Anti-interleukin 6 (IL-6) receptor antibody suppresses Castleman's disease like symptoms emerged in IL-6 transgenic mice

Transgenic mice carrying human IL-6 cDNA fused with a murine major histocompatibility class-I promoter (H-2L(d)) were serially administered with anti-interleukin-6 receptor (IL-6R) monoclonal antibody (mAb), MR16-1, from the age of 4 weeks to estimate its efficacy on a variety of disorders developed in these mice, most of which are similar to the disorders associated with Castleman's disease. In the control mice treated with isotype-matched mAb, a massive and multiple IgG1 plasmacytosis, mesangial proliferative glomerulonephritis, leukocytosis, thrombocytosis, anemia and abnormalities of blood chemical parameters have developed in accordance with the elevation of serum IL-6, and 50% of mice have died of renal failure by 18 weeks of age. In contrast, the treatment with MR16-1 prevented all these symptoms and prolonged the lifetime of the majority of the mice. Thus, the constitutive overexpression of IL-6 caused various disorders, and the treatment with anti-IL-6R mAb completely prevented from these symptoms. These results clearly confirm that IL-6 indeed plays an essential role in the pathogenesis of a variety of disorders. Furthermore, anti-IL-6R mAb could provide novel therapy for Castleman's disease and MR16-1 should be a useful tool to estimate therapeutic potential of IL-6 antagonists in a variety of murine models for human disease.     

TNF-alpha and IL-1 sequentially induce endothelial ICAM-1 and VCAM-1 expression in MRL/lpr lupus-prone mice.

Dysfunctional leukocyte-endothelial interactions are thought to play a key role in systemic lupus erythematosus pathogenesis. We questioned the importance of TNF-alpha and IL-1 for endothelial activation in MRL/lpr lupus-prone mice. Endothelial ICAM-1 and VCAM-1 expression increased significantly with disease evolution in kidney, heart, and brain, as shown by i.v. injected radiolabeled Ab uptake. Lung endothelial VCAM-1 also increased, while lung endothelial ICAM-1 did not rise above a high basal level. Immunoassays showed a significantly raised circulating level of TNF-alpha by 14 wk, with levels of circulating IL-1alpha and IL-1beta being additionally raised by 20 wk. With 14-wk-old MRL/lpr, anti-TNF-alpha antiserum inhibited expression of ICAM-1 and VCAM-1 by endothelial cells cultured with sera in vitro, and uptake of anti-ICAM-1 and anti-VCAM-1 mAb in lung, kidney, brain, and heart in vivo. In contrast, both anti-TNF-alpha and anti-IL-1 antisera were required for maximal inhibition in vitro and in vivo at 20 wk. These data indicate that TNF-alpha is largely responsible for the early up-regulation of endothelial ICAM-1 and VCAM-1, but that IL-1 enhances expression in late disease. Our observations provide novel insights of possible relevance to understanding endothelial activation in systemic lupus erythematosus, and highlight an approach that can be extended to dissecting other chronic inflammatory diseases.     

IL-12 contributes to allergen-induced airway inflammation in experimental asthma

Lack of sufficient IL-12 production has been suggested to be one of the basic underlying mechanisms in atopy, but a potential role of IL-12 in established allergic airway disease remains unclear. We took advantage of a mouse model of experimental asthma to study the role of IL-12 during the development of bronchial inflammation. Administration of anti-IL-12p35 or anti-IL-12p40 mAb to previously OVA-sensitized BALB/c mice concomitantly with exposure to nebulized OVA, abolished both the development of bronchial hyperresponsiveness to metacholine as well as the eosinophilia in bronchoalveolar lavage fluid and peripheral blood. Anti-IL-12 treatment reduced CD4(+) T cell numbers and IL-4, IL-5, and IL-13 levels in the bronchoalveolar lavage fluid and the mRNA expression of IL-10, eotaxin, RANTES, MCP-1, and VCAM-1 in the lung. Anti-IL-12p35 treatment failed to show these effects in IFN-gamma knockout mice pointing to the essential role of IFN-gamma in IL-12-induced effects. Neutralization of IL-12 during the sensitization process aggravated the subsequent development of allergic airway inflammation. These data together with recent information on the role of dendritic cells in both the sensitization and effector phase of allergic respiratory diseases demonstrate a dual role of IL-12. Whereas IL-12 counteracts Th2 sensitization, it contributes to full-blown allergic airway disease upon airway allergen exposure in the postsensitization phase, with enhanced recruitment of CD4(+) T cells and eosinophils and with up-regulation of Th2 cytokines, chemokines, and VCAM-1. IFN-gamma-producing cells or cells dependent on IFN-gamma activity, play a major role in this unexpected proinflammatory effect of IL-12 in allergic airway disease.     

