Hausman G J
Richard B. Russell Research Center, ARS, USDA, Athens, GA 30604-5677, USA.
J Anim Sci. 2000 May;78(5):1227-35. doi: 10.2527/2000.7851227x.
Expression of CAAT/enhancer binding protein (C/EBP) isoforms was examined in primary cultures of adipose tissue stromal vascular (S-V) cells before and during preadipocyte differentiation. Immunocytochemistry showed that the proportions and numbers of C/EBPalpha-, C/EBPbeta-, and C/EBPdelta-reactive cells were maximized after seeding and plating from d 0 to 3 in fetal bovine serum (FBS). However, there were few preadipocytes (AD-3+) and fewer cells with lipid and the number of C/EBPalpha-reactive cells clearly exceeded the number of preadipocytes. Seeding and plating in dexamethasone (DEX) + FBS from d 0 to 3 markedly increased the proportions and numbers of preadipocytes and C/EBPalpha-reactive cells compared to seeding and plating in FBS, d 0 to 3. The number of C/EBPalpha- and C/EBPbeta-reactive cells and preadipocyte reactivity for C/EBPbeta decreased with insulin or DEX treatment, d 3 to 6, following FBS, d 0 to 3. However, insulin + DEX treatment, d 3 to 6, maintained the number of C/EBPalpha-reactive cells and either maintained or increased preadipocyte reactivity for C/EBPalpha and C/EBPbeta. DEX and DEX + insulin treatment induced recruitment of a similar number of preadipocytes, but preadipocytes were not reactive for C/EBPalpha and C/EBPbeta in DEX-treated cultures. The number of C/EBPdelta reactive cells did not change from d 3 to 6 and was not influenced by hormone treatment. After DEX + FBS, d 0 to 3, the high numbers of C/EBPalpha-reactive cells and preadipocytes were maintained by insulin treatment alone. Western blot analysis for C/EBPalpha confirmed the immunocytochemical results. Double staining demonstrated that expression of C/EBPalpha protein was maximized before or at the onset of lipid accretion, whereas expression of C/EBPbeta protein was correlated with lipid accretion. These results indicate that coupling or integration of preadipocyte recruitment with C/EBPalpha expression may be a critical step in glucocorticoid-induced adipogenesis.
在脂肪组织基质血管（S-V）细胞原代培养物中，在脂肪前体细胞分化之前及分化期间，检测了CAAT/增强子结合蛋白（C/EBP）亚型的表达。免疫细胞化学显示，在含有胎牛血清（FBS）的情况下，从第0天到第3天接种和铺板后，C/EBPα、C/EBPβ和C/EBPδ反应性细胞的比例和数量达到最大值。然而，脂肪前体细胞（AD-3+）很少，含脂质的细胞也较少，且C/EBPα反应性细胞的数量明显超过脂肪前体细胞的数量。与在第0天到第3天用FBS接种和铺板相比，在第0天到第3天用地塞米松（DEX）+FBS接种和铺板显著增加了脂肪前体细胞和C/EBPα反应性细胞的比例和数量。在第0天到第3天用FBS培养后，在第3天到第6天用胰岛素或DEX处理，C/EBPα和C/EBPβ反应性细胞的数量以及脂肪前体细胞对C/EBPβ的反应性降低。然而，在第3天到第6天用胰岛素+DEX处理，维持了C/EBPα反应性细胞的数量，并维持或增加了脂肪前体细胞对C/EBPα和C/EBPβ的反应性。DEX和DEX+胰岛素处理诱导的脂肪前体细胞数量相似，但在DEX处理的培养物中，脂肪前体细胞对C/EBPα和C/EBPβ无反应。从第3天到第6天，C/EBPδ反应性细胞的数量没有变化，且不受激素处理的影响。在第0天到第3天用DEX+FBS处理后，仅用胰岛素处理就能维持大量的C/EBPα反应性细胞和脂肪前体细胞。对C/EBPα的蛋白质印迹分析证实了免疫细胞化学结果。双重染色表明，C/EBPα蛋白的表达在脂质积聚之前或开始时达到最大值，而C/EBPβ蛋白的表达与脂质积聚相关。这些结果表明，脂肪前体细胞募集与C/EBPα表达的偶联或整合可能是糖皮质激素诱导脂肪生成的关键步骤。
