Intracellular trafficking pathways in silver nanoparticle uptake and toxicity in Caenorhabditis elegans

We used the nematode Caenorhabditis elegans to study the roles of endocytosis and lysosomal function in uptake and subsequent toxicity of silver nanoparticles (AgNP) in vivo. To focus on AgNP uptake and effects rather than silver ion (AgNO3) effects, we used a minimally dissolvable AgNP, citrate-coated AgNPs (CIT-AgNPs). We found that the clathrin-mediated endocytosis inhibitor chlorpromazine reduced the toxicity of CIT-AgNPs but not AgNO3. We also tested the sensitivity of three endocytosis-deficient mutants (rme-1, rme-6 and rme-8) and two lysosomal function deficient mutants (cup-5 and glo-1) as compared to wild-type (N2 strain). One of the endocytosis-deficient mutants (rme-6) took up less silver and was resistant to the acute toxicity of CIT-AgNPs compared to N2s. None of those mutants showed altered sensitivity to AgNO3. Lysosome and lysosome-related organelle mutants were more sensitive to the growth-inhibiting effects of both CIT-AgNPs and AgNO3. Our study provides mechanistic evidence suggesting that early endosome formation is necessary for AgNP-induced toxicity in vivo, as rme-6 mutants were less sensitive to the toxic effects of AgNPs than C. elegans with mutations involved in later steps in the endocytic process.本研究以秀丽隐杆线虫（Caenorhabditis elegans）为模式生物，探究内吞作用（endocytosis）与溶酶体功能在银纳米颗粒（silver nanoparticles, AgNP）的体内摄取及后续毒性中的作用。为聚焦银纳米颗粒的摄取与效应而非银离子（AgNO3）的作用，本研究选用了低溶解性的银纳米颗粒——柠檬酸盐包被银纳米颗粒（CIT-AgNPs）。研究发现，网格蛋白介导的内吞作用（clathrin-mediated endocytosis）抑制剂氯丙嗪可降低CIT-AgNPs的毒性，但对AgNO3的毒性无影响。本研究还以野生型（N2菌株）为对照，测试了3株内吞缺陷突变体（rme-1、rme-6、rme-8）与2株溶酶体功能缺陷突变体（cup-5、glo-1）的敏感性。相较于N2菌株，其中一株内吞缺陷突变体（rme-6）的银摄取量更低，且对CIT-AgNPs的急性毒性产生了抗性。上述所有突变体对AgNO3的敏感性均未发生改变。溶酶体与溶酶体相关细胞器突变体对CIT-AgNPs和AgNO3的生长抑制效应均表现出更高的敏感性。本研究提供了机制层面的证据，表明早期内体形成（early endosome formation）对于银纳米颗粒诱导的体内毒性是必需的——相较于携带内吞过程后续步骤相关突变的秀丽隐杆线虫，rme-6突变体对银纳米颗粒毒性的敏感性更低。