Cardio-Pulmonary Institute

Clinical and translational research has played a major role in advancing our understanding of pulmonary hypertension (PH), including pulmonary arterial hypertension and other forms of PH with severe vascular remodelling ( e.g. chronic thromboembolic PH and pulmonary veno-occlusive disease). However, PH remains an incurable condition with a high mortality rate, underscoring the need for a better transfer of novel scientific knowledge into healthcare interventions. Herein, we review recent findings in pathology (with the questioning of the strict morphological categorisation of various forms of PH into pre- or post-capillary involvement of pulmonary vessels) and cellular mechanisms contributing to the onset and progression of pulmonary vascular remodelling associated with various forms of PH. We also discuss ways to improve management and to support and optimise drug development in this research field.

Acute myocardial infarction (AMI) and the heart failure that often follows, are major causes of death and disability worldwide. As such, new therapies are required to limit myocardial infarct (MI) size, prevent adverse left ventricular (LV) remodeling, and reduce the onset of heart failure following AMI. The inflammatory response to AMI, plays a critical role in determining MI size, and a persistent pro-inflammatory reaction can contribute to adverse post-MI LV remodeling, making inflammation an important therapeutic target for improving outcomes following AMI. In this article, we provide an overview of the multiple players (and their dynamic roles) involved in the complex inflammatory response to AMI and subsequent LV remodeling, and highlight future opportunities for targeting inflammation as a therapeutic strategy for limiting MI size, preventing adverse LV remodeling, and reducing heart failure in AMI patients.

Angiotensin-converting enzyme 2 (ACE2) and accessory proteases (TMPRSS2 and CTSL) are needed for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) cellular entry, and their expression may shed light on viral tropism and impact across the body. We assessed the cell-type-specific expression of ACE2, TMPRSS2 and CTSL across 107 single-cell RNA-sequencing studies from different tissues. ACE2, TMPRSS2 and CTSL are coexpressed in specific subsets of respiratory epithelial cells in the nasal passages, airways and alveoli, and in cells from other organs associated with coronavirus disease 2019 (COVID-19) transmission or pathology. We performed a meta-analysis of 31 lung single-cell RNA-sequencing studies with 1,320,896 cells from 377 nasal, airway and lung parenchyma samples from 228 individuals. This revealed cell-type-specific associations of age, sex and smoking with expression levels of ACE2, TMPRSS2 and CTSL. Expression of entry factors increased with age and in males, including in airway secretory cells and alveolar type 2 cells. Expression programs shared by ACE2+TMPRSS2+ cells in nasal, lung and gut tissues included genes that may mediate viral entry, key immune functions and epithelial–macrophage cross-talk, such as genes involved in the interleukin-6, interleukin-1, tumor necrosis factor and complement pathways. Cell-type-specific expression patterns may contribute to the pathogenesis of COVID-19, and our work highlights putative molecular pathways for therapeutic intervention. An integrated analysis of over 100 single-cell and single-nucleus transcriptomics studies illustrates severe acute respiratory syndrome coronavirus 2 viral entry gene coexpression patterns across different human tissues, and shows association of age, smoking status and sex with viral entry gene expression in respiratory cell populations.

BACKGROUND: Pulmonary hypertension (PH) is a severe, progressive disease. Although 5 PH subgroups are recognized, reports on survival have focused mainly on pulmonary arterial hypertension (PAH). METHODS: Long-term transplant-free survival and its determinants were investigated in patients with PH (diagnosed by right heart catheterization) within a prospective registry at a single referral center in Giessen, Germany. RESULTS: In total, 2,067 patients were enrolled (PAH, 685 patients [33.1%]; pulmonary venous hypertension, 307 patients [14.9%]; PH due to lung diseases (LD-PH), 546 patients [26.4%; mainly interstitial lung disease and chronic obstructive pulmonary disease]; chronic thromboembolic PH, 459 patients [22.2%]; PH owing to miscellaneous/unknown causes, 70 patients [3.4%]). Median follow-up was 37 months. Differences in transplant-free survival between etiologic groups were highly significant (p < 0.001), with 1-, 3- and 5-year survival rates of 88.2%, 72.2% and 59.4%, respectively, for those with PAH compared with 79.5%, 52.7% and 38.1%, respectively, for patients with LD-PH. Patients' age, gender and 6-minute walk distance (6MWD), but not New York Heart Association (NYHA) functional class, associated significantly with survival across all PH subtypes in multivariate Cox regression analyses. CONCLUSIONS: This is the largest single-center PH cohort described so far. Some parameters used in clinical practice do not independently predict survival. Age, gender and 6MWD outperformed NYHA functional class in predicting survival across all etiologic groups.

