GM2 gangliosidosis variant B1

Variant B1 is a rare type of GM2 gangliosidosis. Clinically, it shows a wide spectrum of forms ranging from infantile to juvenile. We report the first magnetic resonance imaging (MRI) findings from three patients affected by GM2 gangliosidosis variant B1, two presenting with the infantile form and one with the juvenile form. The MRI appearances of the two patients with the infantile form disease are congruent with those reported for the early-onset type of both Tay-Sachs and Sandhoff diseases, and are characterized by early involvement of the basal ganglia and thalamus with cortical atrophy appearing later. In contrast, the patient with the juvenile form of variant B1 showed progressive cortical and white-matter atrophy of the supratentorial structures and, to a lesser extent, the infratentorial structures. No basal ganglia or thalamic anomalies were observed. Because in the adult forms of both Tay-Sachs and Sandhoff diseases a progressive cerebellar atrophy represents the only abnormality detectable, it appears that an MRI pattern peculiar to GM2 gangliosidosis can be defined. This pattern ranges from the basal ganglia injury associated with the early and severe demyelination process noted in the infantile form of the disease, to cerebellar atrophy with no supratentorial anomalies in the adult form. An “intermediate” MRI picture, with cortical atrophy and mild cerebellar atrophy, but without basal ganglia impairment, can be observed in the juvenile form. In addition, our investigations suggest that MRI abnormalities in GM2 gangliosidosis correlate with the clinical form of the disease rather than with the biochemical variant of the enzymatic defect. Received: 9 January 2002, Received in revised form: 26 June 2002, Accepted: 8 July 2002 Correspondence to Paolo Balestri, M. D.     

Does Serum Hexosaminidase Activity Play a Role in the Diagnosis of Strangulated Bowel Obstruction? An Experimental Study

Strangulation is associated with an increased risk of mortality and morbidity in patients with mechanical bowel obstruction. The accurate and early recognition of the presence of strangulation is important to allow safe nonoperative treatment. A number of studies have shown that there was no single and reliable test to detect or exclude the presence of strangulation. The aim of this study was to evaluate the role of serum hexosaminidase (Hex) levels in recognition of strangulation in an experimental model of closed loop small bowel obstruction. Forty-two Wistar albino rats were divided into four groups: I, control (n = 5); II, sham laparotomy (n = 5); III, simple obstruction (n = 16); and IV, strangulation groups (n = 16). Activity levels of total Hex and its fractions (Hex A and B) were assayed in serum samples obtained from rats after 3 and 8 hr. Samples of small bowel were also evaluated histologically. Histological evaluation of bowel sections obtained from the strangulation group after 8 hr, revealed transmural hemorrhagic infarction in all animals with a mean +/- SD total Hex activity of 978.25 +/- 150 nmol/hr/ml, which was significantly higher than that in the other groups (P < 0.001). Although sections of bowel from the strangulation group after 3 hr showed severe ischemic injury, the activities of total Hex, Hex A, and Hex B were not different from those of the control, sham, and simple obstruction groups. Histological examination of these groups did not show any sign of ischemia. Total Hex, Hex A, and Hex B activities in the strangulation group were all significantly greater than the activities seen in the simple obstruction group (P < 0.001, for all). In conclusion, increased serum hex levels indicate irreversible transmural infarction only in the late period of strangulation in the closed loop small bowel obstruction model. It seems unuseful for detecting reversible and/or irreversible ischemia in the early period of strangulation.     

Isoenzyme studies in human leukemia-lymphoma cell lines--III. beta-Hexosaminidase (E.C. 3.2.1.30)

