Objective To detect the expression of metastasis sappressor 1(MTSS1) gene in cervical cancer tissue and to clarify its association with cervical cancer.Methods Totally 103 cases of cervical tissue were collected between Dec 2011 and Dec 2014 and classified according to biopsy and stage .Q-PCR and Western blotting were used to detect the expression of MTSS1 in normal cervical tissue and in different clinical stages of cervical cancer tissue .Results The expression of MTSS1 inⅡB-Ⅳstages of cervical cancer tissue was significantly higher than that of normal tissue or Ⅰ-ⅡA stages through q-PCR (P=0.000).Western blotting results showed that MTSS1 was positively expressed in normal cervical tissue at a rate of 23.3% or 53.3% in cervical cancer tissue.Moreover, the expression of MTSS1 was poorly correlated with age, tumor differentiation and lymphnode metastasis in cervical cancer tissue (P>0.05).The protein level of MTSS1 expressed in ⅡB-Ⅳ stages was significantly higher than that ofⅠ-ⅡA stages(P=0.005).Conclusion The expression of MTSS1 indicates the clinical stage of cervical cancer , suggesting that MTSS1 may play an important role in the development of cervical cancer .

Objective To investigate the effects of cyclopamine (CYP) on endometrial carcinoma (HEC-1A) cell survival and on induction of cell apoptosis .Methods HEC-1A cells were treated with various doses of CYP (0, 5,10, 20 and 40 μmol/L) for 24 h respectively .Then,the inverted microscope was used to observe cell morphology .Cell proliferation and apoptosis were tested by CCK-8 assay and AO/EB bi-labelling assay.The apoptosis rate of HEC-1A was analyzed using flow cytometric analysis , and the key gene expression of Bax and Bcl-2 was detected by quantitative PCR .Results The HEC-1A cells exhibited dramatic morphological changes after treatment with CYP and in a dose-dependent manner .CYP significantly inhibited HEC-1A cell proliferation using CCK8 assays(P<0.05), and induced cell death by AO/EB bi-labelling assay.Moreover,flow cytometry analysis showed that CYP treatment resulted in HEC-1A cell apoptosis, and that a higher concentration of CYP induced severer cell apoptosis (P<0.05).Meanwhile, CYP treated HEC-1A cells exhibited up-regulated expression of Bax and down-regulated expression of Bcl-2 according to Q-PCR.Conclusion Our findings indicatee that CYP can inhibit HEC-1A cell proliferation and induce cell apoptosis .

OBJECTIVE To study the molecular mechanism of cisplatin(DDP)by which HeLa cell growth and proliferation are inhibited. METHODS Cultured HeLa cells were treated with DDP 0.02-75 μmol · L-1 for 24 or 48 h. CCK-8 assay was used to determine the cell proliferation. The wound scratch assay was used to detect the cell migration and invasion. Flow cytometry was used to detect the cell cycle arresting. q-PCR was used to test the expression of metastasis suppressor gene 1 (MTSS1)mRNA. Western blot was used to determine protein levels of MTSS1,phosphorylated-extra?cellular signal-regulated kinase(p-ERK) and phosphorylated-serine-threonine kinase(p-AKT). RESULTS Following the treatment with DDP for 24 or 48 h,the proliferation of HeLa cells was inhibited significantly (P<0.05),the value of the half inhibitory concentration (IC50) of cells was 4.14 and 11.82 μmol · L-1. Migration and invasion activity of HeLa cells were reduced according to the wound scratch assay(P<0.05). Flow cytometry results showed that the cell cycle was arrested at S phase. q-PCR results showed that MTSS1 mRNA expression changed with DDP in a concentration-dependent manner (r24 h=-0.965,P<0.01;r48 h=-0.953,P<0.01). Western blot showed that the protein levels of MTSS1,p-ERK and p-AKT expression declined significantly with the increase in DDP concentrations(p-ERK:r24 h=-0.875,P<0.01;r48 h=-0.966,P<0.01. p-AKT:r24 h=-0.831,P<0.01;r48 h=-0.863,P<0.01. MTSS1:r24 h=-0.969,P<0.01;r48 h=-0.988,P<0.01). CONCLUSION DDP treatment inhibits HeLa growth and proliferation by interfering with the MTSS1 expression and disturbing the activation of ERK and AKT signaling pathways.

