AIM:To observe whether modified epitopes from osteosarcoma high-expressing antigen papilloma-virus-binding factor (PBF) have HLA-A2 restricted antitumor ability,and to develop peptide-based immunotherapy for os-teosarcoma. METHODS:RT-PCR and Western blot were used to determine the expression of PBF in the osteosarcoma cell lines U2OS and Saos-2. HLA-A2 epitopes from PBF protein were predicted by NetCTL 1.2, SYFPEITHI and IEDB. The modified peptides from PBF containing HLA-A2 binding anchor motifs were designed by replacing the anchor residues. The peptides were synthesized by standard solid-phase methods,and the binding affinity of the peptides to HLA-A?0201 was evaluated by T2A2 cell binding assay. ELISPOT assay was used to investigate the seretion of interferon-γ(IFN-γ) from the peptide-induced specific cytotoxic T-lymphocytes (CTLs). The ability of inducing T-cell response was analyzed by lactate dehydrogenase (LDH) release assay and carboxyfluorescein succinimidyl ester (CFSE) cytotoxicity assay in vitro. RE-SULTS:The expression of PBF was observed in the U2OS and Saos-2 cells. The candidate peptides P75-1Y2L,P412-1Y, P416-1Y2L9V, P107-1Y and P435-1Y2L showed moderate affinity toward HLA-A2 molecule. The modified peptides showed significantly higher affinity with HLA-A2 than the native peptide. ELISPOT assay showed that P412, P412-1Y, P416,P416-1Y2L9V and P435-1Y2L induced specific CTLs to secrete IFN-γ, and P412-1Y and P416-1Y2L9V induced more secretion of IFN-γ than the native peptide. The CTLs induced by P412, P412-1Y, P416 and P416-1Y2L9V lysed U2OS cells. P412-1Y and P416-1Y2L9V peptide-specific CTLs showed higher cytotoxicity against U2OS cells than the native peptide-specific CTLs. CONCLUSION:Compared with the native peptide,modified epitopes P412-1Y and P416-1Y2L9V have higher binding affinity with HLA-A?0201 and retain immunogenecity. In addition,the anti-tumor immunity effects of modified epitopes P412-1Y and P416-1Y2L9V are stronger than the native peptide. The peptides P412-1Y and P416-1Y2L9V is excellent HLA-A?0201 restricted CTL epitopes from tumor antigen PBF,which could serve as new can-didates towards antitumor peptide vaccines.

Aged ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Carcinoma, Squamous Cell ; drug therapy ; radiotherapy ; Combined Modality Therapy ; Esophageal Diseases ; etiology ; Esophageal Neoplasms ; drug therapy ; radiotherapy ; Female ; Fluorouracil ; administration & dosage ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Particle Accelerators ; Platinum Compounds ; administration & dosage ; Radiation Injuries ; etiology ; Radiotherapy, High-Energy ; adverse effects

Animals ; Fatty Liver, Alcoholic ; drug therapy ; Female ; Male ; Phytotherapy ; Rats ; Rats, Wistar ; Saponins ; therapeutic use ; Trigonella

