Aim To explore the role of nicotinamide adenine dinucleotide phosphate ( NADPH ) oxidase in high glucose-induced injury in H9c2 cardiac cells. Methods H9 c2 cardiac cells were exposed to differ-ent concentrations of high glucose(5. 5 mmol·L-1 ,11 mmol · L-1 ,22 mmol · L-1 ,33 mmol · L-1 ,44 mmol ·L-1 , 55 mmol · L-1 ) for 24 h and different time pints of high glucose(0 h,12 h,24 h,36 h,48 h,72 h) . Cell viability was measured by MTT colorimetry, the protein expression of Bcl-2 , Bax and NADPH oxi-dase submits such as p22 phox , p47 phox and p67 phox were determined by western blotting. Results H9c2 cardi-ac cells exposure to high glucose for 24 h showed on decrease in cell survival and the Bcl-2 expression while an increase in the Bax expression ( P<0. 05 ) . Moreo-ver, high glucose could markedly up-regulate the activ-ity of NADPH oxidase characterized by the enhanced expression of p22 phox , p47 phox and p67 phox ( P<0. 05 ) . Conclusion Activating NADPH oxidase may play an important role in high glucose-induced injury in H9 c2 cells.

Objective To explore the immunogenicity of recombinant chimeric 6Aβ15-T including the Aβ1-15 epitope and a T-helper epitope formulated with different adjuvants and to evaluate its feasibility as a candidate vaccine for Alzheimer disease (AD).Methods The recombinant chimeric antigen 6Aβ15-T formulated with Al adjuvant, Freund′s adjuvant or MF59 adjuvant was administered to two strains of mice .The 6Aβ15-T-immunized group without adjuvants ( Mock) and non-immunized group (Control) were included in this study as control groups .The specific antibody and cellular immune response of the chimeric antigen were evaluated .Results In BALB/c strain mice, three types of adjuvants could substan-tially boost the immunogenicity of chimeric antigen 6Aβ15-T and produce a high level of specific-Aβ(β-amyloid) antibod-ies.In C57BL/6 strain mice, the existence of adjuvants enhanced the immune response of 6Aβ15-T antigen, but the mice in Mock group also produced a strong antibody response .In two strains of mice, prevalence of anti-AβIgG1, which was an indicator of Th2 polarization, was observed in the 6Aβ15-T-immunized mice.Additionally, the Al adjuvant induced a high-er level of IgG1 antibody titers, and the ratio of IgG1/IgG2a was the largest.As expected, the 6Aβ15-T antigen formulated with or without adjuvants induced PADRE-specific, but not Aβ42-specific T cellular immune response .Conclusion The 6Aβ15-T antigens formulated with different types of adjuvants could induce strong Th 2-polarized Aβ42-specific antibody re-sponses without activating self-reactive Aβ42-specific T cells in two strains of mice .The results suggested that the recombi-nant chimeric antigen 6Aβ15-T is a good candidate vaccine for AD .

To enhance the immunogenicity of DNA and adenoviral vector vaccines expressing HIV-1 subtype B gp160, human interleukin 15 (hIL15) DNA adjuvant (pVR-hIL15) was constructed. BALB/c mice received DNA prime/protein boost immunization with pVR-HIVgp160/Ad5-HIVgp160 alone or combined with pVR-hIL15. Cellular and humoral immune responses were evaluated by IFN-gamma enzyme-linked immunosorbent spot assay and enzyme-linked immunosorbent assay, respectively. Compared with those immunized with vaccines alone, the mice immunized with vaccines combined with pVR-hIL15 had significantly increased specific cellular response and antibody titer (P < 0.05). It suggests that the IL15 DNA adjuvant can enhance the immune responses induced by prime-boost regimen using DNA and adenoviral vector encoding HIV-1 subtype B gp160.
Adenoviridae ; genetics ; Adjuvants, Immunologic ; Animals ; Antibodies, Viral ; immunology ; Antibody Specificity ; Female ; Genetic Vectors ; genetics ; HIV Envelope Protein gp120 ; immunology ; HIV Envelope Protein gp160 ; genetics ; immunology ; HIV Envelope Protein gp41 ; immunology ; Humans ; Immunity, Cellular ; Immunity, Humoral ; Interleukin-15 ; genetics ; Mice ; Mice, Inbred BALB C ; Vaccines, DNA ; genetics ; immunology

