To evaluate the safety of the therapeutic DNA vaccine against the hepatitis B virus. Plasmid DNA was detected by PCR in tissues of BALB/c mice immunized with the plasmids combined with electrotransfer in doses of 10 and 50?g. In long term toxicity study, HBV DNA vaccine was administered by repeated intramuscular injections combined with electrotransfer of pDNA to NIH mice in doses of 30, 60 and 120?g, twice a week for four consecutive weeks. Morbid manifestations, behaviors, hematology, blood chemistry, anti nuclear antibodies, and histopathology were analyzed. Results showed that plasmid DNA was detected primarily in the muscle at the site of injection, where it remained for up to 8 weeks. Eight repeated intramuscular injections of HBV DNA vaccine showed no adverse effects on behaviors, hematology, blood chemistry, and histopathology. No evidences of autoimmune mediated pathology and anti nuclear antibodies were observed in the mice. These results suggested that the therapeutic HBV DNA vaccine was safe and well tolerated.

Objective To study the prognostic value of procalcitonin (PCT) level in the outcome of patients with paraquat poisoning (PQ).Methods The clinical data of 128 patients with acute PQ admitted to emergency department were collected from March 2013 through March 2014.The patients were divided into two groups:the death group and the survival group (survival of 28 days).Poisoning doses,urine concentration of PQ,time elapsed from poisoning to admission,and time elapsed from poisoning to gastrolavage were documented.And on the 1 st day,the 3rd day and the 7th day after poisoning,serum PCT were detected.The level of PCT was used to investigate the prognostic values in patients with acute PQ in the death group and survival group.Results Of 128 cases,72 (56.3％) survived and 56 died in 28 days.Among them,the level of PCT increased to some extent in the first day in 90 cases,and 48 patients died.According to trend analysis,the levels of PCT in death group on the 1st day,the 3rd day and the 7th day after PQ were significantly higher than those in survival group [ld:(0.96 ±0.13) vs.(0.08 ±0.01),3d:(1.12 ±0.14) vs.(0.28 ±0.05),7d:(1.22 ±0.14) vs.(0.20 ±0.03),P ＜0.01].There was a trend of escalating PCT levels in death group,whereas the PCT level reached the peak on the 3st day and decreased gradually in the following days in survival group.The early PCT level was obviously related to poisoning doses,urine concentration,CRP,WBC,ALT,CR (the coefficient of association were 0.794,0.723,0.724,0.332,0.700,0.414,respectively,P＜0.01).Conclusions The serum level of PCT increased in patients with acute PQ was significantly positively correlated with the oral dose and urine concentration of paraquat,and it can be used as an indicator for PQ severity.There is important clinical significance in detecting the change of serum level of PCT for estimating the condition of patients and evaluating the prognosis.

Objective To explore the role of microRNA-206 (miR-206) in peripheral blood mononuclear cells (PBMCs) in infantile bronchiolitis caused by respiratory syncytial virus (RSV).Methods Thirty-five cases of infantile bronchiolitis and 25 cases of healthy controls were enrolled into the current study.PBMCs were isolated from the peripheral blood of both healthy subjects and those with infantile bronchiolitis in the acute and the convalescent stages.Total RNAs were extracted from PBMCs which were stimulated by phorbol-12-myristate-13-acetate (PMA) and Ionomycin, and then the RNA was transcribed reversely into cDNA.The expressions of miR-206 and Kruppel-like transcription factor 4 (KLF4) were detected by real-time quantitative polymerase chain reaction (qRT-PCR) method.Plasma interleukin-17 (IL-17) was determined by enzyme linked immunosorbent assay (ELISA).Results There was a significant difference in miR-206 levels of children with RSV bronchiolitis in the acute stage(0.055 ±0.018) and the convalescent stage(0.187 ±0.069) as well as the healthy controls(0.204 ± 0.075).Through pairwise comparison, the miR-206 levels in the children in the acute stage were significantly lower than those in the convalescent stage and healthy control group (P ＜ 0.01), but no statistical significance was found between the convalescent stage group and healthy control group(P ＞ 0.05).The levels of KLF4 mRNA of children in the acute stage,convalescent stage as well as the healthy subjects were 0.588 ± 0.161,0.086±0.024,0.075 ±0.019, respectively,which was significantly difference (P ＜ 0.01).The levels of IL-17 were (58.26 ±25.88) ng/L, (9.87 ± 3.01) ng/L, (7.65 ± 2.16) ng/L, respectively (P ＜ 0.01).Compared to the convalescent and the normal control group,both the KLF4 mRNA and IL-17 levels were markedly higher in the acute stage (P ＜ 0.01), but there were no significant differences between children with RSV bronchiolitis in convalescent stage and in the healthy controls (P ＞ 0.05).Furthermore, the result of this study showed a negative correlation between the expression of miR-206 and KLF4(r =-0.624 ,P ＜0.01)and IL-17 (r =-0.609 ,P ＜0.01) in children in the acute stage and a positive correlation between KLF4 mRNA and IL-17 in children in the acute stage (r =0.662, P ＜ 0.01).Conclusion The levels of miR-206 may play a role in the onset of RSV associated post-bronchiolitis (PB) and the low expression of miR-206 in children infected with RSV may increase the susceptibility to PB.

