Mechanism of hepatorenal syndrome in rats of Long–Evans Cinnamon strain, an animal model of fulminant Wilson's disease

Abstract

Rats of Long–Evans Cinnamon (LEC) strain were used as a hepatorenal syndrome model of fulminant Wilson's disease. Copper levels in the kidneys increased markedly from 16 to 126 μg Cu/g from 12 to 16 weeks, and remained at the same level at 16 and 19 weeks when the rats suffered from severe renal dysfunction and also at 20 weeks in some other normal rats. The above findings imply that the renal dysfunction may have been induced independently of the copper level in the kidneys. The present study suggested the following mechanism: immediately after copper-induced hepatic dysfunction, plasma copper–metallothionein (CuMT), which was released from the liver, became elevated. The elevation was closely related to the increases in alkaline phosphatase, glucose and amino acids, all in the urine. The above findings suggest that plasma CuMT, which was released from the liver into the blood upon copper-induced hepatic dysfunction, was subsequently filtered at the glomeruli due to its smaller molecular weight, and then caused dysfunction of the brush border membrane of the renal proximal tubules probably after splitting into radical copper and amino acids in acidic vesicles close to the membrane. The critical concentration of plasma CuMT required to induce renal dysfunction was estimated as 1 μg Cu/l.

Introduction

Wilson's disease has been characterized as a hereditary disease involving excessive copper accumulation in the liver due to a decreased excretion of copper from the liver, and childhood patients frequently suffer from fatal hepatic disorder (Yarze et al., 1992). Almost half of Wilson's disease patients develop renal dysfunction (Guan et al., 1986), but information is not yet sufficiently available on the renal dysfunctions in Wilson's disease. A search revealed only one paper (Tochimaru et al., 1991) on renal dysfunction in rats of the Long–Evans Cinnamon strain (LEC rat), an animal model of fulminant Wilson's disease.

It was found recently that LEC rats can serve as a model of Wilson disease because the proximal breakpoint of the gene is located in a position equivalent to human Atp 7b in intron 15, and the deletion appears to extend to include the 3′ end of the gene (Wu et al., 1997). The present study, therefore, focused on the mechanism of the hepatorenal syndrome in fulminant Wilson's disease.

Our hypothesis for the mechanism of copper-induced renal dysfunction in Wilson's disease is identical to that for cadmium-induced renal dysfunction (Nomiyama and Nomiyama, 1998) as seen in Fig. 1: copper accumulates in the liver mostly in the chemical form of copper–metallothionein (CuMT). Once the copper accumulation in the liver exceeds the ability to induce sufficient MT for copper detoxification, the copper radical Cu²⁺ induces hepatic dysfunction, and the massive CuMT accumulated in the liver is then released into the blood from the liver. Plasma CuMT passes readily through the glomeruli into the tubular lumen because of its smaller molecular weight, and then directly injures the brush border membrane of the renal proximal tubular cells resulting in enzymuria, proteinuria, aminoaciduria and glucosuria, following splitting of CuMT into the copper radical Cu²⁺ and amino acids in the acidic vesicles probably close to the brush border membrane, independently of the copper concentration in the renal cortex.

We undertook the present study to verify the above hypothesis for the mechanism of hepatorenal dysfunction in Wilson's disease using LEC rats. We then evaluated abnormal functions of the liver and kidneys in relation to the total copper, CuMT, copper not bound to MT (nonMTCu), lipid peroxide, nitrite and nitrate, superoxide dismutase in plasma, liver or kidneys.