AIM： To investigate genetic diversity of Helicobacter pylori （H. pylorl） cell division-related gene A （cdrA） and its effect on the host response.METHODS： Inactivation of H. py/ori cdrA, which is involved in ceil...AIM： To investigate genetic diversity of Helicobacter pylori （H. pylorl） cell division-related gene A （cdrA） and its effect on the host response.METHODS： Inactivation of H. py/ori cdrA, which is involved in ceil division and morphological elonga- tion, has a role in chronic persistent infections. Ge- netic property of H. pylori cdrA was evaluated using polymerase chain reaction and sequencing in 128 （77 American and 51 Japanese） clinical isolates obtained from 48 and 51 patients, respectively. Enzyme-linked immunosorbent assay was performed to measure in- terleukin-8 （IL-8） secretion with gastric biopsy speci- mens obtained from American patients colonized with cdrA-positive or -negative strains and AGS cells co- cultured with wild-type HPK5 （cdrA-positive） or its de- rivative HPKT510 （cdrA-disruptant）. Furthermore, the cytotoxin-associated gene A （cagA） status （transloca- tion and phosphorylation） and kinetics of transcription factors [nuclear factor-kappa B （NF-~：B） and inhibition kappa B] were investigated in AGS cells co-cultured with HPK5, HPKT510 and its derivative HPKSCA （cagA- disruptant） by western blotting analysis with immuno- precipitation. RESULTS： Genetic diversity of the H. pylori cdrA gene demonstrated that the cdrA status segregated into two categories including four allele types, cdrA-positive （al- lele types, I and 11 ） and cdrA-negative （allele types; 111 and IV） categories, respectively. Almost all Japanese isolates were cdrA-positive （ 1 ： 7.8% and 11 ： 90.2%）, whereas 16.9% of American isolates were cdrA-positive （11） and 83.1% were cdrA-negative （nl： 37.7% and IV： 45.5%）, indicating extended diversity of cdrA in individual American isolates. Comparison of each isolate from different regions （antrum and corpus） in the stomach of 29 Americans revealed that cdrA status was identical in both isolates from different regions in 17 cases. However, 12 cases had a different cdrA al- lele and 6 of them exhibited a different cdrA category between two regions in the stomach. Furthermore, in 5 of the 6 cases possessing a different cdrA category, cdrA-negative isolate existed in the corpus, suggesting that cdrA-negative strain is more adaptable to coloni- zation in the corpus. IL-8 secretions from AGS revealed that IL-8 levels induced by a cdrA-disrupted HPKT510 was significantly lower （P 〈 0.01） compared to wild- type HPK5： corresponding to 50%-60% of those of wild-type HPK5. These data coincided with in vivo data that an average value of IL-8 in biopsy specimens from cdrA-positive and cdrA-negative groups was 215.6 and 135.9 pg/mL, respectively. Western blotting analysis documented that HPKT510 had no effect on CagA translocation and phosphorylation, however, nuclear accumulation of NF-κB was lower by HPKT510 com- pared to HPK5. CONCLUSION： Colonization by a cdrA-negative or cdrA-dysfunctional strain resulted in decreased IL-8 production and repression of NF-κB, and hence, atten- uate the host immunity leading to persistent infection.展开更多
Summary: To investigate the role of NF-κB in endotoxic shock in rats, the model of endotoxin-shock rats was induced by intravenous infusion of lipopolysaccharide (LPS). 1 h, 2 h, 4 h and 6 h after LPS injection, the...Summary: To investigate the role of NF-κB in endotoxic shock in rats, the model of endotoxin-shock rats was induced by intravenous infusion of lipopolysaccharide (LPS). 1 h, 2 h, 4 h and 6 h after LPS injection, the activation of NF-κB in blood mononuclear cells and the content of TNF-α and IL-6 in plasma was detected by enzyme-linked immunoadsordent assay (ELISA). The level of mean arterial pressure (MAP) and the histopathological changes of lung and liver were also observed. The activation of NF-κB in mononuclear cells increased 1 h after LPS injection and reached its peak 2 h after the injection, and its level was higher than that of normal group. The level of TNF-α was increased 1 h after the infusion and peaked 2 h after the injection, and its level was higher than that of normal group after LPS infusion. The content of IL-6 increased gradually with time, the IL-6 level was higher than that of normal group after LPS injection. MAP was decreased gradually with time and its level was lower than that of normal group after LPS injection. Pathological examination showed that endotoxic shock could cause pulmonary alveolar hemorrhage, edema and infiltration of inflammatory cell in lung tissue and congestion, edema, capillary dilation and inflammatory cell infiltration in liver tissue. It is concluded that NF-κB can up-regulate the expression of TNF-α and IL-6 in plasma and play an important role in endotoxin-induced shock in rats.展开更多
