Objective To investigate the role of a cis-acting element,cell cycle genes homology region (CHR),in the transcriptional regulation of human NFBD1 gene (a nuclear factor with BRCT domain 1) by conducting a site-directed mutagenesis analysis on the element in human NFBD1 promoter region.Methods Wild type of NFBD1 promoter reporter,NFBD1-PS1-325,was used as template to make CHR mutant by using PCR based site-directed mutagenesis.Dual luciferase reporter assay was used to determine promoter reporter activity.Adiamycin treatment was employed to investigate the role of CHR in the transcriptional downregulation of NFBD1 after DNA damage.Results Site-directed mutagenesis of CHR caused a significant decrease in NFBD1 promoter activity,and also attenuated the transcriptional down-regulation of NFBD1 after DNA damage.Conclusion CHR element might be involved in both basic transcriptional regulation and transcriptional downregulation of NFBD1 after DNA damage.

Objective To explore the possible factors associated with twice human brucellosis epidemics in Inner Mongolia during 1952 to 2007 to provide scientific tactics for prevention and control brucellosis. Methods Surveillance data and literature about human brucellosis during 1952 to 2007 in Inner Mongolia was collected, descriptive analysis of human brucellosis incidence on distribution in the regions and among occupations was carried out during 1952 to 2007. Results In Inner Mongolia, the first epidemic of human brucellosis peak appeared in the early 1960s, spreading to 12 regions, at an incidence of 55.28/100 000 in 1961, 72.9% of the Brucella infected people were herdsman;another epidemic peak seriously hit middle and eastern regions after 2000, the incidence being 38.44/100 000 in 2005;51.9% and 28.7% of the new brucellosis cases were respectively peasant and herdsman. Conclusions In Inner Mongolia, animal husbandry industry has been rapid developed since the early 1990's, resulting frequent livestock trade without quarantine, at the same time the public health system doesn't match the development, so the epidemic situation of brucellosisbecomes more and more serious after mid-90's, and has reached the peak during 2004 and 2007.

Purpose To investigate the relationship of infundibular bladder neck formation with pelvic floor support structure injury and urethral sphincter defect and its significance in female stress urinary incontinence.Materials and Methods The pelvic floor images of seventy-four female patients with stress urinary incontinence treated in the outpatient Department of Lanzhou University Second Hospital from April 2015 to August 2016 were analyzed retrospectively.The location of the bladder neck,posterior vesicourethral angle and the infundibular bladder neck formation were observed by the transperineal ultrasound under the resting state and the maximum Valsalva status.Meanwhile the thickness of middle urethral sphincter was measured under resting state.At the same time,eighty-one women visiting our hospital for regular physical examination were enrolled as control group.Results The infundibular urinary bladder neck formation rate (66.2％) in the stress urinary incontinence group was significantly higher than that in the control group (4.9％) under maximum Valsalva state,the difference was statistically significant (P＜0.05).The extent of the bladder neck descending and posterior vesicourethral angle in the stress urinary incontinence group were notably higher than those in the control group,both of the difference was statistically significant (P＜0.05).Stress urinary incontinence was confirmed with urethral sphincter defect by urodynamics in nine patients,in whom the infundibular bladder neck occurred.The thickness of the middle urethral sphincter in these nine patients showed no obvious difference with that in patients without sphincter defect and subjects in normal control group (P＞0.05).Conclusion The infundibular bladder neck formation,which is closely related to the pelvic floor support structure dysfunction and urethral sphincter defect,is an important indication of stress urinary incontinence.However,the assessment of urethral sphincter defect through urethral sphincter thickness need to be further studied.

