Differentiation Of The Mouse Induced Pluripotent Stem Cells Into Olfactory Receptor Neuron Cells And Mitral/Tufted Cells In Vitro

Background:although the olfactory nerve has the ability to regenerate,when the damage involves nerve,only a small part of patients can restore the olfactory function.For sensorineural dysosmia,there is no satisfactory treatment.i PSCs can not only be obtained from somatic cell reprogramming,but also are extremely similar to embryonic stem cells(ESCs)and can be differentiated into all kinds of cells without immune rejection or ethical issues.In this study,we will investigate whether mouse induced pluripotent stem cells(i PSCs)can be differentiated into olfactory receptor neurons or mitral/tufted cells,which may provide an experimental basis for the treatment of sensorineural olfactory disorder through cell transplantation.Objective:This study was to investigate the differentiation potential of i PSCs into olfactory receptor neurons and mitral/tufted cells in vitro.Methods:1.Mouse i PSCs culture and pluripotency gene identification:we extracted mouse embryonic fibroblast and prepared feeding layer,where mouse i PSCs were inoculated on;RT-PCR were used to identify i PSCs pluripotency genes Oct4,Nanog,Sox2.2.Co-culturing i PSCs with olfactory epithelium cells and immunological identification:we separated the olfactory epithelium from mice,which contained olfactory receptor neurons,co-cultured i PSCs with olfactory epithelium cell and then identified differentiated cells with the olfactory receptor neurons makers(OMP,GAP43,NCAM)after 14 days co-culturing by immunofluorescence and RT-PCR.3.Co-culturing i PSCs with olfactory bulb cells and immunological identification:we separated the olfactory bulb from mice,which contained mitral/tufted cells,co-cultured i PSCs with olfactory bulb cells and then identified differentiated cells with mitral/tufted cells markers(TBX21,Iba1)after 14 days co-culturing by immunofluorescence and RT-PCRResults:1.we successfully established a stable culture system of mouse i PSCs and RT-PCR showed that pluripotency genes(Oct4,Nanog,Sox2)were expressed in mouse i PSCs2.Immunocytochemical analysis or RT-PCR results indicated that the differentiated i PSCs can expressed olfactory receptor neurons markers(OMP,GAP43,NCAM)and mitral/tufted cells markers(TBX21,Iba1)after being co-culture with olfactory epithelium or olfactory bulb.Conclusions:Our results suggest that mouse i PSC possess the potency to differentiate into olfactory receptor neuron-like cells and mitral/tufted-like cells.