Transcriptomic Landscape of Circulating Extracellular Vesicles in Heart Transplant Ischemia-Reperfusion.

Ischemia-reperfusion injury (IRI) is an inevitable event during heart transplantation, which is known to exacerbate damage to the allograft. However, the precise mechanisms underlying IRI remain incompletely understood. Here, we profiled the whole transcriptome of plasma extracellular vesicles (EVs) by RNA sequencing from 41 heart transplant recipients immediately before and at 12 h after transplant reperfusion. We found that the expression of 1317 protein-coding genes in plasma EVs was changed at 12 h after reperfusion. Upregulated genes of plasma EVs were related to metabolism and immune activation, while downregulated genes were related to cell survival and extracellular matrix organization. In addition, we performed correlation analyses between EV transcriptome and intensity of graft IRI (i.e., cardiomyocyte injury), as well as EV transcriptome and primary graft dysfunction, as well as any biopsy-proven acute rejection after heart transplantation. We ultimately revealed that at 12 h after reperfusion, 4 plasma EV genes (ITPKA, DDIT4L, CD19, and CYP4A11) correlated with both cardiomyocyte injury and primary graft dysfunction, suggesting that EVs are sensitive indicators of reperfusion injury reflecting lipid metabolism-induced stress and imbalance in calcium homeostasis. In conclusion, we show that profiling plasma EV gene expression may enlighten the mechanisms of heart transplant IRI.

Cancer risk and mortality after solid organ transplantation: A population-based 30-year cohort study in Finland.

Cancer is a significant cause of morbidity and mortality after solid organ transplantation (SOT) and related to lifelong immunosuppression. This retrospective registry study assessed for the first time in Finland population-based cancer risk and cancer mortality after all SOTs (lung and childhood transplantations included) as standardized incidence ratios (SIRs) and standardized mortality ratios (SMRs). Data from transplant registries were linked with the data of Finnish Cancer Registry and Statistics Finland. We followed 6548 consecutive first SOT recipients from 1 January 1987 to 31 December 2016 translating to 66 741 person-years (median follow-up time 8.9 years [interquartile range 4.0-15.1]). In total, 2096 cancers were found in 1483 patients (23% of all patients). Majority of cancers (53%) were nonmelanoma skin cancers (NMSCs). The overall SIR was 3.6 (95% confidence interval [CI]: 3.5-3.8) and the SIR excluding NMSCs was 2.2 (95% CI: 2.0-2.3). SIR for all cancers was highest for heart (5.0) and lowest for liver (2.7) recipients. Most common cancer types after NMSCs were non-Hodgkin lymphoma (NHL), SIR 9.9 (95% CI: 8.5-11.4) and kidney cancer, SIR 7.3 (95% CI: 6.0-8.8). Cancer-related deaths were 17% (n = 408) of all deaths after first month post transplantation. SMR for all cancers was 2.5 (95% CI: 2.2-2.7) and for NHL 13.6 (95% CI: 10.7-16.8). Notably, overall SIR for cancer was lower in later period (2000-2016), 3.0 (95% CI: 2.8-3.2), than in earlier period (1987-1999), 4.3 (95% CI: 4.0-4.5), P < .001. Decrease in cancer incidence was temporally associated with major changes in immunosuppression in the 2000s.

Monogenic gene variants in lung transplant recipients with usual interstitial pneumonia.

