Objective: To investigate the effect of protein transduction domain-oligomerization domain-hemagglutinin (PTD-OD-HA) fusion proteins on tumorigenic ability of chronic myeloid leukemia (CML) BaF3-P210 cells in mice. Methods: The untreated BaF3-P210 cells and BaF3-P210 cells treated with 40 μmol/L PTD-OD-HA for 48 h were injected into BALB/c mice through the tail vein, respectively. The general status and survival time of mice in each group were observed. The number of peripheral white blood cells (WBCs) was counted. The Wright-Giemsa-stained blood and bone marrow smears were examined. The pathological changes of liver, spleen and lung tissues were observed by HE staining. The expression level of bcr/abl protein in bone marrow cells from mice was determined by Western blotting. Results: The incidence rates of CML in mice in the untreated BaF3-P210 cell group and the PTD-ODHA-treated BaF3-P210 cell group were 90% (9/10) and 80% (8/10), respectively; WBCs counts in the two groups were (44.3±4.8)×109/L and (20.6±3.2) ×109/L (P<0.05), respectively; the expression levels of bcr/abl protein in bone marrow cells in the two groups were 5.13±0.46 and 1.32±0.29 (P<0.05), respectively. The average survivals in the untreated BaF3-P210 cell group and the PTD-OD-HA-treated BaF3-P210 cell group were (101.3±6.2) d and (185.4±8.7) d (P<0.05), respectively. Wright-Giemsa staining showed that the infiltration degree of leukemic cells in bone marrow, liver and spleen was lower in the PTD-OD-HA-treated BaF3-P210 cell group than that in the untreated BaF3-P210 cell group. Conclusion: PTD-OD-HA can inhibit the tumorigenic ability of CML with BaF3-P210 cells in BALB/c mice. Copyright© 2011 by TUMOR.

Objective: To clone hGLP-1 cDNA in the pBS SK(+/-)vector and construct the expression vector of pGEX-4T-3/hGLP-1cDNA to express GST-hGLP-1 fusion protein. Methods: The hGLP-1 cDNA was constructed by 6 synthetic oligonucleotides fragments, followed by the procedure of annealing and ligation with oligonucleotides fragments. The hGLP-1 cDNA was cloned into the pBS SK(+/-) vector, and was selected by α-complementation. It was confirmed by DNA sequening, then inserted into the MCS of the fusion expression vector pGEX-4T-3. The recombinant vector was transformed into E. coli TG1. Results: The recombinant plasmid DNA was digested with restrictive endonuclease BamHⅠand XhoⅠ. The result demonstrated that the hGLP-1 cDNA was successfully inserted into the pGEX-4T-3 vector and fusion protein GST-hGLP-1 had been expressed in SDS-PAGE. Conclusion： Expression of GST-hGLP-1 fusion protein can provide foundation for obtaining a larger quantity of recombinant hGLP-1 for experimental and clinic studies.

Objective: To clone hGLP-1 cDNA in the pBS SK(+/-)vector and construct the expression vector of pGEX-4T-3/hGLP-1cDNA to express GST-hGLP-1 fusion protein. Methods: The hGLP-1 cDNA was constructed by 6 synthetic oligonucleotides fragments, followed by the procedure of annealing and ligation with oligonucleotides fragments. The hGLP-1 cDNA was cloned into the pBS SK(+/-) vector, and was selected by α-complementation. It was confirmed by DNA sequening, then inserted into the MCS of the fusion expression vector pGEX-4T-3. The recombinant vector was transformed into E. coli TG1. Results: The recombinant plasmid DNA was digested with restrictive endonuclease BamHⅠand XhoⅠ. The result demonstrated that the hGLP-1 cDNA was successfully inserted into the pGEX-4T-3 vector and fusion protein GST-hGLP-1 had been expressed in SDS-PAGE. Conclusion： Expression of GST-hGLP-1 fusion protein can provide foundation for obtaining a larger quantity of recombinant hGLP-1 for experimental and clinic studies.

OBJECTIVETo optimize the solid-phase extraction method by comparison of the extraction recovery of ginsenoside Re plasma samples.