In vitro analysis of interferon gamma (IFN-gamma) and interleukin-12 (IL-12) production and their effects in ileal Crohn's disease

Crohn's disease is an inflammatory disease of the gut in which tumour necrosis factor (TNF) and T helper 1 (Th1) cytokines (interleukin (IL)-12, interferon (IFN)-gamma) are thought to play a major role. After the successes obtained with neutralisation of TNF, interest is now growing for therapy aiming at neutralisation of Th1-associated cytokines. Since cytokines are linked in a delicate network, in vitro cultures of ileal lamina propria mononuclear cells (LPMC) were set up for evaluation of a) IFN-gamma and IL-12 production, b) effects of rhIFN-gamma and rhIL-12 and c) effects of anti-IFN-gamma and anti-IL-12 on pro-inflammatory cytokines and IL-10 production. LPMC were isolated from surgical specimens of a total of 27 Crohn's disease and 17 caecum carcinoma (control) patients. Cells were stimulated with CD40L (which triggers myeloid CD40-expressing cells) or anti-CD3 +CD80 (which triggers T cells). LPMC from involved ileal, Crohn's disease produced, in both non-stimulated and stimulated conditions, more IFN-gamma and IL-12p70 than LPMC from non-involved tissue or from control patients. rhIFN-gamma significantly enhanced TNF production in both controls and in ileal Crohn's disease patients, while rhIL-12 enhanced IFN-gamma but not TNF production. LPMC from involved tissue were more sensitive to IL-12 than control LPMC. LP-T cell-dependent activation of monocytes was then studied by co-culture of anti-CD3/CD80-stimulated LPMC with fresh monocytes, which resulted in high IL-12, IFN-gamma, TNF and IL-10 production. The data show that neutralisation of either IL-12 or IFN-gamma with mAb in these cultures also affects secretion of the reciprocal cytokine and (in the case of anti-IL-12) also that of the anti-inflammatory cytokine IL-10. However, no effect of anti-IL-12 or anti-IFN-gamma on production of TNF, a cytokine with an important pathogenic role in Crohn's disease, could be found. Therapies aiming at neutralisation of IFN-gamma or IL-12 are therefore unlikely to replace anti-TNF, but they might provide an additive or synergistic effect.     

Treatment of T cell-dependent experimental colitis in SCID mice by local administration of an adenovirus expressing IL-18 antisense mRNA.

Recent studies have shown that IL-18, a pleiotropic cytokine that augments IFN-gamma production, is produced by intestinal epithelial cells and lamina propria cells from patients with Crohn's disease. In this study, we show that IL-18 is strongly expressed by intestinal epithelial cells in a murine model of Crohn's disease induced by transfer of CD62L+ CD4+ T cells into SCID mice. To specifically down-regulate IL-18 expression in this model, we constructed an E1/E3-deleted adenovirus expressing IL-18 antisense mRNA, denoted Ad-asIL-18, and demonstrated the capacity of such a vector to down-regulate IL-18 expression in colon-derived DLD-1 cells and RAW264.7 macrophages. Local administration of the Ad-asIL-18 vector to SCID mice with established colitis led to transduction of epithelial cells and caused a significant suppression of colitis activity, as assessed by a newly developed endoscopic analysis system for colitis. Furthermore, treatment with Ad-asIL-18 induced a significant suppression of histologic colitis activity and caused suppression of mucosal IFN-gamma production, whereas IFN-gamma production by spleen T cells was unaffected. Taken together, these data indicate an important role for IL-18 in the effector phase of a T cell-dependent murine model of colitis and suggest that strategies targeting IL-18 expression may be used for the treatment of patients with Crohn's disease.     