Recent studies indicated that regulatory B cells(Bregs)and nuclear factor erythroid 2-related factor 2(Nrf2)antioxidant signaling pathway play important roles in the pathogenesis of chronic graft-versus-host disease(c...Recent studies indicated that regulatory B cells(Bregs)and nuclear factor erythroid 2-related factor 2(Nrf2)antioxidant signaling pathway play important roles in the pathogenesis of chronic graft-versus-host disease(cGVHD).Mangiferin(MA),a polyphenol compound,has been reported to activate Nrf2/antioxidant-responsive element(ARE)signaling pathway.This study was aimed to investigate the effects of MA on Bregs and Nrf2 antioxidant signaling in murine splenic mononuclear cells(MNCs)in vitro.Our results revealed that MA could increase the Bregs level in murine splenic MNCs.Moreover,MA up-regulated the expression of Bregs-associated immunosuppressive factor interleukin-10(IL-10)by activating the Janus kinase 2(JAK2)/signal transducer and activator of transcription 3(STAT3)and extracellular signal-regulated kinase(ERK)signaling in murine splenic MNCs.Meanwhile,MA inhibited the proinflammatory cytokines IL-2 and interferon-y(INF-y)at both mRNA and protein levels.MA also enhanced the transcription and protein expression of Nrf2 and NADPH quinine oxidoreductase 1(NQOl),whereas decreased that of Kelch-like ECH-associated protein 1(Keapl)in murine splenic MNCs.Moreover,MA promoted the proliferation and inhibited the apoptosis of murine splenic MNCs.These results suggested that MA exerts immunosuppressive effects by upregulating the Bregs level,activating the Nrf2 antioxidant pathway,and inhibiting the expression of pro-immunoinflammatory factors.MA,as a natural immunomodulatory and anti-inflammatory agent,may have a potential role in the prophylaxis and treatment of cGVHD.展开更多
目的:探讨p38丝裂原活化蛋白激酶(p38 mitogen-activated protein kinase,p38MAPK)、核因子-κB(nuclearfactor-κB,NF-κB)和环氧化酶-2(cyclooxygenase-2,COX-2)之间的关系,从而研究p38MAPK和NF-κB、COX-2在糖尿病肾病中的作用机制...目的:探讨p38丝裂原活化蛋白激酶(p38 mitogen-activated protein kinase,p38MAPK)、核因子-κB(nuclearfactor-κB,NF-κB)和环氧化酶-2(cyclooxygenase-2,COX-2)之间的关系,从而研究p38MAPK和NF-κB、COX-2在糖尿病肾病中的作用机制。方法:分别以一定浓度的高葡萄糖(25mmol/L)、高胰岛素(100mmol/L)、过氧化氢(100μmol/L)和糖基化终产物(100mg/L)孵育大鼠肾小球系膜细胞系HBZY-1一定时间;先分别以一定浓度的p38MAPK特异抑制剂SB203580(10μmol/L)预处理细胞系HBZY-1,再给予上述4种因素孵育细胞系HBZY-1,观察细胞系HBZY-1p38MAPK、NF-κB和COX-2的表达。结果:高葡萄糖、高胰岛素、过氧化氢和糖基化终产物均可独立激活p38MAPK,使其磷酸化表达量增加,NF-κB、COX-2表达也明显增加,与对照组比较有显著性差异(P<0.01);SB203580预处理后,NF-κB、COX-2表达显著降低,与相应刺激组比较差异有统计学意义(P<0.01)。结论:p38MAPK可通过激活NF-κB、COX-2而诱导DM时肾脏的损害,p38MAPK和NF-κB、COX-2在糖尿病肾病的发生发展过程中可能起重要作用。展开更多
目的:研究人牙囊细胞白细胞介素-10(interleukin-10,IL-10)蛋白的表达及其对破骨细胞分化因子(receptor activator of nuclear factor kappa B ligand,RANKL)表达的影响。方法:采用组织块加胶原酶消化法培养人牙囊细胞,进行IL-10免疫组...目的:研究人牙囊细胞白细胞介素-10(interleukin-10,IL-10)蛋白的表达及其对破骨细胞分化因子(receptor activator of nuclear factor kappa B ligand,RANKL)表达的影响。方法:采用组织块加胶原酶消化法培养人牙囊细胞,进行IL-10免疫组化染色。25ng/mlIL-10作用于牙囊细胞0、1、3、6 h,用RT-PCR检测RANKL mRNA表达的变化。结果:人牙囊细胞IL-10表达阳性。25ng/ml IL-10降低人牙囊细胞RANKL的表达,且具有时间依赖性。结论:人牙囊细胞表达IL-10,IL-10通过降低人牙囊细胞RANKL的表达,在牙齿萌出过程中起重要的调控作用。展开更多
Background Experimental autoimmune myocarditis （EAM） in rats is a T-cell-mediated disorder. The initiation and maintenance of autoimmune responses in EAM depend on the maturation state of dendritic cells. IL-10 is a...Background Experimental autoimmune myocarditis （EAM） in rats is a T-cell-mediated disorder. The initiation and maintenance of autoimmune responses in EAM depend on the maturation state of dendritic cells. IL-10 is a pleiotrophic immunomodulatory cytokine that functions at different levels of the immune response, so it has emerged as a promising therapeutic factor for the treatment of autoimmune/inflammatory diseases. This study was designed to test the hypothesis that IL-10 gene modified bone marrow-derived immature dendritic cells （iDCs） ameliorate EAM and to explore the underlying mechanisms. Methods EAM was induced using the methods of cardiac myosin immunization on day 0 and day 7. Immature and mature bone marrow-derived dendritic cells （BMDCs） were generated without or with the stimulation by lipopolysaccharide （LPS） and the phenotype was analyzed by flow cytometry. Some of the iDCs were transfected by pcDNA3-IL-10 plasmid. 