Objective To develop a rapid method for construction of RNAi vector.Methods After inversely inserting the U6 and H1 polymerase Ⅲ promoters into the pBluescriptⅡ backbone vector,only with two short reverse complementary oligo nucleotides,the RNAi vector with one interference cassette could be constructed.Using two isoaudamers MunⅠ and EcoRⅠ with sticky ends,several cassettes could be fused together to form the ultimate multiple-site targeting RNAi vector.Using this method,we constructed RNAi vectors targeting green fluorescent protein(GFP) and firefly luciferase(LUC) gene separately or both for the test of knock down property of these vectors.Results Constructed single and two site targeting RNAi vectors got desirable knockdown effects,in which single site targeting vector could get about 90 percent knock down efficiency whereas two site targeting vector reached about 87.5 percent knock down efficiency either.Conclusion Our RNAi vector construction strategy is efficient and time-saving so that it can be used for knocking down several genes simultaneously in the same cell.

Objective National external quality assessment (EQA) results from 2005 to 2008 for HLA class Ⅰ (A, B) and class Ⅱ (DRB1) low-resolution DNA typing were summarized with the goal of exploring strategies to assure and improve HLA DNA typing performance in clinical testing. Methods HLA allele results from the four consecutive years EQA events were analysed. Different kinds of errors were described and classified, and the possible causes were discussed. Results Participant laboratories were increasing in the four consecutive years with the number of 22, 28, 47 and 61 in 2005, 2006, 2007 and 2008,respectively. 2 844 HLA DNA typings were returned from the participants during the 4 years EQA surveys, and overall 30 errors (1.05%) were identified. These 30 errors were classified into two major types of errors including 25 technical genotyping errors and 5 human errors. The proportion of laboratory participants which made mistakes was 13. 6%, 10. 7%, 10. 6% and 16.4% in 2005, 2006, 2007 and 2008, respectively. Conclusions Above 10% of participant laboratories exhibited errors in the four consecutive years HLA molecular typing EQA surveys. Relevant important attentions should be greatly paid to clinical HLA molecular typing test.

Objective To investigate the relationship between the X-ray,CT findings and its pathological basis of nonossifying fibroma(NOF).Methods X-ray and CT features of NOF confirmed by surgical pathology in 17 cases,and correlative study of imaging andpathology was done.Results 17 lesions were showed as eccentric and expanded destruction located in the cortex,and the remains of bone could be found inside the tumor,the lesions projecting into marrow cavity with a thin bone between the tumor and marrow.The tumors were grey-yellow or brown tissue around by the thin bone found in operation.NOF was consisted of a lot of shuttle cell and collagen fiber under microscope,some of macrophagocyte with multiple nucleuses and foam cell dropped disseminating in them,and there was not osteogenesis in the tumor.Conclusion The pathological changes of NOF can be reflected by X-ray and CT.

Objective: To investigate the role of hypoxia-inducible factor-1α (HIF-1α) in arsenite-induced epithelial-mesenchymal transition (EMT) and malignant transformation of human liver epithelial cells (L-02 cells). Methods: After the L-02 cells were chronic treated with 2.0 μmol/L NaAsO₂ for 0 (reference), 10, 20, or 30 passages, con siRNA or HIF-1α siRNA was transfected into arsenite-transformed L-02 (T-L-02) cells by lipofectamine^TM2000 and were set as T-L-02+con siRNA group and T-L-02+HIF-1α siRNA group as well as L-02 group and T-L-02 group, EMT index and levels of HIF-1α were detected by western blots. The reporter assays were performed to determine if HIF-1α directly regulate Snail transcriptional activity, and soft agar colony formation and Transwell assay were used to detect the malignancy, invasion, and migration ability of cells. Results: When L-02 cells were treated for 10 generations with 2 μmol/L NaAsO₂, relative expressions of E-cadherin were gradually increased compared to control cells, while the levels of N-cadherin, Snail, and HIF-1α were gradually increased in the L-02 cells compared to control cells, showing the longer the treatment time was, the more obvious the change was (P<0.05) . Down regulating the level of HIF-1α by siNRA caused E-cadherin levels to rise compared to T-L-02 group, while the levels of N-cadherin and Snail fall back compared to T-L-02 group (P<0.05) . Double luciferase reporter gene assays showed that HIF-1α directly targeted Snail to regulate its expression. Soft agar colony formation and Transwell assays showed that the numbers of formed colonies, invasion cells, and metastasis cells of cells in T-L-02 group were all lower than those in L-02 group (P<0.05) . Conclusion HIF-1α is involved in arsenite-induced EMT and malignant transformation of human liver epithelial cells via regulating Snail.

