Preliminary Evaluation of Nanobacteria on Crystal Retention, CaSR, and Claudin-14 Expression in HK-2 Cells

Abstract

To evaluate cell morphology, crystal adhesion, cell damage, Calcium sensitive receptor (CaSR), and Claudin protein-14 (Claudin-14) expression at different time intervals and explore the role of nanobacteria in the formation of urinary calculi. In this experiment, HK-2 cells were cocultured with nanobacteria (NB) in the absence or presence of tetracycline (Tet). Cells treated with calcium oxalate monohydrate (COM) crystals were used as a positive control of urinary stone-induced cell damage. After which, cell morphology was evaluated by hematoxylin-eosin staining in comparison to untreated HK-2 cells (negative control). Use different methods to assess cell damage, crystal adhesion, and protein expression. (The degree of cell damage, crystal adhesion, and protein expression were evaluated by various methods). It was found that the degree of cell damage observed in Tet + NB-treated cells was significantly lower than that in NB-treated cells. Lactate dehydrogenase (LDH) leakage was higher in COM-exposed than in control cells ( $P<0.05$ ). However, LDH release from both NB- and Tet + NB-treated cells was significantly lower than from COM-treated cells ( $P<0.05$ ). The relative expression of CaSR and Claudin-14 proteins was higher in NB, COM, and TET + NB cells than in control cells ( $P<0.001$ ) and was lower in Tet + NB than in NB cells ( $P<0.01$ ). And $P<0.05$ means that the difference was statistically significant, and $P<0.001$ means that there was a significant difference between the both things. From the cell morphology, the cell damage in the COM group was greater than that in the NB group, and the cell damage markers in the COM group and the NB group were elevated. NB caused damage to HK-2 cells by inducing lipid peroxidation, and the degree of damage was increased in processing time. The adhesion of HK-2 cells to COM crystals increased after injury and was proportional to the duration of NB coculture. NB upregulated the expression of CaSR and Claudin-14 in HK-2 cells.