Effects of hydrogen on oxidative stress injury induced by high glucose in rat Schwann cells: the relationship with parthanatos

Objective To evaluate the effects of hydrogen on oxidative stress injury induced by high glucose in Schwann cells and its relationship with PARP-1-dependent cell death(parthanatos). Methods Primary rat Schwann cells were cultured in 96-well plate(1×104 cells/ml, 200 μl/well)or in 6-well plate(1×106 cells/ml, 2 ml/well)with RSM culture medium and were randomly divided into 5 groups(n=30 each): control group(group C), hydrogen group(group H2), high glucose group(group HG), high glucose plus hydrogen group(group HG+ H2)and high osmotic control group(group M). The cells were cultured in the common culture medium in C, HG and M groups.The cells were cultured in hydrogen-rich culture medium in H2 and HG+ H2 groups.In HG and HG+ H2 groups, 50 mmol/L of glucose was added to the culture medium.In C and H2 groups, the equal volume of normal saline was added to the culture medium.In M group, mannitol 44.4 mmol/L was added to the culture medium.The cells were then incubated for 48 h. After 48 h of incubation, the cell viability was measured using CCK-8 assay, intracellular reactive oxygen species(ROS)level was detected by flow cytometry, the concentration of 8-hydroxy-2-deoxy Guanosine(8-OHdG)was determined by ELISA, and the expression of poly(ADP-ribose)-polymerase-1(PARP-1), cleaved-PARP-1, poly(ADP-ribose)(PAR), and apoptosis-inducing factor(AIF)in the total protein and nucleus was measured by Western blot.PARP-1 activity(cleaved-PARP-1/PARP-1)and AIF nuclear translocation were recorded. Results Compared with C and H2 groups, the cell viability was significantly decreased, and PARP-1 activity, expression of ROS, 8-OhdG and PAR, and AIF nuclear translocation were increased in HG and HG+ H2 groups.Compared with HG group, the cell viability was significantly increased, and PARP-1 activity, expression of ROS, 8-OhdG and PAR, and AIF nuclear translocation were decreased in HG+ H2 group.There was no significant difference in each parameter between M and C groups. Conclusion Hydrogen can reduce oxidative stress injury induced by high glucose in Schwann cells, and the mechanism is related to inhibition of parthanatos. Key words: Diabetic neuropathies; Hydrogen; Oxidative stress; Cell death