Nrf2 attenuates the innate immune response after experimental myocardial infarction

Objectives: We aimed to investigate the contribution of the transcription factor nuclear factor erythroid-derived 2-like 2 (Nrf2) to the inflammatory response after experimental myocardial infarction (MI. Background: There is compelling evidence implicating dysregulated inflammation in the mechanism of ventricular remodeling and heart failure (HF) after MI. The transcription factor Nrf2 (encoded by Nfe2l2) is a promising target in this context. It impedes transcriptional upregulation of pro-inflammatory cytokines and is anti-inflammatory in various murine models. Methods: We subjected Nrf2-/- mice and wild type (WT) controls to permanent left coronary artery (LCA) ligation. The inflammatory response was investigated with fluorescence-activated cell sorting (FACS) analysis of peripheral blood and heart cell suspensions, together with qRT-PCR of infarcted tissue for chemokines and their receptors. To investigate whether Nrf2-mediated transcription is a dedicated function of leukocytes, we interrogated publicly available RNA-sequencing (RNA-seq) data from mouse hearts after permanent LCA ligation for Nrf2-regulated gene (NRG) expression. Results: FACS analysis demonstrated a profoundly inflamed phenotype in the hearts of global Nrf2-/- mice as compared to WT mice after MI. Moreover, infarcted tissue from Nrf2-/- mice displayed higher expression of inflammatory cytokines, chemokines, and their receptors, including IL6, Ccl2, and Cxcr4. RNA-seq analysis showed upregulated NRG expression in WT mice after MI compared to untreated mice, which was significantly higher in bioinformatically isolated CCR2+ cells. Conclusions: Taken together, the results suggest that Nrf2 signalling in leukocytes, and possibly CCR2+ monocyte-derived cardiac resident macrophages, may be potential targets to prevent post-MI ventricular remodeling.

Effects of Bone Marrow Stromal Cells (BMSCs) on Behavior, Infarct Size and HIF-1α Expression in Stroke Rats

This study aims to assess BMSCs’ effect on the behavior, infarct size and HIF-1α expression in stroke rats. Rats were separated into sham group, CVA group and BMSCs group with 10 rats in each group followed by analysis of neuroethology scores, brain tissue pathology and infarct size, and HIF-1α level in brain tissues. No difference of neurological scores was found between CVA group and BMSCs group after 3 hours (P > 0.05). After BMSCs transplantation, the nerve score was significantly reduced (P < 0.05) and cognitive function was significantly improved compared to CVA group. Compared with sham rats, CAV rats had a larger area of infarction and the infarcted tissue cells showed degeneration or necrosis with reduced cell number and obvious edema, which were all improved in BMSCs group. CVA group showed a larger area of infarct tissue (P < 0.05), which was reduced in BMSCs group (P < 0.05). Compared with sham group, CVA group showed significantly upregulated HIF-1α level (P < 0.05) which was reduced in BMSCs group (P < 0.05). BMSCs has a certain repair effect on the ethology of stroke rats possibly via inhibition of HIF-1α level in cerebral infarction and brain tissue.

Longitudinal hippocampal volumetric changes in mice following brain infarction

Hippocampal atrophy is increasingly described in many neurodegenerative syndromes in humans, including stroke and vascular cognitive impairment. However, the progression of brain volume changes after stroke in rodent models is poorly characterized. We aimed to monitor hippocampal atrophy occurring in mice up to 48-weeks post-stroke. Male C57BL/6J mice were subjected to an intraluminal filament-induced middle cerebral artery occlusion (MCAO). At baseline, 3-days, and 1-, 4-, 12-, 24-, 36- and 48-weeks post-surgery, we measured sensorimotor behavior and hippocampal volumes from T2-weighted MRI scans. Hippocampal volume—both ipsilateral and contralateral—increased over the life-span of sham-operated mice. In MCAO-subjected mice, different trajectories of ipsilateral hippocampal volume change were observed dependent on whether the hippocampus contained direct infarction, with a decrease in directly infarcted tissue and an increase in non-infarcted tissue. To further investigate these volume changes, neuronal and glial cell densities were assessed in histological brain sections from the subset of MCAO mice lacking hippocampal infarction. Our findings demonstrate previously uncharacterized changes in hippocampal volume and potentially brain parenchymal cell density up to 48-weeks in both sham- and MCAO-operated mice.

