Hypoxia inducible factor 2α promotes tolerogenic macrophage development during cardiac transplantation through transcriptional regulation of colony stimulating factor 1 receptor.

Solid organ transplantation mobilizes myeloid cells, including monocytes and macrophages, which are central protagonists of allograft rejection. However, myeloid cells can also be functionally reprogrammed by perioperative costimulatory blockade to promote a state of transplantation tolerance. Transplantation tolerance holds promise to reduce complications from chronic immunosuppression and promote long-term survival in transplant recipients. We sought to identify different mediators of transplantation tolerance by performing single-cell RNA sequencing of acute rejecting or tolerized cardiac allografts. This led to the unbiased identification of the transcription factor, hypoxia inducible factor (HIF)-2α, in a subset of tolerogenic monocytes. Using flow cytometric analyses and mice with conditional loss or gain of function, we uncovered that myeloid cell expression of HIF-2α was required for costimulatory blockade-induced transplantation tolerance. While HIF-2α was dispensable for mobilization of tolerogenic monocytes, which were sourced in part from the spleen, it promoted the expression of colony stimulating factor 1 receptor (CSF1R). CSF1R mediates monocyte differentiation into tolerogenic macrophages and was found to be a direct transcriptional target of HIF-2α in splenic monocytes. Administration of the HIF stabilizer, roxadustat, within micelles to target myeloid cells, increased HIF-2α in splenic monocytes, which was associated with increased CSF1R expression and enhanced cardiac allograft survival. These data support further exploration of HIF-2α activation in myeloid cells as a therapeutic strategy for transplantation tolerance.

Resolving inflammatory links between myocardial infarction and vascular dementia.

Myocardial infarction is associated with increased risk for vascular dementia. In both myocardial infarction and vascular dementia, there is evidence that elevated inflammatory biomarkers are associated with worsened clinical outcomes. Myocardial infarction leads to a systemic inflammatory response, which may contribute to recruitment or activation of myeloid cells, including monocytes, microglia, and perivascular macrophages, within the central nervous system. However, our understanding of the causative roles for these cells linking cardiac injury to the development and progression of dementia is incomplete. Herein, we provide an overview of inflammatory cellular and molecular links between myocardial infarction and vascular dementia and discuss strategies to resolve inflammation after myocardial infarction to limit neurovascular injury.

CCR2+ monocytes promote white matter injury and cognitive dysfunction after myocardial infarction.

Survivors of myocardial infarction are at increased risk for vascular dementia. Neuroinflammation has been implicated in the pathogenesis of vascular dementia, yet little is known about the cellular and molecular mediators of neuroinflammation after myocardial infarction. Using a mouse model of myocardial infarction coupled with flow cytometric analyses and immunohistochemistry, we discovered increased monocyte abundance in the brain after myocardial infarction, which was associated with increases in brain-resident perivascular macrophages and microglia. Myeloid cell recruitment and activation was also observed in post-mortem brains of humans that died after myocardial infarction. Spatial and single cell transcriptomic profiling of brain-resident myeloid cells after experimental myocardial infarction revealed increased expression of monocyte chemoattractant proteins. In parallel, myocardial infarction increased crosstalk between brain-resident myeloid cells and oligodendrocytes, leading to neuroinflammation, white matter injury, and cognitive dysfunction. Inhibition of monocyte recruitment preserved white matter integrity and cognitive function, linking monocytes to neurodegeneration after myocardial infarction. Together, these preclinical and clinical results demonstrate that monocyte infiltration into the brain after myocardial infarction initiate neuropathological events that lead to vascular dementia.

Monocytes prime autoreactive T cells after myocardial infarction.

