Objective To transfect PCDNA3.0-HIF1-? into the endothelial progenitor cells(EPCs),in order to provide the foundation for exploring myocardial vascular repairing capacity and angiogenesis of the EPCs transfected with HIF gene.Methods The EPCs were separated by the method of density gradient separation in the human bone marrow,and then the cells were inoculated in the fibronectin-packaged petri dishes complete medium with the concentration of 1?106?mL-1,and cultivated for 14 d. The expressions of the endothelial cell-specific components ecNOS and flk-1 were detected with RT-PCR; the cells were stained with acLDL-Dil and FITC-UEA-1; the PCDNA3.0-HIF1-? plasmid was transfected into the EPCs mediated by LipofectamineTM 2000,the transcription of HIF1-? was determined with RT-PCR; the expression of VEGF was detected with enzyme-linked immunosorbent assay (ELISA); the expression of HIF1-? was detected with Western blotting.Results After cultivated for 5 d,some round monocytes transformated into spindled inherent EPCs;7 d later,there were some cell colonies containing dozens of cells;10 d later,there were obvious cell clones; RT-PCR showed ecNOS band in 548 bp,flk-1 band in 819 bp; acLDL-Dil and FITC-UEA-1 double-staining was positive; RT-PCR showed specific band in 383 bp; ELISA showed the content of VEGF in the supernatant of EPCs transfected with HIF1-? was (20.53?2.33) ?g?L-1,and it was (3.96?1.67) ?g?L-1 in the EPCs transfected without HIF1-?,there was significant difference between two groups (P

Objective To discuss the feasibility and effect of labeled antibody on targeting therapy of systemic lupus erythematosus(SLE) associated thrombocytopenia.Methods MTX-IVIG conjugate was prepared by indirect and direct cross linking,and the binding ability of it to Fc fragment was detected by indirect immunofluor-escence.The killing activity of the conjugate was detected by MTT method using murine macrophage with Fc receptor and strain U937 of human monocytic leukemia cell as targets.Results Conjugation showed stronger cytotoxicity upon target cells than free MTX,and it showed less cytotoxic effect on Fc receptor negative cells compared with the positive ones.IVIG-HSA-MTX restrained the phagocyte of macrophage.The killing effect of IVIG-HSA-MTX was significantly stronger than that of IVIG-MTX.Conclusion The conjugation can show a highly specific cytotoxicity upon mononuclear-macrophage in vitro.

Objective To construct the eukaryotic expression vector pEGFP-N1/IL-37b and analyze the expression of IL-37 gene in THP-1 cells. Methods Total RNA was extracted from human peripheral blood mononuclear cells ( PB-MCs) and the coding region of IL-37b gene was amplified by RT-qPCR. Then, the gene was cloned into pEGFP-N1 eu-karyotic expression vector. After transfected the recombinant plasmid into THP-1 cells, the expression of IL-37 was detec-ted by RT-qPCR and Western blot. Results Double restriction enzyme digestion and gene sequencing showed that IL-37b gene was correctly inserted into the eukaryotic expression vector pEGFP-N1. RT-qPCR and Western blot showed that the IL-37 expression level was increased significantly (P<0. 01) after transfection in THP-1 cells. Conclusions We successful-ly constructed a novel anti-inflammatory cytokine IL-37 eukaryotic expression vector pEGFP-N1/IL-37b, which lays a foun-dation for further study on IL-37 functions and its association with related diseases.

The over-expression of Polo-like kinase 1(Plk1)is critical in the producing and progressing of multi-ple human tumors and is recognized as an effective target for the development of novel anti-cancer drugs.Currently a variety of small molecules targeting ATP or substrates binding sites have entered different stages of clinical trials.Polo-box domain(PBD)is a unique domain of Plks which plays an important role in the sub-cellular location of Plks and also in the recognition of their substrates,therefore it has become an attractive target for the development of novel target-directed Plk1 inhibitors.In this paper,PBD function of Plk1 was intro-duced,the progress of small molecule and phosphoserine /phosphothreonine contained short peptide Plk1 inhibi-tors targeting PBD was summarized.Further development of this kind of inhibitors was also proposed.

