Effects Of Inhibiting PGC-1? Expression On Inflammatory Response In Mice Model Of PD

Parkinson's disease(PD)is the second most common neurological degenerative disease following Alzheimer's disease.It is also a neurological degenerative disease with a higher incidence in the elderly.The main features of PD are manifestations of slowness,stiffness,tremor and postural instability.These symptoms are already a classic description of PD,but recently it has been recognized as a more complex disease,including motor symptoms and non-motor symptoms(NMS),such as depression,sleep disorders,paresthesia,autonomic dysfunction and the decline in cognitive ability.Recent studies have found that close factors with PD are mainly mitochondrial dysfunction,oxidative stress,?-synuclein gene mutation and other genetic factors,environmental factors,inflammatory response and so on,but its specific pathogenesis is not entirely clear.Among the above factors,mitochondrial dysfunction and oxidative stress are the main factors.Mitochondria are organelles that provide direct energy in the individual of life,and it is play a very important role in maintaining the normal function of neurons.Mitochondrial complex?is the largest complex in the mitochondrial respiratory chain,which is the key to mitochondrial oxidative phosphorylation.Abnormal expression of mitochondrial complex I can cause more and more reactive oxygen species generation,which can cause cell damaged and death.In addition,the glial cell induced inflammatory response to Parkinson's disease has gradually become a hot topic.Under normal circumstances,glial cells are in resting state.When the neuronsare damaged leading to the release of immunogenic molecules,the glial cells exhibit morphological changes and show the corresponding activation phenotype.The most prominent feature of neuroinflammation is the activation and proliferation of microglia and astrocytes.Glial cells activation can produce a large number of harmful ROS and RNS,at the same time,they can also secrete a large number of proinflammatory cytokines such as TNF-?,IL-1?,IL-6 and IFN-?,which combined with neuronal receptors and then started the cell death pathways.In recent years,a variety of physiological function of the peroxisome proliferator-activated receptor ?(PPAR?)coactivator 1?(PGC-1?)is payed more and more attention.PGC-1? plays an important role in the biological and metabolic processing of mitochondria.It has been found that PGC-1? may inhibit the inflammatory response by regulating the activation and proliferation of glial cells.This progress is regulated by the NF-?B signaling pathway,thus protecting the neurons.But what is the role of PGC-1? in the PD process? What is the relationship between changes in PGC-1? expression and inflammation and PD? This is a question worthy of further studing.In this study,we reduced the expression of PGC-1? and observed the relationship between its expression and inflammatory response.Does it protect neurons? Whether it can improve the symptoms of PD mice and so on.In this study,we used C57BL/6 mice as the object,we injected the lentiviral vector of inhibiting PGC-1? expression into the substantia nigra of mice by stereotactic injection.After injection 28 days,MPTP was injected intraperitoneally.We reproduce the subacute model of PD mice by this method.The experiment was divided into four groups: blank control group(stereotaxiced with saline,intraperitoneal injectioned of saline,recorded as saline+saline group),experimental control group(stereotacticed with saline,intraperitoneal injectioned of MPTP 30 mg/kg,recorded as MPTP+saline group),experimental empty group(stereotaxiced with lentiviral vector NCKD,intraperitoneal injectioned of MPTP,recorded as MPTP+NCKD group)and the experimental group(stereotaxiced with lentiviral vector,which could inhibit the expression of PGC-1?,intraperitoneal injectioned of MPTP,recorded as MPTP+KD).We used the behavioral methods to detect the movement symptoms of PD.Immunofluorescence staining was used to detect the positive cells such as TH and PGC-1? in substantia nigra and the number of microglia markers Iba1 and the IOD of astrocyte marker GFAP;FJC staining method was used to detect the number of apoptotic cells;Western blot was used to detect the protein expression of TH and PGC-1? in substantia nigra,and to investigate the effect of PGC-1? on the behavior of PD mice and the proliferation of microglia and astrocytes associated with.Thus,revealing the relationship between PGC-1?,inflammation and PD.We have got the following experimental results:1.We successfully replicated the PD mouse model by intraperitoneal injection of MPTP(30 mg/kg).With the increase of MPTP injection days,the number of viable dopaminergic neurons and PGC-1? positive cells decreased gradually,and the number of apoptotic cells increased gradually;TH protein and PGC-1? protein expression was less and less;inflammatory reaction was getting heavier and heavier,we finally determined the MPTP damaged optimal time window were 3 days.2.We successfully constructed a lentiviral vector that inhibits PGC-1? expression.We used the qPCR method to detect the best interference target,and the optimal titer of the virus was 5×108 TU/ml by fluorescence.3.The number of dopaminergic neurons in MPTP+KD group was significantly lower than that in MPTP+saline group(P<0.05);The number of autonomic activities and standing in MPTP+KD group was lower than that in MPTP+saline group,but the number of drop from the rotarod and swimming pause were increasing compared with MPTP+saline group,but there was no significant difference.4.The number of PGC-1? positive cells in MPTP+KD group was significantly lower than that in MPTP+saline group(P<0.05),which indicated that lentiviral vector that inhibited PGC-1? expression by stereotactic injection played a role in reducing PGC-1? expression.5.The expression of microglia and astrocytes in MPTP+KD group was significantly lower than that in MPTP+saline group(P<0.05).This suggested that the inflammatory response is reduced.Through the analysis of the above results,we have got the following conclusions:1.We identified the MPTP damaged optimal time window were 3 days by inflammation and survival and apoptosis of DA neurons.2.We screened the lentiviral vector that inhibited the expression of PGC-1? with a virus titer of 5 × 108 TU/ml.3.If the expression of PGC-1? is inhibited,the number of dopaminergic neurons in substantia nigra will be significantly reduced,the number of apoptotic cells will increase,and the activation of microglia and astrocytes will be reduced.The above results indicate that inhibition of PGC-1? expression can reduce the inflammatory response by affecting the activity of microglia and astrocytes.This study provides a basis information for the prevention and treatment of Parkinson's disease,but the relevant mechanism remains to be further studied.