Carboxymethylated Desmodium styracifolium polysaccharide reduces the risk of calcium oxalate kidney stone formation by inhibiting crystal adhesion and promoting crystal endocytosis.

The inhibition of cell surface crystal adhesion and an appropriate increase in crystal endocytosis contribute to the inhibition of kidney stone formation. In this study, we investigated the effects of different degrees of carboxymethylation on these processes. An injury model was established by treating human renal proximal tubular epithelial (HK-2) cells with 98.3 ± 8.1 nm calcium oxalate dihydrate (nanoCOD) crystals. The HK-2 cells were protected with carboxy (-COOH) Desmodium styracifolium polysaccharides at 1.17% (DSP0), 7.45% (CDSP1), 12.2% (CDSP2), and 17.7% (CDSP3). Changes in biochemical indexes and effects on nanoCOD adhesion and endocytosis were detected. The protection of HK-2 cells from nanoCOD-induced oxidative damage by carboxymethylated Desmodium styracifolium polysaccharides (CDSPs) is closely related to the protection of subcellular organelles, such as mitochondria. CDSPs can reduce crystal adhesion on the cell surface and maintain appropriate crystal endocytosis, thereby reducing the risk of kidney stone formation. CDSP2 with moderate -COOH content showed the strongest protective activity among the CDSPs.

Carboxymethylation of Desmodium styracifolium Polysaccharide and Its Repair Effect on Damaged HK-2 Cells.

Objective: Desmodium styracifolium is the best traditional medicine for treating kidney calculi in China. This study is aimed at increasing the carboxyl (-COOH) content of D. styracifolium polysaccharide (DSP0) and further increasing its antistone activity. Methods: DSP0 was carboxymethylated with chloroacetic acid at varying degrees. Then, oxalate-damaged HK-2 cells were repaired with modified polysaccharide, and the changes in biochemical indices before and after repair were detected. Results: Three modified polysaccharides with 7.45% (CDSP1), 12.2% (CDSP2), and 17.7% (CDSP3) -COOH are obtained. Compared with DSP0 (-COOH content = 1.17%), CDSPs have stronger antioxidant activity in vitro and can improve the vitality of damaged HK-2 cells. CDSPs repair the cell morphology and cytoskeleton, increase the cell healing ability, reduce reactive oxygen species and nitric oxide levels, increase mitochondrial membrane potential, limit autophagy level to a low level, reduce the eversion of phosphatidylserine in the cell membrane, weaken the inhibition of oxalate on DNA synthesis, restore cell cycle to normal state, promote cell proliferation, and reduce apoptosis/necrosis. Conclusion: The carboxymethylation modification of DSP0 can improve its antioxidant activity and enhance its ability to repair damaged HK-2 cells. Among them, CDSP2 with medium -COOH content has the highest activity of repairing cells, whereas CDSP3 with the highest -COOH content has the highest antioxidant activity. This difference may be related to the active environment of polysaccharide and conformation of the polysaccharide and cell signal pathway. This result suggests that Desmodium styracifolium polysaccharide with increased -COOH content may have improved potential treatment and prevention of kidney calculi.

Sulfated Undaria pinnatifida polysaccharide inhibits the formation of kidney stones by inhibiting HK-2 cell damage and reducing the adhesion of nano­calcium oxalate crystals.