AIM: The prevalence of monogenic disease-causing gene variants in lung transplant recipients with idiopathic pulmonary fibrosis is not fully known. Their impact on clinical outcomes before and after transplantation requires more evidence. PATIENTS AND METHODS: We retrospectively performed sequence analysis of genes associated with pulmonary fibrosis in a cohort of 23 patients with histologically confirmed usual interstitial pneumonia that had previously undergone double lung transplantation. We evaluated the impact of confirmed molecular diagnoses on disease progression, clinical outcomes and incidence of acute rejection or chronic lung allograft dysfunction after transplantation. RESULTS: 15 patients out of 23 (65%) had a variant in a gene associated with interstitial lung disease. 11 patients (48%) received a molecular diagnosis, of which nine involved genes for telomerase function. Five diagnostic variants were found in the gene for Telomerase reverse transcriptase. Two of these variants, p.(Asp684Gly) and p.(Arg774*), seemed to be enriched in Finnish lung transplant recipients. Disease progression and the incidence of acute rejection and chronic lung allograft dysfunction was similar between patients with telomere-related disease and the rest of the study population. The incidence of renal or bone marrow insufficiency or skin malignancies did not differ between the groups. CONCLUSION: Genetic variants are common in lung transplant recipients with pulmonary fibrosis and are most often related to telomerase function. A molecular diagnosis for telomeropathy does not seem to impact disease progression or the risk of complications or allograft dysfunction after transplantation.

Inhibition of Vascular Endothelial Growth Factor Receptors 1 and 2 Attenuates Natural Killer Cell and Innate Immune Responses in an Experimental Model for Obliterative Bronchiolitis.

Obliterative bronchiolitis (OB) after lung transplantation is a nonreversible, life-threatening complication. Herein, the role of vascular endothelial growth factor receptor (Vegfr)-1 and -2 was investigated in the development of obliterative airway disease (OAD), an experimental model for OB. The nonimmunosuppressed recipients underwent transplantation with fully major histocompatibility complex mismatched heterotopic tracheal allografts and received Vegfr1 and -2-specific monoclonal antibodies either alone or in combination, or rat IgG as a control. The treatment with Vegfr1- or -2-blocking antibody significantly decreased intragraft mRNA expression of natural killer cell activation markers early after transplantation. This was followed by reduced infiltration of Cd11b+ cells and Cd4+ T cells as well as down-regulated mRNA expression of proinflammatory chemokines and profibrotic growth factors. However, blocking of both Vegfr1 and -2 was necessary to reduce luminal occlusion. Furthermore, concomitant inhibition of the calcineurin activation pathway almost totally abolished the development of OAD. This study proposes that blocking of Vegf receptors blunted natural killer cell and innate immune responses early after transplantation and attenuated the development of OAD. The results of this study suggest that further studies on the role of Vegfr1 and -2 blocking in development of obliterative airway lesions might be rewarding.

Heart Transplantation for Early-Onset Anthracycline-Induced Cardiomyopathy Within 5 Months of Chemotherapy Completion.

A 9-year-old boy developed progressive anthracycline-induced cardiomyopathy three months after completion of chemotherapy for osteosarcoma. Five months after completion of chemotherapy, at the age of 10 years, heart transplantation was performed. At 29 months since transplantation, the patient remains free of rejection and recurrence of osteosarcoma. (Level of Difficulty: Intermediate.).

Epitranscriptomics of Ischemic Heart Disease-The IHD-EPITRAN Study Design and Objectives.

Epitranscriptomic modifications in RNA can dramatically alter the way our genetic code is deciphered. Cells utilize these modifications not only to maintain physiological processes, but also to respond to extracellular cues and various stressors. Most often, adenosine residues in RNA are targeted, and result in modifications including methylation and deamination. Such modified residues as N-6-methyl-adenosine (m6A) and inosine, respectively, have been associated with cardiovascular diseases, and contribute to disease pathologies. The Ischemic Heart Disease Epitranscriptomics and Biomarkers (IHD-EPITRAN) study aims to provide a more comprehensive understanding to their nature and role in cardiovascular pathology. The study hypothesis is that pathological features of IHD are mirrored in the blood epitranscriptome. The IHD-EPITRAN study focuses on m6A and A-to-I modifications of RNA. Patients are recruited from four cohorts: (I) patients with IHD and myocardial infarction undergoing urgent revascularization; (II) patients with stable IHD undergoing coronary artery bypass grafting; (III) controls without coronary obstructions undergoing valve replacement due to aortic stenosis and (IV) controls with healthy coronaries verified by computed tomography. The abundance and distribution of m6A and A-to-I modifications in blood RNA are charted by quantitative and qualitative methods. Selected other modified nucleosides as well as IHD candidate protein and metabolic biomarkers are measured for reference. The results of the IHD-EPITRAN study can be expected to enable identification of epitranscriptomic IHD biomarker candidates and potential drug targets.

