Establishment of human osteoblastic cells derived from periosteum in culture

Abstract

We isolated osteoblastic cells derived from human periosteum and established them in culture. Their growth depended on the presence of ascorbic acid, and the doubling time was 40 to 60 h. The requirement for ascorbic acid was used to high production of collagen. These cells produced mainly type I collagen and only small amounts of type III collagen determined by reducing sodium dodecyl sulfate SDS-polyacrylamide gel electrophoresis. The total collagen yield was about 10 mg from 2 X 10(7) cells. The cells could be continuously cultured in alpha-minimum essential medium supplemented with 10% fetal bovine serum for 18 to 40 population doubling levels, depending on the age of the donated periosteum. These cells have the ability to calcify when incubated with 2 mM alpha-glycerophosphate-Na2. Calcification as viewed by the naked eye appeared from Day 15 after treatment. Treatment with the active formed vitamin D3, 1, 25 dihydroxyvitamin D3 enhanced calcification significantly and stimulated osteocalcin production. By electron microscopy, cells with many projections on their surfaces showed well-developed rough endoplasmic reticulum and actinlike fibers, and larger numbers of lysosomes, mitochondria, and secretion granules. Many matrix vesicles, in which minerals were initially localized, and well-banded collagen fibrils were seen in the intercellular spaces. These observations demonstrate typical osteoblastic morphology. The above results indicate that cultured cells from human periostem are osteoblastic cells that have the capacity to differentiate into osteocytes and to deposit calcified minerals in response to 1,25 dihydroxyvitamin D3.

Osteoblastic differentiation of periosteum‐derived cells is promoted by the physical contact with the bone matrix in vivo

“…Periosteal cells have been shown to possess the osteogenic potential in vitro (Koshihara et al, 1989;Tenenbaum et al, 1986) and in vivo (Breitbart et al, 1998;Nakahara et al, 1990b). The phenotype of periosteal cells, however, may be altered through being removed from the original site, dissected for cell culture, or suffering from surroundings that are different from in situ.…”

“…Cultured cells derived from the periosteum exhibit osteoblastic differentiation in the presence of ascorbic acid and 1,25 dihydroxy vitamin D3 in vitro (Fang and Hall, 1996;Koshihara et al, 1989;Tenenbaum et al, 1986). Periosteal explants or cells derived from the periosteum loaded in diffusion chambers or combined with porous ceramics showed the osteogenic potential in implantation experiments in vivo (Breitbart et al, 1998;Gally et al, 1994;Jaroma and Ritsila, 1988;Nakahara et al, 1990aNakahara et al, , 1991.…”

Antisense Suppression of Collagen VI Synthesis Results in Reduced Expression of Collagen I in Normal Human Osteoblast-like Cells

“…7,19) More than 90z of the collagen they produced was type I. 19) SaM-1 cells were cultured with or without 3.0 ng W ml rhIL-4 (kindly supplied by Ono Pharmaceutical, Japan). Transfection with an antisense oligonucleotide (ASODN) for the a1(VI) collagen chain was done as follows.…”

1, 25-Dihydroxyvitamin D₃, Recombinant Human Transforming Growth Factor-β₁, and Recombinant Human Bone Morphogenetic Protein-2 Induce In Vitro Differentiation of Canine Osteosarcoma Cells

“…With respect to ALP activity, the effects of 1,25(OH)2D3 depend on cell type, the time of addition to the culture, and the stage of the cellular maturation cycle [2,12,13,15,16,20,26,30]. In this study, ALP activities of all clonal cells except POS14A were decreased by the addition of 1,25(OH)2D3 to the medium.…”