OBJECTIVE: To investigate the feasibility of inducing in vitro mesenchymal stem cells (MSCs) derived from adult human bone marrow differentiate into osteoblasts and potential applicability of the MSCs as the seed cells in tissue engineering. METHODS: Adult human bone marrow was collected from the healthy adult volunteers to obtain the MSCs, which, after in vitro culture in DMEM supplemented with 10% fetal bovine serum and incubation under standard condition, were induced to differentiate into osteoblasts in DMEM containing dexamethasone (1x10(-8) mol/L), beta-sodium glycerophosphate (10 mmol/L) and ascorbic acid (50 mg L). Proliferation and differentiation of the MSCs were observed continually under inverted phase-contrast microscope and transmission electron microscope. The collagen typeI was detected by immunohistochemistry, alkaline phosphatase (AP) in the MSCs stained by Gomori, the calcified nodules were stained by von Kossa method, and the changes in the content of AP were measured. RESULTS: The MSCs proliferated rapidly in in vitro culture and after a 2- to 3-week induction, the cells began to generate large amount of enlarged endoplasmic reticulum, Golgi complexes and mitochondria, with immature cell nuclei. Positive staining for collagen typeIand strong reaction for AP and calcified nodules were observed. Increasing AP secretion by the MSCs was seen as the time of induction prolonged (P<0.01). CONCLUSIONS: Human bone marrow-derived MSCs can be induced to differentiate into osteoblasts through relatively simple procedures, which provide ideal autogenous source of seed cells for bone tissue engineering. The method adopted in this experiment may be used for routine culture of the seed cells for bone tissue engineering.
OBJECTIVE: To investigate the feasibility of inducing in vitro mesenchymal stem cells (MSCs) derived from adult human bone marrow differentiate into osteoblasts and potential applicability of the MSCs as the seed cells in tissue engineering. METHODS: Adult human bone marrow was collected from the healthy adult volunteers to obtain the MSCs, which, after in vitro culture in DMEM supplemented with 10% fetal bovine serum and incubation under standard condition, were induced to differentiate into osteoblasts in DMEM containing dexamethasone (1x10(-8) mol/L), beta-sodium glycerophosphate (10 mmol/L) and ascorbic acid (50 mg L). Proliferation and differentiation of the MSCs were observed continually under inverted phase-contrast microscope and transmission electron microscope. The collagen typeI was detected by immunohistochemistry, alkaline phosphatase (AP) in the MSCs stained by Gomori, the calcified nodules were stained by von Kossa method, and the changes in the content of AP were measured. RESULTS: The MSCs proliferated rapidly in in vitro culture and after a 2- to 3-week induction, the cells began to generate large amount of enlarged endoplasmic reticulum, Golgi complexes and mitochondria, with immature cell nuclei. Positive staining for collagen typeIand strong reaction for AP and calcified nodules were observed. Increasing AP secretion by the MSCs was seen as the time of induction prolonged (P<0.01). CONCLUSIONS:Human bone marrow-derived MSCs can be induced to differentiate into osteoblasts through relatively simple procedures, which provide ideal autogenous source of seed cells for bone tissue engineering. The method adopted in this experiment may be used for routine culture of the seed cells for bone tissue engineering.