To obtain the tervalent fusion toxin gene (named FT),three toxin gene fragments from three species of Vibrio parahaemolyticus,Vibrio vulnificus and Vibrio mimicus were connected with the flexible linker (GGGGS) using overla Pextension PCR. The three toxin gene fragments respectively encode the mature proteins of the thermostable direct hemolysin (TDH) of V. parahaemolyticus,the cytotoxin (VVC) of V. vulnificus and the heat-labile hemolysin (VMH) of V. mimicus. The identity of FT nucleic acid sequence was 99.6% with the corresponding toxin gene fragments. The open reading frame of FT was 3225 bp,encoding 1074 amino acid residues with the predicted molecular weight (MW) of 120.4 kDa. Then,FT was subcloned into the expression vector pET-22b(+). The construction of recombinant expression vector pET-22b-FT was followed by transforming into E. coli BL21(DE3) for expression. The SDS-PAGE electrophoresis results indicated that the MW of the fusion toxin protein was matched to the predicted MW. After induction by 1 mmol/L IPTG at 37℃,the fusion toxin protein was effectively expressed in E. coli BL21(DE3) with the amount of 11.49% through thin layer chromatography scanning (TLCS) analysis. Cavia cobaya was immunized using the purified cytorrhyctes to produce the anti-serum. Through the determination of the optimum working conditions,the sensitivity test,the specificity test,repeatability test and sample simulation test,the indirect ELISA method was established,which is a broad-spectrum,rapid and specific to detect various of food-poisoning Vibrio simultaneously.

OBJECTIVETo investigate the association of interleukin 6 gene (IL-6) promoter region 634C/G (rs1800796) single nucleotide polymorphism (SNP) with the genetic susceptibility to endometriosis (Ems) in south Han Chinese women.

METHODSA case-control study was performed in 432 Ems patients and 499 control women to evaluate the SNP of IL-6 634C/G by using a fluorescent quantitative PCR-based high resolution melting (HRM) method.

RESULTSThere were statistical significances in the IL-6 634C/G alleles, whether or not to carry allele G and genotype distributions between Ems patients and control women (P=0.032, 0.014 and 0.045, respectively). Allele C enhanced the risk of Ems 1.057 times while allele G reduced the risk of Ems 0.835 time. Carrying allele G reduced the risk of Ems 0.822 time, whereas not carrying allele G enhanced the risk of Ems 1.143 times. Compared with genotype CC, the risk of Ems with genotype CG reduced 0.704 time (95% CI: 0.533-0.931). There was no significant difference in whether or not carrying allele G distribution between Ems patients and control women (P=0.729).

CONCLUSIONThe present study demonstrated significant association between the SNP of IL-6 634C/G and genetic susceptibility to Ems in south Han Chinese women.

Alleles ; Case-Control Studies ; Endometriosis ; genetics ; Female ; Genetic Predisposition to Disease ; genetics ; Genotype ; Humans ; Interleukin-6 ; genetics ; Polymorphism, Single Nucleotide ; genetics

iASPP can prompt the cell proliferation and inhibit the apoptosis of many cells. There are putative binding sites of transcription factor GATA-2 upstream of iASPP transcription start site. GATA-2 plays an important role in the proliferation and differentiation of hematopoietic stem cells (HSC) and progenitors. This study was aimed to explore the role of GATA-2 protein in iASPP gene transcription. Firstly, the expression of iASPP and GATA-2 protein in some leukemia cell lines was detected by Western blot. Second, The expressive vector of pCMV5-GATA2 and the luciferase reporter vectors containing possible binding sites of GATA-2 were constructed and co-transfected into HEK293 and CV-1 cells. Then the luciferase activity was assayed by luminometer. Also, ChIP assays were performed to further confirm the specific binding of GATA-2 to iASPP promoter. The results showed that GATA-2 was overexpressed in most cell lines with high level of iASPP. GATA-2 exhibited a significant effect on luciferase activity of reporter gene iASPP and in a dose-dependant manner. The relative luciferase activity was up-regulated to about two-fold of the empty vector control when the transfection dose of pCMV5-GATA2 plasmid was increased to 100 ng. While the effect was more significant in CV-1 cells and showed a 6.7-fold increase. The ChIP assay demonstrated the in vivo specific binding of GATA-2 to iASPP. The binding sites of GATA2 were located between nt -361 ∼ -334 in upstream of iASPP gene transcription start site. It is concluded that transcription factor GATA-2 can bind with the cis-regulatory region of the iASPP promoter and up-regulate iASPP expression.
Animals ; Cell Line ; Cercopithecus aethiops ; GATA2 Transcription Factor ; genetics ; Gene Expression Regulation, Leukemic ; Humans ; Intracellular Signaling Peptides and Proteins ; genetics ; K562 Cells ; Repressor Proteins ; genetics ; Transcription, Genetic ; Transcriptional Activation ; Transfection

