Protective Effect Of Secretory Factors Of Mesenchymal Stem Cells To Aβ₁₄₀ Induced Apoptosis In PC12 Cells

PrefaceAlzheimer's disease,AD is a kind of chronic progress nervous system recessive disease.The main pathologic characteristic is the cholinergic neuronal recession in substantia nigra striatum,and appear senile plaque(SP),neu-rofibrillary tangle(NFT) and blood vessel amyloidosis following neuronal losing in cerebral cortex and hippocampus.Aβis a kind of protein of AD'patient abnormal releasing and sedimentation;Aβplay a important part in neuronal apoptosis. Excessive neuronal apoptosis may be one of important reason on AD neuronal degeneration.The Recently studydiseovered,Bone marrow stromal cells(BMSCs),or be called mesenchyma lstem cells(MSCs) have stem-cell character and the potential for multidirectional differentiation,MSCs is easy to get,develop and propagate in vitro. Autotransplanting avoids immune rejecting reaction too.As a result, mesenehyma stem cells have vast of applied foreground as the cytology treatment foundation used for the central nervous system disease and its trauma repair.The pheochromocytoma(PC12) cells are the cells of addicted chrome cell tumors for the big rat adrenal glands, its receivers and transmitters are similar to cholinergic neuron,they can illuminate consistent problem with original generation nerve cell, so PC12 cell with Aβ_1-40 model is often the cell model by way of studying AD.The MSCs can secrete cell factors considered to be connected with they having the protective function to the cholinergic neuron.Their conditional culturable liquid was cultured,Pured and collected in this trial,to investige the effect of the mesenehymal sterne cells(MSCs) secretary factors to Aβ_1-40 induced aoptosis and cell vitality in PC12 cells,preliminarily probe into MSCs secretion having protective function to cholinergic neuron.Methods1.The collections and the inspissation of the original MSCs supernatant2.MSCs surface antigen measurement(immunoeytochemistry) The first antibody is theCD44,CD45 and the second antibody is CY3.3.Culture and medical management of PC12 cells.PC12 were disparted into 4group①control group:it is not joined any medicine in the cell cultul-esystem:②Simple supematant of MSCs group:it is joined the supematant of MSCs in the cell culture system after inoculating 24h,the concentration is 30μl,60μl and 120μl;③Aβ_1-40 group,it is joined the Aβ_1-40(10μg/ml) in the cell culture system after inoculating 24h and eventual concentration 5μg/ml, 10μg/ml,20μg/ml respectively;④unite group:it is joined the 30μl, 60μl and 120μl supematant of MSCs after inoculating 24h following joining the 10μg/ml Aβ_1-40,Examination of the following index after joined Aβ_1-40 24h and 48h.4.The measurement of cell vitality(MTT)5.The morphological changes of apoptosis were observed under transmitting electronmicroscopy(TEM)6.Apoptosis ratio was analyzed by PI dyeing Flow Cytometric(FCM)7.Statistical analysis:All the results were transofmred to dates thorugh image analysis system,which were express as X±SD.the values were analyzed using ANOVA and t test of spss sowtfare.A value of P＜0.05 was considered significant,obvious significance was set at P＜0.01.Results1.MSCs may separate and proliferate in vitro,were positive for CD44 and Negative for CD45.2.The results of MTT showed the secretary factors of MSCs can advance the cell viability and resist the toxic function of Aβ_1-40 to PC12 cells.(1)Aβ_1-40 can induce apoptosis in PC12 cells,The more Aβ_1-40 concentration was,the more the toxicity was,the neighbor concentration of the difference have notable significance(P＜0.05);The more the time acted,the more the toxicity was to PC12 cells in the same concentration(p＜0.05).(2)MSCs can advance the cell viability and resist the toxic function of Aβ_1-40 to PC12 cells;The more the concentration was,the more the cell viability was,the neighbor concentration of the difference have notable significance(p＜0.05).3.TEM can observe the apoptosis of the PC12 cells.4.The results of FCM showed that secretary factors of the MSCs can decrease the ratio of apoptosis of the PC12 cells treated with Aβ_1-40.(1) The ratio of apoptosis of the PC12 cells increased in file along with the increase of concentration of secretary factors of Aβ_1-40 (P＜0.05).(2) the MSCs can decrease the ratio of apoptosis of the PC12 cells treated with Aβ_1-40;The ratio of apoptosis of the PC12 cells of different concentration of secretary factors of unite group have notable difference,the neighbor concentration of the difference have notable significacen(P＜0.05)ConclusionMSCs may rapidly proliferate in vitro,were positive for CD44 and negative for CD45.Aβ_1-40 can induce apoptosis in PC12 cells,MSCs have potencial in the secretion neurotrophicfactors of GDNF,BDNF and so on to benefit for recessive cholinergic neuron,so MSCs may exert a protective effect against Aβ_1-40 induced apoptosis in PC12 cells.Its strong and weak was connected with different concentrations. The research provided definitely experimental basis to AD's treatmemt.