Adenovirus-based vascular endothelial growth factor gene delivery to human pancreatic islets
Adenovirus-based vascular endothelial growth factor gene delivery to human pancreatic islets
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DOI：
10.1038/sj.gt.3302267
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发表时间：
2004-07-01
期刊：
影响因子：
5.1
通讯作者：
Mahato, RI
中科院分区：
文献类型：
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作者：
Cheng, K;Fraga, D;Mahato, RI
由于血运重建不良、宿主免疫排斥和非特异性炎症反应导致胰岛移植失败，胰岛移植受到限制。将人血管内皮生长因子（hVEGF）基因输送到胰岛可能会促进胰岛血运重建和存活。我们使用编码 hVEGF 和无 CpG 的绿色荧光蛋白等位基因 (Adv-GFP-hVEGF) 的双顺反子腺病毒载体，并通过转染将其引入人胰岛。我们发现转染效率和细胞凋亡取决于感染复数（MOI）。与Adv-GFP转染和未转染的胰岛相比，Adv-GFP-hVEGF转染的胰岛分泌的hVEGF水平较高，并且hVEGF表达与MOI（10-5000）之间呈现线性关系。还观察到来自未转染胰岛的 hVEGF 持续但低水平的表达。这可能是由于内源性 hVEGF 基因在缺氧条件下表达所致。通过 ELISA 测定胰岛裂解物的 DNA 片段化水平取决于 Adv-GFP-hVEGF 的 MOI。在葡萄糖激发时，转染胰岛的胰岛素释放与未转染胰岛相当。 Adv-GFP-hVEGF 转染的胰岛中 hVEGF 的免疫组织化学染色非常高。在转染和未转染的胰岛中也观察到 hCD31 的弱染色。这些发现表明 Adv-GFP-hVEGF 是促进胰岛血运重建的潜在候选者。
Islet transplantation is limited by islet graft failure due to poor revascularization, host immune rejection and nonspecific inflammatory response. Delivery of human vascular endothelial growth factor (hVEGF) gene to the islets is likely to promote islet revascularization and survival. We used a bicistronic adenoviral vector encoding hVEGF and CpG-free allele of green fluorescent protein (Adv-GFP-hVEGF) and introduced into human pancreatic islets by transfection. We found that transfection efficiency and apoptosis were dependent on the multiplicity of infection (MOI). Compared to Adv-GFP transfected and nontransfected islets, the levels of hVEGF secreted from Adv-GFP-hVEGF transfected islets were higher and exhibit a linear relationship between hVEGF expression and MOI (10-5000). Persistent, but low level expression of hVEGF from nontransfected islets was also observed. This may be due to expression of the endogenous hVEGF gene under hypoxic conditions. The levels of DNA fragmentation determined by ELISA of islet lysates were dependent on the MOI of Adv-GFP-hVEGF. On glucose challenge, insulin release from transfected islets was comparable to nontransfected islets. Immunohistochemical staining for hVEGF was very high in Adv-GFP-hVEGF transfected islets. Weak staining was also observed for hCD31 in both transfected and nontransfected islets. These findings suggest that Adv-GFP-hVEGF is a potential candidate for promoting islet revascularization.