Biomedical subjects

J Ng

Publications and source records attributed to J Ng.

At least 91 records · Page 5Linked to original sources

PubMed1997

Analysis of HLA-A and -B serologic typing of bone marrow registry donors using polymerase chain reaction with sequence-specific oligonucleotide probes and DNA sequencing.

Unrelated volunteer donors (69) recruited by the National Marrow Donor Program were HLA typed by DNA-based methods for both the HLA-A and -B loci. Each donor had been previously typed by serology by at least two independent laboratories. Of the 69 samples, all serologic laboratories were in concordance for HLA-A in 62 typed samples and for HLA-B in 48 typed samples. Of the serologically concordant samples, 5 samples typed for HLA-A and 7 samples typed for HLA-B received DNA and serology types differing in their level of resolution. One sample typed for HLA-A and 3 samples typed for HLA-B by DNA methods gave different results from their serologic assignments. Of the samples exhibiting disparities among the different serologic typing laboratories, the DNA-defined types of 7 samples typed for HLA-A and 18 samples typed for HLA-B were consistent with at least one of the serologic assignments. The DNA types for the remaining 3 HLA-B typed samples did not agree with the serologic assignments and their alleles were subsequently sequenced. One of these sequences was a previously undefined allele, B*1537. Sharing of polymorphic sequences among HLA allelic products creates difficulties for consistent serologic assignments of some types complicating the process of identifying potential donors from bone marrow registries. Thus, the use of DNA-based typing techniques for characterization of donor class I types should allow a more consistent definition of types and should speed the donor selection process.

Bone Marrow Transplantation↗

PubMed1997

Evolutionary conservation and predicted structure of the Drosophila extra sex combs repressor protein.

The Drosophila extra sex combs (esc) protein, a member of the Polycomb group (PcG), is a transcriptional repressor of homeotic genes. Genetic studies have shown that esc protein is required in early embryos at about the time that other PcG proteins become engaged in homeotic gene repression. The esc protein consists primarily of multiple copies of the WD repeat, a motif that has been implicated in protein-protein interaction. To further investigate the domain organization of esc protein, we have isolated and characterized esc homologs from divergent insect species. We report that esc protein is highly conserved in housefly (72% identical to Drosophila esc), butterfly (55% identical), and grasshopper (56% identical). We show that the butterfly homolog provides esc function in Drosophila, indicating that the sequence similarities reflect functional conservation. Homology modeling using the crystal structure of another WD repeat protein, the G-protein beta-subunit, predicts that esc protein adopts a beta-propeller structure. The sequence comparisons and modeling suggest that there are seven WD repeats in esc protein which together form a seven-bladed beta-propeller. We locate the conserved regions in esc protein with respect to this predicted structure. Site-directed mutagenesis of specific loops, predicted to extend from the propeller surface, identifies conserved parts of esc protein required for function in vivo. We suggest that these regions might mediate physical interaction with esc partner proteins.

Amino Acid Sequence↗

PubMed1996

Targeted disruption of the mouse apobec-1 gene abolishes apolipoprotein B mRNA editing and eliminates apolipoprotein B48.

A site-specific C to U editing reaction modifies nuclear apolipoprotein B100 (apoB100) mRNA, producing apolipoprotein B48 in the mammalian small intestine. This reaction is mediated by a multicomponent enzyme complex, which contains a catalytic subunit, Apobec-1. We have used gene targeting to disrupt mouse apobec-1 in order to establish its requisite importance in apoB mRNA editing and also, in view of its widespread tissue distribution in rodents, as a preliminary indication of other potential roles. Both heterozygous (apobec-1+/-) and homozygous (apobec-1-/-) gene-targeted mice appear healthy and fertile with no alterations in serum cholesterol or triglyceride concentrations. The apobec-1+/- mice demonstrated reduced levels of hepatic apoB mRNA editing. By contrast, levels of small intestinal apoB mRNA editing were indistinguishable in wild-type and apobec-1+/- animals, suggesting that Apobec-1 is expressed in limited quantities in the liver but not in the small intestine. The apobec-1-/- mice lacked detectable levels of Apobec-1 mRNA, expressed only unedited apoB mRNA in all tissues, and contained no apoB48 in their serum, demonstrating that there is no functional duplication of this gene.