Objective To establish an extensively drug-resistant Acinetobacter baumannii (XDR-AB) infection model using Caenorhabditis elegans (C.elegans),and evaluate the effect of efflux pump inhibitors(EPIs) on reversal of ciprofloxacin resistance in XDR-AB.Methods XDR-AB infection model of C.elegans was established,six EPIs(CCCP,PAβN,NMP,omeprazole,reserpine,and verapamil)combined with ciprofloxacin were used to treat the infected model,the survival rate of C.elegans was recorded to evaluate the in vivo activities of drugs,toxicity test and in vitro drug susceptibility test were also performed.Results Lethal effect of different concentrations of XDR-AB on C.elegans was varied,5 × 106 CFU/mL of XDR-AB was selected to infect C.elegans.C.elegans survival test showed that survival curves of C.elegans infected with XDR-AB for 3 hours and curves of control group (polymixin B was added) were not significantly different (x2 =3.154,P＞0.05);compared with control group,survival curves of C.elegans infected with XDR-AB for 6 hours or 9 hours were significantly different (both P＜0.001),but 6 hours and 9 hours were not significantly different(x2 =0.669,P＞0.05),6 hours was chosen as the duration of infection,36 hours was appropriate for the duration of antimicrobial therapy.Ciprofloxacin with EPIs for infection model revealed that low concentration of PAβN,NMP,omeprazole,and reserpine could improve the survival rate of C.elegans by 30％-40％,15％-20％,20％-30％,and 20％ respectively,high concentration of verapamil could improve the survival rate of infected C.elegans by about 30％.In vitro susceptibility test and toxicity test results showed that ciprofloxacin combined respectively with CCCP,omeprazole,and verapamil could reduce minimum inhibitory concentration(MIC) to the original 1/4,combined respectively with PAβN,NMP,and reserpine could reduce MIC to the original 1/2,CCCP had the best bacterial inhibitory effect in vitro,but the toxicity was large,and was not suitable for the study of pharmacodynamics in vivo.Conelusion The infection model of C.elegans XDR-AB is initially and successfully established,which is used to evaluate the efficiency of six EPIs for reversing ciprofloxacin resistance.

Objective To investigate the effects of intravenous immunoglobulin(IVIG) on apoptosis and necrosis of myocytes in mice with viral myocarditis.Methods Three hundreds and twenty Balb/c mice were randomly divided into 8 groups.Different courses of IVIG were given in varying time after virus inoculation,Chinese medicine Huangqi given in control group.The virus titer in myocardium、percentage of apoptosis and necrosis of myocytes were detected, myocardial histopathologic scores were counted.Results In every IVIG treatment group,the above 3 items were all significantly lower than that in virus control group and Huangqi group,as IVIG early long course group had the best effect.Conclusion IVIG may reduce the percentage of apoptosis and necrosis of myocytes and virus titer in myocardium in mice with viral myocarditis,the effects are better than that of Huangqi.

In order to study the development characteristics of Rehmannia glutinosa tuberous root expansion and reveal the regulation mechanism of the genes related to hormones in this process, R. glutinosa "wen-85" was used as the experimental material in this study. R. glutinosa tuberous roots of different developmental stages were collected to observe phenotype and tissue morphology using resin semi-thin sections method. The genes related to hormone biosynthesis and response were chosen from the transcriptome of R. glutinosa, which was previously constructed by our laboratory, their expression levels at different development stages were measured by real-time quantitative PCR. The results showed that the root development could be divided into six stages: seeding, elongation, pre-expanding, mid-expanding, late-expanding and maturity stage. The anatomic characteristics indicated that the fission of secondary cambium initiated the tuberous root expansion, and the continuous and rapid division of secondary cambium and accessory cambium kept the sustained and rapid expansion of tuberous root. In addition, a large number oleoplasts were observed in root on the semi-thin and ultra-thin section. The quantitative analysis suggested that the genes related to biosynthesis and response of the IAA, CK, ABA,ethylene, JA and EB were up-regulated expressed, meanwhile, GA synthesis and response genes were down-regulated expressed and the genes of GA negative regulation factors were up-regulated expressed. The maximum levels of most genes expression occurred in the elongation and pre-expansion stage, indicating these two stages were the key periods to the formation and development of tuberous roots. Oleoplasts might be the essential cytological basis for the formation and storage of the unique medicinal components in R. glutinosa. The results of the study are helpful for explanation of development and the molecular regulation mechanism of the tuberous root in R. glutinosa.
Gene Expression Regulation, Developmental ; drug effects ; genetics ; Gene Expression Regulation, Plant ; drug effects ; genetics ; Lipid Droplets ; metabolism ; ultrastructure ; Microscopy, Electron, Transmission ; Plant Growth Regulators ; biosynthesis ; pharmacology ; Plant Proteins ; genetics ; metabolism ; Plant Roots ; genetics ; growth & development ; metabolism ; Rehmannia ; genetics ; growth & development ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Time Factors

Atrioventricular Block ; etiology ; Cardiac Catheterization ; adverse effects ; Heart Septal Defects, Ventricular ; therapy ; Hematologic Diseases ; etiology ; Hemolysis ; Humans

OBJECTIVEThis study was to simultaneously identify Campylobacter jejuni and Campylobacter coli isolates in China by Multi-PCR assay and to study the prevalence of six virulence and toxin genes on them.