Predicting Infarct Core From Computed Tomography Perfusion in Acute Ischemia With Machine Learning

Background and Purpose: The ISLES challenge (Ischemic Stroke Lesion Segmentation) enables globally diverse teams to compete to develop advanced tools for stroke lesion analysis with machine learning. Detection of irreversibly damaged tissue on computed tomography perfusion (CTP) is often necessary to determine eligibility for late-time-window thrombectomy. Therefore, the aim of ISLES-2018 was to segment infarcted tissue on CTP based on diffusion-weighted imaging as a reference standard. Methods: The data, from 4 centers, consisted of 103 cases of acute anterior circulation large artery occlusion stroke who underwent diffusion-weighted imaging rapidly after CTP. Diffusion-weighted imaging lesion segmentation was performed manually and acted as a reference standard. The data were separated into 63 cases for training and 40 for testing, upon which quality metrics (dice score coefficient, Hausdorff distance, absolute lesion volume difference, etc) were computed to rank methods based on their overall performance. Results: Twenty-four different teams participated in the challenge. Median time to CTP was 185 minutes (interquartile range, 180–238), the time between CTP and magnetic resonance imaging was 36 minutes (interquartile range, 25–79), and the median infarct lesion size was 15.2 mL (interquartile range, 5.7–45). The best performance for Dice score coefficient and absolute volume difference were 0.51 and 10.1 mL, respectively, from different teams. Based on the ranking criteria, the top team’s algorithm demonstrated for average Dice score coefficient and average absolute volume difference 0.51 and 10.2 mL, respectively, outperforming the conventional threshold-based method (dice score coefficient, 0.3; volume difference, 15.3). Diverse algorithms were used, almost all based on deep learning, with top-ranked approaches making use of the raw perfusion data as well as methods to synthetically generate complementary information to boost prediction performance. Conclusions: Machine learning methods may predict infarcted tissue from CTP with improved accuracy compared with threshold-based methods used in clinical routine. This dataset will remain public and can be used to test improvement in algorithms over time.

Postinfarction posterobasal ventricular septal defect closure with a triple-layer patch

We present the case of a 65-year-old patient who developed a large posterobasal ventricular septal defect resulting from an extensive acute myocardial infarction involving the inferior and basal septum and wall. We repaired the interventricular lesion by verticalizing the cardiac apex to perform a left posterobasal ventriculotomy. We removed a great part of the residual infarcted tissue, leaving the residual scar in place. Our technique first involved creating a double-layer patch comprising heterologous pericardium and a non-collagen-impregnated Sauvage Dacron patch, fixed with single pledgeted U-stitches from the right side of the anterior septum; then we applied a third layer of heterologous pericardium on the left side of the septum in order to have only a pericardial surface in contact with blood on both ventricular sides. A running suture was used to complete the procedure from the middle to the posterior rim of the ventricular septal defect.

Effect of infarct stiffness on non-infarcted left and right ventricular tissue remodeling: a computational study based on myocardial mechano-transduction

Background/Introduction The mechanical properties of infarcted myocardium are important determinants of cardiac pump function and risk of developing heart failure following myocardial infarction (MI). Purpose To better understand the effects of infarct stiffness on compensatory hypertrophy and dilation of non-infarcted tissue in the left (LV) and right ventricle (RV), by using a computational model. Methods The CircAdapt computational model of the human heart and circulation was applied to simulate an acute MI involving 20% of LV wall mass. The simulation was validated using previously published experimental data. Subsequently, two degrees of increased infarct stiffness were simulated. In all three simulations, a model of structural myocardial adaptation of the non-infarcted tissue was applied, based on sensing of mechanical loading of myocytes and extracellular matrix (ECM). Results Mild and severe stiffening of the infarct reduced the increase of LV end-diastolic volume (EDV) from +23 mL to +17 mL and +16 mL, respectively, and the increase of LV non-infarcted tissue mass from +31% to +21% and +18%. RV EDV decreased after adaptation, and mild and severe infarct stiffening reduced the decrease of RV EDV from −21 mL to −12 mL and −10 mL, respectively. Increase of RV tissue mass was reduced from +13% to +8% and +7% with mild and severe infarct stiffening. In the LV, reduced dilation and hypertrophy were driven mainly by a reduction of maximum stress in the ECM and a higher stress between the myocytes and ECM following infarct stiffening. The decreased RV hypertrophy, but not EDV reduction, was caused by a reduction of maximum RV ECM stress and maximum RV active myofiber stress. Conclusions Model simulations predicted that a stiffened LV infarct reduces both LV and RV non-infarcted tissue hypertrophy as well as LV dilation. In LV remodeling, maximum ECM stress and stress between myocyte and ECM played a more prominent role than in RV remodeling, while maximum active stress was more important in the RV. Overview of all model simulations Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): This work was funded by the Netherlands Organisation for Scientific Research and the Dutch Heart Foundation.