APOBEC-1 Deaminase↗

PubMed1996

Interleukin-2 activation of STAT5 requires the convergent action of tyrosine kinases and a serine/threonine kinase pathway distinct from the Raf1/ERK2 MAP kinase pathway.

Interleukin-2 (IL-2) induces DNA binding of STAT5, a member of the family of cytokine-regulated transcription factors termed 'signal transducers and activators of transcription'. IL-2-stimulated STAT5-DNA complexes include two tyrosine phosphoproteins which exhibit distinct mobilities in SDS-PAGE gels. Our studies have shown that IL-2 rapidly induces both tyrosine phosphorylation and serine phosphorylation of STAT5 and that the two STAT5 tyrosine phosphoproteins detected in IL-2-activated cells differ in their levels of phosphorylation on serine residues. The two different phosphoforms of STAT5 have identical in vitro DNA binding specificity and reactivity with tyrosine phosphopeptides, but differ in their cellular localization. As well, the present data indicate that the transcriptional activity of STAT5 is regulated by serine kinases in T lymphocytes. Two previously characterized serine kinases activated by IL-2, MAP kinase/ERK2 and p70 S6 kinase, do not appear to be involved in STAT5 regulation by this cytokine. Accordingly, STAT5 activation in T cells requires the convergent action of tyrosine kinases and a distinct serine/threonine kinase which has not previously been implicated in IL-2 signalling.

Amino Acid Sequence↗

PubMed1996

Clustering of colorectal cancer in families of probands under 40 years of age.

UNLABELLED: Although sporadic colorectal cancer (CRC) is relatively uncommon in the young, it may constitute an elevated genetic risk for CRC in these individuals. PURPOSE: This study was designed to determine extent of colorectal cancer in families of probands under 40 years of age. METHODS: Medical records of all consecutive patients, 40 years of age or younger at the time of CRC surgery, during the time period 1986 to 1994 were examined. Cases of familial adenomatous polyposis and ulcerative colitis were excluded. Via interviews of surviving probands or nearest relatives, dates of birth and death, causes of death, and diagnosis of cancer were recorded on all first-degree relatives (parents, siblings, and offspring), second-degree relatives (grandparents, aunts, and uncles), and any other relatives. RESULTS: A total of 128 patients, 40 years of age or less at time of CRC resection, were identified. Of these, 45 probands/families were reached by phone, and 45 detailed family histories were obtained. Age range of these 45 probands was 19 to 40 (mean, 33.1) years. In 25 families there was no history of CRC in first-degree, second-degree, or third-degree relatives. Eight of 45 probands (17.8 percent) had at least one first-degree relative with CRC, and three of these eight families fulfilled the Amsterdam criteria for hereditary nonpolyposis colorectal cancer (HNPCC). In all three families, inheritance of CRC appeared to segregate with the maternal side of the family. In addition, 5 of 43 non-HNPCC probands had at least one first-degree, second-degree, or third-degree relative less than 40 years of age, at time of CRC diagnosis. CONCLUSION: Ascertainment of a detailed family history in early age of onset CRC patients identifies frequent familial clustering of CRC and HNPCC in 17.8 percent of cases.

Adult↗

PubMed1996

Residual postoperative paralysis. Pancuronium versus mivacurium, does it matter?