This study was aimed to investigate the expression of c-MPL in acute myeloid leukemia (AML) and the correlation of the c-MPL expression with CD34 and CD38, so as to define the expression of c-MPL in leukemic stem cells. The expression levels of CD34, CD38 and c-MPL were detected by flow cytometry in bone marrow cells from 29 newly diagnosed AML patients. The relationship of c-MPL positive cell ratio with clinical parameters and correlation of c-MPL with CD34 and CD38 expression in AML patients were analyzed. The results showed that expression level of c-MPL in AML patients was significantly higher than that of normal controls (P < 0.05), and the expression level of c-MPL did not correlate with age, sex, white blood cell count, AML1-ETO fusion gene and remission after chemotherapy, but the expression of c-MPL in M2 and M5 patients was higher than that of normal control (P < 0.05). Expression of c-MPL in CD34 positive AML patients was obviously higher than that in CD34 negative AML patients (P < 0.01). c-MPL was significantly higher expressed in CD34(+) cells than that in CD34(-) cells (P < 0.001), while c-MPL expression was not significantly different between CD34(+)CD38(-) and CD34(+)CD38(-) cell groups. Positive correlation between c-MPL and CD34 expression was observed (r = 0.380, P = 0.042). It is concluded that expression of c-MPL is higher in AML patients, and positively correlates with the expression level of CD34. The c-MPL expresses in leukemic stem cells.
Adolescent ; Adult ; Aged ; Case-Control Studies ; Child ; Female ; Humans ; Leukemia, Myeloid, Acute ; metabolism ; pathology ; Male ; Middle Aged ; Neoplastic Stem Cells ; metabolism ; Receptors, Thrombopoietin ; metabolism ; Young Adult

Objective To understand the worry situation about ill consequences in cancer patients and its relationship with depression.Methods The inpatients with cancer were extracted from 4 general hospitals of Wuhan and Beijing city by the cluster sampling.The self-designed questionnaires were adopted to survey the worry situation about ill consequences in cancer patients.The self-rating depression scale (SDS) was used to assess the patients' depression status.Results A total of 485 cancer patients were surveyed,including 204 males (42.1 ％),281 females(57.9 ％),and the mean age was (56.59 ± 11.19)years old.More than half of the patients usually or always worried about medical expenses.Worries about not taking care of their children,losing their lives,physical disability,leaving sequela and losing life ability also happened a lot.Patients worried more about losing their life,the possibility appearing depression was greater (P＜0.05).The younger the patients were,the greater the depression possibility was;the company staffs had lower risk of depression compared with those whose occupation was others (P＜0.05).Conclusion Cancer patients have different degrees of worries about ill consequences such as medical expenses,not taking care of their children and losing their lives and their worries about losing their lives could affect their depression status.

OBJECTIVETo investigate the association of single nucleotide polymorphisms in cytochrome P450 17 (CYP17) and estrogen receptor alpha (ERα ) genes with the risk of endometriosis among southern Chinese women.

METHODSTwo SNPs rs743572 (CYP17 gene 34T/C) and rs9322331 (ERα gene -397T/C) were genotyped by high resolution melting curve in 432 endometriosis patients and 499 matched controls.

RESULTSThere was no significant difference in the genotype frequencies of the two loci between endometriosis patients and the control subjects (P> 0.05). And there was no significant interaction effect of these two genes on the disease either.

CONCLUSIONCYP17 gene and ERα gene may not be genetic risk factors for endometriosis among southern women in China.

Asian Continental Ancestry Group ; genetics ; Endometriosis ; genetics ; Estrogen Receptor alpha ; genetics ; Female ; Genetic Predisposition to Disease ; Genotype ; Humans ; Polymorphism, Single Nucleotide ; Risk Factors ; Steroid 17-alpha-Hydroxylase ; genetics

OBJECTIVETo explore the association between the arylhydrocarbon receptor gene (AhR) 1661G/A or arylhydrocarbon nuclear translocatorgene (ARNT) 567G/C polymorphism and endometriosis in southern Han Chinese women.

METHODSThe polymorphisms of AhR gene 1661G/Aand ARNT gene 567G/C in 431 cases of endometriosis and 499 healthy women were genotyped by fluorescence quantitative PCR-based high resolution melting.

