CAJ | 학술논문

目的探讨壳聚糖-pCrmA纳米粒对白细胞介素(IL)-1β诱导的软骨细胞凋亡的影响.方法制备壳聚糖-pDNA纳米粒并进行表征.荧光显微镜观察壳聚糖介导pIRES2-增强型绿色荧光蛋白(EGFP)在原代软骨细胞中的转染,采用四甲基偶氮唑蓝(MTT)实验分析壳聚糖-pIRES2-EGFP纳米粒的细胞毒性,蛋白印迹法分析壳聚糖介导的CrmA在软骨细胞中的表达,用原位末端脱氧核苷酸转移酶标记(TUNEL)法检测壳聚糖-pCrmA纳米粒对IL-1β诱导的软骨细胞凋亡的影响.采用单因素方差分析进行统计学分析.结果壳聚糖-pDNA纳米粒的粒径为50 nm,壳聚糖-pDNA纳米粒中的pDNA在pH 2.0和pH 7.4缓冲液中呈双相释放.壳聚糖能够介导基因CrmA在软骨细胞中表达.壳聚糖-pDNA纳米粒无细胞毒性.壳聚糖-pcrmA纳米粒处理组软骨细胞凋亡率明显低于壳聚糖组(P＜0.05)与磷酸盐缓冲液组(P＜0.01).结论壳聚糖是一种有效的非病毒基因载体,壳聚糖-pCrmA纳米粒能够显著抑制IL-1β诱导的软骨细胞凋亡.
목적 탐토각취당-pCrmA납미립대백세포개소(IL)-1β유도적연골세포조망적영향.방법 제비각취당-pDNA납미립병진행표정.형광현미경관찰각취당개도pIRES2-증강형록색형광단백(EGFP)재원대연골세포중적전염,채용사갑기우담서람(MTT)실험분석각취당-pIRES2-EGFP납미립적세포독성,단백인적법분석각취당개도적CrmA재연골세포중적표체,용원위말단탈양핵감산전이매표기(TUNEL)법검측각취당-pCrmA납미립대IL-1β유도적연골세포조망적영향.채용단인소방차분석진행통계학분석.결과 각취당-pDNA납미립적립경위50 nm,각취당-pDNA납미립중적pDNA재pH 2.0화pH 7.4완충액중정쌍상석방.각취당능구개도기인CrmA재연골세포중표체.각취당-pDNA납미립무세포독성.각취당-pcrmA납미립처리조연골세포조망솔명현저우각취당조(P＜0.05)여린산염완충액조(P＜0.01).결론 각취당시일충유효적비병독기인재체,각취당-pCrmA납미립능구현저억제IL-1β유도적연골세포조망.
Objective To study the effect of chitosan-pCrmA nanoparticles on the apoptosis of chondrocytes induced by interleukin-1 beta (IL-1β).Methods Chitosan-pDNA nanoparticles were prepared and characterized.The transfection efficiency of chitosan-mediated pIRES2-EGFP was evaluated using fluorescence microscope.The cytotoxicity of chitosan-pIRES2-EGFP nanoparticles in primary rabbit chondrocytes was analyzed by MTT assay.The expression of chitosan-mediated pCrmA in primary rabbit chondrocytes was verified by Western blotting.The effect of chitosan-mediated CrmA on chondrocytes apoptosis induced by IL-1β were analyzed by TUNEL assay.One-way ANOVA was used to analysis.Results The size of chitosan-pDNA nanoparticles was 50 nm.The pDNA release of chitosan-pDNA nanoparticles appeared as biphasic release at pH 2.0 and pH 7.4 buffer.The expression of CrmA in rabbit primary chondrocytes mediated by chitosan could be detected.The chitosan-pIRES2-EGFP nanoparticles had no cytotoxicity.The apoptosis rate of chondrocytes in the chitosan-pCrmA nanoparticles treated group was significantly lower than that of the chitosan treated group (P＜0.05) and PBS group (P＜0.01).Conclsion Chitosan is an effective non-viral gene transfer vector.The CrmA mediated by chitosan can significantly inhibit chondrocytes apoptosis induced by IL-1β,suggesting that chitosan-pCrmA nanoparticles may be the treatment of osteoarthrifis.