降钙素基因相关肽转基因预防小鼠自身免疫性糖尿病发病及其抗氧化应激机制(英文)

Calcitonin gene-related peptide gene therapy suppresses reactive oxygen species in the pancreas and prevents mice from autoimmune diabetes

有证据表明,活性氧(reactive oxygen species,ROS)参与了胰岛β细胞自身免疫损伤,是自身免疫性糖尿病发病的重要原因之一。一氧化氮(NO)和过氧化氢(H2O2)介导了致炎性细胞因子对胰岛β细胞的损害,引起脂质过氧化反应,进而损伤胰岛细胞。降钙素基因相关肽(calcitonin gene-related peptide,CGRP)在心肌细胞中可抑制ROS生成而具有细胞保护作用。通过CGRP裸质粒肌肉注射体细胞电针强化转基因方法,使骨骼肌持续表达CGRP,观察其对小剂量多次注射链脲佐菌素(streptozotocin,STZ)造成的小鼠自身免疫性糖尿病发病的影响,进一步测定胰腺局部活性氧含量以及抗氧化酶的改变,探讨其抗氧化应激机制。结果发现,CGRP裸质粒直接注射入小鼠双后肢胫前肌,继以程控电针刺激导入(体内电穿孔法),可使血浆和骨骼肌组织CGRP表达水平显著增高,且持续4周以 上;注射STZ同时给予CGRP转基因治疗可减轻胰岛β细胞损伤,显著降低自身免疫性糖尿病的发病率及血糖水平;CGRP转基因可显著抑制自身免疫性糖尿病小鼠胰腺局部活性氧和丙二醛的生成,增加过氧化氢酶(CAT)及超氧化物歧化酶(superoxide dismutase,SOD)的活性。结果提示,CGRP裸质粒直接注射、电针辅助导入转基因可获得CGRP持续高水平的表达,能够预防小鼠自身免疫性糖尿病的发病。

Reactive oxygen species ( ROS) is involved in autoimmune destruction of isletβ cells, which has been proven to be an important underlying pathogenesis for insulin dependent diabetes mellitus ( ID-DM). Calcitonin gene-related peptide (CGRP) is a widely distributed neuropeptide, which has been found to play an important role in protecting myocytes from ROS. We hypothesized that exogenous CGRP gene administration before the pathogenic stage of insulitis might suppress the production of ROS and provide a hopeful therapeutic intervention for autoimmune diabetes. We performed CGRP gene transfer by injecting naked plasmid directly into skeletal muscles of mice with electroporation enhancement to achieve a continuous expression of CGRP in skeletal muscles, and thereby its secretion into the circulation. The effect of CGRP gene transfer on the pathogenesis of diabetes was studied in autoimmune diabetic mice induced by multiple low dose streptozotocin ( MLDS). The CGRP gene therapy decreased morbidity of autoimmune diabetes , and significantly ameliorated hyperglycemia in these mice. CGRP gene transfer inhibited the production of ROS and malondialdehyde (MDA). In addition, it enhanced the activity of catalase ( CAT) and su-peroxide dismutase ( SOD) significantly. The data suggest that intramuscular CGRP gene transfer ameliorates autoimmune destruction of islet βcells, resulting in significant reduction in diabetes incidence of MLDS diabetes mice. CGRP benefits might be mediated at least in part by inhibiting the oxidative stress in islet β cells of these mice.