2× 10^6/per rat mature DC （mDC）, immature DC （iDC）, pcDNA3 transfected iDC, pcDNA3-IL-10 transfected iDC or phosphate buffered saline （PBS） were injected intravenously for treatment 5 days after the first immunization. On day 21, HE staining was performed to detect the myocardial inflammation and T lymphocyte proliferation assay was used to determine the effects of IL-10 gene transfected iDC on autoreactive T cell proliferation. Expression of IκB, the inhibitor of NF-κB pathway, was determined by Western blot. Results BMDCs generated in a medium supplemented with granulocyte-macrophage-colony-stimulating factor （GM-CSF） were relatively immature, as determined by flow cytometry. However, stimulation with LPS induced these cells to become mature （m）DCs with higher levels of surface major histocompatibility complex （MHC）-Ⅱ and costimulatory molecules. Intravenous administration of iDCs, especially pcDNA3-IL-10 transfected iDC, ameliorated the histopathological severity of the myosin induced-EAM, and the effect was lost after the DCs underwent maturation induced by in vitro exposure to LPS. IL-10 gene modified iDC inhibited the antigen specific T cell responses towards cardiac myosin. IκB protein was up-regulated significantly in the IL-10 gene modified iDC group. Conclusions IL-10 gene modified iDC induced antigen-specific tolerance in EAM. The underlying mechanisms may be related to costimulatory molecules down-regulation and NF-κB pathway inhibition.展开更多
基金Supported by The Project Research Fund from Kochi University,to Takeuchi Ha Grant-in-Aid for Scientific Research from the Ministry of Education,Science and Culture of Japan,No. 21590631 and 21590629,in part
文摘AIM： To investigate genetic diversity of Helicobacter pylori （H. pylorl） cell division-related gene A （cdrA） and its effect on the host response.METHODS： Inactivation of H. py/ori cdrA, which is involved in ceil division and morphological elonga- tion, has a role in chronic persistent infections. Ge- netic property of H. pylori cdrA was evaluated using polymerase chain reaction and sequencing in 128 （77 American and 51 Japanese） clinical isolates obtained from 48 and 51 patients, respectively. Enzyme-linked immunosorbent assay was performed to measure in- terleukin-8 （IL-8） secretion with gastric biopsy speci- mens obtained from American patients colonized with cdrA-positive or -negative strains and AGS cells co- cultured with wild-type HPK5 （cdrA-positive） or its de- rivative HPKT510 （cdrA-disruptant）. Furthermore, the cytotoxin-associated gene A （cagA） status （transloca- tion and phosphorylation） and kinetics of transcription factors [nuclear factor-kappa B （NF-~：B） and inhibition kappa B] were investigated in AGS cells co-cultured with HPK5, HPKT510 and its derivative HPKSCA （cagA- disruptant） by western blotting analysis with immuno- precipitation. RESULTS： Genetic diversity of the H. pylori cdrA gene demonstrated that the cdrA status segregated into two categories including four allele types, cdrA-positive （al- lele types, I and 11 ） and cdrA-negative （allele types; 111 and IV） categories, respectively. Almost all Japanese isolates were cdrA-positive （ 1 ： 7.8% and 11 ： 90.2%）, whereas 16.9% of American isolates were cdrA-positive （11） and 83.1% were cdrA-negative （nl： 37.7% and IV： 45.5%）, indicating extended diversity of cdrA in individual American isolates. Comparison of each isolate from different regions （antrum and corpus） in the stomach of 29 Americans revealed that cdrA status was identical in both isolates from different regions in 17 cases. However, 12 cases had a different cdrA al- lele and 6 of them exhibited a different cdrA category between two regions in the stomach. Furthermore, in 5 of the 6 cases possessing a different cdrA category, cdrA-negative isolate existed in