Objective To study the receptors for the axon guidance factor Netrin-4 and whether the known Netrin-1 receptors, DCC and UNC5H1, are coreceptors for Netrin-4. Methods The receptor for Netrin-4 was detected by affinitycytochemistry and in situ staining with the alkaline phosphatase (AP) tagged Netrin-4 fusion protein. The dynamic character of the binding between Netrin-4 and its receptor was evaluated through ligand-receptor binding assay and competitive binding assay. Results In situ staining of AP4-Netrin-4 protein bound to COS7 cells transiently transfected with eukaryotic expression coding DCC and UNC5H1 showed that the Netrin-4 could bind to the transfected cells. Competitive binding assays showed that Netrin-1 could compete with Netrin-4 for the same receptors. Conclusion DCC and UNC5H1 are coreceptors for Netrin-4. However, because the binding affinity of Netrin-4 with its receptor was lower than that of Netrin-1,it could be inferred that there should exist unique receptor(s) for Netrin-4 other than DCC and UNC5H1.

Autophagy is a self-protecting cell catabolic pathway. That macrophages involved in lipid metabolism dis-orders are the basis of atherosclerotic lesions. Autophagy plays an important role in the inhibition of inflammation and apoptosis and the promotion of cholesterol efflux. The macrophage autophagy can promote lipid metabolism, re-duce the formation of foam cells and inflammation signal, thereby inhibit atherosclerosis. Induction of macrophage autophagy may have potential significance in the treatment of atherosclerosis.

OBJECTIVETo explore apoptosis-inducing effects of realgar nanoparticle (nano-realgar) on drug-sensitive leukemia cells.

METHODPreparation of nano-realgar was mechanical milled using a high-energy planetary ball mill. Using drug-sensitive leukemia cells (K562) as target cells, MTT assay was used to detect the proliferating activity of K562 cells, and the cellular apoptosis was investigated with double staining of FITC-Annexin V and propidium iodide (PI) by flow cytometry. Flow cytometry (FCM) was employed to detect expression of intracellular Bax, Bcl-2, P-53 protein and the activity of Caspase-3.

RESULTThe raw realgar was made to ultra-fine powder by ball milling, and the average diameter of the nanoparticle was (72.72 +/- 22.18) nm measured with electron microscopes. Nano-realgar significantly inhibited the proliferation of K562 cells, Treated for 24, 48 and 72 hours, the 50% inhibitory concentration (IC50) was 43.48, 20.52, 16.07 mg x L(-1). After exposure to 20 mg x L(-1) and 50 mg x L(-1) nano-realgar for 48 hours, the apoptosis of K562 cells detected by Annexin V/PI staining was increased, the apoptotic rate of K562 cells was 10. 52% and 73.25%. After the target cells were treated with 20 mg x L(-1) and 50 mg x L(-1) nano-realgar for 48 h, the expression of P-53, Bax, Bcl-2 markedly increased in a time and dose-dependent manner. After administration of 20 mg x L(-1) and 50 mg x L(-1) nano-realgar for 48 h, the percentage of BCRP+, P-gp+ and co-expressing P-gp and BCRP cell population in K562 cells incrased dramatically.

CONCLUSIONNano-Realgar significantly induced apoptosis of drug-sensitive leukemia cells.

