OBJECTIVETo evaluate the reliability, validity, and responsiveness of traditional Chinese medicine (TCM) clinical outcomes rating scale for heart failure (HF) based on patients' report.

METHODSTCM clinical outcomes rating scale for HF (TCM-HF-PRO) were evaluated based on 340 HF patients' report from multiple centers. The completion of the investigation was recorded. Cronbach's α coefficient and split-half reliability were used for reliability analysis, and factor analysis was used to assess the construct validity of the rating scale. Pearson correlation analysis was then used for criterion validity analysis. Discriminant analysis was used to assess the responsiveness of the scale. All 340 HF patients having complete TCM-HF-PRO data were assigned to the treatment group and the control group by central randomization. The total TCM-HF-PRO scores of the two groups were compared using paired t-test to reflect the longitude responsiveness of the scale before treatment and at week 2 after treatment.

RESULTS(1) The recycling rate of the scale was 100.0%. One of them was not filled completely, which was rejected thereby. So the completion rate was 99.7%. The completion time for TCM-HF-PRO scale ranged 15 to 25 min. (2) The Cronbach's α coefficient of rating scale was 0.903, split-half reliability was 0.844 and 0.849. (3) Confirmatory factor analysis showed that 7 factors and items formed according to maximum load factor basically coincided with the construct of the rating scale, 7 factors accumulated contribution rate was 43.8%. TCM clinical outcomes rating scale for HF based on patients' report was relatively better correlated with the Minnesota living with HF questionnaire (r = 0.726, P < 0.01). (4) Discriminant analysis showed that the rating scale correctly classified more than 78.8% of case studies having confirmed initial differential diagnosis by experts. The total scale of the rating scale decreased more in the two group after treatment, with significant difference as compared with before treatment (P < 0.01.

CONCLUSIONTCM clinical outcomes rating scale for HF based on patients' report had good reliability, validity and responsiveness, hence it could be used to assess clinical efficacy for HF patients.

Diagnosis, Differential ; Discriminant Analysis ; Factor Analysis, Statistical ; Heart Failure ; diagnosis ; Humans ; Medicine, Chinese Traditional ; methods ; standards ; Reproducibility of Results ; Surveys and Questionnaires

OBJECTIVETo clone Helicobacter pylori flagellin B gene (flaB) to construct prokaryotic expression system of the gene and to identify immunity of the fusion protein.

METHODSThe flaB gene from a clinical isolate Y06 of H.pylori was amplified by high fidelity PCR. The nucleotide sequence of the target DNA amplification fragment was sequenced after T-A cloning. The expression vector pET32a with inserted flaB gene was constructed. FlaB fusion protein was expressed in E.coli strain BL21DE3 inducted by IPTG at different dosages. Western blot was applied to determine immunoreactivity and immunogenicity of the fusion protein and antibody against whole cell of H.pylori and rabbit antiserum immunized with the fusion protein, respectively.

RESULTSIn comparison with the reported corresponding sequences, the homology of nucleotide sequence of the cloned flaB gene was from 96.31% approximate, equals 97.73%, while the homology of its putative amino acid sequence was as high as 99.41% approximate, equals 100%. The expression output of FlaB fusion protein in pET32a-flaB-BL21DE3 system was approximately 40% of the total bacterial proteins. FlaB fusion protein was able to combine with antibody against whole cell of H.pylori and induce rabbit to produce specific antibody with high titer after the animal was immunized with the protein.

CONCLUSIONA prokaryotic expression system of H. pylori flaB gene with high efficiency has been established successfully. The expressed FlaB fusion protein with satisfactory immunogenicity and immunoreactivity can be used as antigen in H.pylori vaccine and detect kit.

