The research on autophagy of human renal tubular epithelial cells induced by calcifying nanoparticles (CNP) / 中华泌尿外科杂志

ABSTRACT

Objective To observe the effects of different concertrations of calcifying nanoparticles (CNP) on growth and proliferation of human renal tubular epithelial cells (HK-2),and to explore the effects of autophagy of human renal tubular epithelial cells induced by CNP. MethodsNormal growth HK-2 cells were cultured in vitro and the culture medium was changed to serum-free medium after cell growth to confluence,CNP with different concentrations were then added and the cell proliferation inhibition was measured by MTT assay after 12,24,48 and 72 h co-incubation.EGFP-LC3 express plasmid was transfected into HK-2 cells,CNP with certain concentration was then added after 24h incubation.The formation of autophagosomes and their adherence to cells were observed by electron microscopy,and EGFP-LC3 staining was used to detect the formation of autophagosome by flourescence microscopy after 3,6,24 and 48 h co-incubation. ResultsCNP presented a concentration-dependent cell proliferation inhibition on HK-2 cells,the inhibitory ratio in cell proliferation was 14.5 ％ after 48 h co-incubation when CNP density was 0.05 mg/ml,and the inhibitory ratio was 21.5％ after 72 h co-incubation when CNP density was 0.1 mg/ml.Autophagosome formation and CNP adherence to cell surface could be observed by electron microscopy 6 h after CNP was added to the DMEM medium.The LC3-positive dots were observed in HK-2 cells incubated with CNP.Conclusions Autophagy of HK-2 could be activated by co-incubation with CNP,which may play an important role in the formation of nephrolithiasis.