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A Study of Dose Effect of Human Bone Mesenchymal Stem Cells on Bone Tissue Engineering

Author: WuHuanHuan
Tutor: CaoYiLin；XiaoZuo；WangQian；LiuXia；YanZuo
School: Beijing Union Medical College
Course: Surgery
Keywords: Bone marrow mesenchymal stem cells Osteogenic induction Tissue Engineering Building
CLC: R329
Type: Master's thesis
Year: 2011
Downloads: 35
Quote: 0
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Abstract


Objective: Bone marrow charge quality stem cells (human bone mesenchymal stem cells, BMSCs) tissue engineered bone is currently the most commonly used seed cells, of BMSCs construct tissue engineering bone (tissue engineering bone, TEB) the dose-effect relationship is TEB clinical application of important problem, but has yet to see clearly reported. This study hBMSCs in vitro into bone induction at different time points of osteoblast-related gene expression and body rumor-generation hBMSCs dry cell surface flag expression, proliferation and differentiation, and by different concentrations hBMSCs build TEB the in vitro observation and different number of hBMSCs build TEB of ectopic bone formation in vivo studies, the dose-effect relationship of hBMSCs build TEB were discussed. Method: 1, density gradient centrifugation hBMSCs, the application of flow cytometry, osteogenic and adipogenic differentiation of stem cells identified. 2 of hBMSCs osteogenic differentiation in vitro into bone induction at different time points in vitro osteoblast gene expression by RT-PCR method to detect. 3 hBMSCs were cultured in vitro generation of application of flow cytometry, MTT assay and cell counting detection a different algebraic hBMSCs stem cell surface markers and proliferative capacity; applications Real-time PCR method and oil red 0 staining algebra hBMSCs The osteogenic and adipogenic differentiation capacity. 4 hBMSCs inoculation of different concentrations of β-TCP scaffold, using scanning electron microscope (SEM), histological staining (HE) and Real-time PCR method to detect different concentrations hBMSCs β-TCP scaffold on the growth and osteogenic differentiation. 5, the same concentration of different volume the hBMSCs inoculated β-TCP scaffold building group TEB and implanted subcutaneously into nude mice, application histology detected in vivo ectopic bone formation. Results: 1 biological characteristics of hBMSCs: density gradient centrifugation, adherent screening combined digestion control method isolated hBMSCs express stem cell surface markers: CD90, CD29, CD166, CD44, CD73, CD1O5 differentiate into bone cells in vitro. and fat cells. 2, hBMSCs in vitro osteoblast induction: osteogenesis-related genes in the induction of the expression of the early part of the mid-term expression, most of the gene expression reached a peak at 14 days, mineralization-related gene expression reached a peak at 21 days. 3, hBMSCs body rumor cultured: P4 on behalf of future of hBMSCs the proliferation capacity, the adipogenic and adipogenic differentiation diminished capacity. 4, hBMSCs build TEB vitro studies: high concentration (10 × 106 and 20 × 106cell/ml) hBMSCs were better than in the β-TCP scaffold growth and matrix secretion of low concentration (1 × 106 and 5 × 106cell/ml ). Different concentration groups, no significant differences in level of osteogenic differentiation. 5 hBMSCs were constructed TEB in vivo studies: hBMSCs inoculated with the same concentration of β-TCP stent constructed TEB, inoculation volume increase, and increase the success rate of osteogenesis in vivo. Conclusion: density gradient centrifugation, adherent screening combined digestion control method can be successfully isolated hBMSCs were. hBMSCs in vitro into osteoblasts in bone induction process related gene expression is dynamic, its expression timing and physiological development of osteoblasts; the hBMSCs proliferation and differentiation before Passage 5 (P5) on behalf of a strong, suitable for clinical use; 10 × 106 and 20 × 106cell/ml suitable concentration is the hBMSCs inoculated β-TCP stent; of the TEB vivo osteogenic success rate increased with increasing number of inoculation with hBMSCs. The hBMSCs construct tissue engineered bone dose parameters to provide a reference for further research and clinical application of tissue engineered bone.

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CLC: > Medicine, health > Basic Medical > Human morphology > Human histology
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