Antibodies, Bacterial ; blood ; Bacterial Vaccines ; immunology ; Base Sequence ; Cloning, Molecular ; Enzyme-Linked Immunosorbent Assay ; Flagellin ; genetics ; immunology ; Helicobacter pylori ; immunology ; Humans ; Polymerase Chain Reaction ; Recombinant Fusion Proteins ; immunology ; Vaccines, Synthetic ; immunology

OBJECTIVETo clone Helicobacter pylori adhesin (hpaA) gene,to construct the expression vector of the gene and to identify immunogenicity of the fusion protein.

METHODSThe hpaA gene from a clinical isolate Y06 of H.pylori was amplified by high fidelity PCR. The nucleotide sequence of the target DNA amplification fragment was sequenced after T-A cloning. The expression vector pET32a with inserted hpaA gene was constructed. hpaA fusion protein was expressed in E.coli strain BL21DE3 induced by IPTG at different dosages. Western blot using antibody against whole cell of H.pylori as well as immunodiffusion assay using antiserum of rabbit against the fusion protein was applied to determine immunogenicity of the fusion protein.

RESULTSIn comparison with the reported corresponding sequences, the homology of nucleotide sequence of the cloned hpaA gene was from 94.25% approximate, equals 97.32%, while the homology of its putative amino acid sequence was as high as 95.38% approximate, equals 98.46%. The expression output of HpaA fusion protein in pET32a-hpaA-BL21DE3 system was approximately 40% of the total bacterial proteins. HpaA fusion protein was able to combine with antibody against whole cell of H.pylori and induce rabbit to preduce high titer antibody after the animal was immunized with the protein.

CONCLUSIONAn expression system with high efficiency of H.pylori hpaA gene has been established successfully. The expressed HpaA fusion protein with satisfactory immunogenicity and immunoreactivity can be used as antigen in H.pylori vaccine.

Adhesins, Bacterial ; Animals ; Antibodies, Bacterial ; blood ; Bacterial Vaccines ; immunology ; Base Sequence ; Cloning, Molecular ; Enzyme-Linked Immunosorbent Assay ; Helicobacter pylori ; immunology ; Hemagglutinins ; genetics ; immunology ; Humans ; Polymerase Chain Reaction ; Rabbits ; Recombinant Fusion Proteins ; immunology ; Vaccines, Synthetic ; immunology

OBJECTIVETo clone Helicobacter pylori ureB gene, to construct prokaryotic expression system of the gene and to identify immunogenicity of the fusion protein.

METHODSThe ureB gene from a clinical isolate Y06 of H.pylori was amplified by high fidelity PCR. The nucleotide sequence of the target DNA amplification fragment was sequenced after T-A cloning. The expression vector pET32a with inserted ureB gene was constructed. ureB fusion protein was expressed in E.coli strain BL21DE3 induced by IPTG at different dosages. Western blot using antibody against whole cell of H.pylori as well as immunodiffusion assay using antiserum of rabbit against the fusion protein was applied to determine immunogenicity of the fusion protein.

RESULTSIn comparison with the reported corresponding sequences, the homology of nucleotide sequence of the cloned ureB gene was from 96.88% approximate, equals 97.82%, while the homology of its putative amino acid sequence was as high as 99.65% approximate, equals 99.82%. The expression output of UreB protein in pET32a-ureB-BL21DE3 system was approximately 40%of the total bacterial proteins. UreB protein was able to combine with antibody against whole cell of H.pylori and induce rabbit to produce high titer antibody after the animal was immunized with the protein.

CONCLUSIONAn expression system with high efficiency of H.pylori ureB gene has been established successfully. The expressed UreB protein with satisfactory immunogenicity and immunoreactivity can be used as antigen in H.pylori vaccine.

Animals ; Bacterial Vaccines ; immunology ; Base Sequence ; Helicobacter pylori ; enzymology ; immunology ; Humans ; Molecular Sequence Data ; Polymerase Chain Reaction ; Rabbits ; Recombinant Fusion Proteins ; immunology ; Urease ; genetics ; immunology ; Vaccines, Synthetic ; immunology

OBJECTIVETo determine the immunoprotective effects of rUreB and rHpaA bivalence genetic engineering vaccine with inner adjuvant of rLTB on BALB/c mice against H.pylori strain SS1 infection.

