Effect Of MGluR3 On The Expression Of Cyclin D1 Of Proliferation Of Human Embryonic Cortical NSCs And Molecular Mechanism

Neural stem cells(NSCs),which exist in the central nervous system of mammals(CNS),are self renewing and pluripotent,and can be isolated from mammalian embryos and adult brains.There are two primary germ zone:the subventicular zone(SVZ)beneath the lateral wall of the lateral ventricles and the subgranulae zone(SGZ)in the dentate gyrus of the hippocampus.They can be stimulated to proliferate and differentiate by some physiological and various areas of adult brain and kept their multiple potential in vitro.The nerve injury is one of the most common disease in the clinical,that is cause serious damage to human health.The major pathological changes of which is mass mortality and loss of neurons,It can lead to a series of neural dysfunction.However,there is no efficient therapy yet until now.That gives the family and society a heavy burden.Recent years,Many central nervous system diseases are widely accepted by neural stem cell replacement therapy.It can improve neurological function after cerebral ischemia,nerve injury and neurodegenerative disorder.These cells are differentiated and have potential to differentiate into different neurons,such as astrocytes,oligodendrocytes and neurons.The replacement therapy of neural stem cells is the hope for the treatment of nerve injury diseases.However,this series of molecular mechanisms is still unclear.It is providing to a theoretical foundation to treatment for CNS diseases.Glutamate excitatory poisoning is involved in clinical nerve injury,such as traumatic brain injury,epilepsy and cerebral ischemia.The metabolic glutamate receptor(mGluRs)belongs to the G-protein coupled receptor family,and the G-protein activates the MAPK signaling pathway by activating the PKC pathway.mGluRs includes mGluR1-8,and mGluR can activate phospholipase C through the lotus root G-protein.It is reported that mGluR5/7 can promote the survival,proliferation and differentiation of rat NSCs in vitro by activating the MAPKs signal transduction pathway.The main members of the MAPKs family include ERK,JNK and p38 MAPKs,which are involved in regulating cell proliferation and apoptosis.Acute and chronic severe brain dysfunction may occur in human and animal models after brain damage,which is a very complex cascade of cellular activity.Recently,a series of research evidence suggests that brain damage can promote the proliferation of NSCs,but its molecular mechanism is still unclear.Previous studies have found that type 5/7 in the mGluR family can promote the survival,proliferation and differentiation of NSCs in rats,and mGluR3 is expressed in the human embryonic cerebral cortex NSCs.Therefore,we hypothesized that mGluR3 plays a role in regulating the proliferation of NSCs in human embryonic cerebral cortex during development.In this study,we studied the effect and molecular mechanism of mGluR3 on the expression of Cyclin D1 in human fetal cerebral cortex by NSCs,which will provide new targets and theoretical basis for our clinical treatment of neurologic diseases.Methods: 1.To isolate and culture NSCs: reagent preparation?embryo preparation?primary culture?subculturing.2.Real time PCR was used to analyze Cyclin D1 mRNA in human NSCs.3.The effect of mGluR3 on the expression of Cyclin D1 in human embryonic cerebral cortex NSCs and the activation of ERK,JNK and p38 phosphorylation was detected by Western blot method.4.Real time PCR was used to analyze mGluR3 activation of MARKs signal pathways on proliferation of NSCs.Results: 1.Human NSCs were isolated from 14-week fetuses cortex.neurospheres were formed by monocell culture.It was proved by Nestin staining,self-renewal and multipotential differentiation potential as NSCs.2.The expression of NSCs Cyclin D1 in the brain cortex of normal cultured human embryo has a certain level.mGluR3 can increase the level of Cyclin D1 mRNA and protein expression significantly.3.mGluR3 activator can promote the significant increase in the expression of p-ERK1/2 and p-JNK2,activate the MAPK signaling pathway,and thus promote the effect of Cyclin D1 mRNA expression in NSCs.4.mGluR3 decreased the phosphorylation level of p38,reduced the activity of p38 signaling pathway to inhibit the apoptosis of NSCs,and could induce the proliferation of embryonic neural stem cells and inhibit the apoptosis of neural cells.1.mGluR3 promotes the proliferation of NSCs in vitro.Conclusion: 1.mGluR3 can upregulate the expression level of NSCs Cyclin D1 in human fetal cerebral cortex,and then induce cells to cross the G1/S phase node,promote DNA replication and mitosis,and NSCs proliferation in human fetal cerebral cortex.2.mGluR3 can increase the level of phosphorylation of ERK and JNK in the NSCs of human embryonic cerebral cortex.By activating the ERK and JNK signaling pathways,the expression of Cyclin D1 mRNA in NSCs is promoted,thus promoting the proliferation of NSCs in the human embryonic cerebral cortex.3.mGluR3 can activate the MAPK signal transduction pathway,and can significantly promote the expression of Cyclin D1 protein,thus promoting the proliferation of NSCs in human embryonic cerebral cortex.