Physiological stress exacerbates murine colitis by enhancing proinflammatory cytokine expression that is dependent on IL-18

Psychological stress is an environmental factor considered to be a precipitating factor of inflammatory bowel disease. Interleukin (IL)-18 plays a role in stress-induced aggravation in some diseases. The aim of this study was to establish a model of murine colitis exacerbated by psychological stress and to clarify the role of IL-18 in this model. Male C57Bl/6 mice and IL-18(-/-) mice were used for this study. The mice received dextran sulfate sodium (DSS) for induction of colitis. Some mice were exposed to psychological stress using a communication box. Body weight, colonic length, and histological inflammation were measured for assessment of colitis. Tumor necrosis factor (TNF)-α and IL-18 expression in the colon and IL-18 expression in the adrenal gland were analyzed using real-time PCR. The effect of anti-IL-18 antibody was also investigated. Effects of TNF-α and IL-18 on cytokine expressions were studied using the colonic epithelial cell line LS174T. Induction of psychological stress in DSS-treated wild-type mice significantly exacerbated colitis with enhanced expression of proinflammatory cytokines and IL-18. However, induction of psychological stress in DSS-treated IL-18(-/-) mice did not aggravate colitis compared with that in the IL-18(-/-) group given only DSS treatment. Stress-induced aggravation of colitis was ameliorated significantly by anti-IL-18 antibody treatment. IL-18 did not enhance TNF-α-induced expression of intercellular adhesion molecule-1 or IL-8 in LS174T. We established a model of colitis exacerbated by psychological stress. Psychological stress enhanced IL-18 expression and plays a proinflammatory role in stress-induced aggravation of colitis.     

Probiotics ameliorate recurrent Th1-mediated murine colitis by inducing IL-10 and IL-10-dependent TGF-beta-bearing regulatory cells

Recent studies of murine models of mucosal inflammation suggest that, whereas some kinds of bacterial microflora are inducers of disease, others, known as probiotics, prevent disease. In the present study, we analyzed the regulatory cytokine and cell response to probiotic (VSL#3) administration in the context of the Th1 T cell colitis induced by trinitrobenzene sulfonic acid treatment of SJL/J mice. Daily administration of probiotics for 3 wk to mice during a remission period between a first and second course of colitis induced by trinitrobenzene sulfonic acid, resulted in a milder form of recurrent colitis than observed in mice administered PBS during this same period. This protective effect was attributable to effects on the lamina propria mononuclear cell (LPMC) population, because it could be transferred by LPMC from probiotic-treated mice to naive mice. Probiotic administration was associated with an early increase in the production of IL-10 and an increased number of regulatory CD4+ T cells bearing surface TGF-beta in the form of latency-associated protein (LAP) (LAP+ T cells). The latter were dependent on the IL-10 production because administration of anti-IL-10R mAb blocked their appearance. Finally, the LAP+ T cells were essential to the protective effect of probiotics because administration of anti-IL-10R or anti-TGF-beta at the initiation of recurrent colitis induction or depletion of LAP+ T cells from LPMC abolished the latter's capacity to transfer protection to naive recipients. These studies show that probiotic (VSL#3) administration during a remission period ameliorates the severity of recurrent colitis by inducing an immunoregulatory response involving TGF-beta-bearing regulatory cells.     

Cultured human follicular dendritic cells. Growth characteristics and interactions with B lymphocytes.

