OBJECTIVES: (1) To determine whether a peroxidase deficiency exists in Huntington's Disease (HD) which could lead to accumulation of brain cell damaging lipopigments; (2) to assay PPD-peroxidase levels in donor age and passage level - paired serially cultured skin fibroblasts in patients with HD, Batten's Disease and genetic disease-free controls; (3) to compare total protein content in age-paired, serially cultured fibroblasts from subjects with HD, Batten's and controls by the method of high resolution, two-dimensional electrophoresis;(4) to open the possibility for fetal diagnosis of HD by amniocentesis which would be made feasible by finding a peroxidase deficiency in cultured fibroblasts; (5) to assess, by light microscopy and histochemistry, the capability of centrophenoxine (CF) to induce a diminution of lipopigment which normally accumulates in serially propagated late passage IMR-90 human cells; (6) to correlate the influence of CF on lipopigment content of aging IMR-90 cells with PPD-peroxidase and catalase levels in these drug-treated and control cells; (7) to attempt the rescue of aging IMR-90 cells by a regime of CF treatmnt in those late passage cultured cells which otherwise are programmed for mitotic decline and death; (8) to demonstrate cellular lipopigment dissolution by CF, perhaps associated with altered peroxidase levels and, thereby, provide a rationale for the eventual treatment of HD and perhaps other patients with this drug. METHODS: (a) Spectrophotometric assay of PPD-peroxidase in HD, Batten's and control cultured skin fibroglasts; (b) determination of total protein content of cells as in (a) by high resolution, two-dimensional electrophoresis; (c) fluorometric control lysosomal enzyme assays on cultured fibroblasts as in (a); (d) peroxisomal catalase determinations in cultured fibroblasts as in (a); (e) assessment, by light microscopy and histochemistry, of CF effects on lipopigment in aging IMR-90 cells; (f) determination of PPD-peroxidase and catalase in cultured youthful and aging IMR-90 cells.
NIH ExPorter
NIH ExPorter