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Lack of retinoblastoma (Rb) protein causes aggressive intraocular retinal tumors in children. Recently, Rb tumors have been shown to have a distinctly altered metabolic phenotype, such as reduced expression of glycolytic pathway proteins alongside altered pyruvate and fatty acid levels. In this study, we demonstrate that loss of hexokinase 1(HK1) in tumor cells rewires their metabolism allowing enhanced oxidative phosphorylation-dependent energy production. We show that rescuing HK1 or retinoblastoma protein 1 (RB1) in these Rb cells reduced cancer hallmarks such as proliferation, invasion, and spheroid formation and increased their sensitivity to chemotherapy drugs. Induction of HK1 was accompanied by a metabolic shift of the cells to glycolysis and a reduction in mitochondrial mass. Cytoplasmic HK1 bound Liver Kinase B1 and phosphorylated AMP-activated kinase-α (AMPKα Thr172 ), thereby reducing mitochondria-dependent energy production. We validated these findings in tumor samples from Rb patients compared to age-matched healthy retinae. HK1 or RB1 expression in Rb-/- cells led to a reduction in their respiratory capacity and glycolytic proton flux. HK1 overexpression reduced tumor burden in an intraocular tumor xenograft model. AMPKα activation by AICAR also enhanced the tumoricidal effects of the chemotherapeutic drug topotecan in vivo. Therefore, enhancing HK1 or AMPKα activity can reprogram cancer metabolism and sensitize Rb tumors to lower doses of existing treatments, a potential therapeutic modality for Rb., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Dilated cardiomyopathy is conventionally defined as the presence of left ventricular or biventricular dilatation or systolic dysfunction in the absence of abnormal loading conditions (eg, primary valve disease) or significant coronary artery disease sufficient to cause ventricular remodelling. This definition has been recognised as overly restrictive, as left ventricular hypokinesis without dilation could be the initial presentation of dilated cardiomyopathy. The causes of dilated cardiomyopathy comprise genetic (primary dilated cardiomyopathy) or acquired factors (secondary dilated cardiomyopathy). Acquired factors include infections, toxins, cancer treatment, endocrinopathies, pregnancy, tachyarrhythmias, and immune-mediated diseases. 5-15% of patients with acquired dilated cardiomyopathy harbour a likely pathogenic or pathogenic gene variant (ie, gene mutation). Therefore, the diagnostic tests and therapeutic approach should always consider both genetic and acquired factors. This Seminar will focus on the current multidimensional diagnostic and therapeutic approach and discuss the underlying pathophysiology that could drive future treatments aiming to repair or replace the existing gene mutation, or target the specific inflammatory, metabolic, or pro-fibrotic drivers of genetic or acquired dilated cardiomyopathy., Competing Interests: Declaration of interests SH receives personal fees for independent scientific advice on early development in the field of heart failure for AstraZeneca, Ribocure, and CSL Behring, and receives research support from AstraZeneca and CSL Behring; acknowledges support from the Netherlands Cardiovascular Research Initiative, an initiative with support of the Dutch Heart Foundation, Dutch Cardiovascular Alliance CVON2016-Early HFPEF 2015–10, CVON She-PREDICTS grant 2017–21, CVON Arena-PRIME 2017–18, and DCVA-Double Dosis 2020-B005; receives funding from IMI2-CARDIATEAM (821508); and is a member of the European Reference Network for rare, low prevalence, and complex diseases of the heart (ERN GUARD-Heart). NKL receives unrestricted research support from Pfizer and the O’Hare and Steggall Family Foundations, and receives modest consulting incomes from Pfizer, Bristol Myers Squibb, Cytokinetics, and Tenaya. CT has received speaker fees or contributions to congresses from Bristol Myers Squibb and Pfizer. KK received moderate speaker fees from Pfizer and Alnylam. We acknowledge funding from the German Research Foundation (Deutsche Forschungsgemeinschaft) with the Collaborative Research Center 1470 (Project B02) to CT and from the Deutsche Herzstiftung (COVID-19) to KK., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Objectives: This study sought to further understand the mechanisms underlying effect of spironolactone and assessed its impact on multiple plasma protein biomarkers and their respective underlying biologic pathways., Background: In addition to their beneficial effects in established heart failure (HF), mineralocorticoid receptor antagonists may act upstream on mechanisms, preventing incident HF. In people at risk for developing HF, the HOMAGE (Heart OMics in AGEing) trial showed that spironolactone treatment could provide antifibrotic and antiremodeling effects, potentially slowing the progression to HF., Methods: Baseline, 1-month, and 9-month (or last visit) plasma samples of HOMAGE