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Objective:To construct a bivalent DNA vaccine against SARS-CoV-2 and influenza A virus H3N2 and to evaluate its immunogenicity in mice.Methods:The coding sequences for spike 1 (S1) protein of SARS-CoV-2 Beta variant and hemagglutinin (HA) of influenza A virus Cambodia (H3N2) strain were codon-optimized and synthesized. The two coding genes were ligated by the self-cleaving 2A peptide using over-lapping PCR to construct S1-2A-HA fragment, which was inserted into pVRC vector to construct the bivalent DNA vaccine, named as pVRC-S1-2A-HA. Indirect immunofluorescence assay (IFA) and Western blot were performed to detect the expression of S1 and HA proteins. BALB/c mice were immunized with pVRC-S1-2A-HA by intramuscular injection and electroporation. The humoral immune responses induced in mice were detected by indirect ELISA, pseudovirus neutralization assay and hemagglutination inhibition assay. Cellular immune responses were detected by IFN-γ ELISPOT, intracellular cytokine staining (ICS) and cytometric bead array (CBA).Results:The bivalent DNA vaccine pVRC-S1-2A-HA could express S1 and HA proteins in vitro. Specific cellular immune responses against S1 protein and specific IgG antibody against HA protein were significantly induced in mice with single-dose immunization. The antigen-specific immunity was significantly enhanced after booster immunization. The geometric mean titer (GMT) of specific IgG antibody increased to 3 251 for S1 protein and 45 407 for HA protein after two-dose immunization. Moreover, the S1-specific T cells increased to 1 238 SFC/10 6 cells. ICS results indicated that the booster vaccination induced CD4 + T and CD8 + T cells to produce IL-2, IFN-γ and TNF-α in mice. The secretion of various cytokines including IL-2, IL- 4, IL-6, IL-10 and IFN-γ in mouse splenocytes was induced after single-dose immunization. Conclusions:A bivalent DNA vaccine against SARS-CoV-2 and influenza A virus H3N2 was constructed and could induce S1- and HA-specific humoral and cellular immune responses in mice, suggesting the great potential of it for further development and application.
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Objective@#To determine the first dengue fever case in Taizhou and trace probable transmission sources.@*Methods@#Collected serum of three patients for antigen, antibody and nucleic acid detection. Dengue viruses were isolated and cultured in C6/36 cell. The whole length of E gene was amplified by reverse transcriptase-polymerase chain reaction(RT-PCR) and then sequenced. The phylogenetic tree was drawn.@*Results@#Three cases were positive in nucleic acid detection. Two cases were IgM positive. One case was NSI antigen postive. Three strains of type I dengue virus were isolated from samples. The phylogenetic trees shown that the three strains were on the same branch. The identities of nucleotide were 99.87%. The identities of amino acid were 99.6%-99.8%.@*Conclusions@#The dengue virus strains isolated in Taizhou was imported from Guangdong or Southeast Asia and caused location infection.
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Objective To explore the expression of microRNA-1285 (miR-1285) in ovarian cancer tissues,and to analyse the effects of miR-1285 expression on proliferation and apoptosis of human ovarian cancer cell lines in vitro.Methods The expressions of miR-1285 in cancer tissues and corresponding adjacent tissues from patients with ovarian cancer were detected by using fluorescence quantitative PCR.At the same time,human ovarian cancer cell lines,including SKOV3 and OVCAR3,were transfected with miR-1285 mimics in vitro,the proliferation activity of cells successfully transfected with miR-1285 was detected by MTT assay,and the apoptotic cells were detected by flow cytometry.Results The results of fluorescence quantitative PCR showed that,for patients with ovarian cancer,the expression level of miR-1285 in ovarian cancer tissues was lower than that in the corresponding adjacent tissues,the difference was statistically significant (P＜0.05).The results of experiment in vitro showed that compared with the control group,the proliferation activity of SKOV3 cells transfected with miR-1285 mimics was decreased,and the rate of apoptosis was significantly increased,there were statistically significant differences (P＜0.05);similarly,compared with the control group,the proliferation activity of OVCAR3 cells transfected with miR-1285 mimics was decreased,and the rate of apoptosis was significantly increased,there were statistically significant differences (P＜0.05).Conclusion The expression level of miR-1285 in tumor tissues of patients with ovarian cancer is decreased,which indicates that miR-1285 may be involved in the occurance and progression of ovarian cancer;the experiment in vitro shows that miR-1285 may be closely related to the proliferation and apoptosis of ovarian cancer cells.