Minced human tonsils were digested with DNase and collagenase, and lymphoid cell-depleted low density cells were cultured and grown in granulocyte-macrophage-CSF. Large, morphologically homogenous adherent cells with elongated extensions grew continuously in culture. These nonphagocytic cells appear to be related to follicular dendritic cell (FDC) as they do not have properties of monocytic lineage cells or dendritic cells and because, like FDC, 1) they express CD11b, CD14, CD29, CD40, CD54, CD73, CD74, and VCAM-1, and do not express CD11c, CD22, T cell markers, CD18, CD25 and CD45; and 2) they bind human B lymphocytes and B cell lines, but not T lymphocytes by an adhesion blocked in part by mAb to VLA-4 (CD49d). The cultured FDC also augmented B cell proliferation stimulated by anti-mu sera and/or CD40 mAb. Cultured FDC spontaneously produced low levels of IL-6, but did not produce IL-1 alpha or TNF-alpha; however, after treatment with either IFN-gamma or LPS, they produced more IL-6. The expression of CD54 (ICAM-1) was elevated by treating the cultured FDC with either TNF-alpha, IL-1 beta, IFN-gamma or granulocyte-macrophage-CSF; in contrast, IL-4 had no effect on CD54 but rather up-regulated expression of VCAM-1. IFN-gamma, unlike the other cytokines tested, increased expression of a set of markers on cultured FDC (CD54, VCAM-1, and CD14) and converted these class II-negative cells into class II+ cells. The fact that various T cell-derived cytokines have different effects on FDC suggests that the T cell products may influence the manner by which FDC stimulate B cell proliferation and maturation.     

A novel monoclonal antibody against murine IL-2 receptor beta-chain. Characterization of receptor expression in normal lymphoid cells and EL-4 cells

A mAb specific for the murine IL-2R beta-chain (IL-2R beta) was produced by immunizing a rat with a rat transfectant cell line expressing a large number of cDNA-encoded murine IL-2R beta. The mAb, designated TM-beta 1, is specifically reactive with the murine IL-2R beta cDNA-transfectant but not with the recipient cell, and immunoprecipitates murine IL-2R beta of Mr 75 to 85 kDa. TM-beta 1 mAb completely abolished the high affinity IL-2 binding by inhibiting the ligand binding to IL-2R beta. Murine IL-2R beta was found to be constitutively expressed on a subpopulation of CD8+ T cells and almost all NK1.1+ NK cells in the spleen, whereas TM-beta 1 mAb inhibited the proliferation of spleen cells induced by 1 nM of IL-2. Interestingly, EL-4 cells that express murine IL-2R beta as detected by TM-beta 1 mAb can bind neither human nor murine IL-2 under the intermediate affinity conditions, although cDNA-directed human IL-2R beta expressed in the same EL-4 cells has been previously shown to manifest the intermediate affinity IL-2 binding. These results may imply that functional expression of IL-2R beta is differentially regulated between humans and mice. Finally, our neutralizing anti-IL-2R beta mAb TM-beta 1 will be useful not only for various in vitro studies but also for in vivo studies to directly investigate the possible involvement of the IL-2/IL-2R pathway in the generation and differentiation of T lymphocytes and NK cells.     

Differential utilization of ICAM-1 and VCAM-1 during the adhesion and transendothelial migration of human T lymphocytes.

The comparative roles of the endothelial cell (EC) adhesion receptors VCAM-1 and ICAM-1 during the adhesion and transendothelial migration of T cells were examined. The adhesion of T cells to IL-1-activated EC was markedly, but not completely, inhibited by mAb to VCAM-1 as well as to its counter-receptor, VLA-4, whereas, T cell binding to IL-1-activated EC was not blocked by mAb to ICAM-1 or to its counter-receptor, LFA-1. In contrast, LFA-1/ICAM-1, but not VLA-4/VCAM-1, mediated much, but not all, of the binding of T cells to unstimulated EC. Activation of T cells with phorbol dibutyrate and ionomycin alter the receptor-counter-receptor pairs used for binding to EC. Regardless of the activation status of the EC, the binding of activated T cells was not blocked by mAb to VLA-4 or VCAM-1. Moreover, the binding of activated T cells to EC was blocked to a lesser degree by mAb to LFA-1 than that of resting T cells, and mAb to ICAM-1 blocked binding only modestly. The role of VCAM-1 and ICAM-1 during the transendothelial migration of T cells was also examined. Regardless of the activation status of the T cells or the EC, VCAM-1 was never found to function during transendothelial migration, even when it mediated the binding of resting T cells to IL-1-activated EC. In contrast, ICAM-1 played an important role in transendothelial migration under all of the conditions examined, including situations when T cell-EC binding was not mediated by ICAM-1. Immunoelectron microscopic analysis of transendothelial migration supported the conclusion that ICAM-1 but not VCAM-1 played a central role in this process. Thus, ICAM-1 was prominently and uniformly expressed at all EC membrane sites that were in contact with bound and migrating T cells, whereas VCAM-1 was localized to the luminal surface of IL-1-activated EC, but was often absent from the surface of the EC in contact with T cells undergoing transendothelial migration. These studies confirm that ICAM-1 and VCAM-1 play reciprocal roles in the binding of resting T cells to resting and IL-1-activated EC, respectively, but a less prominent role in the binding of activated T cells. Moreover, ICAM-1 but not VCAM-1 plays a role in transendothelial migration, regardless of the receptor-counter-receptor pairs used for initial binding.     

