Experimental study of apoptosis induced by rhFasL gene transferred into primary cultured synoviocytes from patients with rheumtoid arthritis / 中华风湿病学杂志

ABSTRACT

Objective To observe apoptosis of primary cultured synoviocytes from patients with rheumtoid arthritis (RA) which induced by transfected recombinant plasmid carring Fas ligand (FasL) gene, aimed to detect the aim targets and develope gene therapy for RA by intra-articular injection. Methods FasL cDNA was introducted into the plasmid pcDNA3.1-neo by reverse transcriptase-polymerse chain reaction (RT-PCR) to set up the recombinated human FasL gene expression carrier, Then the pcDNA3.1-FasL was transducted into the primary cultured synovial cells by catholyte liposomes. After screened by G418, transgenic synovial cells were choosen. Finally through the microscope, TUNEL, Annexin Ⅴ-FITC and PI staining with flow cytometry, we observed the apoptosis state of each synovial cell. Results Prokaryotic and eukaryon carrier with FasL gene were established successfilly. The gene sequencing had completely coincident with the lierature. When eukaryon carrier with FasL (pcDNA3.1-FasL) was transducted into the index number livinging RA synovial cells after 15 h, most synovial cells presented distortion transformation, and the minority sheded from the bottom wall. The transgenic RA synovial cells after screened by G418, only a little amount survived about 1~2 per sight, and the cells grew slowly, the normal matched cells were cloned after 2~4 weeks. The FasL gene transgenic RA synovial cells presented positive findings by TUNEL staining. Early apoptosis cells by Annexin Ⅴ/PI staining, but the difference wasn′t significant. Conclusion The eukaryon carrier with FasL (pcDNA3.1-FasL) can be transferred into RA synovial cells. It can induce synovial cell apoptosis.