Objective:To establish an HPLC method for determining nectandrin B in Uygur medicine Arillus Myristicae. Meth-ods:The analysis was performed on a SunFireTM C18 column (150 mm × 4. 6 mm, 5 μm) at 30°C using methanol /water (52 ∶48) as the mobile phase at a flow rate of 1. 0 ml·min-1 with the UV detection wavelength at 228. 4 nm. Results:The linear range of nectan-drin B was 3. 98 -79. 60 μg·ml-1(r=0. 999 2). The average recovery was 99. 89% with RSD of 1. 11%(n=6). Conclusion:The method is accurate, convenient and reproducible in the determination of nectandrin B in Uygur medicine Arillus Myristicae.

The web-based PBL teaching plan in pediatrics embodies the network learning environment and learning requirements.The basic design unit is a complete real case with certain width and depth,which can stimulate the students' interests of participation and guide students into a predetermined learning area to achieve desired learning objectives.The article dwelled on the compilation and implementation of the teaching plan of children bronchopneumonia.The first part:teachers showed the case in PBL site and proposed the questions.Students made the Powerpoint after studying and discussing in groups by PBL teaching website,QQ group,SMS platform,etc.The second part:students reported and discussed in groups; the teacher provided the new information and put forward the following-up problems; students consulted the documents and materials through the network again; finally teachers made comments on the results.During the teaching,network should be fully utilized and the questions should be proposed progressively at different levels.Teacher should play a guiding role and emphasize the students' autonomous learning.

Zebrafish toxicity assessment system is one of the important vertebrate model systems. Zebrafish is playing an increas-ingly important role in the field of toxicology studies because of its small size, short generation cycle, the transparent embryo and high reproductive rate. Now it is widely used in the embryonic derelopmental toxicology, pathological toxicology, environmental toxicology and other areas of toxicology studies with its unique advantages.

