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. 2023 Feb 28;12(5):1031.
doi: 10.3390/foods12051031.

Antioxidant Activities and Cytotoxicity of the Regulated Calcium Oxalate Crystals on HK-2 Cells of Polysaccharides from Gracilaria lemaneiformis with Different Molecular Weights

Jing-Hong Liu  1 , Yu-Yun Zheng  1 , Jian-Ming Ouyang  1
Affiliations

Antioxidant Activities and Cytotoxicity of the Regulated Calcium Oxalate Crystals on HK-2 Cells of Polysaccharides from Gracilaria lemaneiformis with Different Molecular Weights

Jing-Hong Liu et al. Foods. .

Abstract

The antioxidant activities of seven degraded products (GLPs) with different molecular weights (Mw) of polysaccharides from Gracilaria lemaneiformis were compared. The Mw of GLP1-GLP7 were 106, 49.6, 10.5, 6.14, 5.06, 3.71 and 2.42 kDa, respectively. The results show that GLP2 with Mw = 49.6 kDa had the strongest scavenging capacity for hydroxyl radical, DPPH radical, ABTS radical and reducing power. When Mw < 49.6 kDa, the antioxidant activity of GLPs increased with the increase in Mw, but when Mw increased to 106 kDa, their antioxidant activity decreased. However, the ability of GLPs to chelate Fe2+ ions increased with the decrease in polysaccharide Mw, which was attributed to the fact that the polysaccharide active groups (-OSO3- and -COOH) were easier to expose, and the steric hindrance was smaller when GLPs chelated with Fe2+. The effects of GLP1, GLP3, GLP5 and GLP7 on the crystal growth of calcium oxalate (CaOx) were studied using XRD, FT-IR, Zeta potential and thermogravimetric analysis. Four kinds of GLPs could inhibit the growth of calcium oxalate monohydrate (COM) and induce the formation of calcium oxalate dihydrate (COD) in varying degrees. With the decrease in Mw of GLPs, the percentage of COD increased. GLPs increased the absolute value of the Zeta potential on the crystal surface and reduced the aggregation between crystals. Cell experiments showed that the toxicity of CaOx crystal regulated by GLPs to HK-2 cells was reduced, and the cytotoxicity of CaOx crystal regulated by GLP7 with the smallest Mw was the smallest, which was consistent with the highest SOD activity, the lowest ROS and MDA levels, the lowest OPN expression level and the lowest cell necrosis rate. These results suggest that GLPs, especially GLP7, may be a potential drug for the prevention and treatment of kidney stones.

Keywords: Gracilaria lemaneiformis polysaccharide; antioxidant activity; calcium oxalate; cytotoxicity; molecular weight.

