Objective To develop a simple high-resolution melting ( HRM) analysis method for differentiation of Pc and P2 variants in class 1 integron.Methods DNA fragments containing Pc and P2 variants were amplified from plasmids pACW ( PcW ) and pACWP2 ( PcW-P2 ) respectively , then these purified PCR products and P 2 promoters were analyed full-length amplicon by HRM .Eight DNA fragments containing different Pc promoters were amplified and site-specific mutated from plasmids pACS ( PcS ) , pACH2 ( PcH2 ) , pACH1 ( PcH1 ) , pACW ( PcW ) , genomic DNA of Klebsiellar pneumonia HS07-68 (PcWTGN-10)and HS05-1792(PcH2TGN-10)respectively.The purified PCR products and eight Pc variants were characterized by HRM analyses of an unlabeled probe and full-length amplicon.This assay was applied to the differentiate Pc and P2 variants in 109 class 1 integrons from 95 urine clinical Escherichia coli isolates in Huashan Hospital during 2004 -2007.The differentiation results were compared with that determined by direct sequencing .Results P2 promoter with a significant higher melting temperature ( Tm ) can be identified by HRM analysis clearly .P2 promoters were identified in 2 class 1 integrons and consistent with direct sequencing results .Eight Pc variants were classified into three groups: PcS, PcSTGN-10 , PcW, PcWTGN-10, PcH1, PcH1TGN-10.Using direct HRM analysis.PcH2, PcH2TGN-10 were classified into four groups:PcS, PcH1, PcH2, PcW, PcSTGN-10 , PcH1TGN-10 , PcH2TGN-10 , PcWTGN-10 according to the melting curves of the unlabeled probe .Combined the HRM analyses of the whole amplicon and unlabeled probe , the eight Pc variants can be differentiated from each other .Five different Pc variants, PcS, PcW, PcH1, PcH2TGN-10 and PcWTGN-10 , were identified and consistent with direct sequencing results .Conclusions This developed a simple Pc and P 2 variants differentiation method via simultaneous HRM analyses of an unlabeled probe and full-length amplicon .This method is cost-effective and accurate , could be used in differentiation of Pc and P2 variants of class 1 integrons in clinical isolates .

Objective To investigate the changes in intracellular Ca2+ in the ureter smooth muscle cells (USMC) of rats with neuropathic urinary tract dysfunction (NUTD) and their significance.Methods Forty-five rats were randomly and averagely divided into NUTD group,experimental control (EC) group and blank control (BC) group.The NUTD group was operated with a spinal cord transection at the first lumbar level and the sacral cord was destroyed;in EC group the spinal process was partly bitten at the same position,but the spinal cord was not transected;BC group was given no operations.One week later,the video-urodynamic was performed to observe the acontractile detrusor (ACD),vesicoureteral reflux (VUR) and urinary tract dysfunction in rats among the NUTD group,EC group and the BC group.Video-urodynamic assessment was performed at the sixth week after operation.Ureter smooth muscle cells (USMC) were obtained by collagenase digestion.Intracellular Ca2 + in the USMC were observed by laser scanning confocal microscope.Then the effects of Bay K8644(10-8 mol/L,10-7 mol/L,10-6 mol/L) on cytosolic Ca2+ concentrations([Ca2+] i) in NUTD group were studied by calculating the fluorescence intensity.Results ACD and no detrusor overactivity were found in all rats in NUTD group and without vesicoureteral reflux.Immunofluorescence method confirmed that the cells were USMC.Compared with BC group (31.44 ± 2.82) and EC group (32.06 ± 3.67),the fluorescence intensity (FI) of intracellular Ca2 + in USMC was much lower in the NUTD group (9.80 ± 1.11),and there was significant difference(P ＜ 0.05).Bay K8644 (10-8 mol/L,10-7 mol/L,10-6 mol/L) increased the FI of [Ca2 +] i in a concentration-dependent manner,which were 3.80 ± 1.30,10.04 ± 2.15,19.89 ± 2.06,respectively,and there was significant difference (P ＜ 0.05).Conclusions The decrease in Ca2 + concentration in the ureter smooth muscle cells may be one of the important factors for the primary ureteral dysfunction of NUTD.And calcium channel agonist can be meaningful for adjusting abnormal Ca2+ concentration in USMC of NUTD.