OBJECTIVETo understand the syphilis infection and its high risk factors among men who have sex with men (MSM) recruited from different channels.

METHODSSupported by the China-Gates Foundation HIV program from July to December 2011, we cooperated with community based organizations to conduct syphilis testing intervention among MSM from 14 cities (Beijing, Shanghai, Tianjin, Chongqing, Harbin, Shenyang, Qingdao, Xi'an, Nanjing, Wuhan, Hangzhou, Changsha, Kunming, and Guangzhou) and one province (Hainan province). Participants were recruited from different channels by the staff of local CBOs, Demographic (e.g. age, marital status, and education) and behavioral (e.g. condom use and sexual partners) data were collected using anonymous questionnaires. Blood samples were also collected to test for syphilis. A total of 34 100 MSM participated in the survey. Participants were excluded from data analysis (1 399, 4.1%) if they did not receive syphilis tests or they completed less than 80.00% of the key questions in the survey. Chi-square tests were used to understand the socio-demographic and behavioral differences between each group. Results of syphilis tests were also compared. Logistic regression models were used to test the statistical significance of these differences.

RESULTSA total of 32 701 MSM were enrolled and received syphilis testing. The average age of participates was 30.96 ± 9.57. And among them, 2 284 cases (7.0%) were recruited from gay bathhouses, 4 774 (14.6%) from gay bars, 6 266 (19.2%) from the internet, 1 997 (6.1%) from the parks/toilets and 17 380 (53.1%) from other channels. MSM recruited from the bathhouses had the highest syphilis infection rate than other 4 groups: gaybars (4.5%, 216/4 774), internet (6.7%, 422/6 266), parks/toilets (8.3%, 166/1 997), other channels (6.4%, 1 103/17 380) (χ² = 164.58, P < 0.001). The multivariate logistic regression analysis showed that being > 20 years of age (P < 0.001), having more than 2 homosexual partners in recent 3 months (8.0% (1 408/17 714), OR (95% CI) = 1.44 (1.04-1.98)), having no sex with females in past 3 months (6.8% (1 446/21 276), OR (95% CI) = 1.25 (1.07-1.46)), and not using condom at last anal sex (8.0% (769/9 668), OR (95% CI) = 1.13 (1.03-1.25)) were associated with a higher probability of being infected with syphilis. Whereas MSM married (7.2% (456/6 305), OR (95% CI) = 0.84 (0.73-0.98)), having a college or a higher education (5.3% (829/15 684), OR (95% CI) = 0.60 (0.53-0.67)), being local residents (6.5% (1 843/28 185), OR (95% CI) = 0.73 (0.61-0.87)) and living in the local province of project cities (6.6% (170/2 593), OR (95% CI) = 0.67 (0.53-0.85)) were protective factors.

CONCLUSIONMSM who were recruited from gay bathhouses have a higher rate of syphilis infection than those in channels relatively. They are older, with low education levels and high-risk sexual behaviors. Tailored interventions are required in the future, especially for MSM from gay bathhouses.

Adult ; China ; Condoms ; Demography ; Homosexuality, Male ; Humans ; Logistic Models ; Male ; Risk Factors ; Safe Sex ; Sexual Behavior ; Sexual Partners ; Surveys and Questionnaires ; Syphilis ; Universities

