Quantitative candidate gene association studies of metabolic traits in Han Chinese type 2 diabetes patients.

Recent genome-wide association studies have identified many loci associated with type 2 diabetes mellitus (T2DM), hyperuricemia, and obesity in various ethnic populations. However, quantitative traits have been less well investigated in Han Chinese T2DM populations. We investigated the association between candidate gene single nucleotide polymorphisms (SNPs) and metabolic syndrome-related quantitative traits in Han Chinese T2DM subjects. Unrelated Han Chinese T2DM patients (1975) were recruited. Eighty-six SNPs were genotyped and tested for association with quantitative traits including lipid profiles, blood pressure, body mass index (BMI), serum uric acid (SUA), glycated hemoglobin (HbA1c), plasma glucose [fasting plasma glucose (FPG)], plasma glucose 120 min post-OGTT (P2PG; OGTT = oral glucose tolerance test), and insulin resistance-related traits. We found that CAMTA1, ABI2, VHL, KAT2B, PKHD1, ESR1, TOX, SLC30A8, SFI1, and MYH9 polymorphisms were associated with HbA1c, FPG, and/or P2PG; GCK, HHEX, TCF7L2, KCNQ1, and TBX5 polymorphisms were associated with insulin resistance-related traits; ABCG2, SLC2A9, and PKHD1 polymorphisms were associated with SUA; CAMTA1, VHL, KAT2B, PON1, NUB1, SLITRK5, SMAD3, FTO, FANCA, and PCSK2 polymorphisms were associated with blood lipid traits; CAMTA1, SPAG16, TOX, KCNQ1, ACACB, and MYH9 polymorphisms were associated with blood pressure; and UBE2E3, SPAG16, SLC2A9, CDKAL1, CDKN2A/B, TCF7L2, SMAD3, and PNPLA3 polymorphisms were associated with BMI (all P values <0.05). Some of the candidate genes were associated with metabolic and anthropometric traits in T2DM in Han Chinese. Although none of these associations reached genome-wide significance (P < 5 x 10(-8)), genes and loci identified in this study are worthy of further replication and investigation.

Effect of all-trans retinoic acids (ATRA) on the expression of α-smooth muscle actin (α-SMA) in the lung tissues of rats with pulmonary arterial hypertension (PAH).

The effect of all-trans retinoic acid (ATRA) on the expression of α-smooth muscle actin (α-SMA) in rats with pulmonary arterial hypertension (PAH) was studied, and the mechanism of the effect of ATRA on PAH was proposed. Thirty male SD rats were randomly divided into normal control, monocrotaline (MCT) model, and ATRA [30 mg/(kg.day)]intervention groups (N = 10 each). The mean pulmonary arterial pressure was recorded. Right ventricular hypertrophy index (RVHI) was calculated (weight of right ventricle: total weight of left ventricle and interventricular septum). The percentages of wall thickness of pulmonary arteriole (WT) to external diameter of artery (WT%) and vascular wall area (WA) to total vascular area (WA%) were determined. Real-time fluorescence-based quantitative PCR and western blot analyses were employed to detect the α-SMA mRNA and protein expressions. The mean pulmonary arterial pressure, RVHI, WT%, and WA% were all obviously higher in the model group than in the control and intervention groups. The values of these indicators in the intervention group were also higher than those in the control group (P < 0.01). The mRNA and protein expression levels of α-SMA were significantly higher in the lung tissue of model rats than those in the control and intervention groups. However, the intervention group showed no statistically significant differences in α-SMA mRNA and protein expression levels compared to the control (P < 0.05). ATRA inhibited the α-SMA mRNA and protein expressionin the lung tissues of rats with MCT-induced PAH, and could be used to treat PAH.

Interferon-α-2b as an adjuvant therapy prolongs survival of patients with previously resected oral muscosal melanoma.