In humans, loss of central tolerance for the cardiac self-antigen α-myosin heavy chain (α-MHC) leads to circulation of cardiac autoreactive T cells and renders the heart susceptible to autoimmune attack after acute myocardial infarction (MI). MI triggers profound tissue damage, releasing danger signals and self-antigen by necrotic cardiomyocytes, which lead to recruitment of inflammatory monocytes. We hypothesized that excessive inflammation by monocytes contributes to the initiation of adaptive immune responses to cardiac self-antigen. Using an experimental model of MI in α-MHC-mCherry reporter mice, which specifically express mCherry in cardiomyocytes, we detected α-MHC antigen in myeloid cells in the heart-draining mediastinal lymph node (MLN) 7 days after MI. To test whether monocytes were required for cardiac self-antigen trafficking to the MLN, we blocked monocyte recruitment with a C-C motif chemokine receptor type 2 (CCR2) antagonist or immune modifying nanoparticles (IMP). Blockade of monocyte recruitment reduced α-MHC antigen detection in the MLN after MI. Intramyocardial injection of the model antigen ovalbumin into OT-II transgenic mice demonstrated the requirement for monocytes in antigen trafficking and T-cell activation in the MLN. Finally, in nonobese diabetic mice, which are prone to postinfarction autoimmunity, blockade of monocyte recruitment reduced α-MHC-specific responses leading to improved tissue repair and ventricular function 28 days after MI. Taken together, these data support a role for monocytes in the onset of pathological cardiac autoimmunity following MI and suggest that therapeutic targeting of monocytes may mitigate postinfarction autoimmunity in humans.NEW & NOTEWORTHY Our study newly identifies a role for inflammatory monocytes in priming an autoimmune T-cell response after myocardial infarction. Select inhibition of monocyte recruitment to the infarct prevents trafficking of cardiac self-antigen and activation of cardiac myosin reactive T cells in the heart-draining lymph node. Therapeutic targeting of inflammatory monocytes may limit autoimmune responses to improve cardiac remodeling and preserve left ventricular function after myocardial infarction.

Mechanism of Enhanced MerTK-Dependent Macrophage Efferocytosis by Extracellular Vesicles.

OBJECTIVE: Extracellular vesicles secreted by cardiosphere-derived cells (CDCev) polarize macrophages toward a distinctive phenotype with enhanced phagocytic capacity (MCDCev). These changes underlie cardioprotection by CDCev and by the parent CDCs, notably attenuating the no-reflow phenomenon following myocardial infarction, but the mechanisms are unclear. Here, we tested the hypothesis that MCDCev are especially effective at scavenging debris from dying cells (ie, efferocytosis) to attenuate irreversible damage post-myocardial infarction. Approach and Results: In vitro efferocytosis assays with bone marrow-derived macrophages, and in vivo transgenic rodent models of myocardial infarction, demonstrate enhanced apoptotic cell clearance with MCDCev. CDCev exposure induces sustained MerTK expression in MCDCev through extracellular vesicle transfer of microRNA-26a (via suppression of Adam17); the cardioprotective response is lost in animals deficient in MerTK. Single-cell RNA-sequencing revealed phagocytic pathway activation in MCDCev, with increased expression of complement factor C1qa, a phagocytosis facilitator. CONCLUSIONS: Together, these data demonstrate that extracellular vesicle modulation of MerTK and C1qa expression leads to enhanced macrophage efferocytosis and cardioprotection.

Receptor tyrosine kinase MerTK suppresses an allogenic type I IFN response to promote transplant tolerance.

Recipient infusion of donor apoptotic cells is an emerging strategy for inducing robust transplantation tolerance. Daily clearance of billions of self-apoptotic cells relies on homeostatic engagement of phagocytic receptors, in particular, receptors of the tyrosine kinase family TAM (Tyro3, Axl, and MerTK), to maintain self-tolerance. However, an outstanding question is if allogeneic apoptotic cells trigger the same receptor system for inducing allogeneic tolerance. Here, we employed allogeneic apoptotic splenocytes and discovered that the efferocytic receptor MerTK on recipient phagocytes is a critical mediator for transplantation tolerance induced by this strategy. Our findings indicate that the tolerogenic properties of allogeneic apoptotic splenocytes require MerTK transmission of intracellular signaling to suppress the production of inflammatory cytokine interferon α (IFN-α). We further demonstrate that MerTK is crucial for subsequent expansion of myeloid-derived suppressor cells and for promoting their immunomodulatory function, including maintaining graft-infiltrating CD4+ CD25+ Foxp3+ regulatory T cells. Consequently, recipient MerTK deficiency resulted in failure of tolerance by donor apoptotic cells, and this failure could be effectively rescued by IFN-α receptor blockade. These findings underscore the importance of the efferocytic receptor MerTK in mediating transplantation tolerance by donor apoptotic cells and implicate MerTK agonism as a promising target for promoting transplantation tolerance.

MerTK Cleavage on Resident Cardiac Macrophages Compromises Repair After Myocardial Ischemia Reperfusion Injury.

