Objective:To study the interaction of the inflammatory cytokines expression between CD4+ T cells and primary human umbilical vein endothelial cells (HUVECs) infected by dengue virus (DENV-2).Methods:PBMC was extracted from white blood cells by density gradient centrifugation,CD4+ T cells were sorted by immunomagnetic beads.The expression of CD3 and CD4 molecules on the surface of cells was detected by flow cytometry to identify the purity of CD4+ T cells.First,HUVECs were pretreated by specific-S1 P1 receptor agonist CYM-5442 for 24 h,second,infected by DENV-2 on the titer of 103 TCID50,then co-culturing with CD4+ T cells,The relative expression of NS1 partial sequence and IL-6,L-8 mRNA of HUVECs,and IL-4,IL-17,TNF-α,IFN-γ of CD4+ T cells detected by Real-time RT-PCR.IL-6 and IL-8 secreted in cultured supematant analyzed by ELISA.Results:The purity of CD4+ T cells was (98.02±0.32) ％.The expression of NS1 gradually increased to 24 h (3.03±0.26,P＜0.001),decreased after reaching the peak.The relative expression of NS1 in the group of co-cultured with CD4+ T cells was lower than other groups.After infection,the expression of IL-6 and IL-8 were up-regulated,and the expression of IL-6 and IL-8 at each time point was significantly increased after co-culturing with CD4+ T (P＜0.01).IL-6 of CYM-5442 pretreatment group,in 24 h (28.91±2.34,P＜0.05),36 h(19.36±0.1,P＜ 0.05) and 72 h(13.84±0.82,P＜0.05) was significantly decreased,the expression of IL-8 also decreased significantly.The mRNA expression of IL-4,IL-17,TNF-α and IFN-γ in CD4+ T cells was significantly increased after co-culturing with HUVECs.After the treatment with CYM-5442 group,the expression was decreased.Conclusion:DENV-2 could infect the primary HUVECs,and the expression of NS1 was inhibited after co-culturing with CD4+ T cells.CD4+ T cells can not only enhance the activation of HUVECs infected by DENV-2,but also can be activated by the infected HUVECs infected with DENV-2.

Objective To analyze the changes in the expression of Toll-like receptors 4 (TLR4) and 7 on the surface of murine bone marrow-derived dendritic cells following dengue virus type 2 (DEN2) infection.Methods DEN2 NGC strain was infected into BALB/c suckling mice through intracranial injection and injected into C6/36 cells to induce the in vivo and in vitro proliferation of DEN2, respectively.RT-PCR was performed to identify DEN2 RNA.Reed-Muench method was used to determine the 50% tissue culture infective dose (TCID50) of DEN2.Dendritic cells (DCs) were prepared by stimulating bone marrow cells isolated form C57BL/6 mice with IL-4 and GM-CSF and then identified by flow cytometry.The prepared murine bone marrow-derived DCs were infected with DEN2 and observed with direct immunofluorescence assay.Dynamic changes in the expression of CD11c, CD86 and I-A/I-E molecules on DCs after DEN2 infection were detected by flow cytometry.Levels of DEN2 RNA and the expression of TLR4 and TLR7 at mRNA level were dynamically detected by real-time quantitative PCR.Results The TCID50 of DEN2 to C6/36 cells was 10-5.8.Murine bone marrow-derived DCs were acquired with a purity of 70% and could be infected with DEN2 in vitro.The percentages of CD86 and I-A/I-E molecules on the surface of DCs infected with 1×105 TCID50 of DEN2 were statistically different from those of the negative control group.Neither of the two groups showed a significant difference in the percentages of membrane molecules over time.However, the percentages of membrane molecules on DCs increased with increasing viral load.Compared with the negative control group, the levels of DEN2 RNA in infection groups were increased with increasing virus load, while the expression of TLR4 and TLR7 on DEN2 infected-DCs at mRNA level was decreased with increasing viral load.Conclusion DEN2 infection promotes the maturation of DCs.Expression of TLR4 and TLR7 on DEN2 infected-DCs at mRNA level decreases with increasing viral load, which suggests that TLR4 and TLR7 are closely related to viral infection and play a certain role in the pathogenesis of DEN infection.

