MSC移植对视网膜缺血—再灌注损伤后神经节细胞中HIF-1α及Caspase-3表达的影响

Effect of mesenchymal stem cell transplantation on HIF-1α and caspase-3 Expression after retinal ischemia-reperfusion injury

目的探讨间充质干细胞(mesenchymal stem cells,MSC)移植对视网膜缺血-再灌注损伤低氧诱导因子-1α(hypoxia inducible factor-1α,HIF-1α)及Caspase-3表达及神经节细胞凋亡的影响,为MSC移植治疗视网膜缺血-再灌注损伤提供实验依据。方法将65只健康SD大鼠随机分为3组:正常组5只、模型组30只、MSC移植组30只,各组均将左眼作为实验眼制作视网膜缺血-再灌注损伤模型。贴壁筛选法体外培养SD大鼠MSC,模型组于缺血-再灌注1h予以玻璃体内注射PBS缓冲液,MSC移植组于缺血-再灌注后1h予以玻璃体腔注射MSC。各组SD大鼠分别于缺血-再灌注损伤后1h、6h、12h、24h、48h及72h处死,并立即摘除左眼眼球,固定包埋并切片,免疫组织化学法检测各组SD大鼠视网膜缺血-再灌注后HIF-1α及Caspase-3的表达情况。结果正常组未检测到HIF-1α及Caspase-3有表达,在视网膜缺血-再灌注后1h、6h、12h、24h、48h及72h模型组HIF-1α的表达分别为0.620±0.025、1.889±0.099、7.591±0.359、4.756±0.156、2.564±0.164、1.023±0.144;MSC移植组分别为0.222±0.058、0.774±0.162、1.831±0.124、1.567±0.059、0.930±0.010、0.583±0.065。模型组Caspase-3表达分别为0.106±0.093、0.593±0.083、1.760±0.087、3.855±0.142、2.429±0.070、1.329±0.080;MSC移植组分别为0.028±0.004、0.180±0.032、0.887±0.0075、1.857±0.050、1.554±0.079、0.957±0.087。与模型组相比,MSC移植组HIF-1α及Caspase-3的表达明显减少,差异均有统计学意义(均为P<0.05)。结论视网膜缺血-再灌注损伤后行MSC玻璃体内注射可抑制神经节细胞HIF-1α及Caspase-3的表达,减少神经节细胞的凋亡,对视网膜缺血-再灌注损伤有一定的保护作用。

Objective To investigate the effects of mesenchymal stem cells （MSC） transplantation on neural cell apoptosis and expressions of hypoxia inducible factor-1α （HIF-1α） and caspase-3 after retinal ischemia-reperfusion injury, and provide the experimental basis of MSC transplantation for retinal ischemia and reperfusion. Methods A total of 66 healthy SD rats were randomly divided into three groups：nor- mal group （5 cases） ,model group （30 cases） and MSC transplanting group （30 cases）, and retinal ischemia-reperfusion injury model was established in left eye of each group. MSC of SD rat were cultured by adherent culture method in vitro ,PBS buffer was intra- vitreal injected at 1 hour after injury in model group ,MSC was intravitreal injected at 1 hour after injury in MSC transplanting group. SD rats of each group were sacrificed at 1 hour,6 hours, 12 hours, 24 hours,48 hours and 72 hours after injury, and the left eyeballs were removed immediately ,then fixed embedding and sectioning,and the expression of HIF-1α and caspase-3 in each group were detected by immunohistochemistry assay. suits There was no expression of HIF-1α and caspase-3 in normal group. The levels of HIF-1α at 1 hour,5 hours, 12 hours,24 hours,48 hours,72 hours after injury in model group were 0. 620 ± 0.025,1. 889 ± 0. 099,7. 591 ± 0. 359,4. 756 ± 0. 155,2.564 ± 0. 164 and 1. 023± 0. 144, respectively, which in MSC transplanting group were 0. 222 ± 0. 058, 0. 774± 0. 152,1.831 ± 0. 124,1. 567± 0. 059,0. 930± 0.010 and 0.583 ±0.065, respective- ly;The levels of caspase-3 at 1 hour,6 hours,12 hours,24 hours,48 hours,72 hours after injury in model group were 0. 106 ± 0.093,0. 593± 0. 083,1. 760 ± 0. 087,3. 855 ±0. 142, 2.429± 0. 070 and 1. 329±0. 080, respectively, which in MSC transplanting group were 0.028±0. 004,0. 180 ± 0. 032,0. 887 ± 0. 075,1. 857± 0. 050,1. 554± 0. 079 and 0. 957 ±0. 087, respectively. Compared with model group ,the expression of HIF-1α and caspase- 3 in MSC transplanting group were reduced obviously, there were statistical differences （both P 〈 0.05 ）. Conclusion After retinal ischemia-reperfusion injury, the intravit- real injection of MSC can inhibit the expression of HIF-1α and caspase-3 in ganglion cells, reduce the apoptosis of ganglion cells and have a protective effect on retinal ische- mla-reoerfusion injury.