Tomoyuki Fujita

Egr-1, a master switch coordinating upregulation of divergent gene families underlying ischemic stress.

Activation of the zinc-finger transcription factor early growth response (Egr)-1, initially linked to developmental processes, is shown here to function as a master switch activated by ischemia to trigger expression of pivotal regulators of inflammation, coagulation and vascular hyperpermeability. Chemokine, adhesion receptor, procoagulant and permeability-related genes are coordinately upregulated by rapid ischemia-mediated activation of Egr-1. Deletion of the gene encoding Egr-1 strikingly diminished expression of these mediators of vascular injury in a murine model of lung ischemia/reperfusion, and enhanced animal survival and organ function. Rapid activation of Egr-1 in response to oxygen deprivation primes the vasculature for dysfunction manifest during reperfusion. These studies define a central and unifying role for Egr-1 activation in the pathogenesis of ischemic tissue damage.

Paradoxical rescue from ischemic lung injury by inhaled carbon monoxide driven by derepression of fibrinolysis

Carbon monoxide (CO) can arrest cellular respiration, but paradoxically, it is synthesized endogenously by heme oxygenase type 1 (Ho-1) in response to ischemic stress. Ho-1–deficient (Hmox1−/−) mice exhibited lethal ischemic lung injury, but were rescued from death by inhaled CO. CO drove ischemic protection by activating soluble guanylate cyclase and thereby suppressed hypoxic induction of the gene encoding plasminogen activator inhibitor-1 (PAI-1) in mononuclear phagocytes, which reduced accrual of microvascular fibrin. CO-mediated ischemic protection observed in wild-type mice was lost in mice null for the gene encoding PAI-1 (Serpine1). These data establish a fundamental link between CO and prevention of ischemic injury based on the ability of CO to derepress the fibrinolytic axis. These data also point to a potential therapeutic use for inhaled CO.

Elucidation of the thromboregulatory role of CD39/ectoapyrase in the ischemic brain.

Endothelial CD39 metabolizes ADP released from activated platelets. Recombinant soluble human CD39 (solCD39) potently inhibited ex vivo platelet aggregation in response to ADP and reduced cerebral infarct volumes in mice following transient middle cerebral artery occlusion, even when given 3 hours after stroke. Postischemic platelet and fibrin deposition were decreased and perfusion increased without increasing intracerebral hemorrhage. In contrast, aspirin did not increase postischemic blood flow or reduce infarction volume, but did increase intracerebral hemorrhage. Mice lacking the enzymatically active extracellular portion of the CD39 molecule were generated by replacement of exons 4-6 (apyrase-conserved regions 2-4) with a PGKneo cassette. Although CD39 mRNA 3 of the neomycin cassette insertion site was detected, brains from these mice lacked both apyrase activity and CD39 immunoreactivity. Although their baseline phenotype, hematological profiles, and bleeding times were normal, cd39(-/-) mice exhibited increased cerebral infarct volumes and reduced postischemic perfusion. solCD39 reconstituted these mice, restoring postischemic cerebral perfusion and rescuing them from cerebral injury. These data demonstrate that CD39 exerts a protective thromboregulatory function in stroke.

PKCβ regulates ischemia/reperfusion injury in the lung

Activation of PKCβII is associated with the response to ischemia/reperfusion (I/R), though its role, either pathogenic or protective, has not been determined. In a murine model of single-lung I/R, evidence linking PKCβ to maladaptive responses is shown in the following studies. Homozygous PKCβ-null mice and WT mice fed the PKCβ inhibitor ruboxistaurin subjected to I/R displayed increased survival compared with controls. In PKCβ-null mice, phosphorylation of extracellular signal–regulated protein kinase-1 and -2 (ERK1/2), JNK, and p38 MAPK was suppressed in I/R. Expression of the immediate early gene, early growth response-1 (Egr-1), and its downstream target genes was significantly increased in WT mice in I/R, particularly in mononuclear phagocytes (MPs), whereas this expression was attenuated in PKCβ-null mice or WT mice fed ruboxistaurin. In vitro, hypoxia/reoxygenation-mediated induction of Egr-1 in MPs was suppressed by inhibition of PKCβ, ERK1/2, and JNK, but not by inhibition of p38 MAPK. These findings elucidate key roles for PKCβII activation in I/R by coordinated activation of MAPKs (ERK1/2, JNK) and Egr-1.