METHODAfter extracted by different solid-phase cartridges with water, acetonitrile, and different content methanol elution, the plasma samples were analyzed on an Zorbax SB-C18 column with acetonitrile-water gradient elution. From the recovery achieved, the best solid phase cartridge was found.

RESULTThis method consists of using 40% methanol as the wash solvent, and 80% methanol for the elution. Among the three kinds of solid-phase being tested, Waters Oasis HLB cartridge was found to be the best one.

CONCLUSIONThe average extraction recovery of the Waters Oasis HLB cartridges was between 103%-113%, it can be used in the analysis of ginsenoside Re in plasma samples.

Chromatography, High Pressure Liquid ; methods ; Ginsenosides ; blood ; isolation & purification ; Humans ; Panax ; chemistry ; Plants, Medicinal ; chemistry

OBJECTIVETo investigate reason and the management of portal vein thrombosis in patients with portal hypertension postoperatively.

METHODS329 patients with portal hypertension in liver cirrhosis who had splenectomy was reviewed from 1992 to 2001. In whom 43 (13.1%) patients with portal vein thrombosis postoperative were analyzed.

RESULTSIn these patients, except 1 died for portal vein phlebitis, all patients were recovered. There are 138 patients who underwent splenectomy or splenectomy and devascularization, 26 (18.8%) of them had thrombosis. 191 patients underwent splenectomy and portacaval or portasplenic shut, 17 (8.9%) of them had thrombosis. The data of these two groups have significant difference (chi(2) = 8.44, P < 0.01).

CONCLUSIONSThrombocytosis postsplenectomy as well as the changes of portal hemodynamics is the main reason of portal vein thrombosis. Portal vein thrombosis is also in association with the operative ways. Operation standardization, dynamic examining platelet count, routine color ultrasonography examining and early anticoagulation therapy are the effective methods in preventing and managing portal thrombosis postoperation for portal hypertension.

Adult ; Budd-Chiari Syndrome ; etiology ; prevention & control ; therapy ; Female ; Follow-Up Studies ; Humans ; Hypertension, Portal ; surgery ; Male ; Middle Aged ; Portal Vein ; pathology ; Postoperative Complications ; Vascular Surgical Procedures ; adverse effects ; methods

Objective To observe the curative effect of laparoscopic and open radical gastrectomy for gastric cancer patients and the impact on patients' CD4+CD25+T cells, oxidative stress, IL-2 and soluble receptors. Methods 120 cases from November 2009 to September 2012 were enrolled. 60 patients received laparascopic radical gastrectomy (LRG group) and 60 patients received open radical gastrectomy (ORG group) as research object, the two kinds of operation mode were compared and the impact of operation on patients' CD4+cells, oxidative stress, CD25+T IL-2 and its soluble receptor were recorded. Results The operation time, blood loss, number of lymph node dissection n LRG group were (285.8 ± 98.6), (343.3 ± 125.5), (7.8 ± 1.2), all the indexes were significantly superior to the ORG group and LRG group of patients with anastomotic leakage, intestinal obstruction, bile duct injury and pulmonary infection were 1, 1, 0, and 0 cases no difference in ORG group than MDA and AOPP, after the operation were increased, but the LRG group was lower than that of ORG group patients; postoperative antioxidant GSH-PX, SOD decreased, but LRG was higher than that of ORG group, and LRG group after treatment increased CD4+CD25+T cells in patients with the content of IL-2 of two groups after treatment, s-IL-2R increased than before LRG, group IL-2, s-IL-2R levels were (32.6 ± 4.2) and (23.6 ± 4.6) μg/ml was significantly lower than that of ORG group, P < 0.05, There was no difference in the 1, 2, 3, and 5 year survival rates between the two groups of patients. Conclusion Laparoscopic and open surgery for gastric cancer is equivalent, but also help to stabilize T cells, inhibit oxidative stress and inflammatory factors.

OBJECTIVETo prepare cytochrome (CYP)2E1-silenced hepatocytes by lentivirus-mediated RNA interference technology and to investigate the hepatotoxicity of trichloroethylene (TCE) in CYP2E1-silenced hepatocytes.