Apoptosis ; drug effects ; Arsenicals ; pharmacology ; Cell Proliferation ; drug effects ; Humans ; K562 Cells ; Leukemia ; drug therapy ; pathology ; Nanotechnology ; Proto-Oncogene Proteins c-bcl-2 ; analysis ; Sulfides ; pharmacology ; Tumor Suppressor Protein p53 ; analysis

Endoplasmic reticulum (ER) stress occurs in macrophage-rich areas of advanced atherosclerotic lesions and contributes to macrophage apoptosis and subsequent plaque necrosis. The purpose of the present study was to investigate the effects of caveolin-1 (Cav-1) on ER stress-induced apoptosis in cultured macrophages and the underlying mechanisms. RAW264.7 cells were incubated with thapsigargin (TG) to establish ER stress model. And Cav-1 expression was detected by Western blot. After being pretreated with filipin(III), a caveolae inhibitor, RAW264.7 cells were assayed with flow cytometry and confocal laser scanning microscopy to detect cell apoptosis. Moreover, p38 mitogen-activated protein kinase (MAPK) phosphorylation and C/EBP homologous protein (CHOP) expression were detected with Western blot. The results showed that Cav-1 expression was markedly increased at early stage of TG treatment (P < 0.05) and then decreased with prolonged or high dose TG treatments. The increasing of Cav-1 expression induced by TG in RAW264.7 cells was abolished under inhibition of caveolae by filipin(III) (P < 0.05). The effect of TG on apoptosis of RAW264.7 cells was further augmented after pretreatment with filipin(III) (P < 0.05). Western blotting showed that MAPK phosphorylation induced by TG was inhibited by filipin(III) in RAW264.7 cells (P < 0.05), whereas CHOP remained unchanged (P > 0.05). These results suggest that Cav-1 may play a critical role in suppressing ER stress-induced macrophages apoptosis in vitro, and one of the mechanisms may be correlated with the activation of p38 MAPK prosurvival pathway.
Animals ; Apoptosis ; drug effects ; Caveolin 1 ; genetics ; metabolism ; Cell Line ; Endoplasmic Reticulum Stress ; physiology ; Filipin ; pharmacology ; MAP Kinase Signaling System ; Macrophages ; cytology ; drug effects ; Mice ; Thapsigargin ; pharmacology ; Transcription Factor CHOP ; metabolism ; p38 Mitogen-Activated Protein Kinases ; metabolism

BACKGROUNDPolymorphisms of microRNA (miRNA), as a novel mechanism, are closely associated with disease states by interfering with miRNA function. Direct correlations have been identified between single-nucleotide polymorphisms (SNPs) in miRNA, but the effect on type 2 diabetes mellitus (T2DM) onset among Chinese population remains unclear. Therefore, the aim of this study was to identify correlations between common SNPs in miR-27a, miR-146a, and miR-124a with T2DM among a Chinese population, as well as to explore diabetic pathological mechanisms and the impact of environmental factors.

METHODSSNPscan technology was used to genotype 995 patients newly diagnosed with T2DM and 967 controls. Logistic regression analysis was performed to compare mutation frequencies between cases and controls.

RESULTSWe found no significant correlations between all genotypes of these miRNAs and T2DM in our research. However, stratification analysis identified a lower risk of T2DM associated with the rs531564GC genotype among younger subjects (age < 45 years) (adjusted P = 0.043; odds ratio [OR] = 0.73; 95% confidence interval [CI] = 0.54-0.99). Furthermore, the rs895819CC genotype in overweight people (24 ≤ body mass index [BMI] < 28) was significantly associated with an increased risk of T2DM (adjusted P = 0.042; OR = 1.73; 95% CI = 1.02-2.94), while the rs2910164 genotype in miR-146a was not significantly correlated with T2DM. The genetic risk score was calculated based on the number of risk alleles of the three SNPs and was found to be correlated to total cholesterol (adjusted P = 0.021).

CONCLUSIONSThe rs531564GC genotype acted as a protective factor to decrease the risk of T2DM in younger subjects (age < 45 years), while the presence of the rs895819CC genotype increased the risk of illness among overweight subjects (24 ≤ BMI < 28 kg/m 2 ). The presence of SNPs in miRNA might promote disease by affecting miRNA expression and gene function. Thus, miRNA mimics or inhibitors that directly regulate miRNA expression present novel and promising therapeutic targets.

