Yuan Zhi-xiang, Zhang Zhi-rong, Zhu Di, Sun Xun, Gong Tao, Liu Jie, Luan Chang-tao
West China School of Pharmacy, Sichuan University, Chengdu, Sichuan, People's Republic of China.
Mol Pharm. 2009 Jan-Feb;6(1):305-14. doi: 10.1021/mp800078a.
In our previous studies, randomly 50% N-acetylated low molecular weight chitosan (LMWC) has been confirmed as a potential carrier for the site-specific delivery of prednisolone to kidney, suggesting specific uptake of LMWC in kidney. Interestingly, aminoglycoside, a well-known ligand of megalin receptor, shares a similar glucosamine unit level with LMWC. Based on these, we proposed that the specific renal uptake of LMWC might also be mediated by the megalin receptor. To test this hypothesis, we performed the present study to further investigate the renal uptake process of LMWC and its possible mechanism as well. First, LMWC was found by fluorescent microscopy to selectively accumulate in the kidneys, especially in the renal proximal tubules after iv injection in mice. Then, our research also revealed that LMWC was internalized into renal tubular cells (RTCs) through endocytic pathway, with a concentration-dependent and saturable pattern. The uptake of LMWC could be competitively inhibited in the presence of gentamycin, a kind of aminoglycosides. In addition, cytotoxicity assay showed that there were no obvious effects of LMWC on the viability of L929 and RTCs lines. Finally, megalin-shedding mouse models were established and the distribution of LMWC in tissues of normal and megalin-shedding mice was evaluated. Consistent with gentamycin inhibition assay, in vivo results also suggested the role of megalin in the uptake of LMWC in kidney. In conclusion, LMWC could be specifically taken up by RTCs, where the megalin receptor would likely mediate its binding and uptake.
在我们之前的研究中，随机选取的50% N-乙酰化低分子量壳聚糖（LMWC）已被证实是将泼尼松龙特异性递送至肾脏的潜在载体，这表明LMWC在肾脏中存在特异性摄取。有趣的是，氨基糖苷类药物是巨膜蛋白受体的一种知名配体，其葡糖胺单位水平与LMWC相似。基于这些，我们推测LMWC在肾脏中的特异性摄取可能也由巨膜蛋白受体介导。为了验证这一假设，我们进行了本研究，以进一步探究LMWC的肾脏摄取过程及其可能的机制。首先，通过荧光显微镜观察发现，静脉注射LMWC后，其在小鼠肾脏中选择性蓄积，尤其是在肾近端小管中。然后，我们的研究还表明，LMWC通过内吞途径内化进入肾小管细胞（RTCs），呈现浓度依赖性和饱和性模式。在庆大霉素（一种氨基糖苷类药物）存在的情况下，LMWC的摄取可被竞争性抑制。此外，细胞毒性试验表明，LMWC对L929和RTCs细胞系的活力没有明显影响。最后，建立了巨膜蛋白脱落小鼠模型，并评估了LMWC在正常小鼠和巨膜蛋白脱落小鼠组织中的分布。与庆大霉素抑制试验结果一致，体内结果也表明巨膜蛋白在LMWC肾脏摄取过程中发挥作用。总之，RTCs可特异性摄取LMWC，其中巨膜蛋白受体可能介导其结合与摄取。