METHODSA multi-PCR method with three sets of primers specifically designed for application of a 16S rRNA as a universal control, mapA, ceuE based on the specific sequence of C. jejuni and C. coli, was applied to detect 65 Campylobacter isolates from China. Another two separately PCR Primers were directed towards the hippuricase gene (hipO) characteristic of C.jejuni and glyA gene characteristic of C. coli were performed for further confirmation. The presence of the cadF, virB11, flaA, cdtA, cdtB, cdtC genes among these 65 strains were investigated by PCR.

RESULTSFrom multi-PCR detection, 42 isolates belonged to C. jejuni, other 23 isolates belong to C. coli. Data showing the identification were 100% in concordance with the separated PCR for hipO and glyA amplification. The efficiency (100%) of identification by these three primers multi-PCR method was higher than the biochemical test (83.1%). The cadF and flaA genes were detected from 100% (65/65) of the isolates and the PCR product of each gene were identical with each isolate. Only 10.8% (7/65) of the isolates were positive for virB11. The cdtA gene was found in 92% (60/65) of the isolates. 97.6% (41/42) of C. jejuni had cdtB gene, whereas no PCR product with this primers for all the C. coli isolates. cdtC was presented in all the isolates but the lengths of PCR products were different. For C. jejuni, it was 555 bp, for C. coli, it was about 465 bp.

CONCLUSIONThis three primers simultaneous multi-PCR method seemed to be useful for the identification of C. jejuni and C. coli isolates from China since cadF and flaA genes were widely spread in Campylobacter isolates in this country. The present report on virB11 was similar to previous reports from other countries, but the distribution of cdt gene cluster in Campylobacter species isolated from China might be different.

Campylobacter coli ; genetics ; isolation & purification ; pathogenicity ; Campylobacter jejuni ; genetics ; isolation & purification ; pathogenicity ; China ; DNA Primers ; Genes, Bacterial ; Polymerase Chain Reaction ; Virulence ; genetics

Objective To set up the reference value of serum glyeated albumin (CA) in Chinese for using in clinical practice through a multi-center clinical trial. Methods Three hundred and eighty individuals with normal weight and normal glucose regulation, including 183 males and 197 females ranging from 20 to 69 years, were recruited from 10 hospitals in China. Serum GA levels were measured with liquid enzymatic method. Results (1) The GA level of the 380 subjects was (14. 5±1.9)%. When dividing these subjects by age into 3 subgroups, there was no difference in the GA levels among the 3 subgroups (P>0.05). Compared with the women, the men had higher GA level in the first subgroup aging from 20 to 39 (P =0.028). However, no significant difference was detected after adjusting with BMI as confounder.(2) When dividing those subjects by BMI into 3 subgroups, with BMI ranging from 18. 5-20. 9 kg/m2,21.0-22. 9 kg/m<'2>and 23.0-24. 9 kg/m<'2>respectively, we came to the following results: for men, there was no difference in the GA levels among the 3 subgroups (P>0.05), but for women, the GA level of the first subgroup was higher than that of the second subgroup (P =0.024). (3) The level of GA in the 2. 5th to 97.5th percentile was 10. 8%-17. 1%. (4) Sixty normal subjects were chosen to repeat evaluation of GA levels after 2-3 weeks and the GA levels were of no difference (P>0.05).Conclusion The normal range of serum GA for the Chinese population could be suggested at 11%-17%.

Animals ; beta Catenin ; Blotting, Western ; Breast Neoplasms ; Breast ; Cell Self Renewal ; Flow Cytometry ; Fluorescent Antibody Technique ; Genome ; Humans ; In Vitro Techniques ; Kinesin ; Luciferases ; Mice ; Neoplasm Metastasis ; Neoplastic Stem Cells ; Prognosis ; Real-Time Polymerase Chain Reaction ; Stem Cells