Randomised sham-controlled double-blind trial evaluating remote ischaemic preconditioning in solid organ transplantation: a study protocol for the RIPTRANS trial.

INTRODUCTION: Remote ischaemic preconditioning (RIPC) using a non-invasive pneumatic tourniquet is a potential method for reducing ischaemia-reperfusion injury. RIPC has been extensively studied in animal models and cardiac surgery, but scarcely in solid organ transplantation. RIPC could be an inexpensive and simple method to improve function of transplanted organs. Accordingly, we aim to study whether RIPC performed in brain-dead organ donors improves function and longevity of transplanted organs. METHODS AND ANALYSES: RIPTRANS is a multicentre, sham-controlled, parallel group, randomised superiority trial comparing RIPC intervention versus sham-intervention in brain-dead organ donors scheduled to donate at least one kidney. Recipients of the organs (kidney, liver, pancreas, heart, lungs) from a randomised donor will be included provided that they give written informed consent. The RIPC intervention is performed by inflating a thigh tourniquet to 300 mm Hg 4 times for 5 min. The intervention is done two times: first right after the declaration of brain death and second immediately before transferring the donor to the operating theatre. The sham group receives the tourniquet, but it is not inflated. The primary endpoint is delayed graft function (DGF) in kidney allografts. Secondary endpoints include short-term functional outcomes of transplanted organs, rejections and graft survival in various time points up to 20 years. We aim to show that RIPC reduces the incidence of DGF from 25% to 15%. According to this, the sample size is set to 500 kidney transplant recipients. ETHICS AND DISSEMINATION: This study has been approved by Helsinki University Hospital Ethics Committee and Helsinki University Hospital's Institutional Review Board. The study protocol was be presented at the European Society of Organ Transplantation congress in Copenhagen 14-15 September 2019. The study results will be submitted to an international peer-reviewed scientific journal for publication. TRIAL REGISTRATION NUMBER: NCT03855722.

Cholesterol lowering with EVOLocumab to prevent cardiac allograft Vasculopathy in De-novo heart transplant recipients: Design of the randomized controlled EVOLVD trial.

BACKGROUND: Cardiac allograft vasculopathy (CAV) is characterized by diffuse thickening of the arterial intima. Statins reduce the incidence of CAV, but despite the use of statins, CAV remains one of the leading causes of long-term death after heart transplant. Inhibitors of proprotein convertase subtilisin-kexin type 9 (PCSK9) substantially reduce cholesterol levels but have not been tested in heart transplant recipients. METHODS: The Cholesterol lowering with EVOLocumab to prevent cardiac allograft Vasculopathy in De-novo heart transplant recipients (EVOLVD) trial (ClinicalTrials.gov Identifier: NCT03734211) is a randomized, double-blind trial designed to test the effect of the PCSK9 inhibitor evolocumab on coronary intima thickness in heart transplant recipients. Adults who have received a cardiac transplant within the past 4-8 weeks are eligible. Exclusion criteria include an estimated glomerular filtration rate < 20 mL/min/1.73 m2 , renal replacement therapy, or contraindications to coronary angiography with intravascular ultrasound. 130 patients will be randomized (1:1) to 12-month treatment with evolocumab or matching placebo. The primary endpoint is the coronary artery intima thickness as measured by intravascular ultrasound. CONCLUSION: The EVOLVD trial is a randomized clinical trial designed to show whether treatment with the PCSK9 inhibitor evolocumab can ameliorate CAV over the first year after heart transplant.

