中国骨质疏松杂志
中國骨質疏鬆雜誌
중국골질소송잡지
CHINESE JOURNAL OF OSTEOPOROSIS
2009年
2期
115-122
,共8页
丝裂原活化蛋白激酶%核心结合因子α1%淫羊藿甙%成骨细胞
絲裂原活化蛋白激酶%覈心結閤因子α1%淫羊藿甙%成骨細胞
사렬원활화단백격매%핵심결합인자α1%음양곽대%성골세포
Mitogen-activated protein kinase%Core binding factor-1%Icariin%Osteoblasts
目的 观察淫羊藿甙对体外培养大鼠成骨细胞丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)信号通路的影响以及MAPK信号通路在淫羊藿甙促成骨细胞核心结合因子α1(core binding factor-1,Cbfa1)蛋白表达中的作用,以探讨淫羊藿甙对成骨细胞作用的信号传导机制.方法 用酶消化法分离24 h内新生SD大鼠颅盖骨成骨细胞,进行原代培养.在培养液中加入淫羊藿甙(10 ng/mL),作用于成骨细胞5 min、10 min、30 min、60 min,抽提总蛋白,用Western blot法检测细胞中ERK、p-ERK、P38和p-P38蛋白的表达.用雌二醇(10-8 mol/L)作用于成骨细胞5 min、10 min、30 min,同上法检测细胞中ERK、p-ERK、P38和p-P38蛋白的表达.用淫羊藿甙(10 ng/mL)、雌二醇(10-8 mol/L)和u0126或SB203580单独或共同干预成骨细胞24 h,抽提核蛋白,用Western blot法检测Cbfa1蛋白的表达.结果 ①淫羊藿甙作用于成骨细胞,30 min时可促进ERK蛋白的磷酸化,并可持续至60 min,和空白组比较,差异有显著性(P<0.05);淫羊藿甙作用于成骨细胞,5 min时即可促进P38蛋白的磷酸化,在30 min时达高峰,并可持续至60 min,和空白组比较,差异有显著性(P<0.05).②雌二醇作用于成骨细胞,在30 min时可促进ERK蛋白的磷酸化,和空白组比较,差异有显著性(P<0.05);雌二醇作用于成骨细胞,在10 min时可促进P38蛋白的磷酸化,并可持续至30 min,和空白组比较,差异有显著性(P<0.05).③淫羊藿甙和雌二醇均能促进成骨细胞中Cbfa1蛋白的表达,和空白组相比,差异有显著性(P<0.05).u0126和SB203580可以抑制淫羊藿甙和雌二醇促进Cbfa1蛋白表达的作用.结论 ①淫羊藿甙和雌二醇均可以激活成骨细胞中ERK/MAPK和P38/MAPK信号通路;②淫羊藿甙和雌二醇均能促进成骨细胞中核转录因子Cbfa1的表达,并且ERK/MAPK和P38/MAPK信号通路参与了此过程.
目的 觀察淫羊藿甙對體外培養大鼠成骨細胞絲裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)信號通路的影響以及MAPK信號通路在淫羊藿甙促成骨細胞覈心結閤因子α1(core binding factor-1,Cbfa1)蛋白錶達中的作用,以探討淫羊藿甙對成骨細胞作用的信號傳導機製.方法 用酶消化法分離24 h內新生SD大鼠顱蓋骨成骨細胞,進行原代培養.在培養液中加入淫羊藿甙(10 ng/mL),作用于成骨細胞5 min、10 min、30 min、60 min,抽提總蛋白,用Western blot法檢測細胞中ERK、p-ERK、P38和p-P38蛋白的錶達.用雌二醇(10-8 mol/L)作用于成骨細胞5 min、10 min、30 min,同上法檢測細胞中ERK、p-ERK、P38和p-P38蛋白的錶達.用淫羊藿甙(10 ng/mL)、雌二醇(10-8 mol/L)和u0126或SB203580單獨或共同榦預成骨細胞24 h,抽提覈蛋白,用Western blot法檢測Cbfa1蛋白的錶達.結果 ①淫羊藿甙作用于成骨細胞,30 min時可促進ERK蛋白的燐痠化,併可持續至60 min,和空白組比較,差異有顯著性(P<0.05);淫羊藿甙作用于成骨細胞,5 min時即可促進P38蛋白的燐痠化,在30 min時達高峰,併可持續至60 min,和空白組比較,差異有顯著性(P<0.05).②雌二醇作用于成骨細胞,在30 min時可促進ERK蛋白的燐痠化,和空白組比較,差異有顯著性(P<0.05);雌二醇作用于成骨細胞,在10 min時可促進P38蛋白的燐痠化,併可持續至30 min,和空白組比較,差異有顯著性(P<0.05).③淫羊藿甙和雌二醇均能促進成骨細胞中Cbfa1蛋白的錶達,和空白組相比,差異有顯著性(P<0.05).u0126和SB203580可以抑製淫羊藿甙和雌二醇促進Cbfa1蛋白錶達的作用.結論 ①淫羊藿甙和雌二醇均可以激活成骨細胞中ERK/MAPK和P38/MAPK信號通路;②淫羊藿甙和雌二醇均能促進成骨細胞中覈轉錄因子Cbfa1的錶達,併且ERK/MAPK和P38/MAPK信號通路參與瞭此過程.
