Synergistic inhibition of calcium oxalate crystal formation and synergistic protection of HK-2 cells from crystal damage by sulfated Laminarin polysaccharide and potassium citrate

Abstract

Objective: The first objective is to study the synergistic inhibition of calcium oxalate (CaOx) formation by Laminarin polysaccharides (DLP and SDLP, before and after sulfation) and potassium citrate (K₃cit) and determine the synergistic protection of renal epithelial cells (HK-2 cells) caused by CaOx crystal damage. The second objective is to explore new ways to prevent and treat kidney stones. Methods: The CaOx crystals regulated by five additives (K₃cit group, DLP group, SDLP group, DLP-K₃cit synergistic group and SDLP-K₃cit synergistic group) were characterized by FT-IR, XRD, SEM, zeta potential, ICP, and TGA. The protective effect of each additive group on HK-2 cells damaged by nano-calcium oxalate monohydrate (nano-COM) was compared by detecting cell viability, the cell reactive oxygen species level, the cell survival rate, and mitochondrial membrane potential. Results: When DLP or SDLP acted synergically with K₃cit, the synergistic group induced the same amount of COD at a lower concentration or more COD formation at the same concentration, highlighting the synergistic enhancement effect of 1 + 1 > 2. At 0.3 g L^-1, the COD contents induced by DLP, SDLP, K₃cit, DLP-K₃cit, and SDLP-K₃cit synergistic groups were 20.3%, 75.8%, 75.4%, 87.3%, and 100%, respectively. The synergistic group increased the concentration of soluble Ca²⁺ ions in the supernatant, increased the absolute value of the zeta potential on the surface of CaOx crystals, and inhibited the aggregation among the crystals. TGA and DTG analyses established the adsorption of polysaccharides in the crystals. Cell experiments showed the ability of the synergistic group to significantly inhibit the damage of nano-COM crystals on HK-2 cells, reduce the level of reactive oxygen species and mortality, and improve cell viability and the mitochondrial membrane potential. Conclusions: The synergistic group can more effectively induce COD formation and cell protection than the standalone polysaccharide group or K₃cit group. The synergistic groups, especially SDLP-K₃cit, may be a potential drug for inhibiting the formation of CaOx kidney stones.