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Biomedical subjects

E Ong

Publications and source records attributed to E Ong.

At least 19 recordsLinked to original sources
PubMed2002

Identification and characterization of novel mutations of the aspartoacylase gene in non-Jewish patients with Canavan disease.

Canavan disease, an inherited leukodystrophy, is caused by mutations in the aspartoacylase (ASPA) gene. It is most common among children of Ashkenazi Jewish descent but has been diagnosed in many diverse ethnic groups. Two mutations comprise the majority of mutant alleles in Jewish patients, while mutations in the ASPA gene among non-Jewish patients are different and more diverse. In the present study, the ASPA gene was analysed in 22 unrelated non-Jewish patients with Canavan disease, and 24 different mutations were found. Of these, 14 are novel, including five missense mutations (E24G, D68A, D249V, C152W, H244R), two nonsense mutations (Q184X, E214X), three deletions (923delT, 33del13, 244delA), one insertion mutation (698insC), two sequence variations in one allele ([10T>G; 11insG]), an elimination of the stop codon (941A>G, TAG-->TGG, X314W), and one splice acceptor site mutation (IVS1 - 2A>T). The E24G mutation resulted in substitution of an invariable amino acid residue (Glu) in the first esterase catalytic domain consensus sequence. The IVS1 - 2A>T mutation caused the retention of 40 nucleotides of intron 1 upstream of exon 2. The results of transient expression of the mutant ASPA cDNA containing these mutations in COS-7 cells and assays for ASPA activity of patient fibroblasts indicated that these mutations were responsible for the enzyme deficiency. In addition, patients with the novel D249V mutation manifested clinically at birth and died early. Also, patients with certain other novel mutations, including C152W, E214X, X314W, and frame shift mutations in both alleles, developed clinical manifestations at an earlier age than in classical Canavan disease.

Amidohydrolases↗
PubMed2001

Rapid detection of the two common mutations in Ashkenazi Jewish patients with mucolipidosis type IV.

Among Ashkenazi Jewish individuals with mucolipidosis IV (ML IV), two mutations in the ML IV gene, IVS3-1A --> G and delEX1-EX7, account for more than 95% of disease alleles. The reported method of genotyping for the delEX1-EX7 mutation involves a cumbersome multistep procedure. In the present study, a new simplified one-step procedure is described that detects this mutation in both patients and carriers. An improved procedure is also described for detection of the IVS3-1A --> G mutation. Using these improved procedures, we have characterized the ML IV mutant alleles in 27 patients and 95 of their relatives from 22 families, and in 123 unrelated and unaffected Ashkenazi Jewish controls. Of the 27 ML IV patients, 16 patients (59.3%) were found to be homozygous for the IVS3-1A --> G mutation and 1 patient (3.7%) homozygous for the delEX1-EX7 mutation. Additionally, 9 patients (33.3%) were compound heterozygotes for IVS3-1A --> G/delEX1-EX7. Among the 123 Ashkenazi Jewish controls, two individuals were identified as heteroallelic with one IVS3-1A --> G mutation (carrier frequency: approximately 1 in 61); none showed the delEX1-EX7 mutation. The modifications described here provide a more facile means of genotyping patients and carriers and expand the possibilities for screening at-risk populations.

Adult↗
PubMed2001

Polysialyltransferase ST8Sia II (STX) polysialylates all of the major isoforms of NCAM and facilitates neurite outgrowth.

The neural cell adhesion molecule (NCAM) has different isoforms due to different sizes in its polypeptide and plays a significant role in neural development. In neural development, the function of NCAM is modified by polysialylation catalyzed by two polysialyltransferases, ST8Sia II and ST8Sia IV. Previously, it was reported by others that ST8Sia II polysialylates only transmembrane isoforms of the NCAM, such as NCAM-140 and NCAM-180, but not NCAM-120 and NCAM-125 anchored by a glycosylphosphotidylinositol. In the present study, we first discovered that ST8Sia II polysialylates all isoforms of the NCAM examined, and we demonstrated that polysialylation of NCAM expressed on 3T3 cells facilitates neurite outgrowth regardless of isoforms of NCAM, where polysialic acid is attached. We then show that neurite outgrowth is significantly facilitated only when polysialylated NCAM is present in cell membranes. Moreover, the soluble NCAM coated on plates did not have an effect on neurite outgrowth exerted by soluble L1 adhesion molecule coated on plates. These results, taken together, indicate that ST8Sia II plays critical roles in modulating the function of all major isoforms of NCAM. The results also support previous studies showing that a signal cascade initiated by NCAM differs from that initiated by L1 molecule.

