Objective To evaluate and compare the cellular immune response induced by live at-tenuated and inactivated hepatitis A vaccines. Methods A total of 18 healthy volunteers were enrolled in this study. They were divided randomly into 2 groups and injected by inactivated hepatitis A vaccine and at-tenuated hepatitis A vaccine, respectively. All volunteers' heparinized venous blood was separately collected for testing anti-HAV antibody and the responses of PBMC. The level of IFN-γsecreted by effect T cell was tested by ELISPOT. The percentage of lymphocytes secreting IFN-γ from CD+ and CD8+ T cell was tested by intracellular cytokine staining (ICS) technique. The concentrations of IFN-γ, IL-2, IL-10 and IL-4 in the culture supernatants(grown in the presence or absence of HAV) of the in vitro HAV-primed PBMC were determined by Luminex. Results These two types of vaccines can elicit specific anti-HAY antibodies and no statistical significance between them were observed. At the early stage after inoculation, T cell-mediated immune responses with the secretion of IFN-γ were detected in vaccines inoculated with either type of vac-cines. There was a tendency that the cellular immune responses level induced by inactivated hepatitis A vac-cine was higher than that by live attenuated hepatitis A vaccine during 1 to 3 weeks post-injection. The booster inoculation could significantly increase the level of cellular immune responses induced by the inacti-vated vaccine. Conclusion Both live attenuated and inactivated HAV vaccines can elicit an earlier specific humoral and cellular immune responses, and booster inoculation of inactivated HAV vaccine can rapidly and intensively evoke memory immune responses.

Objective: To investigate the effect of bilateral superior cervical ganglionectomy on cardiac remodeling and function in pressure-overloaded heart failure (HF) mice. Methods: Pressure-overloaded HF mouse model was produced by severe thoracic aorta banding (sTAB). Bilateral superior cervical ganglionectomy (SCGx) was performed 2 weeks after sTAB. Twenty four 6-week-old male C57BL/6 mice were randomized divided into 4 groups (n=6 each): control group: sham sTAB+sham SCGx; denervated group: sham sTAB+SCGx; HF group: sTAB+sham SCGx; denervated HF group: sTAB+SCGx. Cardiac function was measured by echocardiography at week 0, 1, 2, and 4 after sTAB, respectively. All mice were sacrificed at the end of week 4 and heart tissues were harvested. HE and Masson staining were performed. Immunohistochemical staining (IHC) for tyrosine hydroxylase (TH), adrenergic receptor β1 (AR-β1) and CD68 was performed. Western blot was used to determine the protein expression level of TH, B type natriuretic peptide (BNP), and AR-β1. Results: Left ventricular ejection fraction (LVEF) declined continuously in HF group. LVEF was similar between denervated HF group and control group at various time points (P>0.05). LVEF was significantly higher in denervated HF group than in HF group at the end of week 4 (P<0.05). HE staining showed that cross sectional cardiomyocyte area was significantly larger in HF group than in control group and denervated HF group (P<0.05), which was similar between denervated HF group and control group (P>0.05). Masson staining showed that fibrosis level was significantly lower in denervated HF group than in HF group (P<0.05). IHC showed that TH+nerves and CD68⁺ macrophages were significantly increased in HF mice as compared to control mice (P<0.05), whereas this change was abolished in denervated HF group. AR-β1 was significantly down-regulated in HF group compared with control group (P<0.05), which was not affected by denervation (P>0.05). Western blot demonstrated that the expression level of TH and BNP was significantly higher in HF group compared with the control group (P<0.05), whereas this difference was diminished in denervated HF group (P>0.05). Conclusion: Bilateral superior cervical ganglionectomy can reduce sympathetic innervation and macrophage infiltration in pressure overloaded failure heart, thus attenuate cardiac remodeling and improve cardiac function.
Animals ; Cross-Sectional Studies ; Ganglionectomy ; Heart Failure ; Male ; Mice ; Mice, Inbred C57BL ; Stroke Volume ; Ventricular Function, Left ; Ventricular Remodeling