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Objective To explore the effects of aerobic exercise on 6-minute walk distance and serum level of omentin-1 in maintenance hemodialysis (MHD) patients. Methods A total of 89 MHD patients (MHD group) in the 5th Central Hospital of Tianjin were included in the study from December 2013 to November 2014. Patients were randomly divided into two groups:routine treatment group (n=43) and aerobic exercise treatment group (aerobic exercise group, n=46). Fifty healthy subjects were selected as normal control group. Both groups received the same hemodialysis and regular treatments, and the aerobic exercise group received 6-month interdialytic aerobic exercise, and the routine group did not take any exercise intervention. The 6-minute walk distances (6MWDs),body mass,height and blood pressure were recorded, and serum levels of omentin-1, C-reactive protein (CRP), blood routine test and biochemistry test were determined in two groups. The relationship between 6 MWDs, age, CRP, left ventricular ejection fraction (LVEF) and omentin-1 were analyzed. Results The serum levels of omentin-1 and CRP weresignificantly higher, LVEF was significantly lower, in MHD group than those of healthy control group (P<0.01). After 6 months of aerobic exercise, 6MWDs, LVEF, hemoglobin and serum level of omentin-1 were significantly increased than those before treatment in aerobic exercise group. The 6MWDs, LVEF and hemoglobin were significantly increased after treatment than routine group. The levels of CRP, fasting glucose and blood pressure were significant decreased after treatment. The levels of CRP, fasting glucose and systolic blood pressure were decreased in treatment group than those of routine group. Before treatment, 6MWDs was negatively related with age and CRP, and positively related with LVEF and serum level of omentin-1 in MHD patients (r=-0.418,-0.229, 0.252 and 0.234, P<0.05). Conclusion Aerobic exercise can significantly increase values of 6MWDs, LVEF, hemoglobin and serum level of omentin-1, and reduce values of CRP, fasting blood glucose and systolic blood pressure in MHD patients. Aerobic exercise can improve physical performance,heart function and micro inflammatory state, thereby reduce cardiovascular events and mortality risk in MHD patients.
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Hemagglutinin (HA) contains a head domain with a high degree of variability and a relatively conserved stem region. HA is the major viral antigen on the surface of the influenza virus. To define the biologic activities of chimeric HA bearing different head domains and stem regions or their potential use, a HA chimeric gene containing the head domain of the H7 subtype virus and stem region of the H3 subtype virus was modified and expressed using a baculovirus expression vector. Then, the secreted protein was purified and its biologic activities characterized. Approximately 1.4 mg/mL cH7/3 HA could be obtained, and its molecular weight was ≈ 70 kD. The trimer form of cH7/3 protein had hemagglutination activity and could be recognized by specific antibodies. The method described here can be used for further studies on the screening of HA stem-reactive antibodies or the development of vaccines with conserved epitopes.
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Humans , Antibodies, Viral , Allergy and Immunology , Baculoviridae , Genetics , Metabolism , Gene Expression , Genetic Vectors , Genetics , Metabolism , Hemagglutination , Hemagglutinin Glycoproteins, Influenza Virus , Genetics , Allergy and Immunology , Influenza Vaccines , Genetics , Allergy and Immunology , Influenza, Human , VirologyABSTRACT
Objective To evaluate the effect of clinic-pathological variables on prognosis and recurrence of the aged women suffered from borderline epithelial ovarian tumors. Methods The clinical and follow-up data of 102 cases were analyzed retrospectively. Univariate and multivariate logistic regression analysis were used to assess independent prognosis and recurrence factors.Results The postoperative accessory treatment, peritoneal implantation and lymph nodes metastasis could influence the 5-year survival rate(χ2=19.311, 11.054, 18. 559, all P＜0.01). The surgery procedure, peritoneal implantation, lymph nodes metastasis and microinvasion could influence the recurrence rate(χ2=23.606, 23.583, 25.774, 23.583, all P＜0.01). The logistic regression model showed that the postoperative accessory treatment and peritoneal implantation were the two independent prognosis factors (OR=0.060, 35.319, all P＜0.01). The surgery procedure,peritoneal implantation and microinvasion were the three independent factors affecting recurrence(OR =0.001, P＜0.05;OR=60.952, P＜0.05;OR=88.874, P＜0.01). Conclusions The early standard treatment on borderline epithelial ovarian tumors of the aged women have an excellent prognosis, the risk of recurrence remains in some patients. We emphasize that these patients with highrisk factors should be main surveillance population.
