The effect of pH and lysosomal extract on rat liver metallothionein (Cd-thionein)

Meulenaar, H.; Hamer, C.J.A. van den; Ingh, T.S.G.A.M. van den; Stolk, Th.M.W.

doi:10.1007/bf02916546

Cited by 8 publications

(1 citation statement)

References 10 publications

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“…Lysosomes probably represent an important subcellular compartment that might be involved in the degradation of MT in vivo: Previous studies on the degradation of MT in rat liver indicated that lysosomes can degrade apo-MT (11,12). However, apo-MT was also shown to be degraded by trypsin, which is a neutral cytosolic protease (11).…”

Section: Resultsmentioning

confidence: 99%

In Vitro and In Vivo Studies on the Degradation of Metallothionein

Klaassen¹,

Choudhuri²,

McKim³

et al. 1994

Environmental Health Perspectives

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Degradation of metallothionein (MT) from rat liver was examined. Degradation of apo-MT by liver homogenate was greater than that by cytosol. At pH 5.5, degradation by homogenate was more than that at pH 7.2. These findings suggest that proteases that function at acidic pH are probably involved in MT degradation. Because lysosomes are the principal subcellular organelles that contain acid proteases (cathepsins), we compared the degradation of apo-MT by lysosomes and cytsosol. Apo-MT was degraded about 400 times faster by lysosomal fraction than by cytosolic fraction. To determine the relative importance of different cathepsins, we used different inhibitors. Leupeptin, which inhibits cathepsins B and L, inhibited the degradation of apo-MT by 80%, implying that cathepsins B and/or L might be very important in the intracellular turnover of MT. Cathepsin D appeared to be the least significant, because apo-MT degradation was reduced by about 20% by inhibiting cathepsin D. When we extended this study with purified cathepsins, we obtained the same answer, i.e., the ability of different cathepsins to degrade apo-MT was in the following order: cathepsin B >> cathepsin C > cathepsin D. While apo-MT was susceptible to degradation, ZnMT and CdMT were highly resistant to degradation. Coincubation of ZnMT or CdMT with either lysosomal extract or purified cathepsins did not result in any appreciable degradation even after 16 hr. However, longer incubations did result in some degradation, especially by purified cathepsin B. Interestingly, CdMT degraded little faster than ZnMT by both lysosomal extract as well as purified cathepsin B. These data suggest that metals protect MT from rapid proteolysis. By metal-titration study, we found that at least 5 moles of Zn equivalent/mole of MT dramatically reduced the degradation, by both lysosomal fraction and purified cathepsin B. This indicates that metal release might be a prerequisite for the initiation of degradation. Release of metals is possible in the lysosomes because the intralysosomal pH is close to 5.0. Therefore, we determined the displacement of metals (Zn) as a function of pH. We found that at a pH of 4.5, nearly 70% Zn was removed, and at pH 4.0 nearly all Zn was displaced from MT. We propose, therefore, that lysosomes are probably important in the in vivo degradation of MT, and that metal release is a prerequisite for degradation. With the release of metals in the acidic pH of lysosomes, MT becomes more susceptible to degradation, which is probably accomplished by the cathepsins, in particular cathepsin B and/or L. The in vivo study, however, suggests that there are other factors, too, in addition to metal composition and pH, that may have profound effect on determining the half-lives of various forms of MT, at various stages of development. -Environ Health Perspect 102(Suppl 3): 141-146 (1994).

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Section: Resultsmentioning

confidence: 99%

In Vitro and In Vivo Studies on the Degradation of Metallothionein

Klaassen¹,

Choudhuri²,

McKim³

et al. 1994

Environmental Health Perspectives

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Histochemical staining of cadmium thiolate clusters in livers of rats treated chronically with cadmium

Morselt

Straalen

1986

Histochemistry

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In livers of rats exposed to varying doses of CdCl2 80-90% of the cadmium content present in the fresh tissue is retained if these livers are fixed with a neutral or acid formalin fixative. Cadmium assays during different stages of the staining procedure for protein bound disulphides show the ability of this staining to demonstrate cadmium thiolate clusters next to disulphides. The methods described may also be useful in gaining more insight in the mechanism involved in fixation and staining procedure of some other metals.

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Effects of cadmium in freshwater clams. II. Ultrastructural changes in the renal system ofAnodonta cygnea

Hemelraad

Herwig

Donselaar

et al. 1990

Arch. Environ. Contam. Toxicol.

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An ultrastructural study was made of the renal system of freshwater clams,Anodonta cygnea, that had been exposed to cadmium chloride (50 μg Cd/L) for 12 weeks. By stereological analysis an extended lysosomal system and a decreased number of mitochondria was apparent in the epithelial cells lining the proximal compartment of the kidney. The increase of the lysosomal system was mainly accountable to the appearance of a distinct type of lysosome, that accumulated in the apical cell region. The decrease of the mitochondrial population was accompanied by a considerable swelling of the individual mitochondria. Finally, a severe reduction of the glycogen stores was noticed. Similar, but less obvious, changes occurred in the distal kidney compartment. The results suggest that long-term exposure ofAnodonta cygnea to cadmium stimulates the lysosomal system and disturbs the function of organelles involved in the energy metabolism of resorptive kidney cells.

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The effect of pH and lysosomal extract on rat liver metallothionein (Cd-thionein)

Cited by 8 publications

References 10 publications

In Vitro and In Vivo Studies on the Degradation of Metallothionein

In Vitro and In Vivo Studies on the Degradation of Metallothionein

Histochemical staining of cadmium thiolate clusters in livers of rats treated chronically with cadmium

Effects of cadmium in freshwater clams. II. Ultrastructural changes in the renal system ofAnodonta cygnea

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