Adult ; Aged ; Alleles ; Asian Continental Ancestry Group ; genetics ; Diabetes Mellitus, Type 2 ; genetics ; Female ; Genetic Predisposition to Disease ; genetics ; Genotype ; Humans ; Male ; MicroRNAs ; genetics ; Middle Aged ; Polymorphism, Single Nucleotide ; genetics ; Young Adult

AIM To investigate the dose-proportion relation of Tibetan medicine Siwei Jianghuang Prescription (SJP) for protective effects on diabetic nephropathy (DN),and the underlying mechanism.METHODS Diabetes mellitus rats induced by intraperitoneal injection of streptozotocin (STZ) (60 mg/kg) were randomly divided into model group,metformin support group,and eight SJP groups with dose-proportion variation (with reference to the uniform design method) for corresponding drug administration once a day,for four weeks.Measurement of fasting blood glucose (FBG) by a blood glucose meter,the concentrations of blood urea nitrogen (BUN),uric acid (UA),serum creatinine (SCr) and total protein (TP) by chemical methods,serum transforming growth factor-β1 (TGF-β1) and vascular endothelial growth factor (VEGF) by ELISA kits were conducted,the pathological morphology observation and glomerular basement membrane thickness detection by electron microscope were accomplished as well.Principal components analysis (PCA) and multivariate progressive regression analysis (MSRA) were employed to analyze the relationship between the dose-proportion to pharmacodynamics.RESULTS The resultant indexes revealed variant pharmacological improvement in each treatment group.MSRA results showed that the levels of BUN,renal index,FBG,glomerular basement membrane thickness,VEGF,Scr,and UA had correlative relations with a multiple linear or a multiple non-linear in all groups,which regression equation had a statistical significance (P ＜ 0.05);TGF-β1 level and total protein index with the dose-proportion had no linear or non-linear relation,which the regression equation statistical showed non-significance (P ＞ 0.05).In the global optimization comparison around the range of uniform design,the optimal dosage of the rats model was Curcumae Longae Rhizoma ∶ Berberidis dictyophyllae Cortex ∶ Phyllanthi Fructus ∶ Tribuli Fructus =1 ∶ 2 ∶ 1 ∶ 2.CONCLUSION Siwei Jianghuang Prescription shows better therapeutic effects on DN,which may be related to reducing the levels of BUN,renal index,FBG,glomerular basement membrane thickness,VEGF,Scr and UA.

BACKGROUNDOddi sphincter plays an important role in preventing reflux cholangitis. There exists the controversy on application of choledochoduodenostomy in hepatolithiasis management. The present study aimed at evaluating long-term outcomes of choledochoduodenostomy for the treatment of hepatolithiasis.

METHODSForty-six consecutive cases of hepatolithiasis who underwent choledochoduodenostomy were analyzed retrospectively. The pre- and postoperative rates of recurrent cholangitis and acute cholangitis severe type were compared. Paired chi-square test was applied.

RESULTSThe mean follow-up time was 17.3 years ranging from 1.6 to 40 years with a follow-up rate of 97.8% (45/46). High rates of remnant stones (39.1%, 18/46), recurrent stones (31.1%, 14/45), uncorrected strictures (85%, 17/20), and mortality (24.4%, 11/45) were observed in this group. Regurgitation of food debris and duodenal content into the biliary tract through the anastomosis was observed. The rate of recurrent cholangitis was equal to the preoperative period (93.3%, 42/45). The rate of acute cholangitis severe type after choledochoduodenostomy (46.7%, 21/45) increased significantly (P<0.01) when compared to the preoperative period (20.0%, 9/45).

CONCLUSIONSCholedochoduodenostomy did not entirely achieve the goal of clearance of stones, correction of strictures, and removing of hepatobiliary lesions by itself. Choledochoduodenostomy without cholangioplasty resulted in an increase of severe reflux cholangitis due to the loss of the anti-reflux function of the sphincter of Oddi. Therefore, choledochoduodenostomy is not an ideal approach to reduce cholangitis in hepatolithiasis and is not the best choice in the management of hepatolithiasis.

Adolescent ; Adult ; Aged ; Choledochostomy ; Female ; Humans ; Lithiasis ; surgery ; Liver Diseases ; surgery ; Male ; Middle Aged ; Retrospective Studies ; Treatment Outcome ; Young Adult