Quantitation of α2-microglobulin after administration of structurally divergent chemical compounds

Abstract

α₂-Microglobulin-induced nephropathy is a phenomenon which is exclusively found in adult male rats. Various chemicals are able to bind to α₂-microglobulin thus inhibiting its proteolytic degradation in lysosomes of the P2 segment of the rat nephron. The accumulation of this protein in `protein droplets' or `hyaline droplets' leads to necrosis, followed by regeneration which possibly later results in the formation of tumours. Here we report the development of a monoclonal antibody which is specific for α₂-microglobulin. It was utilized to measure α₂-microglobulin concentrations in plasma and tissues, and to stain α₂-microglobulin in fixed tissue slides. In two studies we administered to adult male Wistar rats two groups of compounds: (1) one group of structurally diverse compounds, which give an overview of chemical entities capable of inducing the accumulation of α₂-microglobulin; and (2) another group of structurally closely related compounds (i.e. substituted benzene derivatives) for the purpose of elucidating possible structure-activity relationships. The degree of α₂-microglobulin-induced nephropathy was determined by immunohistochemical staining of kidney sections. In addition, liver and kidney tissue and plasma concentrations of α₂-microglobulin were measured by competitive ELISA. Liver tissue and plasma concentrations of α₂-microglobulin were not found to be elevated whereas kidney tissue concentrations were higher than the controls. The increase over control values ranged␣from 154% (1,4-dichloromethyl-benzene) to 321% [α-methyl-4-(1-methylethyl)-cyclohexanemethanol]. Comparing structurally related benzene derivatives, the hyaline droplet accumulating (HDA) potential was found to depend both on the type of substituent and its position at the aromatic ring. In general HDA activity increased in the order benzene ≅ phenol ≅ alkylated phenols < halogenated phenols < halogenated␣benzenes. Further QSAR studies are needed to provide a theoretical base for these observations.