The quality of SCI papers is one of the objective indexes of evaluation on scientific and technological strength and research capabilities.This paper introduced a comprehensive management strategy to promote the publication of SCI papers with high impact factors,in terms of such dimensions ass research orientation,financial and technical support,personnel training,and scientific research management platform.The short and long term effects of the comprehensive management strategy system were analyzed using the SCI papers publication data and IF data from 201 1 to 2014 at the hospital,as a reference for building a scientific management system of SCI papers for the administrators.

Objective To study the clinical value of whole －body magnetic resonance diffusion weighted imaging(WB－DWI) in evaluating the chemotherapy response for lung cancer,thus to provide evidence for optimizing clinical imaging examination. Methods From October 2017 to May 2018,60 patients with lung cancer confirmed by histopathology in Linfen Central Hospital were selected. The patients underwent DWI examinations before chemotherapy and after two cycles of chemotherapy. The change of tumor size,distant metastasis and apparent diffusion coefficient (ADC) value were compared before and after chemotherapy. The correlation between the change rate of ADC value and the shrinkage rate of tumor size in the effective group was analyzed. Results Of 60 cases,1 case had new cerebral metastases after chemotherapy. There were statistically significant differences in ADC value [(1. 12 ± 0.33) ×10 －3mm2/svs.(1.56±0.40) ×10 －3mm2/s]andtumorsize[(4.63±2.75)cmvs.(2.28±1.45)cm] between before and after chemotherapy in the effective group(t= －3. 954,4. 711,all P＜0. 01). There was correlation between the change of ADC value and tumor size(r=0. 34,P＜0. 05). Conclusion WB－DWI can not only detect the change of tumor size and distant metastasis quickly and effectively,but also can observe the microscopic changes of tumor cells by measuring ADC value. So it can predict the early therapeutic response of the tumor and make effective evaluation for the staging and chemotherapy response of lung cancer.

Objective: To elucidate the clinicopathologic characteristics of atypical epithelioid trophoblastic lesions with cyst and fistula formation after cesarean section. Methods: The clinical and pathological data of 4 cases of post-cesarean atypical epithelioid trophoblastic lesions with cyst and fistula formation diagnosed at Women′s Hospital, School of Medicine, Zhejiang University during April 2007 to June 2018 were evaluated by hematoxylin and eosin stain and EnVision two-step immunohistochemical staining technique. Results: The age of the 4 patients ranged from 32 to 41 years, with a mean age of 36.5 years. Three patients recieved cystectomy and one underwent subtotal hysterectomy. Histologically, the lesions were well circumscribed and consisted of uniform cells of medium size, irregularly enlarged with hyperchromatic nuclei and 1 to 2 inconspicuous nucleoli embedded in abundant hyalinized matrix with fibrinoid material in the center. The cells exhibited immunohistochemical feature of chorionic-type intermediate trophoblastic cells (CK18+, p63+ and CD146-). All patients were alive without recurrence during follow-up of 1 to 40 months (mean=22 months). Conclusion Atypical epithelioid trophoblastic lesion with cyst and fistula formation after cesarean section has unique histological features, and its biological behavior and prognosis are still unclear, which need further exploration.

Objective To investigate the situation of sexual partners and related factors among men who have sex with men (MSM) in college students.Methods Snowball sampling and Convenience sampling were both used to recruit MSM from colleges in Tsingtao in 2016.Questionnaire-based interviews were conducted to collect data of socio-demographic and situation of sexual partners.Sample Size was estimated based on cross-sectional study,and theoretical 267.SPSS 17.0 software was used for statistical analysis.Results A total of 300 MSM,average aged 20.7,were analyzed.Both first sex partner and the last same-sex sexual partner were mct instantly,with proportions as 58.7％ (176/300) and 62.3％ (187/300) respectively.Among all the MSM,88.3％ (265/300) preferred selecting men as sex partners and 42.7％ (128/300) enjoyed finding sex partners in college,while 86.0％ (258/300) preferred finding their sex partners through intemet.Conclusions Intemet had been the major way of looking for sex partner among MSM in college students,the male sexual partner were met instantly.We should focus on men who have sex with men and their sexual health among college students to prevent and control HIV/AIDS.

