Abstract：
Delivering functional gene into targeted skin cells or tissues to modulate the genes expression, has the potential to treat various hereditary cutaneous disorders. Nevertheless, the lack of safe and effective gene delivery vehicles greatly limits the clinical translation of gene therapy for inherited skin diseases. Herein, we developed a facile elution fractionation strategy to isolate eight HPAEs with Mw ranging from 7.6 to 131.8 kg/mol and Đ < 2.0 from the one crude HPAE23.7k, and investigated the expression efficiency for TGM1 and COL7A1 plasmids. Gene transfection results revealed that the intermediate MW HPAEs, HPAE20.6k, exhibited the highest gene transfection efficiency (46.4%) and the strongest mean fluorescence intensity (143,032 RLU), compared to other isolated components and the crude product. Importantly, best-performing isolated HPAE effectively delivered COL7A1 (15,974 bp) and TGM1 (7181 bp) plasmids, promoting the efficient expression of type VII collagen (C7) and transglutaminase-1 proteins in cutaneous cells. Our study establishes a straightforward step-by-step elution fractionation strategy for the development of HPAEs gene delivery vectors, expediting their clinical translation in inherited skin diseases.
摘要：
将功能基因传递到目标皮肤细胞或组织以调节基因表达，有可能治疗各种遗传性皮肤疾病。然而，缺乏安全有效的基因传递载体极大地限制了遗传性皮肤病基因治疗的临床应用。在这里，WedevelopedafacileeletrationfractionationstrategytoisolateeightHPAEswithMwrangefrom7.6to131.8kg/moland检修<2.0fromtheonecrudeHPAE23.7k,并研究了TGM1和COL7A1质粒的表达效率。基因转染结果表明，中间体MWHPAEs，HPAE20.6k,基因转染效率最高(46.4%),平均荧光强度最强(143,032RLU),与其他分离的组分和粗产物相比。重要的是,性能最佳的分离HPAE有效递送COL7A1（15,974bp）和TGM1（7181bp）质粒，促进VII型胶原（C7）和转谷氨酰胺酶-1蛋白在皮肤细胞中的有效表达。我们的研究为开发HPAEs基因递送载体建立了一个简单的分步洗脱分级策略，加快他们在遗传性皮肤病的临床翻译。
