Prelimilary Investigation Of Reconstruction Of Innervated Tissue-engineered Cardiac Sheet In Vitro

Tissue engineering is one of the fast developing subjects aims to reconstruct, repair or replace the damaged tissues or organs. Heart tissue engineering is the hot spot of this field. In recent years, heart tissue engineering was developing quickly, but to guarantee the survival and function of tissue-engineered cardiac tissue after transplantation, the sufficient blood supply and neural innervation were the most important keys. Until now, there have feasible protocols to make the vascularized tissue-engineered cardiac tissues, but there was no report about the innervated tissue-engineered cardiac tissue because heart innervation was very complex and there was no model for the study of heart innervation in vitro. This research was the first study that focuses on the reconstruction of innervated tissue-engineered cardiac sheet in vitro.There are three parts in this research. Firstly, we co-cultured the NGF induced sympathetic neuron-like PC12 cells and the neonatal rat primary cardiacmyocytes to investigate the possibility of neuronal PC12 cells been used as models to investigate the innervation of tissue-engineered cardiac tissue. In the second part, we explored a feasibility method to construct tissue-engineered cardiac sheet using liquid collagen as scaffold. In the third part, we using constructed tissue-engineered cardiac sheet uing neonatal rat primary cardiacmyocytes plus type I collagen gel and at the same time supplemented PC12 cells in the sheet. After that, NGF was used to induce the neuronal differentiation of PC12 cells in the cardiac sheet. After that, histological an TEM observation were performed to investigate the possible innervation of tissue engineered cardiac sheet.The resuts showed that in the 2-D co-culture model, NGF induced PC12 cells outgrew processes which reached the surface of the cardiacmyocytes and moved along with the beat of cardiomyocytes. Their varicosities also touched with cariamyocytes. The tissue engineered cardiac sheet we reconstructed using newborn cardiomyocytes and type I collagen possessed the structure and character of native heart muscle. In the 3-D co-culture model, PC12 cells could be induced by NGF to outgrow processeds which also reached the surface of cardiacmyocytes, and formed synapt -like strctures. The results of our study suggested that PC12 cells is a feasible model to be used in the study of neural innervation of tissue engineered cardiac muscle. And the cardiomyocytes may be innervated in this model.The contents are as follows:Part 1: co-culture of the NGF induced sympathetic neuron-like PC12 cells and the neonatal rat primary cardiacmyocytesPart 2: Reconstruction of tissue- engineered cardiac sheet in vitroPart 3: Initial study of reconstruction of innervated tissue-engineered cardiac sheet in vitro...