Child ; Codon, Nonsense ; genetics ; DNA Mutational Analysis ; DNA Primers ; genetics ; Dent Disease ; diagnosis ; genetics ; Exons ; genetics ; Humans ; Infant ; Male ; Mutation ; Oculocerebrorenal Syndrome ; diagnosis ; genetics ; Phosphoric Monoester Hydrolases ; genetics ; Polymerase Chain Reaction

Objective To investigate whether leukotriene D4 (LTD4) regulates eotaxin-3 (Eot-3) expression in bronchial epithelial cells, and study effect of pranlukst on the regulation.Methods BEAS-2B cells and normal human bronchial epithelia cells were pre- treated with LTD4 for 1 hour,stimulated with interleukin-4, the cells were incubated for 24 hours. Eot-3 protein in supernatant were measured by enzyme linked immunosorbent assay(ELISA). The cells were pretreated with pranlukast in different concentration, then the above procedure was repeated. Results The untreated bronchial epithelial cell expressed Eot-3 protein on a very low level. After stimulating with IL-4 and incubating for 24 hours, Eot-3 production increased significantly. Pretreating the cells with LTD4 enhanced the inducing effect of IL-4. Pranlukast inverted the upregulation of LTD4. Conclusions Upregulating the expression of Eot-3 induced by IL-4 on bronchial epithelial cells may explain partially the mechanism of leukotrienes involving airway allergic inflammation of asthma. The invertion impact on upregulation of LTD4 by pranlukast may be one of mechanisms that leukotrienes receptor antagonist cure asthma.

OBJECTIVETo evaluate the clinical value of perioperative adjuvant chemotherapy in prevention of tumor recurrence and improvement of patient survival after liver transplantation for advanced hepatocellular carcinoma (HCC).

METHODSTwenty patients with advanced HCC (pTNM stages III and IV a) receiving liver transplantation with preoperative transcatheter arterial chemoembolization (TACE) and postoperative adjuvant chemotherapy (ADM+5-Fu+CDDP) were retrospectively reviewed in comparison with 16 patients receiving liver transplantation only for tumor recurrence, cumulative and tumor-free survivals. The feasibility and side-effects of the treatments were also studied.

RESULTSThe recurrence rate was lower in the perioperative treatment group than in non-treatment group (12/20, 60.0% vs 11/16, 87.5%, P<0.05). The 1- and 2-year overall survival rates were 70.8% and 47.1% for the chemotherapy group and 43.8% and 20.5% for the non-chemotherapy group respectively, showing significant differences between them (P<0.05). The 1- and 2-year tumor-free survival rates were 60.6%, 40.5% and 33.6%, 15.6% in the two groups, respectively, with also significant differences (P<0.05).

CONCLUSIONSPerioperative adjuvant treatment may significantly decrease the likeliness of tumor recurrence and prolong the survival of patients with advanced HCC after liver transplantation. Chemotherapy with ADM+5-Fu+CDDP can be effective and safe with only mild side-effects.

Adult ; Carcinoma, Hepatocellular ; drug therapy ; Chemotherapy, Adjuvant ; Female ; Humans ; Liver Neoplasms ; drug therapy ; Liver Transplantation ; Male ; Middle Aged ; Neoplasm Recurrence, Local ; Perioperative Care ; Retrospective Studies ; Survival Rate ; Treatment Outcome

OBJECTIVESepsis remains a serious clinical problem because of high morbidity and mortality. The importance of Toll-like receptors (TLRs) for the induction of immune responses against sepsis was demonstrated in humans. The present study aimed to probe the gene polymorphisms of TLR4 (Asp299Gly and Thr399Ile) and TLR2 (Arg753Gln) in patients with sepsis among Chinese Han children in Wenzhou, and investigate the correlation with sepsis.

METHODThis study was conducted as a case-control study. Using polymerase chain reaction and DNA sequencing, gene polymorphisms of TLR4 (Asp299Gly and Thr399Ile) and TLR2 (Arg753Gln) in 59 children with sepsis, 38 children with severe sepsis (including 20 septic shock) and 57 healthy controls were analyzed. Hardy-Weinberg method of statistics was used to compare the frequency of genotypes alleles among three groups.

RESULTThe mutant genotypes of TLR4 gene (Asp299Gly and Thr399Ile) were not found among sepsis, septic shock and control groups. In severe sepsis group, the Arg753Gln TLR2 polymorphism occurred in 2 out of 38 severe sepsis patients and both of the subjects with the TLR2 Arg753Gln polymorphism had fatal staphylococcal infections.