Objective: To investigate the alteration of bcl- 2 gene family in the rat brain and the molecular mechanism of neuronal apoptosis following traumatic brain injury. Methods: Male Sprague -Dawley rats were subjected to lateral fluid percussion brain injury(FPI) of mo derate severity. Bcl-2, Bcl-x and Bax protein expression was detected by immun ohistochemistry. Results: (1) The immunoreactivity of Bcl-2 and Bcl-x protein decreased in the hippocampus ipsilateral impact site as early as 6 h post-injury, and this was the main cause of down-regulation of the ratio of Bcl-2+Bcl-x to Bax. (2) During 1-3 d after injury, the Bax protein express i on increased significantly, while the Bcl-2 and Bcl-x protein expression decre ased relatively slow. The decreased ratio of Bcl-2+Bcl-x to Bax was mainly due to the Bax up-regulation. Conclusion: The bcl-2 gene family is involved in neuronal apoptosis after FBI, and the protein expression alteration of the family members leads the neuronal cell to apoptosis.

ObjectiveTo retrospectively analyze clinical data of patients who has left-side valvular disease combined with severe tricuspid regurgitation and evaluate the effect of our modified tricuspid annuloplasty with enforcement of artificial felt strip.Methods76 patients who had left-side valvular disease combined with severe tricuspid regurgitation received operations between Jan.2008 and Jun.2010.The average age of the patients was 53.5 years old (32 male and 44 female).Besides the severe tricuspid regurgitation, other combined cardiac impairments included mitral valvar disease (52 cases), aortic valvar disease(5 cases), double valvar disease(19 cases) and left atrial thrombosis(22 cases).6 patients had grade II cardiac function according to the NYHA criteria, while 47 and 23 were in grade III and IV, respectively.Other signs included cyanosis(5cases), jaundice(11 cases), neck vein engorgement(48 cases) , ascites(22 cases), hepatomegaly(41 cases) and pitting edema in the lower limbs(68 cases).The concomitant operative procedures included mitral valve replacement in 52 patients,aortic valve replacement in 5 patients, double valve replacement in 19 patients, removal of left atrial thrombus in 22 patients,left atrium folding in 21 patients and left atrium appendage suture in 68 patients.Left-sided valve disease were corrected first,TAP was performed on the beating heart after the heart had been defibrillated.The anteroseptal commissure was plicated first.A double-armed 3-0 pledgeted suture was taken through the base of the septal leaflet, 5-6 mm from the commissure, extending along the annulus, and out from the point in the anterior annulus 10-12 mm from the anteroseptal commissure.Both ends of the suture was tied until the two Teflon pledgets approximated each other near the commissure.Then a semicircular De Vega type of plicating with a 3-0 prolene was taken, starting just from the anterior annulus near the anteroposterior commissure, and extending clockwise to a point just cephalad to the posteroseptal commissure.The suture was tied with positioning a 27-29 mm valve siser across the tricuspid valve.At last, a 3-5 mm width felt strip was prepared and was sutured to the plicated posterior annulus region with interrupted mattress sutures of 2 to 3 2-0 prolene.A favorable result was considered when TR was not marked by saline injection.Echocardiography was routinely examined one week postoperatively and patients were followed up 6 month after discharge.ResultsThere is no death in all patients.The CVP diminished significantly from 16mmHg preoperatively to 8mmHg postoperatively (P = 0.0021).The systomic pulmonary pressure diminished from 59 mmHg preoperatively to 41 mmHg postoperatively (P = 0.038).Echo one week postoperative showed no tricuspid regurgitation in 56 patients and mild in 18 patients, while 2 had moderate tricuspid regurgitation.The diameter of right atrium diminished significantly postoperatively, too.The ejection fraction was improved even though there was not significant difference as compared with preoperative data.The cardiac function of all patients improved and the signs of right heart failure were alleviated or disappeared.Follow up 1 to 36 months showed no change of the regurgitation except for one become moderate from mild when discharged.No hepatic congestion or edema was observed in all patients.ConclusionThese new modifications make the technique more selective in the remodeling of the tricuspid annulus.It could achieve better coaptation of the anterior leaflet with the others, successful annular reduction, better maintenance of the contractile property of the tricuspid ring, better distribution of pursing force in the more dilated region.It could prevent the tear of the endocardium in the posteroseptal region in the long period of time postoperatively.