Objective To explore the therapeutic effect of olmesartan (OLM) on acute lung injury induced by paraquat (PQ) in rats in order to study its action mechanism.Methods A total of 70 Wister rats wererandomly (random number) divided into 5 groups,namely control group (C group,n =10),poisoning group (PQ group,n =15),rats treated by OLM with low dose (LD group,n =15),moderate dose (MD group,n=15) and high dose (HD group,n =15).PQ (80 mg/kg) was administered by gavage route in PQ group and in OLM groups for paraquat poisoning modelling,while in C group,equivalent normal saline was given instead.The OLM was administered by gastric instillation in OLM treatment groups (LD group:5 mg/kg; MD group:10 ng/kg; HD group:15 mg/kg) 6 hours after paraquat gavage and once a day for 7 days,while in C group and PQ group,normal saline was used instead.All rats were sacrificed 12 hours after the last dose treatment.The levels of glutathione peroxidase (GSH-Px,energy units),superoxide dismutase (SOD,U/mg pro),malondialdehyde (MDA,nmol/mg pro) in lung tissue,and the levels of serum transforming growth factor beta-1 (TGF-β1,pg/mL) and pH,oxygen partial pressure (PaO2) and bicarbonate ions concentration (HCO3-) were determined.Further,the lung coefficient and lung fibrous tissue hyperplasia grading were calculated.Correlation analysis was carried out to explore the correlation among GSH-Px,SOD,MDA,lung coefficient,lung hyperplasia of fibrous tissue and TGF-β1.The lung tissue were prepared for microscopy observation after Hematoxylin-eosin staining method (HE staining) as well.The difference between groups was compared by one-way analysis of variance,and correlation analysis carried out by using Pearson and Spearman rank correlation coefficient.Results The levels of GSH-Px and SOD in lung tissue of PQ and OLM treatment groups were significantly lower than those in C group,while in OLM treatment groups,those were higher than those of PQ group,and the HD group showed most obvious (all P ＜ 0.05).The level of MDA in lung tissue in PQ and OLM treatment groups were significantly highcr than that in C group while in OLM treatment groups,that was lower than that in PQ group,and the HD group showed most obvious (all P ＜ 0.05),and there were no differences between the LD group and MD group (all P ＞ 0.05).The lung coefficient and lung fibrous tissue hyperplasia grading in PQ and OLM treatment groups were significantly higher than those in C group,while in OLM treatment groups,those were lower than those in PQ group,and the HD group showed most obvious (all P ＜ 0.05).The level of serum TGF-β1 in PQ and OLM treatment groups were significantly higher than that inC group,while in OLM treatment groups,that was lower than that in PQ group,and the HD group showed most evident (all P ＜ 0.05),and there were no differences between the LD group and MD group (all P ＞0.05).The pH,PaO2 and HCO3-in PQ and OLM treatment groups were significantly lower than those in C group,while difference between LD and HD groups was also statistical significance (all P ＜ 0.05),while there were no differences between the PQ group and LD group as well as the LD group and MD group (all P ＞ 0.05).The correlation analysis showed GSH-Px and SOD had negative correlation with TGF-β1 [the correlation coefficient (r) were respectively-0.860 and-0.856,all P＜0.05],while MDA,lung coefficient and lung fibrous tissue hyperplasia grading had positive correlation with TGF-β1 (r were respectively 0.800,0.830 and 0.656,all P ＜ 0.05).Lung tissue section showed the degree of alveolar septa widened,alveolar collapse and inflammatory cells infiltration in OLM treatment groups were milder than those in PQ group,and the mildest in HD group.Conclusions OLM can attenuate the pulmonary edema and pulmonary fibrosis caused by paraquat poisoning and maybe it is associated with reducing the expression of TGF-β1 and inhibiting oxidative stress reaction.

Objective:To explore the effect of microRNA (miRNA)-3126-5p on the proliferation and migration of colorectal cancer cells by inhibiting the expression of LIM and SH3 protein 1 ( LASP1). Methods:Real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression levels of miR-3126-5p in colorectal cancer cell lines (HT-29, HCT116, LoVo, SW480) and normal intestinal mucosal epithelial cells (HIEC). The cell line with the lowest expression level was selected as the experimental object. The experiment was divided into 2 groups: the negative control group (transfected with miR-NC) and the miR-3126-5p group (transfected with miR-3126-5p). Cells of each group were collected 48h after transfection. qRT-PCR method was used to detect the expression level of miR-3126-5p in each group. The MTS method and the scratch healing experiment were used to detect the proliferation level and migration ability of the cells in each group. The bioinformatics software microRNA.org and the dual-luciferase reporter gene experiment were used to predict and verify the target genes of miR-3126-5p, respectively. qRT-PCR and Western blot were used to detect the expression levels of target genes in each group of cells. Measurement data were expressed as mean±standard deviation ( Mean± SD), t test was used for comparison between two groups, and one-way analysis of variance was used for comparison between multiple groups. Results:Compared with normal intestinal mucosal epithelial cells (HIEC), the expression level of colorectal cancer cell line miR-3126-5p was significantly reduced ( P<0.05), and the cell line with the lowest expression level was HCT116 cells ( P<0.01). The expression of miR-3126-5p in HCT116 cells in the negative control group and miR-3126-5p group were (1.05±0.16) and (7.91±1.26) respectively, and the difference was statistically significant ( t=5.40, P<0.01). Compared with the negative control group, the proliferation ability of HCT116 cells in the miR-3126-5p group was significantly reduced ( t=4.52, P<0.05), and the migration ability was significantly reduced ( P<0.01). microRNA.org shows that miR-3126-5p has complementary binding sites with LIM and SH3 protein 1 ( LASP1) gene mRNA. miR-3126-5p can target LASP1 mRNA ( P<0.01). Compared with the negative control group, the expression of LASP1 gene in HCT116 cells of the miR-3126-5p group was significantly reduced ( t=4.56, P<0.01). Conclusion:The expression of miR-3126-5p in colorectal cancer cell lines is low, and miR-3126-5p can reduce the proliferation and migration ability of colorectal cancer HCT116 cells by inhibiting the expression of the target gene LASP1.