participants were measured for protein biomarkers (n = 276) by using Olink Proseek-Multiplex cardiovascular and inflammation panels (Olink, Uppsala, Sweden). The effect of spironolactone on biomarkers was assessed by analysis of covariance and explored by knowledge-based network analysis., Results: A total of 527 participants were enrolled; 265 were randomized to spironolactone (25 to 50 mg/day) and 262 to standard care ("control"). The median (interquartile range) age was 73 years (69 to 79 years), and 26% were female. Spironolactone reduced biomarkers of collagen metabolism (e.g., COL1A1, MMP-2); brain natriuretic peptide; and biomarkers related to metabolic processes (e.g., PAPPA), inflammation, and thrombosis (e.g., IL17A, VEGF, and urokinase). Spironolactone increased biomarkers that reflect the blockade of the mineralocorticoid receptor (e.g., renin) and increased the levels of adipokines involved in the anti-inflammatory response (e.g., RARRES2) and biomarkers of hemostasis maintenance (e.g., tPA, UPAR), myelosuppressive activity (e.g., CCL16), insulin suppression (e.g., RETN), and inflammatory regulation (e.g., IL-12B)., Conclusions: Proteomic analyses suggest that spironolactone exerts pleiotropic effects including reduction in fibrosis, inflammation, thrombosis, congestion, and vascular function improvement, all of which may mediate cardiovascular protective effects, potentially slowing progression toward heart failure. (HOMAGE [Bioprofiling Response to Mineralocorticoid Receptor Antagonists for the Prevention of Heart Failure]; NCT02556450)., Competing Interests: FUNDING SUPPORT AND AUTHOR DISCLOSURES HOMAGE was funded by a grant from the European Union 7th Framework Programme for Research and Technological Development (HEALTH-F7-305507 HOMAGE (EU FP7 305507 http://www.homage-hf.eu). Drs. Ferreira, Rossignol, Girerd, and Zannad are supported by the RHU Fight-HF, a public grant overseen by the French National Research Agency (ANR) as part of the second “Investissements d’Avenir” program (reference: ANR-15-RHUS-0004); the French PIA project “Lorraine Université d’Excellence” (reference: ANR-15-IDEX-04-LUE); Contrat de Plan Etat Lorraine IT2MP; and FEDER Lorraine. All other authors have reported that they have no relationships relevant to the contents of this paper to disclose., (Copyright © 2021 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.)

Myocardial damage as a consequence of cardiotropic viruses leads to a broad variety of clinical presentations and is still a complicated condition to diagnose and treat. Whereas the extracellular matrix protein Secreted Protein Acidic and Rich in Cysteine or SPARC has been implicated in hypertensive and ischemic heart disease by modulating collagen production and cross-linking, its role in cardiac inflammation and endothelial function is yet unknown. Absence of SPARC in mice resulted in increased cardiac inflammation and mortality, and reduced cardiac systolic function upon coxsackievirus-B3 induced myocarditis. Intra-vital microscopic imaging of the microvasculature of the cremaster muscle combined with electron microscopic imaging of the microvasculature of the cardiac muscle uncovered the significance of SPARC in maintaining endothelial glycocalyx integrity and subsequent barrier properties to stop inflammation. Moreover, systemic administration of recombinant SPARC restored the endothelial glycocalyx and consequently reversed the increase in inflammation and mortality observed in SPARC KO mice in response to viral exposure. Reducing the glycocalyx in vivo by systemic administration of hyaluronidase, an enzyme that degrades the endothelial glycocalyx, mimicked the barrier defects found in SPARC KO mice, which could be restored by subsequent administration of recombinant SPARC. In conclusion, the secreted glycoprotein SPARC protects against adverse cardiac inflammation and mortality by improving the glycocalyx function and resulting endothelial barrier function during viral myocarditis., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)

Acute viral myocarditis (VM), characterised by leukocyte infiltration and dysfunction of the heart, is an important cause of sudden cardiac death in young adults. Unfortunately, to date, the pathological mechanisms underlying cardiac failure in VM remain incompletely understood. In the current study, we investigated if acute VM leads to cardiac metabolic rewiring and if this process is driven by local inflammation. Transcriptomic analysis of cardiac biopsies from myocarditis patients and a mouse model of VM revealed prominent reductions in the expression of a multitude of genes involved in mitochondrial oxidative energy metabolism. In mice, this coincided with reductions in high-energy phosphate and NAD levels, as determined by Imaging Mass Spectrometry, as well as marked decreases in the activity, protein abundance and mRNA levels of various enzymes and key regulators of cardiac oxidative metabolism. Indicative of fulminant cardiac inflammation, NF-κB