Effects Of Lycium Barbarum Polysaccharides On Oxidative Stress-induced Retinal Ganglion Cells Apoptosis

Objectives:In this study we investigated the protective effects of lycium barbarum polysaccharides on oxidative stress-induced RGC-5 apoptosis and elucidated the related mechanisms, providing a new idea for new therapy strategies to protect visual performance.Methods:RGC-5 were treated with different concentrations of H2O2 (50,100,200,400μmol·L-1),200μmol·L-1 H2O2 clearly induced cell death in 49.68%of the cells within 24 hours,so apoptosis models of RGC-5 induced by 200μmol·L-1 H2O2 was established.Cell apoptosis inhibition ratio of RGC-5 treated by different concentrations of LBP(10 mg·L-1,20mg·L-1,40mg·L-1) for 24 hours was measured with MTT assay,The experiment were divided randomly into the control group,LBP group,H2O2 group and H2O2+LBP group. The apoptotic morphology and apoptotic ratio of RGC-5 was observed by DAPI staining and flow cytometry.In additon, c-Myc,Caspase3,Bcl-2 and Bax expression in RGC-5 were caculated real-time PCR and Western-blot.Results:The results of MTT colorimetry showed that survival ratio of RGC-5 were treated with different concentrations of H2O2 were 91.33±10.0, 87.67±8.78,49.68±7.48,26.50±4.89; H2O2 induce cell death of RGC-5 cells in a concentration-dependent manner.The survival ratio of RGC-5 in (10 mg≮L-1.20mg≮L-1,40mg≮L-1) LBP group were 52.53±10.10,62.88±10.20, 68.62±5.39, Compared with the H2O2 group, treatment of LBP significantly decreased cell apoptosis(P< 0.05). DAPI staining revealed that H2O2 canr e d uc e R G C- 5 c e l l s a pp e a r e d a p op t ot i c c ha n ge s, su c h a s c hr o ma t i n c o n d e n s a t i o n, c o n v o l u t e d n u cl e i w i t h c a v i t a t i o n s, fr a g m e n t a t i o n o f t h e n u c l e u s a n d a p o p t o t i c b o d i e s, L B P c a n d e c r e a s e d c el l mo r p h o l o g i c i mpa i r me n t. Fl o w c yt o me t r y s h o we d a p o p t o s i s r a t i o o f R G C- 5 i n c o n t r o l g r o u p, L BP g r o u p, H2O2 g r o u p a n d H2O2+ L BP g r o u p we r e 4. 9 0 %, 4. 2 5 %, 3 2. 3 5 % a n d 2 4. 2 1 % r e s p e c t i v e l y; T h e r e w e r e s t a t i st i c a l d i ff e r e n c e s b e t we e n c o n t r ol g r o u p, H2O2 g r o u p a n d H2O2+ L BP g r o u p( a l l P < 0. 0 5). Q u a nt i t a t i ve r ea l- t i me P C R a n d We s t e r n- b l o t r e s u l t s s h o w e d t ha t t h e e x pr e ssi o n o f B a x, C a spa se- 3, c- My c i n c r e a s e d a n d B c l- 2 d e c r e a s e d i n H2O2 g r o u p c o mp a r e d wi t h c o n t r o l g r o u p. C o m p a r e d w i t h H2O2 g r o u p, e x p r e s s i o n o f Ba x, C a s p a s e- 3, C- m y c d e c r e a s e d a n d t he e x pr e ssi o n o f B c l- 2 i n c r e a s e d i n H2O2+ L B P g r o u p, t h e d i ff e r e n c e wa s s t a t i st i c a l l y s i g n i f i c a n t( P< 0. 0 5). Conclusion: 1. Our study demonstrated that LBP was able to effectively p r o t e c t r e t i n a l g a n g l i o n c e l l a g a i n st o xi d a t i v e- st r e s se d i nj ur y. 2. T h e p r o t e c t i ve e ff e c t m a y b e a s s o c i a t e d wit h th e do wn-r eg u l a t i o n o f c- M y c, Ca s p a s e- 3 a n d Ba x p r o t e i n s, u p- r eg u l a t i o n o f Bcl- 2 e x p r e s s i o n.