Various morphologies of calcium oxalate dihydrate (COD) crystals containing cross-shaped, flower-like, thin-bipyramid, thick-bipyramid, and elongated-bipyramid structures, with a size of approximately 5 μm, were prepared by varying the reactant concentration, reaction temperature, stirring speed, and additive.X-ray diffraction (XRD) and infrared spectroscopy (FT-IR) showed that the synthesized crystals were purephase COD crystals. The factors affecting the morphology of COD and the mechanism of COD formation were discussed. Physicochemical properties such as specific surface area, pore structure, pore size, zeta potential, and conductivity of the crystals were identified. The cytotoxicity of five different shapes of COD crystals in human kidney proximal tubular epithelial (HK-2) cells was investigated using the Cell Counting Kit-8 (CCK-8) assay and propidium iodide (PI) staining assay. The cytotoxicity was ranked in the following order: COD-elongated-bipyramid > COD-thick-bipyramid ≈ COD-cross-shaped > COD-thinbipyramid > COD-flower-like. The toxicity of tetragonal bipyramid COD crystals was closely correlated with the area of the Ca 2+ ion-rich (100) face; the crystals with large (100) faces showed a higher toxicity to HK-2 cells. The cytotoxicity of cross-shaped COD and flower-like COD was affected by the sharpness of their crystal edges; COD-CS with sharp corners caused more serious injuries to cell membranes. These findings are helpful in understanding the effects of mineralization conditions and additives on crystal morphology as well as to identify the relationship between crystal morphology and cytotoxicity. The results also provide a reference for the injury difference of various shapes of crystals from normal subjects and stone patients to the renal epithelial cells and the formation mechanism of calcium oxalate stones.

Corn silk polysaccharide (CSP0; molecular weight=124 kDa) was degraded by ultrasonication to obtain five degraded polysaccharides, namely, CSP1, CSP2, CSP3, CSP4, and CSP5, with molecular weights of 26.1, 12.2, 6.0, 3.5, and 2.0 kDa, respectively. The structures of these polysaccharides were characterized by FT-IR, 1H NMR, and 13C NMR analyses. The antioxidant activities, including scavenging ability for hydroxyl radicals and DPPH free radicals, chelation ability for Fe2+ ions, and reducing ability of CSP increased with decreased molecular weight of CSPs within 6.0 to 124 kDa. However, antioxidant activity weakened when the molecular weight of CSPs reached 3.5 and 2 kDa. CSP3 with a molecular weight of 6.0 kDa exhibited the strongest antioxidant activity. After protection with 60 μg/mL CSPs, the viability of human renal proximal tubular epithelial cells (HK-2) damaged by nano-COM crystals increased, the level of reactive oxygen species decreased, and the amount of COM crystal adhered onto the cell surface decreased. The ability of CSPs to protect cells from CaOx crystal damage was consistent with their antioxidant activity. CSPs can specifically combine with CaOx crystal to inhibit the conversion of calcium oxalate dihydrate crystal to calcium oxalate monohydrate crystal. All these results showed that the activity of CSPs was closely correlated with molecular weight. A very high or low molecular weight of CSPs was not conducive to their activity. CSPs, especially CSP3 with a molecular weight of 6.0 kDa, can be used as a potential antistone drug.

Abstract:The structure-activity relationships and repair mechanism of six low-molecular-weight seaweed polysaccharides (SPSs) on oxalate-induced damaged human kidney proximal tubular epithelial cells (HK-2) were investigated. These SPSs included Laminaria japonica polysaccharide, degraded Porphyra yezoensis polysaccharide, degraded Gracilaria lemaneiformis polysaccharide, degraded Sargassum fusiforme polysaccharide, Eucheuma gelatinae polysaccharide, and degraded Undaria pinnatifida polysaccharide. These SPSs have a narrow difference of molecular weight (from 1968 to 4020 Da) after degradation by controlling H 2 O 2 concentration. The sulfate group (-SO 3 H) content of the six SPSs was 21.7%, 17.9%, 13.3%, 8.2%, 7.0%, and 5.5%, respectively, and the -COOH contents varied between 1.0% to 1.7%. After degradation, no significant difference was observed in the contents of characteristic -SO 3 H and -COOH groups of polysaccharides. The repair effect of polysaccharides was determined using cell-viability test by CCK-8 assay and cell-morphology test by hematoxylin-eosin staining. The results revealed that these SPSs within 0.1-100 µg/mL did not express cytotoxicity in HK-2 cells, and each polysaccharide had a repair effect on oxalate-induced damaged HK-2 cells. Simultaneously, the content of polysaccharide -SO 3 H was positively correlated with repair ability. Furthermore, the low-molecular-weight degraded polysaccharides showed better repair activity on damaged HK-2 cells than their undegraded counterpart. Our results can provide reference for inhibiting the formation of kidney stones and for developing original anti-stone polysaccharide drugs.

This study aims to investigate the repair effect of subcellular structure injuries of the HK-2 cells of four degraded seaweed polysaccharides (DSPs), namely, the degraded Porphyra yezoensis, Gracilaria lemaneiformis, Sargassum fusiform, and Undaria pinnatifida polysaccharides. The four DSPs have similar molecular weight, but with different content of sulfate groups (i.e., 17.9%, 13.3%, 8.2%, and 5.5%, resp.). The damaged model was established using 2.8 mmol/L oxalate to injure HK-2 cells, and 60 μg/mL of various DSPs was used to repair the damaged cells. With the increase of sulfate group content in DSPs, the scavenging activity of radicals and their reducing power were all improved. Four kinds of DSPs have repair effect on the subcellular organelles of damaged HK-2 cells. After being repaired by DSPs, the release amount of lactate dehydrogenase was decreased, the integrity of cell membrane and lysosome increased, the Δψm increased, the cell of G1 phase arrest was inhibited, the proportion of S phase increased, and cell apoptotic and necrosis rates were significantly reduced. The greater the content of sulfate group is, the stronger is the repair ability of the polysaccharide. These DSPs, particularly the polysaccharide with higher sulfate group content, may be a potential drug for the prevention and cure of kidney stones.

Porphyra yezoensis polysaccharide repaired oxalate-injured renal epithelial cells and decreased COM crystal adhesion on the cell surface.