Objective To study the effects of russel viper venom X(RVV X)on blood coagulation protein.Methods We divide diluted protein into control and RVV-X groups,then use chromogenic substract assay to detect the activation effect of RVV-Ⅹ on coagulation factor Ⅶ,Ⅸ,Ⅹ and antithrombin,plasminogen,with or without activator.Results In RVV-Ⅹ group,the coagulation factor Ⅶ, Ⅸ and plasminogen displayed weakly enhanced chromogenesis,all P

OBJECTIVETo observe the effect of Polygonatum sibiricum on Yin deficiency model rats induced by long-term overload swimming.

METHODExcept for the normal group, all of the remaining rats performed the long-term overload swimming for eight weeks, with five days every week and once every day, to establish the Yin deficiency model. The daily swimming time increased from 10 min to 180 min at the end of the 7th week, with the water depth of 60 cm and the water temperature at 30 degrees C. After the success of the modeling, the rats were orally administered with different doses of aqueous extracts from P. sibiricum (2.5, 10 g x kg(-1)) for eight weeks. After the final administration, their blood were collected from orbits to measure immunoglobulin A, G and M (IgA, IgG, IgM), interleukin 2 and 6 (IL-2, IL-6) and cAMP, cGMP contents in plasma General behavioral indicators (weight, facial temperature, pain threshold and holding power) of rats were observed during the drug administration.

RESULTCompared with the model control group, aqueous extracts from P. sibiricum was given for eight weeks to significantly increase the rat weight and holding power of Yin deficiency model rats, decrease the facial temperature and the sensitivity of pain threshold, and increase IgA, IgG, IgM and IL-6 content and IgG content in serum, but without statistical difference. Aqueous extracts from P. sibiricum (10 g x kg(-1)) could also increase IL-2 content in serum, and decrease cAMP content and cAMP/cGMP ratio.

CONCLUSIONP. sibiricum could improve the general behavioral indicators (weight, holding power, pain threshold and facial temperature), immunologic functions (IgA, IgG, IgM) and cyclic nucleotide (cAMP, cAMP/cGMP), so as to ameliorate such Yin deficiency symptoms as dysphoria in chestpalms-soles, weight loss, soreness and weakness of waist and knees, immunologic dysfunction and cyclic nucleotide system disorders.

Animals ; Body Weight ; drug effects ; Drugs, Chinese Herbal ; administration & dosage ; Female ; Humans ; Male ; Polygonatum ; chemistry ; Rats ; Rats, Sprague-Dawley ; Swimming ; Yin Deficiency ; drug therapy ; physiopathology

Objective To evaluate the effectiveness of MGIT liquid medium fluorescence instrument manual interpretation method for rapid detection of Mycobacterium. Methods Two hundred sputa with newly diagnosed tuberculosis patients were collected from October 2008 to January 2009 in the district hospitals in Shanghai. Of these 200 sputa, 67 sputa were positive AFB, 133 were negative. All the sputa were isolated by L-J, BacT / Alert 3D system and MGIT liquid medium methods. Results Of the 200 sputa specimens,105(52. 5% ) were isolated as Mycobacterium strains. The positive culture rate of the MGIT, BacT/Alert 3D and L-J method was 49. 5% ( 99/200 ), 48. 0% (96/200) and 45.0% ( 90/200), respectively. The MGIT culture positive rate was significantly higher than that of L-J method (x2 = 5.40, P = 0. 020 1 ). Of the 133 sputa with negative AFB, the positive culture rate was 24. 8% ( 33/133 ), 23. 3% ( 31/133 ) and 18. 8% (25/133) with MGIT, BacT/Alert 3D and L-J method, respectively. The MGIT culture positive rate with the AFB negative sputum was significantly higher than that of L-J method (x2 = 5. 33, P = 0. 020 9 ).The median time of detection with MGIT, BacT/Alert 3D system and L-J method was 11 days, 15 days and 22 days, respectively. Comparing the median time of detection of MGIT with BacT/Alert 3D, the difference was statistically significant ( Z = 3.414 ,P ＜ 0. 01 ). Comparing the median time of detection of MGIT with L-J method, the difference was statistically significant (Z =7.083,P＜0. 01).Conclusions MGIT liquid medium manual method is a rapid detection method of Mycobacterium with a high positive detection rate, and do not need expensive equipment This method may suitable to resource limited medical institutions due to its low cost and short round time.

