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Defective glucuronic acid transport from lysosomes of infantile free sialic acid storage disease fibroblasts.

ABSTRACT: Separation by h.p.l.c. and pulsed amperometric detection were employed to measure glucuronic acid (GlcUA) and other acidic monosaccharides in fibroblasts from patients with infantile free sialic acid storage disease (ISSD) and Salla disease. These lysosomal storage disorders result from defective carrier-mediated transport of free N-acetylneuraminic acid (NeuAc) out of cellular lysosomes. Three Salla disease fibroblast strains stored approx. 0.4 nmol of free GlcUA/mg of cell protein, whereas four ISSD strains stored approx. 5 nmol GlcUA/mg (normal is undetectable). The GlcUA content of the mutant cell strains, which by differential centrifugation and Percoll gradient fractionation was localized to the lysosomes, averaged 5% of the free NeuAc content of the cells. N-Glycolylneuraminic acid (NeuGc) also accumulated in ISSD cells, but only when they were grown in the presence of fetal calf serum, which contains abundant NeuGc. No other acidic monosaccharides were detected in any of the mutant cell strains. GlcUA egress studies revealed that 56% of the initial GlcUA content was lost from normal granular fractions after 2 min at 37 degrees C. For similarly loaded ISSD granular fractions, virtually no GlcUA was lost even after 6 min. The results indicate that GlcUA is recognized and transported by the lysosomal NeuAc carrier, and that GlcUA transport is impaired in the lysosomal disorders of free NeuAc storage.

SUBMITTER: Blom HJ

PROVIDER: S-EPMC1131484 | biostudies-other | 1990 Jun

REPOSITORIES: biostudies-other

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Project description:Fibroblasts from patients with sialic acid storage disease (SASD), sialidosis, mucolipidosis II, and from normal controls, were incubated in the presence of the glycoprotein fetuin that was tritium-labelled in its sialic acid residues by the periodate/[3H]borohydride reduction method, and the fate of the intracellular radioactive sialic acid (C7-sialic acid) followed in pulse-chase experiments. The model glycoprotein was readily endocytosed and degraded, more than 90% of the radioactivity being trichloroacetic acid (TCA)-soluble after 4 days of incubation. In all of the patients' fibroblasts, there was an increased accumulation of TCA-soluble radioactivity and, upon chase, a much lower rate of elimination than in normal controls. Gel chromatography of the material from the chase experiment showed that, in normal cells, most of the radioactivity at zero time behaved as free C7-sialic acid. This, as well as material of larger size (sialyloligosaccharides), was very much diminished by 48 h. In cells from two patients with SASD, there were large peaks both in the sialic acid and oligosaccharide positions; whereas the oligosaccharides were somewhat decreased by the end of the chase period, the sialic acid was essentially unchanged. In sialidosis fibroblasts, the radioactive material consisted of oligosaccharides, but very little C7-sialic acid; the elimination of the oligosaccharides was retarded. In normal cells, about 80% of the radioactivity released into the medium after 48 h chase behaved as free C7-sialic acid upon gel chromatography and t.l.c. Subcellular fractionation in Percoll gradients showed that the radioactive C7-sialic acid remaining in normal cells after 48 h of chase was mainly localized in the cytosol. In SASD cells, on the other hand, it was associated with lysosomal fractions which, unexpectedly, exhibited an abnormally low density. Our findings demonstrate that SASD fibroblasts degrade the sialoglycoprotein but, unlike normal cells, accumulate the liberated C7-sialic acid along with sialyloligosaccharides in their lysosomes. The results therefore support the concept of a defective transport system for sialic acid in the lysosomal membrane of patients with SASD.

Project description:Lysosomal free sialic acid storage disorder (FSASD) is an extremely rare, autosomal recessive, neurodegenerative, multisystemic disorder caused by defects in the lysosomal sialic acid membrane exporter SLC17A5 (sialin). SLC17A5 defects cause free sialic acid and some other acidic hexoses to accumulate in lysosomes, resulting in enlarged lysosomes in some cell types and 10-100-fold increased urinary excretion of free sialic acid. Clinical features of FSASD include coarse facial features, organomegaly, and progressive neurodegenerative symptoms with cognitive impairment, cerebellar ataxia and muscular hypotonia. Central hypomyelination with cerebellar atrophy and thinning of the corpus callosum are also prominent disease features. Around 200 FSASD cases are reported worldwide, with the clinical spectrum ranging from a severe infantile onset form, often lethal in early childhood, to a mild, less severe form with subjects living into adulthood, also called Salla disease. The pathobiology of FSASD remains poorly understood and FSASD is likely underdiagnosed. Known patients have experienced a diagnostic delay due to the rarity of the disorder, absence of routine urine sialic acid testing, and non-specific clinical symptoms, including developmental delay, ataxia and infantile hypomyelination. There is no approved therapy for FSASD. We initiated a multidisciplinary collaborative effort involving worldwide academic clinical and scientific FSASD experts, the National Institutes of Health (USA), and the FSASD patient advocacy group (Salla Treatment and Research [S.T.A.R.] Foundation) to overcome the scientific, clinical and financial challenges facing the development of new treatments for FSASD. We aim to collect data that incentivize industry to further develop, obtain approval for, and commercialize FSASD treatments. This review summarizes current aspects of FSASD diagnosis, prevalence, etiology, and disease models, as well as challenges on the path to therapeutic approaches for FSASD.

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Defective glucuronic acid transport from lysosomes of infantile free sialic acid storage disease fibroblasts.

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