Objective: To analyze the mutation of autoimmune regulator ( AIRE ) gene in a pedigree with autoimmune polyendocrine syndrome type Ⅰ (APS-Ⅰ). Methods: The peripheral blood samples from family members were collected for DNA extraction, and then the mutation sites on AIRE gene were screened by PCR and Sanger sequencing. The mutation sites were further verified in 100 healthy persons by the created restriction site PCR (CRS-PCR) and PCR restriction fragment length polymorphism (PCR-RFLP). The effects of mutation on the structure and function of AIRE protein were analyzed with SIFT, PolyPhen-2, Mutation Taster and Antheprot Editor softwares. The effects of mutation on the splicing sites of AIRE mRNA were predicted with Alternative Splice Site Predictor, FruitFly Splice Predictor and SplicePort softwares, and further verified by Sanger sequencing. Results: Two novel heterozygous mutations c.47 C＞G T16R and c.1631-2 A＞T were found in the proband. The c.47 site is highly conserved and homologous in different species. The missense mutation of c.47C＞G changed the secondary structure and hydrophobicity of AIRE protein, and affected its function. The c.1631 -2 A＞T mutation changed the splicing site of AIRE mRNA, and led to the deletion of exon 13. Conclusion Two novel pathogenic mutations c.47 C＞G T16R and c.1631-2 A＞T are identified in a pedigree with autoimmune polyendocrine syndrome type Ⅰ.

Based on the recently proposed Chinese ischemic stroke subclassification (CISS) system, intracranial branch atheromatous disease (BAD) is divided into large artery atherosclerosis (LAA) and penetrating artery disease (PAD). In the current retrospective analysis, we compared the general characteristics of BAD-LAA with BAD-PAD, BAD-LAA with non-BAD-LAA and BAD-PAD with non-BAD-PAD. The study included a total of 80 cases, including 45 cases of BAD and 35 cases of non-BAD. Subjects were classified using CISS system: BAD-LAA, BAD-PAD, non-BAD-LAA and non-BAD-PAD. In addition to analysis of general characteristics, the correlation between the factors and the two subtypes of BAD was evaluated. The number of cases included in the analysis was: 32 cases of BAD-LAA, 13 cases of BAD-PAD, 21 cases of non-BAD-LAA, and 14 cases of non-BAD-PAD. Diabetes mellitus affected more non-BAD-LAA patients than BAD-LAA patients (P=0.035). In comparison with non-BAD-PAD, patients with BAD-PAD were younger (P=0.040), had higher initial NIHSS score (P<0.001) and morbidity of ischemic heart disease (P=0.033). Within patients with BAD, the PAD subtype was associated with smoking (OR=0.043; P=0.011), higher low-density lipoprotein (OR=5.339; P=0.029), ischemic heart disease (OR=9.383; P=0.047) and diabetes mellitus (OR=12.59; P=0.020). It was concluded that large artery atherosclerosis was the primary mechanism of BAD. The general characteristics showed no significant differences between the CISS subtypes of LAA and PAD within BAD, as well as between the BAD and non-BAD within LAA subtype. Several differences between PAD subtypes of BAD and non-BAD were revealed.
Aged ; Brain Ischemia ; pathology ; China ; Female ; Humans ; Male ; Middle Aged ; Stroke ; pathology

OBJECTIVETo study the possible mechanism of protective effect for Baicalin on Bacillus pertussis (BP) infected brain tissue and the dose-effect relationship.

METHODSBrain tissues slices were divided into 7 groups: (1) the normal group; (2) the model group: infected by 10% BP; (3) the baicalin group, which was pretreated with baicalin, infected by BP and subdivided into 5 sub-groups according to different doses of baicalin used; (4) the glutamic acid group: cultured with glutamic acid; (5) the baicalin plus glutamic acid group; (6) the peroxide group: cultured with hydrogen peroxide; and (7) the baicalin plus peroxide group. The lactate dehydrogenase (LDH) content in the supernatant of culture was determined and quantitative protein determination was conducted.

RESULTSThe LDH releasing was higher in the model group, glutamic acid group and peroxide group as compared with that in the normal group, 15.10 +/- 4.89 u/g. protein (the same unit below), 15.49 +/- 5.66 and 16.54 +/- 5.47 vs 6.10 +/- 2.87 respectively (P < 0.01). After being pretreated with 0.25 mmol/L baicalin, LDH level decreased significantly to 8.65 +/- 2.43, which was significantly different from that in the model group (P < 0.01), LDH was also decreased in the baicalin plus glutamic acid group (9.93 +/- 2.89) and baicalin plus peroxide group (9.54 +/- 2.82), which was significantly lower than that in the glutamic acid group and the peroxide group respectively (P < 0.01).

CONCLUSIONPretreatment of baicalin has protective effect on BP caused nerve cell injury in rat brain slices, the protection is possibly related with the reduction of glutamic acid and hydrogen peroxide induced damage on nerve cells in vitro.

