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首页> 外文期刊>ACS Omega >Intracellular Delivery of β-Galactosidase Enzyme Using Arginase-Responsive Dextran Sulfate/Poly-l-arginine Capsule for Lysosomal Storage Disorder
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Intracellular Delivery of β-Galactosidase Enzyme Using Arginase-Responsive Dextran Sulfate/Poly-l-arginine Capsule for Lysosomal Storage Disorder

机译：β-半乳糖苷酶酶的细胞内传递使用精氨酸酶反应性硫酸葡聚糖/聚-1-精氨酸胶囊治疗溶酶体贮积症

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β-Galactosidase (β-gal) is one of the important lysosomal enzymes that is involved in the breakdown of glycosphingolipids (e.g., GM1 ganglioside), and its deficiency leads to GM1 Gangliosidosis, a lysosomal storage disorder (LSD). Intracellular delivery of β-gal is one of the preferable methods to treat this kind of LSDs. However, it cannot permeate the cell membrane due to its intricate macromolecular nature, low stability, and degradation by endogenous proteases. To this end, we report efficient intracellular delivery of β-gal via arginase-responsive dextran sulfate/poly-l-arginine polymer capsules (DS/PA capsules). The therapeutic activity of β-gal enzyme has been assessed in two gene-deficient diseased cell lines, SV (β-galactosidase gene-deficient mouse fibroblast) and R201C (deficient human β-galactosidase gene-introduced mouse fibroblast), and in wild-type mouse fibroblast immortalized cell lines. The activity of β-gal enzyme has been estimated within cells by using fluorescein isothiocyanate-cholera toxin B as a florescent probe that illustrates the level of GM1 ganglioside, the β-gal substrate. We found 1.8-, 3.4-, and 2.8-fold reduction in the substrate level in R201C, SV, and wild-type mouse fibroblast, respectively, which confirms the release and therapeutic activity of β-gal enzyme inside the cells. Moreover, enzyme delivery in gene-deficient diseased cell lines (SV and R201C) via DS/PA capsules reduced the level of enzyme substrate to a normal endogenous level, which is present in untreated wild-type mouse fibroblast cells. We note that loading of β-gal enzyme within DS/PA capsules was estimated to be 3 mU per hundred capsules and more than 77% of β-gal is released within 12 h. Overall, these results highlight the potential of DS/PA capsules as an efficient delivery carrier for therapeutic enzyme.
机译：β-半乳糖苷酶（β-gal）是一种重要的溶酶体酶，参与糖鞘糖脂的分解（例如GM1神经节苷脂），其缺乏会导致GM1神经节苷脂病，一种溶酶体贮积病（LSD）。 β-gal的细胞内递送是治疗这种LSD的优选方法之一。然而，由于其复杂的大分子性质，低稳定性和被内源蛋白酶降解，它不能渗透细胞膜。为此，我们报道了通过精氨酸酶反应性葡聚糖硫酸盐/聚-1-精氨酸聚合物胶囊（DS / PA胶囊）有效的β-gal细胞内递送。已经在两种基因缺陷的病变细胞系SV（β-半乳糖苷酶基因缺陷的小鼠成纤维细胞）和R201C（人类β-半乳糖苷酶基因缺陷的小鼠成纤维细胞）以及野生动物中评估了β-gal酶的治疗活性。型小鼠成纤维细胞永生细胞系。已经通过使用异硫氰酸荧光素-霍乱毒素B作为荧光探针评估了细胞内β-gal酶的活性，该探针说明了β-gal底物GM1神经节苷脂的水平。我们发现R201C，SV和野生型小鼠成纤维细胞的底物水平分别降低了1.8倍，3.4倍和2.8倍，这证实了β-gal酶在细胞内的释放和治疗活性。此外，通过DS / PA胶囊在基因缺陷型病变细胞系（SV和R201C）中的酶递送将酶底物的水平降低到正常的内源水平，该水平存在于未经处理的野生型小鼠成纤维细胞中。我们注意到，DS / PA胶囊中β-gal酶的负载量估计为每100粒3 mU，超过12％的β-gal在12小时内释放。总体而言，这些结果突出了DS / PA胶囊作为治疗性酶的有效递送载体的潜力。

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