HETEROGENEITY OF DNA AND RNA IN HUNTER PATIENTS

Genomic DNA and total RNA from lymphoblasts of nine unrelated Italian patients affected with Hunter syndrome were analyzed using a human cDNA clone coding for the lysosomal enzyme iduronate-2-sulphatase (IDS). Southern blot analysis resulted in patterns similar to the normal control for seven of the patients analyzed; an aberrant pattern was observed in two patients (F.N. and P.D.), suggesting deletions/rearrangement in the IDS gene. Northern blot analysis showed in seven patients, a pattern similar to the normal control; for patients F.N. and P.D. the pattern was atypical, i.e., normal RNA species were absent whereas two different transcripts occurred. These data confirm the heterogeneity of the molecular defects causing Hunter disease. © 1993 Springer-Verlag

Evidence for secretion of cytosolic CuZn superoxide dismutase by hep G2 cells and human fibroblasts

The role so far ascribed to intracellular CuZn superoxide dismutase is that of an intracellular scavenger of oxygen radicals. However, other functions of cytosolic CuZn superoxide dismutase have been hypothesized. For example, CuZn superoxide dismutase incubated with rat hepatocyte cells in culture inhibits 3-hydroxy-3methylglutaryl CoA reductase, thereby reducing cholesterol synthesis. We recently demonstrated the presence of surface membrane receptors for CuZn superoxide dismutase, suggesting possible autocrine or paracrine activities. The aim of the present study was to investigate whether cytosolic CuZn superoxide dismutase can be secreted by human hepatocarcinoma and fibroblast cells lines. Proteins in human hepatocellular carcinoma (Hep G2) cells and human fibroblasts were biosynthetically labelled with [S-35]-cysteine; then cell lysates and media were immunoprecipitated with rabbit polyclonal anti-human CuZn superoxide dismutase antibodies and separated by 12% polyacrylamide gel electrophoresis. Both Hep G2 cells and human fibroblasts produce and secrete CuZn superoxide dismutase which was detectable in cells and medium as a single protein band with the same electrophoretic mobility as human erythrocyte CuZn superoxide dismutase. These data suggest that CuZn superoxide dismutase, an enzyme thus far considered to be located exclusively intracellularly is secreted by at least two cell lines. This is consistent with autocrine or paracrine roles for CuZn superoxide dismutase. (C) 1996 Elsevier Science Lt

Identification of the carrier status in Hunter syndrome by molecular analyses

X-linked recessive Hunter syndrome (mucopolysaccharidosis type II) is a heritable disorder characterized by a deficiency of the lysosomal enzyme iduronate-2-sulphatase. Carrier detection of the disease was studied in two families by molecular analyses. In the first family unequivocal evidence of the carrier status was obtained both by Southern and Northern blot techniques, demonstrating that these methods are a valuable diagnostic tool in genetic counselling of cases where an aberrant pattern is present. In the second family, the intragenic RFLP analysis allowed an accurate genotype diagnosis