It is well documented that altered biosynthesis of cell surface N-linked oligosaccharides is associated with the transformed cells and tumors.N-acetylglucosaminyltransferase Ⅱ(GnT-II;EC 2.4.1.143)is a medial Golgi enzyme that catalyses the incorporation of a GlcNAc residue in ?-1,2 linkage to the Man-?-1,6 arm of the N-glycan core.This is an essential step in the biosynthetic pathway leading from hybrid to complex N-glycans.Because functional GnT-Ⅱ is an prerequisite of N-Acetylglucosaminyltransferase V performance,It was speculated that GnT-Ⅱ was involved in cancer development and progression.The expression of GnT-Ⅱ in mouse breast cancer cells 67NR and 4T1 which have different behavior of metastasis was analysed using RT-PCR.The amounts of GnT-Ⅱ in the highly metastatic cell 4T1 increased to 1.53 times of the lowly metastatic cell 67NR.To determine the association of GnT-Ⅱ with tumor progression,the GnT-Ⅱ encoding gene was amplified with RT-PCR and cloned into retrovirus vector pMSCV,resulting in pMSCV-GnT-Ⅱ.The recombinant plasmid was transfected into 4T1 and the transfected cells were selected in the medium containing puromycin,which were harvested to detect the adhesion ability to fibronection and the migration potential by transwell system.The cell adhesion to fibronectin was weakened by 67% and migration potential was increased by 82%.The data indicates that GnT-Ⅱ mediates cell adhesion and migration,thus may play an important role in cancer metastasis.

Previous studies have revealed Twist,a basic helix-loop-helix transcription factor,plays an important role in breast cancer metastasis.To clarify the molecular mechanism of its involvement in cancer metastasis,Twist was silenced by RNAi in highly metastatic 4T1 cells.Then microarray chips were used to investigate the gene-expression pattern of the Twist-knockdown 4T1 cells and the normal 4T1 cells.The results indicated that silencing of Twist significantly suppressesed lung metastasis of 4T1 cells in vivo.Direct comparison of gene-expression profiles showed that 167 genes in Twist-knockdown cells differed dramatically in expression levels from those in control cells.Among the 167 genes,26 well-known tumor-associated genes,including 15 up-regulated and 11 down-regulated genes were found.These genes appear to be regulated by Twist during breast tumorigenesis.The findings provide new insights into the mechanism by which Twist is involved in tumorigenesis.

BACKGROUND: Properties of cancellous bone is significant for metabolic diseases, fracture and joint retrograde diseases.OBJECTIVE: To study the biomechanical properties of cancellous bone.DESIGN: Experimental observation based on femur specimens as the single sample.SETTING: Laboratory of Orthopaedic Biomechanics Ninth Affiliated Hospital Shanghai Second Medical University.PARTICIPANTS: The study was completed in the Laboratory of Orthopaedic Biomechanics, Shanghai Ninth People's Hospital from May to July in 2001,totally 19 male cadaveric femoral heads were selected.METHODS: Totally 19 cadeveric cancellouce bone specimens(about 30years old) from the bearing area of fresh femur head were tested by compression test.MAIN OUTCOME MEASURES: Yield load limiting stress, limiting strain, energy value and elastic modulus of specimens.RESULTS: The compressive data of cancellous bone were measured, the average value of yield load was (410.64 ± 190.29) N , limiting stress was (8.69 ± 3.75) MPa, limiting strain was (10. 84 ±6.58)%, energy value was (2.54 ± 1.89)J, modulus of elasticity was (40.77 ± 32. 12)MPa, At the early time of loading, the cancellous bone showed elastic deformation, then CONCLUSION: The trial indicates that biomechanical properties lighten the pressure of external compression.

Chills ; prevention & control ; Cold Temperature ; Female ; Humans ; Occupational Exposure ; prevention & control ; Occupational Health Services ; Water