Elevated Circulating Activin A Levels in Patients With Malignant Pleural Mesothelioma Are Related to Cancer Cachexia and Reduced Response to Platinum-based Chemotherapy.

BACKGROUND: Previous preclinical studies have shown that activin A is overexpressed in malignant pleural mesothelioma (MPM), associates with cancer cachexia, and is observed in in vitro resistance to platinum-based chemotherapy. We evaluated circulating activin levels and their endogenous antagonists' follistatin/follistatin-like 3 in intrathoracic tumors. MATERIALS AND METHODS: Patients suspected of thoracic malignancy were recruited prior to surgery. Serum samples were collected from 21 patients with MPM, 59 patients with non-small-cell lung cancer (NSCLC), and 22 patients with benign lung lesions. Circulating activin/follistatin levels were measured using enzyme-linked immunosorbent assay and compared with clinicopathologic parameters. RESULTS: Circulating activin A levels were elevated in patients with MPM when compared with patients with NSCLC or benign lung lesion samples (P < .0001). Also, follistatin and follistatin-like 3 levels were the highest in MPM, although with less difference compared with activin A. Receiver operating characteristic analysis for activin A for separating NSCLC from benign lung lesion showed an area under the curve of 0.856 (95% confidence interval, 0.77-0.94). Activin A levels were higher in patients with cachexia (P < .001). In patients with MPM, activin A levels correlated positively with computed tomography-based baseline tumor size (R = 0.549; P = .010) and the change in tumor size after chemotherapy (R = 0.743; P = .0006). Patients with partial response or stable disease had lower circulating activin A levels than the ones with progressive disease (P = .028). CONCLUSION: Activin A serum level could be used as a biomarker in differentiating malignant and benign lung tumors. Circulating activin A levels were elevated in MPM and associates with cancer cachexia and reduced chemotherapy response.

Donor Simvastatin Treatment in Heart Transplantation.

BACKGROUND: Ischemia-reperfusion injury may compromise the short-term and long-term prognosis after heart transplantation. Experimental studies show that simvastatin administered to the organ donor is vasculoprotective and inhibits cardiac allograft ischemia-reperfusion injury. METHODS: Eighty-four multiorgan donors were randomly assigned to receive 80 mg of simvastatin (42 donors) via nasogastric tube after declaration of brain death and upon acceptance as a cardiac donor, or to receive no simvastatin (42 donors). The primary efficacy end point was postoperative plasma troponin T and I levels during the first 24 hours after heart transplantation. Secondary end points included postoperative hemodynamics, inflammation, allograft function, rejections and rejection treatments, and mortality. Results: Organ donor simvastatin treatment significantly reduced the heart recipient plasma levels of troponin T by 34% (14 900 ± 12 100 ng/L to 9800 ± 7900 ng/L, P=0.047), and troponin I by 40% (171 000 ± 151 000 ng/L to 103 000 ± 109 000 ng/L, P=0.023) at 6 hours after reperfusion, the levels of NT-proBNP (N-terminal pro-B-type natriuretic peptide) by 36% (32 800 ± 24 300 ng/L to 20 900 ± 15 900 ng/L; P=0.011) at 1 week, and the number of rejection treatments with hemodynamic compromise by 53% within the first 30 days (P=0.046). Donor simvastatin treatment did not affect donor lipid levels but was associated with a specific transplant myocardial biopsy gene expression profile, and a decrease in recipient postoperative plasma levels of CXCL10 (C-X-C motif chemokine 10), interleukin-1α, placental growth factor, and platelet-derived growth factor-BB. Postoperative hemodynamics, biopsy-proven acute rejections, and mortality were similar. No adverse effects were seen in recipients receiving noncardiac solid organ transplants from simvastatin-treated donors. CONCLUSIONS: Donor simvastatin treatment reduces biomarkers of myocardial injury after heart transplantation, and-also considering its documented general safety profile-may be used as a novel, safe, and inexpensive adjunct therapy in multiorgan donation. CLINICAL TRIAL REGISTRATION: URL: https://www.clinicaltrials.gov. Unique identifier: NCT01160978.

