Objective: To investigate the regulatory role of LKB1 gene on expression of vascular endothelial growth factor (VEGF) in lung cancer cell line A549. Methods: DNA fragment encoding LKB1 protein was amplified by Nest-PCR from human fetal brain cDNA library and was sub-cloned into the eukaryotic expression vector pcDNA3. 1. The recombinant vector was transferred into A549 cells by Lipofectamine and screened by G418. RT-PCR and Western blot were used to study the expression of LKB1, SP1 and VEGF in A549 cells. Transcription factor SP1 was silenced by small interference RNA (siRNA); RT-PCR and Western blot were applied to examine the changes of SP1 and VEGF gene. Results: DNA sequencing analysis revealed that the open reading frame of LKB1 gene was successfully cloned into the expression vector. Stable cell line of A549 expressing exogenous LKB1 was constructed. LKB1 remarkably suppressed SP1 and VEGF expression. SiRNA targeting SP1 effectively decreased the expression of SP1 and SP1 silencing caused remarkable down-regulation of VEGF expression. Conclusion: LKB1 can negatively regulate the expression of VEGF by negatively regulate the expression of transcription factor SP1.

BACKGROUNDThe transcription factor, repressor of GATA-3 (ROG), can simultaneously suppress the expression of T helper cells (Th1 and Th2) cytokines. Since the suppression of Th2 cytokines by GATA-3 is well understood, it is postulated that there are other molecular targets of ROG that can suppress the expression of the Th1 cytokines. We hypothesized that ROG might suppress the stimulators of T lymphocyte cytokines such as CD3, CD28, and inducible costimulator (ICOS), or indirectly enhance the expression of cytokine suppressors such as T lymphocyte-associated antigen-4 (CTLA-4) and CD45. The objective of this study was to clarify the molecular targets of ROG involved in suppressing Th1 or Th2 cytokines.

METHODSReal-time quantitative PCR (RT-PCR) and Western blotting were performed to evaluate the mRNA and protein levels of CD3, CD28, ICOS, CTLA-4, and CD45 in Th1 and Th2 cells during various levels of ROG expression. Enzyme-linked immunosorbent assay (ELISA) was used to measure the levels of interferon-γ (IFN-γ) and interleukin (IL)-4 in culture media of Th1 and Th2 cells.

RESULTSThe results showed that the mRNA and protein levels of ROG were relatively low in Th1 and Th2 cells (P < 0.01). After ROG-pcDNA3.1 transfection, the mRNA and protein level of ROG was significantly elevated, while the expression of ICOS, IFN-γ, and IL-4 was markedly down-regulated (P < 0.01). Conversely, transfection of ROG-siRNA led to inhibition of ROG expression and up-regulation of ICOS, IFN-γ and IL-4 (P < 0.01). However, the expression levels of CD3, CD28, CTLA-4 and CD45 did not change in either ROG-pcDNA3.1 or ROG-siRNA-transfected Th1 and Th2 cells (P > 0.05).

CONCLUSIONIt is concluded that ROG can inhibit the expression of Th1 and Th2 cytokines by down-regulating the expression of ICOS, which might be a potential molecular target for asthma treatment.

Animals ; Blotting, Western ; CD28 Antigens ; metabolism ; CD3 Complex ; metabolism ; CD4-Positive T-Lymphocytes ; metabolism ; CTLA-4 Antigen ; metabolism ; Cells, Cultured ; Cytokines ; metabolism ; Enzyme-Linked Immunosorbent Assay ; Inducible T-Cell Co-Stimulator Protein ; metabolism ; Interferon-gamma ; metabolism ; Interleukin-4 ; metabolism ; Leukocyte Common Antigens ; metabolism ; Male ; Mice ; Mice, Inbred BALB C ; Real-Time Polymerase Chain Reaction ; Repressor Proteins ; genetics ; metabolism ; T-Lymphocytes ; metabolism ; Th1 Cells ; metabolism ; Th2 Cells ; metabolism

OBJECTIVETo study the expression of mPGES-1 in hepatocellular carcinoma (HCC), observe the effect of MK886 on down-regulation of mPGES-1 gene expression on the biology of human hepatocarcinoma cell line HepG2 and to investigate its significance in the occurrence, progression, metastasis and invasion.

METHODSHCC tissues, para-carcinoma tissues, far-carcinoma tissues and normal liver tissues were collected. The expressions of mPGES-1 were determined by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. The proliferation, adherence, migration and invasion abilities of HepG2 cells interfered by MK886 were assessed by MTT and transwell technique respectively.

