Kenichiro Yamashita

Hokkaido University, Sapporo, Hokkaidō, Japan

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Publications (15)117.96 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Ischemia and reperfusion contribute to substantial organ damage in transplantation. Clinically feasible measures for the prevention thereof are scarce. We tested if rinsing rodent hearts with the antioxidant bilirubin ameliorates IRI. Left ventricular end diastolic pressure (LVEDP), left ventricular developed pressure (LVDevP), rate per pressure product (RPP), coronary flow, maximum (+dP/dt) and minimum (-dP/dt) rate of contraction were analyzed in Lewis rat hearts rinsed with bilirubin prior to reperfusion on a Langendorff apparatus after 12h of cold ischemia. In vivo, isogenic C57Bl/6 mouse hearts rinsed with bilirubin were transplanted after 12h of cold ischemia. Cardiac function and apoptosis was assessed 24h after reperfusion. Heart lysates recovered 15min after reperfusion were probed for the total and the phosphorylated forms of ERK, JNK, p38-MAPK and Akt. In isolated perfused hearts, bilirubin rinse resulted in significantly lower LVEDP and improved LVDevP, RPP, coronary flow, +dP/dt and -dP/dt. In vivo, after reperfusion all MAPKs were suppressed significantly by bilirubin pre-treatment. Bilirubin rinse improved cardiac scores (3,4±0,5 vs. 2,0±1,0 in controls, p<0,05) and significantly suppressed apoptosis. Ex vivo administration of bilirubin to heart grafts prior reperfusion ameliorates IRI and provides a simple and effective tool to ameliorate outcome in heart transplantation. This article is protected by copyright. All rights reserved.
    Transplant International 01/2014; · 3.16 Impact Factor
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    ABSTRACT: Bilirubin is the end product of heme catabolism by heme oxygenases. The inducible form of these enzymes is heme oxygenase-1 (HO-1), which is the rate-limiting enzyme that can degrade heme into equimolar quantities of carbon monoxide (CO), biliverdin, and free iron. Biliverdin is very rapidly converted to bilirubin by the enzyme biliverdin reductase, and free iron upregulates the expression of ferritin. HO-1 is a ubiquitous stress protein and is induced in many cell types by various stimuli. Induced HO-1 exerts antiinflammatory effects and modulates apoptosis. Expression of HO-1 in vivo suppresses the inflammatory responses in endotoxic shock, hyperoxia, acute pleurisy, and organ transplantation, as well as ischemia-reperfusion injury, and thereby provides salutary effects in these conditions. Accumulating evidence indicates that biliverdin/bilirubin can mediate the protective effects of HO-1 in many disease models, such as IRI and organ transplantation, via its antiinflammatory, antiapoptotic, antiproliferative, and antioxidant properties, as well as its effects on the immune response. This review attempts to summarize these protective roles as well as the molecular mechanisms by which biliverdin/bilirubin benefit IRI and solid-organ transplantation, including chronic rejection, and islet transplantation.
    Antioxidants and Redox Signaling 01/2008; 9(12):2175-85. · 7.67 Impact Factor
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    ABSTRACT: We have recently shown that the natural bile pigment bilirubin has antiproliferative effects on vascular smooth muscle cells (VSMCs). Bilirubin is the end product of heme catabolism mediated by heme oxygenases and has for decades been considered a toxic waste product of our bodies. However, 14 separate studies and a meta-analysis have documented an inverse correlation between atherosclerosis and the levels of bilirubin in normal individuals. Having high normal or supranormal levels of bilirubin is associated with less atherosclerotic-type disease as compared with that in individuals with low normal levels of bilirubin. This combined with experimental data showing anti-atherosclerotic properties of the enzyme heme oxygenase-1 encouraged us to hypothesize that bilirubin and its precursor biliverdin, would act to ameliorate components of atherosclerosis, in a manner similar to what has been shown with HO-1. Both did so in an animal model of restenosis in which vascular smooth muscle cell proliferation leads to intimal proliferation and causes narrowing of the vessels. We also analyzed the antiproliferative effects of the bile pigments in an in vitro system where bilirubin/biliverdin caused p53 dependent cell cycle arrest by hypophosphorylation of the retinoblastoma tumor suppressor protein in growth factor stimulated VSMCs.
