OBJECTIVETo observe beta-catenin expression of Wnt signaling pathway in rats with knee osteoarthritis, and influence of Bushen Huoxue decoction on beta-catenin and MMP-7 expression of synoviocytes in rats with knee osteoarthritis (OA).

METHODSRats model with knee osteoarthritis were established by Hulth method. Primary synoviocytes and OA synoviocytes were cultured with collagenase digestion method. The cultured synoviocytes were divided into normal group, OA model group and Bushen Huoxue decoction group. Western blotting method was used to detect beta-catenin, MMP-7 protein expression of synoviocytes after acting by Bushen Huoxue decoction for 48 h; ELISA method was used to detect MMP-7 expression of synovial supernatant.

RESULTSOA synoviocytes were cultured successfully. Western blotting showed that beta-catenin, MMP-7 expression in OA synoviocytes was significantly higher than normal group, Bushen Huoxue decoction could significantly reduce beta-catenin, MMP-7 expression; ELISA results showed that MMP-7 expression of OA synovial supernatant was significantly higher than normal synoviocytes supernatant, Bushen Huoxue decoction significantly regulated the level MMP-7 down.

CONCLUSION(1) High expression of beta-catenin in OA synoviocytes indicates that Wnt classical signal pathway is activated in rat with knee osteoarthritis; (2) High expression of MMP-7 expression in OA synoviocytes confirms the MMP-7 is downstream genes of Wnt/beta-catenin signal pathway; (3) Activation of Wnt signal pathway and increase of MMP-7 may cause degradation of articular cartilage, and promote the formation of osteoarthritis; (4) Bushen Huoxue decoction can reduce expression of MMP-7, and promote cartilage repair, which may be one of mechanisms of osteoarthritis.

Animals ; Blotting, Western ; Drugs, Chinese Herbal ; pharmacology ; Enzyme-Linked Immunosorbent Assay ; Male ; Matrix Metalloproteinase 7 ; analysis ; Osteoarthritis, Knee ; drug therapy ; metabolism ; Rats ; Rats, Wistar ; Synovial Fluid ; chemistry ; cytology ; beta Catenin ; analysis

OBJECTIVETo study the relationship between ambient air pollution and daily mortality of SARS in Beijing.

METHODSThe approach of time-series Poisson regression was used to assess the relationship between daily SARS mortality, ambient air pollution, and other factors from April 25 to May 31, 2003 in Beijing.

RESULTSAn increase of each 10 microg/m3 over a 5-day moving average of PM10, SO2 and NO2 corresponded to 1.06 (1.00-1.12), 0.74 (0.48-1.13) and 1.22 (1.01-1.48) relative risks (RRs) of daily SARS mortality, respectively. The relative risks (RRs) values depended largely on the selection of lag days.

CONCLUSIONThe daily mortality of SARS might be associated with certain air pollutants in Beijing.

Air Pollutants ; adverse effects ; analysis ; toxicity ; Air Pollution ; adverse effects ; analysis ; China ; epidemiology ; Cities ; Dust ; analysis ; Environmental Monitoring ; Epidemiological Monitoring ; Humans ; Nitrogen Dioxide ; analysis ; Particle Size ; Retrospective Studies ; Risk ; Severe Acute Respiratory Syndrome ; epidemiology ; mortality ; Sulfur Dioxide ; analysis

BACKGROUNDIt has been known that the growth of solid tumors is dependent on angiogenesis, and neoangiogeneses of tumor become main target to control tumor growth. The aims of this study are to investigate the inhibition effect of replicate-deficient adenovirus encoding the soluble form of mouse vascular endothelial growth factor receptor 1 (sFlt1-Adv) on angiogenesis and tumor growth in established tumor model.

METHODSMouse Lewis lung cancer cells were inoculated subcutaneously into C57 mice. sFlt1-Adv, GFP-Adv and normal saline were injected twice intravenously after establishing Lewis cancer model. Diameters of tumors were measured every other day. Tumors were resected, weighed and fixed in 3% paraformadehyde. Microvessel density of tumors was determined by immunohistochemical staining with anti-CD31 antibody.

RESULTSThe planted tumor volume and weight in sFlt1-Adv group were significantly lower compared with the two controls (P < 0.01). Its inhibition rate was 71.8%. The microvessel density in sFlt1-Adv group decreased markedly compared with that of the control groups (P < 0.01).