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Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Figure 1
Figure 1
Antioxidant activities of Gracilaria lemaneiformis polysaccharides with different molecular weights. (A) Scavenging ·OH free radicals; (B) Scavenging DPPH radical; (C) Scavenging ABTS free radicals; (D) Reducing power; (E) Fe2+ chelating ability; (F) Comparison of antioxidant activity of each polysaccharide at 3.0 mg/mL. Values are means ± S.D (n = 3).
Figure 1
Figure 1
Antioxidant activities of Gracilaria lemaneiformis polysaccharides with different molecular weights. (A) Scavenging ·OH free radicals; (B) Scavenging DPPH radical; (C) Scavenging ABTS free radicals; (D) Reducing power; (E) Fe2+ chelating ability; (F) Comparison of antioxidant activity of each polysaccharide at 3.0 mg/mL. Values are means ± S.D (n = 3).
Figure 2
Figure 2
XRD spectrum (A) and percentage content of COD in CaOx crystals (B) regulated by GLPswith concentration of 1.0 g/L. (a) Crystal control group without polysaccharides; (b) GLP1; (c) GLP3; (d) GLP5; (e) GLP7. The crystal plane with asterisk shows COD, and the crystal plane without asterisk shows COM. The Mw of GLP1, GLP3, GLP5 and GLP7 was 106, 10.5, 5.06 and 2.42 kDa respectively.
Figure 3
Figure 3
SEM images of the CaOx crystals regulated by GLPs with concentration of 1.0 g/L. (a) Crystal control group without polysaccharides; (b) GLP1; (c) GLP3; (d) GLP5; (e) GLP7. c (CaOx) = 10 mmol/L. The red arrows refer to COM crystals, and the white arrows refer to COD crystals.
Figure 4
Figure 4
Zeta potential of the CaOx crystals regulated by GLPs with concentration of 1.0 g/L. Compared with DC, * p < 0.05; ** p < 0.01.
Figure 5
Figure 5
TGA (A) and DTG (B) curves of the CaOx crystals regulated by GLPs with concentration of 1.0 g/L. Stage I corresponds to the loss of bound water in COM or COD; Stage II corresponds to the loss of polysaccharides; Stage III corresponds to the loss of CO in CaC2O4 and Stage IV corresponds to the loss of CO2 in CaCO3.
Figure 6
Figure 6
Effects of CaOx crystals regulated by GLPs at a concentration of 1.0 g/L on the viability of HK-2 cells. Crystal concentration: 200 μg/mL; time: 6 h. NC: normal control group; DC: crystal control group without polysaccharides. Compared with DC, * p < 0.05; ** p < 0.01.
Figure 7
Figure 7
Effects of CaOx crystals regulated by GLPs at a concentration of 1.0 g/L on the SOD activity (a) and the amount of MDA (b) in HK-2 cells. GLPs concentration: 1.0 g/L. Compared with DC, * p < 0.05; ** p < 0.01.
Figure 8
Figure 8
Effects of CaOx crystals regulated by GLPs at a concentration of 1.0 g/L on the expression of OPN molecules on the surface of HK-2 cells. Scale: 20 μm. In the figure, blue fluorescence represents the nucleus, and green fluorescence represents the OPN molecule. (A) OPN fluorescence map; (B) fluorescence intensity quantitative diagram. The Mw of GLP1, GLP3, GLP5 and GLP7 was 106, 10.5, 5.06 and 2.42 kDa, respectively. NC: normal control group; DC: crystal control group without polysaccharides. Compared with DC, ** p < 0.01.
Figure 9
Figure 9
Effects of CaOx crystals regulated by GLPs at a concentration of 1.0 g/L on ROS level in HK-2 cells. (A) ROS fluorescence map; (B) fluorescence intensity quantitative diagram. The Mw of GLP1, GLP3, GLP5 and GLP7 was 106, 10.5, 5.06 and 2.42 kDa, respectively. NC: normal control group; DC: crystal control group without polysaccharides. Scale: 50 μm. Compared with DC, ** p < 0.01.
Figure 10
Figure 10
Effects of CaOx crystals regulated by GLPs at a concentration of 1.0 g/L on apoptosis and necrosis of HK-2 cells. (A) PI fluorescence map; (B) fluorescence intensity quantitative diagram. The Mw of GLP1, GLP3, GLP5 and GLP7 was 106, 10.5, 5.06 and 2.42 kDa respectively. NC: normal control group; DC: crystal control group without polysaccharides. Scale: 20 μm. Compared with DC, ** p < 0.01.
Figure 11
Figure 11
Model diagram of GLPs antioxidant reducing the cytotoxicity of CaOx crystals on HK-2 cells. The left side of the diagram is the toxic effect of CaOx regulated by high Mw GLP1 on the cells, where the formed COM crystals are more toxic and cause cell damage and cell contraction. The right side is the toxic effect of CaOx regulated by low Mw GLP7 on the cells, where the mainly formed COD crystals have less toxicity and less damage to the cells. The cell morphology is basically normal.
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References

    1. Zeng G., Mai Z., Xia S., Wang Z., Zhang K., Wang L., Long Y., Ma J., Li Y., Wan S.P., et al. Prevalence of kidney stones in China: An ultrasonography based cross-sectional study. BJU Int. 2017;120:109–116. doi: 10.1111/bju.13828. - DOI - PubMed
    1. Lai Y., Zheng H., Sun X., Lin J., Li Q., Huang H., Hou Y., Zhong H., Zhang D., Fucai T., et al. The advances of calcium oxalate calculi associated drugs and targets. Eur. J. Pharmacol. 2022;935:175324. doi: 10.1016/j.ejphar.2022.175324. - DOI - PubMed
    1. Kvsrg P., Sujatha D., Bharathi K. Herbal drugs in urolithiasis-a review. Pharmacogn. Rev. 2007;1:175–178.
    1. Wang Z., Zhang Y., Zhang J., Deng Q., Liang H. Recent advances on the mechanisms of kidney stone formation. Int. J. Mol. Med. 2021;48:149. doi: 10.3892/ijmm.2021.4982. - DOI - PMC - PubMed
    1. Sun X.Y., Ouyang J.M., Li Y.B., Wen X.L. Mechanism of cytotoxicity of micron/nano calcium oxalate monohydrate and dihydrate crystals on renal epithelial cells. RSC Adv. 2015;5:45393–45406. doi: 10.1039/C5RA02313K. - DOI