OBJECTIVETo study the effect of vacuum-assisted closure (V.A.C) on the expression of Urokinase-type plasminogen activator (uPA) and Urokinase-type plasminogen activator receptor(uPAR) protein in margin tissue of pigs with acute wounds and patients with chronic wounds.

METHODSAcute wounds were created on the two side of five male pigs' back, the experiment wounds on one side received V. A. C treatment and the control side received traditional treatment. Punch biopsies were taken from margin tissue of the wounds in 0, 1, 3, 6, 9, 12, 18, 25 days after the V.A.C treatment. The uPA and uPAR positive cells were stained with immunohistochemical technique . Six human chronic wounds were also treated with the V. A. C treatment, and the samples of extravasate from those wounds were collected in 0, 1, 3, 5, 7 days after the treatment, and the levels of uPA and uPAR expression were examined by enzyme-linked immunosorbent assay (ELISA).

RESULTSThe expression of uPA and uPAR protein in margin tissues of pigs with acute wounds increased and peaked in 3 days after the treatment with V. A. C, then it presented rapidly downtrend, but the expression and staining in the experiment group were obviously higher than that of the control group. In the six chronic wounds, the high level expression of uPA and uPAR protein was decreased after the treatment with V. A. C.

CONCLUSIONThe V. A. C may increase the expression of uPA and uPAR protein in acute wound keratinocytes and decrease the high expression of uPA and uPAR in chronic wounds.

Adult ; Animals ; Female ; Humans ; Male ; Middle Aged ; Negative-Pressure Wound Therapy ; Receptors, Urokinase Plasminogen Activator ; metabolism ; Swine ; Urokinase-Type Plasminogen Activator ; metabolism ; Wound Healing

OBJECTIVETo study the effects of VAC on starting the process of wound healing and decreasing apoptosis.

METHODSTo examine the variations in expression of proto-oncogenes c-myc, c-jun and Bcl-2 in pig wound model with acute full-thickness skin defect and human chronic wounds by immunohistochemistry, calculate the numbers of expressive positive cells and the labelling index (LI), and observe the process of wound healing.

RESULTS(1) In pig experiment, the wound in experimental group was very clean and without obvious exudates, many neoepiderm and granulation tissue rapidly appeared or formed after 6 days, and healed completely by the 25th day. On the contrary, in the wound of control group, more exudates and blood crust could be seen and fewer neoepiderm and granulation tissue appeared after 6 days and was healed by 30th day. Immediately after the wound was created, the expression of c-myc, c-jun and Bcl-2 was lower and mainly situated in nucleus or cytoplasma of the basilar cells. After the wound was created in control group, or after starting the VAC treatment in experimental group, their expression rapidly and obviously increased, the distribution of the positive cells also became enlarged, but the amount of expression decreased rapidly after the expressive peak have reached. In the successive 12 days following the wound was created, the expression of c-myc, c-jun and Bcl-2 in the experimental group was constantly higher than that of the control group. (2) In human chronic wounds, there wasn't obvious secretions and more healthy granulation tissue was rapidly formed after VAC treatment. The expression of c-jun was mainly located in cytoplasma of basilar cells of epithelium, dermal fibroblasts and inflammatory cells, and the positive cell and labelling index obviously decreased. The expression of c-myc and Bcl-2 was mainly in cytoplasma of basilar cells, but the amount of expression and the labelling index became obviously increased after VAC treatment.

CONCLUSIONSVAC could rapidly start the healing course of the pig' s acute skin wound and human chronic wound, decrease apoptosis of the reparative cells, so as to accelerate wound healing.

Adult ; Animals ; Apoptosis ; Female ; Humans ; Male ; Middle Aged ; Negative-Pressure Wound Therapy ; Proto-Oncogene Proteins c-jun ; metabolism ; Proto-Oncogenes ; Swine ; Wound Healing

Abstract: Our previous work revealed berberine can significantly enhance the susceptibility of fluconazole against fluconazole-resistant Candida albicans, which suggested that berberine has synergistic antifungal activity with fluconazole. Preliminary SAR of berberine needs to be studied for the possibility of investigating its target and SAR, improving its drug-likeness, and exploring new scaffold. In this work, 13-substitutited benzyl berberine derivatives and N-benzyl isoquinoline analogues were synthesized and characterized by 1H NMR and MS. Their synergetic activity with fluconazole against fluconazole-resistant Candida albicans was evaluated in vitro. The 13-substitutited benzyl berberine derivatives 1a-1e exhibited comparable activity to berberine, which suggested that the introduction of functional groups to C-13 can maintain its activity. The N-benzyl isoquinolines, which were designed as analogues of berberine with its D ring opened, exhibited lower activity than berberine. However, compound 2b, 2c, and 4b showed moderate activity, which indicated that berberine may be deconstructed to new scaffold with synergistic antifungal activity with fluconazole. The results of our research may be helpful to the SAR studies on its other biological activities.
Antifungal Agents ; pharmacology ; Berberine ; pharmacology ; Candida albicans ; drug effects ; Drug Resistance, Fungal ; Drug Synergism ; Fluconazole ; pharmacology ; Isoquinolines ; pharmacology ; Microbial Sensitivity Tests

