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S Chong

Publications and source records attributed to S Chong.

At least 19 recordsLinked to original sources
PubMed2006

Detection of severe acute respiratory syndrome coronavirus in stool specimens by commercially available real-time reverse transcriptase PCR assays.

Three commercially available real-time reverse transcriptase PCR assays (the Artus RealArt HPA coronavirus LightCycler, the Artus RealArt HPA coronavirus Rotor-Gene, and the EraGen severe acute respiratory syndrome coronavirus POL assay) and three RNA extraction methodologies were evaluated for the detection of severe acute respiratory syndrome coronavirus RNA from 91 stool specimens. The assays' sensitivities were highest (58% to 75%) for specimens obtained 8 to 21 days after symptom onset. The assays were less sensitive when specimens were obtained less than 8 days or more than 21 days after the onset of symptoms. All assays were 100% specific.

Feces↗
PubMed2006

Serial magnetic resonance imaging in a rat pup filament stroke model.

Neonatal stroke is increasingly recognized in preterm and term infants but the ability to study this condition has been limited by the technical challenges in developing suitable animal models. In the current study we report the use of transient filament middle cerebral artery occlusion for 1.5 h in 10-day-old rat pups in which we were able to perform serial magnetic resonance imaging (MRI) studies. Serial MRI was performed immediately after the onset of stroke until 28 days after injury in an 11.7 T scanner using diffusion weighted and T2-weighted images. At 28 days the rat pups were sacrificed and standard histological stains were performed to validate stroke area. Serial behavioral assessments were also performed on the day of each imaging study. The anatomical distribution of stroke was similar to that expected from occlusion of the middle cerebral artery in adult models and represents a specific model of neonatal stroke in contrast to the commonly used model of carotid artery occlusion with 8% hypoxia. The initial stroke volume from MR measurements was 39% of the ipsilateral hemisphere at 0 h post-occlusion, reached a maximum at 24 h (44%) and then decreased in size (17%) with subsequent cavitation by 28 days. Infarction was more visible early with diffusion weighted imaging whereas T2-mapping provided more accurate infarct volumes at later time points. Despite the relatively large infarct volume, we saw little evidence of behavioral neurological deficit suggesting that this may also serve as a model of developmental plasticity and recovery.

Analysis of Variance↗
PubMed2006

Increased DNA methylation at the AXIN1 gene in a monozygotic twin from a pair discordant for a caudal duplication anomaly.

The AXIN1 gene has been implicated in caudal duplication anomalies. Its coding region was sequenced in both members of a monozygotic (MZ) twin pair discordant for a caudal duplication anomaly, but no mutation was found. Using bisulfite sequencing, we examined methylation at the promoter region of the AXIN1 gene in these twins and in twin and age-matched singleton controls. Methylation of the promoter region in peripheral blood mononucleated cells was variable among individuals, including MZ pairs. In the MZ pair discordant for the caudal duplication, this region of the affected twin was significantly more methylated than that of the unaffected twin (P < .0001), which was significantly more methylated than those of the controls (P = .02). We have confirmed that this CpG island does function as a promoter in vitro and that its activity is inversely proportional to the extent of methylation. This finding raises the possibility that hypermethylation of the AXIN1 promoter, by mechanisms as yet undetermined, is associated with the malformation. This case may be paradigmatic for some cases of MZ discordance.

Axin Protein↗
PubMed2006

Benign localized fibrous tumour of the pleura: CT features with histopathological correlations.

AIM: To assess the CT features of benign localized fibrous tumour of the pleura, with histopathological correlations. MATERIALS AND METHODS: CT and histopathological findings of 18 patients with surgically resected benign localized fibrous tumour of the pleura were retrospectively assessed. RESULTS: Tumours were pleura or fissure based, semilunar, lentiform or oval in shape, classified according to their homogeneous, slightly heterogeneous or heterogeneous enhancement pattern. Of the 18 tumours, 5 (28%) demonstrated > or = 55 HU increment or higher attenuation than muscles on contrast-enhanced CT, and histopathologically showed a haemangiopericytoma-like pattern with rich vascularity. CONCLUSION: CT analysis of the shape of a mass and the enhancement pattern can be helpful in the diagnosis of benign localized fibrous tumour of the pleura.

Adult↗
PubMed2006

Multicenter comparison of nucleic acid extraction methods for detection of severe acute respiratory syndrome coronavirus RNA in stool specimens.

