Association of the POU class 2 homeobox 1 gene (POU2F1) with susceptibility to Type 2 diabetes in Chinese populations.

AIMS: POU class 2 homeobox 1 (POU2F1), also known as octamer-binding transcription factor-1 (OCT-1), is a ubiquitous transcription factor that plays a key role in the regulation of genes related to inflammation and cell cycles. POU2F1 is located on chromosome 1q24, a region with linkage for Type 2 diabetes in Chinese and other populations. We examined the association of POU2F1 genetic variants with Type 2 diabetes in Hong Kong Chinese using two independent cohorts. METHODS: We genotyped five haplotype-tagging single nucleotide polymorphisms at POU2F1 in 1378 clinic-based patients with Type 2 diabetes and 601 control subjects, as well as 707 members from 179 families with diabetes. RESULTS: We found significant associations of rs4657652, rs7532692, rs10918682 and rs3767434 (OR = 1.26-1.59, 0.0003 < P(unadjusted) < 0.035) with Type 2 diabetes in the clinic-based case-control cohorts. Rs3767434 was also associated with Type 2 diabetes (OR = 1.55, P(unadjusted) = 0.013) in the family-based cohort. Meta-analysis revealed similar associations. In addition, the risk G allele of rs10918682 showed increased usage of insulin treatment during a mean follow-up period of 7 years [hazard ratio = 1.50 (1.05-2.14), P = 0.025]. CONCLUSIONS: Using separate cohorts, we observed consistent results showing the contribution of multiple variants at POU2F1 to the risk of Type 2 diabetes.

Pharmacogenetics of esomeprazole or rabeprazole-based triple therapy in Helicobacter pylori eradication in Hong Kong non-ulcer dyspepsia Chinese subjects.

OBJECTIVE: Our study aimed to assess the effectiveness of esomeprazole or rabeprazole in combination with amoxicillin and clarithromycin for the eradication of Helicobacter pylori in Hong Kong non-ulcer dyspepsia (NUD) patients. METHODS: A prospective clinical trial was conducted at the Alice Ho Miu ling Nethersole Hospital outpatient endoscopy center from June 2004 to December 2005. Participants received amoxicillin 1 g, clarithromycin 500 mg, and, esomeprazole 20 mg (EAC) or rabeprazole 20 mg (RAC), all given twice daily for 1 week. The H. pylori status was determined by the [13C] urea breath test at least 4 weeks after completion of the treatment. Mutation status of CYP2C19 in exon 4 and exon 5 associated with the poor metabolizer phenotype was determined. RESULTS: The intention-to-treat eradication rates in patients treated with RAC and EAC were 77% and 84.6% respectively, and per protocol-based eradication rates were 83.7% and 88.9% respectively. The eradication rates did not vary with CYP2C19 phenotype found. For clarithromycin-sensitive strains, the cure rates were statistically significant regardless of CYP2C19 polymorphism (P < 0.0001). CONCLUSION: Triple therapy with either EAC or RAC is effective for Hong Kong Chinese NUD patients with H. pylori infection. Success eradication was related to clarithromycin resistance and not CYP2C19 genotype.

Associations of insulin-like growth factor binding protein-3 gene polymorphisms with IGF-I activity and lipid parameters in adolescents.