OBJECTIVETo assess if the modulating effect of platelet-derived growth factor (PDGF)-BB on p21(WAF1) was mediated by upregulating transforming growth factor (TGF)-beta(1) expression in vascular smooth muscle cells (VSMC).

METHODSTGF-beta(1) mRNA and protein expressions were measured by reverse transcription-PCR and ELISA, the protein expressions of p21(WAF1) and the downstream TGF-beta signalling including TGF-beta type I receptor (ALK-5 in VSMC), Smurf2, pSmad2/3, Smad4, Smad7 were detected by Western blot.

RESULTSPDGF-BB significantly upregulated the expressions of TGF-beta(1) at mRNA (0.79-fold) and protein (1.98-fold) levels in VSMC, significantly inhibited the expression of p21(WAF1) (-67 +/- 12)%, and enhanced the expressions of ALK-5, pSmad2/3, Smad4, Smurf2 protein by 1.21-fold, 0.95-fold, 0.69-fold and 2.55-fold respectively, inhibited Smad7 expression (-65 +/- 9)%, these alterations were partially restored by anti-TGF-beta(1) neutralizing antibody.

CONCLUSIONSThese findings suggested that PDGF-BB inhibited p21(WAF1) expression in VSMC partially through upregulating TGF-beta(1) expression via PDGF-BB and TGF-beta signalling pathways.

Animals ; Cell Division ; Cells ; Cyclin-Dependent Kinase Inhibitor p21 ; metabolism ; Male ; Muscle, Smooth, Vascular ; cytology ; metabolism ; Myocytes, Smooth Muscle ; drug effects ; metabolism ; Platelet-Derived Growth Factor ; pharmacology ; Proto-Oncogene Proteins c-sis ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; drug effects ; Transforming Growth Factor beta1 ; metabolism

OBJECTIVETo investigate the clinical effects and prevent the complications of posterior and anterior decompression and internal fixation in the revision of cervical anterior internal fixation failure.

METHODSFrom 2008 January to 2011 December, 17 patients with cervical anterior internal fixation failure were treated with posterior and anterior decompression and internal fixation. There were 12 males and 5 females, aged from 26 to 68 years old with an average of 44.1 years. The lower screw loosening was found in 6 cases, the upper screw loosening in 5 cases, titanium mesh caving in 3 cases, the upper screw breakage in 2 cases, the lower screw breakage in 1 case. Informations of bone fusion were observed by X-ray, CT, MRI. Clinical effects were evaluated by modified JOA score.

RESULTSAll the revision operations were successfully completed. One case with poor blood coagulation function before operation resulted in postoperative hematoma and occurred neurological symptoms; after hematoma removal and fresh frozen plasma infusion later, neurological symptoms of the patient disappeared. All patients were followed up from 6 to 38 months with an average of (22.4±10.0) months. Postoperative at 2 weeks, 3 months, and final follow-up, JOA score had obviously improved and respectively was 13.1±1.6, 13.4±1.6, 14.2±1.5. All internal fixation locations were good after revision,and obtained bone fusion at 10 months after operation, with an average fusion time of 6 months.

CONCLUSIONThe combined posterior and anterior decompression and internal fixation in the revision of cervical anterior internal fixation failure is safe, can achieve thoroughly decompression, maintain the cervical curvature, reconstruct the three column stability, and it may be used for the patients of cervical anterior fixation failure.

Adult ; Aged ; Cervical Vertebrae ; surgery ; Decompression, Surgical ; methods ; Female ; Fracture Fixation, Internal ; methods ; Humans ; Male ; Middle Aged

OBJECTIVETo study the cytotoxic T lymphocyte (CTL) response induced by dendritic cells (DC) transduced with recombinant adenovirus vector bearing hepatitis B virus surface antigen (HBsAg) gene in hepatocellular carcinoma HepG2. 2. 15 cells in vitro.

