Objective To predict and identify T cell epitopes of major group 3 allergen derived from Dermatophagoides fari-na(Der f 3). Methods The T cell epitopes of Der f 3 were analyzed through the sequence analysis by using the bioinformatics online tools. The five predicted peptides of T-cell epitopes were artificially synthesized. The spleen lymphocytes were co-cultured with the five T cell epitopes by using the modified MTT method and the levels of IL-2,IFN-γ,IL-4 and IL-5 in the supernatant of the cultures were detected by ELISA. Results Five T cell epitopes of Der f 3 were predicted and three of which could pro-mote the proliferation of the mouse spleen lymphocytes. The secretions of IL-2 and IFN-γwere significantly induced and the se-cretions of IL-4 and IL-5 were significantly decreased by three of five prediction epitopes of Der f 3:37GDCPYQISLQSSSHFC-GG54,98IYQHENYDSMTIDNDVALIKLKTPMT123 and 164SELQRVDIDVVSREQCDQLYS184. Conclusion Three T cell epitopes of Der f 3 have been initially identified,which lays the foundation of the diagnosis and treatment of asthma.

Objective To predict and identify the linear B-cell epitopes in the major group 3 allergen derived from Derma-tophagoides farina(Der f 3). Methods The linear B-cell epitopes of Der f 3 allergen were analyzed based on the physicochemi-cal properties of amino acids including antigenicity,surface accessibility,flexibility,hydrophilicity,beta-turn by online bioinfor-matics softwares. The eight predicted peptides of linear B-cell epitopes were artificially synthesized and incubated with three aller-gic serum pools(4 serum samples in each),which were consisted of total 12 serum samples from the allergic individuals,and the strong positive epitopes were selected. Results Eight B-cell epitopes from Der f 3 were predicted successfully. Five of eight B-cell epitopes were identified with strong IgE-binding abilities followed by specific IgE assay. The amino acid sequences of them were following:33KAKAGDCP40, 86HASGGEKIQVAEIYQHENYDSMTID110, 118LKTPMTLDQTNAKPVPLPPQGSDVKVG144, 156QEGSYSLP163 and 199DVANGGVDSCQGDSGGPVVD218. Conclusions Five linear B-cell epitopes of Der f 3 allergen have been identified successfully. This result might provide a basis of the diagnosis and treatment for asthma.

OBJECTIVE: To identify sarcosaphagous flies and their larvae, pupa. METHODS: Sarcosaphagous flies and their larvae, pupas were collected from human corpses and their surroundings in the Weifang city. A 304 bp region in COI gene was analyzed by mtDNA sequencing. RESULTS: The studied region showed no sequence divergence within same species and significant difference were found between different species in all samples. CONCLUSION It is a practical approach to identify these Sarcosaphagous flies and their larvae, pupas by sequence analysis of the 304bp region of the COI in mtDNA.
Animals ; Base Sequence ; China ; DNA Primers ; DNA, Mitochondrial/genetics* ; Diptera/genetics* ; Electron Transport Complex IV/genetics* ; Forensic Medicine ; Genes, Insect ; Humans ; Larva/genetics* ; Phylogeny ; Polymerase Chain Reaction/methods* ; Pupa/genetics* ; Sequence Analysis, DNA/methods* ; Species Specificity

OBJECTIVE: To improve the correct rate of ABO genotyping by meliorating AS-PCR primer. METHODS: The primer P1 was changed into primer P1' by substituting the fifth base G for C of 3' end and the ABO genotyping results of primer P1 and P1' was compared and analysed. RESULTS: The non-specific product of OO typing is reducing and the wrong genotyping of OO and AO was avoided by meliorating AS-PCR primer. CONCLUSION The rates of wrong ABO genotyping results could be effectively reduced by using altering primer P1'.
ABO Blood-Group System/genetics* ; Alleles ; Base Sequence ; DNA Primers/genetics* ; Genotype ; Humans ; Molecular Sequence Data ; Polymerase Chain Reaction/methods* ; Sensitivity and Specificity

OBJECTIVETo construct a vector encoding T-cell epitopes of major allergen group 1 of Dermatophagoides pteronyssinus as a vaccine delivered by MHC class II pathway.

METHODSThe nucleotide sequences of the 3 target genes were synthesized, including TAT, IhC and the recombinant fragment of Der p 1 encoding 3 T-cell epitopes. After amplification of the 3 target fragments by PCR and digestion with corresponding restriction endonucleases, the recombinant gene TAT-IhC-Der p 1-3T was ligated using T4 DNA ligase and inserted into the prokaryotic expression vector pET28a(+) to construct the recombinant plasmid pET-28a(+)-TAT-IhC-Der p 1-3T, which was confirmed by digestion with restriction endonucleases and sequencing. The recombinant vector was transformed into E. coli strain BL21 (DE3) and induced with IPTG, and the induced protein TAT-IhC-Der p 1-3T was detected by SDS-PAGE. After purification, the recombinant protein was confirmed by Western blotting and its allergenicity tested using IgE-binding assay.