METHODSShort hairpin RNA fragments were designed and synthesized and were then ligated into the lentiviral vector; single colonies were screened; the plasmid was extracted after PCR and sequence identification and then transferred into L02 hepatocytes; the CYP2E1-silenced hepatocytes were selected; real-time quantitative PCR and Western blot were used to evaluate the interference effects. The obtained CYP2E1-silenced hepatocytes, as well as normal L02 hepatocytes, were treated with TCE (0, 0.25, 0.50, 1.00, 2.00, and 4.00 mmol/L). The cell viability and half maximal inhibitory concentration (IC50) of TCE were measured; the apoptotic rate of cells was measured by flow cytometry; the mRNA expression levels of apoptosis genes and oncogenes were measured by real-time quantitative PCR.

RESULTSThe IC50s of TCE for L02 hepatocytes and CYP2E1-silenced hepatocytes were 15.1 mmol/L and 23.6 mmol/L, respectively. The apoptotic rate increased as the dose of TCE rose in the two types of cells; the CYP2E1-silenced hepatocytes hada significantly lower apoptotic rate than L02 hepatocytes when they were exposed to 2.0 and 4.0 mmol/L TCE (P < 0.05 or P < 0.01). The mRNA expression level of bcl-2 (anti-apoptosis gene) in CYP2E1-silenced hepatocytes was 15% ∼ 60% higher than that in L02 hepatocytes (P < 0.01), while the mRNA expression levels of caspase-3 and caspase-9 (apoptosis genes) in CYP2E1-silenced hepatocytes were 30% ∼ 60% lower than those in L02 hepatocytes (P < 0.01). The mRNA expression level of p53 (cancer suppressor gene) in CYP2E1-silenced hepatocytes was 81 - 278% higher than that in L02 hepatocytes (P < 0.01), while the mRNA expression levels of c-fos and k-ras (oncogenes) in CYP2E1-silenced hepatocytes were 20-68% lower than those in L02 hepatocytes (P < 0.01).

CONCLUSIONCYP2E1-silenced cells can be successfully prepared by lentivirus-mediated RNA interference technology. Silencing CYP2E1 gene can reduce the hepatotoxicity of TCE and inhibit the expression of some apoptosis genes and oncogenes, suggesting that CYP2E1 gene plays an important role in TCE metabolism and is related to the hepatotoxicity of TCE.

Apoptosis ; drug effects ; genetics ; Cell Line ; Cell Survival ; drug effects ; genetics ; Cytochrome P-450 CYP2E1 ; genetics ; metabolism ; Genetic Vectors ; Hepatocytes ; drug effects ; metabolism ; Humans ; Lentivirus ; genetics ; RNA Interference ; Trichloroethylene ; toxicity