Objective:To explore the mechanism of action of Coptis chinensis in the treatment of dental caries using a network pharmacology approach and animal experiments. Methods:The active ingredients of C. chinensis and their targets were screened by the traditional Chinese medicine systems pharmacology (TCMSP) database and analysis platform, and the targets were searched online through the GeneCards database. The intersecting targets of C. chinensis and dental caries were screened at Venny 2.1, and the intersection targets were analyzed online for protein-protein interaction analysis and gene ontology (GO) and kyoto encyclopedia of genes and genomics (KEGG) enrichment. Then, Cytoscape was used to create a "component-target-pathway" network diagram. Rats were randomly divided into the model group and the C. chinensis group to establish a rat model of dental caries. Rats in the model group were repeatedly rubbed with a cotton ball soaked in 150 μl of 0.9% NaCl solution for 5 min, and rats in the C. chinensis group were repeatedly rubbed with a cotton ball soaked in C. chinensis (5.8 mg of C. chinensis in 150 μl of 0.9% NaCl solution) for 5 min. The two groups of rats were treated once a week for four consecutive weeks. The number of Streptococcus mutans colonies was counted, and serum serine/threonine protein kinase 1 (AKT1), JUN, interleukin-6 (IL-6), tumor necrosis factor (TNF), and B-cell lymphoma-2 (Bcl-2) were detected by enzyme immunoassay. Results:A total of 11 active ingredients in C. chinensis were found, which regulate multiple molecular pathways by intervening in 54 targets, thereby treating dental caries. Quercetin, berberine, flavodoxin, berberine infusion, and tetrahydroberberine were the core components, and AKT1, JUN, IL-6, TNF, and Bcl-2 were the core targets. GO analysis showed that BP mainly included cytokine activity, signaling receptor activator activity, signaling receptor modulator activity, cytokine receptor binding, and receptor ligand activity, etc.; and CC mainly included the response to lipopolysaccharides, the response to bacterial molecules, cellular responses to lipids, inflammatory responses, and negative regulation of cell population proliferation; MF mainly includes membrane rafts, membrane microregions, extracellular matrix, external encapsulated structures, and plasma membrane protein complexes, etc. KEGG analysis showed that advanced glycosylation end product-receptor for advanced glycosylation end products (AGE-RAGE), TNF, IL-17, Toll-like receptor, hypoxia-inducible factor-1 (HIF-1), mitogen-activated protein kinase (MAPK), nuclear factor-κB (NF-κB), epidermal growth factor receptor (EGFR), Janus kinase-signal transducer and activator of transcription (JAK-STAT), and phosphatidylinositol 3-kinase-protein kinase B (PI3K-Akt) signaling pathways have been associated with C. chinensis treatment. The results of animal experiments showed that serum Bcl-2 protein expression increased and serum AKT1, JUN, IL-6, TNF, and other proteins decreased after the C. chinensis treatment. Conclusions:C. chinensis can be involved in regulating the targets of dental caries through multiple pathways, with good therapeutic effects and a wide range of mechanisms of action, and is expected to be an important component in the development of proprietary Chinese medicines for the treatment of dental caries.

OBJECTIVETo assess the association between -1296T/C and -915A/G polymorphisms in the promoter region of matrix metalloproteinase inhibitor-3 gene (TIMP-3) and atherosclerotic cerebral infarction in an ethnic Han Chinese population.

METHODSPeripheral blood samples were collected from 485 patients with atherosclerotic cerebral infarction and 525 healthy controls. Serum levels of TIMP-3 were measured with an enzyme-linked immunosorbent assay (ELISA). The polymorphisms of the TIMP-3 gene were analyzed with DNA sequencing.

RESULTSThere were significant differences in genotype and allele frequencies in -1296T/C and -915A/G between the patients and healthy controls (chi-square: 5.227 and 5.869; P: 0.022 and 0.015, respectively). Besides, there was a strong linkage disequilibrium between -1296T/C and -915A/G (D'=1.0, r(2)=0.991). The serum levels of TIMP-3 in patients were significantly higher than the control group [(248.90 ± 97.10) pg/mL vs. (200.17 ± 79.70) pg/mL, t=2.098, P=0.039].

CONCLUSIONThe -1296T/C and -915A/G polymorphisms of the TIMP-3 gene are associated with increased risk for atherosclerotic cerebral infarction in ethnic Han Chinese and may be used as molecular markers for the disease. There is also strong linkage disequilibrium between the two loci.