목적 관찰음양곽대대체외배양대서성골세포사렬원활화단백격매(mitogen-activated protein kinase,MAPK)신호통로적영향이급MAPK신호통로재음양곽대촉성골세포핵심결합인자α1(core binding factor-1,Cbfa1)단백표체중적작용,이탐토음양곽대대성골세포작용적신호전도궤제.방법 용매소화법분리24 h내신생SD대서로개골성골세포,진행원대배양.재배양액중가입음양곽대(10 ng/mL),작용우성골세포5 min、10 min、30 min、60 min,추제총단백,용Western blot법검측세포중ERK、p-ERK、P38화p-P38단백적표체.용자이순(10-8 mol/L)작용우성골세포5 min、10 min、30 min,동상법검측세포중ERK、p-ERK、P38화p-P38단백적표체.용음양곽대(10 ng/mL)、자이순(10-8 mol/L)화u0126혹SB203580단독혹공동간예성골세포24 h,추제핵단백,용Western blot법검측Cbfa1단백적표체.결과 ①음양곽대작용우성골세포,30 min시가촉진ERK단백적린산화,병가지속지60 min,화공백조비교,차이유현저성(P<0.05);음양곽대작용우성골세포,5 min시즉가촉진P38단백적린산화,재30 min시체고봉,병가지속지60 min,화공백조비교,차이유현저성(P<0.05).②자이순작용우성골세포,재30 min시가촉진ERK단백적린산화,화공백조비교,차이유현저성(P<0.05);자이순작용우성골세포,재10 min시가촉진P38단백적린산화,병가지속지30 min,화공백조비교,차이유현저성(P<0.05).③음양곽대화자이순균능촉진성골세포중Cbfa1단백적표체,화공백조상비,차이유현저성(P<0.05).u0126화SB203580가이억제음양곽대화자이순촉진Cbfa1단백표체적작용.결론 ①음양곽대화자이순균가이격활성골세포중ERK/MAPK화P38/MAPK신호통로;②음양곽대화자이순균능촉진성골세포중핵전록인자Cbfa1적표체,병차ERK/MAPK화P38/MAPK신호통로삼여료차과정.
Objective To investigate the effect of icariin (ICA) on mitogen-activated protein kinase (MAPK) signal pathway in rat osteoblasts cultured in vitro and the role of MAPK signal pathway in the icariin-promoting expression of core binding factor-1 (Cbfa1) in osteoblasts, and to elucidate the signal mechanism of icariin on osteoblasts. Methods Calvarial osteoblasts were obtained from newborn (<24 h) SD rats by trypsin-collagenase digestion method. After 5 min, 10 min, 30 min, 60 min of treatment with icariin (10 ng/mL) or estrodial (E2) (10-8 mol/L), total protein was isolated from osteoblasts and proteins of ERK, p-ERK, P38 and p-P38 were detected by western-blot analysis. Then calvarial osteoblasts were cultured in the medium containing icariin (10 ng/mL), estrodial (10-8 mol/L) with or without u0126, SB203580 for 24 h respectively, nucleus protein was isolated from osteoblasts and protein of Cbfa1 was detected by western-blot analysis. Results 1.The protein of p-ERK in calvarial osteoblasts increased at 30 min and lasted for 60 min (P<0.05, contrast to the blank group) when treating osteoblasts with icariin(10 ng/mL); the protein of p-P38 in calvarial osteoblasts increased at 5 min and was at the peak at 30 min, and lasted for 60 min (P<0.05, contrast to the blank group) when treating osteoblasts with icariin(10 ng/mL). 2.The protein of p-ERK in calvarial osteoblasts increased at 30 min (P<0.05, contrast to the blank group) when treating osteoblasts with estrodial (10-8 mol/L); the protein of p-P38 in calvarial osteoblasts increased at 10 min and continuously increased to 30 min(P<0.05, contrast to the blank group) when treating osteoblasts with estrodial (10-8 mol/L). 3. Both icariin and estrodial could promote the expression of Cbfa1 protein (P<0.05, contrast to the blank group) and this effect could be weakened by SB203580 or u0126. Conclusion Icariin and estrodial can activate ERK/MAPK and P38/MAPK signal pathways and promote the expression of Cbfa1 protein in osteoblasts respectively. ERK/MAPK and P38/MAPK signal pathway are involved in the processes of icariin-promoting and estrodial-promoting Cbfa1 expression.