Objective:To investigate the effect of long non-coding RNA (lncRNA) HAGLR on the proliferation and migration of gastric cancer cells by inhibiting the expression of microRNA (miRNA, miR)-93-5p.Methods:Real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of HAGLR in gastric cancer cell lines (HS-746T, BGC823, SGC7901, MGC803) and normal gastric mucosal epithelial cells (GES-1). Selected the cell line with the lowest HAGLR expression and transfected with the negative control plasmid (negative control group) or HAGLR-high-expression plasmid (HAGLR group) respectively. The MTS method and the scratch healing test were used to detect the proliferation and migration ability of the cells after transfection. The bioinformatics software miRcode database was used to predict the target gene of HAGLR, and the dual luciferase reporter gene experiment was used to verify the binding of HAGLR to the target gene. qRT-PCR was used to detect the expression of the target gene. Western blot was used to detect the expression of Hippo signaling pathway. The software SPSS 21.0 was used to conduct statistical analysis. The t test was used for comparison between two groups, and the one-way analysis of variance was used for comparison between multiple groups. Results:Compared with GES-1 cells, the expression level of HAGLR in gastric cancer cell lines was lower (all P<0.05), and the cell line with the lowest HAGLR expression was SGC7901 cells ( P<0.01). The HAGLR expression in SGC7901 cells in the HAGLR group and the negative control group were 1.03±0.13 and 9.75±1.10, respectively. The expression level of HAGLR in the negative control group was significantly lower than that in the HAGLR group ( t=7.87, P<0.01). Compared with the negative control group, the absorbance of SGC7901 cells in the HAGLR group was significantly reduced ( P<0.05), and the scratch healing rate was significantly reduced ( P<0.01). The miRcode database showd that HAGLR and miR-93-5p have complementary binding sites. The dual luciferase reporter gene experiment showed that HAGLR can complement miR-93-5p ( P<0.01). Compared with the negative control group, the expression of miR-93-5p in SGC7901 cells in the HAGLR group was significantly reduced ( P<0.01), and the expression of Hippo signaling pathway protein was significantly reduced (all P<0.01). Conclusions:HAGLR is low expressed in gastric cancer cell lines. HAGLR inhibits the proliferation and migration of gastric cancer SGC7901 cells by negatively regulating miR-93-5p.

Objective To investigate early diagnosis and treatment of Paget disease of breast. Methods The clinicopathological data of 26 cases of breast Paget disease in the First Affiliated Hospital of Wenzhou Medical University from January 1995 to June 2018 was analyzed. Results The main clinical manifestation of 26 patients was eczema like papillae in 19 cases (73.1%), and associated with nipple discharge in 9 cases (34.6% ) and breast mass in 6 cases (23.1% ). The diagnosis of this disease was based on curettage cytology in 4 cases (15.4% ), biopsy in 8 cases (30.8% ), and needle aspiration cytology or post resection pathological examination in 14 cases (53.8% ). Paget disease consisted of simple papillary Paget disease in 3 cases (11.5%), ductal carcinoma in 18 cases (69.2%) and invasive ductal carcinoma in 5 cases (19.2%). Pathological TNM staging was 0 stage in 19 cases (73.1%), stageⅠin 3 cases (11.5%), stageⅡin 2 cases (7.7%), stageⅢin 2 cases (7.7%) and no stage inⅣcase. In this group, 2 cases underwent radical mastectomy, 18 cases underwent modified radical mastectomy, 4 cases underwent simple mastectomy, and 2 cases underwent mastectomy combined with low axillary lymph node dissection and intraoperative rapid frozen examination. Fourteen patients received adjuvant chemotherapy with CEF(cyclophosphamide + pharmorubicin + tegafur)/EC (pharmorubicin + cyclophosphamide) + T(docetaxel) or TP(docetaxel + cis- platinum)regiment after operation, 2 cases were treated with trastuzumab targeted therapy and 5 cases with adjuvant radiotherapy. Twenty-five of 26 patients were followed up for 8-108 months except one patient lost in follow-up. The 5-year survival rate was 96.0% (24/25), and the 10-year survival rate was 52.0% (13/25). Conclusions The diagnosis of Paget disease of the breast depends on cytology or pathology, and multidisciplinary treatment based on surgery is judged of by tumor stage and coincident other types of breast cancer and axillary lymph node involvement.