丹参酮ⅡA对软骨细胞Ⅱ型胶原及Wnt/β-catenin信号通路的影响
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摘要
目的:探讨不同浓度丹参酮ⅡA对软骨细胞Ⅱ型胶原及Wnt/β-catenin信号通路的影响。方法:提取新生SD大鼠原代软骨细胞,并通过Ⅱ型胶原蛋白免疫荧光进行鉴定,取P1代细胞进行实验,共设5组,其中1组设为空白对照组,另外4组分别用6.25μmol/L,12.5μmol/L,25μmol/L及50μmol/L浓度丹参酮ⅡA对其进行干预,24h后通过蛋白印迹分析法(Western Blot)检测软骨细胞Ⅱ型胶原及β-catenin蛋白的表达情况。结果:随着丹参酮ⅡA浓度的增加,Ⅱ型胶原蛋白表达增加,12.5μmol/L,25μmol/L及50μmol/L丹参酮ⅡA组Ⅱ型胶原蛋白量均较对照组增高(P<0.05),且Ⅱ型胶原蛋白量与丹参酮ⅡA浓度呈正相关(r=0.949,P<0.001),各丹参酮ⅡA组β-catenin蛋白量均较对照组降低(P<0.05),且β-catenin蛋白量与丹参酮ⅡA浓度呈负相关(r=-0.949,P<0.001)。结论:丹参酮ⅡA可抑制Wnt/β-catenin信号通路,促进软骨细胞Ⅱ型胶原表达,延缓软骨细胞退变。
Objective:To investigate the effect of Tanshinone ⅡA(TAN ⅡA)on collagen Ⅱ and Wnt/β-catenin signal pathway of rat chondrocytes in vitro.Methods:SD rat articular chondrocytes were isolated,and immunofluorescence was performed to identify the cells.A total of 5 groups were set up,1 of which was a blank control group,and the other 4 groups were treated with different concentrations of TAN ⅡA(6.25μmol/L,12.5μmol/L,25μmol/L and 50μmol/L).The expression ofβ-catenin and collagen Ⅱ were examined by Western-Blot after 24 h.Results:The groups of12.5μmol/L,25μmol/L,50μmol/L concentration of TAN ⅡA expressed higher collagenⅡthan the control group(P<0.05).The expression of collagenⅡ was increased as the concentration of TAN ⅡA had a rising trend(r=0.949,P<0.001).All the groups treated with TAN ⅡA expressed lowerβ-catenin than the control group(P<0.05).The expression ofβ-catenin was decreased as the concentration of TAN ⅡA had a rising trend(r=-0.949,P<0.001).Conclusion:TAN ⅡA can inhibit the Wnt/β-catenin signaling pathway,promote the expression of type Ⅱ collagen in chondrocytes,and delay the degeneration of chondrocytes.
Objective:To investigate the effect of Tanshinone ⅡA(TAN ⅡA)on collagen Ⅱ and Wnt/β-catenin signal pathway of rat chondrocytes in vitro.Methods:SD rat articular chondrocytes were isolated,and immunofluorescence was performed to identify the cells.A total of 5 groups were set up,1 of which was a blank control group,and the other 4 groups were treated with different concentrations of TAN ⅡA(6.25μmol/L,12.5μmol/L,25μmol/L and 50μmol/L).The expression ofβ-catenin and collagen Ⅱ were examined by Western-Blot after 24 h.Results:The groups of12.5μmol/L,25μmol/L,50μmol/L concentration of TAN ⅡA expressed higher collagenⅡthan the control group(P<0.05).The expression of collagenⅡ was increased as the concentration of TAN ⅡA had a rising trend(r=0.949,P<0.001).All the groups treated with TAN ⅡA expressed lowerβ-catenin than the control group(P<0.05).The expression ofβ-catenin was decreased as the concentration of TAN ⅡA had a rising trend(r=-0.949,P<0.001).Conclusion:TAN ⅡA can inhibit the Wnt/β-catenin signaling pathway,promote the expression of type Ⅱ collagen in chondrocytes,and delay the degeneration of chondrocytes.
引文
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[2]徐西林,张晓峰,吕航,等.丹参注射液对骨性关节炎模型兔膝关节软骨细胞p-IκBα表达影响的实验研究[J].中国中医药科技,2017,24(4):438-440.
[3]袁长深,梅其杰,段戡,等.丹参加玻璃酸钠对兔膝关节软骨保护的效应与机制研究[J].时珍国医国药,2012,23(10):2452-2453.
[4]马勇,郭杨,顾一煌,等.丹参关节腔注射与针刺修复膝关节全层软骨缺损[J].中国组织工程研究,2012,16(42):7837-7842.
[5]石印玉,徐荣善,陈友红.养血软坚方治疗膝骨关节炎的临床报告[J].中国中医骨伤科杂志,1994,2(4):33-36.
[6]胡建华.丹参酮ⅡA磺酸钠诱导乳兔BMSCs向软骨细胞分化中的作用及细胞密度对诱导的影响[D].南宁:广西医科大学,2015.
[7]缪明星,黄莉莉,王欣.丹参酮ⅡA在Beagle犬体内的血药浓度测定及其药动学研究[J].中国药房,2015,26(19):2632-2635.
[8]庄园.丹参酮对膝骨性关节炎模型细胞活性及氧自由基代谢的影响[D].广州:暨南大学,2007.
[9]Yates KE,Shortkroff S,Reish RG.Wnt influence on chondrocyte differentiation and cartilage function[J].DNA Cell Biol,2005,24(7):446-457.
[10]Blom AB,Brockbank SM,van Lent PL,et al.Involvement of the Wnt signaling pathway in experimental and human osteoarthritis:prominent role of Wnt-induced signaling protein 1[J].Arthritis Rheum,2009,60(2):501-512.
[11]Velasco J,Zarrabeitia MT,Prieto JR,et al.Wnt pathway genes in osteoporosis and osteoarthritis:differential expression and genetic association study[J].Osteoporos Int,2010,21(1):109-118.
[12]Yuasa T,Otani T,Koike T,et al.Wnt/β-catenin signaling stimulates matrix catabolic genes and activity in articular chondrocytes:its possible role in joint degeneration[J].Laboratory Investigation,2008,88(3):264-274.
[13]Martijn H,van den Bosch,Arjen B,et al.Induction of canonical wnt signaling by synovial overexpression of selected wnts leads to protease activity and early osteoarthritislike cartilage damage[J].The American Journal of Pathology,2015,185(7):1970-1980.
[14]Delgado C,Arozamena I,Casafont,et al.Wnt activity,osteocytes and sclerostin expression in hip fractures and osteoarthritis[J].Bone,2011,48(S5):138-140.
[15]王德刚,许传勇,姜玉祥,等.黄芪甲苷对Wnt/β-catenin信号通路活化的人滑膜及软骨细胞共培养体系相关因子的调控作用[J].广东医学,2017,38(3):354-357.