RATIONALE: Clinical benefits of reperfusion after myocardial infarction are offset by maladaptive innate immune cell function, and therapeutic interventions are lacking. OBJECTIVE: We sought to test the significance of phagocytic clearance by resident and recruited phagocytes after myocardial ischemia reperfusion. METHODS AND RESULTS: In humans, we discovered that clinical reperfusion after myocardial infarction led to significant elevation of the soluble form of MerTK (myeloid-epithelial-reproductive tyrosine kinase; ie, soluble MER), a critical biomarker of compromised phagocytosis by innate macrophages. In reperfused mice, macrophage Mertk deficiency led to decreased cardiac wound debridement, increased infarct size, and depressed cardiac function, newly implicating MerTK in cardiac repair after myocardial ischemia reperfusion. More notably, Mertk(CR) mice, which are resistant to cleavage, showed significantly reduced infarct sizes and improved systolic function. In contrast to other cardiac phagocyte subsets, resident cardiac MHCIILOCCR2- (major histocompatibility complex II/C-C motif chemokine receptor type 2) macrophages expressed higher levels of MerTK and, when exposed to apoptotic cells, secreted proreparative cytokines, including transforming growth factor-ß. Mertk deficiency compromised the accumulation of MHCIILO phagocytes, and this was rescued in Mertk(CR) mice. Interestingly, blockade of CCR2-dependent monocyte infiltration into the heart reduced soluble MER levels post-ischemia reperfusion. CONCLUSIONS: Our data implicate monocyte-induced MerTK cleavage on proreparative MHCIILO cardiac macrophages as a novel contributor and therapeutic target of reperfusion injury.

HIF-2α in Resting Macrophages Tempers Mitochondrial Reactive Oxygen Species To Selectively Repress MARCO-Dependent Phagocytosis.

Hypoxia-inducible factor (HIF)-α isoforms regulate key macrophage (MΦ) functions during ischemic inflammation. HIF-2α drives proinflammatory cytokine production; however, the requirements for HIF-2α during other key MΦ functions, including phagocytosis, are unknown. In contrast to HIF-1α, HIF-2α was not required for hypoxic phagocytic uptake. Surprisingly, basal HIF-2α levels under nonhypoxic conditions were necessary and sufficient to suppress phagocytosis. Screening approaches revealed selective induction of the scavenger receptor MARCO, which was required for enhanced engulfment. Chromatin immunoprecipitation identified the antioxidant NRF2 as being directly responsible for inducing Marco Concordantly, Hif-2α-/- MΦs exhibited reduced antioxidant gene expression, and inhibition of mitochondrial reactive oxygen species suppressed Marco expression and phagocytic uptake. Ex vivo findings were recapitulated in vivo; the enhanced engulfment phenotype resulted in increased bacterial clearance and cytokine suppression. Importantly, natural induction of Hif-2α by IL-4 also suppressed MARCO-dependent phagocytosis. Thus, unlike most characterized prophagocytic regulators, HIF-2α can act as a phagocytic repressor. Interestingly, this occurs in resting MΦs through tempering of steady-state mitochondrial reactive oxygen species. In turn, HIF-2α promotes MΦ quiescence by blocking a MARCO bacterial-response pathway. IL-4 also drives HIF-2α suppression of MARCO, leading to compromised bacterial immunosurveillance in vivo.

Soluble, but not transmembrane, TNF-α is required during influenza infection to limit the magnitude of immune responses and the extent of immunopathology.

TNF-α is a pleotropic cytokine that has both proinflammatory and anti-inflammatory functions during influenza infection. TNF-α is first expressed as a transmembrane protein that is proteolytically processed to release a soluble form. Transmembrane TNF-α (memTNF-α) and soluble TNF-α (solTNF-α) have been shown to exert distinct tissue-protective or tissue-pathologic effects in several disease models. However, the relative contributions of memTNF-α or solTNF-α in regulating pulmonary immunopathology following influenza infection are unclear. Therefore, we performed intranasal influenza infection in mice exclusively expressing noncleavable memTNF-α or lacking TNF-α entirely and examined the outcomes. We found that solTNF-α, but not memTNF-α, was required to limit the size of the immune response and the extent of injury. In the absence of solTNF-α, there was a significant increase in the CD8(+) T cell response, including virus-specific CD8(+) T cells, which was due in part to an increased resistance to activation-induced cell death. We found that solTNF-α mediates these immunoregulatory effects primarily through TNFR1, because mice deficient in TNFR1, but not TNFR2, exhibited dysregulated immune responses and exacerbated injury similar to that observed in mice lacking solTNF-α. We also found that solTNF-α expression was required early during infection to regulate the magnitude of the CD8(+) T cell response, indicating that early inflammatory events are critical for the regulation of the effector phase. Taken together, these findings suggest that processing of memTNF-α to release solTNF-α is a critical event regulating the immune response during influenza infection.