Cancer incidence and mortality after heart transplantation - A population-based national cohort study.

Background: Cancer is currently one of the most important factors affecting the long-term health and survival of heart transplant patients. Material and methods: We calculated the standardized incidence ratios (SIR) for different cancer sites and the cancer-specific standardized mortality ratio (SMR) by linking a cohort of 479 adult heart transplant recipients transplanted in 1985-2014 (4491.6 person-years of follow-up) with data from the national Finnish Cancer Registry until the end of 2015, and with the data from the Statistics Finland's national registry of causes of death. Results: A total of 267 cancers occurred in 143 patients (SIR 6.0; 95% confidence interval (CI) 5.3-6.7). The SIR for overall cancer was considerably higher for men (SIR 6.7; 95% CI 5.9-7.5) than for women (1.4; 95% CI 0.6-2.6). Most frequent cancers were non-melanoma skin cancers (basal cell carcinoma 83 cases, squamous cell skin cancer (SCC) 56 cases), followed by Non-Hodgkin lymphoma (NHL) (36 cases), lung cancer (17), cancer of prostate (16) and cancer of kidney (12). SIRs were highest for SCC (51.9; 95% CI 39.2-67.4), lip cancer (47.4; 95% CI 19.1-97.7), cancer of tongue (26.3; 95% CI 7.2-67.4), and NHL (25.7; 95% CI 18.0-35.6). For most cancers, SIRs increased steadily by time since transplantation. Cancer mortality was three times higher for heart transplant recipients than for the population (SMR 3.1; 95% CI 2.1-4.1). Conclusions: Both cancer incidence and mortality are remarkably increased after heart transplantation, with the relative incidence most elevated for SCC, lip and other oral cancers, and for NHL.

Hypoxia-inducible factor controls immunoregulatory properties of myeloid cells in mouse cardiac allografts - an experimental study.

Hypoxia-inducible factors (HIFs) play a critical role in inflammatory properties of myeloid-derived cells. The effect of HIFs on myeloid-derived cell functions in organ transplantation remains unknown, however. We transplanted hearts into transgenic mice with myeloid cell-targeted deletions of HIF-1α or its negative regulator von Hippel-Lindau (VHL) to investigate the effects of HIF-1α inactivation or HIF pathway activation, respectively, on ischemia-reperfusion injury (IRI) and acute rejection. Deletion of VHL in myeloid cells enhanced mRNA expression of anti-inflammatory genes IDO, Arg-1, and HO-1 in vitro. In vivo, VHL-/- myeloid-derived cells of allograft recipients alleviated IRI and acute rejection, evidenced by reduced cardiomyocyte damage, decreased proinflammatory cytokine mRNA levels, and absence of inflammatory infiltrate at 5 days after transplantation. Ultimately, allograft survival was significantly prolonged. In vitro, VHL-/- myeloid-derived cells dose-dependently inhibited T-cell proliferation. Myeloid cells with HIF-1α-deletion retained proinflammatory qualities in vitro and in vivo. Deletion of VHL in myeloid cells of nonimmunosuppressed cardiac allograft recipients reduced myocardial injury and acute rejection. We suggest that HIF transcription factors induce a regulatory phenotype in myeloid-derived cells, which may be harnessed as a novel therapeutic strategy to regulate immune responses after heart transplantation.

Extracorporeal membrane oxygenation for refractory cardiogenic shock: patient survival and health-related quality of life.