signalling and inflammatory cytokine expression were potently induced in the heart during human and mouse VM. In cultured cardiomyocytes, cytokine-mediated NF-κB activation impaired cardiomyocyte oxidative gene expression, likely by interfering with the PGC-1 (peroxisome proliferator-activated receptor (PPAR)-γ co-activator) signalling network, the key regulatory pathway controlling cardiomyocyte oxidative metabolism. In conclusion, we provide evidence that acute VM is associated with extensive cardiac metabolic remodelling and our data support a mechanism whereby cytokines secreted primarily from infiltrating leukocytes activate NF-κB signalling in cardiomyocytes thereby inhibiting the transcriptional activity of the PGC-1 network and consequently modulating myocardial energy metabolism., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

The small leucine-rich proteoglycan osteoglycin has been implicated in matrix homeostasis in different organs, including the ischemic heart. However, whether osteoglycin modulates cardiac hypertrophy, fibrosis or inflammation in hypertensive heart disease and during aging remains unknown. Angiotensin-II-induced pressure overload increases cardiac osteoglycin expression, concomitant with the onset of inflammation and extracellular matrix deposition. Interestingly aging led to decreased cardiac levels of osteoglycin, yet absence of osteoglycin did not affect organ structure or cardiac function up to the age of 18months. However, Angiotensin-II infusion in combination with aging resulted in exaggerated cardiac fibrosis and inflammation in the osteoglycin null mice as compared to wild-type mice, resulting in increased diastolic dysfunction as determined by magnetic resonance imaging. In vitro, stimulation of bone marrow derived macrophages from osteoglycin null mice with Angiotensin-II resulted in significantly higher levels of ICAM-1 as well as pro-inflammatory cytokines and chemokines IL-1β and MCP-1 as compared to WT cells. Further, stimulation of human cardiac fibroblasts with osteoglycin reduced cell proliferation and inhibited TGF-β induced collagen gene expression. In mouse cardiac tissue, osteoglycin expression inversely correlated with TGF-β expression and in cardiac biopsies of aortic stenosis patients, osteoglycin expression is significantly higher than in control biopsies. Interestingly, osteoglycin levels were higher in patients with less severe myocardial fibrosis and overall in the aortic stenosis patients osteoglycin levels negatively correlated with collagen content in the myocardium. In conclusion, osteoglycin expression is increased in the heart in response to pressure overload and its absence results in increased cardiac inflammation and fibrosis resulting in increased diastolic dysfunction., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Objective: Although lymphocytic myocarditis (LM) clinically can mimic myocardial infarction (MI), they are regarded as distinct clinical entities. However, we observed a high prevalence (32%) of recent MI in patients diagnosed post-mortem with LM. To investigate if LM changes coronary atherosclerotic plaque, we analyzed in autopsied hearts the inflammatory infiltrate and stability in coronary atherosclerotic lesions in patients with LM and/or MI., Methods: The three main coronary arteries were isolated at autopsy of patients with LM, with MI of 3-6h old, with LM and MI of 3-6h old (LM+MI) and controls. In tissue sections of atherosclerotic plaque-containing coronary segments inflammatory infiltration, plaque stability, intraplaque hemorrhage and thrombi were determined via (immuno)histological criteria., Results: In tissue sections of those coronary segments the inflammatory infiltrate was found to be significantly increased in patients with LM, LM+MI and MI compared with controls. This inflammatory infiltrate consisted predominantly of macrophages and neutrophils in patients with only LM or MI, of lymphocytes in LM+MI and MI patients and of mast cells in LM+MI patients. Moreover, in LM+MI and MI patients this coincided with an increase of unstable plaques and thrombi. Finally, LM and especially MI and LM+MI patients showed significantly increased intraplaque hemorrhage., Conclusions: This study demonstrates prevalent co-occurrence of LM with a very recent MI at autopsy. Moreover, LM was associated with remodeling and inflammation of atherosclerotic plaques indicative of plaque destabilization pointing to coronary spasm, suggesting that preexistent LM, or its causes, may facilitate the development of MI., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: There is a clinical need for immunosuppressive therapy that can treat myocarditis patients in the presence of an active viral infection. In this study we therefore investigated the effects of colchicine, an immunosuppressive drug which has been used successfully as treatment for pericarditis patients, in a mouse model of coxsackievirus B3(CVB3)-induced myocarditis., Methods: Four groups of C3H