Cardiomyocytes induce macrophage receptor shedding to suppress phagocytosis.

BACKGROUND: Mobilization of the innate immune response to clear and metabolize necrotic and apoptotic cardiomyocytes is a prerequisite to heart repair after cardiac injury. Suboptimal kinetics of dying myocyte clearance leads to secondary necrosis, and in the case of the heart, increased potential for collateral loss of neighboring non-regenerative myocytes. Despite the importance of myocyte phagocytic clearance during heart repair, surprisingly little is known about its underlying cell and molecular biology. OBJECTIVE: To determine if phagocytic receptor MERTK is expressed in human hearts and to elucidate key sequential steps and phagocytosis efficiency of dying adult cardiomyocytes, by macrophages. RESULTS: In infarcted human hearts, expression profiles of the phagocytic receptor MER-tyrosine kinase (MERTK) mimicked that found in experimental ischemic mouse hearts. Electron micrographs of myocardium identified MERTK signal along macrophage phagocytic cups and Mertk-/- macrophages contained reduced digested myocyte debris after myocardial infarction. Ex vivo co-culture of primary macrophages and adult cardiomyocyte apoptotic bodies revealed reduced engulfment relative to resident cardiac fibroblasts. Inefficient clearance was not due to the larger size of myocyte apoptotic bodies, nor were other key steps preceding the formation of phagocytic synapses significantly affected; this included macrophage chemotaxis and direct binding of phagocytes to myocytes. Instead, suppressed phagocytosis was directly associated with myocyte-induced inactivation of MERTK, which was partially rescued by genetic deletion of a MERTK proteolytic susceptibility site. CONCLUSION: Utilizing an ex vivo co-cultivation approach to model key cellular and molecular events found in vivo during infarction, cardiomyocyte phagocytosis was found to be inefficient, in part due to myocyte-induced shedding of macrophage MERTK. These findings warrant future studies to identify other cofactors of macrophage-cardiomyocyte cross-talk that contribute to cardiac pathophysiology.

Inflammatory impact of IFN-γ in CD8+ T cell-mediated lung injury is mediated by both Stat1-dependent and -independent pathways.

Influenza infection results in considerable pulmonary pathology, a significant component of which is mediated by CD8(+) T cell effector functions. To isolate the specific contribution of CD8(+) T cells to lung immunopathology, we utilized a nonviral murine model in which alveolar epithelial cells express an influenza antigen and injury is initiated by adoptive transfer of influenza-specific CD8(+) T cells. We report that IFN-γ production by adoptively transferred influenza-specific CD8(+) T cells is a significant contributor to acute lung injury following influenza antigen recognition, in isolation from its impact on viral clearance. CD8(+) T cell production of IFN-γ enhanced lung epithelial cell expression of chemokines and the subsequent recruitment of inflammatory cells into the airways. Surprisingly, Stat1 deficiency in the adoptive-transfer recipients exacerbated the lung injury that was mediated by the transferred influenza-specific CD8(+) T cells but was still dependent on IFN-γ production by these cells. Loss of Stat1 resulted in sustained activation of Stat3 signaling, dysregulated chemokine expression, and increased infiltration of the airways by inflammatory cells. Taken together, these data identify important roles for IFN-γ signaling and Stat1-independent IFN-γ signaling in regulating CD8(+) T cell-mediated acute lung injury. This is the first study to demonstrate an anti-inflammatory effect of Stat1 on CD8(+) T cell-mediated lung immunopathology without the complication of differences in viral load.

Phagocyte-myocyte interactions and consequences during hypoxic wound healing.