OBJECTIVE: Childhood obesity is a growing global epidemic. Recent studies indicate that obesity and related metabolic traits are highly heritable. Increasing evidence suggests that growth hormone (GH) and the insulin-like growth factor-I (IGF-I) axis have important functions in regulating adiposity and insulin sensitivity. Five single-nucleotide polymorphisms (SNPs) at IGF-binding protein-3 (IGFBP3) were genotyped to find their associations with IGF-1 activity level and common clinical metabolic traits. PATIENTS AND METHODS: We examined the associations of five SNPs at IGFBP3 with serum IGF-I and IGFBP-3 levels, as well as with obesity-related metabolic traits in 981 Hong Kong Chinese adolescents. Factor analysis was used to reduce the intercorrelated variables to five factor scores indicating body composition, blood pressure, IGF-I activity, triglyceride (TG)+high-density lipoprotein cholesterol (HDL-C) and total cholesterol (TC)+low-density lipoprotein cholesterol (LDL-C) factor scores. RESULTS: There was a strong association between the -202A/C polymorphism (rs2854744) and IGF-I activity (P=1.2 x 10(-6)) and TC+LDL-C factor scores (P=0.0085), corrected for age and sex. The C allele was associated with decreased IGFBP-3 levels (P=1.21 x 10(-13)), increased IGF-I/IGFBP-3 molar ratio (P=5.22 x 10(-6)) and decreased LDL-C (P=0.020). There was also a significant association between a G/A polymorphism at the 3' flanking sequence (rs13223993) of the IGFBP3 gene and the TG+HDL-C factor score (P=0.0013). The minor A allele carriers of rs13223993 had a lower HDL-C (P=0.0067) level and a tendency toward a high TG level. Haplotype analysis did not increase the significance of associations between single SNPs and phenotypes. CONCLUSION: Our results support the function of IGFBP3 gene polymorphisms in modulating IGF-I activity and lipid levels in adolescents. Given the prognostic significance of IGF-I, IGFBPs and lipids on risk of diabetes, obesity and cancer, long-term studies are required to clarify the clinical meaning of these findings.

Asthma and atopy are associated with chromosome 17q21 markers in Chinese children.

BACKGROUND: Single-nucleotide polymorphism (SNP)-based genome-wide association study revealed that markers on chromosome 17q21 were linked to childhood asthma but not atopy in Caucasians, with the strongest signal being detected for the SNP rs7216389 in the ORMDL3 gene. Such association was unknown in Chinese. This study delineated the allele and genotype frequencies of 10 SNPs at chromosome 17q21, and investigated the relationship between these SNPs and asthma and plasma IgE in southern Chinese children. METHODS: Asthmatic children and non-allergic controls were recruited from pediatric clinics. Their plasma total and aeroallergen-specific IgE concentrations were measured by immunoassay. Ten SNPs on 17q21 region were genotyped by multiplex SNaPshot, and their genotype associations with asthma traits analyzed using multivariate regression. RESULTS: 315 patients and 192 controls were enrolled. The allele frequency for C allele of rs7216389 varied significantly from 0.232 in our controls, 0.389 in Han Chinese to 0.536 in Caucasians. Asthma diagnosis was associated with rs11650680 and five other SNPs including rs7216389 (P = 0.019-0.034), whereas atopy was associated only with rs11650680 (P = 0.0004). Linear regression revealed the covariates for plasma total IgE to be significant for rs11650680 (P = 0.008-0.0002). Haplotypic associations were found with atopy and increased plasma total IgE, with the respective odds ratios and 95% confidence intervals for TTTCCGTT haplotype to be 0.21 and 0.09-0.52 (P = 0.0002) and 0.41 and 0.18-0.90 (P = 0.025). CONCLUSION: Childhood asthma and atopy are associated with chromosome 17q21 in Chinese, but such association may involve genes other than ORMDL3 in this region.

Transmission of multidrug-resistant tuberculosis in Shanghai: roles of residential status.

SETTING: Shanghai is a mega city where 39% of the population comprises internal migrants. OBJECTIVE: To examine the different roles played by migrants and permanent residents in the transmission of multidrug-resistant tuberculosis (MDR-TB). DESIGN: We conducted a population-based cohort study to assess MDR-TB transmission in Shanghai between 1 January 2009 and 31 December 2012 using genotyping and geospatial analyses. RESULTS: A total of 367 MDR-TB cases formed the study cohort. Significant differences between MDR-TB cases who were internal migrants and those who were permanent residents were found with regard to age, sex, region, genetic characteristics and treatment outcomes. Permanent residents had a higher transmission rate than internal migrants (OR 3.36, 95%CI 1.86-6.09). Permanent residents and genotypic clustering cases had similar clusters in central downtown and some parts of suburban areas. Most of the clusters of internal migrants were found in rural areas bordering suburban areas. Clusters of genotypic non-clustering cases showed patterns that closely matched those of internal migrants, suggesting acquired drug resistance in migrants. CONCLUSION: In Shanghai, permanent residents were significantly associated with recent transmission of MDR-TB in central downtown areas. Clustered cases of internal migrants in rural areas were most likely to have contracted MDR-TB through acquired resistance.

Downregulation of E-cadherin by hepatitis B virus X antigen in hepatocellullar carcinoma.

