Differentiation of muscle, fat, cartilage, and bone from progenitor cells present in a bone-derived clonal cell population: effect of dexamethasone.

Abstract

Abstract. RCJ 3.1, a clonally derived cell population isolated from 21-d fetal rat calvaria, expresses the osteoblast-associated characteristics of polygonal morphology, a cAMP response to parathyroid hormone, synthesis of predominantly type I collagen, and the presence of 1,25-dihydroxyvitamin D3-regulated alkaline phosphatase activity. When cultured in the presence of ascorbic acid, sodium 13-glycerophosphate, and the synthetic glucocorticoid dexamethasone, this clone differentiated in a time-dependent manner into four morphologically distinct phenotypes of known mesenchymal origin. Multinucleated muscle cells were observed as early as 9-10 d in culture, lipid-containing adipocytes formed after 12 d, chondrocyte nodules were observed after 16 d, and mineralized bone nodules formed after 21 d in culture. The differentiated cell types were characterized morphologically, histochemically, and immunohistochemically. The formation of adipocytes and chondrocytes was dependent upon the addition of dexamethasone; the muscle and bone phenotypes were also expressed at low frequency in the absence of dexamethasone. The sex steroid hormones progesterone and 1713-estradiol had no effect on differentiation in this system, suggesting that the effects of dexamethasone represent effects specific for glucocorticosteroids. Increasing concentrations of dexamethasone (10-9-10 -6 M) increased the numbers of myotubes, adipocytes, and chondrocytes; however, when present continuously for 35 d, the lower concentrations appeared to better maintain the muscle and adipocyte phenotypes. Bone nodules were not quantitated because the frequency of bone nodule formation was too low. Single cells obtained by plating RCJ 3.1 cells at limiting dilutions in the presence of dexamethasone, were shown to give rise to subclones that could differentiate into either single or multiple phenotypes. Thus, the data suggest that this clonal cell line contains subpopulations of mesenchymal progenitor cells which can, under the influence of glucocorticoid hormones, differentiate in vitro into four distinct cell types. It is, therefore, a unique cell line which will be of great use in the study of the regulation of mesenchymal stem cell differentiation.

An immortalized osteogenic cell line derived from mouse teratocarcinoma is able to mineralize in vivo and in vitro.

“…Hopefully, some of these clones may correspond to immortalized stages of bone commitment earlier than the C1 stage. This idea is supported by a recent report (13) showing that progenitor cells in a bonederived clonal cell population from rat can differentiate into bone as well as into muscle, fat, or cartilage. Therefore, our immortalized teratocarcinoma-derived system appears to be of interest to study also the regulation of mesenchymal stem cell differentiation.…”

Mesenchymal stem cells reside within the connective tissues of many organs

“…Similarly, Young et al (1993) demonstrated the existence of clonal populations of both lineage-committed progenitor mesenchymal stem cells and lineage-uncommitted pluripotent mesenchymal stem cells located within the connective tissues of avian limb skeletal muscle, dermis, and heart. The lineage-uncommitted pluripotent mesenchymal stem cells had the potential to form mesodermal phenotypes similar to those observed by Grigoriadis et al (1988). Young et al (1993) proposed the hypothesis that populations of both lineage-committed progenitor mesenchymal stem cells and lineage-uncommitted pluripotent mesenchymal stem cells are located within connective tissue compartments associated with multiple organs and organ systems.…”

“…For example, cartilage and bone induction occurs in the associated connective tissues of various nonchondrogenic and non-osteogenic tissues and organs exposed to demineralized bone matrix (Urist et al, 1978;Nathanson et al, 1978;Nathanson and Hay, 1980). Grigoriadis et al (1988) induced the appearance of muscle, cartilage, bone, and fat from a clonal population of cells derived from the periosteal connective tissue surrounding embryonic rat calvarial bone. Similarly, Young et al (1993) demonstrated the existence of clonal populations of both lineage-committed progenitor mesenchymal stem cells and lineage-uncommitted pluripotent mesenchymal stem cells located within the connective tissues of avian limb skeletal muscle, dermis, and heart.…”

Chondrogenic potential of chick embryonic calvaria: I. Low calcium permits cartilage differentiation

“…For example, Bellows and Aubin (1989) identified only 0.3% of the cells in isolated rat calvarial populations as osteoprogenitor cells. Grigoriadis et al (1988) also described a clonally derived cell population of fetal rat calvarial cells which expressed osteoblast-associated characteristics, but also contained a subpopulation of mesenchyma1 progenitor cells which could differentiate into four distinct cell types, including chondrocytes, under various culture conditions. The presence of cartilage markers in calvaria and other membrane bones, as well as in the periosteum has also been observed previously under different environmental conditions (Hall and Jackobson, 1975;Terashima and Urist, 1975;Thorogood, 1979;Tyler, 1983;Hall, 1987).…”