Animals ; Anti-Bacterial Agents ; pharmacology ; Bordetella pertussis ; Brain ; cytology ; microbiology ; Coculture Techniques ; Culture Techniques ; Dose-Response Relationship, Drug ; Female ; Flavonoids ; pharmacology ; Glutamic Acid ; pharmacology ; Hydrogen Peroxide ; pharmacology ; Male ; Neuroprotective Agents ; pharmacology ; Random Allocation ; Rats ; Rats, Sprague-Dawley

Objective To construct the eukaryotic expression recombinant PIAS3 plasmid fused with Myc protein and express the fusion protein Myc-PIAS3.Methods The full length PIAS3 fragment of 1 851bp was amplified by PCR and ligated into pMD18-T vector. The full length PIAS3 fragment was subcloned into eukaryotic pCMV-Myc vector between SalⅠ and NotⅠ sites.The recombinant pCMV-Myc-PIAS3 plasmid was trandfected into PC3 cells.The eukaryotic expression Myc-PIAS3 protein was checked by Western blotting with Myc antibody.Results The recombinant plasmid showed right sequence by the full length sequencing.The recombinant plasmid of pCMV-Myc-PIAS3 was identified by enzyme digestion.As expected,by EcoRⅠ digestion,it showed two bands of 4 357bp and 1 318 bp. By XbaⅠdigestion,it showed two bands of 3 291 bp and 2 384 bp.The sequencing result showed a N-terminal Myc of 13 amino acids followed with PIAS3 gene sequence in right reading frame.The pCMV-Myc-PIAS3 plasmid was transfected into prostate cancer PC3 cells.A specific protein expression band at relative molecular mass 68 000 was obtained by using Myc-antibody with Western blotting method.Conclusion The recombinant plasmid of pCMV-Myc-PIAS3 is sucssesefully constructed,and Myc-PIAS3 fusion protein is sucssesefully expressed.

Objective:To study the antitumor mechanism of W40,a monomer purified from Wedelia prostrate (Hook.et Arn.) Hemsl.Methods:The effects of W40 on the cell proliferative of GLC-82 cells were detected by MTT assay and colony formation assay.The migratory abilities of GLC-82 cells were observed by wound healing assay.Cell apoptosis was evaluated by Annexin V-FITC/PI staining analysis.The levels of apoptosis-relative proteins and cell proliferation-related proteins,such as Caspase-3,PARP,Stat3 and ERK,were detected by Western blotting.Results:MTF assay showed that W40 had a significant cytotoxic effect on non-small cell lung cancer GLC-82 cells.Colony formation assays showed that W40 significantly inhibited GLC-82 cells proliferation.The migration of GLC-82 cells was inhibited by W40 in a dose-dependent manner.Flow cytometry showed that the apoptotic rate increased gradually in a concentration-dependent manner.W40 down-regulated Stat3 as decreasing p-Stat3 and downstream proteins of Bcl-2 and Mcl-1.At the same time,W40 up-regulated the expression of pro-apoptotic protein Bax,and increased the cleavaged Caspase-9,Caspase-3 and PARP.W40 also down-regulated BRAF / MAPK / ERK signal pathway as decreasing p-BRAF,p-MEK and p-ERK.Conclusions:W40 induced apoptosis by inhibiting BRAF / MAPK / ERK and Stat3 signaling pathways.

BACKGROUNDTo investigate the expression and its clinical significance of MMP-2 and MMP-9 in non-small cell lung cancer (NSCLC), so as to provide reference in diagnosis, treatment and determining prognosis of NSCLC.

METHODSMMP-2 and MMP-9 expression was detected in 32 lung cancer tissues, 32 paracancerous lung tissues and 10 benign pulmonary lesion tissues by immunohistochemical method with anti-MMP-2 and anti-MMP-9 antibody.

RESULTS(1)Expression of MMP-2 (90%, 36/40) and MMP-9 (83%, 33/40) in cancer tissues was significantly higher than that in paracancerous tissues (22% and 13%) and benign pulmonary disease tissues (0) (P < 0.01). (2)Expression level of MMP-2 and MMP-9 was significantly related to lymph node metastasis and TNM staging, and histologic classification and differentiation (P < 0.05).

CONCLUSIONSMMP-2 and MMP-9 expression in NSCLC tissue is remarkably higher than that in paracancerous tissues and benign pulmonary tissues. Detaction of MMP-2 and MMP-9 expression in lung cancer tissue might be helpful to determine metastasis, staging of the cancer, and predict the prognosis of patients with NSCLC.