Hepatitis B virus (HBV)-encoded X antigen (HBxAg) contributes to the development of hepatocellular carcinoma (HCC). A frequent characteristic of HCC is reduced or absent expression of the cell adhesion protein, E-cadherin, although it is not known whether HBxAg plays a role. To address this, the levels of E-cadherin were determined in HBxAg-positive and -negative HepG2 cells in culture, and in tumor and surrounding nontumor liver from a panel of HBV carriers. The results showed an inverse relationship between HBxAg and E-cadherin expression both in tissue culture and in vivo. In HBxAg-positive cells, E-cadherin was suppressed at both the mRNA and protein levels. This was associated with hypermethylation of the E-cadherin promoter. Depressed E-cadherin correlated with HBxAg trans-activation function, as did the migration of HepG2 cells in vitro. Decreased expression of E-cadherin was also associated with the accumulation of beta-catenin in the cytoplasm and/or nuclei in tissues and cell lines, which is characteristic of activated beta-catenin. Additional work showed that HBxAg-activated beta-catenin. Together, these results suggest that the HBxAg is associated with decreased expression of E-cadherin, accumulation of beta-catenin in the cytoplasm and nucleus, and increased cell migration, which may contribute importantly to hepatocarcinogenesis.

The DCDC2 deletion is not a risk factor for dyslexia.

Dyslexia is a specific impairment in learning to read and has strong heritability. An intronic deletion within the DCDC2 gene, with ~8% frequency in European populations, is increasingly used as a marker for dyslexia in neuroimaging and behavioral studies. At a mechanistic level, this deletion has been proposed to influence sensory processing capacity, and in particular sensitivity to visual coherent motion. Our re-assessment of the literature, however, did not reveal strong support for a role of this specific deletion in dyslexia. We also analyzed data from five distinct cohorts, enriched for individuals with dyslexia, and did not identify any signal indicative of associations for the DCDC2 deletion with reading-related measures, including in a combined sample analysis (N=526). We believe we conducted the first replication analysis for a proposed deletion effect on visual motion perception and found no association (N=445 siblings). We also report that the DCDC2 deletion has a frequency of 37.6% in a cohort representative of the general population recruited in Hong Kong (N=220). This figure, together with a lack of association between the deletion and reading abilities in this cohort, indicates the low likelihood of a direct deletion effect on reading skills. Therefore, on the basis of multiple strands of evidence, we conclude that the DCDC2 deletion is not a strong risk factor for dyslexia. Our analyses and literature re-evaluation are important for interpreting current developments within multidisciplinary studies of dyslexia and, more generally, contribute to current discussions about the importance of reproducibility in science.

Immunomodulatory effects of a traditional Chinese medicine with potential antiviral activity: a self-control study.

Traditional Chinese medicine (TCM) has been used for prevention and treatment of severe acute respiratory syndrome (SARS) in Hong Kong during the outbreak in spring 2003. We investigated the immunomodulating effects of an innovative TCM regimen derived from two herbal formulas (Sang Ju Yin and Yu Ping Feng San) for treating febrile diseases. Thirty-seven healthy volunteers were given the oral TCM regimen daily for 14 days. Peripheral venous blood samples were taken on days 0, 15 and 29 for hematology, biochemistry and immunology tests, including the measurement of blood lymphocyte subsets and plasma T-helper lymphocyte types 1 and 2 cytokines and receptor. After 3 months, 23 of the volunteers participated in a control study without TCM treatment for the same time course of blood tests. Two volunteers withdrew on day 2, due to headache and dizziness. All others remained well without any side effects. No participants showed significant changes in their blood test results, except that the T-lymphocyte CD4/CD8 ratio increased significantly from 1.31 +/- 0.50 (mean +/- SD) on day 0 to 1.41 +/- 0.63 on day 15 (p < 0.02), and reduced to 1.32 +/- 0.47 on day 29 (p < 0.05). In the control study, there were no changes in the CD4/CD8 ratio. The transient increase in CD4/CD8 ratio was likely due to the TCM intake. We postulate that the administration of the innovative TCM may have beneficial immunomodulatory effects for preventing viral infections including SARS.

Role of asparaginase synthetase and asparagyl-transfer RNA synthetase in the cell-killing activity of asparaginase in Chinese hamster ovary cell mutants.

