Gentaro Hirokata

Long-Term Hepatic Allograft Acceptance Based on CD40 Blockade by ASKP1240 in Nonhuman Primates

Blockade of the CD40–CD154 costimulatory signal is an attractive strategy for immunosuppression and tolerance induction in organ transplantation. Treatment with anti‐CD154 monoclonal antibodies (mAbs) results in potent immunosuppression in nonhuman primates (NHPs). Despite plans for future clinical use, further development of these treatments was halted by complications. As an alternative approach, we have been focusing on the inhibition of the counter receptor, CD40 and have shown that a novel human anti‐CD40 mAb, ASKP1240, markedly prolongs renal allograft survival in NHPs, although allografts eventually underwent chronic allograft nephropathy. On the basis of our previous findings that a CD40–CD154 costimulation blockade induces tolerance to hepatic, but not cardiac, allografts in rodents, we tested here our hypothesis that a blockade of CD40 by ASKP1240 allows acceptance of hepatic allografts in NHPs. A 2‐week ASKP1240 induction treatment prolonged liver allograft survival in NHPs; however, the graft function deteriorated due to chronic rejection. In contrast, a 6‐month ASKP1240 maintenance monotherapy efficiently suppressed both cellular and humoral alloimmune responses and prevented rejection on the hepatic allograft. No serious side effects, including thromboembolic complications, were noted in the ASKP1240‐treated monkeys. We conclude that CD40 blockade by ASKP1240 would be a desirable immunosuppressant for clinical liver transplantation.

Inhibition of macrophage activation and suppression of graft rejection by DTCM-glutarimide, a novel piperidine derived from the antibiotic 9-methylstreptimidone

ObjectiveWe have previously synthesized a novel piperidine compound, 3-[(dodecylthiocarbonyl)methyl]glutarimide (DTCM-glutarimide), that inhibits LPS-induced NO production, and in the present research we studied further the anti-inflammatory activity of DTCM-glutarimide in a macrophage cell line and in mice bearing transplanted hearts.Materials and methodsMouse macrophage-like RAW264.7 cells were employed for the evaluation of cellular inflammatory activity. DTCM-glutarimide was synthesized in our laboratory. The AP-1 activity was measured by nuclear translocation and phosphorylation. For the heart transplantation experiment, male C57BL/6 (H-2b) and BALB/c (H-2d) mice were used as donor and recipient, respectively. DTCM-glutarimide was administered intraperitoneally.ResultsDTCM-glutarimide inhibited the LPS-induced expression of iNOS and COX-2 in macrophages; but, unexpectedly, it did not inhibit LPS-induced NF-κB activation. Instead, it inhibited the nuclear translocation of both c-Jun and c-Fos. It also inhibited LPS-induced c-Jun phosphorylation. Moreover, it inhibited the mixed lymphocyte reaction in primary cultures of mouse spleen cells; and furthermore, in mice it prolonged the graft survival in heart transplantation experiments.ConclusionThe novel piperidine compound, DTCM-glutarimide, was found to be a new inhibitor of macrophage activation, inhibiting AP-1 activity. It also inhibited graft rejection in mice, and thus may be a candidate for an anti-inflammatory agent.

Risk factors for portal vein complications in pediatric living donor liver transplantation.

Shibasaki S, Taniguchi M, Shimamura T, Suzuki T, Yamashita K, Wakayama K, Hirokata G, Ohta M, Kamiyama T, Matsushita M, Furukawa H, Todo S. Risk factors for portal vein complications in pediatric living donor liver transplantation.
Clin Transplant 2010: 24: 550–556.
© 2009 John Wiley & Sons A/S.

Successful transplantation of rat hearts subjected to extended cold preservation with a novel preservation solution

Since prolonged cold preservation of the heart deteriorates the outcome of heart transplantation, a more protective preservation solution is required. We therefore developed a new solution, named Dsol, and examined whether Dsol, in comparison to UW, could better inhibit myocardial injury resulting from prolonged cold preservation. Syngeneic heterotopic heart transplantation in Lewis rats was performed after cold preservation with UW or Dsol for 24 or 36 h. In addition to graft survival, myocardial injury, ATP content, and Ca2+ ‐dependent proteases activity were assessed in the 24‐h preservation group. The cytosolic Ca2+ concentration of H9c2 cardiomyocytes after 24‐h cold preservation was assessed. Dsol significantly improved 7‐day graft survival after 36‐h preservation. After 24‐h preservation, Dsol was associated with significantly faster recovery of ATP content and less activation of calpain and caspase‐3 after reperfusion. Dsol diminished graft injury significantly, as revealed by the lower levels of infarction, apoptosis, serum LDH and AST release, and graft fibrosis at 7‐day. Dsol significantly inhibited Ca2+ overload during cold preservation. Dsol inhibited myocardial injury and improved graft survival by suppressing Ca2+ overload during the preservation and the activation of Ca2+ ‐dependent proteases. Dsol is therefore considered a better alternative to UW to ameliorate the outcome of heart transplantation.

Dendritic cells conditioned with NK026680 prolong cardiac allograft survival in mice.