METHODSrUreB, rHpaA, rLTB-rUreB-rHpaA, rLTKA63, rLTB and rLTB were collected by NTA-Ni affinity chromatography. Western blot was applied to demonstrate the immunoreactivity of the recombinant protein antigens. Adjuvant activities of rLTB, rCTB and rLTB-rUreB-rHpaA were determined by GM1-ELISA. H.pylori strain SS1-infected BALB/c mouse modal was established to measure immunoprotective effects of different compositions of antigens and adjuvants. H.pylori in gastric biopsy specimens was examined by routine isolation method and silver staining method. Two ELISAs were established to detect specific S-IgA in gastric juices and specific IgA in sera of the immunized mice.

RESULTSrUreB, rHpaA and rLTB-rUreB-rHpaA were recognized by commercial antibody against whole cell of H.pylori and were able to combine to the bovine GM1. The protective rate in the mice immunized with single rUreB or rHpaA was lower than 70%. When using rUreB or rHpaA plus rLTB or rCTB, the positive rates increased to 75.0%-83.3%. With different combination of antigens and adjuvants, the immunoprotective rate of rLTB-jrUreB-rHpaA was as high as 100%, and then was 91.7% for rUreB+rHpaA+rCTB+rLTKA63 and was 90.9% for rUreB+rHpaA+rLTB. Both the rLTB and rCTB showed remarkable effects to induce specific IgA in sera and specific S-IgA in gastric juices of the immunized mice, and the former showed stronger S-IgA-inducing ability than the latter. The positive rates of specific IgA were basically identical to the corresponding mouse immunoprotective rates. S-IgA positive rates specific to rUreB and rHpaA in rLTB-rUreB-rHpaA immunized mice were 100% and 91.7%, respectively.

CONCLUSIONThe rUreB and rHpaA possess qualified immunoreactivity and antigenicity. The rLTB and rCTB can show adjuvant activity of mucosal immunization. The high immunoprotective rate of Helicobacter pylori UreB and HpaA bivalence recombinant vaccine with inner adjuvant in mice is associated with high dosage of rLTB, larger molecular weight of the antigen and the high levels of local specific S-IgA.

Adjuvants, Immunologic ; biosynthesis ; genetics ; Animals ; Bacterial Proteins ; biosynthesis ; genetics ; immunology ; Bacterial Vaccines ; immunology ; Carrier Proteins ; biosynthesis ; genetics ; immunology ; Female ; Genetic Engineering ; Helicobacter Infections ; prevention & control ; Helicobacter pylori ; immunology ; Immunoglobulin A ; blood ; metabolism ; Mice ; Mice, Inbred BALB C ; Recombinant Proteins ; biosynthesis ; genetics ; immunology ; Vaccines, Synthetic ; immunology

OBJECTIVETo determine vacA dominant genotypes of Helicobacter pylori in patients with peptic ulcer (PU) or chronic gastritis (CG).

METHODSH.pylori strains were isolated from mucosa samples of gastric antrum and corpus of patients suffering from PU (n=29) or CG (n=34), 126 strains of H.pylori were selected for PCR to detect s and m regions in vacA gene of the isolates. Parts of the amplification products were sequenced after T A cloning. The correlation between infection or coinfection with different vacA genotypes of H.pylori and different gastroduodenal diseases was further analyzed.