Vitamine E in peanut oil was determined by means of f Juorometric method (Ex = 285nm, Em=324nm) and TLC scanning method (? = 292nm). Both methods include saponification of the oil, isolation o'f the unsaponifiable matters and the respective final measurements. The result of the former method indicates the total vitamine E in terms of ?-vitamine E and the latter method may indicate their ?,?,? and 6 analogues separately. These two methods are simple, time-saving and reliable. The recovery of VE and the standard deviation of replication of fluorometric method and TLC scanning method are 99.3%, ? 0.587mg/100g sample and 103.04%, ?0.959mg/100g sample respectively. Both methods are also applicable to other foods such as fortified cakes, confectionaries, infant foods etc., but the oil in these samples must be extracted before saponification.

To study the value of high-fluorescent cell for the screening of malignant hydrothorax or ascites. The clinical diagnosis results and hydrothorax or ascites specimens of 400 outpatients registered from January to December in 2014 were collected, and high-fluorescent cells were counted with Sysmex XE-5000 blood cell analyzer, and then stained for microscopy. Receiver operating characteristic (ROC) curve was used to explore the sensitivity, speci-ficity, positive likelihood ratio and negative likelihood ratio of high-fluorescent cell for screening malignant hydrothorax or ascites. The numbers of high-fluorescent cells of the cancer patients were significantly higher than those of the non-cancer patients, with Z equal to -7.372 and P less than 0.05. The values of the area under curve (AUC), sensitivity, specificity, positive likelihood ratio and negative likelihood ratio were 0.801, 70.67%, 84.31%, 4.50 and 0.35 respective-ly. Detection of high-fluorescent may be used to screen malignant hydrothorax or ascites, and further exami-nation and follow-up have to be performed in case the number of high-fluorescent cells in the hydrothorax or ascites is not less than 68.

Twenty-three patients with pain from osteoporotic vertebral fractures,aged 65-90 yr,weighing 51-78 kg,received an image intensifier-assisted nerve-root block with a 6-8 ml mixture of 0.5 ％ lidocaine,mecobalamine 0.5 mg and betamethasone sodium phosphate injection 5.26 mg in a prone or lateral position.The VAS scores before operation,at 0,1 week,1 and 3 months after operation were 8.6 ± 0.9,1.5 ± 0.7,2.8 ± 0.9,1.6 ± 0.5 and 2.5 ± 0.7,respectively.VAS scores were significantly lower at each time pint after operation than before operation (P ＜ 0.05).According to modified MacNab standard,the effectiveness of treatment was rated as excellent/good in 87％ of the patients.No complication such as bleeding,hematoma,infection,pneumothorax,hemopneumothorax,headache was found during or after operation.Selective nerve-root block is effective in the treatment of pain resulting from osteoporotic vertebral fractures in patients.

Objective To investigate the effects of interaction between human umbilical vein endothelial cells (HUVECs) which were infected with dengue virus type 2 (DENV-2) and CD4+T cells on the expression of ICAM-1 (intercellular adhesion molecule 1),VCAM-1 (vascular cell adhesion molecule 1),IL-10 and TGF-β1 at mRNA level for further understanding the immunological mechanism of DENV infection.Methods HUVECs were treated with CYM-5442,a selective agonist for sphingosine-1-phosphate receptor 1 (S1P1),for 24 hours and then infected with 103 TCID50 (50% tissue culture infective dose) of DENV-2 before co-culturing with CD4+T cells.Changes in the expression of NS1 (DENV-2 nonstructural protein),SPHK1 (sphingosine kinase 1,phosphorylating sphingosine to S1P),ICAM-1,VCAM-1,IL-10 and TGF-β1 at mRNA level were detected by real-time PCR after 4,8,12,24,48 and 72 hours of co-culturing.Results There was a certain timeliness in the expression of NS1 at mRNA level after infecting HUVECs with DENV-2 and the expression reached a peak at 24 h.Treating HUVECs with or without CYM-5442 had no significant influence on the expression of DENV-2 NS1 at mRNA level.The expression of SPHK1 at mRNA level was significantly increased after treating HUVECs with CYM-5442 and DENV-2 (P<0.05).Compared with DENV-2-infected or untreated HUVECs,Co-culturing DENV-2-infected HUVECs with CD4+T cells increased the expression of ICAM-1 and VCAM-1 in HUVECs at mRNA level (P<0.01) as well as the expression of IL-10 in CD4+T cells at mRNA level (P<0.05),but had no significant influence on the expression of TGF-β1 in CD4+T cells at mRNA level.Conclusion This study shows that DENV-2 can replicate and proliferate in HUVECs,but CD4+T cells inhibit the replication and proliferation.CD4+T cells play an important role in promoting the expression of VCAM-1 and ICAM-1 in DENV-2-infected HUVECs at mRNA level,activating HUVECs and increasing inflammation,which may be associated with increased vascular permeability induced by DENV-2 infection.Co-culturing CD4+T cells with DENV-2-infected HUVECs promotes the expression of IL-10 in CD4+T cells at mRNA level,but has no significant effect on TGF-β1.