    Cell cycle (Georgetown, Tex.) 02/2007; 6(1):39-43. · 5.24 Impact Factor
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    ABSTRACT: This investigation focused on obtaining a further understanding of the role of heme oxygenase-1 (HO-1) in tolerance induction. Hearts from C57BL/6 (H-2b) mice survived long-term when transplanted into BALB/c (H-2d) recipients treated with the tolerance-inducing regimen of anti-CD40L antibody (MR-1) plus donor-specific transfusion (DST). Grafts did not, however, survive long-term in (HO-1-/-) recipients given the same treatment. Similarly, long-term survival induced by DST was ablated when HO-1 activity was blocked by zinc protoporphyrin IX (ZnPPIX). We further asked whether modulation of HO-1 expression/activity could be used to promote the induction of graft tolerance. DST alone (day 0) failed to promote any prolongation of survival of DBA/2 (H-2d) hearts transplanted into B6AF1 (H-2(b,k/d)) recipients. However, long-term survival and (dominant peripheral) tolerance were readily induced when DST was combined with induction of HO-1 expression by cobalt protoporphyrin IX (CoPPIX). HO-1 induction plus DST led to a significant up-regulation of Foxp3, TGF-beta, IL-10, and CTLA4, which suggests a prominent role for CD4+CD25+ regulatory T cells (Tregs). In fact, the tolerogenic effect of HO-1 plus DST was dependent on CD4+CD25+ Tregs as suggested by adoptively transferring these cells into irradiated recipients under various regimens. Taken together, these findings show that expression of HO-1 in a graft recipient can be essential for long-term graft survival and for induction of tolerance and that modulation of HO-1 expression/activity can be used therapeutically to synergize in the generation of graft tolerance.
    The FASEB Journal 05/2006; 20(6):776-8. · 5.48 Impact Factor
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    ABSTRACT: Bilirubin, a natural product of heme catabolism by heme oxygenases, was considered a toxic waste product until 1987, when its antioxidant potential was recognized. On the basis of observations that oxidative stress is a potent trigger in vascular proliferative responses, that heme oxygenase-1 is antiatherogenic, and that several studies now show that individuals with high-normal or supranormal levels of plasma bilirubin have a lesser incidence of atherosclerosis-related diseases, we hypothesized that bilirubin would have salutary effects on preventing intimal hyperplasia after balloon injury. We found less balloon injury-induced neointima formation in hyperbilirubinemic Gunn rats and in wild-type rats treated with biliverdin, the precursor of bilirubin, than in controls. In vitro, bilirubin and biliverdin inhibited serum-driven smooth muscle cell cycle progression at the G1 phase via inhibition of the mitogen-activated protein kinase signal transduction pathways and inhibition of phosphorylation of the retinoblastoma tumor suppressor protein. Bilirubin and biliverdin might be potential therapeutics in vascular proliferative disorders.
    Circulation 09/2005; 112(7):1030-9. · 14.95 Impact Factor
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    ABSTRACT: Treatment of animals or certain cells with carbon monoxide (CO), a product of heme degradation by heme oxygenase-1 (HO-1), has potent anti-inflammatory and antiapoptotic effects that contribute to the survival of transplanted organs. We report here that inducing HO-1 in, or administering CO to, only the donor can be used in a therapeutic manner to sustain the survival of transplanted allogeneic islets. Similar treatments of only the islets or only the recipient are also salutary. Administering CO only to the donor frequently leads to long-term survival of those islets in untreated allogeneic recipients, which are then antigen-specifically tolerant. Several proinflammatory and proapoptotic genes that are strongly induced in islets after transplantation in the untreated situation were significantly suppressed after administering CO to the donor without further treatment. These included tumor necrosis factor-alpha, inducible nitric oxide synthase, monocyte chemoattractant protein-1, granzyme B, and Fas/Fas ligand, all of which contribute to the pathogenesis of the rejection of transplanted islets. This correlated with a lesser infiltration of recipient macrophages into the transplanted islets. Our present findings show that induction of HO-1 in, or administration of CO to, only the donor, islets, or the recipient or combinations of such treatments improve allogeneic islet survival.