The chemical investigation on Ganoderma philippii led to the isolation of sixteen compounds by silica gel and Sephadex LH-20 column chromatography. On the basis of spectroscopic data analyses, their structures were elucidated as 2, 5-dihydroxyacetophenone (1), methyl gentisate (2), (S) -dimethyl malate (3), muurola-4, 10 (14) -dien-11beta-ol (4), dihydroepicubenol (5), 5-hydroxymethylfuran carboxaldehyde (6), ergosta-7, 22E-dien-3beta-ol (7), ergosta-7, 22E-dien-3-one (8), ergosta-7, 22E-diene-2beta, 3alpha, 9alpha-triol (9), 6/beta-methoxyergo-sta-7, 22E-dien-3beta, 5alpha-diol (10), ergosta-4, 6, 8(14), 22E-tetraen-3-one (11), ergosta4, 6, 8-(14), 22E-etetraen-3beta-ol (12), 5alpha, 8alpha-epidioxy-ergosta-6, 22E-dien-3beta-ol (13), 7alpha-methoxy-5alpha, 6alpha-epoxyergosta-8-(14), 22E-dien-3beta-ol (14), ergosta-8, 22E-diene-3beta, 5alpha, 6beta, 7alpha-tetraol (15), and ergosta-5, 23-dien-3beta-ol, acetate (16). All the compounds were obtained from this fungus for the first time, and compounds 4 and 5 were isolated from the Ganoderma genus for the first time.
Ganoderma ; chemistry ; Medicine, Chinese Traditional ; Organic Chemicals ; analysis ; isolation & purification

This study is to evaluate the therapeutic effect of fibroblast growth factor 21 (FGF21) on type 2 diabetic mice model and to provide mechanistic insights into its therapeutic effect. Type 2 diabetic animal model was established with high calorie fat diet and low dose streptozotocin (STZ) injection. Mice were then randomized into 5 groups: model control, FGF21 0.25 and 0.05 μmol x kg(-1) x d(-1) groups, insulin treatment group. Ten age-matched normal KM mouse administered with saline were used as normal controls. Serum glucose, insulin, lipid products and the change of serum and liver tissue inflammation factor levels between five groups of mouse were determined. The results showed that blood glucose, insulin, free fatty acids (FFAs), triglycerides, and inflammatory factor average FGF-21 of type 2 diabetes model group and normal control group were significantly higher (P < 0.01), while compared with insulin group, no difference was significant. Average blood glucose, insulin, blood lipid and inflammatory factor of FGF-21 treatment group compared with type 2 diabetes group was significantly lower (P < 0.01) and insulin group has no difference with the model control group. The results of OGTT and HOMA-IR showed that insulin resistance state was significantly relieved in a dose-dependent manner. Thus, this study demonstrates that FGF-21 significantly remits type 2 diabetic mice model's insulin resistance state and participates in the regulation of inflammatory factor levels and type 2 diabetes metabolic disorders.
Animals ; Blood Glucose ; Diabetes Mellitus, Experimental ; drug therapy ; Diabetes Mellitus, Type 2 ; drug therapy ; Diet, High-Fat ; Fatty Acids, Nonesterified ; blood ; Fibroblast Growth Factors ; pharmacology ; Insulin ; blood ; Insulin Resistance ; Mice ; Streptozocin ; Triglycerides ; blood

Research progress ofpolysorbate 80 was summarized,including standards,chemical composition and impurity test.Assay methods of polysorbate 80 in preparations were also summarized.In addition,characteristics of the current technologies were discussed.Compositions of polysorbate 80 could be detected directly by spectrophotometry,SEC-ELSD or LC-MS methods.In addition,they could be analyzed indirectly by determing the hydrolysates with HPLC-UV or GC methods.In recent years,remarkable progress has been achieved in chemical composition determination and impurity test ofpolysorbate 80.But assay method of the excipient in related drugs need further study,due to large difference between purities ofpolysorbate 80 raw materials.

