Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, California, United States of America.
PLoS One. 2011;6(10):e25910. doi: 10.1371/journal.pone.0025910. Epub 2011 Oct 3.
The rotenone-insensitive internal NADH-quinone oxidoreductase from yeast, Ndi1, has been shown to work as a replacement molecule for complex I in the respiratory chain of mammalian mitochondria. In the so-called transkingdom gene therapy, one major concern is the fact that the yeast protein is foreign in mammals. Long term expression of Ndi1 observed in rodents with no apparent damage to the target tissue was indicative of no action by the host's immune system.
METHODOLOGY/PRINCIPAL FINDINGS: In the present study, we examined rat skeletal muscles expressing Ndi1 for possible signs of inflammatory or immune response. In parallel, we carried out delivery of the GFP gene using the same viral vector that was used for the NDI1 gene. The tissues were subjected to H&E staining and immunohistochemical analyses using antibodies specific for markers, CD11b, CD3, CD4, and CD8. The data showed no detectable signs of an immune response with the tissues expressing Ndi1. In contrast, mild but distinctive positive reactions were observed in the tissues expressing GFP. This clear difference most likely comes from the difference in the location of the expressed protein. Ndi1 was localized to the mitochondria whereas GFP was in the cytosol.
CONCLUSIONS/SIGNIFICANCE: We demonstrated that Ndi1 expression did not trigger any inflammatory or immune response in rats. These results push forward the Ndi1-based molecular therapy and also expand the possibility of using foreign proteins that are directed to subcellular organelle such as mitochondria.
已证实酵母中对鱼藤酮不敏感的内部 NADH-醌氧化还原酶 Ndi1 可作为哺乳动物线粒体呼吸链中复合体 I 的替代分子。在所谓的跨物种基因治疗中，一个主要关注点是酵母蛋白在哺乳动物中是外来的这一事实。在没有明显靶组织损伤的啮齿动物中观察到 Ndi1 的长期表达表明宿主免疫系统没有作用。
方法/主要发现：在本研究中，我们检查了表达 Ndi1 的大鼠骨骼肌，以寻找可能的炎症或免疫反应迹象。同时，我们使用相同的病毒载体进行 GFP 基因的传递，该载体用于 NDI1 基因。对组织进行 H&E 染色和免疫组织化学分析，使用针对标志物 CD11b、CD3、CD4 和 CD8 的特异性抗体。数据显示，表达 Ndi1 的组织没有检测到免疫反应的迹象。相比之下，在表达 GFP 的组织中观察到轻微但特征性的阳性反应。这种明显的差异很可能来自于表达蛋白位置的差异。Ndi1 定位于线粒体，而 GFP 位于细胞质中。
结论/意义：我们证明 Ndi1 的表达不会在大鼠中引发任何炎症或免疫反应。这些结果推动了基于 Ndi1 的分子治疗的发展，并扩展了将靶向亚细胞器（如线粒体）的外源蛋白用于治疗的可能性。