    Diabetes 06/2005; 54(5):1400-6. · 8.47 Impact Factor
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    ABSTRACT: Heme oxygenase-1 (HO-1) seems to have an important protective role in acute and chronic inflammation. The products of heme catalysis, biliverdin/bilirubin, carbon monoxide (CO), and iron (that induces apoferritin) mediate the beneficial effects of HO-1. Blockade of HO-1 activity results in exacerbation of experimental colitis. We tested whether HO-1 has protective effects in the development of colitis and determined that specific enzymatic products of HO-1 are responsible for these effects. Colitis was induced by oral administration of dextran sodium sulfate (5%) to C57BL/6 mice for 7 days. HO-1 was up-regulated by cobalt-protoporphyrin (5 mg/kg, intraperitoneally). Biliverdin, exogenous CO, or the iron chelator desferrioxamine was administered to other groups. Cobalt-protoporphyrin treatment resulted in significant up-regulation of HO-1 protein in mucosal and submucosal cells. Induction of HO-1 was associated with significantly less loss of body weight in mice with induced colitis (-12% versus -22% in the control animals, P < 0.001). Development of diarrhea and gastrointestinal hemorrhage was substantially delayed in animals in which HO-1 was induced, and mucosal injury was significantly attenuated. Administration of CO or desferrioxamine alone had no significant effects, whereas enhanced protection with lesser evidence of bowel inflammation was observed with systemic biliverdin administration (50 micromol/kg, 3 times per day, intraperitoneally). We conclude that heightened HO-1 expression or administration of biliverdin ameliorates dextran sodium sulfate-induced experimental colitis. Novel therapeutic strategies based on HO-1 and/or biliverdin administration may have use in inflammatory bowel disease.
    Inflammatory Bowel Diseases 04/2005; 11(4):350-9. · 5.48 Impact Factor
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    ABSTRACT: Heme oxygenase-1 (HO-1), which degrades heme into three products (carbon monoxide, free iron, and biliverdin), plays a protective role in many models of disease via its anti-inflammatory, anti-apoptotic, and anti-proliferative actions. Overexpression of HO-1 has been shown to suppress immune responses and prolong the survival of allografts; however, the underlying mechanism is not clear. We demonstrate two "new" properties of HO-1 that mediate activation induced cell death (AICD) of allo-antigen-responsive murine CD4+ T cells, resulting in immunomodulation. First, it functions in vivo and in vitro to "boost" the proliferative response of CD4+ T cells to allo-antigens in the early phase of allo-antigen-driven immune responses. This "boosting" effect is accompanied with a significant increase of activation markers and IL-2 production. Second, it exerts a pro-apoptotic effect in those activated T cells after the initial burst of proliferation. We further show that the AICD effect is mediated through the Fas/CD95-FasL signal transduction pathway. Correlating with the above-mentioned findings is the observed prolongation of mouse heart graft survival when HO-1 is expressed in vivo in both donor and recipient. In conclusion, induction of HO-1 expression accelerates clonal deletion of peripheral alloreactive CD4+ T cells by promoting AICD, which is presumably a key mechanism for its immunomodulatory effects such as in prolonging the survival of transplanted organs.
    The FASEB Journal 04/2005; 19(3):458-60. · 5.48 Impact Factor
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    ABSTRACT: Heme oxygenase (HO-1) provides a cellular defense mechanism during oxidative stress and catalyzes the rate-limiting step in heme metabolism that produces biliverdin (BV). The role of BV and its potential use in preventing ischemia/reperfusion injury (IRI) had never been studied. This study was designed to explore putative cytoprotective functions of BV during hepatic IRI in rat liver models of ex vivo perfusion and orthotopic liver transplantation (OLT) after prolonged periods of cold ischemia. In an ex vivo hepatic IRI model, adjunctive BV improved portal venous blood flow, increased bile production, and decreased hepatocellular damage. These findings were correlated with amelioration of histological features of IRI, as assessed by Suzuki's criteria. Following cold ischemia and syngeneic OLT, BV therapy extended animal survival from 50% in untreated controls to 90% to 100%. This effect correlated with improved liver function and preserved hepatic architecture. Additionally, BV adjuvant after OLT decreased endothelial expression of cellular adhesion molecules (P-selectin and intracellular adhesion molecule 1), and decreased the extent of infiltration by neutrophils and inflammatory macrophages. BV also inhibited expression of inducible nitric oxide synthase and proinflammatory cytokines (interleukin 1beta, tumor necrosis factor alpha, and interleukin 6) in OLTs. Finally, BV therapy promoted an increased expression of antiapoptotic molecules independently of HO-1 expression, consistent with BV being an important mediator through which HO-1 prevents cell death. In conclusion, this study documents and dissects potent cytoprotective effects of BV in well-established rat models of hepatic IRI. Our results provide the rationale for a novel therapeutic approach using BV to maximize the function and thus the availability of donor organs.
