In vitro toxicity of glyphosate in Atlantic salmon evaluated with a 3D hepatocyte-kidney co-culture model

Abstract

A novel 3D Atlantic salmon co-culture model was developed using primary hepatocytes and kidney epithelial cells isolated from the same fish. Mono and co-cultures of primary hepatocytes and kidney epithelial cells were exposed for 48 h to glyphosate (5, 50 and 500 μM). For comparison, cells were also exposed to chlorpyrifos, benzo(a)pyrene and cadmium. Cell staining, cell viability assessments, RT-qPCR and global metabolomic profiling were used to examine the toxicological effects on liver and renal function and to compare responses in 3D and 2D cultures. The 3D hepatocyte cell culture was considered superior to the 2D culture due to the ATP binding cassette subfamily B member 1 (Abcb1) response and was thus used further in co-culture with kidney cells. Metabolomic analysis of co-cultured cells showed that glyphosate exposure (500 μM) altered lipid metabolism in both hepatocytes and kidney cells. Elevated levels of several types of PUFAs and long-chain fatty acids were observed in exposed hepatocytes, owing to increased uptake and phospholipid remodelling. Glyphosate suppressed the expression of estrogen receptor 1 (Esr1) and vitellogenin (Vtg) and altered histidine metabolism in exposed hepatocytes. Increased levels of cholesterol and downregulation of clusterin (Clu) suggest that glyphosate treatment affected membrane stability in Atlantic salmon kidney cells. This study demonstrates the usefulness of applying 3D co-culture models in risk assessment.