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1.
Chin Med J (Engl) ; 129(9): 1047-52, 2016 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-27098789

RESUMO

BACKGROUND: Dermatomyositis (DM) and polymyositis (PM) are common inflammatory myopathies whose immunopathogenic mechanisms remain poorly understood. The NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome is a type of cytoplasmic multiprotein inflammasome and is responsible for the activation of inflammatory reactivations. Responding to a wide range of exogenous and endogenous microbial or sterile stimuli, NLRP3 inflammasomes can cleave pro-caspase-1 into active caspase-1, which processes the pro-inflammatory cytokines pro-interleukin (IL)-1ß and pro-IL-18 into active and secreted IL-1ß and IL-18. The NLRP3 inflammasome is implicated in infectious and sterile inflammatory diseases. However, it remains unclear whether it is involved in the pathogenesis of DM/PM, which we aim to address in our research. METHODS: In this study, 22 DM/PM patients and 24 controls were recruited. The protein and RNA expression of IL-1ß, IL-18, NLRP3, and caspase-1 in serum and muscle samples were tested and compared between the two groups. RESULTS: The serum IL-1ß and IL-18 levels were significantly higher in DM/PM patients than those in the controls by enzyme linked immunosorbent assay (ELISA, DM vs. control, 25.02 ± 8.29 ng/ml vs. 16.49 ± 3.30 ng/ml,P < 0.001; PM vs. control, 26.49 ± 7.79 ng/ml vs. 16.49 ± 3.30 ng/ml,P < 0.001). Moreover, the real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) showed that DM/PM patients exhibited higher RNA expression of IL-1ß, IL-18, and NLRP3 in the muscle (for IL-1ß, DM vs. control, P= 0.0012, PM vs. control, P= 0.0021; for IL-18, DM vs. control, P= 0.0045, PM vs. control, P= 0.0031; for NLRP3, DM vs. control, P= 0.0017, PM vs. control, P= 0.0006). Moreover, the protein expression of NLRP3 and caspase-1 in muscle samples of DM/PM patients were also significantly elevated compared to that in the muscles of the controls. CONCLUSIONS: Our findings demonstrate that the NLRP3 inflammasome is implicated in the pathogenesis of DM/PM. High NLRP3 expression led to elevated levels of IL-1ß and IL-18 and could be one of the factors promoting disease progress.


Assuntos
Dermatomiosite/etiologia , Inflamassomos/fisiologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/fisiologia , Polimiosite/etiologia , Adulto , Caspase 1/análise , Caspase 1/genética , Feminino , Humanos , Interleucina-18/análise , Interleucina-18/genética , Interleucina-1beta/análise , Interleucina-1beta/genética , Masculino , Pessoa de Meia-Idade , Proteína 3 que Contém Domínio de Pirina da Família NLR/análise , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética
2.
Int J Clin Exp Med ; 8(3): 3965-73, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26064298

RESUMO

OBJECTIVE: To investigate the effect of Foxp3 gene modified dendritic cells (Foxp3 + DC) on allogeneic T cells proliferation and to study the effect of Foxp3 + DC on corneal allograft rejection. METHODS: Lentivirus-Foxp3 was transfected into DC2.4 cells, as Foxp3 + DC cells. 42 BALB/c mice were randomly divided into: Group A (n = 6), normal group; Group B (n = 12), Group C (n = 12) and Group D (n = 12), allograft groups, were treated with normal saline, DC2.4, Foxp3 + DC by intraperitoneal injection, respectively. RESULTS: Compared with the control group, Foxp3 protein in the Foxp3 + DC cells increased significantly (P < 0.05); the expressions of CD80 and CD86 immunophenotypes of Foxp3 + DC cells decreased significantly (P < 0.05); IL-12 secretion reduced (P < 0.05), but IL-10 secretion was promoted (P < 0.05). The average transplant survival time in Group B was (14.833 ± 1.472) d, and Group C and Group D led to a statistically significant prolongation of transplant survival to (17.667 ± 1.366, 23.000 ± 2.000) d (P < 0.05) respectively. 14 d after transplantation, as compared with Group C and D, the expressions of IFN-γ in grafts markedly increased in Group B. 14 d after transplantation, as compared with Group B, the expressions of Foxp3 mRNA, IDO mRNA in grafts decreased remarkably in Group C and D (P < 0.05); as compared with Group C, the expressions of Foxp3 mRNA, IDO mRNA in grafts decreased remarkably in Group D (P < 0.05). CONCLUSION: Foxp3 + DC cells reduce the expression of costimulatory factors, reduce the secretion of IL-12, promote IL-10 production and inhibit the stimulation of alloreactive T cell proliferation response capacity. Foxp3 + DC cells play important roles in inhibiting corneal allograft immune response and prolonging graft survival time.