CONCLUSIONTLR4 gene (Asp299Gly and Thr399Ile) polymorphisms may not be correlated with susceptibility to sepsis among Chinese Han children in Wenzhou. The fact that only 2 out of 38 severe sepsis patients had Arg753Gln TLR2 polymorphism suggests that a larger sample size is needed because of the rarity of the TLR2 allele among Chinese Han children in Wenzhou.

Asian Continental Ancestry Group ; genetics ; Case-Control Studies ; Child ; Child, Preschool ; China ; Female ; Genetic Predisposition to Disease ; Humans ; Male ; Polymorphism, Single Nucleotide ; Sepsis ; ethnology ; genetics ; Toll-Like Receptor 2 ; genetics ; Toll-Like Receptor 4 ; genetics

OBJECTIVETo study the relationship between insulin resistance and methylation of insulin receptor (INSR) gene in the endometrium of women with polycystic ovary syndrome (PCOS).

METHODSBased on the HOMA index, 35 patients with PCOS were divided into insulin resistant group (IR group, n=18) and non-resistant group (NIR group, n=18). The patients age, serum estriol, testosterone, FSH and LH, fasting insulin and fasting blood glucose were compared between the two groups. The endometrial samples were obtained from the patients to examine DNA methylation status of INSR gene in the endometrial cells using methylation-specific PCR.

RESULTSThe BMI, WHR, fasting glucose, fasting insulin, and HOMA index differed significantly between the two groups (P<0.05). PCR analysis showed partial methylation in the promoter region of INSR gene in 13 samples in IR group and 11 samples in NIR group, without detection of full methylation of the INSR gene in either group. The methylation status showed no significant difference between the two groups (P=0.328).

CONCLUSIONPartial methylation of the INSR gene occurs in the endometria of PCOS patients, but this study does not provide a strong evidence supporting the relationship between insulin resistance and INSR gene methylation in women with PCOS.

Adult ; DNA Methylation ; Endometrium ; metabolism ; Female ; Humans ; Insulin Resistance ; Polycystic Ovary Syndrome ; genetics ; metabolism ; Receptor, Insulin ; genetics ; metabolism

OBJECTIVETo investigate the expression of T-bet mRNA in peripheral blood mononuclear cells (PBMC) as well as its relations with total IgE (TIgE), eosinophil cationic protein (ECP) in serum and nasal allergic symptoms in patients with allergic rhinitis (AR).

METHODSThe allergen, TIgE and ECP in serum of patients with AR were detected by Unicap CAP system. Blood sample was taken from 8 healthy individuals and 22 patients with allergic rhinitis. PBMC was isolated by density gradient centrifugation and one part of them was cultured with 50 microg/ml mite allergen. PBMC was subjected to analysis of T-bet mRNA expression using semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR).

RESULTSThe ratio of T-bet to beta-actin mRNA levels was 0.381 +/- 0.099 in patients and 0.750 +/- 0.067 in normal individuals, the difference was significantly (P <0.01). The expression intensity of T-bet mRNA had no relation to varying severity of allergic symptoms and concentration of ECP and the correlation coefficient was 0.187 and -0.165 (all P > 0.05). However, there was an inverse correlation between expression intensity of T-bet mRNA and TIgE concentration (r = -0.525, P < 0.05). Mean mRNA level (x +/- s) of T-bet expression before and after being stimulated by allergen was 0.381 +/- 0.099 and 0.365 +/- 0.104 respectively, which indicated no significant differences (P > 0.05).

CONCLUSIONSAmong allergic patients whose allergen was mite, there was a down-regulated expression of T-bet mRNA, which had no relation to ECP concentration and allergic symptoms, but was one of important links in mechanisms of imbalance of Th1/Th2 in AR. There was no effect of specific allergen on T-bet mRNA in patients with AR T-bet was one of indirect factors that affected the level of IgE.

Adolescent ; Adult ; Case-Control Studies ; Child ; Child, Preschool ; Eosinophil Cationic Protein ; blood ; Female ; Humans ; Immunoglobulin E ; blood ; Male ; Rhinitis, Allergic, Seasonal ; blood ; T-Box Domain Proteins ; blood ; Young Adult

OBJECTIVETo study the effect of specific immunotherapy and intranasal glucocorticoid on T help 17 (Th17) cells and RORgammat in peripheral blood in patients with allergic rhinitis (AR).