The hexosaminidase (beta-N-acetylglucosaminidase) isoenzyme profiles of 86 human hematopoietic cell lines grown actively in suspension culture were analysed by isoelectric focusing and by conventional disc electrophoresis on horizontal thin-layer polyacrylamide gels. A maximum of three hexosaminidase (Hex) isoenzymes (A = anodic, I = intermediate, B = basic) could be demonstrated. The immunological phenotyping of 74 leukemia-lymphoma derived cell lines had led to a categorization into four groups with a subclassification of the T- and B-cell lines into several stages of differentiation: 26 T-cell, 34 B-cell, 6 myelomonocytic and 8 Non-T, Non-B cell leukemia-lymphoma cell lines. Twelve so-called 'normal' B-lymphoblastoid cell lines were also available. Distinct isoenzyme profiles were seen in the different stages of differentiation in the T- and B-leukemia-lymphoma cell lines. Among the 12 normal B-lymphoblastoid cell lines heterogeneity in the isoenzymatic phenotypes was detected. Hex isoenzyme expression in normal and neoplastic lymphoid cell lines represents hypothetically sequential stages of T- and B-cell differentiation. Myelomonocytic cell lines displayed strongly stained bands of all three isoenzymes. Heterogeneity was seen in the group of Non-T, Non-B cell lines. Four out of 5 pre B-cell lines and 4 out of 4 Non-T, Non-B cell lines which are comparable to cases of pre B- and common ALL revealed a high Hex I/Hex A ratio in terms of intensity of the isoenzyme bands. The analysis of Hex isoenzymes is useful for characterizing lymphoid and myeloid populations (both normal and malignant, cultured or fresh), particularly with regard to their stage of differentiation. But this enzyme should be part of a multiple enzyme study where the information obtained is complementary. In turn, enzyme marker analysis should be included in the multiple marker analysis for an optimized characterization of leukemic cells.     

Effective mast cell degranulating peptide inhibitors of the IgE/Fc epsilonRI receptor interaction

Previous studies with mast cell degranulating (MCD) peptide have shown that peptide [Ala(12)]MCD 8 was an inhibitor of IgE binding to mast cell receptors. In an attempt to produce increased inhibition, analogs were synthesized that maintained the alanine residue in position 12 in the MCD peptide sequence and were further modified at both termini. Analogs modified at the C-terminus were [Ala(12),desLys(21)]MCD 2 and [Ala(12),D-Lys(21)]MCD 4. N-terminus modifications were [desLys(6)-Arg(7)-His(8),Ala(12)]MCD 1, [Ala(6), Ala(12)]MCD 6, and [Val(6),Ala(12)]MCD 7. To assess the role of the Proline(12), analogs [D-Ala(12)]MCD 3 and [Meleu(12)]MCD 5 were also synthesized. The analogs were tested for binding to the IgE receptor in cultured mast cells. Inhibitory activity of IgE-caused degranulation was measured using a beta-hexosaminidase assay. Circular dichroism (CD) and molecular modeling of selected analogs were used to follow possible structural differences among these analogs. All analogs showed binding affinity to the IgE receptor and inhibition of IgE-induced mast cell degranulation at different levels. Differences in inhibition were most likely because of diverse interactions of the analogs with the receptor as inferred by the CD and modeling studies. Based on the results of the beta-hexosaminidase assay, analog [Val(6), Ala(12)]MCD 7 proved to be an excellent inhibitor of IgE-mediated mast cell degranulation.     

Two mutated HEXA alleles in a Druze patient with late-infantile Tay-Sachs disease

Two affected HEXA alleles were found in an Israeli Druze Tay-Sachs child born to first-cousin parents. His paternal allele contained two adjacent changes in exon 5: Δ496C, which resulted in a frameshift and premature termination codon 96 nucleotides downstream, and 498C→G, a silent mutation. The maternal allele had a 835T→C transition in exon 8 (S279P). Phosphoimaging quantitation of the parents' RNAs showed that the steady-state levels of mRNAs of the mutant exons 5 and 8 were 5% and 50%, respectively, of normal levels. The exon 5 mutated allele with the premature translation termination resulted in severe deficiency of Hex A. Transient expression of the exon 8 mutated α-chain cDNA in COS-1 cells resulted in deficiency of enzymatic activity. The child exhibited a late-infantile-type disease. Hum Mutat 10:451–457, 1997. © 1997 Wiley-Liss, Inc.     

