Effect of different temperatures on the expression of sumoylation related protein in SHSY5Y cells under oxygen-glucose deprivation

Lv Ruoning; Tong Yuanyuan; Liu Jinping

doi:10.13498/j.cnki.chin.j.ecc.2018.01.11

Abstract：Objective To investigate how temperature affects SHSY5Y cells expressing sumoylation related protein under hypoxic-ischemic stress. Methods The SHSY5Y cells were exposed to oxygen - glucose deprivation (OGD) with different temperature (18℃, 26℃, 30℃, 37℃). The morphological change was observed by inverted microscope. Expression of UBC9 was detected by qPCR and western blot, and sumoylation related protein SUMO1 and SUMO2/3 was detected by western blotting at the time we ended oxygen deficit. 24 hours after reperfusion, LDH and caspase-3 released in cell culture supernatant were detected by western blotting. Results LDH was significantly higher after 4 hours OGD, and hypothermia reduced LDH released in culture than that in 37℃ group. Western blotting results showed hypothermia could up-regulate SUMO1-conjugated expression, which was significantly different between 18℃ and 30℃ (P ＜0.05, n = 6), but not between 18℃ and 26℃ (P ＞0.05, n = 6). Hypothermia increased the ratio of conjugated and free SUMO1 (P ＜0.05, n = 6); on the other hand, hypothermia up-regulated expression of both SUMO2/3-conjugated and SUMO2/3-free. OGD increased the ratio of conjugated and free state of SUMO2/3 (P ＜0.05, n = 6). There was no significant difference in UBC9 expression between experimental group and control group (P ＞0.05, n = 6). After 4 hours of OGD and 24 hours reperfusion, caspase-3 protein level increased and hypothermia reduced the expression of caspase-3 (P ＜0.05, n = 6). OGD and hypothermia did not affect UBC9 mRNA expression (P ＞0.05, n=3). Conclusions Nervous cells express fewer SUMO1 and SUMO2/3 protein when lacking nutrition, low temperature could protect nervous cells possibly by global SUMOylation, especially through enhanced level of conjunctive capacity of SUMO1. No difference of protective effect was found in 26℃, 30℃ and 18℃ after 4 hours OGD.