OBJECTIVE: The formation of kidney stone is closely related to cell injury and crystal adhesion. METHOD: The sulfur trioxide-pyridine method was used to sulfate raw Undaria pinnatifida polysaccharide (UPP) with a molecular weight (Mw) of 8.33 kDa. Four polysaccharides with the sulfate group (-OSO3-) contents of 1.59% (UPP0), 6.03% (UPP1), 20.83% (UPP2), and 36.39% (UPP3) were obtained. The antioxidant activity of the four UPPs, the difference in oxidative damage inflicted by nano-CaOx monohydrate (nano-COM) on human proximal tubular epithelial (HK-2) cells before and after protection by UPPs, and the inhibitory effect on nano-COM adhesion were explored. RESULTS: Structural characterization showed that sulfation was successful. As the -OSO3- content in the UPPs was increased, the antioxidant activity and capability of the UPPs to regulate the growth of calcium oxalate (CaOx) crystals gradually increased. The damage caused by nano-COM crystals to HK-2 cells under protection by UPPs was weakened. This effect enhanced cell viability, enabled the maintenance of good cell morphology, reduced reactive oxygen species (ROS) levels, and inhibited the decrease in mitochondrial membrane potential, as well as decreased the eversion of phosphatidylserine (PS) and the expression of the adhesion proteins osteopontin (OPN), heat shock protein (HSP 90), and Annexin A1 (ANXA1). The adhesion of nano-COM to HK-2 cells was inhibited under the protection by UPPs. CONCLUSION: UPP3 with the highest content of -OSO3- presented the best antioxidant activity and crystal regulation ability, while UPP2 with the second highest -OSO3- content showed optimal cell protection ability and crystal adhesion inhibition ability. The biological activity of UPPs was regulated by Mw and -OSO3- content. UPP2 with moderate -OSO3- content may become a potential drug for preventing CaOx stones.

Regulation of Laminaria Polysaccharides with Different Degrees of Sulfation during the Growth of Calcium Oxalate Crystals and their Protective Effects on Renal Epithelial Cells.

The original Laminaria polysaccharide (LP0) was sulfated using the sulfur trioxide-pyridine method, and four sulfated Laminaria polysaccharides (SLPs) were obtained, namely, SLP1, SLP2, SLP3, and SLP4. The sulfated (-OSO3 -) contents were 8.58%, 15.1%, 22.8%, and 31.3%, respectively. The structures of the polysaccharides were characterized using a Fourier transform infrared (FT-IR) spectrometer and nuclear magnetic resonance (NMR) techniques. SLPs showed better antioxidant activity than LP0, increased the concentration of soluble Ca2+ in the solution, reduced the amount of CaOx precipitation and degree of CaOx crystal aggregation, induced COD crystal formation, and protected HK-2 cells from damage caused by nanometer calcium oxalate crystals. These effects can inhibit the formation of CaOx kidney stones. The biological activity of the polysaccharides increased with the content of -OSO3 -, that is, the biological activities of the polysaccharides had the following order: LP0 < SLP1 < SLP2 < SLP3 < SLP4. These results reveal that SLPs with high -OSO3 - contents are potential drugs for effectively inhibiting the formation of CaOx stones.

Carboxymethylation of Corn Silk Polysaccharide and Its Inhibition on Adhesion of Nanocalcium Oxalate Crystals to Damaged Renal Epithelial Cells.

The purpose of this study was to explore the repair effect of carboxymethyl-modified corn silk polysaccharide (CSP) on oxidatively damaged renal epithelial cells and the difference in adhesion between cells and calcium oxalate crystals. The CSP was degraded and modified through carboxymethylation. An oxidatively damaged cell model was constructed by oxalate damage to human kidney proximal tubular epithelial (HK-2) cells. Then, the damaged cells were repaired by modified polysaccharides, and the changes in biochemical indexes and adhesion ability between cells and crystals before and after repair were detected. Four modified polysaccharides with carboxyl group (-COOH) contents of 3.92% (CSP0), 7.75% (CCSP1), 12.90% (CCSP2), and 16.38% (CCSP3) were obtained. Compared with CSP0, CCSPs had stronger antioxidant activity, could repair damaged HK-2 cells, and could reduce phosphorylated serine eversion on the cell membrane, the expression of osteopontin (OPN) and Annexin A1, and crystal adhesion. However, its effect on the expression of hyaluronic acid synthase was not substantial. The carboxymethyl modification of the CSP can improve its ability to repair cells and inhibit crystal adhesion and aggregation. A high carboxymethylation degree results in strong polysaccharide activity. CCSPs are expected to reduce the risk of kidney stone formation and recurrence.