Objective To investigate the reason of persisting positive rapid plasma reagin (RPR) in serofast syphilis patients, and to provide reference for clinical treatment and prognosis.Methods A total of 33 serofast patients and 23 healthy controls were enrolled in this study.The percentages and absolute counts of CD3+, CD4+, CD8+ T lymphocytes and natural killer (NK) cells were detected by flow cytometry.The comparison of two groups was analyzed by independent sample t test, and the correlation between change of lymphocyte subgroups and RPR titer in serofast syphilis patients was analyzed by bivariate linear correlation method.Results Compared with healthy controls, the percentages of CD3+, CD8+ T lymphocytes in serofast syphilis group were both increased significantly (75.75±5.76)% vs (68.37±5.80)%, (t=4.69, P<0.01);(27.34±7.02)% vs (24.33±1.95)%, (t=2.34, P=0.025), while both the percentage and absolute count of NK cells were significantly decreased (7.32±4.48)% vs (14.87±6.26)%, (t=5.269, P<0.01);(136.2±83.4)/μL vs (298.8±166.9)/μL, (t=4.311, P<0.01).RPR titer of the patients was negatively correlated with both percentage and absolute count of CD4+ T lymphocytes (r=-0.476 and-0.515, respectively, both P<0.01), and it was positively correlated of CD8+ lymphocytes (r=0.588 and 0.305, P<0.01 and P=0.804).Conclusion The imbalance of immune response of lymphocyte subsets observed in serofast syphilis may explain the RPR titers change.

A secretive expression vector of Pichia pastoris system which can be used for the direct cloning of PCR products was constructed,and was verified through the expression of recombinant cellobiohydrolase II in Pichia pastoris.A randomly selected fragment was amplified with properly designed primers by PCR.The XhoI and Eam1105Ⅰ restriction sites were included in the 5'end of the fragment,and the Eam1105Ⅰ and XbaI restriction sites were included in its 3'end.The PCR amplified product was inserted into the P.pastoris expression plasmid pPICZ?A through XhoI and XbaI restriction sites and the resultant plasmid was digested with Eam1105Ⅰ,and lastly the big fragment was recovered,generating the P.pastoris expressive Tvector pPICZ?T.Then the cellobiohydrolase II of T.reesei was successfully expressed in P.pastoris with this expressive Tvector.Such results indicated that the constructed expression Tvector was convenient for PCR product cloning,and was effective for heterologous protein expression in P.pastoris.On the other hand,the application of the expression Tvector avoided the introduction of additional amino acids at the Nterminus of the expressed protein,which generally occurred when normal expression vectors were used in secretive expression system.

Akt1 is a serine-threonine protein kinase that has been implicated in the control of cellular metabolism,survival and growth.Elevated expression of Akt1 has been noted in a significant percentage of human tumors,promoting cellular metastasis.Conversely,some studies have revealed hyperactivated Akt1 inhibited the invasiveness and metastasis of breast cancer cells.To clarify the definite effect of Akt1 on tumorigenesis and development,Akt1 was silenced by RNAi in the highly metastatic murine breast cancer 4T1 cells.Akt1 silencing didn't affect the proliferation of breast cancer cells in MTT assay,while reduced the migration in Transwell assay.Consistent with the above results,Akt1 silencing didn't change the primary tumor weight,but significantly suppressed lung metastasis of 4T1 cells.These observations indicated Akt1 plays an important role in murine breast cancer metastasis,and suggested that Akt1 might be a therapeutic target for breast cancer metastasis.

OBJECTIVETo investigate the protective effect of Danxuetong injection (DXT, a combination of Danshen and Xueshuantong injections) against testicular ischemia-reperfusion injury following testis torsion/detorsion in rats.

METHODSThirty-two 4-week-old healthy male SD rats were randomly divided into four groups of equal number: sham operation, normal saline, single DXT injection, and successive DXT injection. The rat models of testicular ischemia-reperfusion injury were established by 2-hour 720-degree torsion/detorsion of the unilateral testis. At 6 weeks after modeling, the rats were killed and their testes were harvested for measure- ment of testicular coefficients, sperm counts, sperm motility, and the levels of total anti-oxidative capacity (T-AOC) , superoxide dismutase (SOD) , nitric oxide synthase (NOS) , and malondialdehyde ( MDA) in the testis tissue.

RESULTSCompared with the rats of the normal saline group, those of the single DXT injection and successive DXT injection groups showed significant increases in the testicular coefficient (0.11 ± 0.03 vs 0.35 ± 0.04 and 0.40 ± 0.06, P < 0.05), sperm count ([0.46 ± 0.10] vs [1.44 ± 0.50] and [3.00 ± 1.28] x10(9)/ml, P < 0.05), sperm motility ([13.63 ± 14.04] vs [39.63 ± 5.04] and [76.31 ± 3.67]%, P < 0.05), the activity of SOD (72.76 ± 5.58 vs 116.25 ± 8.83 and 133.20 ± 13.84, P < 0.05), and the level of T-AOC (5.58 ± 1.07 vs 13.34 ± 5.81 and 19.21 ± 5.69, P < 0.05), but a remarkable decrease in the content of MDA (42.38 ± 8.94 vs 20.94 ± 5.65 and 15.02 ± 1.03, P < 0. 05) in the injured testes.

