Human Amniotic Epithelial Cells And Human Amniotic Mesenchymalstem Cells Can Improve Cognitive Function And Reduce Neuro-pathological Injury On Alzheimer’s Disease In Rats: A Comparative Study

Objectives: Alzheimer’s disease (AD) is a kind of neurodegenerative diseases.ADmanifests its clinical symptoms as cognitive dysfunction,memory loss and personalityaltermation.The neuropathology characteristics include deposition of beta-amyloid (Aβ)and neurofibrillary tangles (NFT).At present, there is no effective treatment way. Stemcell transplantation maybe a nove therapeutic approach.Human amniotic epithelial cells(hAECs) and human mesenchymal stem cells (hMSCs) have been shown possess goodneurobiological cbaracteristic and broad immuneregulatory capabilties. So far, whetherhAECs transplantation has therapeutic effect on AD is unkown. In this study, weinvestigated the treatment effect and its biological principle of hAECs and hAMSCstransplantation on AD rat model induced with lipopolysaccharide (LPS), and evaluatedtheir superior or time, in order to provide experimental basis for cellular therapy ofneurodegenerative diseases to select a suitable donor cells..method：①hAMSCs and hAECs were isolated from amnion with digestion method oftwo enzymes，then the phenotypes of hAMSCs and hAECs were identified by flowcytometry (FCM) and immunocytochemistry (ICC).②AD rat model induced bybilateral intraventricular injection of LPS. The model animals were randomly dividedinto four groups: Model group （n=12）, Medium group (n=12), hAECs transplantationgroup (n=12) and hAMSCs transplantation group (n=12).Rats in cell treatment groupswere injected10μl suspension of hAECs or hAMSCs (5x105cell/10ul) into bilateralhippocampal, medium group injected equivalent serum-free L-DMEM medium into thesame way, and set up normal group as control group.All of the rats were killed at twoweeks and four weeks after hAECs and hAMSCs transplantation,and take the brain andperipheral blood to pathology and Immunology examination.③The memory andlearning function all of the rats were assessed respectively before and after thetransplanttation cells and put to death before the rats.④The pathological change in thebrain was observed by HE staining, and beta amyloid and NFT deposition in injuriedtissue were tested by thioflavin S staining.⑤The expressions of Aβ42, Tau, Ach and CD68（microglia mark） in the damage area were detected byimmunohistochemistry,and their mean integral optical density (IOD) was analyzed byImage-ProPlus6.0image analysis system.⑥Percentages of lymphocyte subsets inperipheral blood mononuclear cells (PBMCs) were observed by FCM, containingCD4+IFN-γ+(Th1),CD4+IL-4+(Th2),CD4+Foxp3+(Treg),CD4+IL-17+(Th17) cells⑦IL-2,IL-10, IL-4, TNF-α and IFN-γ in serum were detected by cytometric beadarray.⑧Employed immunofluorescence technique to detect survival anddifferentiationof donor cells at2weeks and4weeks following transplantation.Results：①The third generation of hAECs did not express CD34,HLA-DR,CD45,CD80and CD86, expressed CD44, CD29, CD73,CD49f, CD326, E-cad andCK19; The third generation of hAMSCs did not express CD34, HLA-DR, CD19, CD14,CD45,CD80and CD86, highly expressed CD73, CD29, CD44, CD105, CD90andvimentin.②At two weeks after cells transplantation, compared with model group andmedium group, the escaping latency of all cells transplantation groups began to shortensignificantly from the begining the third day (all P﹤0.01), the number of going throughthe platform increased obviously (all P﹤0.05); At four weeks after treatment,theescaping latency of hAECs transplantation group began to shorten from the begining thefour day (P﹤0.05), while escaping latency and number of going through the platformof hAMSCs group were not statistically significant（P﹥0.05）; Compared withhAMSCs group, the escaping latency of hAECs group obviously decreased (P﹤0.05).③Compared model group and medium group,cell treated animals showed animprovement inflammatory injury in the brain, and amyloid plaques and NFT decreasedsignificantly.④In all cell transplantation groups, the expression of beta-amyloidreduced notablely(P﹤0.01; P﹤0.05), abnormal phosphorylation of Tau proteindecreased remarkablely（P﹤0.01; P﹤0.05）, Ach increased obviously（P﹤0.01; P﹤0.05）, and CD68increased significantly (P﹤0.01; P﹤0.05); these changes in thehAECs group were more obvious,comparing with hAMSCs group（P﹤0.01; P﹤0.05).⑤At two weeks, the percentages of Th1and Th17subsets in PBMCs of hAECstreated rats were decreased（P﹤0.05, while Th2cells raised (P﹤0.05); IFN-γ and IL-2were decreased（P﹤0.05）,while IL-4was increased（P﹤0.05）.⑥The donor cellsimplanted into brain were survived in situ for four weeks and expressed neuron-specific nuclear protein.Conclusions: hAMSCs and hAECs transplantation can improve the cognitive functionand neuropathological damage of AD rats，and its mechanism may relate to reduce Aβdeposition and Tau protein excessive phosphorylation. Taken together these results thathAECs is more valuable for cell therapy of neurodegenerative diseases,comparing withhAMSCs.