BACKGROUND: Based on a train-of-four (TOF) ratio greater than 0.70 as the standard of acceptable clinical recovery, undetected postoperative residual paralysis occurs frequently in postanesthesia care units. In most published studies, detailed information regarding anesthetic management is not provided. The authors reexamined the incidence of postoperative weakness after the administration of long- and short-acting neuromuscular blockers because few, if any, such comparative studies are available. METHODS: Ninety-one adult patients were studied. In group 1 (mivacurium, n = 35), anesthesia was induced with propofol/ fentanyl and maintained with nitrous oxide, desflurane, and opioid supplementation. The response of the adductor pollicis to ulnar nerve stimulation was estimated by palpating the thumb. Mivacurium (0.20 mg/kg) was administered for tracheal intubation, and an infusion was adjusted to maintain the TOF count at 1. When surgery was completed, the infusion was discontinued. When a second twitch could be detected, 7.0 micrograms/kg atropine and then 0.5 mg/kg edrophonium were administered. At 5 and 10 min, the mechanical TOF response was measured. Additional measurements were recorded if possible. Patients were tracheally extubated and discharged from the operating room when they could respond to verbal commands and no TOF fade was palpable. In group 2 (pancuronium-desflurane anesthesia, n = 29), the protocol was identical to that of group 1, except that 0.07 mg/kg pancuronium was administered for tracheal intubation. Additional increments (0.5 to 1 mg) were given as needed. Antagonism was accomplished with 0.05 mg/kg neostigmine and 0.01 mg/kg glycopyrrolate. In group 3 (pancuronium propofol-opioid, n = 27), the protocol was identical to that of group 2, except that anesthesia was maintained with nitrous oxide and a propofol-alfentanil infusion. In all groups, patients were assessed until a TOF ratio of 0.90 or more was achieved. RESULTS: All of the patients in group 1 had TOF ratios greater than 0.80 on arrival in the postanesthesia care unit. Twenty of 35 patients had TOF ratios 0.90 or more while they were still in the operating room. Thirty-three of 35 patients had TOF ratios 0.90 or more within 30 min of reversal, and this value was reached in all patients by 45 min. Recovery parameters in groups 2 and 3 did not differ from each other. Hence data from these groups were pooled. Fifty-four of 56 patients who received pancuronium had TOF values of 0.70 or more, the remaining two patients had values of 0.6 to 0.7. In contrast to the mivacurium group, however, only four patients achieved a TOF ratio of 0.90 or greater while still in the operating room. Finally, eight of these patients did not achieve this degree of recovery within 90 min of reversal. CONCLUSIONS: These results suggest that if nondepolarizing neuromuscular blockers are administered using tactile evaluation of the TOF count as a guide, critical episodes of postoperative weakness in the postanesthesia care unit should occur infrequently even with long-acting relaxants. Nevertheless, if full recovery is defined as return to a TOF ratio of 0.90 or more, then short-acting agents would appear to offer a wider margin of safety.

Adult↗

PubMed1996

Large-scale DRB and DQB1 oligonucleotide typing for the NMDP registry: progress report from year 2.

DNA typing of HLA class II alleles of the DRB1/3/4/5 and DQB1 loci using sequence-specific oligonucleotide probes and polymerase chain reaction amplified DNA has been used for the large-scale typing of donors for the National Marrow Donor Program unrelated donor registry. The results of quality control analysis for the second year of the project (10/1/939/30/94) show the typing continues to be highly accurate, specific, and reliable. The average percent of correctly classified HLA oligotypes (groups of alleles defined by a hybridization pattern with a panel of sequence-specific oligonucleotide probes) based on 9,244 DRB1 and 7,244 DQB1 assignments was 99.8% (range 99.4%100.0%) for DRB1/DRB3/DRB4/DRB5 and 99.8% (range 99.6%100.0%) for DQB1. This level of accuracy is particularly remarkable because the 4,636 DRB quality control samples were tested blindly and could not be distinguished from 57,580 donor samples tested at the same time by the laboratories.

Alleles↗

PubMed1996

Novel HLA-B alleles, B8201, B3515 and B*5106, add to the complexity of serologic identification of HLA types.

Three class I alleles, B*8201, B*3515 and B*5106, have been described using DNA and cDNA sequencing. The B*8201 allele is most structurally related to B*5602, differing from it by 14 nucleotide substitutions resulting in 5 amino acid differences. The other two alleles, B*3515 and B*5106, differ from their most closely related HLA-B alleles by 2-3 nucleotide substitutions resulting in 1-2 amino acid substitutions, respectively. The majority of nucleotide substitutions marking these new alleles are observed in other HLA-B alleles suggesting that gene conversion and/or reciprocal recombination have created this diversity. All of the amino acid substitutions are predicted to alter the antigen binding site of the HLA-B molecule. The newly defined HLA-B allelic products were originally defined by their unusual serologic reactivity patterns. The B*8201 allelic product is serologically typed as a B "blank" or as a variant of B22 or B45. These patterns and the serologic reactivity of the other newly described allelic products are consistent with the protein sequence homology among specific HLA-B molecules. While serology remains a powerful tool for detecting HLA diversity, alleles generated by events resulting in the sharing of HLA sequence polymorphisms among alleles at a locus will continue to create complexity in the interpretation of typing results.