Objective To compare lateral orbital keyhole approach(LOK) with conventional keyhole approach including supraorbital keyhole approach (SOK) and pterional approach(PTK) for exposuring the sellar region and oper-ation ability, to provide theoretical and practical basis for the clinic. Methods From January, 2017 to Feburary, 2018, 15 cadaver head specimens of Chinese (30 sides) fixed by formalin were randomly divided into 3 groups, simu-lating SOK, LOK and PTK, application of frameless neuronavigation system, intersection of the posterior margin of the optic chiasma and the lamina terminalis served as the base point. Six different reference points were selected to radi-ate into the parasellar region of the skull base. The direction of the 2 adjacent reference points were connected to the base point to form a triangle. Six triangles constituted the sellar region to represent the total area. The supratentorial area, ipsilateral area, inferior area and contralateral area were calculated by stacking triangle. The comparison was made between groups. The Salma operation exposure scale was used to simulate the aneurysms of the common parts in the brain and the quantitative scores were performed. Results The total parasellar regions by SOK, LOK and PTK respectively were:(1641.6±295.6)mm2, (1782.3±294.6)mm2 and (1552.5±307.4)mm2. There was no statistical differ-ence(P>0.05); To compare the supratentorial region, SOK and LOK were both bigger than PTK ( P<0.05); To compare the ipsilateral and infratentorial area, LOK and PTK were both bigger than SOK respectively ( P<0.05);To compare the contralateral area, SOK, LOK and PTK were increased in turn (P<0.05). Salma operation exposure scale was used to get the scores:the score of SOK was 29.7 (39.08％), LOK was 37.0 (48.68％), and PTK was 36.1 (47.50％). Conclusion Anatomical analysis displayed that the 3 keyhole approaches showed different exposure of each part the parasellar re-gions, the LOK had a good exposure to the parasellar region and so as the higher maneuverability. But the clinical appli-cation should be comprehensive analysis, pay attention to specific lesions and make an appropriate choice. It has impor-tant clinical significance to improve the prognosis of patients.

Objectives:To investigate the relationships between CYP2C9 and VKORC1 polymorphism and the steady-state dose of warfarin in Chinese Han patients with nonvalvular atrial fibrillation.Methods:A total 544 Chinese Han patients with atrial fibrillation who received warfarin anticoagulant therapy in Department of Cardiology of Beijing Hospital, Tongren Hospital, Xuanwu Hospital, Anzhen Hospital and Tiantan Hospital were enrolled from January 2016 to may 2019. The genotype and allele frequency in exon 1, 2, 3, 7 of CYP2C9 gene and 1639 site of VKORC1 gene were analyzed; and the general information, warfarin steady-state dose and concomitant medication of patients were recorded.Results:There were four genotypes CYP2C9:CYP2C9*1*1 (93.57%, 509/544), CYP2C9*1 *2 (0.18%, 1/544), CYP2C9*1 *3 (5.88%, 32/544) and CYP2C9*1 *60 (0.37%, 2/544); while VKORC1 had three genotypes: AA (82.72%, 450/544), GA (15.99%, 87/544) and GG (1.29%, 7/544). After reaching the anticoagulation index (INR 2.0-3.0), the steady-state dose of warfarin was the highest in patients with CYP2C9 *1/*1 and VKORC1 GA/GG genotypes, reaching (3.70±1.34) mg/d. The lowest steady-state dose of warfarin was (2.17±0.29)mg/d in patients with both new mutations ( F=22.09, P<0.001). Multiple linear regression analysis showed that body surface area, use of amiodarone, CYP2C9 and VKORC1 genotypes were the independent influencing factors of warfarin steady-state dose ( t=4.44, -2.90, -6.96, 2.14; P<0.05) and the steady-state dose prediction model of warfarin was established. Conclusion:Body weight, height, body surface area, gender, smoking, and combination of amiodarone may significantly affect the steady-state dose of warfarin in patients. CYP2C9 and VKORC1 mutant genotypes were significantly related to the steady-state dose of warfarin. The prediction model based on genetic factors and other influencing factors may effectively predict the steady-state dose of warfarin in Han patients with atrial fibrillation.

Carotid artery stenosis is an important cause of ischemic stroke. Carotid endarterectomy and carotid artery stenting are the effective methods for treating carotid artery stenosis, but postoperative restenosis remains a challenge. The pathogenesis of postoperative restenosis is currently not fully understood. However, multiple factors, including biomarkers, imaging features, and surgical related factors, have been proven to be associated with postoperative restenosis and can predict the occurrence of postoperative restenosis. This article reviews the predictors of restenosis after carotid endarterectomy and carotid artery stenting.

