目的:观察独活寄生汤血清对膝骨性关节炎退变软骨细胞Aggrecan和Collagen X mRNA表达的影响。方法:选择6例患者标本均来源于云南省中医医院骨科膝骨性关节炎住院患者,行手术截取的新鲜退变软骨组织,采用组织块法培养原代细胞;MTT法检测...目的:观察独活寄生汤血清对膝骨性关节炎退变软骨细胞Aggrecan和Collagen X mRNA表达的影响。方法:选择6例患者标本均来源于云南省中医医院骨科膝骨性关节炎住院患者,行手术截取的新鲜退变软骨组织,采用组织块法培养原代细胞;MTT法检测传代第一代细胞生长情况,并绘制生长曲线;制备独活寄生汤大鼠含药血清;均取传代第一代的退变软骨细胞,配制5%、10%、20%独活寄生汤含药血清及相应体积比浓度的生理盐水血清作为对照;MTT法检测不同浓度的独活寄生汤血清对退变软骨细胞增殖的影响;实时荧光定量PCR检测不同给药时间点各组退变软骨细胞Aggrecan和Collagen Ⅹ mRNA的表达。结果:退变软骨细胞传代第一代生长曲线见传代第8天时细胞增殖达到顶峰,故选择细胞传代第8天为给药干预时间点;与含同体积比的生理盐水血清组比较,不同浓度的独活寄生汤血清可对退变软骨细胞增殖产生促进作用(P<0.01),尤以10%独活寄生汤血清更为明显(P<0.05);与含同体积比生理盐水血清组比较,不同时间点的5%、10%、20%独活寄生汤血清干预后的退变软骨细胞Aggrecan mRNA的表达均上调(P<0.01),Collagen Ⅹ mRNA的表达均下调(P<0.01),且以10%独活寄生汤血清组的作用最为明显(P<0.05)。结论:独活寄生汤可能通过调节退变软骨组织的胶原及其蛋白多糖的表达而产生延缓膝骨性关节炎退变软骨组织的部分作用。展开更多
背景:软骨细胞退变常导致骨关节炎、椎间盘退变、半月板退变等骨科常见疾病,沉默信息调节因子1(silent information regulator 1,SIRT1)基因与软骨细胞退变关系密切,当敲减ATDC5小鼠软骨细胞系中SIRT1基因后,可通过软骨细胞退变相关信...背景:软骨细胞退变常导致骨关节炎、椎间盘退变、半月板退变等骨科常见疾病,沉默信息调节因子1(silent information regulator 1,SIRT1)基因与软骨细胞退变关系密切,当敲减ATDC5小鼠软骨细胞系中SIRT1基因后,可通过软骨细胞退变相关信号通路的作用影响细胞功能,进而影响骨科疾病的进程。目的:探讨SIRT1基因敲减后ATDC5小鼠软骨细胞中显著激活的软骨细胞退变相关信号通路情况及差异因子的表达情况。方法:携载或不携载SIRT1基因的慢病毒转染ATDC5细胞后,分为对照组(转染携载阴性对照慢病毒ATDC5细胞组)和实验组(转染携载SIRT1基因慢病毒ATDC5细胞组)。提取不同组别细胞中总RNA并进行质检,基因芯片检测细胞中编码RNA核酸的表达情况,结合生物信息学分析,寻找显著激活的软骨细胞退变相关信号通路,并对其中部分信号通路的差异变化因子进行阐述。结果与结论:①发现SIRT1基因敲减后ATDC5细胞中被显著激活的信号通路有42条;②选择了其中被显著激活的5条与软骨细胞退变相关且研究较少的信号通路:肝细胞生长因子信号通路(ETS1、PIK3CA、NRAS、PIK3C2A、FGFR2、ELF1、FGFR3、CDKN1A、AKT3、FRS2、PRKD3、MAP3K2)(Ratio=0.1390,Z-score=2.673)、神经调节因子信号通路(RPS6KB1、STAT5A、MTOR、BTC、HBEGF、RNF41)(Ratio=0.1360,Z-score=2.309)、胰岛素受体信号通路(TSC1、EIF4EBP1、PRKAR2B、PPP1R12A、TSC2)(Ratio=0.1060,Z-score=2.138)、趋化因子受体4信号通路(PAK4、EGR1、RHOJ、RHOB、PLCB1、GNG5)(Ratio=0.0970,Z-score=2.500)、肌动蛋白细胞骨架信号通路(ABI2、BRK1、PIP4K2B、MYLK、IQGAP1、ARPC1A、ACTA2、EZR、SSH2、ACTG1、IQGAP3)(Ratio=0.0921,Z-score=2.236)进行汇报,从而为研究软骨细胞退变相关疾病提供新颖靶点。展开更多
The inflammatory cytokine interleukin-1 beta （IL-1β） plays a key role in the process of intervertebral disc degenera- tion （IVDD）. In the present study, we aimed to evaluate the effect of pharmaco-serum of ＂Taor...The inflammatory cytokine interleukin-1 beta （IL-1β） plays a key role in the process of intervertebral disc degenera- tion （IVDD）. In the present study, we aimed to evaluate the effect of pharmaco-serum of ＂Taoren-Honghua-herb pair＂ on IL-1β- induced chondrocyte degeneration in vitro. Taoren （Semen persicae） and Honghua （Safflower carthamus） were administered to the rats, and the pharmaco-serum was collected and prepared. Chondrocytes of the third passage, isolated from the rat＇s vertebral endplates, were treated by standard medium only （Group NC）, IL-1β （Group IL） or combination of IL-1β and pharmaco-serum （Group TRHH）. Cell proliferation and apoptosis were determined, and the expression of aggrecan, Col2ul, Coll0ul, IL-6 and SOX9 at the mRNA level in chondrocytes was quantified by real-time PCR. Immunohistochemistry staining of type II and X collagen and Safranine O staining were also used to evaluate the chondrocytes. Compared with the Group NC, IL-1β treatment inhibited the cell proliferation and induced the cell apoptosis （P〈0.05）, and the expression of aggrecan, Col2αl and SOX9 at the mRNA level was down-regulated. In contrast, the expression of Coll0ul and IL-6 was up-regulated after IL-1β treatment （P〈0.05）. Meanwhile, the immune-staining of type II collagen and Safranine O staining were decreased, while the staining of type X collagen was increased. Compared with the Group IL, cell proliferation was increased, and apoptosis of chondrocytes was decreased when cells were treated with the pharmaco-semm of TRHH-herb pair （P〈0.05）. The expression of aggrecan, Col2cd and SOX9 at the mRNA level was up-regulated, while that of Coll0cd and IL-6 was down-regulated （P〈0.05）. Saffanine O staining also showed increased positive staining （P〈0.05）. Taken together, the treatment of pharmaco-serum of TRHH-herb pair could prevent endplate chondrocyte degeneration induced by IL-1β.展开更多