Cytokine-induced alterations of α7 nicotinic receptor in colonic CD4 T cells mediate dichotomous response to nicotine in murine models of Th1/Th17- versus Th2-mediated colitis

Ulcerative colitis (UC) and Crohn's disease (CD) are two forms of chronic inflammatory bowel disease. CD4 T cells play a central role in the pathogenesis of both diseases. Smoking affects both UC and CD but with opposite effects, ameliorating UC and worsening CD. We hypothesized that the severity of gut inflammation could be modulated through T cell nicotinic acetylcholine receptors (nAChRs) and that the exact clinical outcome would depend on the repertoire of nAChRs on CD4 T cells mediating each form of colitis. We measured clinical and immunologic outcomes of treating BALB/c mice with oxazolone- and trinitrobenzene sulfonic acid (TNBS)-induced colitides by nicotine. Nicotine attenuated oxazolone colitis, which was associated with an increased percentage of colonic regulatory T cells and a reduction of Th17 cells. TCR stimulation of naive CD4(+)CD62L(+) T cells in the presence of nicotine upregulated expression of Foxp3. In marked contrast, nicotine worsened TNBS colitis, and this was associated with increased Th17 cells among colonic CD4 T cells. Nicotine upregulated IL-10 and inhibited IL-17 production, which could be abolished by exogenous IL-12 that also abolished the nicotine-dependent upregulation of regulatory T cells. The dichotomous action of nicotine resulted from the up- and downregulation of anti-inflammatory α7 nAChR on colonic CD4 T cells induced by cytokines characteristic of the inflammatory milieu in oxazolone (IL-4) and TNBS (IL-12) colitis, respectively. These findings help explain the dichotomous effect of smoking in patients with UC and CD, and they underscore the potential for nicotinergic drugs in regulating colonic inflammation.     

Intercellular adhesion molecule 1 deficiency leads to impaired recruitment of T lymphocytes and enhanced host susceptibility to infection with Trypanosoma cruzi

In this study, we investigated the involvement of Th1 cytokines in the expression of cell adhesion molecules (CAM) and recruitment of inflammatory cells to the heart of mice infected with Trypanosoma cruzi. Our results show that endogenously produced IFN-gamma is essential to induce optimal expression of VCAM-1 and ICAM-1 on the cardiac vascular endothelium of infected mice. Furthermore, the influx of inflammatory cells into the cardiac tissue was impaired in Th1 cytokine-deficient infected mice, paralleling the intensity of VCAM-1 and ICAM-1 expression on the vascular endothelium. Consistent with the importance of ICAM-1 in host resistance, ICAM-1 knockout (KO) mice were highly susceptible to T. cruzi infection, as assessed by mortality rate, parasitemia, and heart tissue parasitism. The enhanced parasitism was associated with a decrease in the numbers of CD4(+) and CD8(+) T lymphocytes in the heart tissue of ICAM-1 KO mice. Additionally, ICAM-1 KO mice mounted an unimpaired IFN-gamma response and IFN-gamma-dependent production of reactive nitrogen intermediates and parasite- specific IgG2a. Supporting the participation of ICAM-1 in cell migration during T. cruzi infection, the entrance of adoptively transferred PBL from T. cruzi-infected wild-type C57BL/6 mice into the cardiac tissue of ICAM-1 KO mice was significantly abrogated. Therefore, we favor the hypothesis that ICAM-1 plays a crucial role in T lymphocyte recruitment to the cardiac tissue and host susceptibility during T. cruzi infection.     

