CAJ | 학술논문

目的从体外观察ERK信号通路在甲状旁腺激素(PTH)致人近曲小管上皮细胞(HK-2)合成纤溶酶原激活物抑制物1(PAI-1)中的作用.方法以HK-2细胞株为研究对象,用不同浓度PTH(10-8、10-9、10-10、10-11、10-12 mol/L)作用细胞48 h,以及10-10 mol/LPTH 作用细胞不同时间(12、24、36、48、72 h),分别用RT-PCR法检测PAI-1 mRNA表达,Western 印迹法检测PAI-1蛋白表达.以10-10 mol/L PTH作用细胞48 h,分别观察细胞ERK 1/2抑制剂预处理前后磷酸化(p)ERK1/2、PAI-1mRNA及蛋白的变化情况.结果 10-12 mol/L PTH可促进细胞在基因及蛋白水平合成PAI-1,随着PTH浓度逐渐上升,PAI-1 mRNA及蛋白浓度均相应增加,以10-10 mol/L PTH组刺激作用最显著,分别为对照组的4.01倍和3.81倍(均P＜0.01).但随着PTH浓度进一步增加,PAI- 1mRNA及蛋白浓度却随之下降.10-10 mol/L PTH作用细胞,12 h时有少量PAI-1表达,72 h时达峰值,并呈时间依赖性,分别为0h组的4.06倍和4.03倍(均P＜0.01).10-10 mol/L PTH作用细胞48 h,有大量的p-ERK1/2合成(P＜0.01),经ERK 1/2抑制剂预处理后,PAI-1及ERK均显著下降(均P＜0.01),但仍高于对照组(均P＜ 0.05).结论 ERK信号通路部分参与PTH致HK-2细胞合成PAI-1的作用.
목적 종체외관찰ERK신호통로재갑상방선격소(PTH)치인근곡소관상피세포(HK-2)합성섬용매원격활물억제물1(PAI-1)중적작용.방법 이HK-2세포주위연구대상,용불동농도PTH(10-8、10-9、10-10、10-11、10-12 mol/L)작용세포48 h,이급10-10 mol/LPTH 작용세포불동시간(12、24、36、48、72 h),분별용RT-PCR법검측PAI-1 mRNA표체,Western 인적법검측PAI-1단백표체.이10-10 mol/L PTH작용세포48 h,분별관찰세포ERK 1/2억제제예처리전후린산화(p)ERK1/2、PAI-1mRNA급단백적변화정황.결과 10-12 mol/L PTH가촉진세포재기인급단백수평합성PAI-1,수착PTH농도축점상승,PAI-1 mRNA급단백농도균상응증가,이10-10 mol/L PTH조자격작용최현저,분별위대조조적4.01배화3.81배(균P＜0.01).단수착PTH농도진일보증가,PAI- 1mRNA급단백농도각수지하강.10-10 mol/L PTH작용세포,12 h시유소량PAI-1표체,72 h시체봉치,병정시간의뢰성,분별위0h조적4.06배화4.03배(균P＜0.01).10-10 mol/L PTH작용세포48 h,유대량적p-ERK1/2합성(P＜0.01),경ERK 1/2억제제예처리후,PAI-1급ERK균현저하강(균P＜0.01),단잉고우대조조(균P＜ 0.05).결론 ERK신호통로부분삼여PTH치HK-2세포합성PAI-1적작용.
Objective To explore the role of ERK1/2 in the expression of the type-1 plasminogen activator inhibitor (PAI-1) induced by parathonnone (PTH) in human renal tubular epithelial cell line HK-2 cells. Methods Various concentrentions of PTH and manifold durations were applied in the test.The expression of PAI-1 mRNA and protein in HK-2 cells was measured by RT-PCR and Western blotting,respectively.Besides,ERK1/2 protein was detected by Western blotting before the ERK1/2 inhibitor incubated with the HK-2 cells or after. Results The expression of PAI-1mRNA and protein was gradually up-regulatad along with the increasing concentrations of PTH (10-12-10-10 mol/L).The maximum level of PAI-1 mRNA and protein was detected in 10-10 mol/L PTH and was 4.01 and 3.81 times of control group.Otherwise,the decreased expression of PAI-1 was found while the concentrations of PTH were beyond 10-10 mol/L.The levels of PAI-1 mRNA and protein were increased in pace with time from 12 to 72 hour,in time-dependent manner,which was 4.06 (12 h) and 4.03 (72 h) times of 0 hour group.The levels of ERK1/2 and PAI-1 were ascended after 10-10 mol/L PTH incubated with the HK-2 cells (all P＜0.01).Howerver,both of them decended after cells were pretreated by the ERK1/2 inhibitor (all P＜0.01),but were still higher than those of control group (all P＜0.05). Conclusion ERK1/2 kinase system partly participates in the regulation of PAI-1 induced by PTH in HK-2 cells.