Objective:To construct retroveral vectors,containing the expression sequence of vIL-10 and to transfect rabbit synoviocyte in vitro with recombinant retrovirus and detect the expression of target genes.Methods:①The primers with restrictive enzyme were designed spot and/amplify target gene by PCR from plasmid including vIL-10 gene was amplified.②The retroviral vector pLXSN of target gene was cloned,and identify the aquired plasmid by sequencing.③Co-transfecting the packaging cell GP-293 with constructed retroviral vector and assistant plasmid pVSVG by calcium phosphate-DNA co-precipitation.The medium containing recombinant virus was collected and titer of virus was determined.④Rabbit synoviocytes was transfected with acquired virus in vitro.Detect the protein expression by cell immunohistochemistry.Results:①The recombinant retrovirus rRV-vIL-10 was successfully constructed.The viral titer reached 5?106 cfu/ml.②vIL-10 gene were transduced into rabbit synoviocytes by recombinant retrovirus in vitro.The protein expression of genes could be detected by cell immunohistochemistry.Conclusion:①The recombinant retrovirus rRV-vIL-10 was successfully constructed.②vIL-10 gene were transduced successfully into the rabbit synoviocytes by retroviral vector in vitro.The transduced synoviocytes can express vIL-10 protein.

Objective:To establish a local ex-vivo gene transfer method to treat RA through animal experiments in vivo and in vitro using retrovirus(rV) as a vector which carrying rRV-vIL-10 target gene.Methods:①The rabbit RA models were induced by the rabbit synovial fibroblast cell line which could continuously expreesed hIL-1?. ②In vivo, the rabbit synovial fibroblast cell line was transduced with rRV-vIL-10, then adding G418 to pick out the rRV-vIL-10 positive clon and infecting the rabbit synovium through intra-articular injection. ③RT-PCR, IHC methods were performed to prove the success of gene transfer to the rabbit synovium and expressed the target protein. ④The relative cytokins changes were detected by ELISA before and after gene therapy and evaluated treatment efficacy of rRV-vIL-10.Results:①rRV-vIL-10 was a effective vector which could transfect to the rabbit synovium in vivo through RT-PCR and IHC methods. ②Intra-articular local gene therapy could effectly reduce the synovium inflammation level of rabbit joints and expressed mRNA and vIL-10 protein. The level of cytokin such as IL-1? was decline.Conclusion:Retrovirus-mediated transgene of vIL-10 is successfully transfected to the rabbit synovium ex-vivo and can reduce arthritis inflammation levels of the IL-1? induced arthritis.

Tumor necrosis factor alpha(TNF-?) is one of the key proinflammatory factors in driving and attending chronic inflammatory process in rheumatoid arthritis(RA).TNF-? acts on different kinds of cell in synovial membrane,such as synoviocytes,macrophages,osteoclasts and chondrocytes,which can produce metalloproteinase,collagenase,stromelysin and so on,further induce pannus formation,joint inflammation,bone erosion and cartilage degradation. The TNF inhibitors have been proved by a lot of clinical trials to be an important new group of agents that significantly improve symptoms and signs,induce remission,reduce objectively measured damage in chronic inflammatory conditions,such as RA.There are three kinds of TNF inhibitors: Etanercept,Infliximab and Adalimumab.Etanercept is a fusion protein of human IgG and two p75 TNF receptors.Infliximab is an IgG1 monoclonal antibody(a chimera of human constant and mouse variable regions).Adalimumab is a humanized IgG1 monoclonal antibody with fully human constant and variable regions.The side effects include injection site infusion reactions,infection,lymphoproliferative disease,demyelinating disease,and SLE-like syndromes.

Objective To investigate influence of Panax notoginseng saponins ( PNS) on Cyclooxygenase-2 (COX-2),Prostaglandin E2(PGE-2)and Phospholipases A2(PLA-2)in patients of acute exacerbation of chronic ob-structive pulmonary disease ( AECOPD) with respiratory failure .Methods One hundred AECOPD patients with re-spiratory failure were divided into the control group (n=50)and PNS group(n=50).The levels of blood serum COX-2,PGE-2 and PLA-2 of patients in the two groups were detected .Results The levels of blood serum COX-2,PGE-2 and PLA-2 of patients in PNS group were lower than those in control group ,the two groups after treatment significantly lower than those before treatment(P<0.05).Conclusion The levels of COX-2,PGE-2 and PLA-2 have influence on respiratory failure.PNS can reduce the COX-2,PGE-2 and PLA-2 concentrations in blood serum of patients ,mean-while it play a prominent role in inhibiting airway inflammatory and obstruction process .

