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[Copper level and metallothionein-like Cu-binding protein in cultured skin fibroblasts from patients with Menkes' disease and Wilson's disease].

Abstract

Copper concentration, intracellular copper distribution, and inducibility of metallothionein-like metal-binding protein (MLP) by copper or cadmium addition to culture medium were compared among three types of skin fibroblasts derived from patients with Menkes' disease and Wilson's disease, both exhibiting genetic defects of copper metabolism, and from normal subjects (control). Skin fibroblasts were cultivated in Dulbecco's modified Eagle's medium supplemented with 10% fetal calf serum and antibiotics in 5% CO2 at 37 degrees C. Cells were harvested with rubber-policeman, washed twice with phosphate-buffered saline, pH 7.2, suspended in deionized water, and homogenized. The homogenate from each cell type was used to determine the concentration of copper by atomic absorption spectrophotometry employing graphite-rod atomizer after lyophilization, ashing in HNO3, and coprecipitation with zirconium. Intracellular copper concentration was elevated in Menkes' cells (420 ng Cu/mg of protein) and Wilson's cells (217 ng Cu/mg of protein) than in control cells (90.0 ng Cu/mg of protein), although one of four Wilson's strains showed normal copper level (70.5 ng Cu/mg of protein). Cytosol copper concentration was 5.8-fold higher in Menkes' cells but only 1.3-fold in Wilson's cells than in control cells, and cytosol copper accounted for only 35% of total intracellular copper in Wilson's cells as compared with 68% and 52% in Menkes' and control cells, respectively. These suggest that accumulated copper in each cell type is differently distributed within cells; in Menkes' cells exclusively into cytosol, but in Wilson's cells into particulates rather than cytosol. Elution profiles from Sephadex G-75 columns indicated that most of copper had bound to MLP in Menkes' cells, though no Cu-MLP was detectable in Wilson's or control cells under these experimental conditions.(

ABSTRACT

TRUNCATED AT 250 WORDS)

Authors

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Source

No to shinkei = Brain and nerve 36:11 1984 Nov pg 1063-8

MeSH

Adolescent
Adult
Brain Diseases, Metabolic
Carrier Proteins
Cell Line
Cells, Cultured
Child
Chromatography, Gel
Copper
Female
Fibroblasts
Hepatolenticular Degeneration
Humans
Infant
Male
Menkes Kinky Hair Syndrome
Metallothionein
Skin

Pub Type(s)

English Abstract
Journal Article
Research Support, Non-U.S. Gov't

Language

jpn

PubMed ID

6525319

Citation

Sato, M, et al. "[Copper Level and Metallothionein-like Cu-binding Protein in Cultured Skin Fibroblasts From Patients With Menkes' Disease and Wilson's Disease]." No to Shinkei = Brain and Nerve, vol. 36, no. 11, 1984, pp. 1063-8.
Sato M, Hayashi A, Ito H, et al. [Copper level and metallothionein-like Cu-binding protein in cultured skin fibroblasts from patients with Menkes' disease and Wilson's disease]. No To Shinkei. 1984;36(11):1063-8.
Sato, M., Hayashi, A., Ito, H., Tojo, M., & Arima, M. (1984). [Copper level and metallothionein-like Cu-binding protein in cultured skin fibroblasts from patients with Menkes' disease and Wilson's disease]. No to Shinkei = Brain and Nerve, 36(11), pp. 1063-8.
Sato M, et al. [Copper Level and Metallothionein-like Cu-binding Protein in Cultured Skin Fibroblasts From Patients With Menkes' Disease and Wilson's Disease]. No To Shinkei. 1984;36(11):1063-8. PubMed PMID: 6525319.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - [Copper level and metallothionein-like Cu-binding protein in cultured skin fibroblasts from patients with Menkes' disease and Wilson's disease]. AU - Sato,M, AU - Hayashi,A, AU - Ito,H, AU - Tojo,M, AU - Arima,M, PY - 1984/11/1/pubmed PY - 1984/11/1/medline PY - 1984/11/1/entrez SP - 1063 EP - 8 JF - No to shinkei = Brain and nerve JO - No To Shinkei VL - 36 IS - 11 N2 - Copper concentration, intracellular copper distribution, and inducibility of metallothionein-like metal-binding protein (MLP) by copper or cadmium addition to culture medium were compared among three types of skin fibroblasts derived from patients with Menkes' disease and Wilson's disease, both exhibiting genetic defects of copper metabolism, and from normal subjects (control). Skin fibroblasts were cultivated in Dulbecco's modified Eagle's medium supplemented with 10% fetal calf serum and antibiotics in 5% CO2 at 37 degrees C. Cells were harvested with rubber-policeman, washed twice with phosphate-buffered saline, pH 7.2, suspended in deionized water, and homogenized. The homogenate from each cell type was used to determine the concentration of copper by atomic absorption spectrophotometry employing graphite-rod atomizer after lyophilization, ashing in HNO3, and coprecipitation with zirconium. Intracellular copper concentration was elevated in Menkes' cells (420 ng Cu/mg of protein) and Wilson's cells (217 ng Cu/mg of protein) than in control cells (90.0 ng Cu/mg of protein), although one of four Wilson's strains showed normal copper level (70.5 ng Cu/mg of protein). Cytosol copper concentration was 5.8-fold higher in Menkes' cells but only 1.3-fold in Wilson's cells than in control cells, and cytosol copper accounted for only 35% of total intracellular copper in Wilson's cells as compared with 68% and 52% in Menkes' and control cells, respectively. These suggest that accumulated copper in each cell type is differently distributed within cells; in Menkes' cells exclusively into cytosol, but in Wilson's cells into particulates rather than cytosol. Elution profiles from Sephadex G-75 columns indicated that most of copper had bound to MLP in Menkes' cells, though no Cu-MLP was detectable in Wilson's or control cells under these experimental conditions.(ABSTRACT TRUNCATED AT 250 WORDS) SN - 0006-8969 UR - https://www.unboundmedicine.com/medline/citation/6525319/[Copper_level_and_metallothionein_like_Cu_binding_protein_in_cultured_skin_fibroblasts_from_patients_with_Menkes'_disease_and_Wilson's_disease]_ L2 - http://ccr.coriell.org/Sections/Search/PubSamples.aspx?PgId=516&id=6525319&lvl=0 DB - PRIME DP - Unbound Medicine ER -