Constitutive expression of β-N-acetylhexosaminidase in a microglial cell line: Transcriptional modulation by lipopolysaccharide and serum factors

We investigated the expression of the α- and β-subunits of the lysosomal enzyme β-N-acetylhexosaminidase in the BV-2 microglial cell line under different culture conditions. β-N-acetylhexosaminidase from BV-2 microglia cells was separated into its constituent isoenzymes on diethylaminoethyl (DEAE) cellulose, and its activity was monitored with 4-methylumbelliferyl-β-N-acetylglucosamine and 4-methylumbelliferyl-β-N-acetylglucosamine-6-sulphate substrates. Forms corresponding to the mouse isoenzymes A and B were present in the cells incubated in serum-supplemented medium as well as in serum-free medium. Lipopolysaccharide, a well-known activator of microglia in vitro, added to the BV-2 cells in serum-supplemented medium induced a decrease in the specific enzymatic activity determined with the 4-methylumbelliferyl-β-N-acetylglucosamine substrate. Lipopolysaccharide had no effect on hexosaminidase isoenzyme pattern of BV-2 cells in serum-supplemented medium. The level of α-subunit mRNA was increased and the level of β-subunit mRNA was decreased in BV-2 cells incubated in serum-supplemented medium plus lipopolysaccharide. In the cells incubated in a serum-free medium no significant changes in the hexosaminidase-specific activities towards the above substrates were observed. Interestingly, increased expression of α- and β-subunit mRNA was evident in comparison with cultures in serum-supplemented medium. The present results suggest that the BV-2 cell line may be a useful tool to study the possible role of microglia in the metabolism of brain glycolipids. J. Neurosci. Res. 50:44–49, 1997. © 1997 Wiley-Liss, Inc.     

Lysosomal hydrolases in cerebrospinal fluid from subjects with Parkinson's disease.

Recent studies have shown a genetic association between glucocerebrosidase deficiencies and Parkinson's disease (PD). To further explore this issue the activity of beta-glucocerebrosidase and the activities of other lysosomal enzymes, alpha-mannosidase, beta-mannosidase, beta-hexosaminidase, and beta-galactosidase have been evaluated in the cerebrospinal fluid (CSF) of PD patients. The activities of alpha-mannosidase, beta-mannosidase, beta-glucocerebrosidase, and beta-hexosaminidase were substantially decreased in the CSF of PD patients, while levels of beta-galactosidase were essentially identical to controls. This study indicates that in PD several lysosomal hydrolases have decreased activities, further supporting a possible link between pathophysiological mechanisms underlying PD and lysosomal hydrolases.     

The mutation mechanism causing juvenile-onset Tay-Sachs disease among Lebanese

Expression of the hexosaminidase isozymes was evaluated in fibroblast cell lines obtained from two sibs of Lebanese-Christian origin who presented with juvenile-onset Tay-Sachs disease. In the normal control fibroblasts the alpha subunit of hexosaminidase A (hex A) is synthesized as a 67 KD precursor which is cleaved in lysosomes to a mature 54 KD peptide. The patients' fibroblasts were capable of synthesizing the 67 KD precursor but failed to convert it to the mature subunit. The alpha subunit precursor synthesized by patients' cells could not be phosphorylated, nor was the patients' alpha subunit precursor secreted into the medium in response to NH4Cl, which caused accumulation of both alpha and beta subunit precursor in the medium of the normal control fibroblasts. The measurement of residual enzyme activity in the fibroblasts of patients which best correlated with the onset of the illness was the ion exchange chromatographic separation of Hex A-associated hydrolysis of the synthetic substrate 4-methylumbelliferyl N-acetyl-beta-D-glucosamine-6-sulfate (4MUGS). The patients had 0.32% and 0.36% of Hex A-associated 4MUGS cleaving activity compared to normal control fibroblasts as compared to less than 0.016% for infantile Tay-Sachs disease fibroblasts. The residual Hex A activity in patients' cells had a pH optimum identical with normal enzyme (pH 3.9-4.0), a reduced specific activity for 4MUGS (relative to hydrolysis of unsulfated synthetic substrate), and a greatly enhanced thermal stability. The occurrence of this form of Tay-Sachs disease in Lebanon, the fact that the condition has been described in three unrelated Lebanese immigrant families in Canada, together with the fact that the grandparents of the unrelated probands come from villages in both the northern and southern regions of Lebanon, leads us to speculate that a gene causing juvenile-onset Tay-Sachs disease may not be infrequent in Lebanon.