The emergence of a novel coronavirus (CoV) as the cause of severe acute respiratory syndrome (SARS) catalyzed the development of rapid diagnostic tests. Stool samples have been shown to be appropriate for diagnostic testing for SARS CoV, although it has been recognized to be a heterogeneous and difficult sample that contains amplification inhibitors. Limited information on the efficiency of extraction methods for the purification and concentration of SARS CoV RNA from stool samples is available. Our study objectives were to determine the optimal extraction method for SARS CoV RNA detection and to examine the effect of increased specimen volume for the detection of SARS CoV RNA in stool specimens. We conducted a multicenter evaluation of four automated and four manual extraction methods using dilutions of viral lysate in replicate mock stool samples, followed by quantitation of SARS CoV RNA using real-time reverse transcriptase PCR. The sensitivities of the manual methods ranged from 50% to 100%, with the Cortex Biochem Magazorb method, a magnetic bead isolation method, allowing detection of all 12 positive samples. The sensitivities of the automated methods ranged from 75% to 100%. The bioMérieux NucliSens automated extractor and miniMag extraction methods each had a sensitivity of 100%. Examination of the copy numbers detected and the generation of 10-fold dilutions of the extracted material indicated that a number of extraction methods retained inhibitory substances that prevented optimal amplification. Increasing the volume of sample input did improve detection. This information could be useful for the extraction of other RNA viruses from stool samples and demonstrates the need to evaluate extraction methods for different specimen types.

Feces↗
PubMed2004

Studies in NZB IL-10 knockout mice of the requirement of IL-10 for progression of B-cell lymphoma.

NZB mice develop an age-related malignant expansion of a subset of B cells, B-1 cells, with autocrine production of IL-10. IL-10, a pleiotropic cytokine with anti-inflammatory properties, is a potent growth and survival factor for malignant B cells. To further examine the in vivo requirement for IL-10 in the development and expansion of malignant B-1 clones in NZB mice, we developed a strain of homozygous IL-10 knockout (KO) mice on an NZB background. The NZB IL-10 KO mice develop peritoneal B-1 cells with approximately the same frequency as heterozygous and wild-type littermates. In contrast, the development of malignant B-1 cells in the peripheral blood and spleen, observed in wild-type NZB, rarely occurred in the NZB IL-10 KO. Phenotypic analysis of surface marker expression in splenic B cells indicated that, in contrast to the NZB with malignant B-1 splenic lymphoma, the surface marker expression of NZB IL-10 KO splenic B cells indicated that the majority of the B cells were typical B-2 cells. In the absence of IL-10, spontaneously activated B cells and antiapoptotic gene expression were reduced and lymphoma incidence was decreased. These results indicate that IL-10 is a critical factor for the progression of this B-cell malignant disease.

Animals↗
PubMed2003

Phenotype and genotype correlations in two best families.

OBJECTIVE: To evaluate mutations in the Best mascular dystrophy (VMD2) gene in two families with Best disease and to describe the phenotype-genotype correlations of genetically determined affected and unaffected individuals. DESIGN: Family genetic study. PARTICIPANTS: Two families with Best disease were identified, and family members were evaluated by ophthalmologic examination or fundus photography to assess their phenotype. All affected patients and some of the unaffected family members had a blood sample drawn, and the DNA was analyzed for mutations in the VMD2 gene. MAIN OUTCOME MEASURES: Twenty-one subjects in the two pedigrees with Best disease were studied. One amino acid-changing mutation in the VMD2 gene was found to segregate independently in each family (P297S or E300D, respectively). RESULTS: Eleven individuals had some evidence of maculopathy, including retinal pigment epithelial changes, drusen, pigment epithelial irregularities, or cicatricial changes. Ten of these 11 patients (91%) with maculopathy had a mutation in the VMD2 gene, of whom 8 were clinically diagnosed as having Best disease and 2 were diagnosed as having possible Best maculopathy. The one patient without a mutation in the VMD2 gene had age-related macular degeneration (AMD). Ten family members did not have evidence of maculopathy, of whom 6 had no mutation in the VMD2 gene. Four family members (2 in each pedigree) had mutations in the VMD2 gene, abnormal electro-oculogram (EOG) results, but normal maculae at age 40 or older. Of the 7 individuals with no mutation in the VMD2 gene, 6 were phenotypically normal and the other had late-onset visual loss resulting from AMD. CONCLUSIONS: All family members with maculopathy consistent with Best disease (n = 10) had an amino acid-changing mutation in the VMD2 gene. Four individuals who did not have maculopathy, but did have an abnormal EOG, also had mutations in the VMD2 gene. The presence of a VMD2 mutation is associated with abnormal retinal function, which can occur in the absence of phenotypic manifestation of macular disease.