    Hepatology 01/2005; 40(6):1333-41. · 11.19 Impact Factor
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    ABSTRACT: Induction of the heme-degrading enzyme heme oxygenase-1 (HO-1) has been shown to be beneficial in terms of improvement of liver allograft survival and prevention of CD95-mediated apoptosis in the liver. In the present study, we investigated the effects of HO-1, and its products carbon monoxide (CO), biliverdin (BV), and iron/ferritin, in a mouse model of inflammatory liver damage inducible by lipopolysaccharide (LPS) in mice sensitized with the hepatocyte-specific transcription inhibitor D-galactosamine (GalN). Our results show that HO-1 induction by cobalt-protoporphyrin-IX (CoPP) reduced cytokine expression, protected mice from liver injury, and prolonged survival. While in contrast to ferritin overexpression, single administration of the CO donor methylene chloride (MC) or of BV also protected mice from liver damage, only coadministration of both HO products prolonged survival and reduced the expression of cytokines, e.g., tumor necrosis factor (TNF) and interferon gamma (IFN-gamma). In conclusion, HO-1-induced prolongation of survival, but not the protection from liver damage, seems to be dependent on down-regulation of cytokine synthesis.
    Hepatology 12/2004; 40(5):1128-35. · 11.19 Impact Factor
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    ABSTRACT: Heme oxygenase-1 (HO-1) degrades heme into iron, biliverdin, and carbon monoxide (CO). HO-1 expression can be used therapeutically to ameliorate undesirable consequences of ischemia reperfusion injury (IRI), but the mechanism by which this occurs, remains to be established. Rat hearts, exposed to a prolonged period (24 h) of cold (4 degrees C) ischemia, failed to function upon transplantation into syngeneic recipients. Induction of HO-1 expression by administration of cobalt protoporphyrin IX (CoPPIX) to the graft donor restored graft function. Inhibition of HO-1 enzymatic activity, by administration of zinc protoporphyrin (ZnPPIX) at the time of transplantation, reversed the protective effect of HO-1. Exposure of the graft donor as well as the graft (during ischemia) to exogenous CO mimicked the protective effect of HO-1. This was associated with a significant reduction in the number of cells undergoing apoptosis in the graft with no apparent decrease of intravascular fibrin polymerization, platelet aggregation, or P-selectin expression. In conclusion, HO-1-derived CO prevents IRI associated with cardiac transplantation based on its antiapoptotic action. The observation that exposure of the donor and the graft to CO is sufficient to afford this protective effect should have important clinical implications in terms of preventing IRI associated with heart transplantation in humans.
    The FASEB Journal 05/2004; 18(6):771-2. · 5.48 Impact Factor
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    ABSTRACT: Biliverdin, a product of heme oxygenase-1 (HO-1) enzymatic action, is converted into bilirubin, which has been considered a waste product in the past. We now show that administration of biliverdin has a salutary effect in organ transplantation. A brief course of treatment with biliverdin leads to long-term survival of H-2 incompatible heart allografts. Furthermore, those recipients harboring long-surviving (>100 days) allografts were tolerant to donor antigens indicated by the acceptance of second donor strain hearts but not third-party grafts. Treatment with biliverdin decreased intragraft leukocyte infiltration and inhibited T cell proliferation. Likely related to tolerance induction, biliverdin interferes with T cell signaling by inhibiting activation of nuclear factor of activated T cells (NFAT) and nuclear factor kappaB (NF-kappaB), two transcription factors involved in interleukin-2 (IL-2) transcription and T cell proliferation, as well as suppressing Th1 interferon-gamma (IFN-gamma) production in vitro. These findings support the potential use of biliverdin, a natural product, in transplantation and other T cell mediated immune disorders.