Cytokine and endothelial cell adhesion molecule expression in interleukin-10-deficient mice

The objectives of this study were to quantify cytokine mRNA levels and endothelial cell adhesion molecule message and protein expression in healthy wild-type and interleukin-10-deficient (IL-10(-/-)) mice that develop spontaneous and chronic colitis. We found that colonic message levels of IL-1, IL-6, tumor necrosis factor-alpha, interferon-gamma, lymphotoxin-beta, and transforming growth factor-beta were elevated in colitic mice 10- to 35-fold compared with their healthy wild-type controls. In addition, colonic message levels of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and mucosal addressin cell adhesion molecule-1 (MAdCAM-1) were found to be increased 10-, 5-, and 23-fold, respectively, in colitic IL-10(-/-) mice compared with their wild-type controls. Immunoradiolabeling as well as immunohistochemistry revealed large and significant increases in vascular surface expression of colonic ICAM-1, VCAM-1, and MAdCAM-1 in the mucosa as well as the submucosa of the colons of colitic mice. These data are consistent with the hypothesis that deletion of IL-10 results in the sustained production of proinflammatory cytokines, leading to the upregulation of adhesion molecules and infiltration of mononuclear and polymorphonuclear leukocytes into the cecal and colonic interstitium.     

Commensal bacteria exacerbate intestinal inflammation but are not essential for the development of murine ileitis

The pathogenesis of Crohn's disease has been associated with a dysregulated response of the mucosal immune system against intraluminal Ags of bacterial origin. In this study, we have investigated the effects of germfree (GF) conditions in the SAMP1/YitFc murine model of Crohn's disease-like ileitis. We show that the bacterial flora is not essential for ileitis induction, because GF SAMP1/YitFc mice develop chronic ileitis. However, compared with disease in specific pathogen-free (SPF) mice, ileitis in GF mice is significantly attenuated, and is associated with delayed lymphocytic infiltration and defective mucosal expression of Th2 cytokines. In addition, we demonstrate that stimulation with purified fecal Ags from SPF, but not GF mice leads to the generation of IL-4-secreting effector lymphocytes. This result suggests that commensal bacteria drive Th2 responses characteristic of the chronic phase of SAMP1/YitFc ileitis. Finally, adoptive transfer of CD4-positive cells from GF, but not SPF mice induces severe colitis in SCID recipients. These effects were associated with a decreased frequency of CD4(+)CD25(+)Foxp3(+) T cells in the mesenteric lymph nodes of GF mice compared with SPF mice, as well as lower relative gene expression of Foxp3 in CD4(+)CD25(+) T cells in GF mice. It is therefore apparent that, in the absence of live intraluminal bacteria, the regulatory component of the mucosal immune system is compromised. All together, our results indicate that in SAMP1/YitFc mice, bacterial flora exacerbates intestinal inflammation, but is not essential for the generation of the chronic ileitis that is characteristic of these mice.     

Differential costimulatory effects of adhesion molecules B7, ICAM-1, LFA-3, and VCAM-1 on resting and antigen-primed CD4+ T lymphocytes.

