Nanoparticle cytotoxicity, lysosomal membrane permeabilisation and autophagy induction in kidney cells in vitro

Data collected in completion of a Masters of Science degree in Human Physiology.The datasets were aimed to investigate polyethylene glycol carboxyl gold nanoparticles (PEG-COOH AuNPs) uptake, cytotoxicity and possible autophagy induction in human embryonic kidney (HEK-293) cells in vitro. PEG-COOH AuNPs were firstly characterised in terms of their physicochemical properties when suspended in cell culture media and milli-Q water using transmission electron microscopy (TEM), absorbance spectroscopy, zeta potential meter and a pH meter. HEK 293 cells were then exposed to PEG-COOH AuNPs (0.5 nM -10 nM) for 24 hours and cytotoxicity was assessed using the xCELLigence real-time cell analyser (RTCA). Cellular uptake of PEG-COOH AuNPs (0.13 nM) was assessed using CytoViva dark-field microscopy. The internalisation of PEG-COOH AuNPs into vesicles was visualised using TEM. Analytical interference of the PEG?COOH AuNPs with the LysoTracker green staining dye and the Western blot protein quantification assay kit was assessed prior to employing these techniques. Lysosomal membrane permeabilsation (LMP) was investigated using LysoTracker green staining by flow cytometry and Western blotting of microtubule-associated protein 1 light chain 3 (LC3) was quantified to assess autophagy induction. PEG-COOH AuNPs caused cytotoxicity in HEK 293 cells with an IC50 of 1 nM. PEG-COOH AuNPs were internalised by the HEK 293 cells and an increased number of vacuoles were observed. PEG-COOH AuNPs did not induce LMP despite the accumulation of the PEG-COOH AuNPs in vacuoles. No conversion of LC3I-LC3II was observed. However, LysoTracker green staining with flow cytometry is not an optimal assay as the auto-fluorescent properties of PEG-COOH AuNPs caused analytical interference. No interference of the PEG-COOH AuNPs with the Western blot protein quantification assay kit was observed. Investigation into the potential systemic effects of these particles is warranted due to their toxicity.

本数据集为完成人体生理学硕士学位而收集。该数据集旨在体外研究聚乙二醇羧基金纳米颗粒（PEG-COOH AuNPs）在人胚肾细胞（HEK-293）中的摄取、细胞毒性及可能的自噬诱导作用。首先，利用透射电子显微镜（TEM）、吸收光谱法、zeta电位仪和pH计，表征了PEG-COOH AuNPs在细胞培养基和超纯水（milli-Q水）中悬浮时的理化性质。随后，将HEK-293细胞暴露于浓度为0.5 nM至10 nM的PEG-COOH AuNPs中24小时，并使用xCELLigence实时细胞分析仪（RTCA）评估细胞毒性。采用CytoViva暗场显微镜评估浓度为0.13 nM的PEG-COOH AuNPs的细胞摄取情况。通过TEM观察PEG-COOH AuNPs进入囊泡的内化过程。在使用这些技术前，评估了PEG-COOH AuNPs对LysoTracker绿色染色染料和蛋白质印迹（Western blot）定量检测试剂盒的分析干扰。通过流式细胞术检测LysoTracker绿色染色情况以研究溶酶体膜通透性（LMP），并通过定量检测微管相关蛋白1轻链3（LC3）的蛋白质印迹结果来评估自噬诱导作用。PEG-COOH AuNPs对HEK-293细胞具有细胞毒性，半数抑制浓度（IC50）为1 nM。HEK-293细胞可内化PEG-COOH AuNPs，且观察到液泡数量增加。尽管PEG-COOH AuNPs在液泡中积累，但并未诱导LMP。未观察到LC3I向LC3II的转化。然而，由于PEG-COOH AuNPs的自发荧光特性会产生分析干扰，流式细胞术结合LysoTracker绿色染色并非最优检测方法。未观察到PEG-COOH AuNPs对蛋白质印迹定量检测试剂盒的干扰。鉴于这些颗粒的毒性，有必要进一步研究其潜在的全身效应。