RESULTSThe positive amplification products of vacA gene, s and m regions, were found in the DNA samples of all the isolates. In these products, s1a/m1, s1a/m2, s1a/m1b and s1a/m1b m2 genotypes of vacA gene were detected and s1b and s2m1a genotypes absent. Proportions of the s1a/m1, s1a/m2, s1a/m1b and s1a/m1b-m2 genotypes were 7.1% (9/126), 61.9% (78/126), 29.4% (37/126) and 1.6% (2/126), respectively. 17.5% (11/63) of the patients were confirmed to be coinfected with different genotype H.pylori strains. No statistical differences were found in the distribution of different genotype H.pylori strain infection in the gastric diseases (P>0.05). In comparison with the reported sequences of H.pylori strain 60190 with s1a genotype and strain 87-203 with m2 genotype, homologies of the nucleotide sequences of s1a PCR products from 6 strains of H.pylori isolates and m2 PCR products from 4 strains of H.pylori isolates were 93.15% approximate, equals 94.86%and 93.63% approximate, equals 97.61%, respectively.

CONCLUSIONH.pylori with s1a/m2 or s1a/m1b are the dominant genotypes in the PU or GC patients in Zhejiang area. The nucleotide sequences of partial amplification products from the vacA dominant genotypes of H.pylori show high homology compared with the reported sequences. Part of the patients may be coinfected with different vacA genotypes of H.pylori.

Adolescent ; Adult ; Aged ; Bacterial Proteins ; genetics ; Base Sequence ; Child ; Female ; Genotype ; Helicobacter pylori ; genetics ; Humans ; Male ; Middle Aged

OBJECTIVE: To investigate the frequency of transfusion transmitted virus (TTV) infection in healthy blood donors in Hangzhou area and the mutation of TTV genomic fragment. METHODS DNA in serum samples of 203 healthy donors was extracted by phenol-chloroform method to detect TTV by semi-nested polymerase chain reaction and nucleotide sequences of partial amplification products were determined after T-A cloning. RESULTS TTV infection rate in 203 cases of blood donors in Hangzhou area was 15.3%. The homology of the amplified products of partial TTV positive samples compared with thereported nucleotide and putative amino acid sequences of TTV TA278 were 63.51% approximate, equals 67.12% and 59.46% approximate, equals 66.22% respectively. CONCLUSIONS TTV infection rate in the blood donors in Hangzhou is relatively high. The TTV infecting blood donors in the area may be a kind of novel genotype.

OBJECTIVE This work aimed to investigate the anti-rheumatoid arthritic effect of gentio-picroside from Gentiana macrophylla Pall using an animal model of adjuvant induced arthritis. METH-ODS Adjuvant arthritis was induced in fifty SD male rats,which were randomly divided into five groups (n=10):control(0.5% CMC-Na)group,AIA(rats with CFA)group,dexamethasone(1 mg·kg-1)group, gentiopicroside(50 mg·kg-1)group,and gentiopicroside(100 mg·kg-1)group.Rats were administered intragastrically with drugs or CMC-Na once a day for a period of 2 weeks.Paw swelling,arthritic index, histological changes were assessed to evaluate the anti-arthritic effect.Weight growth,spleen and thymus indexes were also investigated in.RESULTS Gentiopicroside at dose of 100 mg·kg-1significantly inhibited the secondary paw swelling(P<0.05)and arthritis index(P<0.05),decreased synovial inflammatory infil-tration, synovial hyperplasia and bone erosion. Furthermore, gentiopicroside showed no immunosup-pressive adverse effects in body weight, index of spleen and thyums compared with dexamethasone administration (P<0.05, P<0.01). CONCLUSION Gentiopicroside possessed anti-arthritic efficacy in AIA rats without immunosuppressive effects.

OBJECTIVETo construct prokaryotic expression systems of ltB/ctB-lipL41/1 fusion genes, identify immunogenic and adjuvant activities of the products as well as to understand the frequencies of lipL41 gene that carrying and expressing in L. interrogans wild strains and specific antibody levels in sera from patients with leptospirosis.

