Peter F. Daniel

Demonstration of glucuronic acid on brain glycoproteins which react with HNK-1 antibody

Ependymins, a family of extracellular glycoproteins of goldfish and mammalian brain, were shown to contain N-linked complex glycan chains. These glycoproteins reacted with a monoclonal antibody, HNK-1 which recognizes a membrane antigen on a subset of human lymphocytes, myelin-associated glycoprotein glycoprotein epitope reacting with HNK-1 antibody was previously shown to include a terminal 3-sulfoglucuronosyl residue present in certain glycolipids of the nervous tissue (Chou et al., Biochem. Biophys. Res. Commun. 1985, 128, 383-388). In this report, the presence of glucuronic acid in ependymins was demonstrated by gas-liquid chromatography and mass spectrometry. We suggest that a 3-sulfoglucuronosyl residue may be the common epitope on HNK-1-reactive glycoproteins.

Infantile sialic acid storage disease associated with renal disease

A child with infantile sialic acid storage disease is reported. Ultrasonography demonstrated fetal ascites. At birth, the infant appeared hydropic and presented with numerous dysmorphic features, including sparse white hair, coarse facies, hypertelorism, epicanthal folds, anteverted nostrils, and a long philtrum. In addition, he had visceromegaly, bilateral inguinal hernias, and a slight gibbus deformity. Lymphocytes were vacuolated and bone marrow contained large numbers of foam cells. There were generalized vacuolations of both reticuloendothelial and parenchymal cells in the examined tissues. Neuropathologic studies revealed wide-spread neuronal storage, myelin loss, axonal spheroids, and gliosis. Neurons, endothelial cells, and Kupffer cells stained with wheat germ agglutinin indicated an accumulation of sialic acid. Free sialic acid was significantly increased in urine and serum, as well as in liver, heart, and brain tissues. The alpha-neuraminidase activity was normal. It is assumed that the basic defect of infantile sialic acid storage disease lies in impaired transport of sialic acid across the lysosomal membrane.

Oligosaccharides from placenta: early diagnosis of feline mannosidosis

High‐pressure liquid chromatography analysis of oligosaccharides from placentas allowed the diagnosis of α‐mannosidosis in three litters of kittens. The chromatography also afforded a detailed comparison of the oligosaccharide pattern and levels in placenta, liver, brain, urine and ocular fluid of the affected animals. In all cases, two series of compounds were observed, with one or two residues of N‐acetylglucosamine at the reducing terminus, respectively, and between two and nine mannose residues. This pattern is unlike that of human mannosidosis, and resembles that of ruminants, except that the major oligosaccharide contains three mannose residues instead of two.

Induced mannosidosis‐excretion of oligosaccharides by locoweed‐intoxicated sheep

Daily urine samples were collected from a locoweed‐fed sheep, and the oligosaccharide content examined by thin‐layer and liquid chromatography. An unusual pattern of urine oligosaccharides was observed, which appears to be characteristic of loco intoxication. Changes in the pattern could be correlated with the onset of visible disease, which occurred approximately 5 weeks after the typical urine sugars were first detected. HPLC showed that these sugars consisted of two homologous series of oligosaccharides containing one and two residues of 2‐acetamido‐2‐deoxy‐D‐glucose, respectively.

Characterization of oligosaccharides from the urine of loco-intoxicated sheep

Two major oligosaccharides were isolated by preparative HPLC from the urine of a locoweed‐fed sheep. Analysis by gas‐liquid chromatography and mass‐spectrometry indicated compositions of (Man)4(GlcNAc)2 and (Man)5(GlcNAc)2, respectively. Structures were determined by digestion with α‐D‐mannosidase and endo‐β‐N‐acetylglucosaminidases D and H, and comparison of the products by HPLC with synthetic standards, and oligosaccharides isolated from human mannosidosis urine. Incubation with an exo‐β‐N‐acetylglucosaminidase was without effect.

