Inhibitory effect of small interfering RNA-TLR9 on high glucose-induced retinal ganglion cell apoptosis / 中华实验眼科杂志

ABSTRACT

Objective@#To investigate the effect of Toll-like receptor 9 (TLR9) on high glucose-induced retinal ganglion cells-5 (RGC-5) apoptosis and the inhibitory effects of small interfering RNA-TLR9 (si-TLR9) on apoptosis.@*Methods@#RGC-5 cells were divided into normal control group, high glucose group, high glucose+ negative control group and high glucose+ si-TLR9 group which cells were respectively dealt with normal culture medium, high glucose medium, transfection of non-specific siRNA under high glucose and transfection of siRNA-TLR9 under high glucose.The expression of TLR9 mRNA was detected by real time PCR; the survival rate of the cells was evaluated by MTT assay; the apoptotic rate of the cells was detected by flow cytometry; the caspase-3 activity in the cells was detected by related kit, and the expressions of TLR9, B cell lymphoma (bcl-2), bcl-2 associated X protein (bax), p38 mitogen-activated protein kinases (p38MAPK) and phosphorylated (p)-p38MAPK proteins were detected by Western blot.@*Results@#The expressions of TLR9 mRNA and protein in the high glucose+ si-TLR9 group were significantly decreased in comparison with the high glucose+ negative control group (both at P<0.05). The cell survival rate of the high glucose group and high glucose+ negative control group was (78.36±5.13)% and (75.12±4.25)%, respectively, which was significantly lower than (95.48±7.25)% in the normal control group, and that in the high glucose+ si-TLR9 group was (86.58±5.32)%, which was significantly reduced in comparison with the high glucose+ negative control group (all at P<0.05). The apoptotic rate of the cells in the high glucose group and high glucose+ negative control group was (13.23±1.22)% and (12.52±1.38)%, respectively, which was significantly higher than (2.26±0.15)% of the normal control group, and apoptotic rate in the high glucose+ si-TLR9 group was (7.15±0.24)%, which was significantly lower than that in high glucose+ negative control group (all at P<0.05). The expression of bax in the cells of the high glucose+ si-TLR9 group was significantly decreased, and the expression of bcl-2 in the cells of the high glucose+ si-TLR9 group was significantly increased in comparison with the high glucose+ negative control group (both at P<0.05). Caspase-3 activity in the cells was significantly decreased in the high glucose+ si-TLR9 group compared with the high glucose+ negative control group (P<0.05). The relative expression of p-p38MAPK in the cells was significantly decreased in the high glucose+ si-TLR9 group compared with the high glucose+ negative control group (P<0.05).@*Conclusions@#Down-regulation of TLR9 expression in the RGC-5 cells can inhibit the apoptosis induced by high glucose probablely by suppressing p38MAPK signaling pathway.