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.

本研究收集于完成人类生理学硕士学位期间的数据。该数据集旨在探究聚乙二醇羧基金纳米颗粒（PEG-COOH AuNPs）在体外人胚胎肾（HEK-293）细胞中的摄取、细胞毒性及其可能诱导的自噬作用。PEG-COOH AuNPs首先通过透射电子显微镜（TEM）、吸光度光谱、ζ电位计和pH计等手段，对其在细胞培养介质和 Milli-Q 水中的物理化学性质进行了表征。随后，将HEK-293细胞暴露于不同浓度的PEG-COOH AuNPs（0.5 nM - 10 nM）中24小时，并使用xCELLigence实时细胞分析仪（RTCA）评估其细胞毒性。利用CytoViva暗场显微镜评估了PEG-COOH AuNPs（0.13 nM）的细胞摄取。通过TEM对PEG-COOH AuNPs进入囊泡的内部化进行了可视化。在应用这些技术之前，评估了PEG-COOH AuNPs与LysoTracker绿色染色染料和Western blot蛋白质定量试剂盒的潜在分析干扰。利用LysoTracker绿色染色通过流式细胞术和微管相关蛋白1轻链3（LC3）的Western blot分析，研究了溶酶体膜通透性（LMP）。PEG-COOH AuNPs在HEK-293细胞中表现出细胞毒性，IC50值为1 nM。PEG-COOH AuNPs被HEK-293细胞摄取，并观察到空泡数量增加。尽管PEG-COOH AuNPs在空泡中积累，但并未诱导LMP。未观察到LC3I-LC3II的转化。然而，由于PEG-COOH AuNPs的自发光特性导致了分析干扰，因此使用流式细胞术进行LysoTracker绿色染色并不是一个理想的检测方法。未观察到PEG-COOH AuNPs对Western blot蛋白质定量试剂盒的干扰。鉴于这些粒子的毒性，有必要调查其潜在的系统性影响。