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Objective To investigate the distribution of different Human papilloma virus(HPV) subtype infection of women cervixes in parts of Guizhou Province and the high risk factors of HPV infection,in order to offer some evidences for the use of vaccine of cervix eaneers in the region, and offer a group of reliable data for the screen-ing statistics of cervix eaneer in the world. Methods A total of 2339 women having sexual life were checked for 21 HPV genotypes by DNA flow-through hybridization technique and liquid-hased pap test(LPT) in their exfoliated cer-vical cells. Cervix tissues were taken in some HPV positive women and (or) LPT positive women among them for bi-opsy by the vaginoscope. The high risk factors of HPV infection and the distribution feature of HPV subtype infection in the different cervix diseases were anlayzed. Results The total positive rate of HPV infection was 30.31% (709/ 2339 ) in the 2339 women. There were 20 HPV subtypes to be detected. HPV subtype infection from high to low were HPVI6,52,18,58 and 11. The rates of HPV subtype infection were 9.87% (231/2339) ,5.64% ( 132/2339), 4.95% (116/2339 ) ,4.19% (98/2339)and 2.99 % (70/2339). The peak age of HPV infection ranged from 31 to 35 years old. The earlier the first sexual life,the higher HPV infeetion (tendeney X2 test,P <0.05). 366 CIN pa-tients and 181 patients of invasive cervix carcinoma( ICC)were diagnosed through pathology examination. The rate of HPV infection were 34.26% (74/216)in CIN Ⅰ ,77.78% (56/72)in CIN Ⅱ ,82.05% (64/78)in CIN Ⅲ. Among the 181 ICC,the rate of HPV infection was 84.52% (131/155)in squamous carcinoma of the cervix (SCC) and 38.46% (10/26) in adenocareinoma of uterine cervix (AUC). The infeetion rate of HPV16 was 61.83% ( 81/131 ) in the SCC of HPV positive. The infection rate of HPV18 was 60% (6/10)of the AUC of HPV positive. HPV16 was related closely with SCC ( X2 = 11. 948 ,P < 0.001 , ORSCC/AUC = 5. 946). There was no significant difference be-tween HPV 18 in SCC and AUC( P > 0.05). Conclusions The major HPV subtypes are 16,52,18,58 in Guizhou province. There is a close correlation between HPV infection and cervix diseases. HPV infection often occurs in young and middle-aged women. The first sexual life is a high factor of HPV infection. High risk HPV infecting would cause cervix cancer. CIN and cervix cancers are caused by high risk HPV infection. HPV16 is related closely with SCC. There is no significant difference between HPV18 in SCC and AUC. HPV16/18 vaccine could be considered to re-duce the incidence of cervical carcinoma in Guizhou province.
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Objective To observe the effect of compound rehmannia on SOD and ATP enzyme in the brain of SAM-P/8 dementia mice and the change of apoptosis neuron cells.Methods SAM-P/8 mice were divided randomly into three groups:experimental group,treatment group and model group.Mice in experimental group were lavaged by compound rehmannia(2ml/day,individual).Mice in treatment group were lavaged by donepezil hydrochloride(400mg/kg,individual),and mice in normal group and model contrast group were lavaged by equal sodium chloride.After 3 weeks,we observed mice′s behavior in every group and investigated the changes of SOD,Na+-K+-ATP enzyme by immunohistochemistry and investigated the changes of apoptosis cells peak by FCM.Results Compound rehmannia could improve learning and memory ability of the SAM-P/8 mice,strengthen SOD,Na+-K+-ATP activity and reduce apoptosis neurons in brain.Conclusion Compound rehmannia can significantly improve the Alzheimer's mice learning and memory ability,strengthen the energy of brain cells,and increase resistance to free radical antioxidant capacity,reduce the neuron apoptosis and has a good relatively protection and care to SAM-P/8 mice brain neurons.