OBJECTIVE: To study the change in P wave on electrocardiogram and its diagnostic value in children and adolescents with cardioinhibitory vasovagal syncope (VVS-CI). METHODS: A total of 43 children and adolescents who were diagnosed with VVS-CI were enrolled as the VVS-CI group, and 43 healthy children and adolescents were enrolled as the control group. P wave duration and P wave voltage were measured by 12-lead electrocardiography in a basal state, and the changes were analyzed. RESULTS: Compared with the control group, the VVS-CI group had a significantly lower heart rate (P<0.05) and significantly longer P wave duration (Pwd), P wave maximum duration (Pmax), and corrected P wave maximum duration (Pcmax), as well as significantly higher P wave dispersion (Pd) and corrected P wave dispersion (Pcd) (P<0.05). Pwd, Pmax, Pd, Pcmax and Pcd had a certain diagnostic value in children and adolescents with VVS-CI (P<0.05): Pwd had a sensitivity of 69.77% and a specificity of 83.72% at the optimal cut-off value of 78.49 ms; Pmax had a sensitivity of 76.74% and a specificity of 90.70% at the optimal cut-off value of 93.39 ms; Pd had a sensitivity of 95.35% and a specificity of 69.77% at the optimal cut-off value of 27.42 ms; Pcmax had a sensitivity of 46.51% and a specificity of 88.37% at the optimal cut-off value of 120.90 ms; Pcd had a sensitivity of 83.72% and a specificity of 72.09% at the optimal cut-off value of 36.37 ms. CONCLUSIONS Children and adolescents with VVS-CI have significantly increased Pwd, Pmax, Pd, Pcmax, and Pcd, which may indicate abnormal atrial electrical activity. The cut-off value of P wave has a certain diagnostic value in VVS-CI.
Adolescent ; Child ; Electrocardiography ; Heart Rate ; Humans ; Syncope, Vasovagal

OBJECTIVE: To evaluate the validity of two abbreviated protocols (AP) of MRI in breast cancer screening of dense breast tissue. MATERIALS AND METHODS: This was a retrospective study in 356 participants with dense breast tissue and negative mammography results. The study was approved by the Nanjing Medical University Ethics Committee. Patients were imaged with a full diagnostic protocol (FDP) of MRI. Two APs (AP-1 consisting of the first post-contrast subtracted [FAST] and maximum-intensity projection [MIP] images, and AP-2 consisting of AP-1 combined with diffusion-weighted imaging [DWI]) and FDP images were analyzed separately, and the sensitivities and specificities of breast cancer detection were calculated. RESULTS: Of the 356 women, 67 lesions were detected in 67 women (18.8%) by standard MR protocol, and histological examination revealed 14 malignant lesions and 53 benign lesions. The average interpretation time of AP-1 and AP-2 were 37 seconds and 54 seconds, respectively, while the average interpretation time of the FDP was 3 minutes and 25 seconds. The sensitivities of the AP-1, AP-2, and FDP were 92.9, 100, and 100%, respectively, and the specificities of the three MR protocols were 86.5, 95.0, and 96.8%, respectively. There was no significant difference among the three MR protocols in the diagnosis of breast cancer (p > 0.05). However, the specificity of AP-1 was significantly lower than that of AP-2 (p = 0.031) and FDP (p = 0.035), while there was no difference between AP-2 and FDP (p > 0.05). CONCLUSION: The AP may be efficient in the breast cancer screening of dense breast tissue. FAST and MIP images combined with DWI of MRI are helpful to improve the specificity of breast cancer detection.
Breast Neoplasms* ; Breast* ; Diagnosis ; Ethics Committees ; Female ; Humans ; Magnetic Resonance Imaging* ; Mammography ; Mass Screening ; Retrospective Studies ; Sensitivity and Specificity ; Transcription Factor AP-1

OBJECTIVETo investigate the adaptor protein CRKL phosphorylation level (p-CRKL) and its significance in chronic myeloid leukemia (CML) treated with imatinib.