Adolescent↗
PubMed2003

Oxidative activation of acylguanidine prodrugs: intestinal presystemic activation in rats limits absorption and can be inhibited by co-administration of ketoconazole.

1. The disposition of acyl prodrugs was studied to improve the delivery of a guanidine-containing parent compound with poor membrane permeability and poor absorption. 2. The prodrugs were evaluated in vitro and in vivo for conversion to drug. Prodrugs were evaluated for hydrolytic or oxidative bioactivation in intestinal homogenate and rat liver S9 or microsomes. The disposition of the prodrugs in vivo was monitored in bile duct-cannulated rats. 3. Compounds with n-alkylacyl groups were efficiently bioactivated, but were hydrolysed before absorption. 4. Hydrolytic bioactivation could be blocked in vitro by branching in the alkyl chain. These compounds showed modest improvements in absorption, despite favourable permeability. Experiments with liver microsomes demonstrated efficient NADPH-dependent oxidative bioactivation, which was proposed to occur through a CYP-mediated side chain oxidation followed by cyclization and release of parent compound. Ketoconazole co-administration yielded approximately a twofold increase in absorption. 5. The hydrolytically stable prodrugs were successful in increasing absorption of parent drug and were efficiently bioactivated, but they did not yield increased systemic levels of drug.

Administration, Oral↗
PubMed2003

Application of structure-metabolism relationships in the identification of a selective endothelin A antagonist, BMS-193884, with favourable pharmacokinetic properties.

1. Based on binding affinity, 2'-amino-N-(3,4-dimethyl-5-isoxazolyl)-4'-(2-methylpropyl)[1,1'-biphenyl]-2-sulfonamide (2) was identified as an initial lead in a programme to identify selective endothelin (ET) receptor antagonists. However, the compound was extensively metabolized in preclinical animal species and human in vitro systems due to oxidative biotransformation. 2. To optimize this structural class, the site of metabolism of 2 was determined. This allowed for focussed structure-activity and structure-metabolism studies aimed at finding more metabolically stable analogues that maintained potency. New analogues were screened for their ET binding characteristics and their stability in rat and human liver microsomes. 3. The use of the microsomal stability screen was tested by the determination of the pharmacokinetic parameters of select analogues. A good correlation was found between reduced rates of rat microsomal metabolism and reduced clearance in the rat. 4. N-(3,4-dimethyl-5-isoxazolyl)-4'-(2-oxazolyl)[1,1'-biphenyl]-2-sulfonamide (3) was identified as an analogue with improved in vitro properties and further studies revealed that the compound had improved pharmacokinetic properties. 5. N-[[2'-[[(3,4-dimethyl-5-isoxazolyl)amino]sulfonyl]-4-(2-oxazolyl)[1,1'-biphenyl]-2-yl]methyl]acetamide (4) was subsequently identified as a compound with superior in vitro properties compared with compound 3, but when tested in vivo it had a substantially increased rate of clearance. Further studies demonstrated that the clearance of this closely related structural analogue was not dictated by metabolic processes, but was mediated by transport-mediated direct biliary excretion. 6. The utility of screening for in vitro liver microsomal stability as part of the lead optimization process for compounds with metabolic liabilities was shown. It was also shown that relatively small molecular changes can dramatically change the disposition of closely related analogues and care must be used when screening for a single property.

Animals↗
PubMed2003

Impact of urine collection order on the ability of assays to identify Chlamydia trachomatis infections in men.

BACKGROUND: Noninvasive urine samples have been used to diagnose Chlamydia trachomatis infections, with the assumption that the first-void urine (FVU), defined as the first 20 to 30 ml at any micturition, would be the optimal collection. We compared testing technologies on first, second, and third volumes for diagnosis. GOAL: The goal was to test in nonculture assays three sequential volumes of urine from men also undergoing urethral swabbing for C trachomatis culture specimens. STUDY DESIGN: A total of 237 men attending an STD clinic (C trachomatis prevalence, 11%) collected three containers of urine (each containing 20-30 mL) for testing in four nonculture assays. A urethral swab specimen was tested in cell culture. RESULTS: The numbers of men positive by testing of FVU with nucleic acid amplification (LCx chlamydia), nucleic acid hybridization (PACE 2), enzyme immunoassay (Chlamydiazyme), and a leukocyte esterase dipstick were 26, 7, 14, and 11, respectively; urethral culture identified 6 of the infected men. Comparative testing of all voids from the 26 men positive by the FVU assays demonstrated a reduction of LCx-positives. Non-amplified-test positivity declined precipitously in subsequent voids, approaching zero in the third void. The presence of symptoms and time of last void up to 8 hours had little effect on the number of positives detected by LCx of FVU. CONCLUSION: Amplified testing of FVU was most effective for diagnosing infection in these men.