METHODSFull length HBsAg cDNAs were subcloned into pIND vector, followed by being cloned into pShuttle vector. The HBsAg gene fragments resulted from the pShuttle-S digested with PI-Sce and I-Ceu were linked to the linear adeno-X virus DNA. After packaged with HEK293 cells, the adenovirus expression vector was obtained. Then the recombinant adenovirus expression plasmid AdVHBsAg was transfected into human monocyte-derived dendritic cells, to construct AdVHBsAg hepatocarcinoma tumor vaccine. The effectiveness of transfection was detected by Western blot. Surface molecules of AdVHBsAg-DC were detected by FACS. Autologous T cell proliferation stimulated by AdVHBsAg-DC was detected by 3H-TdR assay. Cytotoxic CTL activity induced by AdVHBsAg-DC in vitro was detected by LDH assay.

RESULTSHBsAg gene in the inserted DNA of AdVHBsAg was confirmed by PCR, and predictive fragments proved by restriction enzyme digestion analysis were exhibited. Cell pathological changes appear after 10 days HEK293 cells transfected AdVHBsAg. Western blot analysis showed that HBV surface antigen gene was expressed in transfected DC, indicating that the transfection was effective. AdVHBsAg-DC was able to upregulate CD1a, CD11c, CD80, CD86 and HLA-DR. Autologus T cell proliferation induced by AdVHBsAg-DCs was significantly higher than that in DC control group and LacZ-DC group (P < 0.05). AdVHBsAg-DC activated CTL presented the specific killer ability to the hepatocellular carcinoma cells expressing HBsAg.

CONCLUSIONDC transduced with recombinant adenovirus HBsAg can express HBV-related hepatocellular carcinoma antigen (HBsAg), and AdVHBsAg-DC can induce potent immune response against HBsAg-positive hepatocellular carcinoma cells in vitro.

Adenoviridae ; genetics ; Antigens, CD1 ; metabolism ; CD11c Antigen ; metabolism ; Cancer Vaccines ; immunology ; Carcinoma, Hepatocellular ; immunology ; pathology ; virology ; Cell Line, Tumor ; Cell Proliferation ; Cytotoxicity, Immunologic ; immunology ; Dendritic Cells ; cytology ; immunology ; metabolism ; Genetic Vectors ; Hepatitis B Surface Antigens ; genetics ; metabolism ; Humans ; Liver Neoplasms ; immunology ; pathology ; virology ; Plasmids ; Recombinant Proteins ; genetics ; metabolism ; T-Lymphocytes, Cytotoxic ; cytology ; immunology ; Transfection

Objective:To observe the effect of acupuncture in regulating ubiquitin-proteasome pathway (UPP),and discuss the action of acupuncture in intervening heroin-induced brain damage.Methods:Thirty male Sprague-Dawley (SD) rats were divided into a control group,a model group and an acupuncture group by using the random number table.Rats in the model and acupuncture groups received intramuscular heroin injection for successive 8 d at a progressively increased dose.Afterwards,the injection was suspended for 5 d for withdrawal.The heroin relapse rat model was established by repeating the drug addiction and withdrawal process for 3 times.The control group followed the step of the model establishment,but was given intramuscular injection of normal saline at the stage of addiction and no intervention at the stage of withdrawal;the model group was given intramuscular heroin injection at a progressively increased dose at the addiction stage and no intervention at the withdrawal stage;the acupuncture group was dealt in the same way as the model group at the addiction stage,but received acupuncture at Baihui (GV 20) and Dazhui (GV 14) at the withdrawal stage,with the needles retained for 30 min each time,1 session a day,for successive 5 d.On the 39th day,brain tissues were extracted from the hippocampus and ventral tegmental area (VTA) of the three groups of rats.The apoptosis of brain nerve cells was detected by using terminal deoxynucleotidyl transferase-mediated nick and labeling (TUNEL).The mRNA and protein expressions of ubiquitin (Ub),ubiquitin protein ligase (E3) and 26S were examined by immunohistochemistry and quantitative real-time polymerase chain reaction (RT-qPCR).Results:Compared with the model group,rat's hippocampus and VTA in the acupuncture group showed significantly fewer cells positively stained by TUNEL staining (P＜0.01),and its mRNA and protein expressions of Ub,E3,26S were significantly lower (P＜0.01).Conclusion:Reducing nerve cell apoptosis and regulating the mRNA and protein expressions of Ub,E3 and 26S in rat's hippocampus and VTA are possibly one of the action mechanisms of acupuncture in intervening heroin-induced brain damage.

