The Role And Mechanism Of MKP7 In The Regulation Of Glucolipotoxicity In Islet β Cells

Diabetes is characterised by insulin resistance,insufficient insulin secretion or excessive glucagon secretion as a direct result of hyperglycaemia.As of 2019,the prevalence of diabetes in China has reached 10.9%(116 million people),making it the most prevalent country in the world.Of these,type 2 diabetes mellitus accounts for approximately 90% of the total prevalence of diabetes.Type 2 diabetes mellitus is a condition of impaired glucose regulation caused by insulin resistance and progressive damage to the pancreatic β cells.Continued development of hyperglycaemia can further lead to complete islet β cell failure.Evidences from multiple sources,including prospective epidemiological studies,human genetics and experimental validation,suggest that glucolipotoxicity is the primary cause of islet β cell damage in type 2diabetes mellitus,but the underlying mechanisms remain unclear.Mitogen-activated protein kinase phosphatase(MKP),also known as dual specificity phosphatase(DUSP),is a negatively regulated phosphatase of Mitogenactivated protein kinase(MAPK).As a negatively regulated phosphatase of MAPK,the C-terminal catalytic domain of MKP is able to inactivate MAPK by dephosphorylating its threonine and/or tyrosine residues.It has been shown that MKP7 regulates insulin resistance and insulin secretion abnormalities in muscle tissue and alleviates inflammatory responses and dyslipidemia in the liver,suggesting that MKP7 may be involved in regulating the pathological process of diabetes.However,the role of MKP7 in pancreatic tissues is still unknown.Therefore,this study conducted a preliminary study on the regulatory role of MKP7 on glucolipotoxicity injury of pancreatic β cells and its molecular mechanism.Firstly,we found that MKP7 expression was significantly reduced in the pancreatic tissues of HFD(High-fat diet)mice,suggesting an association between MKP7 and islet β cell damage.We then constructed a MIN6-MKP7 stable overexpression cell line and a lentivirus that interfered with MKP7 expression to investigate the effects of overexpression and inhibition of MKP7 expression on the regulation of glycolipotoxic damage in MIN6 cells,respectively.The results of the cell proliferation assay showed that MKP7 overexpression effectively alleviated the inhibitory effect of glucose and palmitic acid(GP)treatment on the proliferation of MIN6 cells.We further found that MKP7 could alleviate GPinduced decrease in relative expression levels of Bcl-2/BAX,increase in Caspase-12,Caspase-9/3 activation levels and increase in CHOP expression levels,indicating that it could inhibit GP-induced apoptosis in MIN6 cells;by assaying kinases p IRE-1α,p EIF-2α in endoplasmic reticulum stress overexpression of MKP7 inhibited the expression of GP-related endoplasmic reticulum stress signaling molecules,while interference with MKP7 expression increased the level of endoplasmic reticulum stress,suggesting that MKP7 could alleviate the level of GP-stimulated endoplasmic reticulum stress in MIN6 cells.The inflammatory response is closely related to islet β cell apoptosis.In this study,we examined the effects of GP treatment on inflammation and oxidative stress in MIN6 cells.Real-time PCR results showed that MKP7 was able to inhibit the gene transcription levels of pro-inflammatory factors IL-6,IL-1β and TNF-α in MIN6 cells under GP stimulation.MKP7 was also able to alleviate GP-induced oxidative stress in MIN6 cells by measuring the transcript levels of ROS production and oxidative stressrelated genes HIF-1α and PDH-3 in GP-treated MIN6 cells.Therefore,this study also verified the regulatory role of MKP7 on the MAPK protein family in GP-treated MIN6 cells.The results showed that overexpression of MKP7 significantly inhibited the phosphorylation levels of JNK,P38 and ERK in GP-stimulated MIN6 cells,but the inhibition of ERK was transient,whereas inhibition of MKP7 expression enhanced the phosphorylation levels of JNK,P38 and ERK simultaneously.In terms of pancreatic β cell function,the results of the Glucose-stimulated insulin secretion(GSIS)assay showed that MKP7 was effective in alleviating the impaired glucose-stimulated insulin secretion in MIN6 cells caused by GP stimulation,while Real-time PCR and Western blot results also showed that MKP7 had a regulatory effect on the levels of p AKT,GLUT2 and PDX-1 in MIN6 cells induced by GP stimulation.The results of Real-time PCR and Western blot also showed that MKP7 could regulate the levels of AKT,GLUT2 and PDX-1 in MIN6 cells under GP stimulation and could resist the GP-induced dysfunction in MIN6 cells.Given that AKT is essential for the survival of islet beta cells,we investigated the regulatory role of AKT in pancreatic islet β-cells using the AKT inhibitor MK-2206.The results showed that treatment with MK-2206 significantly elevated dysfunction,endoplasmic reticulum stress,apoptosis,oxidative stress and inflammatory responses in MIN6 cells,and that inhibition of AKT activation in MIN6 cells resulted in effects that were similar to those caused by GP treatment.Meanwhile,overexpression of MKP7 could alleviate the damage caused by MK-2206 to MIN6 cells by increasing the phosphorylation level of AKT,suggesting that MKP7 may regulate the process of glycolipotoxic damage in MIN6 cells through the AKT signaling pathway.In summary,during GP-induced islet β cell injury,MKP7 inhibited cellular inflammation and oxidative stress,and alleviated apoptosis and dysfunction,suggesting that MKP7 may serve as a potential target for type 2 diabetes mellitus biotherapy.In addition,MKP7 may alleviate glucolipotoxicity injury in pancreatic β cells through MAPK or AKT signaling pathways,but its exact mechanism of action still needs to be investigated in depth through in vivo experiments.