IL-1 receptor accessory protein-Ig/IL-1 receptor type II-Ig heterodimer inhibits IL-1 response more strongly than other IL-1 blocking biopharmaceutical agents.

INTRODUCTION: Interleukin (IL)-1 is a key orchestrator of inflammation and IL-1 inhibitors are expected to be promising pharmaceutical agents for such pathologies. IL-1 is bound to the complex of two receptor components with much higher affinity than with either receptor component alone. MATERIALS AND METHODS: We examined the effect of a heterodimer of IL-1 receptor accessory protein (Acp)-immunoglobulin (Ig) and IL-1R type II (IL1R2)-Ig named AcP-Ig/IL1R2-Ig heterodimer, and compared its effects with other IL-1 inhibitors reported previously. RESULTS AND DISCUSSION: Our results demonstrated that the rat AcP-Ig/IL1R2-Ig heterodimer (IC50=1.95 pM) inhibited IL-1 response to a greater extent than IL1RA (IC50=1,935 pM), Acp-IL1R type I (IL1R1)-Ig homodimer (IC50=73.7 pM) and Acp-IL1R2-Ig homodimer (IC50=72.8 pM). Moreover, human AcP-Ig/IL1R2-Ig heterodimer (IC50=0.14 pM) inhibited it to a greater extent than Acp-IL1R1-Ig homodimer (IC50=4.48 pM) and strongly inhibited responses of both IL-1α and IL-1ß. CONCLUSIONS: The AcP-Ig/IL1R2-Ig heterodimer, which is similar to the original extracellular structure of the Acp/IL1R1 complex, may inhibit the IL-1 response more vigorously than other IL-1 blocking biopharmaceutical agents.

Expression of the peptide hormone hepcidin increases in cardiomyocytes under myocarditis and myocardial infarction.

The micronutrient iron is an essential component that plays a role in many crucial metabolic reactions. The peptide hormone hepcidin is thought to play a central role in iron homeostasis and its expression is induced by iron overloading and inflammation. Recently, hepcidin has been reported to be expressed also in the heart; however, the kinetics of altered hepcidin expression in diseases of the heart remain unknown. In this study, we examined cardiac expression of hepcidin in rat experimental autoimmune myocarditis (EAM), human myocarditis and rat acute myocardial infarction (AMI). In rat EAM and AMI hearts, hepcidin was expressed in cardiomyocytes; ferroportin, which is a cellular iron exporter bound by hepcidin, was also expressed in various cells. Analysis of the time course of the hepcidin to cytochrome oxidase subunit 6a (Cox6a)2 expression ratio showed that it abruptly increased more than 100-fold in hearts in the very early phase of EAM and in infarcted areas 1 day after MI. The hepcidin/Cox6a2 expression ratio correlated significantly with that of interleukin-6/gamma-actin in both EAM and AMI hearts (r=0.781, P<.0001 and r=0.563, P=.0003). In human hearts with histological myocarditis, the ratio was significantly higher than in those without myocarditis (0.0400+/-0.0195 versus 0.0032+/-0.0017, P=.0045). Hepcidin is strongly induced in cardiomyocytes under myocarditis and MI, conditions in which inflammatory cytokine levels increase and may play an important role in iron homeostasis and free radical generation.

Gene expression profiles of cardiomyocytes in rat autoimmune myocarditis by DNA microarray and increase of regenerating gene family.

Cardiomyocytes with myocarditis compared with the normal state are thought to change the expressions of various genes greatly, some of which may be new biomarkers or new biologic medicinal products. However, until now, little comprehensive analysis has been made of gene-expression changes in cardiomyocytes with myocarditis. In this study, we performed a DNA microarray analysis by using cardiomyocytes from rat experimental autoimmune myocarditis (EAM). On day 0, rats were immunized with porcine cardiac myosin and cardiomyocytes were isolated and purified from EAM hearts and normal hearts by a method that is hardly thought to change gene expressions in cardiomyocytes. RNA from normal cardiomyocytes and cardiomyocytes of EAM on day 18 was analyzed for 7711 gene expressions by DNA microarray. Some gene expressions showed over 10-fold changes. In particular, the regenerated gene (Reg)2/pancreatitis-associated protein (PAP)1 messenger RNA (mRNA) level most markedly increased in the genes, which were clearly expressed in cardiomyocytes rather than in noncardiomyocytes, and it was approximately 2000-fold greater in cardiomyocytes under active myocarditis than normal by real-time reverse transcription polymerase chain reaction analysis. Moreover, we demonstrated that Reg2/PAP1 proteins determined by Western blot analysis and immunohistochemistry and other Reg/PAP family gene expressions were remarkably increased in EAM hearts; in addition, interleukin (IL)-6 expression was significantly related to Reg2/PAP1. It seemed that these data were useful as a reference database of gene-expression changes in cardiomyocytes with myocarditis. The Reg/PAP family, which was found to show dramatically increasing gene expressions by DNA microarray analysis, was suspected to play an important role in myocarditis.

Imatinib mesilate inhibits neointimal hyperplasia via growth inhibition of vascular smooth muscle cells in a rat model of balloon injury.

