Biological characteristics of canine bone marrow mesenchymal stem cells in vitro under non-inducing conditions
- VernacularTitle:非诱导条件下犬骨髓基质干细胞的生物特性
- Author:
Jiang CHEN
;
Jin YANG
;
Fuhua YAN
;
Xin ZHAO
- Publication Type:Journal Article
- From:
Chinese Journal of Tissue Engineering Research
2005;9(46):135-137
- CountryChina
- Language:Chinese
-
Abstract:
BACKGROUND: Research on seed cells is the most important aspect in the filed of tissue-engineering research, and because of their various ad vantages, bone marrow mesenchymal stem cells (BMSCs) have been taken as the ideal bone tissue-engineering seed cells. OBJECTIVE: To observes the growth characteristics of in vitro cultured BMSCs and their osteogenetic characteristics under non-inducing conditions. DESIGN: A single sample experiment.SETTING: Implanting Center of Stomatological Hospital of Fujian Medical University MATERIALS: This experiment was conducted at the Central Laboratory of Stomatological Hospital of Fujian Medical University, between May andDecember 2003. BMSCs are derived from the primary passage to the 3rd passage of in vitro cultured cells from 4 one-year old dogs. METHODS: Under the aseptic conditions, bone marrow puncture was made at bilateral femur trochanter and 5 mL of heparin anticoagulatedbone marrow was obtained. Then it was placed in 50 mL of anti-Dulbecco's modified Eagle's culture medium in centrifuge tube for monocyte isolation. The BMSCs single cells were primarily isolated and placed in culture box for subculture. After 48 hours, culture medium was removed and the medium was cha1ged once every 3 days. Then subculture was carried on continuously to observe BMSCs in morphology under inverted the phase contrast microscope with the assistance of HE staining. The number of cells was counted daily to calculate doubling time and to draw a growth curve. Alkaline phosphatase was detected by using calcium-cobalt method,and chinalizarin staining was used to detect the growth state of calcification tubercle. MAIN UTCOME MEASURES: ①Optical microscopic structure of dog BMSCs; ② The growth curve of dog BMSCs; ③Observation of osteogenetic index- alkaline phosphatase and calcification tubercle. RESULTS: ① Morphological observation indicated that BMSCs were adherent to the walls, clonogenic and appearing fibroblastic phenotype, and they presented morphological changes without exposing to osteogenetic inducer. ② The expression of Alkaline phosphatase in primary cells was stronger, and it was strong in the 1st passage cell, but weak in the 2nd and 3rd passage cells. ③Calcium deposition was observed, which was stronger in primary cells than in subcultured cells. CONCLUSION: ①BMSCs massively proliferated during in vitro culture,capable of differentiating into osteoblasts and considered as the optimal seed cells for bone tissue-engineering reconstruction. ② BMSCs derived from the 3rd passage has osteogenic activity, but the osteogenic activity of the primary cultured cells was stronger than thatr of the subcultured cells.
