Tohru Yutaka

I-cell disease: clinical studies of 21 Japanese cases

Clinical pictures of 21 cases with I‐cell disease patients, 12 males and 9 females, were analyzed. Characteristic coarse facial features and shortness of stature were observed in all cases. In general, the motor development was found to be more severely retarded than the mental development of the patients. Rather little involvement of the nervous system seemed to cause somewhat acceptable mental development in some cases, and also cause the absence of epileptic seizures in all cases. Involvement of the cardiovascular system, especially progressive hypertrophic cardiomyopathy, could be highly responsible for frequent sudden death of I‐cell disease patients.

The effects of sucrose loading on lysosomal hydrolases.

SummaryThe addition of 88 mM sucrose to the culture medium of human skin fibroblasts from normal subjects caused remarkable increase in the intracellular lysosomal hydrolase activities. The mechanism of this induction by sucrose loading was carefully studied with several fibroblast strains of different inherited lysosomal storage disorders. In single lysosomal hydrolase defect such as GM1-gangliosidosis, mannosidosis and Sandhoff disease, no induction of the deficient hydrolase was found with 88 mM sucrose loading. In contrast, sucrose loading caused normalization of intracellular lysosomal hydrolase activities in I-cell disease fibroblasts and cytoplasmic inclusion materials disappeared. Subsequent investigations reveal that I-cell disease cells are classified into three subgroups by the degree of hydrolase induction by sucrose loading; a high responding, an intermediate responding and a no-response group. The heterogeneity may be based upon different induction by sucrose loading of the enzyme, probably the residual phosphotransferase which is involved in the processing steps of lysosomal enzyme molecules. With the addition of mannose-6-phosphate and 10 mM NH4Cl to cultured skin fibroblasts, it was shown that sucrose loading caused increased synthesis of lysosomal enzyme proteins. The result of the test with 2,4-dinitrophenol suggests that sucrose is indeed pinocytosed by cultured human skin fibroblasts and localized in lysosomes and that this event is the essential factor to trigger the induction of lysosomal hydrolases. Simultaneous loading of both invertase and sucrose in cultured cells caused no induction of α-mannosidase activity. This result indicates that invertase is also pinocytosed, reaches the lysosomes and hydrolyzes sucrose in the lysosomes. Lysosomal overloading with sucrose resulted in induction of lysosomal hydrolases and invertase blocked the induction of α-mannosidase activity. However, some induction still exists in β-galactosidase and α-fucosidase activity. Thus it is very likely that the induction of lysosomal hydrolases demands a complicated process.In this article, we investigated the effects of sucrose on the lysosomal hydrolases in cultured human skin fibroblasts of several inherited lysosomal storage disorders and normal subjects and discuss the possible mechanism. of the induction of lysosomal hydrolase activities by sucrose loading.

Beta-galactosidase deficient-type mucolipidosis: A complementation study of neuraminidase in somatic cell hybrids

Abstract By cell fusion with polyethylene glycol (PEG) a remarkable increase of neuraminidase activity was observed in the fused cells between ML-II and other two neuraminidase deficient disorders, ML-Gal (mucolipidosis, galactosidase deficient type) and ML-I (mucolipidosis I). No complementation was found in the combination of ML-I and ML-Gal. This result suggests that ML-I and ML-Gal may be allelic mutations and belong to the same genetic disorder as a primary neuraminidase deficiency.

Neuronal depletion of cerebellum in late infantile metachromatic leulcodystrophy

The cerebellums of two children with late infantile metachromatic leukodystrophy were investigated by light and electron microscopy. These observations showed that the morphological abnormalities were comprised not only of severe demyelination and presence of metachromatic granules but also marked loss of Purkinje cells and granule cells. The depletion of Purkinje cells is considered to have resulted from demyelination of their myelinated axons. On the other hand, the reduction of granule cells is considered to be caused by the loss of Purkinje cells and presynaptic elements such as mossy fibers and glomerular collaterals of climbing fibers. The depletion of these neurons induced destruction of synaptic connection in the cerebellum.