Alleles↗

PubMed1996

Typing the HLA-B locus by a nested primer approach and oligonucleotide hybridization.

A system for intermediate level identification of the HLA-B locus alleles was devised. This system can be extended to identify individual alleles in any sample. The first step used primers which amplify all HLA-B alleles. This amplicon was subjected to SSOP hybridization to allow intermediate level typing of samples. In the second step, group-specific primers were utilized to obtain specific amplification of groups consisting of a few alleles. The oligotypes within each group were identified by the use of SSOP. The separation of groups of alleles by amplification allowed the use of a limited number of probes to identify oligotypes present in a sample. Additional probes can be added as new alleles are identified, increasing the flexibility of the system. HLA typing software was developed to determine the resolution of the system and to identify HLA oligotypes. PCR-SSOP methods are in wide use and have been extensively validated. The procedures reported here will be relatively easy to implement for large-scale DNA-based typing of the HLA-B locus.

Base Sequence↗

PubMed1996

Differences in calcium kinetics between adolescent girls and young women.

Rapid bone accretion occurs throughout childhood but peaks during adolescence. The achievement of optimal peak bone mass, which can protect against osteoporosis later in life, is greatly dependent on rates of bone accretion. To identify differences in calcium metabolism during rapid vs. slower bone accretion, calcium kinetics were compared in 14 healthy girls aged 11-14 yr and 11 women aged 19-31 yr. Calcium kinetics were measured while subjects were undergoing a calcium balance study in a camp simulating a free-living environment. After 7 days on a diet containing 1,330 mg Ca/day, two stable isotopes were administered (44Ca orally and 42Ca intravenously), and blood samples and all urine and feces were collected for 14 days. Samples were analyzed for total calcium by atomic absorption spectrophotometry and for enrichment of 42Ca and 44Ca by fast atom bombardment mass spectrometry. Data from serum, urine, and feces were analyzed using the Simulation, Analysis and Modeling (SAAM) software. Data were fitted by a three-compartment model; the first pool was the same size in girls and women (1.6 g) but the second and third pools were larger in girls (2.85 vs. 1.66 g and 12 vs. 5 g). Compared with the women, girls absorbed more calcium from the diet (38 vs. 22% or 494 vs. 283 mg/day), excreted less calcium in urine (100 vs. 203 mg/day), deposited more calcium in bone (1,459 vs. 501 mg/day), and resorbed more calcium from the skeleton (1,177 vs. 542 mg/day), whereas endogenous calcium excretion did not differ between girls and women (112 vs. 121 mg/day). Girls retained more calcium than women (282 vs. -41 mg/day) through increased absorption, lower urine excretion, and higher bone turnover.

Absorption↗

PubMed1995

Penetrating keratoplasty after radial keratotomy. A report of six patients.

BACKGROUND: For more than 15 years, radial keratotomy has increased in popularity as an option for treating myopia in the United States. During this period of time, the procedure has been modified to improve results and decrease complications. Despite these changes, complications from radial keratotomy continue to occur. The authors report six cases of penetrating keratoplasty performed to correct significant loss of vision resulting from complications of radial keratotomy. METHODS: The surgical records of one author (WR) were reviewed retrospectively for penetrating keratoplasties performed for complications of radial keratotomy. RESULTS: Six cases of penetrating keratoplasty performed for complications of radial keratotomy were found. Severe loss of vision was the indication for surgery in each case, and was associated with aggressive and repeated incisional refractive attempts to correct astigmatism, hyperopic overcorrection, residual myopia, or refractive errors associated with keratoconus. Glare associated with subepithelial scarring and irregular astigmatism were the primary findings associated with loss of vision. CONCLUSIONS: Despite advances in technique and instrumentation, radial keratotomy is limited in the amount of myopia it can correct. The risk for loss of vision increases with increasing number of incisions, intersecting incisions, very small optical zones, and keratoconus.

Adult↗

PubMed1995

Direct sequencing of SSP-PCR-amplified cDNA to identify new alleles in the DR52-associated DRB1 group: identification of DRB11115, DRB11117 and DRB1*1319.

