patients undergoing lung partial resection for carcinomas. The cultured HBSMCs were divided into four groups: (1) control group; (2) 4-aminopyridine (4-AP) group: containing 4 mmol/L 4-AP; (3) TEA group: containing 1 mmol/L TEA; (4) Glib group: containing 0.1 mmol/L Glib. The cell cycles were observed by flow-cytometry; Ca(2+) concentration in HBSMCs were investigated by fluorescent quantification using fluorospectrophotometer. The proliferation and apoptosis were detected by MTT methods and TUNEL, respectively. Immunocytochemical staining was used to detect the effects of three potassium channel blockers on the expressions of PCNA and apoptosis related-gene Fas, FasL of HBSMCs.

Results: (1) The K(V) blocker 4-AP was shown to significantly increase the optical density value [the value of A of the 4-AP group was (0.67 +/- 0.14), compared to the control group (0.30 +/- 0.08), P< 0.01] and the expression of proliferating cell nucleus antigen [the positive percentage of PCNA of the 4-AP group was (89 +/- 7)%, compared to the control group (23 +/- 5)]%, P < 0.01). 4-AP significantly increased the Ca(2+) concentration [(255 +/- 17) nmol/L] in cultured HBSMCs, compared to the control group [(98 +/- 7) nmol/L, P < 0.01] and the numbers [(28.8 +/- 2.4)%] of S + G(2)M HBSMCs by flow-cytometry in cultured HBSMCs, compared to the control group [(12.6 +/- 4.8)%, P < 0.01]. The value of A of TEA group and Glib group were all (0.30 +/- 0.07). The positive percentage of PCNA of the two groups were (21 +/- 5)%, (20 +/- 4)%, respectively. Ca(2+) concentration of the two groups were (97 +/- 7) nmol/L, (99 +/- 6) nmol/L, respectively. And the numbers of S + G(2)M HBSMCs were (12.8 +/- 4.4)%, (11.0 +/- 4.4)%, respectively. But no significant differences were found between the two groups and the control group (all P > 0.05). (2) The level of TUNEL and the expressions of Fas and FasL of the control were (4.36 +/- 0.66)%, (2.92 +/- 0.25)%, (4.0 +/- 0.6)%, respectively. 4-AP significantly decrease the apoptosis rate and the expressions of Fas, FasL [(0.84 +/- 0.13)%, (0.92 +/- 0.16)]%, (1.4 +/- 0.6)%, respectively) (compared to control, all P < 0.01). The level of TUNEL of TEA group and Glib group were (4.47 +/- 0.93)%, (4.33 +/- 0.77)%, respectively. And the expressions of Fas, FasL of the two groups were (2.87 +/- 0.23)%, (4.2 +/- 0.8)%, (2.91 +/- 0.26)% and (4.2 +/- 0.9)%, respectively. There were no significant differences between the two groups and the control group (all P > 0.05).

Conclusion: Inhibition of K(V) activity can increase the proliferation and intracellular [Ca(2+)]i, and decrease the apoptosis of HBSMCs, but inhibition of K(Ca) and K(ATP) showed no effects.