Restenosis is a major problem in percutaneous catheter intervention (PCI) for coronary artery stenosis in patients with acute myocardial infarction. Coronary restenosis arises from intimal hyperplasia, i.e., hyperplasia of the vascular smooth muscle cells (SMCs) caused by endothelial cell (EC) damage due to PCI. Drug eluting stent (DES), a novel stent coated with a cell-growth inhibitor, such as rapamycin, has been utilized to block SMC proliferation, but DES also blocks EC repair and thus requires the administration of anti-platelets for a long time to prevent thrombus formation after PCI. Moreover, insufficient prevention of platelet aggregation sometimes induces restenosis after PCI. One of the signal transduction inhibitors, imatinib mesilate, blocks tyrosine kinase activity of platelet-derived growth factor receptor (PDGFR), and therefore it may block the development of neointima through growth inhibition of SMCs without the obstructive effect on EC-repair. We therefore studied the effects of imatinib on neointimal hyperplasia in a balloon injury model of rat carotid arteries. Rats were orally administered with imatinib for 14 days after balloon injury, and sacrificed to analyze the neointimal formation. Intimal hyperplasia was inhibited by imatinib in a dose-dependent manner. Therefore imatinib presumably obstructed the growth of SMCs via interception on growth-signaling of PDGFR. The administration of imatinib after coronary stenting or the use of an imatinib-eluting stent may further reduce the risk of restenosis in patients.

Alteration of IL-17 related protein expressions in experimental autoimmune myocarditis and inhibition of IL-17 by IL-10-Ig fusion gene transfer.

BACKGROUND: T-helper (Th)1/Th2 cytokine balance plays an important role in the pathogenesis of myocarditis. Recently, some studies indicate that interleukin (IL)-17, known as a T cell (Th17)-derived proinflammatory cytokine, is the major mediator of tissue inflammation in inflammatory and autoimmune diseases. Experimental autoimmune myocarditis (EAM) is a T cell-mediated autoimmune disease; however, the pathogenic role of IL-17 in the development of rat EAM remains largely unknown. METHODS AND RESULTS: In the present study, alterations of IL-17-related protein expressions were investigated and then the effect of hydrodynamic-based delivery of plasmid DNA encoding the IL-10-Ig gene on rat EAM and the effect of IL-10-Ig on IL-17 was evaluated. The results showed that IL-17 was expressed more highly than IFN-gamma expressed by Th1 cells in alphabetaT cells and the peaks of IL-17 related protein expression in the heart were the early phase of EAM. Moreover, we observed that IL-10-Ig gene therapy was effective in controlling EAM and that IL-10-Ig significantly suppressed the expression of IL-17 as well as other proinflammatory cytokines, IL-1beta and TNF-alpha, in IL-1-stimulated splenocytes cultured from EAM rats. CONCLUSIONS: IL-17 is highly produced by alphabetaT cells in the early phase of EAM hearts and IL-17 inhibition might be a possible mechanism of the amelioration of EAM by IL-10-Ig treatment. These data suggest that IL-17 produced by Th17 plays an important role in the pathogenesis of rat EAM.

Serum erythropoietin level as a marker of limb ischemia.

Bone marrow implantation (BMI) has been utilized for the treatment of limb ischemia, however, serum markers have not yet been reported to express the degree of limb ischemia. We analyzed the serum levels of several cytokines including erythropoietin (EPO) in the treated legs and the contralateral ones in 11 patients with limb ischemia treated with BMI. The EPO level in the pre-treated legs in the 5 patients with arteriosclerosis obliterans revealed a good correlation with ankle-brachial pressure index. The EPO level, but not the levels of TNF-alpha, VEGF, and bFGF in the pre-treated legs was significantly higher than that in the contralateral legs in the 11 patients, and the EPO level decreased in 4 weeks after BMI. The serum EPO level may express the degree of limb ischemia presumably through the reactive production of EPO in ischemic tissue.

[Prognostic factors for critical limb ischemia after autologous bone marrow implantation].

OBJECTIVES: Autologous bone marrow implantation (BMI) is effective to treat critical limb ischemia, but the long-term prognosis is not clear. The outcome of BMI treatment for ischemic legs was investigated related to the clinical background of the patient, and short-term effects of BMI. The end event was defined as unexpected lower limb amputation. METHODS AND RESULTS: This study included 21 consecutive patients (mean age 60.0 +/- 13.6 years) with peripheral arterial disease who underwent BMI between December 2001 and March 2005. Twelve patients had arteriosclerosis obliterans (ASO), 5 had Buerger disease (thromboangiitis obliterans), 3 had thromboembolism, and 1 had hypereosinophilic syndrome. The patients with ASO had severe complications such as diabetes and hyperlipidemia. The total number of transplanted CD34-positive cells, ankle-brachial pressure index (ABI), and tissue oxygen pressure (TcO2) were lower in ASO patients than non-ASO patients. Significant risk factors for the event were diagnosis of ASO and low TcO2 (< 30 mmHg) according to the Kaplan-Meier survival curve and log rank test. All 6 patients who required limb amputation had ASO simultaneously with low TcO2 (6 of 9, 67%). In contrast, there was no correlation between the end event and short-term effect of BMI such as improvements in ABI and TcO2. CONCLUSIONS: Treatment with BMI could not save legs in some patients with ASO associated with severe leg ischemia.