In order to identify novel genes involved in spermatogenesis, testis cDNA samples from Balb/C mice of different postnatal days were hybridized with the whole mouse genome Affymetrix chip to screen the testis-specific genes. The characteristics of the selected genes were analyzed by RT-PCR as well as other bioinformatic tools. A novel differentially expressed testis-specific gene (GenBank Accession No: NM_029042) in the developmental stages of testes was identified, and named TSCPA. Cellular mapping prediction of TSCPA indicated that its protein was probably expressed in nuclei, and one putative domain (aa 332-377) was anchoring domain of cAMP-dependent type II PK. The result of subcellular localization of GFP-TSCPA fusion protein in Cos-7 cells showed that TSCPA protein was expressed in nuclei. RT-PCR analysis revealed that TSCPA was expressed specifically in mouse and human testis. TSCPA gene was expressed weakly in 21-day-old mouse testis and the expression was increased gradually from 38th day to 6th month of mouse testes. No expression of hTSCPA was found in cryptorchidism and Sertoli-cell-only syndrome patients. It was concluded that the expression profile of TSCPA in human and mice indicated that TSCPA might play an important role in spermatogenesis.

0.05)and no significant statistical difference was found.But the successful rate of first therapy of group A was 71.42%(85/119)and group B was 56.20%(77/137)(P

AIM:The bacillus calmette-guerin(BCG) vaccine is the most widely used Th1-inducing vaccine.In recent years,some studies argued that mycobacterium vaccae can be used as adjuvant to induce regulatory T cells(Treg) and then suppress asthmatic airway inflammation.We previously have engineered recombined BCG that expressed Der p2 of house dust mites(Der p2 rBCG) on the cell wall.The aim of this study is to investigate the immune regulatory mechanisms of Der p2 rBCG.METHODS:Mice were vaccined with PS,BCG or rBCG.The relative proportion and the absolute numbers of related Tregs in spleen cells were analyzed.The suppressive activity of Der p2 rBCG-induced CD4+CD25+ T cells was detected both in vitro and in vivo.RESULTS:(1) Der p2 rBCG induced a CD4+CD25+Foxp3+ T cell subtype.(2) Der p2 rBCG-induced CD4+CD25+ T cells suppressed the proliferation of Th2 effector cells in vitro in an antigen-specific way.(3) Der p2 rBCG-induced CD4+CD25+ T cells mediated Der p2 specific suppression of airway allergy in vivo.CONCLUSION:Der p2 rBCG induces a CD4+CD25+Foxp3+ T cell subtype,which suppresses inflammation in allergic airway in a mouse model.

Objective To observe whether the expression of interferon-regulatory factor genes are re- lated to systemic lupus erythematosus (SLE).Methods The clinical data of 45 SLE patients and 37 normal controls were collected.Total RNA of peripheral blood was extracted and transcripted into cDNA.Sybr green dye based real-time quantitative PCR method was used to compare the expression (indicated as-??Ct value) of IRFI,IRF4,IRF8 in patients with SLE and those in the controls.Results The levels of IRF1,IRF4 and IRF8 mRNA were-3.90?0.19,-8.04?0.25 and 3.60?0.15 respectively in normal controls.In SLE patients, IRF4 mRNA expression was -8.82?0.18,higher than that in normal (P=0.011).But IRF8 mRNA expression was 3.09?0.13,lower than that in normal (P=0.012).Conclusion Abnormal IRF mRNA expression is found in the peripheral blood of SLE patients.IRFs may play roles in the pathogenesis of SLE by affecting the differen- tiation of Th cells.

