Wei Huang

Over-expression of BCAT1, a c-Myc target gene, induces cell proliferation, migration and invasion in nasopharyngeal carcinoma

BackgroundNasopharyngeal carcinoma (NPC) is a common malignant tumor in southern China and Southeast Asia, but its molecular mechanisms of pathogenesis are poorly understood. Our previous work has demonstrated that BCAT1 mRNA is over expressed in NPC and knocking down its expression in 5-8F NPC cell line can potently inhibit cell cycle progression and cell proliferation. However, the mechanism of BCAT1 up-regulation and its functional role in NPC development remain to be elucidated yet.MethodsImmunohistochemistry (IHC) method was utilized to detect the expression of BCAT1 protein in NPC at different pathological stages. The roles of gene mutation, DNA amplification and transcription factor c-Myc in regulating BCAT1 expression were analyzed using PCR-sequencing, quantitative polymerase chain reaction (qPCR), IHC, ChIP and luciferase reporter system, respectively. The functions of BCAT1 in colony formation, cell migration and invasion properties were evaluated by RNA interference (RNAi).ResultsThe positive rates of BCAT1 protein expression in normal epithelia, low-to-moderate grade atypical hyperplasia tissues, high-grade atypical hyperplasia tissues and NPC tissues were 23.6% (17/72), 75% (18/24 ), 88.9% (8/9) and 88.8% (71/80), respectively. Only one SNP site in exon1 was detected, and 42.4% (12/28) of the NPC tissues displayed the amplification of microsatellite loci in BCAT1. C-Myc could directly bind to the c-Myc binding site in promoter region of BCAT1 and up-regulate its expression. The mRNA and protein of c-Myc and BCAT1 were co-expressed in 53.6% (15/28) and 59.1% (13/22) of NPC tissues, respectively, and BCAT1 mRNA expression was also down-regulated in c-Myc knockdown cell lines. In addition, BCAT1 knockdown cells demonstrated reduced proliferation and decreased cell migration and invasion abilities.ConclusionsOur study indicates that gene amplification and c-Myc up-regulation are responsible for BCAT1 overexpression in primary NPC, and overexpression of BCAT1 induces cell proliferation, migration and invasion. The results suggest that BCAT1 may be a novel molecular target for the diagnosis and treatment of NPC.

Measurement of inflammation and oxidative stress following drastic changes in air pollution during the Beijing Olympics: a panel study approach

Ambient air pollution has been linked to cardiovascular and respiratory morbidity and mortality in epidemiology studies. Frequently, oxidative and nitrosative stress are hypothesized to mediate these pollution effects, however precise mechanisms remain unclear. This paper describes the methodology for a major panel study to examine air pollution effects on these and other mechanistic pathways. The study took place during the drastic air pollution changes accompanying the 2008 Olympics in Beijing, China. After a general description of air pollution health effects, we provide a discussion of panel studies and describe the unique features of this study that make it likely to provide compelling results. This study should lead to a clearer and more precise definition of the role of oxidative and nitrosative stress, as well as other mechanisms, in determining acute morbidity and mortality from air pollution exposure.

Increased plasma HDL cholesterol levels and biliary cholesterol excretion in hamster by LCAT overexpression.

Lecithin cholesterol acyltransferase (LCAT) is a key enzyme in the metabolism of high density lipoprotein (HDL), which has been found inversely correlated with atherosclerosis. Adenovirus mediated overexpression of human LCAT (hLCAT) in hamsters resulted in increased levels of plasma total cholesterol, HDL cholesterol, phospholipids and enlarged particle size of HDL. It also increased cholesterol and total bile acid concentrations in bile. Hepatic mRNA level of cholesterol 7α‐hydroxylase increased 2.7‐fold in hamsters. However, such effects were not observed in mice in a parallel experiment. This study suggests that overexpression of hLCAT in hamsters facilitated reverse cholesterol transport. Similar metabolic changes in humans might modify atherogenic risk.

Expression of LPL in Endothelial-Intact Artery Results in Lipid Deposition and Vascular Cell Adhesion Molecule-1 Upregulation in Both LPL and ApoE-Deficient Mice

Objective—Overexpression of lipoprotein lipase (LPL) in deendothelialized artery led to profound localized lipid deposition. In this study the role of LPL in atherogenesis in endothelial-intact carotid arteries was assessed in genetically hyperlipidemic LPL- and ApoE-deficient mice. Methods and Results—Human wild-type LPL (hLPLwt), catalytically inactive LPL (hLPL194), or control alkaline phosphatase (hAP) were expressed in endothelial-intact carotid arteries via adenoviral vectors. Compared with Ad-hAP, lipid deposition in the arterial wall increased 10.0- and 5.1-fold for Ad-hLPLwt and Ad-hLPL194 in LPL-deficient mice, and 10.6- and 6.2-fold in ApoE-deficient mice, respectively. Vascular cell adhesion molecule-1 (VCAM-1) was upregulated in Ad-hLPLwt and Ad-hLPL194 transferred arteries. Conclusions—Endothelial cell associated LPL, either active or inactive, in the arterial wall is a strong proatherosclerotic factor in both LPL- and ApoE-deficient mice.