Optimal proliferation of T cells although initiated via ligation of the CD3/TCR complex requires additional stimulation resulting from adhesive interactions between costimulatory receptors (R) on T cells and their counter-R on APC. At least four distinct adhesion molecules (counter-R) present on APC, B7, ICAM-1 (CD54), LFA-3 (CD58), and VCAM-1 have been individually shown to costimulate T cell activation. Because some of these molecules may be expressed simultaneously on APC, it has been difficult to examine relative contributions of individual counter-R during the induction of T cell proliferation. We have produced soluble IgC gamma 1 fusion chimeras (receptor globulins or Rg) of B7, ICAM-1, LFA-3, and VCAM-1 and compared their relative abilities to costimulate proliferation of resting or Ag-primed CD4+ T cells. When co-immobilized with mAb directed at TCR alpha beta or CD3 but not CD2 or CD28, each Rg induced proliferation of both resting and Ag-primed CD4+ cells. In contrast, similarly co-immobilized CD7 Rg or ELAM-1 Rg were ineffective. Resting CD4+ T cells produced more IL-2, expressed significantly higher levels of IL-2R alpha, and proliferated more efficiently when costimulated with either ICAM-1 Rg or VCAM-1 Rg than with B7 Rg or LFA-3 Rg. CD4+ CD45RO+ memory T cells proliferated more vigorously in response to the costimulation by each of the four Rg than CD4+ CD45RA+ naive T cells. In contrast with the behavior of resting CD4+ T cells, proliferation of Ag-preactivated CD4+ T cells was most efficient when costimulated by B7 Rg. The costimulatory effect of LFA-3 Rg on Ag-primed CD4+ T cells was weaker than that of B7 Rg but was significantly greater than that of either ICAM-1 Rg or VCAM-1 Rg. These results suggest that resting and Ag-primed CD4+ T cells preferentially respond by proliferation to different costimulatory counter-R. ICAM-1 and VCAM-1 may be involved in the initiation of proliferation of Ag-responsive T cells, and B7 and LFA-3 may facilitate sustained proliferation of Ag-primed T cells. The cumulative costimulation by the above counter-R may facilitate optimal expression of various regulatory and effector functions of T cells.     

IL-2 in vivo activities and antitumor efficacy enhanced by an anti-IL-2 mAb

IL-2 is a potent immunostimulant and has been tested for clinical use, including in immunotherapy for cancers and HIV infection. Here we show that a widely used neutralizing anti-murine IL-2 mAb (S4B6) exhibits unexpected activities that enhance the treatment effects of IL-2 in vivo. Coinjection of the anti-IL-2 mAb with a plasmid carrying murine IL-2 cDNA significantly increased the serum IL-2 levels and induced a substantial increase in the division of CD8+ T and NK1.1(high) cells in vivo. Injection of the mAb premixed with recombinant murine IL-2 showed the same enhanced effect. A 5-day treatment with the anti-IL-2 mAb alone gradually increased the CD44(high)CD8+ population, and the increased population was maintained for >300 days, suggesting that the mAb can gradually maintain and potentially enhance the bioactivity of endogenous IL-2 for extended periods. Furthermore, combined treatment with the anti-IL-2 mAb plus the IL-2 plasmid markedly enhanced Ag-specific CTL activity in vivo and partially protected mice from tumor metastasis to the lungs, compared with the anti-IL-2 mAb or IL-2 plasmid alone. These results demonstrated IL-2-enhancing effects of the anti-IL-2 mAb in vivo and suggest that combining a neutralizing anti-IL-2 Ab with IL-2 gene delivery might be used effectively to enhance IL-2 functions in clinical applications.     

白细胞介素23受体、白细胞介素17A在炎症性肠病患者中的表达 及临床意义

目的:通过检测白细胞介素23受体(IL-23R)及白细胞介素17A(IL-17A)在炎症性肠病(IBD)患者肠黏膜及血清中的表达水平,探讨其在IBD发病过程中的作用及意义。方法:收集32例克罗恩病(CD)患者、29例溃疡性结肠炎(UC)患者及27例对照者的内镜肠黏膜活检标本,采用荧光定量PCR技术检测肠黏膜内IL-23R、IL-17AmRNA的表达情况,免疫组化技术分析IL-23R、IL-17A在肠黏膜中的原位表达。结果:与健康对照组相比,CD及UC患者肠黏膜组织内IL-23R mRNA表达显著增高(P<0.05),CD及UC组间的表达量差异无统计学意义(P>0.05)。CD及UC患者肠黏膜组织内IL-17A mRNA表达显著增高(P<0.05),CD组肠黏膜组织内IL-17AmRNA表达显著高于UC组(P<0.05)。免疫组化分析显示IL-23R阳性细胞在CD与UC肠黏膜固有层内有较多表达,较正常黏膜内的肠上皮细胞相比,CD及UC患者肠黏膜IL-23R蛋白表达量最著增高(P<0.05),UC及CD组间的表达量差异无统计学意义(P>0.05)。IL-17A阳性细胞在CD与UC肠黏膜固有层内有较多表达,较正常黏膜内的肠上皮细胞相比,CD及UC患者肠黏膜IL-17A蛋白表达量最著增高(P<0.05)。结论:IL-23R及IL-17A在IBD患者肠黏膜中表达显著增高,提示IL-23R及IL-17A表达异常与IBD的发生发展密切相关,有可能成为IBD治疗的新靶点。     