Two major subtypes of melanoma include cutaneous melanoma and mucosal melanoma. The latter type is rare and usually occurs in the head and neck region. High-dose interferon-α-2b (IFN-α-2b) has proven effective in the treatment of cutaneous melanoma. Recently, a regimen of temozolomide plus cisplatin was reported more likely to improve relapse-free survival and overall survival than high-dose IFN-α-2b for mucosal melanoma. We conducted this study to analyze the therapeutic effect of high-dose IFN-α-2b for patients with oral mucosal melanoma who had received prior chemotherapy. One hundred and seventeen patients with stage III-IVa oral mucosal melanoma who had received chemotherapy were analyzed. The overall survival and relapse-free survival were compared between the patients with/without high-dose IFN-α-2b. The results indicate that the IFN-α-2b treatment group had a longer relapse-free survival rate (P = 0.0169) as compared to the control group. However, the overall survival was not significant between the two groups (P = 0.096), except in patients in stage IVa, whose overall survival increased by 20 months (P = 0.0146). The adverse reactions included a drug-induced influenza-like syndrome, gastrointestinal responses, myelosuppression, and hepatoxicity, which were predominantly of grade 1-2 and reversible. Thus, patients with resected oral mucosal melanoma, even those who have received chemotherapy, could benefit from the treatment of high-dose IFN-α-2b.

Cloning and expression analysis of terpenoid metabolism-related gene NsyCMS in Nicotiana sylvestris.

Terpenoids constitute the main class of secondary metabolites produced in plants with industrial, pharmacological, and agricultural interests. Nicotiana sylvestris has been widely adopted as a diploid model system in plant biology for studies of terpenoid biosynthesis. In this paper, we report the isolation and analysis of the 2-C-methyl-d-erythritol 2,4-cyclodiphosphate synthase (CMS) gene of the MEP (methylerythritol 4-phosphate) pathway from N. sylvestris. We used homologous-based cloning with a RACE method to obtain the full-length coding sequence of the NsCMS. Then, the physical and chemical properties, function, and three-dimensional structure of the NsyCMS protein were predicted. Fluorogenic quantitative PCR was used to conduct an expression analysis at different developmental stages of various tissues of the NsyCMS. The sequence of the NsyCMS consists of a 954-bp open reading frame and encodes a predicted protein of 317 amino acids, with a molecular weight of approximately 49.6 kDa and pi of 6.92. The in vivo localization of the encoded protein was cytoplasmic with no signal peptide, whereas 2 transmembrane regions were found in NsyCMS. The conserved domains of typical 2-C-methyl-d-erythritol 2,4-cyclodiphosphate synthase, aminotransferase, and pyridoxal phosphate-dependent transferase were found in NsyCMS. Differential expression patterns of the NsyCMS were observed throughout the different developmental stages and tissues. NsyCMS messenger RNA was expressed in all tissues, with the highest level of expression in the seedling leaves. NsyMK was expressed at a higher level in the resettling roots. The results from our study set the foundation for exploring the terpenoid biosynthetic pathways in N. sylvestris.

Clinical application of high-sensitivity cardiac troponin T test in acute myocardial infarction diagnosis.

The aim of this study was to investigate the clinical application of a high-sensitivity cardiac troponin T (hs-cTnT) test in the diagnosis of acute myocardial infarction (AMI). Serum levels of hs-cTnT and cardiac troponin I (cTnI) were detected in 240 AMI patients and 200 healthy donors and used to plot receiver operating characteristic (ROC) curves. A clinically applicable diagnostic cut-off value of hs-cTnT was determined from the ROC curve and the diagnostic accuracy of hs-cTnT and cTnI levels in AMI were compared.The serum hs-cTnT levels in the AMI group were higher than 0.014 ng/mL (the 99th percentile of the healthy population), among which hs-cTnT levels in patients with ST-segment elevation myocardial infarction (STEMI) were higher than in patients with non-STEMI (NSTEMI). The area under the ROC curve (AUC) for hs-cTnT was significantly higher than for cTnI, and the detection combining hs-cTnT and creatine kinase isoenzyme (CK-MB) further increased the AUC. When 0.014 ng/mL was set as the cut-off value for hs-cTnT, the diagnostic sensitivity for AMI reached 100% but the specificity was only 45.5%. The diagnostic ability of hs-cTnT for AMI peaked at a cut-off value of 0.035 ng/mL, resulting in the highest Youden index (0.654) and sensitivity and specificity values of 91.8 and 74.9%, respectively. The diagnostic utility of the hs-cTnT test for AMI is superior to the traditional cTnI method. However, since hs-cTnT levels of non-AMI patients can be over the diagnostic cut-off value, further studies are necessary to define clinically applicable cut-off values of hs-cTnT.