Adult↗
PubMed2003

Specimen processing and concentration of Chlamydia trachomatis added can influence false-negative rates in the LCx assay but not in the APTIMA Combo 2 assay when testing for inhibitors.

Inhibitors in clinical specimens can be detected by adding the target of nucleic acid amplification to the sample. Introduction of a Chlamydia trachomatis L2 434 preparation containing 12 elementary bodies (EBs) into first-void urine (FVU) from 225 nonpregnant women and 190 pregnant women before specimen processing by the assays produced false-negative rates of 0.48% (2 of 415 specimens) and 13% (44 of 338 specimens) by the APTIMA Combo 2 and the Chlamydia LCx tests, respectively. Reducing the amount of C. trachomatis added to one EB, a concentration closer to the APTIMA Combo 2 test cutoff, for a subset of 244 FVU specimens increased the number of specimens with false-negative results by the APTIMA Combo 2 assay to 7 (2.9%), suggesting that the strength of the input C. trachomatis per specimen has an influence on the number of specimens with false-negative results. Repeat testing after overnight storage and dilution decreased the APTIMA Combo 2 test false-negative rates to 0% (0 of 415 specimens) with the stronger inoculum and 0.8% (2 of 244 specimens) with the weaker inoculum; the false-negative rate of the LCx assay was reduced to 5.4% (18 of 334 specimens). When an additional 70 FVU specimens from women to which 12 EBs were added before specimen processing were tested by the LCx assay, 34 specimens had false-negative results, whereas 21 specimens had false-negative results when the C. trachomatis EBs were introduced after processing. Nine of the 21 specimens to which EBs were added after processing and all of the 34 urine specimens to which the target was added before processing remained falsely negative on repeat testing at a 1:2 dilution, suggesting that input C. trachomatis DNA was lost during processing by the LCx assay. In contrast, the APTIMA Combo 2 assay appears to have a higher sensitivity and either lost little nucleic acid during processing or demonstrated few problems with inhibitors of transcription-mediated amplification.

Adult↗
PubMed2002

Laparoscopic excision of duodenal tumors.

We present three patients with bleeding duodenal tumors who were managed by laparoscopic excision. This represents the largest study of such an experience. The literature is reviewed, and the various technical approaches are discussed.

Adenoma↗
PubMed2002

The use of an oral fluid immunoglobulin G ELISA for the detection of Helicobacter pylori infection in children.

BACKGROUND: Enzyme linked immunosorbent assay (ELISA) evaluation of oral fluid immunoglobulin G (IgG) antibodies to Helicobacter pylori is a unique approach for both epidemiological studies and the diagnosis of infection, especially in children. The use of oral fluid sampling to evaluate specific H. pylori IgG antibodies has advantages over serum, including reduced biohazard risk and noninvasive collection. Oral fluid sampling is fast and involves minimal patient discomfort. Since children facilitate transmission of H. pylori infection, a simple, accurate, noninvasive diagnostic test is necessary for large epidemiologic studies. The aim of our study was to evaluate a new oral fluid ELISA for detection of IgG antibodies to H. pylori in children. MATERIALS AND METHODS: We compared this new oral fluid ELISA with the HM-CAPTM serum ELISA and gastric biopsy histology using 779 oral fluid samples from children collected at 11 clinical sites across the United States. This cohort included 315 children symptomatic for abdominal pain and 464 asymptomatic. All samples were evaluated in a double blind manner. The oral fluid ELISA demonstrated a sensitivity of 76.2% and a specificity of 94.0% in children 2 months old to 201/2 years, as compared with the HM-CAPTM serologic assay. The assay's sensitivity improved to 81.3% in children aged 5 or greater and the specificity remained at 94.0%. When compared with gastric biopsy histology in the same age group, the oral fluid ELISA demonstrated a sensitivity of 71.7% and a specificity of 90.4%. RESULTS: This new oral fluid ELISA is moderately sensitive and offers a very specific method for detecting H. pylori infection in older children, but it is of little value in children under the age of 5 years. CONCLUSIONS: Overall, we conclude that this oral fluid ELISA does not appear to be a helpful clinical tool for the diagnosis of H. pylori infection in children.

