Skip to main content
Log in

In vitro toxicity of various classes of test agents using the neutral red assay on a human three-dimensional physiologic skin model

  • Regular Papers
  • Published:
In Vitro Cellular & Developmental Biology - Animal Aims and scope Submit manuscript

Summary

A new three-dimensional human skin model consisting of several layers of actively dividing and metabolically active human neonatal foreskin-derived fibroblasts and epidermal keratinocytes grown on nylon mesh has been used to assess the in vitro toxicity of test agents from various classes. Utilizing a slight modification of the published neutral red viability assay for endpoint determination, we have assayed and obtained dose-dependent toxicity curves for test agents from the following classes: detergents (n=15), alcohols (n=5), metal chlorides (n=10), perfumes and colognes (n=5), shampoos (n=4), conditioners (n=3), moisturizers (n=3), pesticides (n=3), and antimicrobial preservatives (n=4). Limited comparisons to in vivo ocular irritancy data with alcohols and detergents are encouraging. We have demonstrated the utility of this metabolically active dermal substrate containing naturally secreted collagen and other extracellular matrix proteins along with the neutral red viability assay for assessing the toxicity of a number of test agents from a variety of different classes with broad industrial applications.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  1. Babich, H.; Sardana, M. K.; Borenfreund, E. Acute cytotoxicities of polynuclear aromatic hydrocarbons determinedin vitro with the human liver tumor cell line, HepG2. Cell Biol. Toxicol. 4:295–309; 1988.

    Article  PubMed  CAS  Google Scholar 

  2. Babich, H.; Borenfreund, E. Structure-activity relationships for diorganotins, chlorinated benzenes and chlorinated anilines established with bluegill sunfish BF-2 cells. Fundam. Appl. Toxicol. 10:295–301; 1988.

    Article  PubMed  CAS  Google Scholar 

  3. Babich, H.; Martin-Alguacil, N.; Borenfreund, E. Comparisons of the cytotoxicities of dermatotoxicants to human keratinocytes and fibroblastsin vitro. In: Goldberg, A. M., ed. Alternative methods in toxicology, vol. 7. New York: MaryAnn Liebert, Inc.; 1989:153–167.

    Google Scholar 

  4. Bohmont, B. L. The new pesticide user’s guide. Reston, VA: Reston Publishing Co., Inc.; 1983.

    Google Scholar 

  5. Booman, K. A.; Cascieri, T. M.; Demetrulias, J., et al.In vitro methods for estimating eye irritancy of cleaning products. Phase I: preliminary assessment. J. Toxicol. Cutaneous Ocular Toxicol. 7(3):173–185; 1988.

    CAS  Google Scholar 

  6. Borenfreund, E.; Puerner, J. A simple quantitative procedure using monolayer cultures for cytotoxicity assays (HTD/NR-90). J. Tissue Cult. Methods 9(1):7–9; 1984.

    Article  Google Scholar 

  7. Borenfreund, E.; Shopsis, C. Toxicity monitored with a correlated set of cell-culture assays. Xenobiotica 15:705–711; 1985.

    Article  PubMed  CAS  Google Scholar 

  8. Borenfreund, E.; Puerner, J. A. Cytotoxicity of metals, metal-metal and metal-chelator combinations assayedin vitro. Toxicology 39:121–124; 1986.

    Article  PubMed  CAS  Google Scholar 

  9. Borenfreund, E.; Babich, H.In vitro cytotoxicity of heavy metals, acrylamide and organotin salts to neural cells and fibroblasts. Cell Biol. Toxicol. 3:63–73; 1987.

    Article  PubMed  CAS  Google Scholar 

  10. Brown, C. D.; Li, Q.; Brabec, M. J. Cytotoxicity of six divalent metals to testicular cellsin vitro. In Vitro Toxicol. 2:129–137; 1988/1989.

    CAS  Google Scholar 

  11. Coin, P. G.; Stevens, J. B. Toxicity of cadmium chloridein vitro: indices of cytotoxicity with the pulmonary alveolar macrophage. Toxicol. Appl. Pharmacol. 82:140–150; 1986.

    Article  PubMed  CAS  Google Scholar 

  12. Gordon, V. C.; Bergman, H. C. EYTEX™, an in vitro method for evaluation of ocular irritancy. In: Goldberg, A. M., ed. Alternative methods in toxicology, vol. 5. New York: MaryAnn Liebert, Inc.; 1987:293–302.

    Google Scholar 

  13. Hammond, M. E.; Goodwin, J.; Dvorak, H. F. Quantitative measurements of neutral red uptake and excretion by mammalian cells. J. Reticuloendothel. Soc. 27:337–346; 1980.