Myocardial infarction (MI), secondary to atherosclerotic plaque rupture and occlusive thrombi, triggers acute margination of inflammatory neutrophils and monocyte phagocyte subsets to the damaged heart, the latter of which may give rise briefly to differentiated macrophage-like or dendritic-like cells. Within the injured myocardium, a primary function of these phagocytic cells is to remove damaged extracellular matrix, necrotic and apoptotic cardiac cells, as well as immune cells that turn over. Recognition of dying cellular targets by phagocytes triggers intracellular signaling, particularly in macrophages, wherein cytokines and lipid mediators are generated to promote inflammation resolution, fibrotic scarring, angiogenesis, and compensatory organ remodeling. These actions cooperate in an effort to preserve myocardial contractility and prevent heart failure. Immune cell function is modulated by local tissue factors that include secreted protease activity, oxidative stress during clinical reperfusion, and hypoxia. Importantly, experimental evidence suggests that monocyte function and phagocytosis efficiency is compromised in the setting of MI risk factors, including hyperlipidemia and ageing, however underlying mechanisms remain unclear. Herein we review seminal phagocyte and cardiac molecular factors that lead to, and culminate in, the recognition and removal of dying injured myocardium, the effects of hypoxia, and their relationship to cardiac infarct size and heart healing.

The BMAL1/HIF2A heterodimer modulates circadian variations of myocardial injury.

Acute myocardial infarction stands as a prominent cause of morbidity and mortality worldwide1-6. Clinical studies have demonstrated that the severity of cardiac injury following myocardial infarction exhibits a circadian pattern, with larger infarct sizes and poorer outcomes in patients experiencing morning onset myocardial infarctions7-14. However, the molecular mechanisms that govern circadian variations of myocardial injury remain unclear. Here, we show that BMAL114-20, a core circadian transcription factor, orchestrates diurnal variability in myocardial injury. Unexpectedly, BMAL1 modulates circadian-dependent cardiac injury by forming a transcriptionally active heterodimer with a non-canonical partner, hypoxia-inducible factor 2 alpha (HIF2A)6,21-23, in a diurnal manner. Substantiating this finding, we determined the cryo-EM structure of the BMAL1/HIF2A/DNA complex, revealing a previously unknown capacity for structural rearrangement within BMAL1, which enables the crosstalk between circadian rhythms and hypoxia signaling. Furthermore, we identified amphiregulin (AREG) as a rhythmic transcriptional target of the BMAL1/HIF2A heterodimer, critical for regulating circadian variations of myocardial injury. Finally, pharmacologically targeting the BMAL1/HIF2A-AREG pathway provides effective cardioprotection, with maximum efficacy when aligned with the pathway's circadian trough. Our findings not only uncover a novel mechanism governing the circadian variations of myocardial injury but also pave the way for innovative circadian-based treatment strategies, potentially shifting current treatment paradigms for myocardial infarction.

Immunometabolism at the Heart of Cardiovascular Disease.

Immune cell function among the myocardium, now more than ever, is appreciated to regulate cardiac function and pathophysiology. This is the case for both innate immunity, which includes neutrophils, monocytes, dendritic cells, and macrophages, as well as adaptive immunity, which includes T cells and B cells. This function is fueled by cell-intrinsic shifts in metabolism, such as glycolysis and oxidative phosphorylation, as well as metabolite availability, which originates from the surrounding extracellular milieu and varies during ischemia and metabolic syndrome. Immune cell crosstalk with cardiac parenchymal cells, such as cardiomyocytes and fibroblasts, is also regulated by complex cellular metabolic circuits. Although our understanding of immunometabolism has advanced rapidly over the past decade, in part through valuable insights made in cultured cells, there remains much to learn about contributions of in vivo immunometabolism and directly within the myocardium. Insight into such fundamental cell and molecular mechanisms holds potential to inform interventions that shift the balance of immunometabolism from maladaptive to cardioprotective and potentially even regenerative. Herein, we review our current working understanding of immunometabolism, specifically in the settings of sterile ischemic cardiac injury or cardiometabolic disease, both of which contribute to the onset of heart failure. We also discuss current gaps in knowledge in this context and therapeutic implications.

Single-cell profiling reveals inflammatory polarization of human carotid versus femoral plaque leukocytes.