3.
Zhongguo Zhong Yao Za Zhi ; 39(2): 278-84, 2014 Jan.
Artigo em Chinês | MEDLINE | ID: mdl-24761646

RESUMO

OBJECTIVE: To evaluate the therapeutic effect of Liujunzi decoction combined with Zuojin pills in treating the radioactive duodenitis and their mechanism, and compare with clinical routine acid suppressants combined with mucous membrane protective preparations to study the mechanism of their efficacy. METHOD: According to the study of Williams J P and characteristics of duodenitis, and by reference to the radiation enteritis modeling standard, we took the lead in establishing the mouse radioactive duodenal injury model. The model mice were randomly divided into the control group (n = 26), traditional Chinese medicine (TCM) group (n = 16) and the western medicine (oral administration with famotidine 0.5 mL + almagate suspension 0.3 mL per mouse, once a day) group (n = 16). After the standard administrating, such objective indexes as general condition, weight, changes in health score, pathology and expression of inflammatory factors were observed to evaluate the efficacy. RESULT: The radioactive duodenitis model of mice was successfully established with 12 Gy. Mice in the control group suffered from weight loss, anorexia, low fluid intake, loose stools, and occasionally mucous bloody stool, poor spirit, dim fur, lack of exercise and arch back. Mice in drug intervention groups were generally better than those in the pure irradiation group. The IL-6, IL-1beta, TNF-alpha mRNA expressions in spleen and mesenteric lymph node tissues in TCM and western medicine groups showed a declining trend compared with the control group. Their concentrations in peripheral blood serum also slightly changed. The TCM group revealed notable advantage in reducing inflammatory factors. The microscopic observation showed that a better mucosa repair in intervention groups than the pure irradiation group. The improved Chiu's scoring method showed a statistical significance in the difference between TCM and western medicine groups (P < 0.05). CONCLUSION: Liujunzi decoction combined with Zuojin pills could treat acute radiation enteritis, regulate organic immunity, and inhibit acute injury, promote local tissue repair, with the potential to resist such adverse effects as radiation intestinal fibrosis. The regulation of inflammatory factor release is one of efficacy generation mechanisms.


Assuntos
Medicamentos de Ervas Chinesas/farmacologia , Duodenite/tratamento farmacológico , Lesões Experimentais por Radiação/tratamento farmacológico , Animais , Radioisótopos de Cobalto/efeitos adversos , Interações de Medicamentos , Medicamentos de Ervas Chinesas/uso terapêutico , Duodenite/sangue , Interleucina-1beta/sangue , Interleucina-6/sangue , Camundongos , Camundongos Endogâmicos BALB C , Membrana Mucosa/efeitos dos fármacos , Membrana Mucosa/efeitos da radiação , Lesões Experimentais por Radiação/sangue , Fator de Necrose Tumoral alfa/sangue
4.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 27(7): 721-4, 2011 Jul.
Artigo em Chinês | MEDLINE | ID: mdl-21722519

RESUMO

AIM: To construct a Foxp3 lentiviral vector and transfer it into DC2.4 cells, which provides Foxp3+DC cells for further study on its immune modulation. METHODS: We cloned mouse Foxp3 gene into lentiviral vector(pGC-FU) and acquired the plasmid pGC-FU-Foxp3. PCR and sequencing analysis were made for verifying the positive clones. The virus packaging plasmids(pGC-FU-Foxp3, pHelper1.0 and pHelper2.0) were contransfected into 293T cells, and the Lentivirus-Foxp3 was harvested from 293T cells. The Lentivirus-Foxp3 was used to infect DC2.4 cells in vitro and the expression of Foxp3 in infected DC2.4 cells was detected with flow cytometry(FCM). RESULTS: PCR and sequencing revealed that the pGC-FU-Foxp3 plasmid was correctly constructed. The Lentivirus-Foxp3 with a titer of 2×10(8); TU/mL was successfully packaged. Foxp3 expression in DC2.4 cells infected with the Lentivirus-Foxp3 was increased significantly compared with negative control lentivirus. CONCLUSION: The pGC-FU-Foxp3 plasmid has been successfully constructed and the Lentivirus-Foxp3 has been successfully packaged. Foxp3 can be enhanced in DC cells infected with the Lentivirus-Foxp3.