    PubMed  CAS  Google Scholar 

  14. Jacques, P. J. Endocytosis. In: Dingle, J. T.; Fell, H. B., eds. Lysosomes in biology and pathology. Amsterdam: North Holland; 1973:395–420.

    Google Scholar 

  15. Muir, C. K. Surfactant-induced opacity of bovine isolated cornea: an epithelial phenomenon? Toxicol. Lett. 38:51–54; 1987.

    Article  PubMed  CAS  Google Scholar 

  16. Naughton, G. K.; Jacob, L.; Naughton, B. A. A physiological skin model forin vitro toxicity studies. In: Goldberg, A. M., ed. Alternative methods in toxicology, vol. 7. New York: MaryAnn Liebert, Inc.; 1989:183–189.

    Google Scholar 

  17. Ng, T. B.; Liu, W. K. Toxic effects of heavy metals on cells isolated from the rat adrenal and testis. In Vitro Cell. Dev. Biol. 26:24–28; 1990.

    Article  PubMed  CAS  Google Scholar 

  18. North-Root, H.; Yackovich, F.; Demetrulias, J., et al. Prediction of the eye irritation potential of shampoos using thein vitro SIRC toxicity test. Food Chem. Toxicol. 23:271–273; 1985.

    Article  PubMed  CAS  Google Scholar 

  19. Sewell, R. B.; Soong Ling, T.; Yeomans, N. D. Ethanol-induced cell damage in cultured rat antral mucosa assessed by chromium-51 release. Dig. Dis. Sci. 31(8):853–858; 1986.

    Article  PubMed  CAS  Google Scholar 

  20. Shopsis, C.; Eng, B. Rapid cytotoxicity testing using a semi-automated protein determination on cultured cells. Toxicol. Lett. 26:1–8; 1985.

    Article  PubMed  CAS  Google Scholar 

  21. Shopsis, C.; Sathe, S. Uridine uptake as a cytotoxicity test: correlations with the Draize test. Toxicology 29:195–206; 1984.

    Article  PubMed  CAS  Google Scholar 

  22. Shopsis, C.; Borenfreund, E.; Stark, D. M. Validation studies on a battery of potentialin vitro alternatives to the Draize test. In: Goldberg, A. M., ed. Alternative methods in toxicology, vol. 5. New York: MaryAnn Liebert, Inc.; 1987:31–44.

    Google Scholar 

  23. Silverman, J.; Pennisi, S.Tetrahymena thermophila (30377) as an indicator of ocular irritancy in rabbits. In: Goldberg, A. M., ed. Alternative methods in toxicology, vol. 3. New York: MaryAnn Liebert, Inc.; 1985:673–680.

    Google Scholar 

  24. Thomson, M. A.; Hearn, L. A.; Smith, K. T., et al. Evaluation of the neutral red cytotoxicity assay as a predictive test for the ocular irritancy potential of cosmetic products. In: Goldberg, A. M., ed. Alternative methods in toxicology, vol. 7. New York: MaryAnn Liebert, Inc.; 1989:297–305.

    Google Scholar 

  25. Triglia, D.; Wegener, P. T.; Harbell, J., et al. Interlaboratory validation study of the keratinocyte neutral red bioassay from Clonetics Corporation. In: Goldberg, A. M., ed. Alternative methods in toxicology, vol. 7. New York: MaryAnn Liebert, Inc.; 1989:357–365.

    Google Scholar 

  26. Verma, S. R.; Tyagi, A. K.; Pal, N., et al.In vitro effect of sodium lauryl sulfate on ATPase system in a few tissues fromChanna punctatus. Toxicol. Lett. 2:355–359; 1978.

    Article  CAS  Google Scholar 

  27. Wallin, R. F.; Hume, R. D.; Jackson, E. M. The agarose diffusion method for ocular irritancy screening: cosmetic products, part I. J. Toxicol. Cutaneous Ocular Toxicol. 6(4):239–250; 1987.

    Google Scholar 

  28. Yu, H-S.; Pastor, S. A.; Lam, K-W., et al. Ascorbate-enhanced copper toxicity on bovine corneal endothelial cellsin vitro. Curr. Eye Res. 9(2):177–182; 1990.

    PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Triglia, D., Braa, S.S., Yonan, C. et al. In vitro toxicity of various classes of test agents using the neutral red assay on a human three-dimensional physiologic skin model. In Vitro Cell Dev Biol - Animal 27, 239–244 (1991). https://doi.org/10.1007/BF02630923

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF02630923

Key words

Navigation