Solomon S S, Palazzolo M R, Elam M B, Green S, Raghow R
Department of Medicine, University of Tennessee, Memphis.
J Lab Clin Med. 1994 Sep;124(3):348-58.
Previously we demonstrated that in streptozotocin-induced or spontaneously diabetic BB rats (BB-SDR), low-Km cyclic AMP (cAMP), phosphodiesterase (PDE), and calmodulin (CaM) are decreased. Isolated fat cells of diabetic animals synthesized less CaM and contained reduced levels of CaM transcripts (Solomon SS, Palazzolo MR, Green SA, Raghow R. Biochem Biophys Res Commun 1990; 168: 1007-12). Treatment of diabetic animals with insulin restores CaM transcripts to normal. RNA was extracted from isolated hepatocytes from BB-SDR rats in primary tissue culture treated with insulin (from 2.8 x 10(4) to 1.4 x 10(6) microU/ml) for 48 hours, was immobilized on nitrocellulose, and was sequentially hybridized with radiolabeled probes for CaM, actin, and tubulin. Insulin stimulates steady state levels of mRNA for calmodulin > actin > tubulin. Furthermore, decreased steady state levels of CaM mRNA in hepatocytes from diabetic animals are restored to normal levels with in vitro insulin incubation. Data from nuclear transcription run-on assays demonstrate that insulin stimulates transcription of mRNA CaM by 80%. In addition, we observed RNA degradation in the untreated diabetic but not insulin-treated liver. These data support transcriptional as well as post-transcriptional effects of insulin on CaM mRNA. We postulate that in uncontrolled diabetes, elevations in levels of cAMP in tissue result in part from decreased activity of the apparently co-regulated PDE and CaM and that PDE inactivation in diabetes results from both insulin insufficiency and CaM down-regulation.
先前我们证明，在链脲佐菌素诱导的或自发糖尿病的BB大鼠（BB-SDR）中，低Km环磷酸腺苷（cAMP）、磷酸二酯酶（PDE）和钙调蛋白（CaM）减少。糖尿病动物的分离脂肪细胞合成的CaM较少，且CaM转录本水平降低（所罗门SS、帕拉佐洛MR、格林SA、拉戈R。生物化学与生物物理研究通讯1990；168：1007-12）。用胰岛素治疗糖尿病动物可使CaM转录本恢复正常。从原代组织培养中用胰岛素（2.8×10⁴至1.4×10⁶微单位/毫升）处理48小时的BB-SDR大鼠分离的肝细胞中提取RNA，将其固定在硝酸纤维素上，然后依次与放射性标记的CaM、肌动蛋白和微管蛋白探针杂交。胰岛素刺激钙调蛋白>肌动蛋白>微管蛋白的mRNA稳态水平。此外，糖尿病动物肝细胞中CaM mRNA稳态水平的降低通过体外胰岛素孵育恢复到正常水平。核转录连续分析数据表明，胰岛素刺激CaM mRNA转录增加80%。此外，我们观察到未治疗糖尿病肝脏中有RNA降解，但胰岛素治疗的肝脏中没有。这些数据支持胰岛素对CaM mRNA的转录及转录后作用。我们推测，在未控制的糖尿病中，组织中cAMP水平升高部分是由于明显共同调节的PDE和CaM活性降低，且糖尿病中PDE失活是由胰岛素不足和CaM下调共同导致的。