Femoral atherosclerotic plaques are less inflammatory than carotid plaques histologically, but limited cell-level data exist regarding comparative immune landscapes and polarization at these sites. We investigated intraplaque leukocyte phenotypes and transcriptional polarization in 49 patients undergoing femoral (n = 23) or carotid (n = 26) endarterectomy using single-cell RNA-Seq (scRNA-Seq; n = 13), flow cytometry (n = 24), and IHC (n = 12). Comparative scRNA-Seq of CD45+-selected leukocytes from femoral (n = 9; 35,265 cells) and carotid (n = 4; 30,655 cells) plaque revealed distinct transcriptional profiles. Inflammatory foam cell-like macrophages and monocytes comprised higher proportions of myeloid cells in carotid plaques, whereas noninflammatory foam cell-like macrophages and LYVE1-overexpressing macrophages comprised higher proportions of myeloid cells in femoral plaque (P < 0.001 for all). A significant comparative excess of CCR2+ macrophages in carotid versus plaque was observed by flow cytometry in a separate validation cohort. B cells were more prevalent and exhibited a comparatively antiinflammatory profile in femoral plaque, whereas cytotoxic CD8+ T cells were more prevalent in carotid plaque. In conclusion, human femoral plaques exhibit distinct macrophage phenotypic and transcriptional profiles as well as diminished CD8+ T cell populations compared with human carotid plaques.

Macrophage-produced VEGFC is induced by efferocytosis to ameliorate cardiac injury and inflammation.

Clearance of dying cells by efferocytosis is necessary for cardiac repair after myocardial infarction (MI). Recent reports have suggested a protective role for vascular endothelial growth factor C (VEGFC) during acute cardiac lymphangiogenesis after MI. Here, we report that defective efferocytosis by macrophages after experimental MI led to a reduction in cardiac lymphangiogenesis and Vegfc expression. Cell-intrinsic evidence for efferocytic induction of Vegfc was revealed after adding apoptotic cells to cultured primary macrophages, which subsequently triggered Vegfc transcription and VEGFC secretion. Similarly, cardiac macrophages elevated Vegfc expression levels after MI, and mice deficient for myeloid Vegfc exhibited impaired ventricular contractility, adverse tissue remodeling, and reduced lymphangiogenesis. These results were observed in mouse models of permanent coronary occlusion and clinically relevant ischemia and reperfusion. Interestingly, myeloid Vegfc deficiency also led to increases in acute infarct size, prior to the amplitude of the acute cardiac lymphangiogenesis response. RNA-Seq and cardiac flow cytometry revealed that myeloid Vegfc deficiency was also characterized by a defective inflammatory response, and macrophage-produced VEGFC was directly effective at suppressing proinflammatory macrophage activation. Taken together, our findings indicate that cardiac macrophages promote healing through the promotion of myocardial lymphangiogenesis and the suppression of inflammatory cytokines.

Macrophage AXL receptor tyrosine kinase inflames the heart after reperfused myocardial infarction.

Tyro3, AXL, and MerTK (TAM) receptors are activated in macrophages in response to tissue injury and as such have been proposed as therapeutic targets to promote inflammation resolution during sterile wound healing, including myocardial infarction. Although the role of MerTK in cardioprotection is well characterized, the unique role of the other structurally similar TAMs, and particularly AXL, in clinically relevant models of myocardial ischemia/reperfusion infarction (IRI) is comparatively unknown. Utilizing complementary approaches, validated by flow cytometric analysis of human and murine macrophage subsets and conditional genetic loss and gain of function, we uncover a maladaptive role for myeloid AXL during IRI in the heart. Cross signaling between AXL and TLR4 in cardiac macrophages directed a switch to glycolytic metabolism and secretion of proinflammatory IL-1ß, leading to increased intramyocardial inflammation, adverse ventricular remodeling, and impaired contractile function. AXL functioned independently of cardioprotective MerTK to reduce the efficacy of cardiac repair, but like MerTK, was proteolytically cleaved. Administration of a selective small molecule AXL inhibitor alone improved cardiac healing, which was further enhanced in combination with blockade of MerTK cleavage. These data support further exploration of macrophage TAM receptors as therapeutic targets for myocardial infarction.

Comparative Risk of Incident Coronary Heart Disease Across Chronic Inflammatory Diseases.