Properties of sulfatases in cultured skin fibroblasts of multiple sulfatase deficient patients

Various sulfatase activities were assayed in cultured skin fibroblasts from patients with multiple sulfatase deficiency (MSD). MSD cell lines displayed deficiencies of arylsulfatase A and iduronate sulfatase, but activities of arylsulfatase B, N‐acetylgalactosamine 6‐sulfate sulfatase and N‐acetylglucosamine 6‐sulfate sulfatase were within normal ranges, but not consistently. Arylsulfatase A, minor anionic arylsulfatase and N‐acetylgalactosamine 6‐sulfate sulfatase in MSD cell lines had similar Km, pH optima, inhibitory or activator sensitivity to that of normal skin fibroblasts. Arylsulfatase B in MSD cell lines also had properties similar to that of normal skin fibroblasts, except an abnormal heat stability. From our results, we conclude that properties of arylsulfatase A, minor anionic arylsulfatase and N‐acetylgalactosamine 6‐sulfate sulfatase in MSD fibroblasts were intact. On the other hand, arylsulfatase B in MSD might be a functionally abnormal enzyme.

Degradation of keratan sulfate by ß‐N‐acetylhexosaminidases in GM2‐gangliosidosis

We have prepared a new substrate from a keratan sulfate‐derived‐oligosaccharide (2‐acetamido‐2‐deoxyglucosyl(1–3)‐[1‐3H] Galactitol), which is necessary to measure β‐N‐acetylhexosaminidase activity. This substrate was prepared from a cornea keratan sulfate by digestion with endo‐β‐galactosidase, followed by isolation of disaccharide on gel filtration chromatography and chemical desulfation. Using this substrate, we found that a striking deficiency of β‐N‐acetylhexosaminidase activity was present in the skin fibroblasts of patients with Sandhoff disease but not in Tay‐Sachs disease. Both β‐N‐acetyl‐hexosaminidase A & B contributed to the catabolism of keratan sulfate.

Electronmicroscopy of conjunctival biopsy in mannosidosis.

Electronmicroscopic examination was performed on conjunctival biopsies from two adolescent siblings with mannosidosis. Fitroblasts and kndothelial cells contained membrane-bound vacuoles and vesicles that contained homogeneous osmiophilic globules. These vesicles seem to be pathognomonic of mannosidosis. Plasma cells also contained membrane-bound vacuoles, suggesting inhibition of the immunoglobulin production.

Diagnosis of Pompe's disease using pyridylamino-maltooligosaccharides as substrates of α-1,4-glucosidase

We have developed a sensitive method for the assay of alpha-1,4-glucosidase in cultured skin fibroblasts and muscle tissue using pyridylamino-maltooligosaccharides as fluorescent substrates. This method is useful for the diagnosis of Pompes disease.

GLYCOSAMINOGLYCAN IN THE SKIN AND URINE OF A HUNTER SYNDROME PATIENT WITH A SPECIFIC SKIN LESION

The glycosaminoglycan (GAG) contents of the cutaneous papules, the non‐eruptive skin, and the urine of a patient with Hunter syndrome were examined. Both skin samples contained hyaluronic acid (HA) and dermatan sulfate (DS) as major components and heparan sulfate (HS) and chondroitin sulfate (CS) as minor components. The HA content of the papules was greatly increased, while that of the non‐eruptive skin was normal. The amounts of DS and HS were increased in all three samples; the papules, non‐eruptive skin, and urine. HS from the patients skin and urine had an electrophoretic mobility different from that of authentic HS. It seemed interesting from a pathogenetic viewpoint that the components of GAG in the cutaneous papules differed from those in the non‐eruptive skin and urine.

Impaired degradation of chondroitin sulfate in GM2-gangliosidosis

We have prepared a new radiolabeled substrate, derived from chondroitin 6‐sulfate oligosaccharide, for the assaying of chondroitin sulfate degradation by beta‐N‐acetylgalactosaminidase. Using this substrate, we found a striking deficiency of beta‐N‐acetylgalactosaminidase activity in the cultured skin fibroblasts of patients with Sandhoff disease and Tay‐Sachs disease. DEAE‐cellulose chromatography at pH 6.0 revealed that both isoenzymes A and B of beta‐N‐acetylgalactosaminidases from normal human liver participated in the catabolism of chondroitin 6‐sulfate. However, there were major differences in substrate specificity between isoenzyme A and isoenzyme B.