Objective To study the clinical effect of gasless-laparoscopic vaginoplasty using sigmoid colon segment.Methods Clinical data of 119 cases undergoing laparoscopic or gasless-laparoscopic vaginoplasty using a vascularized pedicled sigmoid colon segment in Beijing Anzhen Hospital from January 2007 to December 2010 were reviewed retrospectively.Those patients were classified into 57 cases with laparoscopic sigmoid colon vaginoplasty and 62 cases with gasless-laparoscopic sigmoid colon vaginoplasty.The operation time,blood loss in operating,bowel movement after operation,postoperation hospital duration,side effect,and artificial vagina were compared between laparoscopic and gasless-laparoscopic group.Results The vaginoplasty were preformed successfully in 119 cases.The mean operation time of were (159 ± 18) min in laparoscopic group and (146 ± 17) min in gasless-laparoscopic group,respectively,which reached statistical difference (P ＜0.01).The blood loss in operating were (83 ± 14) ml and (86 ± 13)ml,bowel movement after operation were (68 ± 8) hours and (68 ± 11) hours,and postoperation hospital duration were (11.1 ± 1.3) days and (11.4 ± 1.9) days respectively in laparoscopic group and gasless-laparoscopic group.No significant difference were found in the blood loss in operating,bowel movement after operation,and postoperation hospital duration between two groups (P ＞ 0.05).No intraoperative complication occurred.There were two cases with incomplete adhesive intestinal obstruction at 15-20 days postoperatively,which one was in laparoscopic group and one was in gas-less laparoscopic group.At 6-50 months of following up (median time 12 months),all artificial vaginas had a capacity of over two fingers in wideness and 12-15 cm in length.Vaginal discharges resembled a milky white water or mucus without odour.Eighty-five patients with sexual intercourse reported satisfactory feeling.One patients complained vaginal stenosis in laparoscopic group.Conclusion Gasless-laparoscopic vaginoplasty using sigmoid colon segment is an alternative feasible and practical treatment.

OBJECTIVETo analyze the differential proteomics in human umbilical cord mesenchymal stem cells (MSC) induced by chemical hypoxia-mimetic agent cobalt chloride (CoCl(2)) by two-dimensional gel electrophoresis (2-DE) and mass-spectrometry.

METHODS2-DE was performed to separate proteins from treated and untreated human umbilical cord MSC with CoCl(2). 2-DE images were analyzed by ImageMaster 2D Platinum software 6.0. The differential expressed proteins was identified by MALDI-TOF-MS. The differential proteins were classified based on their functions.

RESULTS2-DE reference patterns of CoCl(2) treated human umbilical cord MSC were established. A total of twenty-six differential proteins were identified, of them eleven proteins were up-regulated and fifteen down-regulated. Their biological functions involved in carbohydrate metabolism, protein metabolism and modification, lipid metabolism, coenzyme and prosthetic group metabolism, cell cycle, immunity and defense, cell structure and motility, signal transduction, protein targeting and localization, neuronal activities, muscle contraction, etc. Peroxiredoxin1 (Prdx) was down-regulated, whereas alpha-enolase (ENO1) and vesicle amine transport protein 1 homolog (VAT1) up-regulated.

CONCLUSIONThe effects of hypoxia on human umbilical cord MSC were participated by multiple proteins and involved in multiple functional pathways.

Cobalt ; pharmacology ; Humans ; Mesenchymal Stromal Cells ; drug effects ; metabolism ; Proteome ; analysis ; Proteomics ; Umbilical Cord ; cytology ; drug effects

To investigate the correlation between methylation and expression of multidrug resistance (mdr1) gene, restriction endonuclease HpaII combined with competitive PCR technique was used to quantitatively detect the methylation status of two CCGG sites located at -110 and -50 bp (region I and II) up to the transcription start site in mdr1 promoter in 54 AL and 9 MM patients. Semi-quantitative RT-PCR was used to detect the expression level of mdr1 gene. The results showed that inverse correlation between methylation rate of either region or total methylation rate and expression of mdr1 gene was observed. The correlation in the region I (r = -0.64) was closer than that in the region II (r = -0.4). High expression rate of mdr1 ascended significantly in low methylation group (n = 36) (P < 0.001). In comparison with chemotherapy sensitive group (n = 8), the methylation rate in refractory AL patients (n = 16) was lower (P = 0.05) in the region I, P < 0.05 in the region II and total regions. Comparing with the untreated patients (n = 36), the methylation rate in the region I and total methylation rate were lower in the patients with chemotherapy (n = 14) (P < 0.05). The methylation rate in the region II was also decreased after chemotherapy, however, no statistical significance was shown (P > 0.05). Increased mdr1 expression level accompanying with decreased methylation rate after chemotherapy was found, although no significant difference was shown (P = 0.06). It is concluded that the expression level of mdr1 gene was associated with the methylation status of CCGG in -110 and -50 bp upstream to the transcription start site, especially the -110 site. In both the patients treated with chemotherapy and the refractory patients, the methylation level of mdr1 gene decreased relatively. The rising expression of mdr1 gene after chemotherapy was associated with the decrease of methylation level.
DNA Methylation ; Genes, MDR ; Hematologic Neoplasms ; drug therapy ; genetics ; Humans ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo suppress the expression of CCR5 and CXCR4, the co-receptors for human immunodeficiency virus type 1 ( HIV-1), and thus inhibit HIV-1 from entering cells.