3T3 Cells↗
PubMed2000

Molecular cloning and expression of two distinct human chondroitin 4-O-sulfotransferases that belong to the HNK-1 sulfotransferase gene family.

Using an expression cloning strategy, the cDNA encoding the human HNK-1 sulfotransferase (HNK-1ST) has been cloned. During this cloning we found that HNK-1ST and other Golgi-associated sulfotransferases cloned before share homologous sequences including the RDP motif (Ong, E., Yeh, J.-C., Ding, Y., Hindsgaul, O., and Fukuda, M. (1998) J. Biol. Chem. 223, 5190-5195). Using this conserved sequence in HNK-1ST as a probe, we identified two expressed sequence tags in EST data base which have 31.6 and 30.7% identity with HNK-1ST at the amino acid levels. Expression of these two full-length cDNAs failed to form HNK-1 glycan nor to add sulfate to CD34 or NCAM. Surprisingly, proteins expressed by these cDNAs transferred sulfate to the C-4 position of N-acetylgalactosamine in chondroitin and desulfated dermatan sulfate, thus we named these two enzymes, chondroitin 4-O-sulfotransferase 1 and -2 (C4ST-1 and C4ST-2). Both C4ST-1 and C4ST-2, however, did not form 4, 6-di-O-sulfated N-acetylgalactosamine when chondroitin sulfate C was used as an acceptor. Moreover, analysis of (35)S-labeled dermatan sulfate formed by C4ST-1 indicate that sulfation preferentially took place in GlcA-->GalNAc unit than in IdoA-->GalNAc unit, suggesting that 4-O-sulfation at N-acetylgalactosamine may precede epimerization of glucuronic acid to iduronic acid during dermatan sulfate biosynthesis. Northern analysis demonstrated that the transcript for C4ST-1 is predominantly expressed in peripheral leukocytes and hematopoietic tissues while the C4ST-2 transcript is more widely expressed in various tissues. These results indicate C4ST-1 and C4ST-2 play complementary roles in chondroitin and dermatan sulfate synthesis in different tissues.

Amino Acid Sequence↗
PubMed2000

Plasmapheresis treatment for recurrent focal sclerosis in pediatric renal allografts.

Recurrence of focal segmental glomerulosclerosis (FSGS) in pediatric renal allografts is associated with a poor graft survival. This study reports on plasmapheresis for the treatment of recurrent FSGS in pediatric renal transplant recipients. The records of 100 consecutive pediatric (age <21 years) renal transplants were reviewed. Twenty patients had FSGS as the cause of renal failure. Eight of these (40%) had a recurrence (proteinuria >1 g/m2 per day) within 1 month of transplantation. Five of six patients treated with plasmapheresis went into remission (<0.2 g/m2 per day), receiving a total of 42+/-26 (12-73) sessions, with the mean number of sessions required to achieve a remission being 24+/-17 (8-51). One patient had a second recurrence 1 year following cessation of plasmapheresis and responded to another course of plasmapheresis. The 1 patient who did not respond to plasmapheresis had a delay in initiation of therapy of 42 days. Plasmapheresis initiated within 48 h of recurrence resulted in earlier remissions and improved graft survival among our patients. Plasmapheresis appears to be effective in treating recurrent FSGS following kidney transplantation and should be started as soon as possible. The number of plasmapheresis sessions used to achieve remission should be adjusted according to response rather than adhering to a fixed protocol.

