Intracellular degradation of sulforhodamine-GM1: Use for a fluorescence-based characterization of GM2-gangliosidosis variants in fibroblasts and white blood cells

Vered Agmon, Rami Khosravi, Sergio Marchesini, Tama Dinur, Arie Dagan, Shimon Gatt, Ruth Navon

Research output: Contribution to journalArticlepeer-review

Abstract

A novel fluorescent ganglioside, sulforhodamine-GM1 was administered into cells derived from carriers and patients with different subtypes of GM2 gangliosidosis, resulting from various mutations in the gene encoding the lysosomal enzyme hexosaminidase (Hex) A. The cells used were skin fibroblasts and white blood cells, i.e. lymphocytes, monocytes and macrophages. In the severe infantile form of the GM2 gangliosidosis, Tay-Sachs disease, the sulforhodamine-GM1 was hydrolyzed within the lysosomes to the corresponding sulforhodamine-GM2 which, because of lack of Hex A activity, was not further degraded. In comparison, in the cells derived from GM2 gangliosidoses carriers, as well as pseudodeficient and adult forms of GM2 gangliosidosis, the sulforhodamine-GM2 was further processed and sequentially degraded by the lysosomal glycosidases to sulforhodamme-ceramide. The latter was converted to sulforhodamine-sphingomyelin, which was secreted into the culture medium. The fluorescence of the sulforhodamine ceramide in cell extracts and/or sulforhodamine-sphingomyelin in the culture medium was quantified and related to parallel data obtained using cells of normal individuals. This permitted distinguishing between the various GM2 gangliosidoses subtypes and relating the intracellular hydrolysis of sulforhodamine-GM1 to the genotypes of the respective GM2 gangliosidoses variants.

Original languageEnglish
Pages (from-to)105-120
Number of pages16
JournalClinica Chimica Acta
Volume247
Issue number1-2
DOIs
StatePublished - 29 Mar 1996

Keywords

  • Fluorescent G ganglioside
  • G ganglioside
  • G gangliosidosis
  • Lipid storage diseases
  • Tay-Sachs disease
  • β-N-acetyl hexosaminidase A

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