Assuntos
Células Dendríticas/metabolismo , Fatores de Transcrição Forkhead/genética , Vetores Genéticos/genética , Lentivirus/genética , Animais , Sequência de Bases , Linhagem Celular , Células Dendríticas/imunologia , Expressão Gênica , Imunomodulação , Camundongos , Reação em Cadeia da Polimerase , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
5.
Med Sci Monit ; 17(5): BR125-31, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21525800

RESUMO

BACKGROUND: The aim of this study was to investigate the mechanisms underlying tolerance induction of dexamethasone (Dex)-treated dendritic cells (DCs). MATERIAL/METHODS: Well-grown DC2.4 cells were randomly assigned to receive control, 50 µg/L, 100 µg/L, or 200 µg/L of dexamethasone and then were cultured for 6 days. The expressions of CD80, CD86, galectin-9, and PD-L1 on the surface of DC2.4 cells were analyzed with flow cytometry and the level of IL-12 secreted by DC2.4 cells was determined by ELISA. The stimulating activity of DC2.4 cells on allogeneic T cells was assessed with mixed lymphocyte reaction. Dexamethasone-treated DC2.4 cells were co-cultured with allogeneic splenic lymphocytes and the Foxp3 expression in naive T lymphocytes was determined with flow cytometry. RESULTS: Compared with the control group, the expressions of CD80, CD86, galectin-9, and PD-L1 on the surface of DC2.4 cells exposed to different doses of dexamethasone showed no significant changes; however, dexamethasone treatment significantly reduced IL-12 secretion and inhibited DC2.4's stimulation on the proliferation of allogeneic T lymphocytes. Moreover, dexamethasone-treated DC2.4 cells effectively promoted FOXP3 expression in naive T lymphocytes. CONCLUSIONS: DC2.4 is a stable cell line with high expressions of CD80, CD86, and PD-L1. Dexamethasone does not significantly change the cell phenotype of DC2.4 cells, but inhibits the secretion of IL-12 cytokine and attenuates DC2.4's stimulation of the proliferation of allogeneic T cells. Dexamethasone-treated DC2.4 cells also effectively promote FOXP3 expression in naive T lymphocytes.


Assuntos
Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Dexametasona/farmacologia , Tolerância Imunológica/efeitos dos fármacos , Animais , Antígeno B7-1/metabolismo , Antígeno B7-2/metabolismo , Adesão Celular/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Células Dendríticas/citologia , Citometria de Fluxo , Fatores de Transcrição Forkhead/metabolismo , Galectinas/metabolismo , Interleucina-12/metabolismo , Teste de Cultura Mista de Linfócitos , Camundongos , Frações Subcelulares/efeitos dos fármacos , Frações Subcelulares/metabolismo , Fatores de Tempo
6.
Zhongguo Shi Yan Xue Ye Xue Za Zhi ; 18(3): 766-70, 2010 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-20561447

RESUMO

After treating with chemotherapy or immunosuppressant, malignant diseases of hematopoietic system such as leukemia, malignant lymphoma and aplastic anemia usually induced severe infection such as sepsis. Sepsis which is hard to be diagnosed causes high death rate. This study was purposed to establish an experimental sepsis mouse model so as to provide a basis for pathogenesis and intervention study. A classic caecal ligation and puncture (CLP) was used to establish experimental sepsis model. ELISA was used to detect levels of C5a, IL-6, TNFalpha, and IFN-gamma. Flow Cytometry was applied to measure apoptosis of lymphocytes in thymus and mesentery. The pathologic changes of thymus and spleen were confirmed by HE staining. The results showed that almost 70%-80% mice died at 72 hours after CLP. Only approximate 20% animal survived during finite time, mice in CLP group had significant weight lose. Meanwhile large release of different inflammatory mediators which are related with sepsis (C5a, IL-6, TNF-alpha, and IFN-gamma) was observed after CLP. Apoptosis of lymphocytes in thymus and mesentery lymphonodus was enhanced markedly after CLP. Significantly pathologic injury was also observed in thymus and spleen. It is concluded that a mouse model of experimental sepsis was successfully established by caecal ligation and puncture which can well mimic the clinical symptom of sepsis. The experimental sepsis mouse model provides an excellent tool for exploring the pathogenesis and intervention ways for sepsis accompanied with complicated malignant hematological diseases in vivo.