The accumulation of oligosaccharides in tissues and body fluids of cats with α-mannosidosis

Oligosaccharides were extracted from tissues and body fluids of five kittens with alpha-mannosidosis, three being from the same litter. The kittens were all of different ages at death and were compared to normal and heterozygote cats. The oligosaccharides were analyzed by high-pressure liquid chromatography after perbenzoylation and were identified by comparison with compounds of known structure. This provided a detailed picture of the distribution of oligosaccharides in each tissue, and a method for quantitation of the total oligosaccharides. With the exception of the youngest animal (death at day 2), the oligosaccharide elution profiles were broadly similar for all tissues and fluids, and were typical of feline alpha-mannosidosis. In contrast, concentrations of total oligosaccharides diverged widely from one source to another, from a high of 17.3 mumol/g to a low of 0.04 mumol/g. The results are interpreted in the context of glycoprotein catabolism.

Alpha-Mannosidase Deficiency in Persian Cats: A Model of Human Alpha-Mannosidosis

A family of cats deficient in lysosomal α-mannosidase activity was studied. Analysis of the family pedigree indicated an autosomal recessive mode of inheritance. All affected kittens had frontal bossing, facial coarsening and dysostosis multiplex. They had retarded growth and developed progressive ataxia, tremor, corneal and lenticular opacities, and hepatomegaly. Thin layer chromatography and high pressure liquid chromatography of urine revealed partially degraded N glycoprotein-derived oligosaccharides. Activity of acidic α-mannosidase from plasma and leukocytes of affected kittens was less than 2% of normal, while the enzyme activity of heterozygotes was less than 50%. In blood smears, 90% of circulating lymphocytes were vacuolated. Ultrastructural studies of circulating white blood cells revealed vacuoles containing storage material in various cell types. Affected animals died or were killed at an early age due to the severity of the disease. Most cell types in visceral, neural and skeletal organs were affected. These cells were enlarged, vacuolated and contained fine granular material or appeared empty. The course of this disease in cats and the morphological and biochemical changes, are similar to those seen in the infantile phenotype of human α-mannosidosis.

Biochemical, ultrastructural and histochemical studies of cat placentae deficient in activity of lysosomal α-mannosidase

Lysosomal alpha-mannosidase activity, oligosaccharide profiles, light and electron microscopy and lectin histochemistry studies were performed on full-term placentae obtained from five litters of cats. They resulted from breeding related cats who are obligate heterozygotes for lysosomal alpha-mannosidase deficiency. alpha-Mannosidase activity in placentae from affected kittens was less than 10 per cent of control, while in placentae from presumptive heterozygotes the activity was less than 50 per cent of control. High-pressure liquid chromatographic analysis of oligosaccharides revealed massive accumulation of undegraded oligosaccharides in placentae of affected kittens. A small elevation was found in placentae from presumptive heterozygous kittens, and none was detected in placentae of normal kittens. Light and electron microscopic examinations revealed vacuolization of fetal endothelial and mesenchymal cells only in placentae of affected kittens. Succinylated wheat germ agglutinin and concanavalin A stained the fetal fibroblasts only in placentae of affected kittens.

High Performance Liquid Chromatography of Oligosaccharides from Human Milk and Colostrum

Human milk is unique with respect to the high concentration and complexity of its oligosaccharide fraction. Polonovsky and Lespagnol1 isolated and studied a human milk fraction which they named gynolactose and reported that it contained mostly oligosaccharides, Montreuil and Mullet2 determined that this fraction comprises 2.4% of human colostrum and between 1.2 and 1.3% of mature human milk. Thus, the oligosaccharide fraction is the fourth largest constituent of human milk, after water, lactose, and fat, and represents a significant portion of the non-protein nitrogen found therein. This fraction proved to be a rich source of a wide variety of oligosaccharides, in contrast to the analogous fraction from bovine milk which comprised only 0.1% and consisted of a limited number of oligosaccharide types3