mice were included: control mice (n=8), mice infected with CVB3 (1×10(5) PFU, n=10), mice with colchicine administration (2mg/kg i.p, n=5) and mice with combined CVB3 infection and colchicine administration (n=10). After three days, the heart, pancreas and spleen were harvested and evaluated using (immuno)histochemical analysis and CVB3 qPCR., Results: Mice were terminated at day 3 post-virus infection as colchicine treatment rapidly resulted in severe illness and mortality in CVB3-infected mice. Colchicine significantly decreased the number of macrophages in the heart in CVB3-infected mice (p<0.01) but significantly increased the number of neutrophils (p<0.01). In the pancreas, colchicine caused complete destruction of the acini in the CVB3-infected mice and also significantly decreased macrophage (p<0.01) and increased neutrophil numbers (p<0.01). In the spleen, colchicine treatment of CVB3-infected mice induced massive apoptosis in the white pulp and significantly inhibited the virus-induced increase of megakaryocytes in the spleen (p<0.001). Finally, we observed that colchicine significantly increased CVB3 levels in both the pancreas and the heart., Conclusions: Colchicine treatment in CVB3-induced myocarditis has a detrimental effect as it causes complete destruction of the exocrine pancreas and enhances viral load in both heart and pancreas., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Acute cellular rejection (ACR) is the adverse response of the recipient's immune system against the allogeneic graft. Using human surveillance endomyocardial biopsies (EMBs) manifesting ACR and murine allogeneic grafts, we profiled implicated microRNAs (miRs) and mRNAs. MiR profiling showed that miR-21, -142-3p, -142-5p, -146a, -146b, -155, -222, -223, and -494 increased during ACR in humans and mice, whereas miR-149-5p decreased. mRNA profiling revealed 70 common differentially regulated transcripts, all involved in immune signaling and immune-related diseases. Interestingly, 33 of 70 transcripts function downstream of IL-6 and its transcription factor spleen focus forming virus proviral integration oncogene (SPI1), an established target of miR-155, the most upregulated miR in human EMBs manifesting rejection. In a mouse model of cardiac transplantation, miR-155 absence and pharmacological inhibition attenuated ACR, demonstrating the causal involvement and therapeutic potential of miRs. Finally, we corroborated our miR signature in acute cellular renal allograft rejection, suggesting a nonorgan specific signature of acute rejection. We concluded that miR and mRNA profiling in human and murine ACR revealed the shared significant dysregulation of immune genes. Inflammatory miRs, for example miR-155, and transcripts, in particular those related to the IL-6 pathway, are promising therapeutic targets to prevent acute allograft rejection., (© Copyright 2015 The American Society of Transplantation and the American Society of Transplant Surgeons.)

Background: To investigate the prevalence and prognostic relevance of cardiac involvement in an ANCA-associated vasculitis (AAV) population of eosinophilic granulomatosis with polyangiitis (EGPA) and granulomatosis with polyangiitis (GPA) patients., Methods: Prospective cohort study of fifty EGPA and forty-one GPA patients in sustained remission without previous in-depth cardiac screening attending our clinical immunology outpatient department. Cardiac screening included clinical evaluation, ECG, 24-hour Holter registration, echocardiography and cardiac magnetic resonance imaging (CMR) with coronary angiography and endomyocardial biopsy upon indication. Fifty age-, sex- and cardiovascular risk factor-matched control subjects were randomly selected from a population study. Long-term outcome was assessed using all-cause and cardiovascular mortality., Results: A total of 91 AAV-patients (age 60±11, range 63-87years) were compared to 50-matched control subjects (age 60±9years, range 46-78years). ECG and echocardiography demonstrated cardiac abnormalities in 62% EGPA and 46% GPA patients vs 20% controls (P<0.001 and P=0.014, respectively). A total of 69 AAV-patients underwent additional CMR, slightly increasing the prevalence of cardiac involvement to 66% in EGPA and 61% in GPA patients. After a mean follow-up of 53±18months, presence of cardiac involvement using ECG and echocardiography in AAV-patients showed increased all-cause and cardiovascular mortality (Log-rank P=0.015 and Log-rank P=0.021, respectively)., Conclusion: Cardiac involvement in EGPA and GPA patients with sustained remission is high, even if symptoms are absent and ECG is normal. Moreover, cardiac involvement is a strong predictor of (cardiovascular) mortality. Therefore, risk stratification using cardiac imaging is recommended in all AAV-patients, irrespective of symptoms or ECG abnormalities., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Background: The relation between myocardial scar and different types of ventricular arrhythmias in