Influence Of Pressure On The Viability Of Retinal Ganglion Cells In Vitro

ObjectiveGlaucoma is the second leading cause of blindness globally. It is a group of diseases characterized by apoptosis of retinal ganglion cell. Elevated intra-ocular pressure is the most important risk factor for glaucoma,however,the exact celluar mechanisms remain unclear. Genetic studies have established that POAG is genetically heterogeneous. OPTN is identified as one of the pathogenic genes associated with POAG. This study is to investigate viability, apoptotic progression and the expression of OPTN gene of RGC-5 cultured under the elevated pressure and primarily discuss the direct influence of pressure on RGC.MethodsRGC-5 were cultured and submitted to elevated pressures for 48 hours. Elevated pressure conditions were 20,40,60,80mmHg for experimental groups and OmmHg for normal control. The viability and apoptosis of RGC-5 were detected by MTT method and FITC-AnnexinV labeling.The expression of OPTN in RGC-5 was detected using immunohistological method,Western-blot and Q-Realtime-PCR technique, respectively. Data is presented as the mean±SEM and statistically compared by paired t-test with SPSS 13.0.Results1,Cultured RGC-5 mainly showed a spindle-shaped morphology with an average time of 2 to 3 days for each division.2,Compared with control group, the cell growth and proliferation in RGC-5 were significantly inhibited in pressure-elevated groups (p<0.01)and the rate of apoptosis were significantily higher in pressure-elevated groups(p<0.05). We demonstrated that 40mmHg induce the early stage of apoptosis and 60mmHg induce the late stage of apoptosis and 80mmHg induce cell death. 3,OPTN was highly expressed in RGC-5 in normal control group. Within the 20-40mmHg, the expression of OPTN significantly decreased with the pressure elevation as compared with control group (p<0.05) and within the 60-80mmHg, the expression of OPTN significantly increased with the pressure elevation as compared with control group (p<0.05).Conclusions1,Pressures had direct damages to the viability of RGC-5.2,As the elevation of the pressure, the viability of RGC-5 decreased and the rate of apoptosis and death increased.3,As the elevation of the pressure, the expression of OPTN gene showed a V shaped fluctuation. The fluctuation implied the regulation of OPTN gene on the viability of RGC-5 in vitro changed according to the pressure conditions, and OPTN gene possibily play a protecvive role on late stage of impaired cells.