Myocardial Patch Formation By Three-dimensional 3-hydroxybutyrate-co-4-hydroxybutyrate Cultured With Mouse Embryonic Stem Cells

Objective: Comparatively studying of using the vitamin C as inducer and without vitamin C to promote the mouse embryonic stem cells(ESCs)and poly3-hydroxybutyrate-co-4-hydroxybutyrate[P(3HB-co-4HB)] Co-culture differentiation into cardiac patch.Observed myocardial cell in the growth and surface attachment,Analysis and calculation of the two groups of induced differentiation efficiency curve,get Vitamin C is in the promotion of differentiation of embryonic stem cells and P(3HB-co-4HB)when co-cultured into cardiac patch,this three-dimensional polymer material is suitable for myocardial cell growth and proliferation.P(3HB-co-4HB)has the potential for development as a clinical alternative to current treatment methods for the regeneration of cardiomyocytes in patients with myocardial infarction.Methods:Get embryonic fibrous cells(MEF)in primary culture,proliferation and treatment;recovery,cultured and passaged mouse embryonic stem cells(ESCs);Preparation of mouse embryonic stem cells into Embryonic bodies(EBs);product cell patch;DAPI fluorescence after the dyeing process observed fluorescence labeling cell patch;confocal microscopy [Confocal Laser Scanning Microscope(CLSM or LSCM)],through moving the lens system observe EBs growth on the P(3HB-co-4HB)cells patch three-dimensional scanning,EBs observed on cell growth situation patch;Staining the specific marker cTnT and ?-actin of cardiomyocytes by immoluno fluorescence;a scanning electron microscope to observe myocardial patch.From the7 th day of EB and the patch co-culture,every two days we will observe and count the cardiac myocytes.Mapping the distribution of the differentiation efficiency with the induction time.Results: MEF cells and ESCs adherent in a petri dish growth well,normal morphology;cardiomyocytes were positive to cTnT and ?-actin;staining scanningelectron microscope,P(3HB-co-4HB)materials and cardiomyocytes co-cultured into cardiomyocytes form,we can see a lot of cells on the surface,normal cell morphology,combined with three-dimensional film closely.Draw differentiation efficiency with induction time of distribution,display differentiation efficiency over time gradually increased differentiation vitamin group was significantly higher than the group without the addition of vitamin C,no vitamin C group had the highest induction rate of 17.8%,plus vitamin C group was the highest-induced rate of 71.1%.Conclusions: Polymer biomaterial patches composed of P(3HB-co-4HB)permit cardiomyocytes growth,proliferation and differentiation into cardiac.Add vitamin C can significantly increase the probability of induced into cardiomyocytes.P(3HB-co-4HB)has the potential for development as one of a clinical alternative to current treatment methods for the regeneration of cardiomyocytes in patients with myocardial infarction.