Adult ; Aged ; Aged, 80 and over ; Asian Continental Ancestry Group ; ethnology ; genetics ; Atherosclerosis ; blood ; epidemiology ; ethnology ; genetics ; Base Sequence ; Cerebral Infarction ; blood ; epidemiology ; ethnology ; genetics ; China ; epidemiology ; Female ; Gene Frequency ; Humans ; Male ; Middle Aged ; Molecular Sequence Data ; Polymorphism, Single Nucleotide ; Risk Factors ; Tissue Inhibitor of Metalloproteinase-3 ; blood ; genetics

OBJECTIVETo investigate the association between cerebral infarction (CI) and single nucleotide polymorphism (SNP) in the exon of membrane-type 1 matrix metalloproteinase (MMP-14) gene in Chinese Han population.

METHODSFive hundred seventy four patients with CI and 463 healthy individuals were recruited. Serum MMP-14 level was measured with enzyme-linked immunosorbent assay (ELISA). rs1042704 and rs2236307 polymorphisms of the MMP-14 gene were genotyped with a TaqMan assay. Multivariate logistic regression was carried out to analyze the risk factors of CI.

RESULTSA significant lower risk of CI was found in individuals with MMP-14 rs2236307 TC and CC genotypes (vs. TT genotype: P<0.05). The frequencies of MMP-14 rs2236307 C allele were significantly different between the CI group (37.46%) and the control group (43.95%) (P=0.003). Serum level of MMP-14 was higher in the CI group (P=0.003) and was also higher in the group with MMP-14 rs2236307 TT genotype compared with those with CT and CC genotypes (P=0.000; P=0.009). Logistic regression analysis indicated that the MMP-14 rs2236307 CT+CC genotypes was a protective factor, and that history of hypertension, smoking status, triglycerides, diastolic blood pressure and systolic blood pressure were the independent risk factors of CI (AOR:2.027, 1.302, 1.296, 1.434, 2.087; P<0.05).

CONCLUSIONThe rs2236307 polymorphism of MMP-14 gene is associated with CI, for which the C allele maybe a protective factor. No association of MMP-14 gene rs1042704 polymorphism with CI has been found.

Adult ; Aged ; Alleles ; Asian Continental Ancestry Group ; genetics ; Blood Pressure ; Case-Control Studies ; Cerebral Infarction ; genetics ; physiopathology ; Female ; Genetic Association Studies ; Genotype ; Humans ; Male ; Matrix Metalloproteinase 14 ; genetics ; Middle Aged ; Polymorphism, Single Nucleotide ; Risk Factors

OBJECTIVETo analyze specific expression of blood group genes using nucleated erythroid cells cultured from un-mobilized peripheral stem cells in vitro.

METHODSHematopoietic stem cells(HSC) bearing the CD34 antigen were isolated from peripheral blood by centrifugation and magnetic beads sorting, followed by suspension culture in vitro. Cells were collected from medium on various stages and analyzed by immunofluorescence. The RNA transcription of RH and ABO blood group genes was analyzed using culture cells on day 12.

RESULTSA total of(3.19±0.13) ×10 (4) CD34+cells were isolated from about 50 mL peripheral blood with a recovery rate of 67.3%±2.7%. The cells amount in erythroid-lineage culture system on day 9 reached a plateau of a 237.1±15.5-fold amplification of the initial cell input. The stem cell-specific CD34 antigen was dropped off, while the erythroid-specific CD235a and CD240D antigens were increased in culture period. RHD/CE and ABO genes can be amplified using RNA extracted from culture cells on day 12, and genotypes of Rh and ABO systems by DNA sequencing were consistent with their serologic phenotypes.

CONCLUSIONA method was established to analyze the gene expression of erythroid blood group derived from un-mobilized peripheral stem cells cultured in vitro. It can be used to study the expression of various erythroid-specific genes.

Antigens, CD34 ; analysis ; genetics ; Base Sequence ; Blood Group Antigens ; analysis ; genetics ; Cells, Cultured ; Erythrocytes ; cytology ; Flow Cytometry ; Hematopoietic Stem Cells ; cytology ; Humans ; Molecular Sequence Data