Advanced glycation end products (AGEs) and their receptor (RAGE) induce apoptosis of periodontal ligament fibroblasts

Diabetics have an increased prevalence of periodontitis, and diabetes is one of the causative factors of severe periodontitis. Apoptosis is thought to be involved in this pathogenic relationship. The aim of this study was to investigate apoptosis in human periodontal ligament (PDL) fibroblasts induced by advanced glycation end products (AGEs) and their receptor (RAGE). We examined the roles of apoptosis, AGEs, and RAGE during periodontitis in diabetes mellitus using cultured PDL fibroblasts that were treated by AGE-modified bovine serum albumin (AGE-BSA), bovine serum albumin (BSA) alone, or given no treatment (control). Microscopy and real-time quantitative PCR indicated that PDL fibroblasts treated with AGE-BSA were deformed and expressed higher levels of RAGE and caspase 3. Cell viability assays and flow cytometry indicated that AGE-BSA reduced cell viability (69.80±5.50%, P<0.01) and increased apoptosis (11.31±1.73%, P<0.05). Hoechst 33258 staining and terminal-deoxynucleotidyl transferase-mediated nick-end labeling revealed that AGE-BSA significantly increased apoptosis of PDL fibroblasts. The results showed that the changes in PDL fibroblasts induced by AGE-BSA may explain how AGE-RAGE participates in and exacerbates periodontium destruction.

Advanced glycation end products (AGEs) and their receptor (RAGE) induce apoptosis of periodontal ligament fibroblasts.

Diabetics have an increased prevalence of periodontitis, and diabetes is one of the causative factors of severe periodontitis. Apoptosis is thought to be involved in this pathogenic relationship. The aim of this study was to investigate apoptosis in human periodontal ligament (PDL) fibroblasts induced by advanced glycation end products (AGEs) and their receptor (RAGE). We examined the roles of apoptosis, AGEs, and RAGE during periodontitis in diabetes mellitus using cultured PDL fibroblasts that were treated by AGE-modified bovine serum albumin (AGE-BSA), bovine serum albumin (BSA) alone, or given no treatment (control). Microscopy and real-time quantitative PCR indicated that PDL fibroblasts treated with AGE-BSA were deformed and expressed higher levels of RAGE and caspase 3. Cell viability assays and flow cytometry indicated that AGE-BSA reduced cell viability (69.80 ± 5.50%, P<0.01) and increased apoptosis (11.31 ± 1.73%, P<0.05). Hoechst 33258 staining and terminal-deoxynucleotidyl transferase-mediated nick-end labeling revealed that AGE-BSA significantly increased apoptosis of PDL fibroblasts. The results showed that the changes in PDL fibroblasts induced by AGE-BSA may explain how AGE-RAGE participates in and exacerbates periodontium destruction.

Single nucleotide polymorphisms predisposing to asthma in children of Mauritian Indian and Chinese Han ethnicity.

Our objective was to investigate the distributions of six single nucleotide polymorphisms (SNPs) MS4A2 E237G, MS4A2 C-109T, ADRB2 R16G, IL4RA I75V, IL4 C-590T, and IL13 C1923T in Mauritian Indian and Chinese Han children with asthma. This case-control association study enrolled 382 unrelated Mauritian Indian children, 193 with asthma and 189 healthy controls, and 384 unrelated Chinese Han children, 192 with asthma and 192 healthy controls. The SNP loci were genotyped using polymerase chain reaction (PCR)-restriction fragment length polymorphism for the Chinese Han samples and TaqMan real-time quantitative PCR for the Mauritian Indian samples. In the Mauritian Indian children, there was a significant difference in the distribution of IL13 C1923T between the asthma and control groups (P=0.033). The frequency of IL13 C1923T T/T in the Mauritian Indian asthma group was significantly higher than in the control group [odds ratio (OR)=2.119, 95% confidence interval=1.048-4.285]. The Chinese Han children with asthma had significantly higher frequencies of MS4A2 C-109T T/T (OR=1.961, P=0.001) and ADRB2 R16G A/A (OR=2.575, P=0.000) than the control group. The IL13 C1923T locus predisposed to asthma in Mauritian Indian children, which represents an ethnic difference from the Chinese Han population. The MS4A2 C-109T T/T and ADRB2 R16G A/A genotypes were associated with asthma in the Chinese Han children.