METHODSDNA fragments encoding either CCR5 or CXCR4 were amplified from healthy human peripheral blood mononuclear cells (PBMCs) by reverse transcript polymerase chain reaction (RT-PCR) and sequencing was performed. Correct fragments were inserted into Shuttle plasmid inversely, which was recombined with backbone plasmid containing homologous adenoviral genome in E. coli BJ5183. The recombinant plasmids were transfected into 293 cells in which they were packaged and amplified. Recombinant adenoviruses containing antisense RNA of CCR5 or CXCR4 were obtained and identified by RT-PCR, and the titres of them were determined by cytopathic effect (CPE) method. The U937 and MT4 cells were infected by recombinant adenoviruses containing antisense RNA of CCR5 (multiplicity of infection, MOI = 100) and CXCR4 (MOI = 200), respectively. The expression of co-receptors on infected cell was measured by fluorescence activated cell sorter at 24, 48, 72 hours and 10 days after infection. In addition, the chemotactic activity and proliferation of infected cells were detected with Boyden chamber and 3H incorporation respectively.

RESULTSWe constructed the recombinant plasmids and obtained the recombinant adenoviruses which contained antisense RNA of CCR5 or CXCR4 and were designated as pAd-antiR5 and pAd-antiX4 respectively. The titers of recombinant adenoviruses pAd-antiR5 and pAd-antiX4 were 5 x 10" PFU/ml and 7 x 10(10) PFU/ml, respectively. The expression rate of CCR5 on U937 cells decreased from 82. 10% (blank control) to 1.12% (Ad-antiR5 infected) , and that of CXCR4 on MT4 cells decreased from 42% (blank control) to 1.03% (Ad-antiX4 infected) 24 hours later. The expression rates of CCR5 on Ad-antiR5 infected U937 cells were 1.02% , 1.26% , 1.23% at 48 hours, 72 hours, and 10 days later, respectively. The expression rates of CXCR4 on Ad-antiX4 infected MT4 cells were 1.13%, 1.17%, 1.22% at 48 hours, 72 hours, and 10 days later, respectively. Moreover, the recombinant adenovirus had no effects on chemotactic activity and proliferation of the cells.

CONCLUSIONThe recombinant adenovirus containing antisense CCR5 or CXCR4 can remarkably decrease the expression of co-receptors for HIV-1 on U937 or MT4 cells without affecting their chemotactic activities and proliferative abilities.

Adenoviridae ; genetics ; Cell Line, Transformed ; Cell Proliferation ; Chemotaxis ; Down-Regulation ; Genetic Vectors ; Humans ; RNA, Antisense ; genetics ; RNA, Messenger ; genetics ; Receptors, CCR5 ; biosynthesis ; genetics ; Receptors, CXCR4 ; biosynthesis ; genetics ; Transfection ; U937 Cells

OBJECTIVETo clarify the gene type of Orientia tsutsugamushi (Ot) from Shandong province.

METHODSNested-polymerase chain reaction (nPCR) was used to identify the gene type of 23 isolated Ot strains, 2 pools of homogenized leptotrombidium (L.) scutellare, 10 blood specimens of scrub typhus patients, and at the same time to compare with the international reference strains Gilliam, Karp, Kato. Sequencing analysis of the Sta56 gene was also used to further identify the precise gene types.