Abstract:One H5N1 subtype avian influenza virus, A/duck/Shandong/009/2008 (Dk/SD/009/08), was isolated from apparently healthy domestic ducks in some live bird market in East China during our epidemiological surveillance. To investigate the genetic composition, Dk/SD/009/08 was subjected to genome sequencing. The amino acid motif of cleavage site was "PLRERRRK-R/GL", which was consistent with the characterization of the HPAIV. According to the newest unified nomenclature system of H5N1, Dk/SD/ 009/08 was classified into Clade 2.3.4. The BLAST results showed that four gene segments (HA, NA, NP and NS) had the highest nucleotide identities with H5N1 subtype AIVs whereas the remaining four (PB2, PB1, PA and M) displayed the closest relationship with H9N2 subtype. Therefore, Dk/SD/009/08 might be a natural reassortant virus. The phylogenetic analysis further indicated that G1-like H9N2 subtype AIVs which was prevalent mainly in quails of Southern China might provide the internal genes for Dk/ SD/009/08.
Animals ; Chick Embryo ; Genome, Viral ; Humans ; Influenza A Virus, H5N1 Subtype ; classification ; genetics ; isolation & purification ; Influenza A Virus, H9N2 Subtype ; classification ; genetics ; isolation & purification ; Influenza, Human ; virology ; Molecular Sequence Data ; Phylogeny ; Reassortant Viruses ; classification ; genetics ; isolation & purification ; Recombination, Genetic ; Viral Proteins ; genetics

This study was aimed to investigate the prophylactic effect of CsA, MTX and MMF combined with ATG on graft versus host disease (GVHD) after unrelated donor peripheral blood hematopoietic stem cell transplantation (URD-PBHSCT). 33 patients underwent URD-PBHSCT with unrelated donor of HLA matched or 1 locus mismatched. The clinical data of all cases were retrospectively analyzed. URD-PBHSCT recipients received cyclosporin A+short term MTX+mycophenolate mofetil (MMF)+antithymocyte globulin to prevent GVHD (URD-ATG group), while 13 out of 33 patients were given additionally anti-CD25 antibody (URD-ATG+CD25 group). The results showed that engraftment was successfully achieved in 100% of all patients. In URD-ATG+CD25 group and URD-ATG group, the incidence of aGVHD were 23.07% and 45%, the incidence of cGVHD were 0 and 47.4% respectively. The latter was significantly higher than the former (p<0.05). The relapse rate in URD-ATG+CD25 group and URD-ATG group were 53.84% and 15% respectively, the former was significantly higher than the latter (p<0.05). The analysis on disease status of patients before transplantation displayed that the relapse rate of patients in progression status was significantly higher than that of patients in stable status (p<0.01), while the relapse rate of patients in progression status in URD-ATG+CD25 group reached to 100%. The overall survival (OS) at 1 year of patients in URD-ATG+CD25 group and URD-ATG group were 53.8% and 75% respectively, the OS at 5 years of patients in URD-ATG+CD25 group and URD-ATG group were 38.8% and 65% respectively, the OS of patients in URD-ATG group was higher than that of patients in URD-ATG+CD25 group (p<0.05). Simultaneously the OS of patients with progression status before transplantation declined obviously. It is concluded that adopting CsA+MTX+MMF+ATG as the prophylaxis of GVHD for UDR-PBSCT is effective. Reducing the dose of ATG may be good for patient in progression status.
Adolescent ; Adult ; Antilymphocyte Serum ; therapeutic use ; Cyclosporine ; therapeutic use ; Female ; Graft vs Host Disease ; prevention & control ; Humans ; Immunosuppressive Agents ; therapeutic use ; Male ; Methotrexate ; therapeutic use ; Middle Aged ; Mycophenolic Acid ; analogs & derivatives ; therapeutic use ; Peripheral Blood Stem Cell Transplantation ; Retrospective Studies ; Young Adult

This study was aimed to investigate the curative efficacy of allogenetic hemopoietic stem cell transplantation (allo-HSCT) using FLAG and modified BUCY conditioning regimen for patients with refractory and relapse or high risk hematologic malignancies. The therapeutic effect of allo-HSCT with FLAG and modified BUCY conditioning regimen on 10 patients with hematologic malignancies was analyzed. 10 patients included 2 cases of relapse (1 early relapse, 1 progression) and 7 cases of refractory (2 CR(2), 1 CR(3) and 2 PR and 1 NR) and 1 CR(1) with high risk. Among 10 cases 8 cases was diagnosed as AML, 1 case as pro-T lymphoblast lymphoma/leukemia and 1 case as spleen marginal zone lymphoma. The results showed that implantation of all the patients was successful. The relapse-free median survival time of 8 cases was 164 days (57 - 442 days) and survived up to now, 2 cases died (1 case died of pulmonary infection, 1 case died of fungus pulmonitis). In conclusion, the curative efficacy of allo-HSCT using FLAG and modified BUCY conditioning regimen for patients with refractory and relapsed hematologic malignancies is satisfactory.
Adult ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Female ; Hematologic Neoplasms ; surgery ; therapy ; Hematopoietic Stem Cell Transplantation ; methods ; Humans ; Male ; Middle Aged ; Transplantation Conditioning ; methods ; Young Adult