RESULTSThe frequencies of genotypes AA, AG, GG and alleles A and G in controls were 12.0%, 41.9%, 46.1%, 33.0% and 67.0%, respectively, which were not significantly different from those in patients with endometriosis (9.7%, 44.6%, 45.7%, 32.0% and 68.0%, respectively). The genotype frequencies of GG, GC, CC and alleles C and G in controls (15.6 %, 51.7%, 32.7%, 58.5%, 41.5%) were not significantly different from those in patients with endometriosis (13.5%, 47.8%, 38.7%, 62.6%, 37.4%), either. And no interaction of AhR 1661G/A and ARNT 567G/C on endometriosis was found.

CONCLUSIONNo association between AhR 1661G/A and ARNT 567G/C genetic polymorphisms and endometriosis was found in the southern Han Chinese women in this study.

Alleles ; Aryl Hydrocarbon Receptor Nuclear Translocator ; genetics ; China ; Endometriosis ; genetics ; Female ; Genetic Predisposition to Disease ; Genotype ; Humans ; Polymerase Chain Reaction ; methods ; Polymorphism, Single Nucleotide ; Receptors, Aryl Hydrocarbon ; genetics

BACKGROUND AND OBJECTIVEExpression of Skp2 was related with the prognosis of several tumors. However, there was no intensive study on the relationship between Skp2 and extranodal NK/T cell lymphoma. This study was to explore the role of Skp2 in extranodal NK/T cell lymphoma.

METHODSThe clinicopathological data of 39 patients with extranodal NK/T cell lymphoma were analyzed. The expression of Skp2 was examined by immunohistochemistry on formalin fixed, paraffin embedded tissue sections.

RESULTSAmong the patients with high expression of Skp2, complete remission (CR) rate was only 14.3% (2/14). However, CR rate among the patients with low expression of Skp2 was 68.0% (17/25). Significant difference was shown between these two groups (P < 0.001). In the group of low expression, the median overall survival (OS) was 85.59 months (95% CI: 35.83 135.34 months), the 1 and 2 year OS rates were 81% and 71%, respectively. However, in the group of high expression, the median OS was only 9.73 months (95% CI: 2.05-17.40 months), the 1 and 2 year OS rates were 42% and 14%, respectively. There was statistical difference between these two groups (P < 0.001). Multivariate analysis showed that Skp2 expression (P <0.001), LDH (P = 0.026) and ECOG PS (P = 0.003) were dependent prognostic factors of extranodal NK/T cell lymphoma.

CONCLUSIONHigh expression of Skp2 is an independent unfavorite adverse prognostic factor of extranodal NK/T cell lymphoma.

Adolescent ; Adult ; Aged ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Female ; Follow-Up Studies ; Humans ; L-Lactate Dehydrogenase ; blood ; Lymphoma, Extranodal NK-T-Cell ; drug therapy ; metabolism ; pathology ; radiotherapy ; Male ; Middle Aged ; Neoplasm Staging ; Remission Induction ; S-Phase Kinase-Associated Proteins ; metabolism ; Survival Rate ; Young Adult

Drug resistance is an important character of leukemic stem cells. To explore the mechanism of the chemotherapy resistance of N-cadherin positive leukemia cells, the quiescent state of N-cadherin positive leukemia cells was determined by flow cytometry and the relationship of G(0) phase cell ratio with the chemotherapy resistance was analyzed. After KG1a cells were induced to enter cell cycle, the G(0) phase cell ratio and the sensitivity of cells to VP16 were determined. Finally the quiescent state and drug resistance properties of KG1a cells were determined after inhibiting N-cadherin-mediated cell-cell interaction by EGTA treatment. The results showed that the G(0) phase cell ratio in N-cadherin positive KG1a cells was higher than that in N-cadherin negative KG1a cells. After KG1a cells were induced to enter cell cycle, the G(0) phase cell ratio was decreased significantly and the sensitivity of KG1a cells to VP16 increased. Following EGTA treatment for 24 hours, the G(0) phase cell ratio decreased and the drug-sensitivity was enhanced significantly. It is concluded that N-cadherin-mediated adhesion keeps N-cadherin positive leukemia cells in quiescent state of G(0) phase, thus protect these leukemia cells against VP16 chemotherapy.
Antigens, CD ; metabolism ; Apoptosis ; drug effects ; Cadherins ; metabolism ; Cell Line, Tumor ; Etoposide ; pharmacology ; therapeutic use ; Flow Cytometry ; Humans ; Leukemia, Myeloid, Acute ; drug therapy ; Resting Phase, Cell Cycle ; drug effects