RESULTSOf the 35 samples, 33 had the same products in the amplification of template Ot-DNA. They all belonged to Kawasaki strains endemic in Japan while 2 (FXS4 and LHGM2 strain) belonged to Karp strains. The Sta56 gene sequence homologies to Japan Kawasaki strain of the 2 representative strains (B-16 and FXS2 strain) of the 33 samples were 94.22%, 95.21% respectively, but they were less than 75.87% to other prototype strains; The homologies to Karp strain of FXS4 and LHGM2 strain were 83.03%, 96.45% respectively. B-16 and FXS2 strain were designated as of types strain Japan Kawasaki, FXS4 and LHGM2 as Karp strain.

CONCLUSIONThe results indicated that the dominant Ot strains in Shandong Province were similar to Kawasaki strains, but Karp strains also existed.

Animals ; DNA, Bacterial ; genetics ; Genotype ; Humans ; Mice ; Orientia tsutsugamushi ; genetics ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Scrub Typhus ; epidemiology ; microbiology ; Sequence Homology ; Serotyping

Objective To discuss a real-time fluorescence quantitative polymerase chain reaction (FQ-PCR) wether if can be used to detect Brucella. Methods According to the BCSP31 gene sequences specific for Brucella, one pair of primers and one TaqMan probe were designed. A real-time PCR was developed with the BCSP31 fragments cloned into PMD18-T vector. The standard cure was established and the sensitivity, the species specificity and the stability of the assay were evaluated. The clinical blood specimens were detected by QT-PCR and compared with clinical diagnosis. Results The standard curve was established with the standard template and the relationship between the Ct and the DNA copy number was linear(r=0.999). The sensitivity of the real-time PCR was 5 copies/μl. The sensitivity of the common PCR was 5×102 copies/μl. The sensitivity was about 100 times higher than common PCR. Species specificity of this FQ-PCR assay evaluated using genomic DNA from 6 Bmcella strains and 5 non-Brucella strains and strong fluorescence was detected in all Brucella strains. The CV of intra-assay and inter-assay reproducibility were 0.71%,7.23%, reprectively. Twenty-four specimens from clinical brucellosis cases, 19 showed positive, the positive coincident rate was 79%(19/24). The negative results were obtained for all 31 negative control, and the negative coincident rate was 100%(31/31). Two were positive from all 30 specimens clinically suspected. Conclusions Highly specific, sensitive, repeatable and coincidental with clinic, this FQ-PCR is quite useful for rapid detection of tiny DNA of Brucella in various samples and laboratory diagnosis.

Objective Blood pressure variability (BPV) is an independent risk factor for the death of patients with maintenance hemodialysis (MHD).There is no study on the influencing factors of BPV at home in HD patients in China.The article aimed to investigate MHD patients'BPV at home and related influencing factors in order to provide theoretical basis for reducing home BPV (HBPV) clinically. Methods We chose 103 patients who were treated with MHD in the Renal Medicine Room of Nephrology Department in three upper first -class hospitals including Jiangsu Provincial People 's Hospital, the First Affiliated Hospital of Suzhou University and the Affiliated Hospi -tal of Jiangsu University from March 2017 to October 2017.We col-lected their 7 days'blood pressure monitoring at home and blood pressure before dialysis, average value and standard deviation in sys -tolic blood pressure monitoring at home, and took the coefficient of variation of systolic blood pressure as the expression of HBPV .The patients were divided into high BPV group (BPV≥5.8％) and low BPV group (BPV<5.8％) according to the average BPV.At the same time, we recorded indexes such as sex , age, dialysis age, primary disease, BMI, inter-dialytic weight gain (IDWG), left ven-tricular mass index(LVMI) and analyzed relative influencing factors by multiple linear regression . Results The age, IDWG and LV-MI were positive independent influencing factors of HBPV (R 2 =0.467,F=10.945,P<0.001).According to standardized regression co-efficient, the contribution of each variable to HBPV was as follows : PIBWG >Age>LVMI. Conclusion In clinical nursing, we should actively control the IDWG of patients , encourage patients to monitor their blood pressure at home , and increase their awareness of the importance of home BPV.Meanwhile, HBPV is an important index for predicting left ventricular hypertrophy and can be used as an objective tool to improve patients 'self-management ability.