OBJECTIVES: Our goal was to study the outcome of patients with cardiogenic shock who were treated with venoarterial extracorporeal membrane oxygenation (VA ECMO), including the subsequent long-term health-related quality of life (HRQoL). METHODS: We conducted a retrospective study of 133 consecutive patients treated in a single centre from 2007 to 2016. The HRQoL was studied using the EuroQuol-5 dimensions-3 level questionnaire and the RAND 36-Item Short Form Health Survey at a minimum of 1 year after VA ECMO. RESULTS: Of all patients, 66 (49.6%) were weaned from VA ECMO and 16 (12.0%) patients were bridged directly to a transplant, 15 (11.3%) to a ventricular assist device and 1 (0.8%) to a total artificial heart. Survival to discharge was 63.9% and to 1 year, 60.9%. A higher in-hospital mortality rate was independently associated with lower HCO3 at VA ECMO implantation [odds ratio (OR) 1.2/decrease of 1 mmol/l in HCO3 (95% confidence interval 1.1-1.3, P < 0.001)] and with increased need of red blood cells transfused during intensive care [OR 1.9/unit of red blood cells needed/day (95% confidence interval 1.4-2.6, P < 0.001)]. HRQoL measured with the EuroQuol-5 dimensions-3 level questionnaire was equal to the HRQoL of the general population. In the 36-Item Short Form questionnaire, patients reported better emotional well-being and equal energy, pain and general health perception compared to the general population. Limitations were experienced only in physical health. In total, 56% of the patients ≤ 60 years had returned to work. CONCLUSIONS: VA ECMO can provide acceptable long-term survival with good HRQoL for selected patients with refractory cardiogenic shock. Timing of patient assessment and of VA ECMO implantation is essential because deeper acidosis is associated with a higher in-hospital mortality rate.

Propranolol for gastrointestinal bleeding in two patients with continuous flow left ventricle assist device.

Patients with continuous-flow left ventricle assist devices are at risk for gastrointestinal bleeding from angiodysplastic bowel lesions. Neoangiogenesis secondary to von Willebrand factor degradation and increased vascular endothelial growth factor (VEGF) signalling is likely related to their pathophysiology. We speculated that propranolol, known to downregulate VEGF signalling, could be beneficial in patients with recurrent bleeding episodes and anaemia. In this case report, we present a short-term outcome of 2 patients treated with propranolol.

Dual antiplatelet and anticoagulant APAC prevents experimental ischemia-reperfusion-induced acute kidney injury.

BACKGROUND: Renal ischemia-reperfusion predisposes to acute kidney injury (AKI) and mortality. APAC, mast cell heparin proteoglycan mimetic is a potent dual antiplatelet and anticoagulant inhibiting thrombosis in several vascular models. METHODS: Clinically relevant (0.06 and 0.13 mg/kg) and high (0.32 and 7.3 mg/kg) heparin doses of APAC and unfractionated heparin (UFH) were administered i.v. in pharmacological studies. Antithrombotic action of APAC and UFH was assessed with platelet aggregation to collagen, activated partial thromboplastin (APTT) and prothrombin (PT) times. Pharmacodynamics of [64Cu]-APAC or -UFH were monitored by PET/CT. Next, APAC and UFH doses (0.06 and 0.13 mg/kg) were i.v. administered 10 min prior to renal ischemia-reperfusion injury (IRI) in rats. RESULTS: APAC in contrast to UFH inhibited platelet aggregation. During 0.06 and 0.13 mg/kg dose regimens APTT and PT remained at baseline, but at the high APTT prolonged fourfold to sixfold. Overall bio-distribution and clearance of APAC and UFH were similar. After bilateral 30-min renal artery clamping, creatinine, urea nitrogen and neutrophil gelatinase-associated lipocalin concentrations and histopathology indicated faster renal recovery by APAC (0.13 mg/kg). APAC, unlike UFH, prevented expression of innate immune ligand hyaluronan and tubulointerstitial injury marker Kim-1. Moreover, in severe bilateral 1-h renal artery clamping, APAC (0.13 mg/kg) prevented AKI, as demonstrated both by biomarkers and survival. Compatible with kidney protection APAC reduced the circulating levels of vascular destabilizing and pro-inflammatory angiopoietin-2 and syndecan-1. No tissue bleeding ensued. CONCLUSION: APAC and UFH were similarly eliminated via kidneys and liver. In contrast to UFH, APAC (0.13 mg/kg) was reno-protective in moderate and even severe IRI by attenuating vascular injury and innate immune activation.