The Cardiopulmonary Effects of Ambient Air Pollution and Mechanistic Pathways: A Comparative Hierarchical Pathway Analysis

Previous studies have investigated the associations between exposure to ambient air pollution and biomarkers of physiological pathways, yet little has been done on the comparison across biomarkers of different pathways to establish the temporal pattern of biological response. In the current study, we aim to compare the relative temporal patterns in responses of candidate pathways to different pollutants. Four biomarkers of pulmonary inflammation and oxidative stress, five biomarkers of systemic inflammation and oxidative stress, ten parameters of autonomic function, and three biomarkers of hemostasis were repeatedly measured in 125 young adults, along with daily concentrations of ambient CO, PM2.5, NO2, SO2, EC, OC, and sulfate, before, during, and after the Beijing Olympics. We used a two-stage modeling approach, including Stage I models to estimate the association between each biomarker and pollutant over each of 7 lags, and Stage II mixed-effect models to describe temporal patterns in the associations when grouping the biomarkers into the four physiological pathways. Our results show that candidate pathway groupings of biomarkers explained a significant amount of variation in the associations for each pollutant, and the temporal patterns of the biomarker-pollutant-lag associations varied across candidate pathways (p<0.0001) and were not linear (from lag 0 to lag 3: pu200a=u200a0.0629, from lag 3 to lag 6: pu200a=u200a0.0005). These findings suggest that, among this healthy young adult population, the pulmonary inflammation and oxidative stress pathway is the first to respond to ambient air pollution exposure (within 24 hours) and the hemostasis pathway responds gradually over a 2–3 day period. The initial pulmonary response may contribute to the more gradual systemic changes that likely ultimately involve the cardiovascular system.

Glucose intolerance and decreased early insulin response in mice with severe hypertriglyceridemia

Hypertriglyceridemia (HTG) is one of the key features of dyslipidemia in type 2 diabetes, caused by the overproduction and/or decreased clearance of triglyceride (TG)-rich lipoproteins, and significantly promotes the development of cardiovascular diseases in diabetes. However, the effect of severe HTG on glucose metabolism has not previously been determined. Lipoprotein lipase (LPL) deficiency results in severe HTG in humans. By using LPL-deficient mice with severe HTG, we assessed the impact of severe HTG on insulin secretion and glucose tolerance in the present study. While young LPL-deficient mice (4 months of age) showed higher fasting blood glucose (7.42 ± 0.84 versus 4.8 ± 0.80 mmol/L, P < 0.01) and lower insulin concentrations (0.16 ± 0.03 versus 0.48 ± 0.14 ng/mL, P < 0.05), old mice (12 months of age) had higher insulin (1.70 ± 0.35 versus 0.77 ± 0.04 ng/mL, P < 0.05) but normal fasting blood glucose concentrations. Both young and old mice had elevated free fatty acid (FFA) concentrations and exhibited decreased early insulin response; however, only old mice showed impaired glucose tolerance, as compared with wild-type mice of a similar age. Morphological assessment showed enlarged islets in old LPL-deficient mice. These findings suggest that different tests for glucose homeostasis may be needed for patients with LPL deficiency and severe HTG, even though their glucose concentrations are normal at initial screening.

Urinary polycyclic aromatic hydrocarbon metabolites as biomarkers of exposure to traffic-emitted pollutants

1-Nitro-pyrene has been considered a compound specific to diesel combustion emission, while 1- and 2-nitro-napthalene are mainly produced through photochemical conversion of naphthalene released to the atmosphere. Metabolites of these compounds may serve as biomarkers of exposure to traffic related pollutants. We collected urine samples from 111 healthy and non-smoking subjects within (i.e., during the Beijing Olympics) and outside (i.e., before and after the Olympics) a traffic control regime to improve Beijings air quality. Urines were analyzed for the sum of 1&2-amino-naphthalene (metabolites of 1- and 2-nitro-naphthalene) and 1-amino-pyrene (a metabolite of 1-nitro-pyrene), using an HPLC-fluorescence method. Within the same time periods, PM2.5 mass and constituents were measured, including elemental carbon, sulfate, nitrate, PAHs, carbon monoxide, nitrogen dioxide, sulfur dioxide, ozone, and particle number concentrations. The associations between the urinary metabolites and air pollutants were analyzed using linear mixed-effects models. From the pre- to during-Olympic period, 1&2-amino-naphthalene and 1-hydroxy-pyrene decreased by 23% (p=0.066) and 16% (p=0.049), respectively, while there was no change in 1-amino-pyrene (2% increase, p=0.892). From during- to post-Olympic period, 1&2-amino-naphthalene, 1-amino-pyrene and 1-hydroxy-pyrene concentrations increased by 26% (p=0.441), 37% (p=0.355), and 3% (p=0.868), respectively. Furthermore, 1&2-amino-naphthalene and 1-hydroxy-pyrene were associated with traffic related pollutants in a similar lag pattern. 1-amino-pyrene was associated more strongly with diesel combustion products (e.g. PN and elemental carbon) and not affected by season. Time-lag analyses indicate strongest/largest associations occurred 24-72h following exposure. 1&2-amino-naphthalene and 1-hydroxy-pyrene can be used as a biomarker of exposure to general vehicle-emitted pollutants. More data are needed to confirm 1-amino-pyrene as a biomarker of exposure to diesel combustion emissions. Controlling creatinine as an independent variable in the models will provide a moderate adjusting effect on the biomarker analysis.