文摘背景:软骨细胞退变常导致骨关节炎、椎间盘退变、半月板退变等骨科常见疾病,沉默信息调节因子1(silent information regulator 1,SIRT1)基因与软骨细胞退变关系密切,当敲减ATDC5小鼠软骨细胞系中SIRT1基因后,可通过软骨细胞退变相关信号通路的作用影响细胞功能,进而影响骨科疾病的进程。目的:探讨SIRT1基因敲减后ATDC5小鼠软骨细胞中显著激活的软骨细胞退变相关信号通路情况及差异因子的表达情况。方法:携载或不携载SIRT1基因的慢病毒转染ATDC5细胞后,分为对照组(转染携载阴性对照慢病毒ATDC5细胞组)和实验组(转染携载SIRT1基因慢病毒ATDC5细胞组)。提取不同组别细胞中总RNA并进行质检,基因芯片检测细胞中编码RNA核酸的表达情况,结合生物信息学分析,寻找显著激活的软骨细胞退变相关信号通路,并对其中部分信号通路的差异变化因子进行阐述。结果与结论:①发现SIRT1基因敲减后ATDC5细胞中被显著激活的信号通路有42条;②选择了其中被显著激活的5条与软骨细胞退变相关且研究较少的信号通路:肝细胞生长因子信号通路(ETS1、PIK3CA、NRAS、PIK3C2A、FGFR2、ELF1、FGFR3、CDKN1A、AKT3、FRS2、PRKD3、MAP3K2)(Ratio=0.1390,Z-score=2.673)、神经调节因子信号通路(RPS6KB1、STAT5A、MTOR、BTC、HBEGF、RNF41)(Ratio=0.1360,Z-score=2.309)、胰岛素受体信号通路(TSC1、EIF4EBP1、PRKAR2B、PPP1R12A、TSC2)(Ratio=0.1060,Z-score=2.138)、趋化因子受体4信号通路(PAK4、EGR1、RHOJ、RHOB、PLCB1、GNG5)(Ratio=0.0970,Z-score=2.500)、肌动蛋白细胞骨架信号通路(ABI2、BRK1、PIP4K2B、MYLK、IQGAP1、ARPC1A、ACTA2、EZR、SSH2、ACTG1、IQGAP3)(Ratio=0.0921,Z-score=2.236)进行汇报,从而为研究软骨细胞退变相关疾病提供新颖靶点。
基金National Natural Science Foundation of China(Grant No.81273777,81102606,81072831)the National Basic Re search Program of China(Grant No.2010CB530400)the Program for Changjiang Scholars and Innovative Research Team in University(PCSIRT,IRT1270)
文摘The inflammatory cytokine interleukin-1 beta （IL-1β） plays a key role in the process of intervertebral disc degenera- tion （IVDD）. In the present study, we aimed to evaluate the effect of pharmaco-serum of ＂Taoren-Honghua-herb pair＂ on IL-1β- induced chondrocyte degeneration in vitro. Taoren （Semen persicae） and Honghua （Safflower carthamus） were administered to the rats, and the pharmaco-serum was collected and prepared. Chondrocytes of the third passage, isolated from the rat＇s vertebral endplates, were treated by standard medium only （Group NC）, IL-1β （Group IL） or combination of IL-1β and pharmaco-serum （Group TRHH）. Cell proliferation and apoptosis were determined, and the expression of aggrecan, Col2ul, Coll0ul, IL-6 and SOX9 at the mRNA level in chondrocytes was quantified by real-time PCR. Immunohistochemistry staining of type II and X collagen and Safranine O staining were also used to evaluate the chondrocytes. Compared with the Group NC, IL-1β treatment inhibited the cell proliferation and induced the cell apoptosis （P〈0.05）, and the expression of aggrecan, Col2αl and SOX9 at the mRNA level was down-regulated. In contrast, the expression of Coll0ul and IL-6 was up-regulated after IL-1β treatment （P〈0.05）. Meanwhile, the immune-staining of type II collagen and Safranine O staining were decreased, while the staining of type X collagen was increased. Compared with the Group IL, cell proliferation was increased, and apoptosis of chondrocytes was decreased when cells were treated with the pharmaco-semm of TRHH-herb pair （P〈0.05）. The expression of aggrecan, Col2cd and SOX9 at the mRNA level was up-regulated, while that of Coll0cd and IL-6 was down-regulated （P〈0.05）. Saffanine O staining also showed increased positive staining （P〈0.05）. Taken together, the treatment of pharmaco-serum of TRHH-herb pair could prevent endplate chondrocyte degeneration induced by IL-1β.