Vascular Endothelial Growth Factor-B Overexpressing Hearts Are Not Protected From Transplant-Associated Ischemia-Reperfusion Injury.

OBJECTIVES: Cardiac vascular endothelial growth factor-B transgene limits myocardial damage in rat infarction models. We investigated whether heart transplant vascular endothelial growth factor-B overexpression protected against ischemia-reperfusion injury. MATERIALS AND METHODS: We transplanted hearts heterotopically from Dark Agouti to Wistar Furth rats. To characterize the role of vascular endothelial growth factor-B in ischemia-reperfusion injury, we transplanted either long-term human vascular endothelial growth factor-B transgene overexpressing hearts from Wistar Furth rats or short-term adeno-associated virus 9-human vascular endothelial growth factor-B-transduced hearts from Dark Agouti rats into Wistar Furth rats. Heart transplants were subjected to 2 hours of cold and 1 hour of warm ex vivo ischemia. Samples were collected 6 hours after reperfusion. RESULTS: Two hours of cold and 1 hour of warm ischemia increased vascular endothelial growth factor-B mRNA levels 2-fold before transplant and 6 hours after reperfusion. Transgenic vascular endothelial growth factor-B overexpression caused mild cardiac hypertrophy and elevated cardiac troponin T levels 6 hours after reperfusion. Laser Doppler measurements indicated impaired epicardial tissue perfusion in these transgenic transplants. Recombinant human vascular endothelial growth factor-B increased mRNA levels of cytochrome c oxidase and extracellular ATPase CD39, suggesting active oxidative phosphorylation and high ATP production. Adeno-associated virus 9-mediated vascular endothelial growth factor-B overexpression in transplanted hearts increased intragraft macrophages 1.5-fold and proinflammatory cytokine interleukin 12 p35 mRNA 1.6-fold, without affecting recipient serum cardiac troponin T concentration. CONCLUSIONS: Vascular endothelial growth factor-B expression in transplanted hearts is linked to ischemia and ischemia-reperfusion injury. Cardiac transgenic vascular endothelial growth factor-B overexpression failed to protect heart transplants from ischemia-reperfusion injury.

High relative density of lymphatic vessels predicts poor survival in tongue squamous cell carcinoma.

Tongue cancer has a poor prognosis due to its early metastasis via lymphatic vessels. The present study aimed at evaluating lymphatic vessel density, relative density of lymphatic vessel, and diameter of lymphatic vessels and its predictive role in tongue cancer. Paraffin-embedded tongue and lymph node specimens (n = 113) were stained immunohistochemically with a polyclonal antibody von Willebrand factor, recognizing blood and lymphatic endothelium and with a monoclonal antibody podoplanin, recognizing lymphatic endothelium. The relative density of lymphatic vessels was counted by dividing the mean number of lymphatic vessels per microscopic field (podoplanin) by the mean number of all vessels (vWf) per microscopic field. The high relative density of lymphatic vessels (≥80 %) was associated with poor prognosis in tongue cancer. The relative density of lymphatic vessels predicted poor prognosis in the group of primary tumor size T1-T2 and in the group of non-metastatic cancer. The lymphatic vessel density and diameter of lymphatic vessels were not associated with tongue cancer survival. The relative density of lymphatic vessels might have clinically relevant prognostic impact. Further studies with increased number of patients are needed.

