Vierck J L, McNamara J P, Dodson M V
Department of Animal Sciences, Washington State University, Pullman 99164-6320, USA.
In Vitro Cell Dev Biol Anim. 1996 Oct;32(9):564-72. doi: 10.1007/BF02722983.
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.
在克隆培养系统中测定了绵羊来源的单房脂肪细胞（脂肪成纤维细胞）后代对地塞米松、胰岛素、胰岛素样生长因子I（IGF-I）、生长激素（GH）和碱性成纤维细胞生长因子（FGF）的反应性。通过悬滴培养技术从绵羊的肌间脂肪组织（半膜肌/半腱肌接缝处脂肪库）获得成熟脂肪细胞的原代培养物。在单房脂肪滴退化并重新建立成纤维样脂肪成纤维细胞后，收集附着在上层培养瓶表面的所有脂肪成纤维细胞，并通过Percoll梯度离心将其与成纤维细胞（无多房小泡）分离。分离出单个脂肪成纤维细胞的后代，并测试其增殖、分化和积累脂质的能力。脂肪成纤维细胞的储备培养物在5天内达到汇合，并用地塞米松-甲基异丁基黄嘌呤-胰岛素（DMI）诱导其在7至9天内分化。在汇合前用胰岛素、IGF-I、GH或FGF孵育，然后用DMI诱导，对随后的分化没有直接（启动）作用。在2天的分化/诱导期内，当单独替代DMI时，所有因子均诱导培养的脂肪成纤维细胞分化，这通过脂肪生成（P <.05）和脂蛋白脂肪酶活性（P <.05）得以确定。因此，绵羊肌肉中分离出的脂肪成纤维细胞可能被激素和生长因子诱导，在细胞培养中呈现成熟脂肪细胞的形态。进一步明确脂肪成纤维细胞培养系统可能有助于识别调节绵羊骨骼肌脂肪细胞发育的机制，以及研究脂肪细胞与肌肉细胞之间的相互联系。