RESULTSThe expression of mPGES-1 in HCCs was higher than that in normal liver tissues (P < 0.01), which increased following histological grade. Furthermore, mPGES-1 expression level was higher in the capsule invasion and metastasis tumor than in primary locus. A significant dose-dependent down-regulation of expressions of mPGES-1 gene mRNA and protein were observed in HepG2 cells when MK886 was given for 48 h (F = 140.402, P < 0.01; a'= 0.00714, P < 0.01). Compared with the control group, the growth inhibitory rate of HepG2 cell was observed significantly time and dose-dependent when MK886 was given. The rate of adhesion cells in experimental groups were 85.3% ± 1.3%, 70.5% ± 1.5% and 45.8% ± 2.4%, respectively, less than that in control group 100.0% ± 0 (F = 626.313, P < 0.01). The migration cells was 92.47 ± 1.90, 62.63 ± 1.96 and 37.33 ± 0.83 respectively in the experimental groups after 24 h, lower than that in the control group 128.93 ± 2.60 (F = 1253.805, P < 0.01). The invasion assay revealed that the invading cells were 41.67 ± 1.30, 25.47 ± 1.30 and 13.93 ± 1.66 in the experimental groups, in contrast to 55.67 ± 2.08 in control group after 24 h. The difference between these groups was significant (F = 372.615, P < 0.01). The numbers of adhesion, migration and invasion of HepG2 cells were dose-dependent in MK886 groups.

CONCLUSIONOver-expression of mPGES-1 was associated with the tumorigenesis and progression of HCC. The down-regulation of mPGES-1 gene expression might indicated the decrease of the invasion and metastasis of HCC.

Carcinoma, Hepatocellular ; metabolism ; pathology ; Cell Adhesion ; Cell Movement ; Cell Proliferation ; Female ; Hep G2 Cells ; Humans ; Indoles ; pharmacology ; Intramolecular Oxidoreductases ; metabolism ; Liver Neoplasms ; metabolism ; pathology ; Male ; Microsomes ; metabolism ; Neoplasm Invasiveness ; Neoplasm Metastasis ; Prostaglandin-E Synthases

Aged ; C-Reactive Protein ; analysis ; Coronary Angiography ; Coronary Artery Disease ; blood ; diagnostic imaging ; Cross-Sectional Studies ; Female ; Humans ; Male ; Middle Aged ; Regression Analysis ; Severity of Illness Index

BACKGROUNDNontraumatic osteonecrosis of the femoral head (NONFH) is a debilitating disease that represents a significant financial burden for both individuals and healthcare systems. Despite its significance, however, its prevalence in the Chinese general population remains unknown. This study aimed to investigate the prevalence of NONFH and its associated risk factors in the Chinese population.

METHODSA nationally representative survey of 30,030 respondents was undertaken from June 2012 to August 2013. All participants underwent a questionnaire investigation, physical examination of hip, and bilateral hip joint X-ray and/or magnetic resonance imaging examination. Blood samples were taken after overnight fasting to test serum total cholesterol, triglyceride, and high-density lipoprotein (HDL) and low-density lipoprotein (LDL) levels. We then used multivariate logistic regression analysis to investigate the associations between various metabolic, demographic, and lifestyle-related variables and NONFH.

RESULTSNONFH was diagnosed in 218 subjects (0.725%) and the estimated NONFH cases were 8.12 million among Chinese people aged 15 years and over. The prevalence of NONFH was significantly higher in males than in females (1.02% vs. 0.51%, χ2 = 24.997, P < 0.001). Among NONFH patients, North residents were subjected to higher prevalence of NONFH than that of South residents (0.85% vs. 0.61%, χ 2 = 5.847, P = 0.016). Our multivariate regression analysis showed that high blood levels of triglycerides, total cholesterol, LDL-cholesterol, and non-HDL-cholesterol, male, urban residence, family history of osteonecrosis of the femoral head, heavy smoking, alcohol abuse and glucocorticoid intake, overweight, and obesity were all significantly associated with an increased risk of NONFH.

CONCLUSIONSOur findings highlight that NONFH is a significant public health challenge in China and underscore the need for policy measures on the national level. Furthermore, NONFH shares a number of risk factors with atherosclerosis.

Adult ; Age Distribution ; Aged ; Asian Continental Ancestry Group ; China ; epidemiology ; Female ; Femur Head Necrosis ; epidemiology ; Humans ; Male ; Middle Aged ; Prevalence ; Risk Factors ; Young Adult

The purpose of our study was to assess drinking status in middle school students and to understand the associated factors. The adjusted drinking rates were 50.9%, 39.8%, and 15.1% for lifetime, past-year, and current drinking, respectively. Multivariable logistic regression revealed the following variables positively related to drinking during the past year: family income, maternal educational level, family members who drink, parents who support drinking, best friends who drink, friends who encourage drinking, frequently attending friends' parties, smoking, and actuation trend (P<0.05). The main risk factors were friends (OR=6.77, 95% CI: 6.17, 7.43) and smoking (OR=4.70, 95% CI: 3.44, 6.43). These results contribute to obtaining the potential associated factors and aid in targeted intervention into the drinking behavior of middle school students.
Adolescent ; Alcohol Drinking ; psychology ; China ; Data Collection ; Family ; Female ; Friends ; Humans ; Logistic Models ; Male ; Multivariate Analysis ; Odds Ratio ; Risk Factors ; Smoking ; Socioeconomic Factors