Kellerer M, Machicao F, Berti L, Sixt B, Mushack J, Seffer E, Mosthaf L, Ullrich A, Häring H U
Institut für Diabetesforschung, München, Germany.
Biochem J. 1993 Nov 1;295 ( Pt 3)(Pt 3):699-704. doi: 10.1042/bj2950699.
Inositol phospho-oligosaccharides (IPOs), which are released from liver membranes upon stimulation by insulin, mimic a wide spectrum of insulin effects in different cells, but not the stimulation of glucose transport. We investigated whether other insulin-sensitive tissues release glucose transport-stimulating IPOs and whether this is related to the human insulin receptor isoform-A or -B (HIR-A or HIR-B). Rat1 fibroblasts overexpressing HIR-A or -B (rat1-HIR cells) were labelled with [3H]glucosamine, [3H]mannose or myo-[3H]inositol. IPOs from the cell supernatant were partially purified by an AG1X2 anion-exchange column, and fractions were eluted at different pH values (pH 3, pH 2 and pH 1.3). The label from glucosamine, mannose and myo-inositol appeared predominantly in the pH 2 fraction. The biological activity of the fractions was determined by measuring 3-O-methylglucose transport and lipogenesis in fat cells. Using the pH 2 fraction from the supernatant of rat1-HIR fibroblasts, insulin increased the release of 3-O-methylglucose-transport-stimulating activity (HIR-A: without insulin, 22.4 +/- 5.4%; with insulin 54.0 +/- 8.4%; HIR-B: without insulin 21.6 +/- 7.5%, with insulin, 44.7 +/- 10.6%, given as a percentage of equilibrium glucose transport reached after 4 s) and lipogenesis-stimulating activity (HIR-A: without insulin, 1.24 +/- 0.17; with insulin, 4.69 +/- 0.2; HIR-B: without insulin, 1.34 +/- 0.18; with insulin, 4.98 +/- 0.31, given as nmol of [3H]glucose converted into lipids/min per 10(6) cells). Analogous experiments were performed with isolated rat fat cells expressing the physiological level of insulin receptors. Upon insulin stimulation of fat cells in the presence of 2.5 mM mannose, the release of 3-O-methylglucose-transport-stimulating activity was detected (for purified supernatant of adipocytes without insulin, 6.9 +/- 1.12%; with insulin, 41.0 +/- 3.6%) and lipogenesis-stimulating activity (without insulin, 0.93 +/- 0.17, with insulin 2.96 +/- 0.31 nmol/min per mg). These data suggest (1) that adipocytes and rat1-HIR fibroblasts release IPOs that are able to stimulate glucose transport, (2) that both insulin receptor isoforms (HIR-A and HIR-B) mediate the effect of insulin on IPO release, and (3) that overexpression of insulin receptors increases the basal release of IPOs.
肌醇磷酸寡糖（IPOs）在胰岛素刺激下从肝细胞膜释放，可模拟胰岛素在不同细胞中的多种作用，但不包括对葡萄糖转运的刺激作用。我们研究了其他胰岛素敏感组织是否释放刺激葡萄糖转运的IPOs，以及这是否与人类胰岛素受体同工型A或B（HIR - A或HIR - B）有关。用[³H]葡萄糖胺、[³H]甘露糖或肌醇[³H]肌醇标记过表达HIR - A或 - B的大鼠1成纤维细胞（大鼠1 - HIR细胞）。细胞上清液中的IPOs通过AG1X2阴离子交换柱进行部分纯化，不同pH值（pH 3、pH 2和pH 1.3）洗脱各组分。来自葡萄糖胺、甘露糖和肌醇的标记主要出现在pH 2组分中。通过测量脂肪细胞中3 - O - 甲基葡萄糖转运和脂肪生成来确定各组分的生物活性。使用大鼠1 - HIR成纤维细胞上清液的pH 2组分，胰岛素增加了3 - O - 甲基葡萄糖转运刺激活性的释放（HIR - A：无胰岛素时，22.4±5.4%；有胰岛素时，54.0±8.4%；HIR - B：无胰岛素时，21.6±7.5%，有胰岛素时，44.7±10.6%，以4秒后达到的平衡葡萄糖转运的百分比表示）和脂肪生成刺激活性（HIR - A：无胰岛素时，1.24±0.17；有胰岛素时，4.69±0.2；HIR - B：无胰岛素时，1.34±0.18；有胰岛素时，4.98±0.31，以每10⁶个细胞每分钟转化为脂质的[³H]葡萄糖的纳摩尔数表示）。对表达生理水平胰岛素受体的分离大鼠脂肪细胞进行了类似实验。在2.5 mM甘露糖存在下，胰岛素刺激脂肪细胞后，检测到3 - O - 甲基葡萄糖转运刺激活性的释放（对于无胰岛素的脂肪细胞纯化上清液，6.9±1.12%；有胰岛素时，41.0±3.6%）和脂肪生成刺激活性（无胰岛素时，0.93±0.17，有胰岛素时，2.96±0.31纳摩尔/分钟/毫克）。这些数据表明：（1）脂肪细胞和大鼠1 - HIR成纤维细胞释放能够刺激葡萄糖转运的IPOs；（2）两种胰岛素受体同工型（HIR - A和HIR - B）均介导胰岛素对IPOs释放的作用；（3）胰岛素受体的过表达增加了IPOs的基础释放。