OBJECTIVETo construct a recombinant lentiviral vector for manganese superoxide dismutase (MnSOD) gene expression, and observe its effect on the proliferation of esophageal cancer cells in vitro.

METHODSChemical methods were employed for synthesis of the MnSOD cDNA sequence sections, along with the attB sites. Target gene fragment was constructed on the pMD-18T vector, and the recombinant plasmid pDONR221 was obtained after BP recombination reaction. Sequencing was followed by LR recombination reaction between the plasmid and DEST to obtain the lentiviral vector, which worked with helper plasmid for co-transfection of human embryonic kidney epithelial cells (293T cells). Amplification was done to determine its titer, and both transfection and selection procedures were made to get two stable transfected esophageal cancer TE-1 cell lines with medium MnSOD expression (TE-1Mm cells) and high MnSOD expression (TE-1Mh cell), and empty vector cell (TE-1Mn cells). Reverse transcription polymerase chine reaction (RT-PCR), immunofluorescence, immunocytochemistry and Western blot were used to detect the target gene with respect to its expression in the TE-1 cells. Additionally, colorimetric 3-[4,5-dimethy thiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay, agar colony formation assay, annexin V-FITC/PI staining and flow cytometry experiments were also conducted as to observe the influence of the medium and high MnSOD overexpressions on the proliferation of esophageal cancer cells.

RESULTSRT-PCR indicated that the transfected TE-1 cells showed positive MnSOD expression at different levels. Immunofluorescence, immunocytochemistry and Western blot suggested that TE-1Mm cells and TE-1Mh cells had MnSOD protein expression at different levels. MTT assay indicated that TE-1Mm cells had a significantly decreased survival rate compared with that of the two control cells (TE-1 cells and TE-1Mn cells), and TE-1 Mh cells had an significantly increased survival rate (P<0.05). The colony formation ability of TE-1Mm cells was (23.0 +/- 2.7)%, and that of TE-1Mh cells was (45.3 +/- 4.5)%, significantly different form the (34.7 +/- 4.2)% in TE-1 cells and (33.7 +/- 4.7)% in TE-1Mn cells (P<0.05). Annexin V-FITC/PI double staining experiment of the stably transfected cells cultured for 48 h showed that the early apoptosis rate in TE-1Mm cells was (10.6 +/- 1.0)%, significantly higher than (2.6 +/- 0.2)% in the TE-1 cells, (2.5 +/- 0.6)% in the empty vector cells and (1.0 +/- 0.1)% in the TE-1Mh cels (P<0.05). The fluorescence index (FI) of mitochondrial apoptosis of TE-1Mm cells was 0.948 +/- 0.019, significantly lower than that of TE-1 cell (1.000 +/- 0.022) and empty vector The fluorescence index of TE-1Mn cells (0.997 +/- 0.023) and TE-1 cells (1.000 +/- 0.022) were significant different from that of 0.948 +/- 0.019 in TE-1Mm cells and 1.076 +/- 0.022 in TE-1Mh cells, indicating a significant difference of mitochondrial apoptosis between the cell groups. FCM results indicated that the ROS fluorescence index of TE-1Mm cells was 0.859 +/- 0.040, that of TE-1Mh cells was 0.763 +/- 0.039, significantly lower than that of TE-1 cells (1.000 +/- 0. 042) and empty vector cells (1.002 +/- 0.047) (P<0.05).

CONCLUSIONSStably transfected cell lines with MnSOD expression have been successfully established. MnSOD overexpression shows bidirectional effect on the proliferation of esophageal cancer cells.

