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Subcellular distribution of acidα-glucosidase in fibroblasts and of antigenically cross-reactive material in Pompe's disease fibroblasts

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Summary

From fibroblasts of two cases of Pompe's disease (acidα-glucosidase deficiency), one of the childhood type (RH-SF-1) and one of the adult type (RH-SF-2), and normal fibroblasts, antigenically cross-reactive material and acidα-glucosidase were immunoprecipitated and analysed by immunoelectrotransfer blotting. The acidα-glucosidase and antigenically cross-reactive material (which reacts with antibody raised against normal acidα-glucosidase) revealed a precursor form of molecular weight 97 000 and two major components of 79 000 and 76 000. When monensin was added to the fibroblast culture, the two major components of normal acidα-glucosidase were decreased, whereas the large molecular weight precursor was increased. On the other hand, the 97 000 molecular weight component of cross-reactive material in the Pompe's fibroblasts (RH-SF-1 and RH-SF-2) was only slightly increased on monensin treatment. The fibroblasts were pulse-chase labelled with [2-H3] mannose and32Pi. The cross-reactive material and acidα-glucosidase were precipitated with anti acidα-glucosidase antibody, and after sodium dodecyl sulphate—polyacrylamide gel electrophoresis (SDS—PAGE), fluorography was performed. The radiolabel of3H in the cross-reactive material of RH-SF-1 and -2 was weak, and32P in the cross-reactive material of both fibroblasts was very weak when compared with those of the acidα-glucosidase. The radiolabel of32P in the cross-reactive material of RH-SF-1 was extremely weak.

Immunofluorescence histochemistry revealed a granular localization of acidα-glucosidase in the normal fibroblast cytoplasm, and a diffuse distribution of cross-reactive material in the cytoplasm of RH-SF-1 and -2. Immuno-electron microscopic examinations showed a normal acidα-glucosidase localization on the inner side of the lysosomal membrane and also diffusely in the lysosome; when treated with monensin, it was present on the trans part of the Golgi apparatus. Antigenically cross-reactive material, however, was found in the cytoplasm and endoplasmic reticulum. Some lysosomal localization was observed sporadically. Even after monensin treatment, it was not demonstrated on the Golgi apparatus.

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Authors and Affiliations

  1. Department of Pathology, Ryukyu University School of Medicine, 207 Uehara, 903-01, Nishihara, Okinawa, Japan

    Teruo Iwamasa, Kazutaka Nashiro & Takeyuki Ohshita

  2. Department of Paediatrics, Kumamoto University Medical School, 860, Honjo, Kumamoto, Japan

    Ichiro Matsuda

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  1. Teruo Iwamasa
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  2. Kazutaka Nashiro
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  3. Takeyuki Ohshita
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  4. Ichiro Matsuda
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Iwamasa, T., Nashiro, K., Ohshita, T. et al. Subcellular distribution of acidα-glucosidase in fibroblasts and of antigenically cross-reactive material in Pompe's disease fibroblasts. Histochem J 18, 613–624 (1986). https://doi.org/10.1007/BF01675297

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  • DOI: https://doi.org/10.1007/BF01675297

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