Development of the osteoblast phenotype of serial cell subcultures from human bone marrow

Abstract

Bone marrow cells have been used for testing biocompatibility of bone substitute materials that would be applied in maxillofacial and orthopedic surgeries. However, it remains unclear whether cells in serial subcultures retain the ability to differentiate into osteoblasts. The purpose of this study was to compare the development of osteoblast phenotype of serially passaged cells from human bone marrow. Cells from first to third passage were cultured (2x10 4 cells/well) in supplemented culture medium. Cells were incubated at 37ºC in a humidified atmosphere of 5% CO 2 and 95% air. Cell attachment was assessed at 4 and 24 h. At 7, 14 and 21 days, cell proliferation, cell viability, total protein content and alkaline phosphatase (ALP) activity were evaluated. Bone-like formation was evaluated at 14 and 21 days. Data were compared by two-way ANOVA and Duncan's multiple range test. Cell attachment, cell viability and total protein content were not affected by serial subcultures. However, serial subcultures did interfered negatively with osteoblast differentiation as shown by osteoblast parameters observed in second and third subcultures, such as continuous cell proliferation, lower ALP activity and bone-like formation in comparison to first subculture. Therefore, it is important to evaluate cell ability to growth and differentiate before selecting the cell population for studies that investigate the biocompatibility of materials to replace bone tissue.

The Characteristics Of Human Bone-Derived Cells (HBDCS) during osteogenesis in vitro

“…The time course of ALP gene expression roughly fits with the increase in ALP activity as shown using an osteosarcoma cell line [29]. The increase in ALP activity correlates with the formation of bone-like structures in vitro [30]. Our study showed that ALP activity was detectable 24 h after cell seeding and increased during human osteoblast maturation.…”

Bone cell responses to the composite of Ricinus communis polyurethane and alkaline phosphatase

“…That fewer cells were counted for RCP+ALP/SBF at days 7 and 14 likely indicates that such biomaterial could favor cell differentiation, since ALP activity was enhanced on its surface. Corroborating these findings, a positive correlation between the decrease in cell proliferation and the subsequent induction of cell differentiation in both rat calvarial and alveolar human bone cell cultures was described elsewhere 33, 34. With exception of RCP+ALP that was cytotoxic, in the present work cell viability was around 90% for all other groups, comparable to that observed for tissue culture polystyrene (data not shown).…”

Effect of surface roughness of ground titanium on initial cell adhesion

“…ter Brugge et al [12] found that the surface roughness (R a : 0.60-1.64 m) of Ti can affect the initial interaction of U-2 OS cells with the substrate. However, contrary results have been reported by other researchers [13][14][15]. The experimen- tal study in dogs indicated that the soft tissue attachment on commercial Ti dental implants is not influenced by the surface roughness [13].…”

“…The experimen- tal study in dogs indicated that the soft tissue attachment on commercial Ti dental implants is not influenced by the surface roughness [13]. Rosa and Beloti [14] claimed that the initial attachment of human bone marrow cell is not affected by surface roughness (R a : 0.80-1.90 m) of Ti after 4 h cell culture. They also found that the surface roughness (R a : 0.24-1.92 m) of Ti does not affect significantly the initial attachment of rat bone marrow cell after 2 h incubation [15].…”