Functional Role of Protein Kinase C during Differentiation of Rat Preadipocytes in Serum-free Culture.

为了阐明肥胖的发生机制，我们研究了蛋白激酶C在前脂肪细胞分化过程中的作用。从雄性SD大鼠附睾脂肪垫中获得前脂肪细胞，并在胰岛素、转铁蛋白和T3的作用下体外培养。收集细胞，通过形态和甘油3-磷酸脱氢酶活性评估脂肪转化程度。在免疫印迹研究中，PKC异构体的特异性抗体被用来检测PKC的细胞表达。PKC激活剂TPA抑制脂肪细胞分化。相反，staurosporine和H7， PKC抑制剂，增强分化和逆转TPA的抑制作用。免疫分析显示pkc - α、- δ、-epsilon和-zeta亚型在脂肪前细胞和成熟脂肪细胞中均有表达。pkc - α和-epsilon的表达在分化过程中增加。TPA下调pkc - α和- δ的表达。激活PKC可抑制大鼠脂肪前体细胞的分化。PKC同工异构体表达的差异表明激酶同工异构体作为调节剂的不同作用。

To delineate the mechanism of the generation of obesity, we investigated the role of protein kinase C during differentiation of preadipocyte. Preadipocytes were obtained from epididymal fat pads of male SD rats, and were cultured in vitro in the presence of insulin, transferrin, and T3. Cells were harvested and the degree of adipose conversion was assessed both by morphology and by glycerol 3-phosphate dehydrogenase activity. For immunoblot studies, specific antibodies to PKC isoforms were used to detect cellular expression of PKC.TPA, an activator of PKC suppressed adipocyte differentiation in. Conversely, staurosporine and H7, inhibitors of PKC, enhanced differentiation and reversed the inhibitory effect of TPA.Immumoblot analysis revealed the expression of PKC-alpha, -delta, -epsilon, and -zeta isoforms in both preadipocytes and mature adipocytes. Expression of PKC-alpha and -epsilon increased during the course of differentiation. TPA down regulated the expression of PKC-alpha and -delta. Activation of PKC suppressed differentiation of rat adipocyte precursor cells. Difference in expression of the isoforms of PKC suggests the differential role of the kinase isoforms as a modulator.