METHODSForty patients with allergic rhinitis (group A) were divided randomly into two subgroups (group A1 and A2), and each subgroup had 20 patients. The patients in group A1 were treated with intranasal glucocorticoid (INGS) for one-year. The patients in group A2 were treated with special immunotherapy (SIT) for 4 weeks. Blood samples were respectively taken from 10 healthy individuals (group B), 20 AR patients (group A1) before and after SIT with specific standardized allergen and 20 AR patients (group A2) before and after INGS. The ratio of Th17 cells in peripheral blood monouclear cells (PBMC) were analysed by flow cytometry. The expression of RORgammat mRNA were detected by real-time polymerase chain reaction and the interleukin-23(IL-23), IL-17, IL-6 were detected by enzyme-linked immunosorbent assay.

RESULTSThe ratio of Th17 cells in PBMC and the expression of RORgammat mRNA in group A [(18.97 +/- 1.05)% and (0.604 +/- 0.027)] were respectively higher than those in group B [(15.12 +/- 1.09)% and (0.447 +/- 0.024)] and the difference reached statistical significance (t were respectively -10.056 and -17.986, each P < 0.01). The level of IL-6, IL-17 and IL-23 in group A were respectively higher than those in group B and the difference reached statistical significance (t were respectively -41.149, -17.618 and -26.824, all P < 0.01). The ratio of Th17 cells in PBMC, the expression of RORgammat mRNA, the level of IL-6, IL-17 and IL-23 before INGS did not show significant difference from those of after INGS in group A1 (t were respectively 0.298, 0.240, -1.136, 0.283 and -1.670, all P > 0.05). The ratio of Th17 cells in PBMC and the expression of RORgammat mRNA were respectively (18.99 +/- 1.14)% and (0.603 +/- 0.027) before SIT and were respectively (16.30 +/- 1.63)% and (0.429 +/- 0.023) after SIT in group A2, and the difference reached statistical significance (t were respectively 6.035 and 22.015, all P < 0.01). The level of IL-6, IL-17 and IL-23 before SIT were lower respectively than those of after SIT in group A2 and the difference reached statistical significance (t were respectively 9.235, 11.289, 7.267, all P < 0.01).

CONCLUSIONSThe ratio of Th17 cells in PBMC, the expression of RORgammat mRNA, the level of IL-6, IL-17 and IL-23 were up-regulated in patients with AR. The treatment of SIT could get the 5 items down and the treatment of INGS couldn't.

Allergens ; administration & dosage ; Humans ; Interleukin-17 ; Leukocytes, Mononuclear ; Nuclear Receptor Subfamily 1, Group F, Member 3 ; Rhinitis, Allergic

OBJECTIVETo study the prevalence of hepatitis B virus (HBV) genotype in 5 cities of Fujian province and the clinical implications of distinct genotypes in HBV-related liver diseases.

METHODSHBV genotype was determined by the restriction fragment length polymorphism analysis in patients with chronic HBV infection in 5 cities of Fujian province. The relationship between HBV genotype and its clinical implications was studied by multinomal logistic regression and correspondence analysis.

RESULTSOf the 431 HBV DNA positive patients detected by PCR, 275 (63.8%) belonged to HBV genotype B, 100 (23.2%) to genotype C, 51 (11.8%) to genotype D and D-mixed genotype. Genotype A, E and F were not found. Multinomal logistic regression showed that genotype B was more prevalent in Quanzhou and Sanming cities than in Fuzhou (P = 0.002, P = 0.006), and genotype B appeared significantly more common in asymptomatic carriers (ASC), chronic hepatitis B (CHB) and severe hepatitis (SH). Genotype C was most prevalent in patients with liver cirrhosis (LC) (47.0%) than in those with ASC (14.5%) and SH (14.7%) (P = 0.009, P < 0.001). The positive rate of hepatitis B e antigen was higher in patients with genotype C than in those with genotype B and genotype D (56.0% vs. 52.4%, P = 0.008, and 56.0% vs. 30.8%, P = 0.051, respectively). By correspondence analysis, genotype D and D-mixed genotype seemed to be correlated with hepatocellular carcinoma (HCC).

CONCLUSIONS(1) The major popular genotypes of HBV were B, C and D in Fujian. (2) Data of our study suggested that the geographic distribution of genotype B and C might be different in some cities of Fujian. (3) Genotype B might have a tendency to lead to SH in younger patients with chronic hepatitis B and the development of LC might be associated with genotype C among the elder patients. (4) Genotype D appeared to associate with development of HCC, which called for further study to confirm.