The cell-killing activity of asparaginase on three classes of Chinese hamster ovary cell mutants was examined: a mutant which overproduces asparagine synthetase (AH5); mutants defective in asparagine synthetase (N3 and N4); and mutants conditionally defective in asparagyl-transfer RNA synthetase (Asn 3, Asn 7, and Asn 9). The overproducer was more resistant to the cell-killing activity of asparaginase than wild-type Chinese hamster ovary cells, while mutants defective in asparagine synthetase were more sensitive. Surprisingly, the asparagyl-transfer RNA synthetase mutants were even more sensitive to asparaginase than the asparagine synthetase mutants. In a preliminary survey of four human lymphoid cell lines (RPMI 8402, RPMI 8392, B46M, and Molt-4F) which showed dramatically different asparaginase sensitivity, however, sensitivity to the cell-killing activity of asparaginase was correlated with reduced levels of asparagine synthetase and not with reduced levels of asparagyl-transfer RNA synthetase.

Specific interaction between 14-3-3 isoforms and the human CDC25B phosphatase.

CDC25 dual-specificity phosphatases are essential regulators that activate cyclin-dependent kinases (CDKs) at critical stages of the cell cycle. In human cells, CDC25A and C are involved in the control of G1/S and G2/M respectively, whereas CDC25B is proposed to act both in S phase and G2/M. Evidence for an interaction between CDC25 phosphatases and members of the 14-3-3 protein family has been obtained in vitro and in vivo in several organisms. On the basis of the work performed with CDC25C, it has been proposed that phosphorylation is required to mediate the interaction with 14-3-3. Here we have examined the molecular basis of the interaction between CDC25B phosphatases and 14-3-3 proteins. We show that in the two-hybrid assay all three splice variants of CDC25B interact similarly and strongly with 14-3-3eta, beta and zeta proteins, but poorly with epsilon and Theta. In vitro, CDC25B interacts at a low level with 14-3-3beta, epsilon, zeta, eta, and Theta isoforms. This interaction is not increased upon phosphorylation of CDC25B by CHK1 and is not abolished by dephosphorylation. In contrast, a specific, strong interaction between CDC25B and 14-3-3zeta and eta isoforms is revealed by a deletion of 288 residues in the amino-terminal region of CDC25B. This interaction requires the integrity of Ser 323, although it is independent of phosphorylation. Thus, interaction between 14-3-3 proteins and CDC25B is regulated in a manner that is different from that with CDC25C. We propose that, in addition to a low affinity binding site that is available for all 14-3-3 isoforms, post-translational modification of CDC25B in vivo exposes a high-affinity binding site that is specific for the zeta and eta14-3-3 isoforms.

Isolation and characterization of a human heart cDNA encoding a new member of the small heat shock protein family--HSPL27.

A novel cDNA clone was isolated from a human adult heart cDNA library. This cDNA clone is similar to the small heat shock protein (smhsp) in both DNA and amino acid sequences, especially in the conserved region. Sequence analysis has shown that the putative novel smhsp, named 27 kDa heat-shock-protein-like protein (HSPL27) is a protein of 241 amino acids with a deduced molecular mass of 26.7 kDa and a deduced pI of 8.0. We have expressed the HSPL27 in E. coli and the expressed protein was found to be present in the soluble fraction of the bacterial cell lysate. Chromosomal mapping data shows that the HSPL27 gene is located at human chromosome 5q11.2.

A novel cDNA encoding a human homologue of ribosomal protein L29.

During the large scale partial sequencing of human heart cDNA clones, a novel clone which is very similar to the rat ribosomal protein L29 in both DNA and amino acid sequences was found. The cDNA encodes a protein with a deduced molecular weight of 17751 (159 aa). It shows 80.4% homology to protein L29 from the large ribosomal subunit of rat and is related to yeast YL43. The putative protein was named human ribosomal protein L29 (hRPL29). hRPL29 has a large excess of basic residues over acidic ones. The large amount of charged residues makes the protein very hydrophilic and the protein has a deduced pI of 12.16. Internal repeats have been characterised in many ribosomal proteins and a tandem repeat of KAKAKAKA was found to be unique to hRPL29. Analysis of gene organisation by Southern blotting shows that of the approximate 10 copies of hrpL29, all but one are pseudogenes. Northern analysis indicated that the mRNA that encodes human L29 is approx. 800 base pairs in length. An intron of hrpL29 has also been cloned and sequenced by polymerase chain reaction using human genomic DNA as the template.

