Proliferation and differentiation of progeny of ovine unilocular fat cells (adipofibroblasts)

Abstract

The responsiveness of progency of sheep-derived unilocular fat cells (adipofibroblasts) to dexamethasone, insulin, insulinlike growth factor I (IGF-I), growth hormone (GH), and basic fibroblst growth factor (FGF) was determined in a clonal culture system. Primary cultures of mature adipocytes were obtained from intermuscular adipose tissue (semimembranosus/semitendinosus seam depot) of sheep by ceiling culture techniques. Following degeneration of unilocular fat droplets and re-establishment of fibroblasticlike adipofibroblasts, all adipofibroblasts adhering to upper flask surfaces were collected and isolated away from fibroblasts (which had no multilocular vesicles) by Percoll gradient centrifugation. Progeny derived from a single adipofibroblast were isolated and tested for the ability to proliferate, differentiate, and accumulate lipids. Stock cultures of adipofibroblasts reached confluence in 5 d and were induced to differentiate from 7 to 9 d with dexamethasone-methyl isobutylxanthine-insulin (DMI). Incubation with insulin, IGF-I, GH, or FGF prior to confluence followed by induction with DMI produced no direct (priming) effect on subsequent differentiation. When substituted individually in place of DMI during the 2 d differentiation/induction period, all factors induced differentiation of cultured adipofibroblasts as determined by lipogenesis (P < .05) and lipoprotein lipase activity (P < .05). Thus, isolated adipofibroblasts from sheep muscle may be induced by hormones and growth factors to display mature adipocyte morphology in cell culture. Further definition of the adipofibroblast culture system may aid in the identification of mechanisms regulating adipocyte development in sheep skeletal muscle, as well as in the study of intercommunication between fat and muscle cells.

Regulation of differentiation of sheep subcutaneous and abdominal preadipocytes in culture

“…Previous studies with ovine (Broad & Ham 1983, Casteilla et al 1991, Vierck et al 1996 and bovine preadipocytes (Aso et al 1995) have also found a high concentration of insulin or IGF-I to be essential for differentiation, and dexamethasone to enhance differentiation. Broad & Ham (1983) found a lipid supplement stimulated differentiation of ovine preadipocytes, while Björntorp et al (1980) showed that lipid stimulated differentiation of rat preadipocytes.…”

Clonogenic analysis reveals reserve stem cells in postnatal mammals: I. Pluripotent mesenchymal stem cells

“…Previous studies (Mauro, 1961;Cruess, 1982;Campion, 1984;Owen, 1988;Beresford, 1989;Ailhaud et al, 1992;Grounds et al, 1992;Rickard et al, 1994;Vierck et al, 1996;Caplan et al, 1997;Kadiyala et al, 1997;Prockop, 1997;Johnstone et al, 1998;Palis and Segel, 1998;McGuire, 1998;Ratajczak et al, 1998;Pittenger et al, 1999) established the existence of resident populations of lineage-committed progenitor stem cells and have proposed their involvement in maintenance and repair of body tissues. Studies from our lab and others (Grigoriadis et al, 1988;Young et al, 1992aYoung et al, , 1995Young et al, , 1998aYoung, 2000;Eisenberg and Markwald, 1997; have suggested the presence of both pluripotent stem cells and progenitor stem cells in prenatal animals.…”

“…Examples of unipotent reserve stem cells from the mesodermal germ layer include the unipotent myosatellite myoblasts of muscle (Mauro, 1961;Campion, 1984;Grounds et al, 1992), the unipotent adipoblast cells of adipose tissue (Ailhaud et al, 1992), and the unipotent chondrogenic cells and osteogenic cells of the perichondrium and periosteum, respectively (Cruess, 1982;Young et al, 1995). Bipotent reserve mesenchymal stem cells include the adipofibroblasts of adipose tissue (Vierck et al, 1996) and the bipotent chondrogenic/osteogenic stem cells of marrow (Owen, 1988;Beresford, 1989;Rickard et al, 1994;Caplan et al, 1997;Prockop, 1997). Tripotent reserve mesenchymal stem cells include the chondrogenic/ osteogenic/adipogenic stem cells of marrow (Pittenger et al, 1999).…”

“…Preliminary studies involving postnatal rats, rabbits, and humans suggest that resident stem cells consist of both lineage-uncommitted pluripotent stem cells (Pate et al, 1993;Lucas et al, 1995Lucas et al, , 1996Young et al, 1999) as well as the established populations of lineage-committed progenitor stem cells (Mauro, 1961;Cruess, 1982;Campion, 1984;Owen, 1988;Beresford, 1989;Ailhaud et al, 1992;Grounds et al, 1992;Rickard et al, 1994;Vierck et al, 1996;Caplan et al, 1997;Prockop, 1997;Palis and Segel, 1998;McGuire, 1998;Ratajczak et al, 1998;Pittenger et al, 1999). The current clonogenic study was undertaken to determine whether clonal populations of pluripotent stem cells are present in the connective tissues of postnatal rats, and to examine their functional capabilities.…”

Adult stem cells

“…Bipotent progenitor cells include the adipofibroblasts of adipose tissue (Vierck et al, 1996) and the bipotent chondrogenic-osteogenic stem cells of marrow (Owen, 1988;Owen and Friedenstein, 1988;Beresford, 1989;Caplan et al, 1997;Prockop, 1997;Young, 2000). Tripotent progenitor cells include the chondrogenic-osteogenic-adipogenic stem cells of marrow (Pittenger et al, 1999;Young, 2000).…”