    The FASEB Journal 04/2004; 18(6):765-7. · 5.48 Impact Factor
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    ABSTRACT: Heme oxygenase-1 (HO-1), a stress-responsive enzyme that catabolizes heme into carbon monoxide (CO), biliverdin, and iron, has previously been shown to protect grafts from ischemia/reperfusion injury and rejection. Here we investigated the protective potential of HO-1 in 5 models of immune-mediated liver injury. We found that up-regulation of endogenous HO-1 by cobalt-protoporphyrin-IX (CoPP) protected mice from apoptotic liver damage induced by anti-CD95 antibody (Ab) or d-galactosamine in combination with either anti-CD3 Ab, lipopolysaccharide (LPS), or tumor necrosis factor alpha (TNF-alpha). HO-1 induction prevented apoptotic liver injury, measured by inhibition of caspase 3 activation, although it did not protect mice from caspase-3-independent necrotic liver damage caused by concanavalin A (Con A) administration. In addition, overexpression of HO-1 by adenoviral gene transfer resulted in protection from apoptotic liver injury, whereas inhibition of HO-1 enzymatic activity by tin-protoporphyrin-IX (SnPP) abrogated the protective effect. HO-1-mediated protection seems to target parenchymal liver cells directly because CoPP treatment protected isolated primary hepatocytes from anti-CD95-induced apoptosis in vitro. Furthermore, depletion of Kupffer cells (KCs) did not interfere with the protective effect in vivo. Exogenous CO administration or treatment with the CO-releasing agent methylene chloride mimicked the protective effect of HO-1, whereas treatment with exogenous biliverdin or overexpression of ferritin by recombinant adenoviral gene transfer did not. In conclusion, HO-1 is a potent protective factor for cytokine- and CD95-mediated apoptotic liver damage. Induction of HO-1 might be of a therapeutic modality for inflammatory liver diseases.
    Hepatology 11/2003; 38(4):909-18. · 11.19 Impact Factor
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    ABSTRACT: Heme oxygenase-1 (HO-1) is induced under a variety of pro-oxidant conditions such as those associated with ischemia-reperfusion injury (IRI) of transplanted organs. HO-1 cleaves the heme porphyrin ring releasing Fe2+, which induces the expression of the Fe2+ sequestering protein ferritin. By limiting the ability of Fe2+ to participate in the generation of free radicals through the Fenton reaction, ferritin acts as an anti-oxidant. We have previously shown that HO-1 protects transplanted organs from IRI. We have linked this protective effect with the anti-apoptotic action of HO-1. Whether the iron-binding properties of ferritin contributed to the protective effect of HO-1 was not clear. We now report that recombinant adenovirus mediated overexpression of the ferritin heavy chain (H-ferritin) gene protects rat livers from IRI and prevents hepatocellular damage upon transplantation into syngeneic recipients. The protective effect of H-ferritin is associated with the inhibition of endothelial cell and hepatocyte apoptosis in vivo. H-ferritin protects cultured endothelial cells from apoptosis induced by a variety of stimuli. These findings unveil the anti-apoptotic function of H-ferritin and suggest that H-ferritin can be used in a therapeutic manner to prevent liver IRI and thus maximize the organ donor pool used for transplantation.
    The FASEB Journal 10/2003; 17(12):1724-6. · 5.48 Impact Factor
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    ABSTRACT: Heme oxygenase (HO)-1 catabolizes heme into three products: carbon monoxide (CO), biliverdin (which is rapidly converted to bilirubin) and free iron (which leads to the induction of ferritin, an iron-binding protein). HO-1 serves as a "protective" gene by virtue of the anti-inflammatory, anti-apoptotic and anti-proliferative actions of one or more of these three products. Administration of CO, biliverdin, bilirubin or iron-binding compounds is protective in rodent disease models of ischemia-reperfusion injury, allograft and xenograft survival, intimal hyperplasia following balloon injury or as seen in chronic graft rejection and others. We suggest that the products of HO-1 action could be valuable therapeutic agents and speculate that HO-1 functions as a "therapeutic funnel", mediating the beneficial effects attributed to other molecules, such as interleukin-10 (IL-10), inducible nitric oxide synthase (NOS2; iNOS) and prostaglandins. This Review is the third in a series on the regulation of the immune system by metabolic pathways.
    Trends in Immunology 09/2003; 24(8):449-55. · 12.03 Impact Factor

Publication Stats

2k Citations
117.96 Total Impact Points


  • 2014
    • Hokkaido University
      Sapporo, Hokkaidō, Japan
  • 2008
    • Medizinische Universität Innsbruck
      • Department für Operative Medizin
      Innsbruck, Tyrol, Austria
  • 2003–2006
    • Beth Israel Deaconess Medical Center
      • Department of Surgery
      Boston, Massachusetts, United States
    • University of Pittsburgh
      • Department of Critical Care Medicine
      Pittsburgh, PA, United States