CONCLUSIONSsFlt1-Adv can inhibit the growth of tumor through the inhibition of tumor angiogenesis. sFlt1-Adv may be potentially valuable for clinical treatment of solid tumor.

Osteoarthritis is a degenerative joint disease,with the characters of degradation of articular cartilage, the formation of the joint marginal osteophyte and synovium lesions. Previous studies have focused on the treatment of articular cartilage lesions. In recent years, new research in shows synovial inflammation plays an important role in OA. Synovium lesions and synovial inflammation-related factors induced the degradation and destruction of articular cartilage, and promoted the development of osteoarthritis. The role of synovial lesions in osteoarthritis is increasingly prominent, and the treatment for synovial lesions will become a new target. So this paper reviews the various manifestations of synovial in osteoarthritis.
Humans ; Osteoarthritis ; pathology ; Synovial Membrane ; pathology

Objective To discuss the etiology,clinical features and treatments of patients with pituitary adenomas having onset symptom of hyponatremia.Methods Retrospective analysis was performed on etiology,clinical features and treatment methods of 28 patients with pituitary adenomas having onset symptom ofhyponetramia,admitted to our hospital from Jantary 2003 to December 2012.Results Among 28 patients,27 had non-functional pituitary adenomas and one prolactinoma; 23 had pituitary macroadenomas and 5 giant adenomas; 22 achieved total tumor resection through transsphenoidal approach,2 subtotal removal and 4 received conservative therapy.In total,13 were combined with hypocortisolism,8 with hypothyroidism,5 with both hypocortisolism and hypothyroidism,and 2 with hyponatremia caused by syndrome of inappropriate secretion of antidiuretic hormone in preoperative evaluation.Hyponatremia was corrected by hormone replacement therapy and water limitation during perioperative and postoperative period.Conclusions Pituitary adenomas with onset symptom of hyponatremia are mostly non-functional pituitary macroadenomas.Pituitary adenoma resection is the first-line treatment; hyponatremia and anterior pituitary hypofunction should be corrected in perioperative period,and close postoperative follow-up is necessary.

OBJECTIVETo construct a recombinant adenovirus vector expressing hCD40L gene and explore it in the use of anti-tumor gene therapy.

METHODS1,900 bp gene fragment was obtained form plasmid pORF-hCD40L by Xho I/Swa I cutting and then cloned directionally into the pShuttle plasmid, finally, the resultant plasmid was digested by restriction endonnuclease PmeI and subsequently cotransformtion into BJ5183 cells with the adenoviral backbone pAdEasy-1 to obtain the homologous recombinant and then the recombinant was packaged in the 293 cells. Some methods such as PCR and endonulease digestion were employed to identify the recombinant adenovirus.

RESULTSThe evidences of endonulease digestion and PCR analysis confirmed that recombinant hCD40L gene was correctly inserted into adenovirus vector.

CONCLUSIONThe adenoviral vector which expressed hCD40L gene was constructed. It provides an experimental basis for studies on it expression in the mammalian cells and in tumor gene therapy.

Adenoviridae ; Adenoviruses, Human ; Animals ; CD40 Ligand ; Genetic Therapy ; Genetic Vectors ; Humans ; Plasmids ; Polymerase Chain Reaction

FGF21 (fibroblast growth factor 21) is a recently described member of the FGF family. It has been previously demonstrated that FGF21 is a potent regulator of glucose homeostasis. To improve stability of FGF21 for better efficacy, a new form of recombinant FGF21 was generated by fusion of a full length FGF21 gene and the Fc fragment of human IgG4 with flexible linker sequence. To examine the glucose regulation activity of FGF21-L-Fc, 3T3-L1 pre-adipocytes were differentiated into adipocytes, and glucose uptake activity of FGF21-L-Fc was examined by glucose oxidase and peroxidase (GOD-POD) assay. The results showed that in comparison with wild type FGF21, FGF21-L-Fc was more potent in stimulation of glucose uptake by 3T3-L1. In vivo studies on the modified protein demonstrated that FGF-L-Fc had a better efficacy in lowering blood glucose of the STZ-induced diabetic animals and controlled glucose level for a longer time. The results provided a sound basis for further studies.
3T3-L1 Cells ; Adipocytes ; metabolism ; Animals ; Blood Glucose ; metabolism ; Diabetes Mellitus, Experimental ; drug therapy ; metabolism ; Escherichia coli ; metabolism ; Fibroblast Growth Factors ; genetics ; pharmacology ; Glucose ; metabolism ; Immunoglobulin G ; genetics ; pharmacology ; Male ; Mice ; Recombinant Proteins ; genetics ; pharmacology

