| Objective: Diabetic encephalopathy (DE) is the central nervous system complication of diabetes, which is chronic and progressive. Its clinical symptoms are mild to moderate cognitive impairments, such as the decline of learning, memory and problem-solving abilities. When the diabetic encephalopathy occurs, there are corresponding changes on brain morphology, nerve electrophysiology and molecular biology. Although researchers have studied DE from various angles, the pathogenesis of DE is still unclear. Recent studies found that insulin was a neurotrophic factor of nerve cells. The biopotency changes of insulin in the brain play an important role in the central nervous systems of diabetes patients. The insulin signaling pathway begins with the combination of insulin and insulin receptor substrate. The insulin signaling pathway disturbance will lead to insulin resistance (IR) which means that the biopotency of normal dose insulin is lower than normal. Our previous studies indicated that endoplasmic reticulum stress (ERS) played an important role in Alzheimer's disease (AD) and its pathological products. The concentration of insulin is increased in the cerebrospinal fluid of AD patients, compared to the control ones. The insulin receptor density in the occipital regions is increased and the tyrosine kinase activity is reduced. Furthermore, evidences indicated that there exists a close relationship between endoplasmic reticulum stress and insulin resistance. Based on these, our experiment toke endoplasmic reticulum stress as a starting point to investigate the pathogenesis of diabetic encephalopathy which may be a very important work. Diabetes mellitus is named Xiaokebing in the Traditional Chinese Medicine. Its basic pathogenesis is yin-deficiency and inner heat. Although lung, spleen, and kidney are the organs related to the pathogenesis of Xiaokebing, the point that spleen-deficiency contributes to Xiaokebing draws more and more attention now. Zibu Piyin Recipe (ZBPYR) is created by Professor Libin Zhan. It could reinforce spleen and nourish spleen yin. It was established to treat spleen-yin deficiency syndrome.Previous experiments showed that ZBPYR had notable effects on neuroprotection and anti-aging of brain, and could cure the spleen-yin deficiency in Alzheimer's disease animal models and primary cultured rat hippocampus neurons against Aβ1-40 toxicity. Therefore, this study observed the role of ZBPYR in changing insulin signaling pathway in hippocampus and its possible mechanisms in spleen-yin deficiency diabetic encephalopathy rats.Methods:1.Healthy adult male Sprague Dawley (SD) rats were divided randomly into control group, spleen-yin deficiency group, diabetic encephalopathy group, spleen-yin deficiency diabetic encephalopathy group and spleen-yin deficiency diabetic encephalopathy+ZBPYR group, four rats each group.2.After the acclimatization, the control group and spleen-yin deficiency group were feed with foundational diet. Other groups were feed with high fat diet. 4 weeks later, the rats which were feed with high fat diet intraperitoneal injected with 1% STZ (30mg/kg) once. The rats whose radom blood glucose was more than 16.7mmol/L were identified as diabetes mellitus models after 72h. We detected the level of fasting serum insulin (FSI), oral glucose tolerance test (OGTT) and insulin tolerance test (ITT) after intraperitoneal injection with 1% STZ on the 4th,8th,12nd day. We induced spleen-yin deficiency encephalopathy model rats by defatigation, hungry-full diet and drenching the drug of damaged yin. ZBPYR group were induced by the intervention of ZBPYR in 16 days and decapitated for collecting hippocampus.3.Morris water maze test and step-down test were used to evaluate the behavioral changes.4.Using the Western Blotting method to detect the protein expression of JNK, phospho-JNK, IRE1αand IRS-1, phospho-IRS-1 in hippocampus. Results:1.After STZ injection, the diabetes mellitus rats blood glucose was significantly higher than nondiabetes rats (P<0.01). In the oral glucose tolerance test, the diabetes mellitus rats presented impaired glucose tolerance. In the insulin tolerance test, it suggests that diabetes mellitus rats presented insulin resistance. the rate of decline of blood glucose in diabetes rats was more slowly than nondiabetes rats. However, fasting serum insulin levels in diabetes rats and non-diabetes rats had no significantly difference (P>0.05).2.The spleen-yin deficiency group rats showed increased water drinking (P<0.01), and rectal temperature (P<0.05). The results of Morris water maze test and step-down test showed that the learning and memory abilities of the diabetic encephalopathy group and the spleen-yin deficiency diabetic encephalopathy group were reduced significantly (P<0.05), but that of the ZBPYR treatment group were increased significantly (P<0.05).3.Western blotting results showed that compared with the control group and spleen-yin deficiency group, the hippocampus phospho-IRS-1 phospho-JNK and IRE1αprotein expression of diabetic encephalopathy group and spleen-yin deficiency diabetic encephalopathy group were increased (P<0.05), but these of ZBPYR treatment group were weakened.Conclusion:1.The spleen-yin deficiency diabetic encephalopathy rats model was successfully established through the way of combination of syndrome and disease. The results of Morris water maze test and step-down test showed that the learning and memory of the diabetic encephalopathy group and the spleen-yin deficiency diabetic encephalopathy group were reduced.2.ZBPYR can be significantly improved learning and memory abilities of spleen-yin deficiency diabetic encephalopathy rats .3.The function of ZBPYR improved learning and memory abilities of spleen-yin deficiency diabetic encephalopathy rats may be associated with insulin signaling pathway in hippocampus. ZBPYR can protect and maintenance conduction of insulin signaling pathway.4.ZBPYR can reduce the phospho-JNK and IRE1αprotein expression level and improve the endoplasmic reticulum stress to regulate the insulin signaling pathway. |
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