Objective To study the depressant effect of atorvastatin on arteriosclerosis of aortic allograft in rats.Methods The models of abdominal aorta transplantation were established in rats with the use of(micro-surgery).The recipients were divided into three groups:allograft control group,allograft experimental group and isograft control group.After 60 days of transplant,vascular intimal thickness(VIT) in all of the groups was observed by histological examination.The expression of PCNA and ?-SMA was determined by(immunohistochemistry).Results The degree of VIT in rats of the allograft experimental group was lower than that in the allograft control group;the VIT area ratio in the allograft control group,allograft experimental group and isograft control group was(12.40?2.65)%,(5.20?6.35)%,and(1.2?1.10)%,respectively,A statistical difference between these groups was observed(P

Objective To evaluate the surgical method and curative effect of minimally invasive percutaneous nephrolithotomy(MPCNL) in treatment of impacted proximal ureteric calculi.Methods 45 cases of proximal ureteric calculi treated with MPCNL were studied retrospectively.Percutaneous nephrostomy was established under the guide of X-ray,B-ultrasound and naked eye,and the nephropathy was extended to F16 through invasive percutaneous.Stones were taken out by ureteropyeloscopic pneumatic lithotripsy with help of hydraulic perfusion pump.Results The surgical time was 15～110 min with the average of 42 min after percutaneous nephrostomy.The average blood loss volume was 30 mL;no severe complications were occurred in the intra-or post-operative time.The rate of complete clearance of stones was 100% in first attempt.The mean hospital stay was 9 d,and the mean postoperative hospital stay was 3.5 d.Conclusion MPCNL used to treat the impacted proximal urethral calculi have satisfied effects and a lot of merits.

Objective To evaluate the effect of technical improvement of radical retropubic prostatectomy(RRP) for patients and reduction of complications.Methods 22 patients with clinically prostate cancer(mean age 67 years,stage B 19 cases,stage C 3 cases) underwent radical retropubic prostatectomy.During operation the dorsal vein complex,puboprostatic ligament,puborectalis sling,the neurovascular bundle,external urethral sphincter,reconstruction of the bladder neck were carefully managed.Results All the patients recovered uneventful except 2 deaths,and they survived and were followed up for a mean time of 16 months(range 1～4 years).The mean operative time was 140 minutes(range 90～220minutes) and the mean blood loss was 600mL(range 250～1500mL).16 cases received blood transfusion for 300～1200mL and no ureteral or rectal injury occurred.The urinary catheters were removed 2～3 weeks after operation.2 cases had stress incontinence 3 months later,1 case encountered urethral stricture and became well after urethral dilation,15 cases had satisfactory erectile function after operation and 3 patients had biochemical failure and endured continually chemotherapy during follow-up.Conclusions Accurate maneuver is handled according to correct anatomy can protect the apex of prostate and its surrounding tissue and reconstructing the bladder neck is helpful to reduce complications and achieve better outcome.

Objective To evaluate the surgical method and curative effect of minimally invasive percutaneous nephrolithotomy(MPCNL) in treatment of impacted proximal ureteric calculi and giant renal pelvic calculi with moderate or severe hydronephrosis by non-imaging in localization.Methods 52 cases of calculi treated with MPCNL were studied retrospectively.Percutaneous nephrostomy was established by non-imaging in localization(naked eye) according location of kidneys,length of 12th ribs,degree of hydronephrosis and condition of calculi,and the nephropathy was extended to F16 through invasive percutaneous.Stones were taken out by ureteropyeloscopic pneumatic lithotripsy with the help of hydraulic perfusion pump.Results The operative time was 32~140min with the average of 112min.The average blood loss volume was 45mL.No severe complication occurred in the intra-or post-operative time.The rate of complete clearance of stones was 96 % in first attempt,and the residual stones passed out by themselves after the double "J" tube was pulled out in cases.The mean hospital stay was 9 days,and the mean postoperative hospital stay was 4.2 days.Conclusion To treat the impacted proximal ureteric calculi and giant renal pelvic calculi with moderate hydronephrosis or severe hydronephrosis,some skilled doctors can appropriately use the method of MPCNL by non-imaging in localization,since the method can reduce the harm by X-ray,and save medical expenses.