Adolescent↗
PubMed2000

An aggressive intrasinusoidal lymphoma presenting with marked systemic disturbance but normal imaging studies.

A patient with an aggressive intrasinusoidal non-Hodgkins lymphoma, presenting with marked systemic disturbance but only a mildly raised alkaline phosphatase as a localising sign is described. All imaging studies of the liver were normal and the diagnosis was delayed until a percutaneous liver biopsy was performed. Once diagnosed, the patient responded extremely well to conventional anti-lymphoma chemotherapy.

False Negative Reactions↗
PubMed1999

Structure and function of HNK-1 sulfotransferase. Identification of donor and acceptor binding sites by site-directed mutagenesis.

HNK-1 glycan, sulfo-->3GlcAbeta1-->3Galbeta1-->4GlcNAc-->R, is uniquely enriched in neural cells and natural killer cells and is thought to play important roles in cell-cell interaction. HNK-1 glycan synthesis is dependent on HNK-1 sulfotransferase (HNK-1ST), and cDNAs encoding human and rat HNK-1ST have been recently cloned. HNK-1ST belongs to the sulfotransferase gene family, which shares two homologous sequences in their catalytic domains. In the present study, we have individually mutated amino acid residues in these conserved sequences and determined how such mutations affect the binding to the donor substrate, adenosine 3'-phosphate 5'-phosphosulfate, and an acceptor. Mutations of Lys(128), Arg(189), Asp(190), Pro(191), and Ser(197) to Ala all abolished the enzymatic activity. When Lys(128) and Asp(190) were conservatively mutated to Arg and Glu, respectively, however, the mutated enzymes still maintained residual activity, and both mutant enzymes still bound to adenosine 3',5'-diphosphate-agarose. K128R and D190E mutant enzymes, on the other hand, exhibited reduced affinity to the acceptor as demonstrated by kinetic studies. These findings, together with those on the crystal structure of estrogen sulfotransferase and heparan sulfate N-deacetylase/sulfotransferase, suggest that Lys(128) may be close to the 3-hydroxyl group of beta-glucuronic acid in a HNK-1 acceptor. In contrast, the effect by mutation at Asp(190) may be due to conformational change because this amino acid and Pro(191) reside in a transition of the secondary structure of the enzyme. These results indicate that conserved amino acid residues in HNK-1ST play roles in maintaining a functional conformation and are directly involved in binding to donor and acceptor substrates.

Amino Acid Sequence↗
PubMed1999

Molecular cloning and expression of a novel beta-1, 6-N-acetylglucosaminyltransferase that forms core 2, core 4, and I branches.

Mucin-type O-glycans are classified according to their core structures. Among them, cores 2 and 4 are important for having N-acetyllactosamine side chains, which can be further modified to express various functional oligosaccharides. Previously, we discovered by cloning cDNAs that the core 2 branching enzyme, termed core 2 beta-1,6-N-acetylglucosaminyltransferase-leukocyte type (C2GnT-L), is highly homologous to the I branching beta-1, 6-N-acetylglucosaminyltransferase (IGnT) (Bierhuizen, M. F. A., Mattei, M.-G., and Fukuda, M. (1993) Genes Dev. 7, 468-478). Using these homologous sequences as probes, we identified an expressed sequence tag in dbEST, which has significant homology to C2GnT-L and IGnT. This approach, together with 5'and 3' rapid amplification of cDNA ends, yielded a human cDNA that encompasses a whole coding region of an enzyme, termed C2GnT-mucin type (C2GnT-M). C2GnT-M has 48.2 and 33.8% identity with C2GnT-L and IGnT at the amino acid levels. The expression of C2GnT-M cDNA directed the expression of core 2 branched oligosaccharides and I antigen on the cell surface. Moreover, a soluble chimeric C2GnT-M had core 4 branching activity in addition to core 2 and I branching activities. A soluble chimeric C2GnT-L, in contrast, almost exclusively contains core 2 branching activity. Northern blot analysis demonstrated that the C2GnT-M transcripts are heavily expressed in colon, small intestine, trachea, and stomach, where mucin is produced. In contrast, the transcripts of C2GnT-L were more widely detected, including the lymph node and bone marrow. These results indicate that the newly cloned C2GnT-M plays a critical role in O-glycan synthesis in mucins and might have distinctly different roles in oligosaccharide ligand formation compared with C2GnT-L.

