CULTURED PROXIMAL TUBULAR CELLS SHOW NANOPLASTIC UPTAKE AND IMPAIRMENTS IN GROWTH AND CELL CYCLE
Mr HAYDEN GILLINGS1,2, PROF JONATHAN M GLEADLE2,3, DR DARLING M ROJAS-CANALES2,3, DR SOON WEI WONG3, A/PROF MELANIE MACGREGOR4,5
1College of Science and Engineering, Flinders University, Bedford Park, Australia, 2College of Medicine and Public Health, Flinders University, Bedford Park, Australia, 3Department of Renal Medicine, Flinders Medical Centre, Bedford Park, Australia, 4Nano Microplastics Research Consortium, Bedford Park, Australia, 5Flinders Institute for Nanoscale Science and Technology, Bedford Park, Australia
Aim: To determine effects of short- and long-term exposure to polystyrene nanoplastics (NPs) in human kidney-2 (HK-2) proximal tubular cells (PTC).
Background: NPs are plastic particles less than 100 nm in size, typically generated through the degradation of plastic waste. Knowledge of their exposure effects on the kidney are extremely limited.
Methods: The HK-2 cell line was exposed to 0.2 to 200 µg/mL of 20 nm and 100 nm spherical NPs for 24-hours, or at a concentration of 100 µg/mL every 72-hours for a total 24 days. Cell viability and cell cycle analysis was completed by flow cytometry. Fluorescent and phase contrast microscopy was used to determine NP internalisation and cell morphology changes respectively.
Results: The viability of HK-2 cells was not significantly affected by exposure to 20 nm and 100 nm NPs, with only 1-2% increase in Annexin-V positive cells. However, cell cycle analysis revealed increased entry into S-phase (n=3, P <0.01) with the 20 nm NPs, while no significant changes were seen with the 100 nm particles. Fluorescent NP internalisation studies showed uptake and accumulation within the cytoplasm after 24-hours, whilst little morphological changes were observed. Long-term exposure (24-days) showed gross morphological changes, with decreased total cell numbers and increased doubling times, most evident in cells exposed to 20 nm NPs.
Conclusions: HK-2 PTC are able to internalise NPs. The most significant changes were observed with the 20nm particles, which showed reduced proliferation despite S-phase entry, suggestive of S-phase cell cycle arrest and possible DNA damage. This however did not affect cell viability. Further research is needed to characterise the toxic effects of NPs less than 100nm on kidney health and function.
Biography:
Hayden Gillings is a PhD student at Flinders University studying the health impacts of sub-micron particles called nanoplastics. He has a background in medical biotechnology, with particular interest in cell response, genetics, and bioinformatics, having previously investigated the use of natural extracts, derived from Australian plants and marine invertebrates, as a way to inhibit the invasion ability of glioblastoma. Hayden, now in the second year of his PhD, is currently researching how chronic, repeated exposure to nanoplastics could impact the health and function of kidneys.