Enhanced Vascularization and Survival of Neural Transplants with Ex Vivo Angiogenic Gene Transfer
Enhanced Vascularization and Survival of Neural Transplants with Ex Vivo Angiogenic Gene Transfer
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DOI：
10.3727/000000002783985828
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发表时间：
2002-05
影响因子：
3.3
通讯作者：
D. Casper;Samara J Engstrom;Gautam R Mirchandani;A. Pidel;David Palencia;Paul H Cho;M. Brownlee
中科院分区：
文献类型：
--
作者：
D. Casper;Samara J Engstrom;Gautam R Mirchandani;A. Pidel;David Palencia;Paul H Cho;M. Brownlee
通过神经移植恢复大脑功能在很大程度上取决于供体神经元的存活。不幸的是，在啮齿动物模型和人类帕金森病患者中，移植神经元的存活率都很低。我们采用了一种策略，通过提高血管形成的速度来增加移植物的营养可用性。含有人血管内皮生长因子 (HSVhvegf) cDNA 和细菌 β-半乳糖苷酶基因 (HSVlac) 的复制缺陷型 HSV-1 载体已平行转导至由胚胎第 15 天大鼠中脑细胞制成的非贴壁神经元聚集培养物中。通过用 X-gal 染色来确认固定制剂中 HSVlac 的基因表达。通过夹心 ELISA 检测培养物上清液测定，HSVhvegf 感染的 VEGF 表达比 HSVlac 感染的姐妹培养物高出 322 倍。该肽还具有生物活性，可在体外诱导内皮细胞增殖。成年 Sprague-Dawley 大鼠接受双侧纹状体移植，一侧移植有 HSVlac，另一侧移植有 HSVhvegf。在长达 8 周的规定时间间隔内，处死动物并评估纹状体振动切片的细胞存活和血管形成的各种参数。结果表明，HSVhvegf 转导的移植物中血管密度呈剂量依赖性增加。这些移植物在移植后 4 周内以更快的速度血管化。 8周后，HSVhvegf感染的移植物的平均大小是对照的两倍。特别是，移植的多巴胺能神经元的存活率增加了 3.9 倍。总而言之，这些实验提供了令人信服的证据，表明血管形成率可能是神经元存活的主要决定因素，可以通过 VEGF 基因转导来操纵。
Restoration of brain function by neural transplants is largely dependent upon the survival of donor neurons. Unfortunately, in both rodent models and human patients with Parkinson's disease the survival rate of transplanted neurons has been poor. We have employed a strategy to increase the availability of nutrients to the transplant by increasing the rate at which blood vessels are formed. Replication-deficient HSV-1 vectors containing the cDNA for human vascular endothelial growth factor (HSVhvegf) and the bacterial β-galacto-sidase gene (HSVlac) have been transduced in parallel into nonadherent neuronal aggregate cultures made of cells from embryonic day 15 rat mesencephalon. Gene expression from HSVlac was confirmed in fixed preparations by staining with X-gal. VEGF expression as determined by sandwich ELISA assay of culture supernatant was up to 322-fold higher in HSVhvegf-infected than HSVlac-infected sister cultures. This peptide was also biologically active, inducing endothelial cell proliferation in vitro. Adult Sprague-Dawley rats received bilateral transplants into the striatum, with HSVlac on one side and HSVhvegf on the other. At defined intervals up to 8 weeks, animals were sacrificed and vibratome sections of the striatum were assessed for various parameters of cell survival and vascularization. Results demonstrate dose-dependent increases in blood vessel density within transplants transduced with HSVhvegf. These transplants were vascularized at a faster rate up to 4 weeks after transplantation. After 8 weeks, the average size of the HSVhvegf-infected transplants was twice that of controls. In particular, the survival of transplanted dopaminergic neurons increased 3.9-fold. Taken together these experiments provide convincing evidence that the rate of vascularization may be a major determinant of neuronal survival that can be manipulated by VEGF gene transduction.