Objective:To conjugate IVIG and MTX to produce a specific cytotoxicity upon phagocytes.Methods :MTX was conjugated with IVIG by indirect conjugating methods. HSA was used as an intermedi-ary to conjugate MTX with IVIG. The indirect immunofluorescence was adopted to test the binding abilityof Fc fragment. MTT assay was used to measure the cytotoxicity of conjugation on phagocytes. Results:Conjugation showed stronger cytotoxicity upon target cells than free MTX,and it showed only less cyto-toxic effect on Fc receptor negative cells compared with the positive ones. The specific cytotoxicity of IVIG-HSA-MTX was significantly stronger than that of MTX. Conclusion: In vitro the conjugation showed ahighly specific cytotoxicity upon phagocytes.

Objective To evaluate the effects of different smoking cessation interventions on cigarette consumption for young soldiers.Methods Sixty-eight soldiers were prospectively investigated in this randomly controlled clinical trial and assigned to the psychological intervention group,auricular acupuncture group,and smoking cessation medication group.Results All the participants showed significant reduction(33.3％ to 73.9％)in post-treatment cigarette consumption.The highest quit rate was found at 7 days,although this declining trend was faded over time.One-year follow-up indicated a 6-month quit rate of 16.7％,23.8％ and 30.4％ in three groups,respectively.Conclusion Psychological intervention,auricular acupuncture and smoking cessation medication may be effective methods of reducing cigarette consumption and improving quit rate in young soldiers.

AIM: To examine the expression of miRNA-22 in the ovarian tissues and the effect of miRNA-22 over-expression on the proliferation, migration and invasion in SKOV-3 cells.METHODS: The expression levels of miRNA-22 in different ovarian tissues and SKOV-3 cells were determined by qPCR .miRNA-22 was over-expressed by trans-fection of miRNA-22 mimic.The cell viability was examined by CCK-8 assay.The cell migration was measured by wound healing test .The cell invasion was analyzed by Transwell assay .The protein expression levels of VEGF and P 53 were deter-mined by Western blot .RESULTS: Compared with the normal ovarian tissue , the expression level of miRNA-22 was remarkably decreased in the ovarian tumor tissues .After transfection with miRNA-22 mimic, the expression level of miRNA-22 in the SKOV-3 cells was significantly increased , while the cell viability , migration and invasion were obviously decreased .Moreover , the protein expression of VEGF and P 53 was dramatically inhibited after over-expression of miRNA-22.CONCLUSION:The decreased miRNA-22 expression may be correlated with the development of ovarian can-cer.Over-expression of miRNA-22 decreases the cell viability , migration and invasion by reducing the protein expression of VEGF and P53.

HER2, a member of epidermal growth factor receptor family proteins, is overexpressed in about 30% of human breast cancer. Increased levels of HER2 are associated with poor patient prognosis and enhanced metastasis. RNA interference (RNAi) is developed recently as a new technique which can inhibit gene expression specifically in mammalian cells. On the basis of previous study,in which two target sequences with favorable RNAi effect on HER2 were identified, a series of dual promoter siRNA-expressing vectors containing two opposing U6 and H1 promoters were constructed. After transfection of HER2-overexpressing SKBR3 breast cancer cells with the siRNA-expressing vectors, downregulation of HER2 was identified quantitatively. Subsequently, the siRNA-expressing cassettes were subcloned into lentiviral vectors by LR recombination reaction and lentivirus was prepared successfully. The results from infection of SKBR3 cells with siRNA-expressing lentivirus demonstrated that lentiviral-mediated RNAi could downregulate HER2 expression efficiently through fluorescent quantitative PCR (FQ-PCR), western blot, and FACS analysis. Furthermore, cell growth was inhibited in cell proliferation assay after treatment with siRNA lentivirus.A new tool for clarifying the function of HER2 in cancer metastasis and developing the gene therapy drug was offered.