Infarct-Like Spindle Cell Carcinoma of the Lung: Clinicopathologic, Immunohistochemical, and Molecular Analysis of 4 Cases

Pulmonary spindle cell carcinoma is a rare and aggressive malignancy that often mimics benign conditions. We report 4 cases that simulate a pulmonary infarction, 2 of which were misdiagnosed. Patients were 3 men and 1 woman, smokers, presenting chest pain. All cases appeared as pleural-based, solitary, and rounded nodules. Patients underwent wedge resections followed by adjuvant chemotherapy (3/4) but died of disease. At histology, lesions consisted of widely necrotic nodules surrounded by organizing fibrosis and pleuritis. Examination and immunostains with pan-cytokeratins and epithelial membrane antigen (EMA) revealed atypical spindle cells encircling necrotic tissue and involving the vascular wall. Positive staining with PD-L1 was noted. Molecular analysis showed KRAS (2/4) and TP53 (1/4) mutations, whereas EGFR, ALK, and ROS1 alterations were not detected. Although in a limited series, these cases further evidence the treacherous appearance of spindle cell carcinomas and the need for careful attention when examining pulmonary infarcted tissue, thus requiring extensive sampling, meticulous examination of vascular structures, and immunostaining with cytokeratins.

In vitro models of human cardiac fibrotic tissue on ‘bioartificial’ scaffolds

Cardiac infarction is a global burden worldwide that leads to fibrotic and not contractile myocardial tissue. In this work, in vitro models of infarcted tissue were developed as tools to test novel therapies for cardiac regeneration in the future. Human cardiac fibroblasts were cultured on scaffolds, with different compositions and architectures, as to mimic structural and chemical features of infarcted cardiac tissue. Early findings from in vitro cell tests were reported, showing an enhancement of cell attachment and proliferation in the case of “bioartificial” scaffolds, i.e. scaffolds based on a synthetic and a bioactive polymer.

Abstract TP266: Ischemic Stroke Induces Striking Heterogeneity in the Inflammatory Leukocyte Infiltrate Across the Core and Penumbra

Background: Current therapies for ischemic stroke focus on reperfusion but do not address the acute inflammatory response. Previous clinical trials aimed at modulating the inflammatory milieu by disrupting leukocyte infiltration failed to show clinical efficacy. One possible explanation for this unexpected shortcoming is an incomplete understanding of the precise spatio-temporal underpinnings of leukocyte extravasation and infiltration. Methods: Here we describe the evolution of the inflammatory response in a mouse transient middle cerebral artery occlusion (tMCAO) stroke model at 0, 1, 2 and 3 days post reperfusion. We used wide field and confocal immunofluorescence microscopy to examine the exact nature and location of the invading myelomonocytic populations, with close examination of the leukocyte position with regard to the brain vasculature and the perivascular space. Results: Our findings suggest that the vast majority of infiltrating myelomonocytic cells escape the perivascular compartment and enter the parenchyma. Interestingly, leukocyte extravasation and accumulation in the subcortex occurred over several days. Dramatic heterogeneity in the inflammatory infiltrate was observed across the infarcted tissue, but also in the surrounding penumbra and adjacent cortical surface. In addition, triphenyl tetrazolium chloride staining, a common indicator for infarcted tissue, did not correlate with the amount or location of leukocyte infiltration. Conclusion: Taken together our findings demonstrate that the infiltration of leukocytes dynamically evolves over several days following reperfusion. Furthermore, leukocytes infiltrate in a heterogeneous pattern that does not correlate well with traditional markers of cellular dysfunction. A better understating of the precise spatio-temporal infiltration of inflammatory cells could help inform the next generation of therapeutic interventions.

Abstract WP71: Gradient of Tissue Injury on Diffusion Tensor Imaging After Stroke: Rethinking the Infarct vs Non-infarcted Dichotomy

Objective: To evaluate the degree of variability in microstructural injury within and adjacent to regions identified as infarcted tissue using Diffusion Tensor Imaging (DTI). Methods: Perfusion CT was performed in 18 patients within 12 hours of ischemic stroke onset followed by Fluid-attenuated Inversion recovery (FLAIR) and DTI one month after stroke. Four regions of interest (ROIs) corresponding to the severity of hypoperfusion on CT perfusion within and beyond the radiological infarct lesion defined on FLAIR were segmented. Fractional anisotropy (FA) and mean diffusivity (MD) were quantified for each ROI and compared to a mirror homologue in the contralateral hemisphere. Ipsilateral to contralateral FA and MD ratios were compared across ROIs. Results: Lower FA and higher MD values were observed within both the infarct lesion and the peri-infarct tissue compared with their homologous contralateral brain regions (all comparisons p≤0.01). No difference was observed in FA and MD between remote non-hypoperfused tissue and its contralateral homologous region (FA p=0.42, MD p≥0.99). The magnitude of asymmetry (ipsilateral/contralateral ratios) of FA and MD was greater with increasing severity of hypoperfusion in a dose-response pattern. Asymmetry greatest in the area of infarction with severe hypoperfusion, followed by infarction with moderate hypoperfusion, the peri-infarct hypoperfused tissue and lastly the remote non-hypoperfused normal tissue (median on clustered quantile regression p≤0.01). Conclusion: A gradient of microstructural injury corresponding to the severity of ischemic insult is present within and beyond conventionally-defined infarct boundaries. The traditional dichotomized notion of infarcted versus non-infarcted tissue widely adopted in clinical research and in practice warrants re-examination.