Low and high resolution sequence specific oligonucleotide probe hybridization patterns were used to design an approach to direct sequencing of allele specific amplified cDNA. Several PCR amplifications were used to derive overlapping sequence fragments to define complete first domain sequences for a single allele. This method has been used to characterize three new DRB1 alleles in the DR52 family, DRB1*1115, DRB1*1117, and DRB1*1319. All three alleles carry polymorphisms previously observed in other DRB alleles and underscore the importance of utilizing a directed sequencing approach for obtaining unambiguous typing results in matching for bone marrow transplantation between unrelated donor and recipient.

Alleles↗

PubMed1995

Rumen involvement in sheep tannic acid metabolism.

Tannic acid (TA) metabolism was studied in sheep after p.o., intra-abomasal and i.p. administration. TA and its phenolic metabolites in ruminal fluid, abomasal fluid, plasma and urine were determined using high-performance liquid chromatography at various intervals post-dosing. After po administration, gallic acid (GA) and pyrogallol (PYR) were present in ruminal fluid and plasma, and GA, PYR and 4-O-methyl gallic acid (40MGA) were found in urine. The concentration of GA in ruminal fluid and urine gradually decreased after dosing, while PYR concentration continued to rise. By contrast, after intra-abomasal dosing, GA and ellagic acid (EA) as well as TA were found in abomasal fluid, GA, 40MGA, EA and TA were present in plasma, and GA, 40MGA and PYR were in urine. The latter 3 metabolites were also detected in the urine after ip administration of TA. The plasma concentration of TA after intra-abomasal administration was significantly correlated with liver necrosis, and PYR concentration in ruminal fluid following po administration was significantly correlated to blood methemoglobin level. It was concluded that the methemoglobinemia seen in sheep given TA po is caused by high levels of the phenolic metabolite PYR, which is produced in the rumen. Liver and kidney necrosis, seen in sheep given TA directly into the abomasum, appears caused by unmetabolized TA rather than by metabolites.

Abomasum↗

PubMed1994

Cloning and expression of cyclosporin A- and FK506-sensitive nuclear factor of activated T-cells: NF45 and NF90.

Nuclear Factor of Activated T-cells (NF-AT) is a crucial transcription factor required for T-cell expression of interleukin 2. Purified NF-AT contains 45-kDa and 90-kDa subunits (Corthésy, B., and Kao, P. N. (1994) J. Biol. Chem. 269, 20682-20690). Partial internal amino acid sequences derived from each subunit indicate that these proteins are novel. The amino acid sequences were used to clone the cDNAs encoding each subunit. The cDNAs predict proteins of novel structures: NF45 has limited similarity to prokaryotic transcription factor sigma-54 and to human DNA topoisomerase II; NF90 has limited similarity to Drosophila Staufen in a domain predicted to bind double-stranded RNA. RNA encoding NF45 and NF90 exists in nonstimulated Jurkat T-cells and in all other cell types examined (HeLa, HepG2, K562). Immunofluorescence microscopy was used to demonstrate that both proteins are located in the nucleus of Jurkat T-cells. Clones NF45 and NF90 with a polyhistidine fusion tag were transiently expressed and processed in the native environment of Jurkat T-cells. Histidine-tagged NF45 and NF90 proteins, affinity-purified on nickel chelate columns, encode a NF-AT DNA-binding activity that is enhanced following T-cell stimulation, and this enhancement is blocked when T-cells are stimulated in the presence of cyclosporin A or FK506.

Amino Acid Sequence↗

PubMed1994

No apparent involvement of the FMR1 gene in five patients with phenotypic manifestations of the fragile X syndrome.

Most fragile X patients have a significant increase in the number of CGG repeats in the FMR1 gene. Two patients were described with a deletion and one patient with a point mutation in the FMR1 gene. We describe 5 patients with a fragile X or Martin-Bell phenotype. Two brothers were discordant for the region containing the FMR1 gene; if there is a common cause for the mental retardation this is not located in the FMR1 gene. In the other 3 patients the expression of the FMR1 gene was found to be normal and no abnormalities were noted in the FMR1 mRNA. No amplification was found in the GCC repeat which is associated with the fragile site FRAXE. We conclude that the Martin-Bell phenotype can also be caused by mutations outside the FMR1 gene.