Amino Acid Sequence↗
PubMed1999

Desensitization to co-trimoxazole (trimethoprim-sulphamethoxazole) in HIV-infected patients: is patch testing a useful predictor of reaction?

OBJECTIVE: To establish the safety and efficacy of desensitization to co-trimoxazole in hypersensitive HIV-infected subjects. To assess if delayed hypersensitivity (type IV) to co-trimoxazole predicts those unable to be desensitized. METHOD: desensitization to co-trimoxazole, comprising trimethoprim (T) 0.4 mg and sulphamethoxazole (S) 2 mg initially with doubling dose daily, full strength co-trimoxazole (T/S 160 mg/800 mg) at 10 days. Patch testing with 4.5% and 9% co-trimoxazole in yellow soft paraffin, CMI Multitest. RESULTS: nineteen patients, 18 male and one female, were recruited and completed the desensitization regime. Of these 80%(15) achieved successful desensitization. Three of those who reacted did so within 18 days. All patients were successfully managed in an outpatient setting. There were no major adverse reactions. Of those reacting none gave a positive patch test to co-trimoxazole and all showed absent delayed type hypersensitivity reactions to recall antigens. CONCLUSIONS: co-trimoxazole desensitization is a safe and efficacious procedure, with a success rate of 80% using the above regime. Patch testing with co-trimoxazole gives no useful information about those that reacted.

Adult↗
PubMed1999

Patient satisfaction with HIV service provision in NPMS hospitals: the development of a standard satisfaction questionnaire. NPMS Steering Group.

A self-completion satisfaction questionnaire evaluating the standard of care of HIV outpatient services was developed as part of the National Prospective Monitoring System on the Use, Cost and Outcome of HIV Service Provision in English Hospitals (NPMS). The questionnaire was designed in conjunction with service users and health care professionals, and piloted in three London and three non-London HIV clinics. In addition to testing alternative methods of administering the questionnaire, the pilot provided satisfaction scores on a variety of aspects of service provision for participating clinics. The questionnaire was completed by 548 respondents and was most effectively collected using a sealed box in the clinic waiting area. Mean satisfaction scores for the attitude and skills of staff members was 4.7 (95% CI 4.6-4.7) but satisfaction scores were significantly lower for the clinic environment with a mean of 4.1 (95% CI 4.1-4.2). Satisfaction scores did not differ significantly by gender, age, sexual orientation, ethnic group, employment status or severity of symptoms. London respondents were more satisfied with the clinic environment and seeing preferred members of staff than their non-London counterparts, however there were no other differences between clinics. The questionnaire functioned well in practice and provided meaningful and useful information for individual clinics as well as at aggregate level.

Acquired Immunodeficiency Syndrome↗
PubMed1999

Effects of spectacle intervention on the progression of myopia in children.

The literature on myopigenesis suggests an active emmetropization mechanism regulated by optical defocus. The strongest evidence comes from compensatory ocular growth in response to lens-induced defocus in different species of animals. Based on these results, it has been suggested that, however useful, spectacle intervention for the optical correction of human myopia would lead to its exacerbation. The present study seeks to evaluate the progression of juvenile-onset myopia in children differentiated by their lens wear patterns. Data from 43 myopes from our longitudinal study of refraction were evaluated, with myopia defined as a spherical equivalent of at least -0.50 D. Refractions were obtained in the laboratory by noncycloplegic retinoscopy performed by one experienced optometrist at regular intervals. Information regarding the subjects' prescription lens-wearing history was obtained from the subjects and their eye care providers. Based on their wearing patterns, subjects were divided into four categories: (1) full-time wearers; (2) myopes who switched from distance to full-time wear; (3) distance wearers; and (4) nonwearers. Exponential functions were fit to the individual refraction data. The age of onset of myopia, the mean myopia at onset of spectacle wear, and the refractive shift over a period of at least 3 years were derived from these fits. Results show that the 3-year refractive shifts are not significantly different among the four groups. A comparison of the extreme conditions, i.e., full-time vs. nonwear categories, also revealed no significant difference when the data were corrected for age effects despite the fact that the nonwearers exhibited an age-adjusted 3-year progression approximately one-half that of the full-time wearers. In summary, the present study failed to demonstrate any overall effects of spectacle intervention on the progression of human myopia. Further investigation using a larger sample is warranted.