IL-4 exacerbates disease in a Th1 cell transfer model of colitis

IL-4 is associated with Th2-type immune responses and can either inhibit or, in some cases, promote Th1-type responses. We tested the effect of IL-4 treatment on the development of inflammation in the CD4(+)CD45RB(high) T cell transfer model of colitis, which has been characterized as a Th1-dependent disease. IL-4 treatment significantly accelerated the development of colitis in immunodeficient recipients (recombinase-activating gene-2 (Rag2)(-/-)) of CD4(+)CD45RB(high) T cells. Quantitative analysis of mRNA expression in the colons of IL-4-treated mice showed an up-regulation of both Th1- and Th2-associated molecules, including IFN-gamma, IP-10, MIG, CXCR3, chemokine receptor-8, and IL-4. However, cotreatment with either IL-10 or anti-IL-12 mAb effectively blocked the development of colitis in the presence of exogenous IL-4. These data indicate that IL-4 treatment exacerbates a Th1-mediated disease rather than induces Th2-mediated inflammation. As other cell types besides T cells express the receptor for IL-4, the proinflammatory effects of IL-4 on host cells in Rag2(-/-) recipients were assessed. IL-4 treatment was able to moderately exacerbate colitis in Rag2(-/-) mice that were reconstituted with IL-4Ralpha-deficient (IL-4Ralpha(-/-)) CD4(+)CD45RB(high) T cells, suggesting that the IL-4 has proinflammatory effects on both non-T and T cells in this model. IL-4 did not cause colitis in Rag2(-/-) mice in the absence of T cells, but did induce an increase in MHC class II expression in the lamina propria of the colon, which was blocked by cotreatment with IL-10. Together these results indicate that IL-4 can indirectly promote Th1-type inflammation in the CD4(+)CD45RB(high) T cell transfer model of colitis.     

Targeting IL-12/IL-23 by employing a p40 peptide-based vaccine ameliorates TNBS-induced acute and chronic murine colitis

Interleukin (IL)-12 and IL-23 both share the p40 subunit and are key cytokines in the pathogenesis of Crohn's disease. Previously, we have developed and identified three mouse p40 peptide-based and virus-like particle vaccines. Here, we evaluated the effects and immune mechanisms of the optimal vaccine in downregulating intestinal inflammation in murine acute and chronic colitis, induced by intrarectal administrations of trinitrobenzene sulfonic acid (TNBS). Mice were injected subcutaneously with vaccine, vaccine carrier or saline three times, and then intrarectally administered TNBS weekly for 2 wks (acute colitis) or 7 wks (chronic colitis). The severity of colitis was evaluated by body weight, histology and collagen and cytokine levels in colon tissue. Th1 and Th17 cells in mesenteric lymph nodes (MLN) were determined. Our results showed the vaccine induced high level and long-lasting specific IgG antibodies to p40, IL-12 and IL-23. After administrations of TNBS, vaccinated mice had significantly less body weight loss and a significant decrease of inflammatory scores, collagen deposition and expression of p40, IL-12, IL-23, IL-17, TNF, iNOS and Bcl-2 in colon tissues, compared with carrier and saline groups. Moreover, vaccinated mice exhibited a trend to lower percentages of Th1 cells in acute colitis and of Th17 cells in chronic colitis in MLN than in controls. In summary, administration of the vaccine induced specific antibodies to IL-12 and IL-23, which was associated with improvement of intestinal inflammation and fibrosis. This suggests that the vaccine may provide a potential approach for the long-term treatment of Crohn's disease.