Single nucleotide polymorphisms predisposing to asthma in children of Mauritian Indian and Chinese Han ethnicity

Our objective was to investigate the distributions of six single nucleotide polymorphisms (SNPs) MS4A2 E237G, MS4A2 C-109T, ADRB2 R16G, IL4RA I75V, IL4 C-590T, and IL13 C1923T in Mauritian Indian and Chinese Han children with asthma. This case-control association study enrolled 382 unrelated Mauritian Indian children, 193 with asthma and 189 healthy controls, and 384 unrelated Chinese Han children, 192 with asthma and 192 healthy controls. The SNP loci were genotyped using polymerase chain reaction (PCR)-restriction fragment length polymorphism for the Chinese Han samples and TaqMan real-time quantitative PCR for the Mauritian Indian samples. In the Mauritian Indian children, there was a significant difference in the distribution of IL13 C1923T between the asthma and control groups (P=0.033). The frequency of IL13 C1923T T/T in the Mauritian Indian asthma group was significantly higher than in the control group [odds ratio (OR)=2.119, 95% confidence interval=1.048-4.285]. The Chinese Han children with asthma had significantly higher frequencies of MS4A2 C-109T T/T (OR=1.961, P=0.001) and ADRB2 R16G A/A (OR=2.575, P=0.000) than the control group. The IL13 C1923T locus predisposed to asthma in Mauritian Indian children, which represents an ethnic difference from the Chinese Han population. The MS4A2 C-109T T/T and ADRB2 R16G A/A genotypes were associated with asthma in the Chinese Han children.

EZH2 suppresses hepatocellular differentiation of mouse bone marrow mesenchymal stem cells.

Our previous studies have indicated that mouse bone marrow mesenchymal stem cells (mBMMSCs) have potential to differentiate into hepatocytes with high efficiency. Our study aimed to evaluate the role of the mouse histone methyltransferase enhancer of zeste homolog 2 gene (EZH2) in the hepatocellular differentiation of mBMMSCs. The mBMMSCs isolated from femurs and tibias were cultured in Iscove's modified Eagle's medium (IMEM) supplemented with 10% fetal bovine serum. Hepatocellular differentiation was induced by 20 ng/mL hepatocyte growth factor and 10 ng/mL fibroblast growth factor 4. The mouse histone methyltransferase EZH2 gene was introduced via PLenti-eGFP-EZH2 or PLenti-eGFP-NEO as a control. Hepatocellular-induced mBMMSCs showed lower expression of EZH2 and lower level of histone H3 lysine 27 trimethylation (H3K27me3) in the AFP and FOXa2 gene promoter regions compared to uninduced mBMMSCs. Introduction of EZH2 inhibited hepatocellular induction, reduced both the mRNA and protein levels of AFP and FOXa2, and increased the level of histone H3K27me3 in the AFP and FOXa2 gene promoter regions. In summary, the mouse histone methyltransferase EZH2 gene could suppress hepatocellular differentiation of mBMMSCs by increasing the level of H3K27me3 in the AFP and FOXa2 gene promoter regions.

FGF4 and HGF promote differentiation of mouse bone marrow mesenchymal stem cells into hepatocytes via the MAPK pathway.

Our research demonstrated the potential for mouse bone marrow mesenchymal stem cells (mBMMSCs) to differentiate into hepatocytes in vitro and in vivo. However, the exact mechanism of this process remains unknown. In this study, we investigated the role of the mitogen-activated protein kinase (MAPK) cell-signaling pathway in the differentiation of mBMMSCs into hepatocytes. mBMMSCs were isolated from femurs and tibias, and hepatic differentiation was induced in Isove's modified Eagle's medium supplemented with 10% fetal bovine serum, containing human growth factor and fibroblast growth factor 4. After seven days of induction, morphological characteristics were examined. For inhibition of signaling molecular activities, the inhibitors p38 (SB203580), ERK1/2 (U0126), and MSK1 (H89) were added to the differentiation medium. Real-time polymerase chain reaction and Western blot analysis were used to evaluate the gene expression profiles and protein expression of several markers, including the early specific markers of hepatocytes (AFP and FOXa2), phosphorylated-p38 (p-p38), phosphorylated-ERK1/2 (p-ERK1/2), and phosphorylated- MSK1 (p-MSK1). Expressions of p-p38, p-ERK1/2, and p-MSK1 were effectively inhibited by their respective inhibitors. Expressions of early specific markers, AFP and FOXa2, in the p38, ERK1/2, and MSK1 inhibitor-treated groups were significantly decreased compared to those of the cytokine-induced control. Notably, the expressions of AFP and FOXa2 in the p38 inhibitor group were more obviously reduced than those in the ERK1/2 inhibitor group. The MAPK signaling pathway, especially p38, is sufficient to drive differentiation of mBMMSCs into hepatocytes. This could increase the efficiency of hepatocyte differentiation, which would benefit clinical applications.