Background: Chronic inflammatory diseases (CIDs) are considered risk enhancing factors for coronary heart disease (CHD). However, sparse data exist regarding relative CHD risks across CIDs. Objective: Determine relative differences in CHD risk across multiple CIDs: psoriasis, rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), human immunodeficiency virus (HIV), systemic sclerosis (SSc), and inflammatory bowel disease (IBD). Methods: The cohort included patients with CIDs and controls without CID in an urban medical system from 2000 to 2019. Patients with CIDs were frequency-matched with non-CID controls on demographics, hypertension, and diabetes. CHD was defined as myocardial infarction (MI), ischemic heart disease, and/or coronary revascularization based on validated administrative codes. Multivariable-adjusted Cox models were used to determine the risk of incident CHD and MI for each CID relative to non-CID controls. In secondary analyses, we compared CHD risk by disease severity within each CID. Results: Of 17,049 patients included for analysis, 619 had incident CHD (202 MI) over an average of 4.4 years of follow-up. The multivariable-adjusted risk of CHD was significantly higher for SLE [hazard ratio (HR) 1.9, 95% confidence interval (CI) 1.2, 3.2] and SSc (HR 2.1, 95% CI 1.2, 3.9). Patients with SLE also had a significantly higher risk of MI (HR 3.6, 95% CI 1.9, 6.8). When CIDs were categorized by markers of disease severity (C-reactive protein for all CIDs except HIV, for which CD4 T cell count was used), greater disease severity was associated with higher CHD risk across CIDs. Conclusions: Patients with SLE and SSc have a higher risk of CHD. CHD risk with HIV, RA, psoriasis, and IBD may only be elevated in those with greater disease severity. Clinicians should personalize CHD risk and treatment based on type and severity of CID.

Hypoxia-inducible factors individually facilitate inflammatory myeloid metabolism and inefficient cardiac repair.

Hypoxia-inducible factors (HIFs) are activated in parenchymal cells in response to low oxygen and as such have been proposed as therapeutic targets during hypoxic insult, including myocardial infarction (MI). HIFs are also activated within macrophages, which orchestrate the tissue repair response. Although isoform-specific therapeutics are in development for cardiac ischemic injury, surprisingly, the unique role of myeloid HIFs, and particularly HIF-2α, is unknown. Using a murine model of myocardial infarction and mice with conditional genetic loss and gain of function, we uncovered unique proinflammatory roles for myeloid cell expression of HIF-1α and HIF-2α during MI. We found that HIF-2α suppressed anti-inflammatory macrophage mitochondrial metabolism, while HIF-1α promoted cleavage of cardioprotective MerTK through glycolytic reprogramming of macrophages. Unexpectedly, combinatorial loss of both myeloid HIF-1α and HIF-2α was catastrophic and led to macrophage necroptosis, impaired fibrogenesis, and cardiac rupture. These findings support a strategy for selective inhibition of macrophage HIF isoforms and promotion of anti-inflammatory mitochondrial metabolism during ischemic tissue repair.

Bone marrow-derived AXL tyrosine kinase promotes mitogenic crosstalk and cardiac allograft vasculopathy.

Cardiac Allograft Vasculopathy (CAV) is a leading contributor to late transplant rejection. Although implicated, the mechanisms by which bone marrow-derived cells promote CAV remain unclear. Emerging evidence implicates the cell surface receptor tyrosine kinase AXL to be elevated in rejecting human allografts. AXL protein is found on multiple cell types, including bone marrow-derived myeloid cells. The causal role of AXL from this compartment and during transplant is largely unknown. This is important because AXL is a key regulator of myeloid inflammation. Utilizing experimental chimeras deficient in the bone marrow-derived Axl gene, we report that Axl antagonizes cardiac allograft survival and promotes CAV. Flow cytometric and histologic analyses of Axl-deficient transplant recipients revealed reductions in both allograft immune cell accumulation and vascular intimal thickness. Co-culture experiments designed to identify cell-intrinsic functions of Axl uncovered complementary cell-proliferative pathways by which Axl promotes CAV-associated inflammation. Specifically, Axl-deficient myeloid cells were less efficient at increasing the replication of both antigen-specific T cells and vascular smooth muscle cells (VSMCs), the latter a key hallmark of CAV. For the latter, we discovered that Axl-was required to amass the VSMC mitogen Platelet-Derived Growth Factor. Taken together, our studies reveal a new role for myeloid Axl in the progression of CAV and mitogenic crosstalk. Inhibition of AXL-protein, in combination with current standards of care, is a candidate strategy to prolong cardiac allograft survival.