METHODSABL kinase domain was amplified by nested RT-PCR, domain point mutations analysis by direct sequencing, BCR-ABL mRNA level by real time-PCR, and p-CRKL level by flow cytometry in 52 bone marrow samples from 35 CML patients, and the relationship of p-CRKL level with ABL kinase domain mutation and with BCR-ABL mRNA level was analyzed.

RESULTSIn the 15 imatinib-resistant patients, ABL domain point mutations were detected in 6 with 4 types of nucleotide substitutions: T315I (n = 3), Y253H (n = 1), E255K and F317L. The incidence of mutations in disease chronic phase (CP), accelerated phase (AP) and blast phase (BP) was 25.00%, 40.00% and 30.00%, respectively. The BCR-ABL mRNA level in newly diagnosed CML was higher than that in imatinib-responded patients (P = 0.01); and so did in imatinib-resistant patients than in imatinib-effective patients (P = 0.03). The level of BCR-ABL mRNA was not significantly different between newly diagnosed CML and imatinib-resistant patients. p-CRKL%, MFI showed a high degree of phosphorylation in newly diagnosed CML and imatinib-resistant patients (P = 5.130; P = 3.178). The level of p-CRKL % and MFI in newly diagnosed group was higher than that in imatinib responded group (P = 0.000; P = 0.01) and also higher in imatinib-effective group than in imatinib-resistant group (P = 0.000; P = 0.02). There was a positive correlation between the level of BCR-ABL expression and p-CRKL% (and the MFI of p-CRKL) (P < 0.05).

CONCLUSIONIt seems that p-CRKL detection might be helpful in predicting imatinib treatment outcomes.

Adaptor Proteins, Signal Transducing ; metabolism ; Adult ; Antineoplastic Agents ; therapeutic use ; Benzamides ; Female ; Humans ; Imatinib Mesylate ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; drug therapy ; metabolism ; Male ; Middle Aged ; Nuclear Proteins ; metabolism ; Phosphorylation ; Piperazines ; therapeutic use ; Pyrimidines ; therapeutic use ; Young Adult

OBJECTIVETo establish a simple and practical method for screening of Env-specific monoclonal antibodies from HIV-1 infected individuals.

METHODSHuman B cells were purified by negative sorting from PBMCs and memory B cells were further enriched using anti-CD27 microbeads. Gp120 antigen labbled with biotin was incubated with memory B cells to specifically bind IgG on cells membrane. The memory B cells expressing the Env-specific antibody were harvested by magnetic beads separating, counted and diluted to the level of single cell in each PCR well that loading with catch buffer containing RNase inhibitor to get RNAs. The antibody genes were amplified by single cell RT-PCR and nested PCR, cloned into eukaryotic expression vectors and transfected into 293T cells. The binding activity of recombinant antibodies to Env were tested by ELISA.

RESULTSThree monocolonal Env-specific antibodies were isolated from one HIV-1 infected individual.

CONCLUSIONWe can obtain Env-specific antibody by biotin labbled antigen, magnetic beads separating technique coupled with single cell RT-PCR and expression cloning.

Amino Acid Sequence ; Antibodies, Monoclonal ; isolation & purification ; B-Lymphocytes ; immunology ; HIV Antibodies ; isolation & purification ; HIV Envelope Protein gp120 ; immunology ; HIV-1 ; immunology ; Immunomagnetic Separation ; methods ; Molecular Sequence Data ; Reverse Transcriptase Polymerase Chain Reaction ; methods