IL-1beta sensitizes rat intervertebral disc cells to Fas ligand mediated apoptosis in vitro

来源 :Acta Pharmacologica Sinica | 被引量 : 0次 | 上传用户:HanMa_1978
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Aim:To determine the apoptotic effect of recombinant rat Fas Ligand on rat inter-vertebral disc cells pre-treated with IL-1beta in vitro,and the expression of Fas incultured rat intervertebral disc cells.Methods:Cells were isolated from the innerannulus fibrosus and transition zones of lumbar discs from Sprague-Dawley rats.The cells were grown in monolayer and divided in 5 treatment groups.IL-1 beta (10ng/mL),FasL (5,20 ng/mL) with/without IL-1beta (10 ng/mL) pre-treatment wasrespectively added in Dulbecco’s modified Eagle’s medium and Ham’s F-12 me-dium with 1% fetal bovine serum.After 32 h,the cells were stained with annexin V-FITC and propidium iodide to evaluate apoptosis using flow cytometry and toanalysis transcription of Fas using RT-PCR.Results:Compared with controlgroup,FasL (20 ng/mL),IL-1β(10 ng/mL)+FasL (5 ng/mL),and IL-1β(10 ng/mL)+FasL (20 ng/mL) induced significant apoptosis of the disc cells (P<0.01).Apoptosis was also induced by FasL 5 ng/mL (P<0.05);whereas,apoptosis wasnot induced by 1L-1β(10 ng/mL) (P>0.05).IL-1β(10 ng/mL) enhanced the apoptosis-inducing effects of FasL (5 ng/mL) and FasL (20 ng/mL) in disc cells.Fas genetranscription in all groups and Fas expression in the 5 treatment groups wereapproximately 1.2-2.1-fold greater than control group (respectively,P<0.05).Additionally,Fas expression in FasL with IL-1β pre-treatment groups were signifi-cantly up-regulated than in FasL groups (P<0.01).Conclusion:The results of thisstudy showed disc cells pre-treated with IL-lbeta increased apoptotic rate inresponse to FasL in vitro and provided insights to understand Fas/FasL system-mediated apoptosis in disc cells which would be enhanced due to inflammationfactor in degenerative disc. Aim: To determine the apoptotic effect of recombinant rat Fas Ligand on rat inter-vertebral disc cells pre-treated with IL-1 beta in vitro, and the expression of Fas incultured rat intervertebral disc cells. Methods: Cells were isolated from the innnulusulus fibrosus and transition zones of lumbar discs from Sprague-Dawley rats. The cells were grown in monolayer and divided in 5 treatment groups. IL-1 beta (10 ng / mL), FasL (5,20 ng / mL) with / without IL- 10 ng / mL) pre-treatment wasrespectively added in Dulbecco’s modified Eagle’s medium and Ham’s F-12 me-dium with 1% fetal bovine serum. After 32 h, the cells were stained with annexin V-FITC and propidium iodide to evaluate apoptosis using flow cytometry and toanalysis transcription of Fas using RT-PCR. Results: Compared with controlgroup, FasL (20 ng / mL), IL- 1β (10 ng / mL) (P <0.05); apoptosis, apoptosis induced by apoptotic cells (P <0.05); FasL (10 ng / mL) enhanced the apoptosis-inducing effects of FasL (5 ng / mL) and FasL (20 ng / mL) was induced by 1 L-1β (10 ng / in disc cells. Fas gene transfer in all groups and Fas expression in the 5 treatment groups wereapproximately 1.2-2.1-fold greater than control group (respectively, P <0.05) .Additionally, Fas expression in FasL with IL- Signifi-cantly up-regulated than in FasL groups (P <0.01). Conclusions: The results of this study showed that disc cells pre-treated with IL-lbeta increased apoptotic rate inresponse to FasL in vitro and provided insights to understand Fas / FasL system- mediated apoptosis in disc cells which would be enhanced due to inflammation factor in degenerative disc.
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