the corpus, suggesting that cdrA-negative strain is more adaptable to coloni- zation in the corpus. IL-8 secretions from AGS revealed that IL-8 levels induced by a cdrA-disrupted HPKT510 was significantly lower （P 〈 0.01） compared to wild- type HPK5： corresponding to 50%-60% of those of wild-type HPK5. These data coincided with in vivo data that an average value of IL-8 in biopsy specimens from cdrA-positive and cdrA-negative groups was 215.6 and 135.9 pg/mL, respectively. Western blotting analysis documented that HPKT510 had no effect on CagA translocation and phosphorylation, however, nuclear accumulation of NF-κB was lower by HPKT510 com- pared to HPK5. CONCLUSION： Colonization by a cdrA-negative or cdrA-dysfunctional strain resulted in decreased IL-8 production and repression of NF-κB, and hence, atten- uate the host immunity leading to persistent infection.
基金This project was supported by a grant from Hubei Province Science and Technology Foundation (2003AA301C51).
文摘Summary: To investigate the role of NF-κB in endotoxic shock in rats, the model of endotoxin-shock rats was induced by intravenous infusion of lipopolysaccharide (LPS). 1 h, 2 h, 4 h and 6 h after LPS injection, the activation of NF-κB in blood mononuclear cells and the content of TNF-α and IL-6 in plasma was detected by enzyme-linked immunoadsordent assay (ELISA). The level of mean arterial pressure (MAP) and the histopathological changes of lung and liver were also observed. The activation of NF-κB in mononuclear cells increased 1 h after LPS injection and reached its peak 2 h after the injection, and its level was higher than that of normal group. The level of TNF-α was increased 1 h after the infusion and peaked 2 h after the injection, and its level was higher than that of normal group after LPS infusion. The content of IL-6 increased gradually with time, the IL-6 level was higher than that of normal group after LPS injection. MAP was decreased gradually with time and its level was lower than that of normal group after LPS injection. Pathological examination showed that endotoxic shock could cause pulmonary alveolar hemorrhage, edema and infiltration of inflammatory cell in lung tissue and congestion, edema, capillary dilation and inflammatory cell infiltration in liver tissue. It is concluded that NF-κB can up-regulate the expression of TNF-α and IL-6 in plasma and play an important role in endotoxin-induced shock in rats.
基金the National Natural Science Foundation of China(No.81470347,No.81974003)The authors would like to thank the Department of Central Laboratory,Union Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan,China,for providing relevant experimental facilities and technical support.
文摘Recent studies indicated that regulatory B cells(Bregs)and nuclear factor erythroid 2-related factor 2(Nrf2)antioxidant signaling pathway play important roles in the pathogenesis of chronic graft-versus-host disease(cGVHD).Mangiferin(MA),a polyphenol compound,has been reported to activate Nrf2/antioxidant-responsive element(ARE)signaling pathway.This study was aimed to investigate the effects of MA on Bregs and Nrf2 antioxidant signaling in murine splenic mononuclear cells(MNCs)in vitro.Our results revealed that MA could increase the Bregs level in murine splenic MNCs.Moreover,MA up-regulated the expression of Bregs-associated immunosuppressive factor interleukin-10(IL-10)by activating the Janus kinase 2(JAK2)/signal transducer and activator of transcription 3(STAT3)and extracellular signal-regulated kinase(ERK)signaling in murine splenic MNCs.Meanwhile,MA inhibited the proinflammatory cytokines IL-2 and interferon-y(INF-y)at both mRNA and protein levels.MA also enhanced the transcription and protein expression of Nrf2 and NADPH quinine oxidoreductase 1(NQOl),whereas decreased that of Kelch-like ECH-associated protein 1(Keapl)in murine splenic MNCs.Moreover,MA promoted the proliferation and inhibited the apoptosis of murine splenic MNCs.These results suggested that MA exerts immunosuppressive effects by upregulating the Bregs level,activating the Nrf2 antioxidant pathway,and inhibiting the expression of pro-immunoinflammatory factors.MA,as a natural immunomodulatory and anti-inflammatory agent,may have a potential role in the prophylaxis and treatment of cGVHD.
