The Study Of Adult Degeneration Disc Seed Cells Culture In Intervertebral Disc Tissue Engineering

Objective:To erect the three-dimension culture model of adult degenerative intervertebral disc and find a way of resolving the problem of selecting high-quailty seed cells in tissue engineering throught the determination of morphology, histology , cell biology methed and detecting the extracellular matrix of intervertebral disc cells.Methods:1. Intervertebral disc tissue were obtained from fetus and patients operated and cultivanted as primary cell with monolayer culture and three-dimension culture respectively.2. Growth of degeneration intervertebral disc cells with different groups were detected in morphology and histology by inverted phase contrast microscope; selecting cells and ultramicrocuting, detecting the cell ultra-microstructure with transmission electron microscope; staining cells with Giemsa and HE methed, comparing the different of microscope under the light microscope.3. Detecting the cytobiology index of intervertebral disc cells between three-dimension culture and monolayer culture. Containing cell growth curve, cell inoculation survival rate, proliferating cell nuclear antigen detection, cell growth activity, cell cycle, apoptosis et al. and comparing the effects on cell growth between three-dimension culture and monolayer culture.4. Extra-cellular matrix of degeneration intervertebral disc cells with different groups were detected, Quantitatly detecting the content of different collagen and proteoglycan in intervertebral disc cells by western blot and 35S-sulfate labeling, respectively.Results:1.successfully erect the model of adult intervertebral disc cells with tissue digesting method and on this basis construct the three-dimension culture with microcarrier stiring cultivation system.2. Comparing foetus and adult cataplasia intervertebral disc cells in monolayer culture and three-dimension culture, measuring cell growth curve, cell inoculation...