Objective:To explore the clinical experiences and treatment outcomes of partial complicated posterior communicating artery aneurysms (PCoAs) via Dolenc approach.Methods:The clinical data of 27 patients with partial complicated PCoAs accepted clipping via Dolenc approach in our hospitals from July 2014 to June 2019 were retrospectively reviewed. The clinical efficacies were analyzed and the surgical experiences were summarized in these patients.Results:Among 27 patients with partial complicated PCoAs, 26 accepted clipping successfully via Dolenc approach, and one accepted clipping and wrapping. For clinical ourcomes, 2 out of 3 patients with preoperative oculomotor paralysis got improvement after surgery, and one was without change; in 4 patients with new-onset oculomotor paralysis after surgery, 2 returned to normal after conservative treatment, one got improvement, and one was without change. After surgery, 2 had basal ganglia focal cerebral infarction and one had ventriculoperitoneal shunt due to hydrocephalus. DSA or CTA examination within two weeks after surgery showed all 26 aneurysms were completely clipped and without residual. During the 9-12 months of follow up, Glasgow outcome scale (GOS) showed that 20 patients had GOS 5 scores, 3 had GOS 4 scores, 2 had GOS 3 scores, one had GOS 2 scores, and one died. No patients showed aneurysm recurrent in the imaging examination.Conclusion:Via Dolenc approach, anterior clinoid process and its surrounding bony structures are drilled off via extradural way, and proximal of parent artery is exposed well, therefore, more operation space can be provided to deal with partial complicated PCoAs.

Objective:To verify the accuracy of the International Warfarin Pharmacogenetics Consortium(IWPC)model, identify the effects of genetic and clinical factors on steady-state doses of Warfarin, and establish a Warfarin dose prediction model for the Han-Chinese population aged 75 years and over under the guidance of pharmacogenetics.Methods:A total of 544 Han-Chinese patients receiving Warfarin therapy for atrial fibrillation were divided into two groups: those aged 75 years and over(n=164)and those aged below 75 years(n=380). Data for the whole population and the two age groups were each substituted into the IWPC prediction model for accuracy verification.Demographic and clinical characteristics of 164 patients aged 75 years and over were recorded, and the genotypes of CYP2 C9 and VKORC1- G1639 A were detected by polymerase chain reaction.A new pharmacogenetic-guided dosing algorithm for the elderly was obtained by stepwise multiple linear regression.The accuracy of the new model was compared with that of the IWPC model. Results:The predictive accuracy of IWPC for steady-state dosing of warfarin was 35.47% in all subjects, 33.75% in 164 subjects aged below 75 years, and only 28.70% in subjects aged 75 years and over, respectively.In 164 subjects aged 75 years and over, three genotypes of *1/*1, *1/*3 and *1/*2 were detected in CYP2 C9 polymorphism, and the CYP2 C9*1/*1 genotype was the most common one, with a frequency of 87.80%(144/164), followed by the CYP2 C9*1/*3 genotype, at 11.59%(19/164). GG, GA and AA genotypes were detected in VKORC1 polymorphism, among which the AA genotype accounted for 82.32%(135/164)and the GG genotype accounted for only 1.83%(3/164). The steady state dose for Warfarin in patients with the wild-type CYP2 C9*1/*1 was higher than in those with the heterozygote CYP2 C9*1/*3 and *1/*2(3.18±0.86 mg/d vs.2.27±0.51 mg/d, t=5.637, P<0.05). Patients with a mutant homozygotic AA genotype of VKORC1 required lower maintenance doses than those with the heterozygotic GA and GG genotypes(2.96±0.66 mg/d vs. 3.59±1.43 mg/d, t=-2.092, P<0.05). The steady-state dose for Warfarin in subjects carrying CYP2 C9 (*1/*2 or *3)and VKORC1 (GA and GG)was(2.00±0.63)mg/d, lower than in those carrying other genotype combinations( P<0.05). We established a new Warfarin dosing algorithm for elderly subjects aged 75 years and over containing height, creatinine, amiodarone usage, CYP2 C9 and VKORC1 mutants, and the accuracy of the new model was 56.0%, which could explain 56.0% of individual variability, and the accuracy was higher than that of the IWPC algorithm(56.0% vs. 45.8%, P<0.05). Conclusions:Polymorphisms of CYP2 C9 and VKORC1 clearly affect the steady-state dose for Warfarin in the elderly Han-Chinese population aged 75 years and over.A combination of pharmacogenomics with clinical factors can better guide warfarin medication in Han-Chinese people aged 75 years and over.