Long Non-coding RNA H19 Induces Hippocampal Neuronal Apoptosis Via Wnt Signaling In A Streptozotocin-induced Rat Model Of Diabetes Mellitus

OBJECTIVE:Diabetes with cognitive impairment is associated with decreased ability to learn and memory,comprehension and judgment,and other serious impairments in quality of life.Moreover,cognitive dysfunction can make diabetic patients’ compliance with treatment worse,thereby aggravating diabetes,and both form a vicious circle.Synaptic plasticity in the hippocampus and disorders of memory and learning function are central nervous system complications of diabetes.Here,we used streptozotocin-induced diabetic rat model to study the role of lncRNA H19 in memory,learning and hippocampal apoptosis,and to clarify the role of Wnt signaling pathway.To explore a new way to treat cognitive function decline in patients with diabetes.METHODS:STZ diabetic rat models were established and the experimental animals were divided into 6 groups(n = 10 in each group)by stereo injection: normal group,model group: empty vector group,overexpressed lncRNA H19 group,NC group,and lncRNA H19-sh RNA group.The Morris water maze test was used to test the learning and memory ability of each group of rats.HE staining and electron microscopy were performed to observe the change of neuronal cell number,pathological changes and ultrastructural changes in hippocampal CA1 region in each group.The expression of lncRNA H19 in hippocampus tissues of each group was analyzed by q RT-PCR.The differences in the methylation of the whole unit and the differences in the methylation of the H19 promoter region were examined.Western blot and q RT-PCR were used to study the Wnt signaling pathway and the apoptosis-related representative proteins Wnt3,GSK-3β,β-catenin,TCF-1,Bax,Bcl-2,Caspase-3,Caspase-8 Expression,as well as changes in the expression of the corresponding genes and m RNA.Apoptosis of hippocampal neurons in 6 groups was detected by TUNEL method,and the apoptosis rate was calculated.The data was analyzed using SPSS 18.0 statistical software and the results are expressed as the mean ± s.The t-test was used to test the comparison between the two groups with normal distribution.One-way analysis of variance(ANOVA)was used to test the comparison among groups.P <0.05 indicated that the difference was statistically significant.RESULTS:Compared with the normal group,the hippocampus in the model group was hypomethylated(P < 0.01).The methylation level in the promoter region of model group H19 was lower than that in the normal group and the NC group(all P < 0.01).Expression of lncRNA H19 in hippocampus.Compared with the normal group,the expression of lncRNA H19 was significantly increased in the empty vector model,NC,lncRNA H19 overexpression group.Compared with the normal group,the latency of positioning navigation of the empty vector group,the overexpressed lncRNA H19,the model,and the NC group was significantly increased,and the duration of spatial exploration was significantly shortened.In the empty vector group,the overexpressed lncRNA H19 group,the model,and the NC group,nerve cells were loosely arranged and cell morphology was changed.These changes include nuclear atrophy and cytoplasm reduction,indicating that apoptosis occurs.The number of overexpressed lncRNA H19 mutants decreased,and the interstitial degeneration gap increased with the progression of intrasynaptic mitochondrial vacuolation.The expression of Wnt3,β-catenin and TCF-1 in the lncRNA H19 overexpression group increased,but the expression of GSK-3β decreased(P <0.05).The overexpression of lncRNA in H19 group increased the apoptosis rate of hippocampal neurons,increased the levels of Bax,caspase-3 and caspase-8 proteins,and decreased the expression of Bcl-2(P <0.05).CONCLUSION:The expression of lncRNA H19 increased in DM rats.LncRNA H19 mediates the occurrence of cognitive impairment in diabetic rats,but H19 knockdown markedly improved the learning and memory ability of experimental animals.Wnt signaling is activated by lncRNA H19.Silencing H19 can inhibit the activity of Wnt signaling pathway.Overexpression of lncRNA H19 can induce apoptosis of hippocampal neurons in diabetic rats.Silencing lncRNA H19 can increase the survival of hippocampal neurons.The overexpression of lncRNA can activate the expression of apoptosis-related proteins.On the contrary,the expression of apoptosis-related proteins is decreased when H19 is silenced.lncRNA H19 mediates apoptosis in hippocampal neurons and synaptosomes via Wnt signaling pathways.Inhibition of lncRNA H19 may be a promising new method for treating cognitive decline in DM patients.