Xu Shu-yun, Xu Yong-jian, Zhang Zhen-xiang, Ni Wang, Chen Shi-xin
Department of Respiratory Medicine, Tongji Hospital Affiliated to Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Zhonghua Jie He He Hu Xi Za Zhi. 2003 Dec;26(12):756-60.
To investigate the role of protein kinase C (PKC) signal pathway in airway smooth muscle cell (ASMCs) proliferation in asthmatic rats.
(1) Forty-eight Wistar rats were divided into an asthmatic group (group A) and a control group (group B), and then subdivided into group A(1), A(2), A(3) and group B(1), B(2), B(3) based on the time of challenge (2, 4, 8 weeks). The proliferation of ASMCs isolated from group A and B was examined with cell cycle analysis, MTT colorimetric assay and proliferating cell nuclear antigen (PCNA) immunocytochemical staining. (2) ASMCs from group A(1) and B(1) were treated with PKC agonist PMA and inhibitor Ro-31-8220, and their proliferations were examined. (3) The expression of PKC-alpha in group A(1), A(2), A(3) and B(1) was observed by immunocytochemical staining and RT-PCR, respectively.
(1) The percentage of S phase, absorbance (A) value and the expression rate of PCNA protein in ASMCs from group A were significantly increased compared to those of group B (P < 0.01). (2) Before ASMCs from A(1) group were treated, the percentage of S phase, A value and the expression rate of PCNA were (19 +/- 3)%, 0.459 +/- 0.036, and (80 +/- 10)% respectively; After treated with 10 nmol/L PMA, these values reached (27 +/- 4)%, 0.599 +/- 0.078, and (95 +/- 9)% respectively; After treated with 50 nmol/L PMA, these values were (14 +/- 3)%, 0.346 +/- 0.038, and (53 +/- 8)% respectively; After treatment with Ro-31-8220, these values were (14 +/- 3)%, 0.343 +/- 0.048, and (49 +/- 8)% respectively. Statistical analysis revealed that there were significant differences in the percentage of S phase, A value and the expression of PCNA before and after treatment (P < 0.01). After treatment with 50 nmol/L PMA, there was a significant difference as compared to 10 nmol/L PMA treatment (P < 0.01), whereas there was no statistical significance as compared to Ro-31-8220 treatment (P > 0.05). (3) Compared with that of group B(1) (0.84 +/- 0.07), the ratios of A value of PKC-alpha mRNA in group A(1) (1.08 +/- 0.06), group A(2) (1.12 +/- 0.05), and group A(3) (1.10 +/- 0.08) were significantly increased (P < 0.01). Compared with that of group B(1) (28 +/- 4)%, the expression rates of PKC-alpha protein in group A(1) (54 +/- 5)%, group A(2) (56 +/- 5)%, and group A(3) (53 +/- 6)% were significantly increased (P < 0.01).
Airway smooth muscle cell proliferation increases in asthmatic rats. PKC and it's alpha subtype may contribute to this process.
探讨蛋白激酶C（PKC）信号通路在哮喘大鼠气道平滑肌细胞（ASMCs）增殖中的作用。
（1）将48只Wistar大鼠分为哮喘组（A组）和对照组（B组），然后根据激发时间（2、4、8周）将其细分为A（1）、A（2）、A（3）组和B（1）、B（2）、B（3）组。采用细胞周期分析、MTT比色法和增殖细胞核抗原（PCNA）免疫细胞化学染色检测A组和B组分离的ASMCs的增殖情况。（2）用PKC激动剂PMA和抑制剂Ro-31-8220处理A（1）组和B（1）组的ASMCs，并检测其增殖情况。（3）分别通过免疫细胞化学染色和RT-PCR观察A（1）、A（2）、A（3）组和B（1）组中PKC-α的表达。
（1）与B组相比，A组ASMCs的S期百分比、吸光度（A）值和PCNA蛋白表达率显著升高（P<0.01）。（2）A（1）组ASMCs处理前，S期百分比、A值和PCNA表达率分别为（19±3）%、0.459±0.036和（80±10）%；用10 nmol/L PMA处理后，这些值分别达到（27±4）%、0.599±0.078和（95±9）%；用50 nmol/L PMA处理后，这些值分别为（14±3）%、0.346±0.038和（53±8）%；用Ro-31-8220处理后，这些值分别为（14±3）%、0.343±0.048和（49±8）%。统计学分析显示，处理前后S期百分比、A值和PCNA表达有显著差异（P<0.01）。用50 nmol/L PMA处理后与用10 nmol/L PMA处理相比有显著差异（P<0.01），而与用Ro-31-8220处理相比无统计学意义（P>0.05）。（3）与B（1）组（0.84±0.07）相比，A（1）组（1.08±0.06）、A（2）组（1.12±0.05）和A（3）组（1.10±0.08）中PKC-α mRNA的A值比值显著升高（P<0.01）。与B（1）组（28±4）%相比，A（1）组（54±5）%、A（2）组（56±5）%和A（3）组（53±6）%中PKC-α蛋白的表达率显著升高（P<0.01）。
哮喘大鼠气道平滑肌细胞增殖增加。PKC及其α亚型可能参与了这一过程。
