NEUTRAL RED UPTAKE INHIBITION IN FOUR CELL LINES AS AN ALTERNATIVE SYSTEM FOR IN VIVO NEPHROTOXICITY

HILDE H1. BOEHTS1,2, ETIENNE J1. NOEWEN2, MARC E. DE BROE2 AND PAUL J. DIERICKX1
1Instituut poor Hygiene en Epidemiologie, Afdeling toxicologie, Wytsmanstruat 14, B-1050 Brussel, Belgium; 2UZA-UIA, Dienst Nefrologie-Hypertensie, Wilrijkstraat 1O, B-2650 Edegem, Belgium

Regular article AATEX 2(3):109-114

Abstract
The neutral red uptake inhibition in LLC-PK1 (derived from pig kidney proximal tubules) and MDCK (derived from dog kidney distal tubules) cells was investigated for its use as an indicator for in vivo nephrotoxicity, as quantified by urinary protein content and -glutamyltransferase activity in rats. Both cell lines were investigated for their possible organ specificity by comparing them with cells derived from human liver (Hep G2) and osteoblast (SAOS). The chemicals tested were: cisplatinum, mercury chloride, cupric chloride, potassium bichromate, sodium tetrathionate, ammonium fluoride and the aminoglycosides: gentamicin, neomycin, kanamycin and streptomycin. The relative cytotoxicity of the test compound was established by the determination of the NI50. This is the concentration of test compound inducing a 50% reduction in neutral red uptake. LLC-PK1 cells were more susceptible to HgCI2. than MDCK cells, reflecting the in vivo situation where proximal tubular cells are also more sensitive to HgCI2 than the distal tubular cells.
The in vitro-in vivo comparison was made by calculating the Spearman rank correlation coefficient (rs) between the NI50 ranks and the nephrotoxicity ranks in rats. Potassium bichromate was identified as an outlayer and was therefore omitted from the correlation calculations. The following correlations were found: rs=0.90 for MDCK; rs=0.97 for LLCPK1; rs=0.93 for SAOS and rs=0.95 for Hep G2. The reported results indicate the potential value of the neutral red uptake inhibition assay in the four cell lines for the nephrotoxic ranking of chemicals.


return