Adolescent↗
PubMed2001

Construction of a mini-intein fusion system to allow both direct monitoring of soluble protein expression and rapid purification of target proteins.

Affinity purification of recombinant proteins has been facilitated by fusion to a modified protein splicing element (intein). The fusion protein expression can be further improved by fusion to a mini-intein, i.e. an intein that lacks an endonuclease domain. We synthesized three mini-inteins using overlapping oligonucleotides to incorporate Escherichia coli optimized codons and allow convenient insertion of an affinity tag between the intein (predicted) N- and C-terminal fragments. After examining the splicing and cleavage activities of the synthesized mini-inteins, we chose the mini-intein most efficient in thiol-induced N-terminal cleavage for constructing a novel intein fusion system. In this system, green fluorescent protein (GFP) was fused to the C-terminus of the affinity-tagged mini-intein whose N-terminus was fused to a target protein. The design of the system allowed easy monitoring of soluble fusion protein expression by following GFP fluorescence, and rapid purification of the target protein through the intein-mediated cleavage reaction. A total of 17 target proteins were tested in this intein-GFP fusion system. Our data demonstrated that the fluorescence of the induced cells could be used to measure soluble expression of the intein fusion proteins and efficient intein cleavage activity. The final yield of the target proteins exhibited a linear relationship with whole cell fluorescence. The intein-GFP system may provide a simple route for monitoring real time soluble protein expression, predicting final product yields, and screening the expression of a large number of recombinant proteins for rapid purification in high throughput applications.

Base Sequence↗
PubMed2001

Intein-mediated rapid purification of Cre recombinase.

Cre recombinase produced by bacteriophage P1 catalyzes site-specific recombination of DNA between loxP recognition sites in both prokaryotic and eukaryotic cells and has been widely used for genome engineering and in vitro cloning. Recombinant Cre has been overproduced in Escherichia coli and its purification involves multiple steps. In this report, we used an "intein" fusion system to express Cre as a C-terminal fusion to a modified protein splicing element, i.e., intein. The modified intein contained a Bacillus circulans chitin-binding domain which allowed binding of the fusion protein on a chitin column and could be induced to undergo in vitro peptide bond cleavage which specifically released Cre from the column. Using the intein system, we have obtained highly pure nontagged Cre after just a single chromatographic step, which corresponded to approximately 80% recovery and 27-fold purification. The activity of the purified Cre was determined in an in vitro assay system and was found to remain stable over a period of more than 6 months.

Bacillus↗
PubMed2001

Assessment of mutations in the Best macular dystrophy (VMD2) gene in patients with adult-onset foveomacular vitelliform dystrophy, age-related maculopathy, and bull's-eye maculopathy.

PURPOSE: To study the presence of Best macular dystrophy (VMD2) gene mutations in patients diagnosed with maculopathies other than classic Best disease and to describe the clinical characteristics of these subjects. DESIGN: Case-comparison study of phenotype-genotype correlations. METHODS: Patients with either age-related maculopathy (ARM; n = 259) or maculopathies other than classic Best disease (n = 28) were screened for mutations in the Best gene (VMD2; OMIM 153700). These cases were compared with ethnically similar subjects in the same age range without maculopathy (n = 196). All patients underwent a complete dilated ocular examination, and all affected individuals underwent fundus photography. Phenotype-genotype comparisons were made. MAIN OUTCOME MEASURES: Presence of mutations in the Best gene (VMD2; OMIM 153700) and the clinical phenotype. RESULTS: Three of 259 patients (1%) with ARM and 2 of 28 patients (7%) with other maculopathies including 1 of 3 patients with adult-onset foveomacular vitelliform dystrophy and 1 of 5 patients with a bull's eye maculopathy, but none of the controls, were found to possess amino acid-changing variants in the VMD2 gene. These included a man with confluent drusen and retinal pigment epithelial detachments (variant in exon 6; T216I), a man with geographic atrophy and numerous soft drusen (variant in exon 10; L567F), a woman with drusen and retinal pigment epithelial alterations (variant in exon 10; L567F), a woman with drusen and retinal pigment epithelial alterations resembling bull's-eye maculopathy (variant in exon 4; E119Q), and a woman diagnosed with adult-onset foveomacular vitelliform dystrophy (variant in exon 4; A146K). CONCLUSIONS: Novel mutations in the VMD2 gene were found in patients diagnosed with maculopathies other than classic Best disease. Some cases diagnosed as adult-onset vitelliform foveomacular dystrophy may represent a variant of Best disease with delayed onset. The VMD2 gene does not play a major role in the development of ARM.

Aged↗