Child↗

PubMed1994

Melanoma-specific CD4+ T lymphocytes recognize human melanoma antigens processed and presented by Epstein-Barr virus-transformed B cells.

While much emphasis has been placed on the role of MHC class I-restricted CD8+ T cells in the recognition of tumor-specific antigens (Ag), evidence has accumulated that CD4+ T cells also play a critical role in the anti-tumor immune response. However, little information exists on the nature of MHC class II-restricted human tumor Ag. In an attempt to develop in vitro systems to characterize such Ag, we examined the ability of Epstein-Barr virus (EBV)-transformed B cells to present melanoma-associated Ag to melanoma-specific CD4+ cells. CD4+ T cells cultured from lymphocytes infiltrating a s.c. melanoma metastasis secreted TNF-alpha and GM-CSF specifically in response to autologous cultured melanoma cells expressing MHC class II molecules. These CD4+ cells also recognized MHC class II-compatible EBV-B cells pulsed with extracts of autologous melanoma cells, but failed to recognize EBV-B cells pulsed with autologous non-transformed cells or a variety of allogeneic tumors or normal cells. B cells pre-fixed with paraformaldehyde were incapable of Ag presentation, suggesting that intracellular processing events were occurring. Antibody-blocking studies defined HLA-DR as the dominant if not exclusive restriction locus in this T-B interaction, and HLA-DR genotyping revealed DRBI*0404 to be the probable restriction element. In a second patient, a CD4+ T-cell clone cultured from a melanoma lesion recognized autologous tumor Ag presented by autologous EBV-B; no corss-reactivity was observed with the other tumor system investigated, nor with autologous CD4+ T cells specific for tetanus toxoid. These findings demonstrate that tumor Ag can be processed and presented by EBV-transformed B cells to MHC class II-restricted tumor-specific CD4+ T cells. They also provide a model system for direct identification of these tumor-derived antigens.

Antigen Presentation↗

PubMed1994

Plasma concentrations and effect on testosterone metabolism after single doses of MK-0434, a steroid 5 alpha-reductase inhibitor, in healthy subjects.

A four-period, two-panel, single-rising-dose study (0.1-100 mg) was conducted in healthy males to investigate the pharmacodynamics, tolerability and pharmacokinetics of MK-0434, a steroid 5 alpha-reductase inhibitor. MK-0434 was associated with a significant reduction in dihydrotestosterone, which was maximal at 24 h and maintained through 48 h post treatment. The maximum reduction was approximately 50% and occurred at all doses above 5 mg (10, 25, 50 and 100 mg). MK-0434 appeared to have no effect on serum testosterone at these single doses. Rising single doses of MK-0434 were associated with an increase in Cmax and AUC but the changes were less than proportional to dose, most likely due to nonlinear absorption. MK-0434 given in single doses up to 100 mg was without significant adverse effects in healthy male volunteers. In summary, MK-0434 is a well-tolerated, potent, orally active 5 alpha-reductase inhibitor in man.

5-alpha Reductase Inhibitors↗

PubMed1994

Effect of finasteride on prostate-specific antigen density.

OBJECTIVE: Prostate-specific antigen density (PSAD) has been proposed as a diagnostic marker for prostate cancer. Because treatment with finasteride may affect PSAD differently in men with BPH compared with men with BPH plus prostate cancer, we evaluated the diagnostic utility of PSAD in men treated with finasteride for twelve months. METHODS: Data for this analysis were obtained from 895 men with BPH enrolled in a twelve-month placebo-controlled North American study. Prostate volume was measured by magnetic resonance imaging and PSA was measured by the Hybritech immunoradiometric assay. RESULTS: Treatment with finasteride for twelve months increased the positive predictive value of PSAD for identifying the presence of prostate cancer from 14 percent to 30 percent. Using PSA values alone with a cutoff of 10 ng/mL at baseline and 5 ng/mL after twelve months of treatment, similar specificity and sensitivity could be achieved. CONCLUSION: The data suggest that adjustment of PSA values during treatment with finasteride can be used to maintain the diagnostic accuracy for prostate cancer while PSAD can be used to provide additional reassurance without requiring adjustment for treatment.

Adult↗