Chemerin Deficiency Alleviates Symptoms In PD Model Mice And Inhibits Neuro-inflammation

Parkinson’s disease（PD）is a neurodegenerative disease characterized by the loss of dopaminergic neurons in the substantia nigra of the midbrain and the accumulation of intracellular protein（α-synuclein）.The incidence rate of the disease is increasing,but the current treatment methods are not able to inhibit the progression of the disease.In particular,there is a lack of treatments that have few adverse effects and can halt the progression of PD in its early stages.In the process of PD chronic immune function has been changed,a variety of immune signals are actived and cause neuroinflammation.Macrophage-derived microglia play an important role in the molecular mechanisms of neuro-inflammation.Retinoic acid receptor responder 2（Chemerin）is an adipokine that affects glucolipid metabolism and participates in the regulation of inflammation.Chemerin is expressed in dendritic cells and macrophages and binds with its main receptor ChemR23 to exert immune function,promoting chemotaxis and phagocytosis of immune cells.However,its role in PD is still unclear.In this paper,PD mouse model,cellular inflammatory model and clinical blood samples were used to demonstrate the effect of chemerin on Parkinson’s disease and discuss its mechanism.Objective:Parkinson’s disease（PD）mouse model was established with MPTP and BV-2 cell inflammatory model was established with LPS.The effect of chemerin on DA neurons in PD model was demonstrated,and the mechanism of neuro-inflammation was discussed.Methods:1.In vivo experiment:Each mouse was injected intraperitoneally with MPTP at the dose of 25 mg/kg for 7 consecutive days,and the mice were divided into four groups:wild control（WT）group,Chemerin-/-（KO）mice,PD model of WT mice,and PD model of KO mice.The successful establishment of PD mouse model was confirmed by the following experiments:open field tests and Rotarod tests were used to detect the motor function of mice;The expression of TH in substantia nigra was observed by immunohistochemistry.Determination of antioxidant levels in substantia nigra and striatum was measured.After the model was established,the expression of chemerin receptor（ChemR23 protein）in the substantia nigra was detected by western blot.The activation of microglia in the substantia nigra was observed by immunohistochemical staining of IB A-1.The contents of TNF-α and IL-6 in substantia nigra were determined by ELISA.2.In vitro experiments:Lps-induced inflammatory model conditions were determined by BV-2 cell activity detection and cell morphology observation under inverted microscope.Cell activity was measured by CCK8.Transfected with si-chemerin,BV-2 cells were divided into four groups:blank control group,si-chemerin treated group,LPS-stimulated group and LPS-stimulated group after si-chemerin.The expression of chemeirn and its receptor in BV-2 cells was determined by qRT-PCR and western blot.The expression of pro-inflammatory factors in BV2 cells was determined by ELISA,qRT-PCR and western blot.The expressions of NF-κB and TLR4 in BV-2 cells were determined by qRT-PCR and western blot.The effects of Chemerin on glycolysis of BV-2 cells were studied according to the LDH activity in BV-2 cells.Conditioned medium of BV-2 was prepared and the effect of Chemerin on SH-SY5Y cell viability was detected.3.Clinical Samples:Peripheral blood samples from PD patients in Yangzhou University Affiliated Hospital were collected and the plasma was separated.The levels of Chemerin and ChemR23 were detected by ELISA.Results:1.Neuroprotective effect of Chemerin deficiency on MPTP-induced PD model miceChemerin deficiency improved dyskinesia in PD model mice:Chemerin deficiency increased the ability of autonomous activity,spatial exploration ability and balance ability.Chemerin deficiency increases TH expression in substantia nigra in PD mice model.Chemerin deficiency increased the antioxidant level of substantia nigra striatum in PD mice model:SOD activity increased,MDA content decreased.Chemerin deficiency inhibited the activation of microglia in the substantia nigra and reduced the expression of TNF-α and IL-6 by downregulating ChemR23 in the PD mouse model.2.Chemerin deficiency inhibits LPS-induced neuro-inflammation of BV-2 cellsStimulation of BV-2 cells with LPS at 1 μg/ml for 12 h was a suitable condition for establishing Bv-2 cell inflammatory model.Chemeim is upregulated in inflammatory environments.ChemR23 and GPR1 were up-regulated in BV-2 cell inflammatory model,while was down-regulated after si-chemerin transfection.The expression of TNF-α,IL-6 and IL-1β in BV-2 cells was down-regulated in LPS-induced inflammation model and the activation of TLR4/NF-κB signaling pathway was inhibited after si-chemerin transfection.The intracellular LDH activity of BV-2 cells decreased in LPS-induced inflammatory model after si-chemerin transfection.Conditioned medium of BV-2 cells increased the activity of SHSY5Y cells in LPS-induced inflammatory model after si-chemerin transfection.3.Comparison of Chemerin and ChemR23 in peripheral blood of healthy people and PD patientsThe levels of Chemerin and ChemR23 in peripheral blood of PD patients were higher than those of healthy subjects.Conclusion:Chemerin deficiency has a protective effect on MPTP-induced DA neurons in PD model,which has been preliminarily verified in clinical samples.On the one hand,Chemerin acts through ChemR23 receptor,on the other hand,it may inhibit activation of TLR4/NF-κB signaling pathway,thereby inhibiting neuro-inflammation,inhibit the progress of Parkinson’s disease.