Genomic characterisation of the severe acute respiratory syndrome coronavirus of Amoy Gardens outbreak in Hong Kong.

Severe acute respiratory syndrome (SARS) is a global health concern. In Hong Kong, two major outbreaks, one hospital based and the other in the Amoy Gardens apartments, were identified. The frequency of diarrhoea, admission to intensive care, and mortality differed significantly between the two outbreaks. We did genomic sequencing for viral isolates from five Amoy Gardens patients. The virus sequence was identical in four of these five patients. The sequence data from one hospital case and the four identical community cases had only three nucleotide differences. Alterations in the SARS coronavirus genome are unlikely to have caused the distinctive clinical features of the Amoy Gardens patients, and these results highlight the importance of non-viral genomic factors in this outbreak.

Characterization of nucleosome core particles containing histone proteins made in bacteria.

The four core histone proteins, H2A, H2B, H3, and H4 of Xenopus laevis have been individually expressed in milligram quantities in Escherichia coli. The full-length proteins and the "trypsin-resistant" globular domains were purified under denaturing conditions and folded into histone octamers. Both intact and truncated recombinant octamers, as well as chicken erythrocyte octamer, were assembled into nucleosome core particles using a 146 bp defined-sequence DNA fragment from a 5 S RNA gene. The three types of core particles were characterized and compared by gel electrophoresis, DNase I cleavage, and tyrosine fluorescence emission during stepwise dissociation with increasing ionic strength. Nucleosome core particles containing native and mutant histones made in bacteria have facilitated its X-ray structure determination at 2.8 A resolution.

A novel method for generating a nested set of unidirectional deletion mutants using mixed oligodeoxynucleotides.

A novel method that allows introduction of unidirectional deletions into cloned DNA is described. This method is based on the use of a mixture of oligodeoxynucleotide primers that have fixed 5' ends defining the end point of the deletion and variable 3' ends composed of mixtures of all four nucleotides at six positions. The 5' ends of the oligodeoxynucleotides are hybridized to a fixed location of the M13K11RX templates and the 3' ends are hybridized randomly to the DNA to be analyzed. Such oligodeoxynucleotide primers when extended with DNA polymerase can direct deletions of intervening parts of the single-stranded DNA that by design contains multiple EcoK sites; the deletion products are selected on a host strain with the EcoK restriction system (e.g., using JM101 cells). This method is an efficient way of generating a nested set of deletion mutants useful for dideoxy-sequencing. It can be used for creating a set of deletion mutants with a particular codon at the 5' or 3' end point.

Isolation of a human vimentin cDNA with a long 3'-noncoding region from a human osteosarcoma cell line (MG-63).

Human vimentin cDNA clones were isolated and sequenced from a lambda gt11 library of the human osteosarcoma cell line MG-63. The 3'-noncoding region of the sequence is 325 nucleotides (nt) long. This cDNA sequence includes both the further 104 nt and the polyadenylation signal site at the 3' end which were earlier predicted from the genomic nt sequence, but had not been found in any previously cloned cDNAs.

Molecular cloning and characterization of FHL2, a novel LIM domain protein preferentially expressed in human heart.

A full-length cDNA clone encoding a novel LIM-only protein was isolated and sequenced from a human fetal heart cDNA library. This full-length clone consists of 1416 base pairs and has a predicted open reading frame (ORF) encoding 279 amino acids. The ORF of this polypeptide codes for the human heart-specific four and a half LIM-only protein 2 (FHL2). It possesses an extra zinc finger that is a half LIM domain and four repeats of LIM domain. When the human FHL2 cDNA probe was used to hybridize with poly-A RNA of various human tissues, a very strong signal could be seen in heart tissues, and only moderately low signals could be detected in placenta, skeletal muscle and ovary. Virtually no signal could be detected in brain, lung, liver, kidney, pancreas, spleen, thymus, prostate, testis, small intestine, colon or peripheral blood leukocyte. FHL2 was mapped to chromosome 2q12-q13 by fluorescent in-situ hybridization (FISH).