Comparative In Vitro Cytotoxicity of 20 Potential Food Ingredients in Canine Liver, Kidney, Bone Marrow-Derived Mesenchymal Stem Cells, and Enterocyte-like Cells

Abstract

To begin development of a mechanistically relevant humane alternative platform for safety assessment of dog food ingredients, comparative in vitro cytotoxicity of 20 ingredients was assessed in four canine cell types relevant for toxicity assessments. Previously, we described the toxicity of 13 compounds (clove leaf oil, eugenol, guanosine monophosphate [GMP], GMP plus inosine monophosphate, sorbose, ginger root extract, cinnamon bark oil, cinnamaldehyde, thyme oil, thymol, lemon grass oil, xylitol, and citric acid) using in vitro primary canine cell culture models for liver, kidney, bone marrow-derived mesenchymal stem cells (BMSC), and enterocyte-like cells (ELC). In this report, dose–response cytotoxicity studies and LC50 using alamar blue assays are reported for seven additional compounds: denatonium benzoate, eucalyptol, hexahydroisohumulone, tetrahydroisohumulone, green tea catechin extract, epigallocatechin gallate, and sodium copper chlorophyllin. Data across 20 compounds were compared between different cell types and responses were not parallel, precluding the use of a single cell line for in vitro ingredient hazard assessment. Hepatocytes were most resistant to all compounds, consistent with their xenobiotic detoxification functions. BMSC and ELC showed an increase in sensitivity to the essential oils eucalyptol, eugenol, and thymol compared to renal cells and hepatocytes. These studies provide a baseline of acute cytotoxicity of 4 canine cell types to 20 different food components that begin to illustrate how such an in vitro panel could be used for hazard assessment.