Establishment of a diabetic neural stem cell glycolipid injury model and the influence of ZCL278

GAO Qi; GUO Ya-ru; GU Ting-ting; WU Zhi-xin; DENG Jiao

doi:10.13820/j.cnki.gdyx.20240233

Abstract：Objective To establish an in vitro model of neural stem cell(NSC)injury in the mouse hippocampal dentate gyrus area induced by high-fat high-sugar conditions,mimicking diabetic neuropathy,and to explore the role of CDC42 in high-fat high-sugar-related NSC injury.Methods NSCs from embryonic mice were cultured,and their stemness and purity were confirmed by immunofluorescence staining for Sox2 and Nestin.NSCs were cultured with differ-ent concentrations of glucose and oxidized low-density lipoprotein(ox-LDL)alone or in combination,and cell viability was assessed using the CCK8 assay to determine suitable glucose and lipid concentrations affecting NSC proliferation.NSCs were then cultured in high-fat high-sugar conditions(HHM)or regular medium(RM),and their effects on NSC proliferation were assessed by CCK8 assay,scratch assay,and Transwell migration assay.Protein extracts from the hipp-ocampal tissues of 10-week-old db/db mice and NSCs cultured under different conditions were subjected to Western blot analysis to detect CDC42 expression.The specific CDC42 inhibitor ZCL278(10 µmol/L)was used in vitro to assess its therapeutic effect on high-fat high-sugar-induced NSC proliferation impairment.Results Glucose concentrations of 10-40 mmol/L and ox-LDL concentrations of 25-35 μg/mL inhibited the proliferation of cultured NSCs.A combi-nation of 10 mmol/L glucose and 25 μmol/L oxLDL was selected as the model of HHM-induced NSC injury,which con-tinuously inhibited NSC proliferation within 72 h.Scratch and Transwell migration assays showed that this combination sig-nificantly inhibited NSC migration.Increased expression of CDC42 was observed in the hippocampus and NSCs of diabetic mice.In vitro treatment with the CDC42-specific inhibitor ZCL278 increased the viability of NSCs cultured under HHM conditions,suggesting partial reversal of HHM-induced NSC proliferation impairment.Conclusion A combination of 10 mmol/L glucose and 25 mmol/L ox-LDL can simulate glycolipid injury of neural stem cells in diabetic animals,af-fecting NSC proliferation and migration.Overexpression of CDC42 is involved in high-fat high-sugar-induced inhibi-tion of NSC proliferation.The CDC42-specific inhibitor ZCL278 can partially reverse high-fat high-sugar-induced NSC proliferation impairment.These results provide an in vitro model for studying NSC proliferation and migration impair-ments associated with abnormal glycolipid metabolism,and suggest that the CDC42-specific inhibitor ZCL278 may par-tially reverse high-fat high-sugar-related NSC proliferation impairment.