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《Journal of Clinical Rehabilitative Tissue Engineering Research》 2009-50
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Caspase 3 siRNA inhibits chondrocytes apoptosis

Ye Chun-ting1,2,Huang Yao-xiong1,Yang Xiao-hong2 1Department of Biomedical Engineering,Jinan University,Guangzhou 510632,Guangdong Province,China;2Institute of Traumatic Surgery,Guangzhou Red Cross Hospital,Medical College of Jinan University,Guangzhou 510220,Guangdong Province,China  
BACKGROUND:Up to date,studies concerning capspase 3 inhibitor mainly focus on peptide/non-peptide compounds synthesis and detection.Few reports addressing inhibits chondrocytes apoptosis using silenced caspase 3 gene.OBJECTIVE:To inhibit apoptosis of chondrocytes by blocking the apoptotic cascade reaction,gene silencing of caspase 3,and transduction of caspase 3 siRNA into chondrocytes with lentivirus vector.DESIGN,TIME AND SETTING:A single sample observation was performed at the Institute of Traumatic Surgery,Guangzhou Red Cross Hospital,Medical College of Jinan University from June 2008 to June 2009.MATERIALS:Chondrocytes were harvested from SD rats,and caspase 3 shRNA plasimid was constructed by our laboratory.METHODS:Rattus caspase 3 siRNA was synthesized and cloned into pSIH1-H1-copGFP plasmid.pSIH1-H1-copGFP-caspase 3 siRNA lentivirus was generated in 293TN cells by pPACKH1? Lentivector Packaging Kit and transducted into chondrocytes of rats.MAIN OUTCOME MEASURES:After the lentivirus was transducted into chondrocytes,the caspase 3 mRNA was tested by RT-PCR and the caspase 3 protein was tested by Western blot.Both the transducted cells and untransducted cells were induced apoptosis by tumor necrosis factor α (TNF-α).Cell apoptosis was assessed by flow cytometry,Annevin V/PI.RESULTS:The transduction rate of caspase 3 siRNA was about 90% by lentivirus vector.The expression of caspase 3 mRNA and caspase 3 protein in transducted chondrocytes was lower than the normal chondrocytes (P 0.01).When the cells induced apoptosis by TNF-α,the apoptosis rate of the negative siRNA-chondrocytes was 7 times higher than that of caspase 3 siRNA-chondrocytes.CONCLUSION:The caspase 3 siRNA could inhibit caspase 3 expression and decrease drug-induced apoptosis of the chondrocytes.
【Fund】: 广州市医药卫生重点项目资助(2006-ZDi-04)~~
【CateGory Index】: R329
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