Background Pharmacologically modulated dendritic cells (DCs) can potentially regulate alloimmune responses. We examined the characteristics of immunoregulatory DCs induced by a novel triazolopyrimidine derivative, NK026680, which has been previously shown to inhibit DC maturation. Methods DCs were generated from bone marrow progenitor cells from C57BL/6 (B6, H-2b haplotype) mice with granulocyte-macrophage colony-stimulating factor and interleukin (IL)-4. DCs were cultured with allogeneic BALB/c (H-2d) splenocyte lysates with or without NK026680. DC functions were examined in vitro after stimulation of tumor necrosis factor &agr; and in vivo by the intravenous injection of C3He/J (C3H, H-2k) DCs cultured with B6 cell lysates and NK026680 into C3H mice. Seven days later, DC-treated mice received B6 heart allografts, and graft survival and alloimmune responses were assessed. Results In NK026680-treated DCs (NK-DCs), significant inhibition of the up-regulation of surface activation markers (CD40, CD80, CD86, and major histocompatibility complex class II) and IL-12 p40 production was observed after stimulation of tumor necrosis factor &agr; compared with that of control DCs. Furthermore, NK-DCs suppressed alloreactive T-cell proliferation. The modulation of NK-DCs was likely associated with the inhibition of phosphorylation of p38 mitogen-activated protein kinase and the up-regulation of indolamine 2,3-dioxygenase expression. Compared with both noninjected and control DC–injected mice, mice that received a single in vivo infusion of NK-DCs showed significant increases in splenocyte IL-10 production and the splenic CD4+ IL-10high T-cell population 7 days after injection, a significantly increased splenic CD4+CD25+FoxP3+ T-cell population 14 days after injection, and markedly prolonged cardiac allograft survival. Conclusions Ex vivo NK026680 conditioning allows DCs to acquire immunoregulatory properties that suppress alloimmune responses and prolong cardiac allograft survival.

Immunomodulatory effect of nuclear factor-κB inhibition by dehydroxymethylepoxyquinomicin in combination with donor-specific blood transfusion.

Background. Nuclear factor-&kgr;B (NF-&kgr;B) is a key molecule in alloimmune responses, however, its role in tolerance induction is not clear. We have previously reported that dehydroxymethylepoxyquinomycin (DHMEQ), a novel NF-&kgr;B inhibitor, prolongs cardiac allograft survival. In this study, we evaluated the immunomodulatory effects of DHMEQ when combined with a donor-specific blood transfusion (DST), and assessed whether the treatment induces tolerance in a mouse heart transplantation model. Methods. DST (20×106 splenocytes) was given intravenously at day −7. DHMEQ (30 mg/kg/day) was administered intraperitoneally for 14 days after DST. Graft survival and histology were evaluated. The underlying mechanisms of immunomodulation by DST and DHMEQ treatments were investigated by assessing alloimmune responses after transplantation. Results. In fully mismatched H2d-to-H2b heart transplants, DST alone prolonged allograft median survival time to 15 days, whereas when DST was combined with DHMEQ treatment, the graft median survival time was prolonged to 39.5 days. When the donor-recipient strain combination was reversed, that is, H2b-to-H2d, heart transplants were accepted (>150 days survival) in more than 60% of recipients treated with a DST and DHMEQ, whereas control allografts were all rejected within 8 days. The combined therapy markedly inhibited immune responses by both the direct and indirect allorecognition pathways mainly attributed to promotion of activation-induced cell death and Treg generation. Conclusions. Our results demonstrate the distinctive ability of NF-&kgr;B inhibition in combination with donor alloantigen to promote transplantation tolerance through multiple cellular mechanisms.

NK026680 inhibits T-cell function in an IL-2-dependent manner and prolongs cardiac allograft survival in rats

NK026680 is a triazolopyrimidine derivative that has been shown to inhibit dendritic cell maturation and activation. Here, we examined the immunosuppressive properties of NK026680 on T-cell function and assessed its immunosuppressive efficacy in an ACI (RT1(av1) haplotype) to Lewis (RT1(l)) rat heart transplantation model. The effects of NK026680 on T-cell proliferation, activation, and cytokine production were investigated in vitro. Heart transplant recipient rats were administered NK026680 daily for 14 days post-transplantation. In addition to graft survival time, alloimmune responses and graft histology at 4-10 days post-transplantation were assessed. NK026680 was found to inhibit proliferation, CD25 upregulation, IL-2 production, and cell cycle progression in αCD3/αCD28-stimulated murine T cells. These effects were likely due to suppression of the p38 mitogen-activated protein kinase pathway and the subsequent inhibition of p65, c-Fos, and to a lesser extent, c-Jun. Daily NK026680 treatment suppressed alloimmune responses, prevented cellular infiltration into allografts, and prolonged graft survival. The anti-rejection effects of NK026680 were enhanced by tacrolimus. In conclusion, NK026680 inhibits the activation of T cells and prolongs cardiac allograft survival in rats. These features make it a potential candidate immunosuppressant for the treatment of organ transplant patients in the future.

A rescue case of an idiopathic cecum perforation accompanied with sub-acute fulminant hepatitis

症例は38歳女性. 亜急性型劇症肝炎で生体肝移植の適応評価のため当科に転院しICU管理となった. 高容量持続的血液透析濾過と血漿交換を併用する保存的治療を行ったが, 入院10日目に発症した上行結腸破裂に対し右半結腸切除術を行い回腸に人工肛門を増設した. その後肝機能は順調に回復し入院30日目で一時退院, 5カ月後に再入院し人工肛門を閉鎖し現在社会復帰している. 切除した結腸の穿孔部は虚血性腸炎の病理診断を得た. 亜急性型劇症肝炎の保存的治療中に虚血性腸炎による大腸穿孔の報告はなく, その臨床経過と大腸穿孔の原因に考察を加え報告した.