Mouse colon cancer cells CT26 were transfected with constructed plasmid expressing mouse soluble B lymphocyte stimulator (msBlyS). Single cell clones were selected with 100 microg/ml Zeosin and subcloned by serial limiting dilution. Eight resistant transfectants were isolated and expanded, and five of them displayed the desirable msBlyS cDNA band amplified by semi-quantitative RT-PCR assay. Western blot analysis showed that only msBlyS molecules of the expected size were detected in the cell lysates from transfectants. The supernatant of transfectants could costimulate B cell proliferation in standard costimulation assay. Thus we have successfully screened the stable transfectants expressing high levels of msBlyS in CT26 cells, which could be used as cancer vaccines for further anti-tumor immunotherapy.
Animals ; Antibodies, Monoclonal ; B-Cell Activating Factor ; B-Lymphocytes ; immunology ; Cancer Vaccines ; biosynthesis ; genetics ; Cloning, Molecular ; Colonic Neoplasms ; pathology ; DNA, Complementary ; biosynthesis ; genetics ; Epitopes, B-Lymphocyte ; genetics ; Membrane Proteins ; biosynthesis ; genetics ; Mice ; Recombinant Proteins ; biosynthesis ; genetics ; Recombination, Genetic ; Transfection ; Tumor Cells, Cultured ; Tumor Necrosis Factor-alpha ; biosynthesis ; genetics

A DNA fragment encoding human interleukin 4 was obtained by PCR from pORF-hIL4 plasmid. The amplified fragment was inserted into prokaryotic expression vector PQE60 and recombinant protein was expressed in E. Coli M15 by adding isopropyl-beta-D-thiogalactoside (IPTG). The hIL-4 protein was present as insoluble inclusion bodies in the bacterial extract. After denaturation of inclusion bodies with 5 mol/L guanidine hydrochloride, the supernate was diluted to get renaturized. Then dialysis and Ni chelating chromatography were used for purification. TF-1 proliferation assay of recombinant human interleukin 4 was performed, and then rhIL-4 was fit to be used for proliferation of human dendritic cells from monocyte in vitro.
Escherichia coli ; genetics ; metabolism ; Genetic Vectors ; Humans ; Inclusion Bodies ; metabolism ; Interleukin-4 ; biosynthesis ; genetics ; Protein Folding ; Recombinant Proteins ; biosynthesis ; genetics ; isolation & purification

OBJECTIVETo silence the expression of Raf-1 gene in HNE1 cells using vector-based RNA interference (RNAi) technique.

METHODSThe vector containing the human U6 promoter was used for targeted gene silencing when a dsDNA oligonucleotide encoding an appropriate shRNA was ligated into the vector, and 67nt oligonucleotide fragment was inserted into the downstream of the U6 promoter. Plasmids containing different Raf-1 target sequences [ (1) pshuttle-Raf-1-a( 225), (2) pshattle-Raf-1-b ( 358) and (3) pshuttle-Raf-1-c(474)], were transfected into HNE1 cells. Expression of Raf-1 mRNA was assayed by RT-PCR. Apoptosis were determined by cytometry.

RESULTSVector-based RNAi had advantages over antisense RNA because it could be delivered to the target cell more efficiently, and effect could last longer. Raf-1 expression could be inhibited by plasmid-expressed shRNA. Three different targeting sequences were selected from Raf-1 gene, and the inhibitory effect of pSIREN shuttle-Raf-1-b (358) was biggest.

CONCLUSIONRaf-1 expression in HNE1 cells can be inhibited significantly using plasmid-based RNAi.

Apoptosis ; genetics ; Cell Line, Tumor ; Gene Expression ; Genetic Vectors ; Humans ; Nasopharyngeal Neoplasms ; genetics ; metabolism ; pathology ; Promoter Regions, Genetic ; Proto-Oncogene Proteins c-raf ; genetics ; RNA Interference ; RNA, Messenger ; genetics ; Transfection