Limitation of Water-Soluble Tetrazolium Salt for the Cytocompatibility Evaluation of Zinc-Based Metals

Overview

Journal Materials (Basel)

Publisher MDPI

Date 2021 Nov 13

PMID 34771776

Citations 3

Authors

Peijun Zhu

Jiahao Chen

Ping Li

Shulan Xu

Affiliations

Soon will be listed here.

Abstract

Zinc (Zn) and its alloys have been regarded as promising biodegradable metals. The standardized cytotoxicity evaluation is a mandatory step to screen the biocompatibility of novel Zn and its alloys. Nevertheless, the suitability of the tetrazolium-based assay in the direct contact test for some metallic biomaterials (i.e., magnesium and manganese) is questionable. In this study, our results demonstrate an obvious inconsistency between qualitative observation via fluorescence staining and quantitative assessment using water-soluble tetrazolium salt (CCK-8). Subsequent experiments revealed that Zn and pre-treated Zn can directly convert tetrazolium salts to formazan, falsifying the cytotoxicity results. Therefore, we conclude that the CCK-8 assay is not suitable for evaluating the cytotoxicity of biodegradable Zn-based metals in the direct contact test.

Citing Articles

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References

Lin J, Tong X, Shi Z, Zhang D, Zhang L, Wang K . A biodegradable Zn-1Cu-0.1Ti alloy with antibacterial properties for orthopedic applications. Acta Biomater. 2020; 106:410-427. DOI: 10.1016/j.actbio.2020.02.017. View

Yang H, Wang C, Liu C, Chen H, Wu Y, Han J . Evolution of the degradation mechanism of pure zinc stent in the one-year study of rabbit abdominal aorta model. Biomaterials. 2017; 145:92-105. DOI: 10.1016/j.biomaterials.2017.08.022. View

Pierson D, Edick J, Tauscher A, Pokorney E, Bowen P, Gelbaugh J . A simplified in vivo approach for evaluating the bioabsorbable behavior of candidate stent materials. J Biomed Mater Res B Appl Biomater. 2011; 100(1):58-67. DOI: 10.1002/jbm.b.31922. View

Drynda A, Hassel T, Bach F, Peuster M . In vitro and in vivo corrosion properties of new iron-manganese alloys designed for cardiovascular applications. J Biomed Mater Res B Appl Biomater. 2014; 103(3):649-60. DOI: 10.1002/jbm.b.33234. View

Fu J, Su Y, Qin Y, Zheng Y, Wang Y, Zhu D . Evolution of metallic cardiovascular stent materials: A comparative study among stainless steel, magnesium and zinc. Biomaterials. 2019; 230:119641. PMC: 6934082. DOI: 10.1016/j.biomaterials.2019.119641. View

Zhang W, Li P, Neumann B, Haag H, Li M, Xu Z . Chandler-Loop surveyed blood compatibility and dynamic blood triggered degradation behavior of Zn-4Cu alloy and Zn. Mater Sci Eng C Mater Biol Appl. 2020; 119:111594. DOI: 10.1016/j.msec.2020.111594. View

Su Y, Cockerill I, Wang Y, Qin Y, Chang L, Zheng Y . Zinc-Based Biomaterials for Regeneration and Therapy. Trends Biotechnol. 2018; 37(4):428-441. PMC: 6421092. DOI: 10.1016/j.tibtech.2018.10.009. View

Yang H, Jia B, Zhang Z, Qu X, Li G, Lin W . Alloying design of biodegradable zinc as promising bone implants for load-bearing applications. Nat Commun. 2020; 11(1):401. PMC: 6972918. DOI: 10.1038/s41467-019-14153-7. View

Shearier E, Bowen P, He W, Drelich A, Drelich J, Goldman J . In Vitro Cytotoxicity, Adhesion, and Proliferation of Human Vascular Cells Exposed to Zinc. ACS Biomater Sci Eng. 2016; 2(4):634-642. PMC: 5102339. DOI: 10.1021/acsbiomaterials.6b00035. View

10.

Yang Y, Cheng Y, Peng S, Xu L, He C, Qi F . Microstructure evolution and texture tailoring of reduced graphene oxide reinforced Zn scaffold. Bioact Mater. 2020; 6(5):1230-1241. PMC: 7653236. DOI: 10.1016/j.bioactmat.2020.10.017. View

11.

Guo H, Xia D, Zheng Y, Zhu Y, Liu Y, Zhou Y . A pure zinc membrane with degradability and osteogenesis promotion for guided bone regeneration: In vitro and in vivo studies. Acta Biomater. 2020; 106:396-409. DOI: 10.1016/j.actbio.2020.02.024. View

12.

Scarcello E, Lambremont A, Vanbever R, Jacques P, Lison D . Mind your assays: Misleading cytotoxicity with the WST-1 assay in the presence of manganese. PLoS One. 2020; 15(4):e0231634. PMC: 7161962. DOI: 10.1371/journal.pone.0231634. View

13.

Al Hegy A, Smith R, Gauthier E, Gray-Munro J . Investigation of a cyanine dye assay for the evaluation of the biocompatibility of magnesium alloys by direct and indirect methods. Bioact Mater. 2020; 5(1):26-33. PMC: 6957867. DOI: 10.1016/j.bioactmat.2019.12.002. View

14.

Li P, Zhang W, Dai J, Xepapadeas A, Schweizer E, Alexander D . Investigation of zinc‑copper alloys as potential materials for craniomaxillofacial osteosynthesis implants. Mater Sci Eng C Mater Biol Appl. 2019; 103:109826. DOI: 10.1016/j.msec.2019.109826. View

15.

Bowen P, Drelich J, Goldman J . Zinc exhibits ideal physiological corrosion behavior for bioabsorbable stents. Adv Mater. 2013; 25(18):2577-82. DOI: 10.1002/adma.201300226. View

16.

Li P, Schille C, Schweizer E, Rupp F, Heiss A, Legner C . Mechanical Characteristics, In Vitro Degradation, Cytotoxicity, and Antibacterial Evaluation of Zn-4.0Ag Alloy as a Biodegradable Material. Int J Mol Sci. 2018; 19(3). PMC: 5877616. DOI: 10.3390/ijms19030755. View

17.

capek J, Kubasek J, Pinc J, Fojt J, Krajewski S, Rupp F . Microstructural, mechanical, in vitro corrosion and biological characterization of an extruded Zn-0.8Mg-0.2Sr (wt%) as an absorbable material. Mater Sci Eng C Mater Biol Appl. 2021; 122:111924. DOI: 10.1016/j.msec.2021.111924. View

18.

Li P, Schille C, Schweizer E, Kimmerle-Muller E, Rupp F, Han X . Evaluation of a Zn-2Ag-1.8Au-0.2V Alloy for Absorbable Biocompatible Materials. Materials (Basel). 2019; 13(1). PMC: 6981962. DOI: 10.3390/ma13010056. View

19.

Semisch A, Hartwig A . Copper ions interfere with the reduction of the water-soluble tetrazolium salt-8. Chem Res Toxicol. 2014; 27(2):169-71. DOI: 10.1021/tx400414c. View

20.

Li P, Zhang W, Spintzyk S, Schweizer E, Krajewski S, Alexander D . Impact of sterilization treatments on biodegradability and cytocompatibility of zinc-based implant materials. Mater Sci Eng C Mater Biol Appl. 2021; 130:112430. DOI: 10.1016/j.msec.2021.112430. View