Establishment of Protein Delivery Systems Targeting Podocytes
Establishment of Protein Delivery Systems Targeting Podocytes
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DOI：
10.1371/journal.pone.0011837
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发表时间：
2010-07-29
期刊：
影响因子：
3.7
通讯作者：
Reiser, Jochen
中科院分区：
文献类型：
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作者：
Chiang, Wen Chih;Geel, Tessa M.;Reiser, Jochen
背景：足细胞是结构独特的细胞，对肾脏滤过屏障至关重要。它们在肾小球毛细血管外侧的解剖位置使足细胞暴露于大量的血浆和尿组分，因此可用于药物递送。近年来已经清楚地表明，足细胞特异性疾病通路的干扰可以调节肾小球功能和影响的严重程度和进展的肾小球diseases.Methodology/主要调查结果：在这里，我们描述的研究表明，有效的蛋白质运输到哺乳动物细胞小鼠3T3成纤维细胞和足细胞，利用一种称为profection的方法。我们正在使用合成脂质结构，允许蛋白质或抗体的安全包装，导致随后将蛋白质递送到细胞中。脂质包被的蛋白质的摄取通过细胞（例如足细胞）的内在特性促进，以吞噬生理上保留在细胞外基质中的颗粒。限制性内切酶MunI在3T3小鼠成纤维细胞中的表达引起DNA降解的增加。此外，纯化的蛋白质，如β-半乳糖苷酶和大的GT酶发动蛋白，可以使用两种不同的表达试剂表达到足细胞中，成功率为95 - 100%。递送的β-半乳糖苷酶被适当折叠，并且能够在足细胞中切割其底物X-gal。经嘌呤霉素处理的足细胞也是蛋白质货物的潜在受体，因为我们还将荧光团标记的IgG递送到嘌呤霉素处理的足细胞中。我们目前正在优化我们的协议在vivo profection. Conclusions：蛋白质转移是一个令人兴奋的工具，研究和靶向高度分化的细胞，如足细胞。
Background: Podocytes are uniquely structured cells that are critical to the kidney filtration barrier. Their anatomic location on the outer side of the glomerular capillaries expose podocytes to large quantities of both plasma and urinary components and thus are reachable for drug delivery. Recent years have made clear that interference with podocyte-specific disease pathways can modulate glomerular function and influence severity and progression of glomerular disease.Methodology/Principal Findings: Here, we describe studies that show efficient transport of proteins into the mammalian cells mouse 3T3 fibroblasts and podocytes, utilizing an approach termed profection. We are using synthetic lipid structures that allow the safe packing of proteins or antibodies resulting in the subsequent delivery of protein into the cell. The uptake of lipid coated protein is facilitated by the intrinsic characteristic of cells such as podocytes to engulf particles that are physiologically retained in the extracellular matrix. Profection of the restriction enzyme MunI in 3T3 mouse fibroblasts caused an increase in DNA degradation. Moreover, purified proteins such as beta-galactosidase and the large GTPase dynamin could be profected into podocytes using two different profection reagents with the success rate of 95-100%. The delivered b-galactosidase enzyme was properly folded and able to cleave its substrate X-gal in podocytes. Diseased podocytes are also potential recipients of protein cargo as we also delivered fluorophore labeled IgG into puromycin treated podocytes. We are currently optimizing our protocol for in vivo profection.Conclusions: Protein transfer is developing as an exciting tool to study and target highly differentiated cells such as podocytes.