PURPOSE: Myeloid differentiation-2 (MD-2) has been associated with endotoxin and inflammatory disorders because it can recognize lipopolysaccharide (LPS) binding and attenuate Toll-like receptor 4 (TLR4)-mediated signaling. However, its role in allergic inflammation has yet to be clarified. We examined whether single nucleotide polymorphisms (SNPs) in MD-2 promoter can affect MD-2 expression and aimed to clarify the relationship between Der p 2 allergy and SNPs of MD-2 promoter. METHODS: The function of SNPs of MD-2 promoter and the effects of cytokines and immunoglobulin on the secretion and mRNA expression were investigated in 73 allergic subjects with different MD-2 gene promoter variants. Peripheral blood mononuclear cells were cultured with or without LPS in the presence of Dermatophagoides pteronyssinus group 2 allergen (Der p 2), followed by mRNA extraction and cytokine expression analysis. The culture supernatants were collected for cytokine measurement. RESULTS: Patients with the MD-2 promoter SNPs (rs1809441/rs1809442) had increased mRNA expressions of MD-2, epsilon heavy chain of IgE (Cepsilon), and interleukin (IL)-8; however, only MD-2 and IL-8 were further up-regulated after Der p 2 stimulation. Patients with SNPs of MD-2 promoter tended to have high levels of IL-1beta, IL-6, IL-8, IL-10, and tumor necrosis factor (TNF)-alpha after Der p 2 and LPS stimulation. Increased secretions of IL-6, IL-8, and IL-10 were found to be up-regulated by Der p 2 stimulation, and an increased secretion of IFN-gamma and decreased secretion of IL-4 were noted after LPS stimulation. CONCLUSIONS: The high levels of proinflammatory cytokines secreted by Der p 2 were predetermined by MD-2 promoter SNPs (rs1809441/rs1809442). Through cytokine secretion by Der p 2 and LPS, these SNPs may serve as an indicator of the pathological phenotype of Der p 2-induced allergic inflammation.
Cytokines ; Dermatophagoides pteronyssinus ; Humans ; Hypersensitivity ; Immunoglobulin E ; Immunoglobulins ; Inflammation* ; Interleukin-10 ; Interleukin-4 ; Interleukin-6 ; Interleukin-8 ; Interleukins ; Phenotype ; Polymorphism, Single Nucleotide ; RNA, Messenger ; Toll-Like Receptor 4 ; Tumor Necrosis Factor-alpha

OBJECTIVEErectile dysfunction (ED) is now recognized as a comorbid condition, especially in men with cardiovascular disease or diabetes mellitus. This randomized controlled trial was to examine the effect of long-term small-dose tadalafil in the treatment of ED.

METHODSA total of 98 men older than 18 years with at least a 6-month ED history were enlisted and divided into two groups to receive once-daily treatment with tadalafil at 5 mg (n = 60) and 20 mg (n = 38), respectively, for 12 months. The effects of medication were analyzed and compared using IIEF, Global Assessment Questionnaire (GAQ) and Sexual Encounter Profile (SEP), and so were the safety and tolerance of the two doses.

RESULTSThere were no statistically significant differences in the therapeutical results between the 5 mg and 20 mg groups (P < 0.05). The IIEF-5 score was raised by 8.1 points in the former and 7.9 points in the latter; the YES answers to SEP2 in the two groups were 51.3% and 49.2% before the treatment and 82.6% and 84.9% after it. No serious adverse events were observed, except some common ones, such as rubeosis (11.9% vs 8.7%) and headache (5.3% vs 4.9%) in the 5 mg and 20 mg groups.

CONCLUSIONOral tadalafil at 5 mg once daily is efficacious with good tolerance in the treatment of ED, and it can be an alternative to on-demand medication for some men to eliminate the inconvenience of planned intercourse within a limited timeframe.

Adult ; Carbolines ; administration & dosage ; adverse effects ; therapeutic use ; Erectile Dysfunction ; drug therapy ; Humans ; Male ; Middle Aged ; Phosphodiesterase Inhibitors ; administration & dosage ; adverse effects ; therapeutic use ; Tadalafil ; Treatment Outcome

OBJECTIVETo explore the application of computed tomographic angiography (CTA) and computer aided design\manufacture (CAD\CAM) techniques in mandible reconstruction with free fibula flap.

METHODS7 cases of complex mandible defect were reconstructed with free fibula flap. Patients' lower extremity vasculature was evaluated by preoperative CTA examination; mandible tumor resection and reconstruction were simulated by CAD technique, the surgical template was manufactured with CAM technique preoperatively; and fibula flap was shaped according to surgical template during mandible reconstruction operation.

RESULTSPreoperative CTA examination performed without complication, the desired anatomy was adequately demonstrated in all patients. All of flaps survived, patients' facial contour are symmetry approximately.

CONCLUSIONSCTA and CAD\CAM techniques have important value for complex mandible reconstruction in selection of donor site, planning of operation and shaping of fibula flap.

Adolescent ; Adult ; Bone Transplantation ; methods ; Female ; Fibula ; blood supply ; transplantation ; Humans ; Male ; Mandible ; surgery ; Middle Aged ; Reconstructive Surgical Procedures ; methods ; Surgical Flaps ; blood supply ; Young Adult