Biochemical Characterization with Parathormone and Calcitonin of Isolated Bone Cells: Provisional Identification of Osteoclasts and Osteoblasts

Abstract

Two metabolically distinct types of bone cell populations were isolated from mouse calvaria by a repetitive digestive procedure with a mixture of collagenase and trypsin. Cells released early in the digestion showed approximately two-fold increases in cAMP when treated with either parathormone or calcitonin. These populations were denoted CT type. Later eluting cells showed larger parathormone-induced increases in cAMP but did not respond to calcitonin. These populations were denoted PT type. Six metabolic and enzymatic activites were measured in the two types of populations: acid and alkaline phosphatases, hyaluronate synthesis, citrate decarboxylation, prolyl hydroxylase, and general protein synthesis. Although each of these activites was present in both cell types, the basal levels of acid phosphatase and hyaluronate synthesis were higher in the CT cells, whereas alkaline phosphatase, citrate decarboxylation, and prolyl hydroxylase were higher in te PT cells. Parathormone stimulated acid phosphatase and hyaluronate synthesis by 100-200% only in the CT cells; in inhibited alkaline phosphatase, citrate decarboxylation, and prolyl hydroxylase by 75-90% only in the PT cells. Calcitonin alone had no effect on any of these activities other than cAMP production, but in inhibited the action of parathormone in the CT cells. The sensitivities, time courses of development,and magnitudes of these hormonal effects were similar to those observed previously in intact calvaria, indicating that the isolated cell system is a reliable model for the study of bone metabolism. Based on the metabolic responses of the cells, we postulate that the CT type of populations is enriched in osteoclasts and, possibly, osteocytes, and the PT type of population is enriched in osteoblasts.

Bone formation and calcification by isolated osteoblastlike cells.

“…This means that, although osteoblasts do not appear to come along when the periosteum is stripped of the calvarium, the periosteum must contain preosteoblasts or osteoprogenitor cells that can differentiate into osteoblasts (15) [PTHI in log units/ml FIGURE might be found in the relative short culture time we used for the PF transplants on the quail chorio-alloantoic membrane or in the lack of an inducer needed to stimulate PF cells to differentiate into osteoprogenitor cells and eventually into osteoblasts . If this were the case, OB cells could be considered to be "determined osteogenic cells" (6) in the sense that the cells are in a "competent state of readiness" (11) to form bone if the milieu is favorable. PF cells may be designated as periosteal fibroblastlike cells and possibly inducible osteogenic precursor cells (6).…”

Infection-induced Up-regulation of the Costimulatory Molecule 4-1BB in Osteoblastic Cells and Its Inhibitory Effect on M-CSF/RANKL-induced in Vitro Osteoclastogenesis

“…Cell Cultures-Mouse osteoblasts were obtained from newborn mouse calvaria (24). MC3T3-E1 and RAW264.7, NIH3T3, and J774 cells were maintained in ␣-MEM, Dulbecco's modified Eagle's medium, and RPMI 1640 (Invitrogen, Carlsbad, CA), respectively, containing 10% fetal bovine serum (FBS; Invitrogen) and 100 units/ml of penicillin-G at 37°C in a humidified atmosphere of 5% CO 2 in air.…”

Parathyroid Hormone Rapidly Stimulates Hyaluronan Synthesis by Periosteal Osteoblasts in the Tibial Diaphysis of the Growing Rat

“…PTH stimulates the production of HyA in cultures of calvarial explants (18 -21) and in cultures of primary cells isolated from bone tissues (22)(23)(24). HyA has been detected in bone tissues by several investigators, and these studies suggest that the resident osteogenic cells produce HyA (25,26).…”