Identification, expression and subcellular localization of ESRG

ESRG (embryonic stem cell related gene, also known as HESRG), is a novel human gene first cloned and identified by our group with microarray analysis. Interestingly, it is expressed specifically in undifferentiated human embryonic stem cells (hESCs), while its expression pattern and its role in hESCs remain unclear. Here, full-length 3151nt ESRG cDNA was further identified by RNA ligase mediated rapid amplification of cDNA ends (RLM-RACE) technique. Meanwhile, an alternatively splicing ESRG transcript (ESRG-B) of 2837nt in length was also found. Surprisingly, bioinformatics analyses showed that the open reading frames (ORFs) of ESRG and ESRG-B were identical. Both of them consist of 669nt and encode a 222aa protein with a predicted molecular size of 24 kDa. The ESRG protein was located in the nuclei of hESCs as demonstrated by immunocytochemical staining and Western blotting using ESRG specific antibody generated by us. In contrast, ESRG located in the cytoplasm of COS7 cells when it was forced to be expressed in these cells by gene transfection strategy, suggesting there may be some special proteins present only in hESCs which can help ESRG protein transport into the nuclei of hESCs. By spatial expression analysis, we further discovered that ESRG only expressed in the ovary tissue and hESCs instead of other tissues or cell lines. Our current data provide us with an important basis for conducting further studies on the functions and regulatory mechanisms underlying the role of ESRG in hESCs.

Association of air pollution sources and aldehydes with biomarkers of blood coagulation, pulmonary inflammation, and systemic oxidative stress

Using data collected before, during, and after the 2008 Summer Olympic Games in Beijing, this study examines associations between biomarkers of blood coagulation (vWF, sCD62P and sCD40L), pulmonary inflammation (EBC pH, EBC nitrite, and eNO), and systemic oxidative stress (urinary 8-OHdG) with sources of air pollution identified utilizing principal component analysis and with concentrations of three aldehydes of health concern. Associations between the biomarkers and the air pollution source types and aldehydes were examined using a linear mixed effects model, regressing through seven lag days and controlling for ambient temperature, relative humidity, gender, and day of week for the biomarker measurements. The biomarkers for pulmonary inflammation, particularly EBC pH and eNO, were most consistently associated with vehicle and industrial combustion, oil combustion, and vegetative burning. The biomarkers for blood coagulation, particularly vWF and sCD62p, were most consistently associated with oil combustion. Systemic oxidative stress biomarker (8-OHdG) was most consistently associated with vehicle and industrial combustion. The associations of the biomarkers were generally not significant or consistent with secondary formation of pollutants and with the aldehydes. The findings support policies to control anthropogenic pollution sources rather than natural soil or road dust from a cardio-respiratory health standpoint.

Reduced high-density lipoprotein 2b in non-obese type 2 diabetic patients analysed by a microfluidic chip method in a case-control study.

Background: Alterations in high-density lipoprotein (HDL) subfractions, especially in the HDL2b subfraction, have been reported in type 2 diabetes mellitus (T2DM). However, new methods for convenient and reliable quantitation of HDL2b are yet to be developed. Methods: Thirty-eight patients with T2DM were enrolled and age-, sex- and body mass index (BMI)-matched controls were selected from the same population. A microfluidic chip method was employed to analyse serum HDL subfractions. Results: The microfluidic chip method revealed a significant reduction in HDL2b and its ratio to total HDL in T2DM patients. There was a reverse correlation for total HDL and HDL2b, and its ratio with triglycerides, homeostasis model assessment-insulin sensitivity index (HOMA-IS) and insulin resistance index (HOMA-IR). Conclusions: We have shown a reduction of HDL2b and its ratio to total HDL by a novel chip method in T2DM patients. The significant correlation between HDL2b and HOMA-IS and HOMA-IR may have further predictive value in clinical utility.