d-Limonene induced hyaline droplet nephropathy in alpha 2u-globulin transgenic mice.
d-Limonene is a hyaline droplet inducing agent and produces nephrotoxicity in male rats when the 1,2-epoxide metabolite binds to alpha 2u-globulin. Mice, which do not synthesize alpha 2u-globulin, are resistant to hyaline droplet nephropathy. In this study, the ability of d-limonene to cause hyaline droplet nephropathy in a transgenic mouse engineered to express alpha 2u-globulin was evaluated. The C57BL/6-derived mice excreted 0.4 +/- 0.1 mg alpha 2u-globulin/day, or approximately 16 mg alpha 2u-globulin/kg body wt. This represents about 30% of the amount excreted by adult male rats (11.9 +/- 1.1 mg/day or approximately 48 mg/kg). Transgenic mice excreted less mouse urinary protein (9.3 +/- 1.2 mg/day) than normal mice (15.1 +/- 1.6 mg/day). Unlike normal male rats, untreated transgenic mice did not show significant spontaneous hyaline droplet formation. Liver microsomes from naive transgenic mice oxidized d-limonene to the cis- and transisomers of the 1,2-epoxide, and following oral treatment with [14C]d-limonene reversible binding of d-limonene equivalents to renal cytosolic proteins was observed. Furthermore, with d-limonene treatment, hyaline droplets were observed in the transgenic mouse kidneys. These droplets, however, were much smaller in size than those seen in d-limonene-treated male rats. The accumulation of alpha 2u-globulin in the kidneys of transgenic mice and normal male rats before and after d-limonene treatment was analyzed by Western blotting. These results indicated that alpha 2u-globulin was present in the kidneys of the control transgenic mice, despite the lack of spontaneous hyaline droplet formation. After d-limonene treatment, approximately a three-fold increase in alpha 2u-globulin in the transgenic mouse kidney was observed, a response similar in magnitude to that seen in d-limonene-treated male rats. These results indicate that expression of alpha 2u-globulin in a species that does not normally develop hyaline droplet nephropathy is necessary and sufficient to render that species sensitive to this renal toxicity.
Lehman-McKeeman
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Miami Valley Laboratories, Procter & Gamble Company, Cincinnati, Ohio 45239-8707.
Caudill
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D000510:Alpha-Globulins / Q000096:biosynthesis* / Q000235:genetics*; D000818:Animals; D015153:Blotting, Western; D053138:Cyclohexenes; D007674:Kidney Diseases / Q000139:chemically induced* / Q000378:metabolism / Q000473:pathology; D000077222:Limonene; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D051381:Rats; D011916:Rats, Inbred F344; D013729:Terpenes / Q000378:metabolism / Q000633:toxicity*
D000510:Alpha-Globulins; D053138:Cyclohexenes; D013729:Terpenes; C013803:alpha 2u globulin; D000077222:Limonene