IL-17 is the cytokine secreted by the subgroup of CD4+T cells named Th17.The differentiation,proliferation and cytokine secretion of Th17 are regulated by TGF-?,IL-6,IL-15 and IL-23.IL-17 modulates the production and secretion of proinflammative factors,CXCs,affecting the transfer of neutrophil,the activation and the absorption of bone.It suggests that IL-17 also plays an important role in the pathogenesis of autoimmune diseases.

BAFF is an essential ligand essential for survival and differentiation of peripheral B cells. By interacting with three receptors, BAFF can promote B cell maturation and class switching, enhance humoral immunity and T cell co-stimulation. Over-expression of BAFF leads to autoimmune diseases such as systemic lupus erythematosus (SLE) in mouse model. Treating the mice model with BAFF antagonists can slow-down disease progression and enhance survival rate. Moreover, in some SLE patients serum level of BAFF is elevated and correlated with serum anti-dsDNA titer. The preliminary clinical trial of anti-BAFF monoclonal antibody has shown to be safe and effective. BAFF antagonists are promising therapeutic drugs for SLE.

Objectives To construct two retroviral vectors, one containing human interleukin-1 recep-tor antagonist (hIL-1Ra) gene and the other containing human interleukin-10 (hIL-10) gene and to transfect rabbit synoviocytes in vitro and detect the expression level of target genes. Methods RNA from human peripheral blood mononuclear cells were extracted and target genes were amplified by RT-PCR. The target genes were cloned into retroviral vector pLXSN, which was then transducted into GP2-293 cells to produce recombinant retrovirus. Rabbit synoviocytes were transfected and the expression of target genes was detected by RT-PCR, immunohistochemistry and western-blot. Results The retroviral vector containing hIL-1Ra gene or hIL-10 gene was constructed successfully. The hIL-1Ra gene and hIL-10 gene were transduced respectively into rabbit synoviocytes in vitro. The mRNA level of both genes reached peak in 5 days. In positive cell clones, the protein level of hIL-1Ra reached peak within 30 days and maintained at least 60 days; the protein level of hIL-10 maintained at least 40 days. Conclusion The hIL-1Ra gene and hIL-10 gene can be transduced successfully into rabbit synoviocytes by recombinant retrovirus.

Objective To study the analgesic effects of combination of fortanodyn and verapamil. Methods Used the model of the acetic acid induction writhing in mice and the method of hot-plate in mice, then observe the action of the mice after treatment with combination of fortanodyn and verapamil. Results Low dose Verapamil com-bined AP237 (40 mg/kg, 80 mg/kg)could enhance the threshold of pain of the mouse ache response obviously, and postponed the time which the mouse ache appeared (comparing using simply AP237 P <0.05 with controlled group P <0. 01) ;Low dose Verapamil combined AP237 (20 mg/kg,40 mg/kg) had the obvious inhibitory action in the body turning times of the mouse. Conclusion Verapamil can strengthen the analgesia function of AP237.

Objective:To investigate the expression of Foxp3 and CD25 on CD4+ T cells from patients with new-onset systemic lupus erythematosus(SLE)and assess their clinical significance.Methods:Twenty-one new-onset and untreated SLE patients including ten with active(SLEDAI≥10)and eleven with inactive disease(SLEDAI≤5)were enrolled in this study.Eleven age-and sex-matched healthy volunteers were also included as controls.Peripheral blood samples were collected and mononuclear cells isolated.The expression of CD25,Foxp3 and CD127 by CD4+ T cells was analyzed by flow cytometry(FACS).Proliferation assays were performed with the isolated CD4+CD25+ and/or CD4+CD25-T cells.Results:There was no significant difference in number of CD4+CD25+Foxp3+T cells in subjects with either active(1.91%-6.75%)or inactive(2.74%-7.01%)SLE compared to normal controls(2.11%-9.90%)(P=0.524 and P=0.794 respectively),Moreover,suppressive capacity of CD4+CD25+T cells in new-onset lupus patients was not impaired as measured by the capacity to inhibit proliferation of CD4+CD25-effector T cells(P=0.174,P=0.689).While CD4+CD25-Foxp3+T cells in both active(3.71%-10.94%)and inactive(2.97%-7.69%)patients with new-onset SLE were significantly more frequent than in normal controls(1.01%-3.62%)(P

Objective To investigate the expression of FOXP3 and CD25 in CD4+ T cells of peripheral blood from patients with early new-onset systemic lupus erythematosus(SLE) before and after treatment.Methods Forty-four early new-onset SLE patients including twenty-four with active disease (SLEDAI≥10), twenty with low active disease (SLEDAI