patients with nonischemic dilated cardiomyopathy (NIDCM) is unknown., Objectives: The purpose of this study was to analyze the effect of myocardial scar, assessed by late gadolinium enhancement cardiac magnetic resonance imaging (LGE-CMR), on the occurrence and type of ventricular arrhythmia in patients with NIDCM., Methods: Consecutive patients with NIDCM who underwent LGE-CMR and implantable cardioverter-defibrillator (ICD) implantation at either of 2 centers were included. LGE was defined by signal intensity ≥35% of maximal signal intensity, subdivided into core and border zones (≥50% and 35%-50% of maximal signal intensity, respectively), and categorized according to location (basal or nonbasal) and transmurality. ICD recordings and electrocardiograms were reviewed to determine the occurrence and type of ventricular arrhythmia during follow-up., Results: Of 87 patients (age 56 ± 13 y, 62% male, left ventricular ejection fraction 29% ± 12%), 55 (63%) had LGE (median 6.3 g, interquartile range 0.0-13.8 g). During a median follow-up of 45 months, monomorphic ventricular tachycardia (VT) occurred in 18 patients (21%) and polymorphic VT/ventricular fibrillation (VF) in 10 (11%). LGE predicted monomorphic VT (log-rank, P < .001), but not polymorphic VT/VF (log-rank, P = .40). The optimal cutoff value for the extent of LGE to predict monomorphic VT was 7.2 g (area under curve 0.84). Features associated with monomorphic VT were core extent, basal location, and area with 51%-75% LGE transmurality., Conclusions: Myocardial scar assessed by LGE-CMR predicts monomorphic VT, but not polymorphic VT/VF, in NIDCM. The risk for monomorphic VT is particularly high when LGE shows a basal transmural distribution and a mass ≥7.2 g. Importantly, patients without LGE on CMR remain at risk for potentially fatal polymorphic VT/VF., (Copyright © 2015 Heart Rhythm Society. Published by Elsevier Inc. All rights reserved.)

Background: Diagnosing lymphocytic myocarditis (LM) is challenging because of the large variation in clinical presentation and the limitations inherent in current diagnostic tools. The objective of this study was to analyze infiltration of inflammatory cells in quadriceps skeletal muscle of LM patients and investigate the potential diagnostic value of assaying infiltrating inflammatory cells., Methods: Quadriceps muscle tissue, obtained at autopsy from control patients (n = 9) and LM patients (n = 21), was analyzed using immunohistochemistry for infiltration of lymphocytes (CD45), macrophages (CD68), neutrophilic granulocytes (myeloperoxidase), and several lymphocyte subtypes (CD3, CD4, CD8, CD20) and using polymerase chain reaction for a panel of myocarditis-associated viruses. Additionally, quadriceps muscle from mice with acute coxsackievirus B3-induced myocarditis and control mice was analyzed for presence of lymphocytes and virus., Results: In quadriceps muscle of LM patients the number of infiltrating lymphocytes were significantly increased and LM was diagnosed with specificity of 100% and sensitivity of 71%. Parvovirus B19 was the primary virus found in our patient groups, found in quadriceps tissue of 3 LM patients (although it was also found in 1 control patient). In the mice, enteroviral RNA was present in the quadriceps muscle, although enteroviral capsid proteins and lymphocyte infiltration were found primarily in the adipose tissue within and directly adjacent to the myocyte tissue, rather than in the myocyte tissue itself., Conclusions: LM is associated with lymphocyte infiltration and viral presence in quadriceps muscle. This indicates that skeletal muscle biopsy/lymphocyte quantification might be a potential diagnostic tool for LM patients., (Copyright © 2014 Canadian Cardiovascular Society. Published by Elsevier Inc. All rights reserved.)

Viral myocarditis is a life-threatening disease characterized by severe cardiac inflammation that can result in heart failure or sudden cardiac death in previously healthy adults. In a subset of patients, it may result in the development of dilated cardiomyopathy due to the chronic inflammatory process. Despite its clinical need, specific treatments for myocarditis are currently not available. The extracellular matrix (ECM) under normal conditions, functions to maintain the mechanical and structural integrity of the heart but can adapt under pathological circumstances to preserve cardiac function. Recent studies have revealed a crucial role of the ECM in the reparative process after cardiac insult, not only as a key component in cardiac remodeling but also as a regulator of the inflammatory process. Increasing our understanding of the impact the ECM has in the disease pathogenesis and progression of viral myocarditis, might lead to much needed therapeutic interventions. In this review we will describe the pathology of viral myocarditis and illustrate the interplay between inflammation and the ECM in general terms, and during viral myocarditis., (Copyright © 2011 Elsevier GmbH. All rights reserved.)