Objective: To explore the relationship between HCM pathogenic gene mutations and clinical phenotypes by analyzing the prenatal diagnosis and genetic characteristics of a pregnant woman from a family with hypertrophic cardiomyopathy (HCM). Methods: The clinical data of the proband and her family members was collected. The DNA was extracted from the peripheral blood, amniotic fluid cells and cultured amniotic fluid cells of proband. Next generation sequencing (NGS) was utilized for screening pathogenetic loci of the proband. The suspected mutation sequences of HCM pathogenic candidate genes MYH7 and MYBPC3 were directly sequenced after PCR. Pathogenicity prediction of amniotic fluid cells was performed by using genetic data and bioinformatics software, such as Mutation taster, PolyPhen-2 and ANTHEPROT. Results: The sequencing results showed that heterozygous mutations of MYH7 c.1988G＞A (p.Arg663His) and MYBPC3 c.151G＞A (p.Ala51Thr) were found in the proband. The phenotype of her father was normal, and no abnormal mutations were detectable. Her mother also showed normal phenotype but carried MYBPC3 c.151G＞A heterozygous mutation. Only MYH7 c.1988G＞A heterozygous mutation was found in the fetus and no abnormal variation of MYBPC3 was showed. The prediction of mutation effect and analysis of protein structure and function revealed that the two missense mutations could affect the hydrophobicity and antigenicity of the protein. The genetic data demonstrated MYH7 c.1988G＞A was defined as a pathogenic mutation. Conclusion MYH7 c.1988G＞A should be a newly generated pathogenic mutation in the proband, or caused by reproductive chimerism of her parents. MYBPC3 c.151G＞A mutation may promote the occurrence of HCM. Although the fetus only carries MYH7 c.1988G＞A, her phenotype may still display as HCM.

OBJECTIVE: To evaluate the value of Sequential Organ Failure Assessment (SOFA), Simplified Acute Physiology Score Ⅱ (SAPS-Ⅱ), Oxford Acute Severity of Illness Score (OASIS) and Logistic Organ Dysfunction System (LODS) scoring systems for predicting ICU mortality in patients with sepsis. METHODS: We collected the data of a total of 2470 cases of sepsis recorded in the MIMIC-III database from 2001 to 2012 and retrieved the scores of SOFA, SAPS-Ⅱ, OASIS and LODS of the patients within the first day of ICU admission. We compared with the score between the survivors and the non-survivors and analyzed the differences in the area under the ROC curve (AUC) of the 4 scoring systems. Binomial logistic regression was performed to compare the predictive value of the 4 scoring systems for ICU mortality of the patients. RESULTS: In the 2470 patients with sepsis, 1966 (79.6%) survived and 504 (20.4%) died in the ICU. Compared with the survivors, the non-survivors had a significantly older mean age, higher proportion of patients receiving mechanical ventilation, and higher initial lactate level, creatinine, urea nitrogen, SOFA score, SAPS-Ⅱ score, OASIS score and LODS score ( < 0.05) but with significantly lower body weight and platelet counts ( < 0.05). The AUCs of the SOFA score, SAPS-Ⅱ score, OASIS score, and LODS score were 0.729 ( < 0.001), 0.768 ( < 0.001), 0.757 ( < 0.001), and 0.739 ( < 0.001), respectively. The AUC of SAPS-Ⅱ score was significantly higher than those of SOFA score (=3.679, < 0.001) and LODS score (=3.698, < 0.001) but was comparable with that of OASIS score (=1.102, =0.271); the AUC of OASIS score was significantly higher than that of LODS score (=2.172, =0.030) and comparable with that of SOFA score (=1.709, =0.088). For predicting ICU mortality in patients without septic shock, the AUC of SAPS-Ⅱ score was 0.769 (0.743-0.793), the highest among the 4 scoring systems; in patients with septic shock, the AUCs SAPS-Ⅱ score and OASIS score, 0.768 (0.745-0.791) and 0.762 (0.738-0.785), respectively, were significantly higher than those of the other two scoring systems. Binomial logistic regression showed the corrected SOFA, SAPS-Ⅱ, and OASIS scores, but not LODS scores, were significantly correlated with ICU mortality in patients with sepsis, and their ORs were 1.08 (95% CI: 1.03-1.14, =0.001), 1.04 (95% CI: 1.02-1.05, < 0.001), 1.04 (95% CI: 1.01-1.06, =0.001), 0.96 (95% CI: 0.89-1.04, =0.350), respectively. CONCLUSIONS The scores of SOFA, SAPS-Ⅱ, OASIS, and LODS can predict ICU mortality in patients with sepsis, but SAPS-Ⅱ and OASIS scores have better predictive value than SOFA and LODS scores.
Humans ; Intensive Care Units ; Prognosis ; ROC Curve ; Retrospective Studies ; Sepsis