Adolescent ; Adult ; Aged ; Child ; Child, Preschool ; China ; Female ; Gene Frequency ; Genotype ; Hepatitis B ; virology ; Hepatitis B virus ; genetics ; Humans ; Logistic Models ; Male ; Middle Aged ; Multivariate Analysis ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length

OBJECTIVETo explore the therapeutic feasibility of allogeneic hematopoietic stem cell transplantation (allo-HSCT) from partially HLA-mismatched related donors for hematologic diseases.

METHODSThirty patients with hematologic diseases received allo-HSCT from 1 - 3 loci mismatched related donors conditioning regimen consisting of ATG (thymoglobulin, total dose of 10 mg/kg, intravenously on - 4 d to - 1 d), and only 5 (18%) of 28 recipients from HLA-identical sibling donors were treated with regimen containing ATG. Donors were given G-CSF prior to hematopoietic stem cell harvest and CsA, short-term MTX and mycophenolate mofetil (MMF) were used for GVHD prophylaxis in both group.

RESULTSAll patients were successfully engrafted. There was no significant difference in the incidence of grade II to IV acute graft-versus-host disease (aGVHD) and grade III to IV aGVHD between the mismatched and matched groups (34% vs 32%, and 13% vs 11%, respectively). 3-year overall survival (OS) and disease-free survival (DFS) in mismatched and matched groups were 57% vs 77% (P = 0.14) and 57% vs 69% (P = 0.28), respectively. Multivariate analysis showed that advanced disease pre-transplant (P = 0.006) and CMV infection (P = 0.04) were risk factors for OS. OS for patients with stable disease in mismatched and matched groups were 87% vs 81% (P = 0.65) respectively, and for those with advanced disease were 21% vs 71% (P = 0.02).

CONCLUSIONSIt is feasible to perform allo-HSCT from 1 -3 loci HLA-mismatched related donors for patients with stable disease who lack HLA-identical sibling donors. Nevertheless, for patients with advanced disease optimized conditioning regimen and intensive supporting therapy should be administered to obtain better clinical outcomes.

Graft vs Host Disease ; prevention & control ; HLA Antigens ; Hematopoietic Stem Cell Transplantation ; methods ; Humans ; Siblings ; Tissue Donors ; Transplantation Conditioning

Objective To explore the siRNA as a new antiviral therapy,evaluate the inhibition effect of siRNA based on vector on the HBV of HepG2.2.15 cell,and observe the side effect and toxicity of siRNA vector on cells and the off-target effect of siRNA.Methods Three pairs of siRNA duplexes targeting HBV C gene were designed as double strands,and the duplex were annealed and ligated into the p-Silencer-Cmv 4.1-hygro vector.The ligation products were used to transform JM109 cells.The clones with shRNA were obtained,and the vectors were purified.After the initial identification of the vector with agarose gel and the size of the inserted sequence got examined by native polyacrylamide gel electrophoresis,furthermore the sequencing was further carried out.The recombinant plasmids were purified with ultrapure Midipreps DNA Purification System.Then HepG2.2.15 cells were transfected with the plasmid mixed with siPort XP-1.The expression of HBsAg and HBeAg were detected by immunofluorescence and Western blot,and the HBV RNA was investigated by RT-PCR.Furthermore the real-time quantitive PCR was carried out to detect the changes of HBV DNA.In order to evaluate the toxicity of the shRNA,MTT was used to examine the growth rate and curve of cells.The ELISA was performed to detect the changes of interferon-? (IFN-?).Results The Western blot showed that the HBsAg and HBeAg protein were suppressed with (81.15?0.69)%,(88.12?0.92)% respectively by vector p-C2 on the third day of post-transfection.It had the similar result indicated by immunofluorescence.And the RT-PCR showed that the specific siRNA targeting HBV C gene could markedly suppress the expression of HBV mRNA and the HBV C gene mRNA was inhibited with 96.9%.The real-time quantitive PCR showed that the specific functional siRNA could markedly suppress HBV DNA copy with two orders of magnitude,while the siRNA vector had no effect on the growth of cell showed by MTT detection.Compared with the non-transfected group and p-NC group,the IFN-? level was almost the same with siRNA p-C1,p-C2,p-C3 groups.Conclusions The siRNA based on the expression vector can suppress the expression and replication of HBV in HepG2.2.15 cell.The inhibition effect was specific and had a certain dependency on siRNA concentration.No toxicity effect was found in the study.And the drug resistance wouldn′t happen because the silence was based on the split of gene.