文摘目的:探讨p38丝裂原活化蛋白激酶(p38 mitogen-activated protein kinase,p38MAPK)、核因子-κB(nuclearfactor-κB,NF-κB)和环氧化酶-2(cyclooxygenase-2,COX-2)之间的关系,从而研究p38MAPK和NF-κB、COX-2在糖尿病肾病中的作用机制。方法:分别以一定浓度的高葡萄糖(25mmol/L)、高胰岛素(100mmol/L)、过氧化氢(100μmol/L)和糖基化终产物(100mg/L)孵育大鼠肾小球系膜细胞系HBZY-1一定时间;先分别以一定浓度的p38MAPK特异抑制剂SB203580(10μmol/L)预处理细胞系HBZY-1,再给予上述4种因素孵育细胞系HBZY-1,观察细胞系HBZY-1p38MAPK、NF-κB和COX-2的表达。结果:高葡萄糖、高胰岛素、过氧化氢和糖基化终产物均可独立激活p38MAPK,使其磷酸化表达量增加,NF-κB、COX-2表达也明显增加,与对照组比较有显著性差异(P<0.01);SB203580预处理后,NF-κB、COX-2表达显著降低,与相应刺激组比较差异有统计学意义(P<0.01)。结论:p38MAPK可通过激活NF-κB、COX-2而诱导DM时肾脏的损害,p38MAPK和NF-κB、COX-2在糖尿病肾病的发生发展过程中可能起重要作用。
基金This study was supported by Harbin Medical University Postgraduate Creation Foundation (2005).
文摘Background Experimental autoimmune myocarditis （EAM） in rats is a T-cell-mediated disorder. The initiation and maintenance of autoimmune responses in EAM depend on the maturation state of dendritic cells. IL-10 is a pleiotrophic immunomodulatory cytokine that functions at different levels of the immune response, so it has emerged as a promising therapeutic factor for the treatment of autoimmune/inflammatory diseases. This study was designed to test the hypothesis that IL-10 gene modified bone marrow-derived immature dendritic cells （iDCs） ameliorate EAM and to explore the underlying mechanisms. Methods EAM was induced using the methods of cardiac myosin immunization on day 0 and day 7. Immature and mature bone marrow-derived dendritic cells （BMDCs） were generated without or with the stimulation by lipopolysaccharide （LPS） and the phenotype was analyzed by flow cytometry. Some of the iDCs were transfected by pcDNA3-IL-10 plasmid. 2× 10^6/per rat mature DC （mDC）, immature DC （iDC）, pcDNA3 transfected iDC, pcDNA3-IL-10 transfected iDC or phosphate buffered saline （PBS） were injected intravenously for treatment 5 days after the first immunization. On day 21, HE staining was performed to detect the myocardial inflammation and T lymphocyte proliferation assay was used to determine the effects of IL-10 gene transfected iDC on autoreactive T cell proliferation. Expression of IκB, the inhibitor of NF-κB pathway, was determined by Western blot. Results BMDCs generated in a medium supplemented with granulocyte-macrophage-colony-stimulating factor （GM-CSF） were relatively immature, as determined by flow cytometry. However, stimulation with LPS induced these cells to become mature （m）DCs with higher levels of surface major histocompatibility complex （MHC）-Ⅱ and costimulatory molecules. Intravenous administration of iDCs, especially pcDNA3-IL-10 transfected iDC, ameliorated the histopathological severity of the myosin induced-EAM, and the effect was lost after the DCs underwent maturation induced by in vitro exposure to LPS. IL-10 gene modified iDC inhibited the antigen specific T cell responses towards cardiac myosin. IκB protein was up-regulated significantly in the IL-10 gene modified iDC group. Conclusions IL-10 gene modified iDC induced antigen-specific tolerance in EAM. The underlying mechanisms may be related to costimulatory molecules down-regulation and NF-κB pathway inhibition.
