Emanueli Costanza, Graiani Galliam, Salis Maria B, Gadau Sergio, Desortes Elisa, Madeddu Paolo
Experimental Medicine and Gene Therapy Section, Istituto Nazionale Biostrutture e Biosistemi, Alghero and Osilo, Italy.
Diabetes. 2004 Apr;53(4):1096-103. doi: 10.2337/diabetes.53.4.1096.
Diabetes macro- and microvascular disease causes tissue hypoperfusion. This deficit, together with a failure to mount an adequate angiogenic response, might explain why vascular occlusion evolves more severely among diabetic patients. The present study investigated whether prophylactic gene therapy with human tissue kallikrein (hTK) may protect diabetic limbs from the consequences of supervening ischemia. Vehicle (saline) or an adenovirus carrying the gene for either hTK (Ad.hTK) or luciferase (Ad.Luc) was injected into left adductor muscles of streptozotocin-induced type 1 diabetic mice 2 weeks before operative occlusion of the ipsilateral femoral artery. Saline-injected nondiabetic mice served as controls. Hindlimb blood flow recovery was analyzed sequentially over the 2 weeks after ischemia induction. At necroscopy, microvessel density and endothelial cell proliferation and apoptosis were quantified in skeletal muscles. We found that limb perfusion recovery of saline-injected type 1 diabetic mice is delayed because of insufficient reparative neovascularization and excessive activation of endothelial cell apoptosis. By contrast, prophylactic Ad.hTK renewed the ability to mount an appropriate neovascularization response to ischemia, suppressed apoptosis, and upregulated endothelial nitric oxide synthase expression. Ultimately, correction of diabetic endotheliopathy by Ad.hTK allowed proper perfusion recovery as seen in nondiabetic mice. These discoveries disclose new therapeutic options for the treatment of diabetic complications.
糖尿病的大血管和微血管疾病会导致组织灌注不足。这种不足，再加上无法产生足够的血管生成反应，可能解释了为什么糖尿病患者的血管闭塞会发展得更严重。本研究调查了用人组织激肽释放酶（hTK）进行预防性基因治疗是否可以保护糖尿病肢体免受随后缺血的影响。在同侧股动脉手术闭塞前2周，将载体（生理盐水）或携带hTK基因（Ad.hTK）或荧光素酶基因（Ad.Luc）的腺病毒注射到链脲佐菌素诱导的1型糖尿病小鼠的左内收肌中。注射生理盐水的非糖尿病小鼠作为对照。在缺血诱导后的2周内依次分析后肢血流恢复情况。尸检时，对骨骼肌中的微血管密度、内皮细胞增殖和凋亡进行定量分析。我们发现，注射生理盐水的1型糖尿病小鼠的肢体灌注恢复延迟，原因是修复性新生血管形成不足和内皮细胞凋亡过度激活。相比之下，预防性给予Ad.hTK恢复了对缺血产生适当新生血管形成反应的能力，抑制了凋亡，并上调了内皮型一氧化氮合酶的表达。最终，Ad.hTK纠正糖尿病内皮病变后，实现了如非糖尿病小鼠所见的适当灌注恢复。这些发现揭示了治疗糖尿病并发症的新治疗选择。