RESULTSThe recombinant plasmid pET-28a-TAT-IhC-Der p 1-3T was successfully constructed as confirmed by restriction endonuclease digestion and sequencing and the expression of the recombinant protein TAT-IhC-Der p 1-3T was induced in E. coli. Western blotting verified successfull purification of the target protein, which showed a stronger IgE-binding ability than Der p 1.

CONCLUSIONWe successfully constructed a recombinant expression vector pET-28a-TAT-IhC-Der p 1-3T expressing a T-cell epitope vaccine delivered by MHC II pathway with strong IgE-binding ability, which provides a basis for further study on specific immunotherapy via MHC class II pathway.

Allergens ; immunology ; Animals ; Antigens, Dermatophagoides ; immunology ; Arthropod Proteins ; immunology ; Base Sequence ; Cloning, Molecular ; Cysteine Endopeptidases ; immunology ; Dermatophagoides pteronyssinus ; Epitopes, T-Lymphocyte ; Escherichia coli ; Gene Expression ; Genes, MHC Class II ; Genetic Vectors ; Plasmids ; Polymerase Chain Reaction ; Recombinant Proteins ; immunology ; Vaccines ; immunology

Objective To construct a vector encoding T-cell epitopes of major allergen group 1 of Dermatophagoides pteronyssinus as a vaccine delivered by MHC class II pathway. Methods The nucleotide sequences of the 3 target genes were synthesized, including TAT, IhC and the recombinant fragment of Der p 1 encoding 3 T-cell epitopes. After amplification of the 3 target fragments by PCR and digestion with corresponding restriction endonucleases, the recombinant gene TAT-IhC-Der p 1-3T was ligated using T4 DNA ligase and inserted into the prokaryotic expression vector pET28a(+) to construct the recombinant plasmid pET-28a(+)-TAT-IhC-Der p 1-3T, which was confirmed by digestion with restriction endonucleases and sequencing. The recombinant vector was transformed into E. coli strain BL21 (DE3) and induced with IPTG, and the induced protein TAT-IhC-Der p 1-3T was detected by SDS-PAGE. After purification, the recombinant protein was confirmed by Western blotting and its allergenicity tested using IgE-binding assay. Results The recombinant plasmid pET-28a-TAT-IhC-Der p 1-3T was successfully constructed as confirmed by restriction endonuclease digestion and sequencing and the expression of the recombinant protein TAT-IhC-Der p 1-3T was induced in E. coli. Western blotting verified successfull purification of the target protein, which showed a stronger IgE-binding ability than Der p 1. Conclusion We successfully constructed a recombinant expression vector pET-28a-TAT-IhC-Der p 1-3T expressing a T-cell epitope vaccine delivered by MHC II pathway with strong IgE-binding ability, which provides a basis for further study on specific immunotherapy via MHC class II pathway.

Objective To construct a vector encoding T-cell epitopes of major allergen group 1 of Dermatophagoides pteronyssinus as a vaccine delivered by MHC class II pathway. Methods The nucleotide sequences of the 3 target genes were synthesized, including TAT, IhC and the recombinant fragment of Der p 1 encoding 3 T-cell epitopes. After amplification of the 3 target fragments by PCR and digestion with corresponding restriction endonucleases, the recombinant gene TAT-IhC-Der p 1-3T was ligated using T4 DNA ligase and inserted into the prokaryotic expression vector pET28a(+) to construct the recombinant plasmid pET-28a(+)-TAT-IhC-Der p 1-3T, which was confirmed by digestion with restriction endonucleases and sequencing. The recombinant vector was transformed into E. coli strain BL21 (DE3) and induced with IPTG, and the induced protein TAT-IhC-Der p 1-3T was detected by SDS-PAGE. After purification, the recombinant protein was confirmed by Western blotting and its allergenicity tested using IgE-binding assay. Results The recombinant plasmid pET-28a-TAT-IhC-Der p 1-3T was successfully constructed as confirmed by restriction endonuclease digestion and sequencing and the expression of the recombinant protein TAT-IhC-Der p 1-3T was induced in E. coli. Western blotting verified successfull purification of the target protein, which showed a stronger IgE-binding ability than Der p 1. Conclusion We successfully constructed a recombinant expression vector pET-28a-TAT-IhC-Der p 1-3T expressing a T-cell epitope vaccine delivered by MHC II pathway with strong IgE-binding ability, which provides a basis for further study on specific immunotherapy via MHC class II pathway.

