Experimental Medicine Section, Oral Infection and Immunity Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD 20892, USA.
Diabetologia. 2011 Sep;54(9):2347-57. doi: 10.1007/s00125-011-2221-6. Epub 2011 Jul 6.
AIMS/HYPOTHESIS: Islet antigen 2 (IA-2) and IA-2β are dense core vesicle (DCV) transmembrane proteins and major autoantigens in type 1 diabetes. The present experiments were initiated to test the hypothesis that the knockout of the genes encoding these proteins impairs the secretion of insulin by reducing the number of DCV.
Insulin secretion, content and DCV number were evaluated in islets from single knockout (Ia-2 [also known as Ptprn] KO, Ia-2β [also known as Ptprn2] KO) and double knockout (DKO) mice by a variety of techniques including electron and two-photon microscopy, membrane capacitance, Ca(2+) currents, DCV half-life, lysosome number and size and autophagy.
Islets from single and DKO mice all showed a significant decrease in insulin content, insulin secretion and the number and half-life of DCV (p < 0.05 to 0.001). Exocytosis as evaluated by two-photon microscopy, membrane capacitance and Ca(2+) currents supports these findings. Electron microscopy of islets from KO mice revealed a marked increase (p < 0.05 to 0.001) in the number and size of lysosomes and enzymatic studies showed an increase in cathepsin D activity (p < 0.01). LC3 protein, an indicator of autophagy, also was increased in islets of KO compared with wild-type mice (p < 0.05 to 0.01) suggesting that autophagy might be involved in the deletion of DCV.
CONCLUSIONS/INTERPRETATION: We conclude that the decrease in insulin content and secretion, resulting from the deletion of Ia-2 and/or Ia-2β, is due to a decrease in the number of DCV.
目的/假设：胰岛抗原 2（IA-2）和 IA-2β 是致密核心囊泡（DCV）跨膜蛋白，也是 1 型糖尿病的主要自身抗原。本实验旨在验证以下假说，即这些蛋白编码基因的敲除会通过减少 DCV 的数量而损害胰岛素的分泌。
通过多种技术，包括电子显微镜和双光子显微镜、膜电容、Ca²⁺电流、DCV 半衰期、溶酶体数量和大小以及自噬，评估来自单个基因敲除（IA-2[也称为 Ptprn]KO、IA-2β[也称为 Ptprn2]KO）和双基因敲除（DKO）小鼠的胰岛中的胰岛素分泌、含量和 DCV 数量。
单个和 DKO 小鼠的胰岛均显示出胰岛素含量、胰岛素分泌以及 DCV 的数量和半衰期明显减少（p < 0.05 至 0.001）。双光子显微镜、膜电容和 Ca²⁺电流评估的胞吐作用支持了这些发现。KO 小鼠胰岛的电子显微镜显示溶酶体数量和大小明显增加（p < 0.05 至 0.001），酶学研究显示组织蛋白酶 D 活性增加（p < 0.01）。与野生型小鼠相比，KO 小鼠胰岛中的 LC3 蛋白（自噬的标志物）也增加（p < 0.05 至 0.01），表明自噬可能参与了 DCV 的缺失。
结论/解释：我们得出结论，由于 Ia-2 和/或 Ia-2β 的缺失导致胰岛素含量和分泌减少，是由于 DCV 数量减少所致。