Left ventricular (LV) hypertrophy is a natural response of the heart to increased pressure loading, but accompanying fibrosis and dilatation may result in irreversible life-threatening heart failure. Matrix metalloproteinases (MMPs) have been invoked in various cardiac diseases, however, direct genetic evidence for a role of the plasminogen activator (PA) and MMP systems in pressure overload-induced LV hypertrophy and in heart failure is lacking. Therefore, the consequences of transverse aortic banding (TAB) were analyzed in mice lacking tissue-type PA (t-PA(-/-)), urokinase-type PA (u-PA(-/-)), or gelatinase-B (MMP-9(-/-)), and in wild-type (WT) mice after adenoviral gene transfer of the PA-inhibitor PAI-1 or the MMP-inhibitor TIMP-1. TAB elevated LV pressure comparably in all genotypes. In WT and t-PA(-/-) mice, cardiomyocyte hypertrophy was associated with myocardial fibrosis, LV dilatation and dysfunction, and pump failure after 7 weeks. In contrast, in u-PA(-/-) mice or in WT mice after PAI-1- and TIMP-1-gene transfer, cardiomyocyte hypertrophy was moderate and only minimally associated with cardiac fibrosis and LV dilatation, resulting in better preservation of pump function. Deficiency of MMP-9 had an intermediate effect. These findings suggest that the use of u-PA- or MMP-inhibitors might preserve cardiac pump function in LV pressure overloading.

Site directed mutagenesis (350 variants) of recombinant staphylokinase (SakSTAR), a potent fibrin-selective thrombolytic agent, was undertaken in order to reduce its antigenicity while maintaining its potency. Variants with K35A, (ie, Lys[K] in position 35 substituted with Ala[A]), E65D or E65Q, K74R or K74Q, E80A+D82A, K130T, and K135R displayed increased enzymatic activity or reduced binding of human staphylokinase-specific antibodies. Additive mutagenesis identified 8 variants with intact thrombolytic potencies, which absorbed down to less than a third of SakSTAR-specific antibodies. Intra-arterial administration in 61 patients with peripheral arterial occlusion caused no significant allergic reactions. Median neutralizing antibody titers (with 15 to 85 percentiles), expressed as microgram (microg) compound neutralized per milliliter plasma, were 4.4 (0.3 to 49) for the variants, compared with 12 (4 to 100) in 70 patients given wild-type SakSTAR (P =.002 by Mann-Whitney rank sum test). Overt neutralizing antibody induction (more than 5 microg compound neutralized per milliliter plasma) was observed in 57 of 70 patients (81%) given wild-type SakSTAR, but only in 28 of 60 patients (47%) treated with variants (P <.0001 by Fisher exact test). On the basis of this study, the variant SakSTAR (K35A, E65Q, K74R, D82A, S84A, T90A, E99D, T101S, E108A, K109A, K130T, K135R) (code SY155) has been selected for further clinical development. (Blood. 2000;96:1425-1432)

The plasminogen system plays an important role in the proteolytic degradation of extracellular matrices during wound healing. In the present study we investigated the impact of the plasminogen system on cardiac wound healing and function after myocardial infarction. Myocardial infarction was induced in plasminogen-deficient mice (Plg-/-) and in wild-type controls (Plg+/+). Structural analysis 1, 2, and 5 weeks after infarction revealed that infarct healing was virtually abolished in Plg-/- mice, indicating that the plasminogen system is required for the repair process of the heart after infarction. In the absence of plasminogen, inflammatory cells did not migrate into the infarcted myocardium. Necrotic cardiomyocytes were not removed and the formation of granulation tissue and fibrous tissue did not occur. In these non-healing infarcted hearts, LV dilatation was not altered. In addition, gelatinolytic activity of MMP-2 and MMP-9 was depressed in the Plg-/- infarcted hearts, suggesting that the plasmin effect on infarct healing may be mediated by MMPs. Surprisingly, cardiac function was only attenuated to a rather small extent in the Plg-/- infarcted mice when compared to the wild-types. This study provides direct prove that plasmin-mediated proteolysis plays a central role in cardiac wound healing after myocardial infarction in mice.