Antisense Intercellular Adhesion Molecule-1 (ICAM-1) Oligodeoxyribonucleotide Delivered During Organ Preservation Inhibits Posttransplant ICAM-1 Expression and Reduces Primary Lung Isograft Failure

Transiently increased expression of leukocyte adhesion receptors after lung preservation contributes to early graft demise by recruiting leukocytes, activating complement, and causing microcirculatory stasis. We hypothesized that inhibiting intercellular adhesion molecule-1 (ICAM-1) expression even briefly may significantly improve lung graft function and that the preservation period might provide a unique window to deliver a therapeutic pulse of antisense oligonucleotide ICAM-1 to inhibit ICAM-1 expression after transplantation. Interleukin-1beta-treated rat pulmonary endothelial cells given a 20-mer phosphorothioate oligonucleotide comprising an antisense span targeted to the 3-untranslated region of rat ICAM-1 demonstrated an oligonucleotide dose-dependent reduction in ICAM-1 expression. Using a cationic liposomal carrier, this same antisense oligonucleotide (but not the sense control) instilled into the pulmonary vasculature at the time of preservation reduced subsequent graft ICAM-1 expression and graft leukostasis and markedly improved oxygenation, pulmonary blood flow, and graft survival. These experiments demonstrate that the preservation period presents a window during which to target an anti-ICAM-1 expression strategy to inhibit early adhesion receptor expression and improve functional outcome after lung transplantation.

Clinical Outcomes of Patients with Heartmate II Left Ventricular Assist Device: A Single Center Experience from Japan

BACKGROUNDnLeft ventricular assist device (LVAD) therapy is the gold standard alternative therapy for patients with advanced heart failure. However, LVAD therapy is still uncommon in the Asia-Pacific region. Therefore, we aimed to elucidate the clinical outcomes of patients from Japan supported with the HeartMate II (HM-II) LVAD at our institution.nnnMETHODSnNinety-two patients (mean 44.3 ± 12.1 years, 68 men, average body mass index 1.65 ± 0.28 m2; 81 with nonischemic cardiomyopathy) who underwent HM-II implantation for bridge to transplantation (nxa0= 91) or for destination therapy in a clinical study (nxa0= 1) at the National Cerebral and Cardiovascular Center between April 2013 and October 2017 were enrolled in this analysis. Preoperatively, most patients (nxa0= 73, 79%) had an INTERMACS (Interagency Registry for Mechanically Assisted Circulatory Support) profile of between level 2 and 4. Postoperatively, the average pump speed was 8602 ± 258 rpm and the hemodynamics were well compensated.nnnRESULTSnAdverse events consisted of 38 (41.3%) hemolysis, 30 (32.6%) major infection, 27 (29.3%) major bleeding (6 [6.5%] with gastrointestinal bleeding), and 18 (19.6%) neurologic dysfunction events. Eighteen patients underwent heart transplantation (HTx) after an average of 32.9 ± 8.9 months of VAD support, and overall survival at both 6 months and 3 years was 96.3%.nnnCONCLUSIONnClinical outcome among patients with HM-II at our institution is satisfactory for both survival and adverse events. The HM-II can provide effective hemodynamic support during the extremely long waiting period for HTx in Japan.

Erratum: Egr-1, a master switch coordinating upregulation of divergent gene families underlying ischemic stress (Nature Medicine (2000) 6 (13551-1361))

Egr-1, a master switch coordinating upregulation of divergent gene families underlying ischemic stress Shi-Fang Yan, Tomoyuki Fujita, Jiesheng Lu, Kenji Okada, Yu Shan Zou, Nigel Mackman, David J. Pinsky & David M. Stern Nature Med. 6, 1355–1361 (2000) In Fig. 1, panels h and i are incorrect. The correct panels are shown here.