Assuntos
Modelos Animais de Doenças , Sepse , Animais , Apoptose , Ceco/lesões , Complemento C5a/metabolismo , Interferon gama/metabolismo , Interleucina-6/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Sepse/metabolismo , Sepse/patologia , Baço/patologia , Timo/patologia , Fator de Necrose Tumoral alfa/metabolismo
7.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 21(1): 24-8, 2005 Jan.
Artigo em Chinês | MEDLINE | ID: mdl-15629077

RESUMO

AIM: To develop rAAV2 containing cDNA of GAD-Ig fusion gene [GAD-Ig fusion gene is constructed by inserting glutamic acid decarboxylase(GAD) into N-terminal of murine IgG3 heavy chain(Ig)], and to study whether rAAV2 mediated GAD-Ig fusion gene expression could induce specific tolerance in IDDM animal model-nonobese diabetic (NOD) mice. METHODS: GAD-Ig cDNA was amplified by PCR from the plasmid BSSK-GAD-Ig and inserted into an eukaryotic expression plasmid pSNAV to construct a recombinant expression plasmid pSNAV/GAD-Ig. Plasmid pSNAV-GAD-Ig was then transfected into the rAAV2 packaging cell-BHK-21 cells using LipofectAMINE TM 2000 to produce rAAV-GAD-Ig. Target gene expression in BHK-21 cells infected by rAAV-GAD-Ig was confirmed by RT-PCR, Western blot and ELISA respectively. Then rAAV-GAD-Ig was injected into intramuscularly NOD mice. Antigen-specific tolerance in NOD mice was detected by specific lymphocytic proliferation reaction to GAD antigen. RESULTS: GAD-Ig gene was inserted into the genome of target cells. Target cells transfected by rAAV2-GAD-Ig could secret GAD-Ig. rAAV2-GAD-Ig induced specific tolerance in NOD mice. CONCLUSION: rAAV2-GAD-Ig was obtained, which can be used in IDDM gene therapy in NOD mice.


Assuntos
DNA Recombinante/genética , Dependovirus/genética , Diabetes Mellitus Tipo 1/terapia , Terapia Genética , Glutamato Descarboxilase/genética , Imunoglobulina G/genética , Cadeias Pesadas de Imunoglobulinas/genética , Animais , Fusão Gênica Artificial , Linhagem Celular , Proliferação de Células , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Expressão Gênica , Vetores Genéticos/genética , Glutamato Descarboxilase/imunologia , Tolerância Imunológica , Imunoglobulina G/imunologia , Cadeias Pesadas de Imunoglobulinas/imunologia , Linfócitos/citologia , Linfócitos/imunologia , Camundongos , Camundongos Endogâmicos NOD , Reação em Cadeia da Polimerase , Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
8.
Cell Mol Immunol ; 2(6): 461-5, 2005 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-16426497

RESUMO

CD3-specific monoclonal antibody was the first one used for clinical practice in field of transplantation. Recently, renewed interests have elicited in its capacity to prevent autoimmune diabetes by inducing immune tolerance. In this study, we tested whether this antibody can also be used to treat another kind of autoimmune disease myasthenia gravis (MG) and explored the possible mechanisms. MG is caused by an autoimmune damage mediated by antibody- and complement-mediated destruction of AChR at the neuromuscular junction. We found that administration of CD3-specific antibody (Fab)2 to an animal model with experimental autoimmune myasthenia gravis (EAMG) (B6 mice received 3 times of AChR/CFA immunization) could not significantly improve the clinical signs and clinical score. When the possible mechanisms were tested, we found that CD3 antibody treatment slightly down-regulated the T-cell response to AChR, modestly up-regulating the muscle strength. And no significant difference in the titers of IgG2b was found between CD3 antibody treated and control groups. These data indicated that CD3-specific antibody was not suitable for treating MG, an antibody- and complement- mediated autoimmune disease, after this disease has been established. The role of CD3-specific antibody in treating this kind of disease remains to be determined.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/uso terapêutico , Complexo CD3/imunologia , Miastenia Gravis Autoimune Experimental/tratamento farmacológico , Miastenia Gravis Autoimune Experimental/imunologia , Animais , Proliferação de Células/efeitos dos fármacos , Eletromiografia , Feminino , Linfócitos/citologia , Linfócitos/efeitos dos fármacos , Camundongos , Miastenia Gravis Autoimune Experimental/diagnóstico , Miastenia Gravis Autoimune Experimental/prevenção & controle , Baço/citologia , Baço/efeitos dos fármacos
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