Simvastatin pretreatment reduces caspase-9 and RIPK1 protein activity in rat cardiac allograft ischemia-reperfusion.

BACKGROUND: In transplantation-associated ischemia/reperfusion injury (Tx-IRI), tumor necrosis factor alpha and damage-associated molecular patterns promote caspase-8 and -9 apoptotic and receptor-interacting protein kinase-1 and -3 (RIPK1/3) necroptotic pathway activation. The extent of cell death and the counterbalance between apoptosis and regulated necrosis eventually determine the immune response of the allograft. Although simvastatin prevents Tx-IRI, its role in apoptotic and necroptotic activity remains unsolved. METHODS: Rat allograft donors and recipients were treated with a single-dose of simvastatin 2h prior to allograft procurement and reperfusion, respectively. Intragraft caspase-3, -8, and -9 and RIPK1 and -3 mRNA expression was analysed by quantitative RT-PCR and protein activity measured by immunohistochemistry and luminescent assays 6h after reperfusion. Lactate and lactate dehydrogenase (LDH) levels were analysed from allograft recipient and from hypoxic endothelial cell cultures having treated with activated simvastatin. RESULTS: When compared to without cold ischemia, prolonged 4-hour cold ischemia significantly enhanced intragraft mRNA expression of caspase-3 and -9, and RIPK1 and -3, and elevated protein activity of caspase-9 and RIPK1 in the allografts. Simvastatin pretreatment decreased mRNA expression of caspase-3 and -9, and RIPK1 and -3 and protein activity of caspase-9 and RIPK1 in the allografts. Intragraft caspase-8 mRNA expression remained constant regardless of cold ischemia or simvastatin pretreatment. Simvastatin pretreatment attenuated lactate and LDH levels, both in the allograft recipients and in hypoxic endothelial cell cultures. CONCLUSIONS: The beneficial effects of simvastatin pretreatment in cardiac allograft IRI may involve prevention of apoptosis and necroptosis.

Platelet-derived Growth Factor-B Protects Rat Cardiac Allografts From Ischemia-reperfusion Injury.

BACKGROUND: Microvascular dysfunction and cardiomyocyte injury are hallmarks of ischemia-reperfusion injury (IRI) after heart transplantation. Platelet-derived growth factors (PDGF) have an ambiguous role in this deleterious cascade. On one hand, PDGF may exert vascular stabilizing and antiapoptotic actions through endothelial-pericyte and endothelial-cardiomyocyte crosstalk in the heart; and on the other hand, PDGF signaling mediates neointimal formation and exacerbates chronic rejection in cardiac allografts. The balance between these potentially harmful and beneficial actions determines the final outcome of cardiac allografts. METHODS AND RESULTS: We transplanted cardiac allografts from Dark Agouti rat and Balb mouse donors to fully major histocompatibility complex-mismatched Wistar Furth rat or C57 mouse recipients with a clinically relevant 2-hour cold ischemia and 1-hour warm ischemia. Ex vivo intracoronary delivery of adenovirus-mediated gene transfer of recombinant human PDGF-BB upregulated messenger RNA expression of anti-mesenchymal transition and survival factors BMP-7 and Bcl-2 and preserved capillary density in rat cardiac allografts at day 10. In mouse cardiac allografts PDGF receptor-ß, but not -α intragraft messenger RNA levels were reduced and capillary protein localization was lost during IRI. The PDGF receptor tyrosine kinase inhibitor imatinib mesylate and a monoclonal antibody against PDGF receptor-α enhanced myocardial damage evidenced by serum cardiac troponin T release in the rat and mouse cardiac allografts 6 hours after reperfusion, respectively. Moreover, imatinib mesylate enhanced rat cardiac allograft vasculopathy, cardiac fibrosis, and late allograft loss at day 56. CONCLUSIONS: Our results suggest that PDGF-B signaling may play a role in endothelial and cardiomyocyte recovery from IRI after heart transplantation.