METHODSPolymerase chain reaction (PCR) with linking primer was applied to construct the fusion genes ltB-lipL41/1 and ctB-lipL41/1. By routine molecular biological techniques, prokaryotic expression systems of the two fusion genes were constructed. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was used to examine expression of the target recombinant proteins rLTB-rLipL41/1 and rCTB-rLipL41/1. Both western blot and Ganglioside-enzyme linked immunosorbent assay (GM-ELISA) were used while immunogenic and adjuvant activities of rLTB-rLipL41/1 and rCTB-rLipL41/1 were measured. PCR and MAT were performed to detect lipL41 gene and expression of the gene in 97 wild strains of L. interrogans, respectively. Antibodies against product of lipL41 gene in serum samples from 228 leptospirosis patients were detected by ELISA.

RESULTSIn comparison with reported corresponding sequences, the similarities of ltB-lipL41/1 and ctB-lipL41/1 fusion genes to nucleotide and putative amino acid sequence were 99.6%-99.9% and 99.8%-100%, respectively. Expression outputs of both rLTB-rLipL41/1 and rCTB-rLipL41/1, mainly presenting with inclusion body, consisting approximate 10% of the total bacterial proteins. Both rLTB-rLipL41/1 and rCTB-rLipL41/1 could combine rabbit anti-rLipL41/1 serum as well as bovine GM1, respectively. 87.6% of the L. interrogans wild strains(85/97) having lipL41 gene while 84.5% (82/97) of the wild strains with rLipL41/1 or rLipL41/2 antiserum were positive for MAT with titers of 1:4-1:128. 84.6% (193/ 228), 78.5% (179/228) from the patients' serum samples were positive for rLipL41/1 and rLipL41/2 antibodies, respectively.

CONCLUSIONltB-lipL41/1 and ctB-lipL41/1 fusion genes and their prokaryotic expression systems were successfully constructed in this study. The two expressed fusion proteins showed qualified immunogenic and adjuvant activities. lipL41 gene was extensively distributed and frequently expressed in different serogroups of L. interrogans. rLTB-rLipL41/1 or rCTB-rLipL41/1 seemed to have had good potential to serve as an antigen in L. interrogans genus-specific vaccine.

Amino Acid Sequence ; Animals ; Antibody Specificity ; Antigens, Bacterial ; biosynthesis ; chemistry ; genetics ; immunology ; Cattle ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli Proteins ; genetics ; Genetic Engineering ; methods ; Humans ; Leptospira interrogans ; genetics ; physiology ; Leptospirosis ; immunology ; Polymerase Chain Reaction ; Recombinant Fusion Proteins ; biosynthesis ; chemistry ; genetics ; immunology ; Sequence Analysis, DNA

Small intestinal obstruction is a common complication of primary gastrointestinal cancer or metastatic cancers.Patients with this condition are often poor candidates for surgical bypasses,and placement of self-expanding metal stent (SEMS) can be technically challenging.In this study,we examined the feasibility of combined application of single-balloon enteroscope (SBE) and colonoscope for SEMS placement in patients with malignant small intestinal obstruction.Thirty-four patients were enrolled in this study,among which 22 patients received SEMS placement by using SBE and colonoscope,while the other 12 patients received conservative medical treatment.The patients were followed up for one year.Stent placernent was technically feasible in 95.5％ (21/22).Clinical improvement was achieved in 86.4％ (19/22).For the 19 clinical success cases,the average time of benefits from a gastric outlet obstruction scoring system (GOOSS) increase ≥1 was 111.9±89.5 days.For the 12 patients receiving conservative medical treatment,no significant improvement in GOOSS score was observed.Moreover,a significant increase of Short-Form-36 health survey score was observed in the 19 patients at time of 30 days after stent placement.By Kaplan-Meier analysis,a significant survival improvement was observed in patients with successful SEMS placement,compared with patients receiving conservative medical treatment.Taken together,combined use of SBE and colonoscope makes endoscopic stent placement feasible in patients with malignant small intestinal obstruction,and patients can benefit from it in terms of prolonged survival and improved quality of life.