基于瞬时受体电位香草素亚家族4信号通路探讨牛蒡子苷元对体外培养软骨细胞增殖及Ⅱ型胶原蛋白和软骨蛋白聚糖表达的影响  

An experimental study of effect of arctigenin on proliferation of chondrocyte cultured in vitro and expression of type Ⅱ collagen protein and aggrecan based on the transient receptor potential vanilloid 4 signaling pathway

作　　者：陈世宣 刘益杰[1] 胡笑燊 张增乔 李安琪[1] 厉坤鹏 冯伟[2] 

机构地区：[1]上海中医药大学,上海201203 [2]上海市第七人民医院,上海201137

出　　处：《中医正骨》2017年第10期13-19,共7页The Journal of Traditional Chinese Orthopedics and Traumatology

基　　金：上海市自然科学基金资助项目(15ZR1441000)

摘　　要：目的:基于瞬时受体电位香草素亚家族4(transient receptor potential vanilloid 4,TRPV4)信号通路探讨牛蒡子苷元对体外培养软骨细胞增殖及Ⅱ型胶原蛋白和软骨蛋白聚糖表达的影响。方法:自新生SD大鼠膝关节软骨组织提取软骨细胞,进行体外培养,取第1代软骨细胞分别加入不同培养液进行干预。空白组仅加入常规培养液,牛蒡子苷元组加入含10μmol牛蒡子苷元的培养液,牛蒡子苷元联合阻断剂组加入含10μmol牛蒡子苷元和10μmol GSK205的培养液,激动剂组加入含1μmol 4α-PPD的培养液,阻断剂联合激动剂组加入含10μmol GSK205和1μmol 4α-PPD的培养液,阻断剂组加入含10μmol GSK205的培养液。分别于干预24 h和72 h后以CCK-8法检测软骨细胞增殖情况,48 h后以Western blot法检测软骨蛋白聚糖和Ⅱ型胶原蛋白水平。结果:(1)软骨细胞增殖测定结果。干预24 h后,6组的吸光度比较,差异有统计学意义(1.03±0.72,1.32±0.11,1.01±0.05,1.33±0.34,1.04±0.50,1.10±0.06;F=18.309,P=0.000)。牛蒡子苷元组、激动剂组的吸光度均高于空白组(P=0.000,P=0.000);牛蒡子苷元联合阻断剂组、阻断剂联合激动剂组、阻断剂组的吸光度与空白组比较,组间差异均无统计学意义(P=0.632,P=0.840,P=0.164);牛蒡子苷元组的吸光度高于牛蒡子苷元联合阻断剂组、阻断剂联合激动剂组(P=0.000,P=0.000);牛蒡子苷元组与激动剂组比较,差异无统计学意义(P=0.834);牛蒡子苷元联合阻断剂组的吸光度低于激动剂组(P=0.000),与阻断剂联合激动剂组比较,差异无统计学意义(P=0.498);激动剂组的吸光度高于阻断剂联合激动剂组(P=0.000)。干预72 h后,6组的吸光度比较,差异有统计学意义(1.66±0.02,2.21±0.05,1.84±0.04,1.92±0.07,1.71±0.10,1.74±0.08;F=37.629,P=0.000)。牛蒡子苷元组、牛蒡子苷元联合阻断剂组、激动剂组的吸光度均高于空白组(P=0.000,P=0.001,P=0.000);阻断剂联合激动剂组、阻断剂组Objective: To explore the effect of arctigenin on proliferation of chondrocyte cultured in vitro and expression of typeⅡcollagen protein and aggrecan based on the transient receptor potential vanilloid 4( TRPV4) signaling pathway. Methods: The chondrocytes were extracted from the knee cartilage tissues of newborn SD rats and were cultured in vitro,then the first-generation chondrocytes in blank group,arctigenin group,arctigenin-blocker group,agonist group,blocker-agonist group and blocker group were cultured in conventional culture medium,arctigenin( 10 μmol) culture medium,arctigenin( 10 μmol)-GSK205( 10 μmol) culture medium,4α-PPD( 1 μmol) culture medium,GSK205( 10 μmol)-4α-PPD( 1 μmol) culture medium and GSK205( 10 μmol) culture medium respectively. The chondrocyte proliferation was detected by using CCK-8 method after 24-and 72-hour intervention respectively,and the expression of aggrecan and typeⅡcollagen protein were detected by using Western blot method after 48-hour intervention. Results: After 24-hour intervention,there was statistical difference in the absorbance between the 6 groups( 1. 03 +/-0. 72,1. 32 +/-0. 11,1. 01 +/-0. 05,1. 33 +/-0. 34,1. 04 +/-0. 50,1. 10 +/-0. 06; F = 18. 309,P = 0. 000). The absorbance was higher in arctigenin group and agonist group compared to blank group( P = 0. 000,P = 0. 000). There was no statistical difference in the absorbance between arctigenin-blocker group and blank group and between blocker-agonist group and blank group and between blocker group and blank group( P = 0. 632,P = 0. 840,P =0. 164). The absorbance was higher in arctigenin group compared to arctigenin-blocker group and blocker-agonist group( P = 0. 000,P =0. 000). There was no statistical difference in the absorbance between arctigenin group and agonist group( P = 0. 834). The absorbance was lower in arctigenin-blocker group compared to agonist group( P = 0. 000). There was no statistical difference in the absorbance between arctigenin-blocker group and blocker-agonist group( P = 0. 498).

关 键 词：骨关节炎 牛蒡子甙 软骨细胞 瞬时受体电位香草素亚家族4 细胞增殖 胶原Ⅱ型 软骨蛋白聚糖类 

分 类 号：R285[医药卫生—中药学]