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Development and characterization of a conditionally immortalized human fetal osteoblastic cell line
Author(s) -
Harris Steven A.,
Enger Robert J.,
Riggs Lawrence B.,
Spelsberg Thomas C.
Publication year - 1995
Publication title -
journal of bone and mineral research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.882
H-Index - 241
eISSN - 1523-4681
pISSN - 0884-0431
DOI - 10.1002/jbmr.5650100203
Subject(s) - osteonectin , osteoblast , bone sialoprotein , osteocalcin , biology , cell culture , microbiology and biotechnology , osteopontin , medicine , endocrinology , alkaline phosphatase , in vitro , biochemistry , genetics , enzyme
Abstract We report the establishment of a human fetal osteoblast cell line derived from biopsies obtained from a spontaneous miscarriage. Primary cultures isolated from fetal tissue were transfected with a gene coding for a temperature‐sensitive mutant ( ts A58) of SV40 large T antigen along with a gene coding for neomycin (G418) resistance. Individual neomycin resistant colonies were screened for alkaline phosphatase (AP)‐specific staining. The clone with the highest AP level, hFOB 1.19, was examined further for other osteoblast phenotypic markers. Incubation of hFOB cells at the permissive temperature (33.5°C) resulted in rapid cell division, whereas little or no cell division occurred at the restrictive temperature (39.5°C). Both AP activity and osteocalcin (OC) secretion increased in a dose‐dependent manner following dihydroxyvitamin D 3 (1,25‐D 3 ) treatment when cultured at either temperature. However, AP and 1,25‐D 3 ‐induced OC levels were elevated in confluent hFOB cells cultured at 39.5°C compared with 33.5°C. Treatment of hFOB cells with 1–34 parathyroid hormone (PTH) resulted in an increase in cAMP levels. Upon reaching confluence, hFOB cultures went through programmed differentiation and formed mineralized nodules as observed by von Kossa staining. Further, immunostaining of postconfluent, differentiated hFOB cells showed that high levels of osteopontin, osteonectin, bone sialoprotein, and type I collagen were expressed. Therefore, the clonal cell line hFOB 1.19 provides a homogeneous, rapidly proliferating model system to study certain stages of human osteoblast differentiation.