Molecular characteristics and cloning of two pepper genes AN2 and UPA20.

The complete coding sequences (CDSs) of "Yunnan Purple Pepper No.1" (Capsicum annuum L.) AN2 and UPA20 genes were amplified using the reverse transcriptase polymerase chain reaction on the basis of the conserved sequence information of some Solanaceae plants and known highly homologous pepper expressed sequence tags. The nucleotide sequence analysis of these 2 genes revealed that pepper AN2 gene encoded a protein of 263 amino acids that has high homology with the AN2-like protein of 4 species: tobacco, tomato, potato, and petunia. The UPA20 gene encoded a protein of 341 amino acids that has high homology with the proteins of 3 species: tobacco, petunia, and tomato. The tissue expression analysis indicated that the pepper AN2 gene was overexpressed in the pericarp and placenta; moderately in stems, flowers, and seeds; and weakly in the roots, leaves, and pericarp. The pepper UPA20 gene was overexpressed in the flowers and seeds; moderately expressed in the roots and stems; and weakly expressed in the leaves and placenta. Our findings might form the basis for further research on these 2 pepper genes.

ReSeqTools: an integrated toolkit for large-scale next-generation sequencing based resequencing analysis.

Large-scale next-generation sequencing (NGS)-based resequencing detects sequence variations, constructs evolutionary histories, and identifies phenotype-related genotypes. However, NGS-based resequencing studies generate extraordinarily large amounts of data, making computations difficult. Effective use and analysis of these data for NGS-based resequencing studies remains a difficult task for individual researchers. Here, we introduce ReSeqTools, a full-featured toolkit for NGS (Illumina sequencing)-based resequencing analysis, which processes raw data, interprets mapping results, and identifies and annotates sequence variations. ReSeqTools provides abundant scalable functions for routine resequencing analysis in different modules to facilitate customization of the analysis pipeline. ReSeqTools is designed to use compressed data files as input or output to save storage space and facilitates faster and more computationally efficient large-scale resequencing studies in a user-friendly manner. It offers abundant practical functions and generates useful statistics during the analysis pipeline, which significantly simplifies resequencing analysis. Its integrated algorithms and abundant sub-functions provide a solid foundation for special demands in resequencing projects. Users can combine these functions to construct their own pipelines for other purposes.

Development of fetal nicotine and muscarinic receptors in utero

The role of acetylcholine in the central and peripheral nervous systems is well established in adults. Cholinergic modulation of vascular functions and body fluid balance has been extensively studied. In the embryo-fetus, cholinergic receptors are widespread in the peripheral and central systems, including smooth muscle and the epithelial lining of the cardiovascular, digestive, and urinary systems, as well as in the brain. Fetal nicotine and muscarinic receptors develop in a pattern (e.g., amount and distribution) related to gestational periods. Cholinergic mechanisms have been found to be relatively intact and functional in the control of vascular homeostasis during fetal life in utero at least during the last third of gestation. This review focuses on the development of fetal nicotine and muscarinic receptors, and provides information indicating that central cholinergic systems are well developed in the control of fetal blood pressure and body fluid balance before birth. Therefore, the development of cholinergic systems in utero plays an important role in fetal vascular regulation, gastrointestinal motility, and urinary control.

Development of fetal nicotine and muscarinic receptors in utero.

The role of acetylcholine in the central and peripheral nervous systems is well established in adults. Cholinergic modulation of vascular functions and body fluid balance has been extensively studied. In the embryo-fetus, cholinergic receptors are widespread in the peripheral and central systems, including smooth muscle and the epithelial lining of the cardiovascular, digestive, and urinary systems, as well as in the brain. Fetal nicotine and muscarinic receptors develop in a pattern (e.g., amount and distribution) related to gestational periods. Cholinergic mechanisms have been found to be relatively intact and functional in the control of vascular homeostasis during fetal life in utero at least during the last third of gestation. This review focuses on the development of fetal nicotine and muscarinic receptors, and provides information indicating that central cholinergic systems are well developed in the control of fetal blood pressure and body fluid balance before birth. Therefore, the development of cholinergic systems in utero plays an important role in fetal vascular regulation, gastrointestinal motility, and urinary control.