Age of Onset↗
PubMed1998

A cytoplasmic protein, bystin, interacts with trophinin, tastin, and cytokeratin and may be involved in trophinin-mediated cell adhesion between trophoblast and endometrial epithelial cells.

Trophinin and tastin form a cell adhesion molecule complex that potentially mediates an initial attachment of the blastocyst to uterine epithelial cells at the time of implantation. Trophinin and tastin, however, do not directly bind to each other, suggesting the presence of an intermediary protein. The present study identifies a cytoplasmic protein, named bystin, that directly binds trophinin and tastin. Bystin consists of 306 amino acid residues and is predicted to contain tyrosine, serine, and threonine residues in contexts conforming to motifs for phosphorylation by protein kinases. Database searches revealed a 53% identity of the predicted peptide sequence with the Drosophila bys (mrr) gene. Direct protein-protein interactions of trophinin, tastin, and bystin analyzed by yeast two-hybrid assays and by in vitro protein binding assays indicated that binding between bystin and trophinin and between bystin and tastin is enhanced when cytokeratin 8 and 18 are present as the third molecule. Immunocytochemistry of bystin showed that bystin colocalizes with trophinin, tastin, and cytokeratins in a human trophoblastic teratocarcinoma cell, HT-H. It is therefore possible that these molecules form a complex and thus are involved in the process of embryo implantation.

Amino Acid Sequence↗
PubMed1998

Expression cloning of a human sulfotransferase that directs the synthesis of the HNK-1 glycan on the neural cell adhesion molecule and glycolipids.

The HNK-1 carbohydrate is expressed on various adhesion molecules in the nervous system and is suggested to play a role in cell-cell and cell-substratum interactions. Here we describe the isolation and functional expression of a cDNA encoding a human sulfotransferase that synthesizes the HNK-1 carbohydrate epitope. A mutant Chinese hamster ovary cell line, Lec2, which stably expresses human neural cell adhesion molecule (N-CAM) (Lec2-NCAM), was first established. Lec2-NCAM was co-transfected with a human fetal brain cDNA library, a cDNA encoding the rat glucuronyltransferase that forms a precursor of the HNK-1 carbohydrate, and a vector encoding the polyoma large T antigen. The transfected Lec2-NCAM cells expressing the HNK-1 glycan were enriched by fluorescence-activated cell sorting. Sibling selection of recovered plasmids resulted in a cDNA encoding a sulfotransferase, HNK-1ST, that directs the expression of the HNK-1 carbohydrate epitope on the cell surface. The deduced amino acid sequence indicates that the enzyme is a type II membrane protein. Sequence analysis revealed that there is a short amino acid sequence in the presumed catalytic domain, which is highly homologous to the corresponding sequence in other Golgi-associated sulfotransferases so far cloned. The amount of HNK-1ST transcript is high in fetal brain compared with fetal lung, kidney, and liver. Expression of HNK-1ST resulted in the formation of the HNK-1 epitope on N-CAM and a soluble chimeric form of HNK-1ST was shown to add a sulfate group to a precursor, GlcAbeta1-->3Galbeta1-->4GlcNAcbeta1-->R, forming sulfo-->3GlcAbeta1-->3Galbeta1-->4GlcNAcbeta1-->R. The results combined together indicate that the cloned HNK-1ST directs the synthesis of the HNK-1 carbohydrate epitope on both glycoproteins and glycolipids in the nervous tissues.

Amino Acid Sequence↗