Apoptosis ; Cell Line, Tumor ; Cell Proliferation ; Esophageal Neoplasms ; metabolism ; pathology ; Gene Expression Regulation, Neoplastic ; Genetic Vectors ; HEK293 Cells ; Humans ; Lentivirus ; genetics ; Mitochondria ; pathology ; Plasmids ; RNA, Messenger ; metabolism ; Reactive Oxygen Species ; metabolism ; Recombinant Proteins ; genetics ; metabolism ; Superoxide Dismutase ; genetics ; metabolism ; Transfection

In order to enhance the antitumor efficacy of recombinant Newcastle disease virus, rNDV-IL15 was rescued in this study. Recombinant plasmid prNDV-IL15 was constructed, and BHK21 cells were transfected with the recombinant plasmid. Finally, the recombinant Newcastle disease virus rNDV-IL15 was successfully rescued. The growth curves of these two recombinant viruses were determined. Murine melanoma B16F10 cells were infected with rNDV-IL15 at MOI of 0.1, and the expression level of IL15 in the supernatant was detected by ELISA. The antitumor efficacy of rNDV-IL15 and rNDV was compared in vitro and in vivo. Results showed that prNDV-IL15 was constructed and recombinant virus rNDV-IL15 was successfully rescued. The growth curve of rNDV-IL15 showed that the growth of rNDV-IL15 had not been changed after insertion of IL15 gene. Results showed that there was high level of IL15 expression in the supernatant of rNDV-IL5-infected B16F10 cells (1 044.3 +/- 27.7 ng x mL(-1)). rNDV-IL15 and rNDV significantly inhibited the growth of B16F10 cells in vitro in a time-dependent manner. However, there was no significant difference between them. In animal experiments, rNDV-IL15 efficiently suppressed tumor growth in vivo when compared with rNDV, and the difference was statistically significant. The results suggested that rNDV-IL15 is a more effective antitumor agent.
Animals ; Body Weight ; Cell Line, Tumor ; Cell Proliferation ; Chick Embryo ; Cytotoxicity, Immunologic ; Female ; Genetic Therapy ; Interleukin-15 ; genetics ; metabolism ; Melanoma, Experimental ; pathology ; therapy ; Mice ; Neoplasm Transplantation ; Newcastle disease virus ; genetics ; Plasmids ; Recombinant Proteins ; genetics ; metabolism ; Transfection ; Tumor Burden

To investigate the cell-killing effect and its possible mechanism of rClone30-hDR5 in combination with TRAIL on human hepatic carcinoma (HCC) cell line, first of all, recombinant plasmid pee12.4-hDR5 was introduced into HepG2 cells by liposome transfection. After five rounds of screening by flow cytometry, HepG2 cells expressing high levels of DR5 on cell surface were isolated. The cytotoxicity of TRAIL to selected cells was higher than that of TRAIL to HepG2 cells by MTT method (P < 0.01). The result suggested that the cloned hDR5 gene had biological activity. MTT assay showed that, rClone30- hDR5 in combination with TRAIL more efficiently inhibited the tumor growth of HepG2 cells compared to rClone30-hDR5 or TRAIL in vitro. The results of Annexin V-FITC/PI staining and Quantitative Real-time PCR indicated that rClone30-hDR5 in combination with TRAIL significantly increased the mRNA levels of caspase 3 and caspase 8, and induced the apoptosis of tumor cells. HepG2 cells were infected with rClone30-hDR5 or rClone30 at MOI of 1. The expression of hDR5 on tumor surface increased significantly by rClone30-hDR5 compared to that by rClone30, which contributed to the sensitivity to TRAIL. In conclusion, rClone30-hDR5 in combination with TRAIL has potential application value in cancer treatment.
Apoptosis ; Carcinoma, Hepatocellular ; pathology ; Caspase 3 ; metabolism ; Caspase 8 ; metabolism ; Drug Synergism ; Hep G2 Cells ; Humans ; Liver Neoplasms ; pathology ; Real-Time Polymerase Chain Reaction ; Receptors, TNF-Related Apoptosis-Inducing Ligand ; pharmacology ; TNF-Related Apoptosis-Inducing Ligand ; pharmacology ; Transfection