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BACKGROUND: Frenulum of prepuce of penis contained many nerve terminals is an extremely sensitive region. If the frenulum is injured in circumcision or other operations, the complication, such as postoperative spontaneous pain of penis, sexual disturbance and so on, will occur. But there still is no define explanation for this up to now.OBJECTIVE: To observe the distribution of immunoreactive nerve terminal of calcitonin gene-related peptide (CGRP) in prepuce of penis and frenulum of prepuce of adult SD rats, and look for the source of CGRP immunoreactive nerve terminal in frenulum of prepuce.DESIGN: A single sample trial.SETTING:Department of Anatomy, School of Medicine, Zhejiang University.MATERIALS: The experiment was performed at Department of Anatomy,School of Medicine. Zhejiang University from September 2004 to May 2005. A total of 20 adult male SD rats were selected, and were raised in warm, quiet, photophygous environment for 1 week before the trial so as to make the rats fit for the environment and maintain their basal state.METHODS: The rats were assigned randomly into 2 groups. Ten rats in the first group were treated with the immunohistochemical method to observe the distribution of CGRP immunoreactive nerve terminal in prepuce of penis and frenulum of prepuce of adult rats. Ten rats in the second group were treated with fluorogold (FG) retrograde labeled combined with CGRP immunofluorescence labeled method to look for the source of CGRP immunoreactive nerve terminal in frenulum of prepuce of penis. MAIN OUTCOME MEASURES: ①The morphology and distribution of CGRP immunoreactive nerve terminal in prepuee of penis and frenulum of prepuce of adult SD rats were observed under light microscope. ②The distributive density and difference of CGRP immunoreactive nerve terminal in prepuce of penis and frenulum of prepuce were detected and compared (represented by A). ③Morphology and distribution of FG retrograde labeled -positive, CGRP single-labeled positive and FG/CGRP double-labeled positive neurons in dorsal root ganglion were observe under fluorescence microscope. ④Mean quantity of FG retrograde labeled positive, CGRP single abeled positive and FG/CGRP double-labeled positive neurons in dorsal root ganglion was counted.RESULTS: Totally 20 rats were involved in the analysis of results. ① Amber-coloured CGRP immunoreactive nerve terminal appeared in prepuce of penis and frenulum of prepuce of adult rats. These nerve terminal mainly occurred in basal layer of epidermis and papillary layer of dermis, distributed as twig shape or intestiniform; mostly of them were bundled, different in length, and some of them showed enlarged nodosity. ②The distributive density of CGRP immunoreactive nerve terminal in frenulum of prepuce of penis was significantly larger than that in prepuce of penis (2.15±0.32, 1.02±0.22,t =-2.03,P＜0.01). ③Combined with the FG retrograde labeled method it was found that these nerve terminal was derived from neurons of dorsal root ganglion opposed to the sixth lumbar spinal cord and the neurons of dorsal root ganglion opposed to the first acral spinal cord. FG retrograde labeled positive neurons differed in length. The cell body showed round or orbicular-ovate, without obvious prominence. Bright inaurate fine particle appeared in cytoplasm, no label in nucleus. Most cells arranged in line along nerve tract or diffusedly distributed. Most CGRP single-labeled positive neurons were middle or small cells found by CGRP immunofluorescence labeling. Dyeing was too dark.Reaction product distributed evenly in cytoplasm, which showed bright dark green (FITC labeled color). The same positive section was observed comparatively under different excitation light. It was found that FG/CGRP double-labeled positive cells were middle or small, and its amount accounted for a half of the total number of FG retrograde positive cells.CONCLUSION: CGRP may participate the transmission of sensory information in prepuce of penis and frenulum of prepuce of rats. The CGRP immunoreactive nerve terminal in frenulum of prepuce of penis of rats is sourced from neurons of dorsal root ganglion opposed to the sixth lumbar spinal cord and the neurons of dorsal root ganglion opposed to the first sacral spinal cord.
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Objective To study the effects of hypoxia in uterus on nNOS expression in postnatal mouse parietal cortex and the learning and memory ability of adult mouse. Methods The number and shape of parietal cortex neurons during postnatal development with hypotonic hypoxia in uterus were observed by Nissl staining.The expression of nNOS in parietal cortex neurons during postnatal development with hypoxia in the uterus was studied with immunohistochmistry.The ability to learn and memorize in adult mouse was observed by Morris water maze. Results Compared with the control group,the number of neurons in parietal cortex from mouse during postnatal development in hypoxic group reduced significantly(P