Objective: To elucidate the biological basis of the heart qi deficiency (HQD) pattern, an in-depth understanding of which is essential for improving clinical herbal therapy. Methods: We predicted and characterized HQD pattern genes using the new strategy, TCM-HIN2Vec, which involves heterogeneous network embedding and transcriptomic experiments. First, a heterogeneous network of traditional Chinese medicine (TCM) patterns was constructed using public databases. Next, we predicted HQD pattern genes using a heterogeneous network-embedding algorithm. We then analyzed the functional characteristics of HQD pattern genes using gene enrichment analysis and examined gene expression levels using RNA-seq. Finally, we identified TCM herbs that demonstrated enriched interactions with HQD pattern genes via herbal enrichment analysis. Results: Our TCM-HIN2Vec strategy revealed that candidate genes associated with HQD pattern were significantly enriched in energy metabolism, signal transduction pathways, and immune processes. Moreover, we found that these candidate genes were significantly differentially expressed in the transcriptional profile of mice model with heart failure with a qi deficiency pattern. Furthermore, herbal enrichment analysis identified TCM herbs that demonstrated enriched interactions with the top 10 candidate genes and could potentially serve as drug candidates for treating HQD. Conclusion Our results suggested that TCM-HIN2Vec is capable of not only accurately identifying HQD pattern genes, but also deciphering the basis of HQD pattern. Furthermore our finding indicated that TCM-HIN2Vec may be further expanded to develop other patterns, leading to a new approach aimed at elucidating general TCM patterns and developing precision medicine.

Objective? To explore the status quo of thirst in patients receiving cardiopulmonary bypass (CPB) during fasting and water deprivation after extubation and to analyze its influencing factors. Methods? Totally 120 patients undergoing CPB in a Class Ⅲ Grade A hospital in Nanjing from December 2017 to May 2018 were selected in this prospective study. The general information questionnaire was used to collect the data, and the patients' thirst score and unstimulated salivary flow rate (USF) were measured. SPSS 22.0 was used to analyze the factors affecting postoperative thirst in the patients. Results? The incidence rate of thirst in the CPB patients 6 h post extubation was 100%, and their thirst score was (6.20±1.90); and the patients' thirst intensified over time. Univariate analysis showed that the patients' postoperative thirst was positively correlated with postoperative body temperature, respiratory rate, serum sodium concentration and urinary volume (r=0.172, 0.105, 0.209, 0.258; P＜ 0.05), and negatively correlated with postoperative fluid balance volume and USF (r=-0.222, -0.565; P＜ 0.05). Multivariate analysis showed that USF and fluid balance volume were the main influencing factors to thirst in the patients (P＜0.05). Conclusions? The incidence rate of thirst in CPB patients is relatively high and it is associated with multiple factors. Nurses should pay attention to the patients' thirst and take targeted measures to ameliorate their thirst to improve their comfort in nursing practice.

OBJECTIVETo study methylation of Id4 gene and demethylation effect of arsenic trioxide (ATO) in Raji cells.

METHODSHuman Burkitt's Raji lymphoma cells were cultared and treated with ATO at different concentrations and different time points. Methylated degree of Id4 gene was detected by methylation specificity polymerase chain reaction (MS-PCR), Id4 mRNA expression in Raji cell by reverse transcription polymerase chain reaction (RT-PCR), the growth of cell by MTT assay, and cell apoptosis and cycle distribution by Flow Cytometry (FCM).

RESULTS(1) The Id4 gene exhaustive methylation in control group, and hypermethylation in experimental group were reversed by ATO in a dose-dependent manner. (2) Id4 mRNA expression in Raji cells treated with ATO for 48 h increased gradually with ATO concentration increasing in experimental group. (3) Raji cell growth inhibited rates after different concentrations of ATO treatment for 24, 48, 72 h were 12.15% ∼ 92.17% in the experimental group (P < 0.05). (4) Apoptosis peak emerged after ATO treatment for 48 hours in experimental group, while a much lower apoptosis in control group. (5) After ATO treatment for 48 h in experiment group, the cells were arrested at G(0)/G(1) phase in a dose-dependent manner.

CONCLUSIONId4 gene presents exhaustive methylation in Raji cells. ATO can reverse the hypermethylation of Id4 gene and recover the expression of Id4 mRNA. Hypermethylation of Id4 gene is one of the reasons of Raji cells malignant proliferations.

Apoptosis ; drug effects ; Burkitt Lymphoma ; genetics ; Cell Line, Tumor ; DNA Methylation ; Humans ; RNA, Messenger ; genetics