[骨碎补总黄酮对高糖处理后体外成骨细胞分化及ERK1/2和p38丝裂原活化蛋白激酶表达的影响]

[Effects of total flavonoids in Drynaria fortunei on osteoblasts differentiation and the expression of ERK1/2 and p38 MAPK after treatment by high glucose in vitro].

作者信息

Zhu Xiao-Feng, Wang Ting-Chun, Zhang Rong-Hua, Sun Sheng-Yun, Wang Pan-Pan, Yang Li, Han Li, Jin Ling

机构信息

Traditional Chinese Medicine Department, the First Affiliated Hospital of Jinan University, Guangzhou 510630, China.

出版信息

Zhong Yao Cai. 2012 Mar;35(3):424-9.

PMID:22876683

Abstract

OBJECTIVE

To observe the effects of Total Flavonoids in Drynaria fortunei (TFDF) on osteoblasts differentiation activity after treatment by high glucose and observe the effects on p38MAPK and ERK1/2 signaling protein in osteoblasts.

METHODS

Primary osteoblasts of newborn SD rats was extracted and cultured and its biological characteristics was observed. MTT method was used to observe osteoblasts' cytotoxicity,and to choose a suitable concentration of TFDF in the culture medium. pNPP,ELISA,Alizarin dyeing were used to test ALP,Type I collagen,osteocalcin and mineralization of osteoblasts after treatment by different concentration of glucose respectively and after treatment by TFDF and high glucose. Western-blot was used to detect p38MAPK and ERK1/2 protein phosphorylation after treatment by TFDF and high glucose.

RESULTS

Primary osteoblasts of newborn SD rats could be used well in this experiment. According to the toxicity of TFDF on OB, 25, 50, 100 mg/L of TFDF were selected for the experimental concentration gradient. ALP, Type I collagen,osteocalcin and mineralization of osteoblasts after treatment with glucose (25, 50 mmol/L) were less than those of control group respectively. TFDF could increase ALP, Type I collagen, osteocalcin activity and mineralization of osteoblasts in a dose-dependent manner after treatment by high glucose (25 mmol/L). TFDF(50 mg/L) could increase protein phosphorylation of p38MAPK and ERK1/2 of osteoblasts after treatment by high glucose (25 mmol/L).

CONCLUSION

High glucose can decrease differentiation and mineralization of osteoblasts. TFDF can increase differentiation and mineralization of osteoblasts in a dose-dependent manner after treatment by high glucose. The role of TFDF in the promotion of osteoblasts differentiation is related to protein phosphorylation of p38MAPK and ERK1/2.

摘要

目的

观察骨碎补总黄酮（TFDF）对高糖处理后成骨细胞分化活性的影响，并观察其对成骨细胞中p38丝裂原活化蛋白激酶（p38MAPK）和细胞外信号调节激酶1/2（ERK1/2）信号蛋白的影响。

方法

提取并培养新生SD大鼠原代成骨细胞，观察其生物学特性。采用MTT法观察成骨细胞的细胞毒性，以选择培养基中合适浓度的TFDF。分别用对硝基苯磷酸酯（pNPP）法、酶联免疫吸附测定（ELISA）法、茜素染色法检测不同浓度葡萄糖处理后以及TFDF与高糖共同处理后成骨细胞的碱性磷酸酶（ALP）、Ⅰ型胶原、骨钙素及矿化情况。采用蛋白质免疫印迹法（Western-blot）检测TFDF与高糖共同处理后成骨细胞中p38MAPK和ERK1/2蛋白的磷酸化水平。

结果

新生SD大鼠原代成骨细胞可很好地用于本实验。根据TFDF对成骨细胞的毒性，选择25、50、100 mg/L作为实验浓度梯度。葡萄糖（25、50 mmol/L）处理后成骨细胞的ALP、Ⅰ型胶原、骨钙素及矿化水平分别低于对照组。高糖（25 mmol/L）处理后，TFDF可呈剂量依赖性增加成骨细胞的ALP、Ⅰ型胶原、骨钙素活性及矿化水平。高糖（25 mmol/L）处理后，TFDF（50 mg/L）可增加成骨细胞中p38MAPK和ERK1/2的蛋白磷酸化水平。