CONCLUSIONDXT can effectively rid the testis tissue of oxygen free radicals, improve sperm count and motility by antioxidation, and protect the testis tissue of prepubertal rats against testicular ischemia-reperfusion injury after testis torsion/detorsion. It also has a protective effect on the contralateral testis, and successive injection has a better effect than single injection of DXT.

Animals ; Antioxidants ; therapeutic use ; Drug Therapy, Combination ; methods ; Drugs, Chinese Herbal ; therapeutic use ; Humans ; Male ; Malondialdehyde ; metabolism ; Nitric Oxide Synthase ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Reperfusion Injury ; prevention & control ; Spermatic Cord Torsion ; complications ; therapy ; Superoxide Dismutase ; metabolism ; Testis ; blood supply ; metabolism

Previous studies have shown that ginsenoside Rb1 (Rb1), one of active components in ginseng, can activate insulin signaling pathway and promote translocation of glucose transporters (GLUTs) to increase glucose uptake in adipocytes. However, the effect of Rb1 on the expressions of GLUTs remains unknown. In this study, the effects of Rb1 on GLUT1 and GLUT4 were observed in 3T3-L1 adipocytes and epididymal adipose tissue of db/db obese diabetic mice. Male db/db mice were treated with Rb1 by intraperitoneal injection at the dosage of 20 mg x kg(-1) for 14 d. Rb1 reduced HOMA-IR significantly (P < 0.05, n = 5), and FBG and FINS sowed declining trend after treatment with Rb1. Rb1 recovered the expressions of GLUT1 and GLUT4 and phosphorylation of AKT in adipose tissue of db/db mice. In vitro, glucose consumption in 3T3-L1 adipocytes treated with 10 micromol x L(-1) Rb1 for 24 h was elevated (P < 0.05, n=3), and mRNA of GLUT1 and GLUT4 were up-regulated (P < 0.05, n=3) and proteins of GLUT1 and GLUT4 were also increased. AKT was activated in adipocytes treated with Rb1 for 3 h. It can be concluded that ginsenoside Rb1 can up-regulate the expression of GLUTs in adipose tissue, in addition to activate insulin signalling pathway, which may partially account for its insulin sensitizing activity and regulating effect of glucose metabolism.
3T3 Cells ; Adipocytes ; drug effects ; Animals ; Cell Line ; Diabetes Mellitus, Experimental ; metabolism ; Ginsenosides ; pharmacology ; Glucose ; metabolism ; Glucose Transport Proteins, Facilitative ; metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Inbred NOD ; Up-Regulation ; drug effects

To investigate the therapeutic effect and molecular mechanism of the main flavonoid components of Silybum marianum (S. marianum) on nonalcoholic fatty liver disease (NAFLD), we identified nine flavonoids in S. marianum through TCMSP, PubChem database and corresponding literatures. The potential therapeutic targets of NAFLD were predicted by SwissTargetPrediction, GeneCards and Venny 2.1.0 platform, while the protein-protein interaction (PPI) network of potential targets was analyzed using String platform and Cytoscape software. Then GO and KEGG pathway enrichment analysis were performed using David 6.8 database, followed by molecular docking verification using AutoDock software. In vitro, components with higher degree value in the "components-targets-pathway" network were chosen for further analysis. L02 cells were used to establish lipid accumulation model and treated with different components. Furthermore, the effects of four pure active compounds from S. marianum on lipid accumulation in hepatocytes were analyzed by oil red O staining. The results showed that the main nine flavonoids extracted from S. marianum contained 24 potential NAFLD targets. Several critical pathways closely related to NAFLD process were identified by GO and KEGG enrichment analysis, including phosphatidylinositol 3-kinase-protein kinase B (PI3K-Akt) pathway, type 2 diabetes pathway, tumor necrosis factor (TNF) pathway and insulin resistance pathway. The results of molecular docking further indicated that the core components displayed strong binding abilities with key targets respectively, and silandrin showed better binding activity as compared to other components. The results obtained from L02 cells showed that the lipid accumulation was reduced by treatment with isosilybin A, isosilybin B, silydianin and silychristin, while the activity of isosilybin B was better than that of isosilybin A. Taken together, we concluded that the main flavone components of S. marianum could improve lipid accumulation through multiple signaling pathway in hepatocytes, and this could be a potential new strategy for the treatment of NAFLD.