Endothelial cells play a critical role in the adaptation of tissues to injury. Tissue ischemia induced by infarction leads to profound changes in endothelial cell functions and can induce transition to a mesenchymal state. Here we explore the kinetics and individual cellular responses of endothelial cells after myocardial infarction by using single cell RNA sequencing. This study demonstrates a time dependent switch in endothelial cell proliferation and inflammation associated with transient changes in metabolic gene signatures. Trajectory analysis reveals that the majority of endothelial cells 3 to 7 days after myocardial infarction acquire a transient state, characterized by mesenchymal gene expression, which returns to baseline 14 days after injury. Lineage tracing, using the Cdh5-CreERT2;mT/mG mice followed by single cell RNA sequencing, confirms the transient mesenchymal transition and reveals additional hypoxic and inflammatory signatures of endothelial cells during early and late states after injury. These data suggest that endothelial cells undergo a transient mes-enchymal activation concomitant with a metabolic adaptation within the first days after myocardial infarction but do not acquire a long-term mesenchymal fate. This mesenchymal activation may facilitate endothelial cell migration and clonal expansion to regenerate the vascular network.

The blood-brain barrier (BBB) is a dynamic platform for exchange of substances between the blood and the brain parenchyma, and it is an essential functional gatekeeper for the central nervous system (CNS). While it is widely recognized that BBB disruption is a hallmark of several neurovascular pathologies, an aspect of the BBB that has received somewhat less attention is the dynamic modulation of BBB tightness to maintain brain homeostasis in response to extrinsic environmental factors and physiological changes. In this review, we summarize how BBB integrity adjusts in critical stages along the life span, as well as how BBB permeability can be altered by common stressors derived from nutritional habits, environmental factors and psychological stress.

Idiopathic pulmonary fibrosis (IPF) is a fatal disease in which the intricate alveolar network of the lung is progressively replaced by fibrotic scars. Myofibroblasts are the effector cells that excessively deposit extracellular matrix proteins thus compromising lung structure and function. Emerging literature suggests a correlation between fibrosis and metabolic alterations in IPF. In this study, we show that the first-line antidiabetic drug metformin exerts potent antifibrotic effects in the lung by modulating metabolic pathways, inhibiting TGFβ1 action, suppressing collagen formation, activating PPARγ signaling and inducing lipogenic differentiation in lung fibroblasts derived from IPF patients. Using genetic lineage tracing in a murine model of lung fibrosis, we show that metformin alters the fate of myofibroblasts and accelerates fibrosis resolution by inducing myofibroblast-to-lipofibroblast transdifferentiation. Detailed pathway analysis revealed a two-arm mechanism by which metformin accelerates fibrosis resolution. Our data report an antifibrotic role for metformin in the lung, thus warranting further therapeutic evaluation.

AIMS: Coronavirus disease 2019 is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and has emerged as a global pandemic. SARS-CoV-2 infection can lead to elevated markers of cardiac injury associated with higher risk of mortality. It is unclear whether cardiac injury is caused by direct infection of cardiomyocytes or is mainly secondary to lung injury and inflammation. Here, we investigate whether cardiomyocytes are permissive for SARS-CoV-2 infection. METHODS AND RESULTS: Two strains of SARS-CoV-2 infected human induced pluripotent stem cell-derived cardiomyocytes as demonstrated by detection of intracellular double-stranded viral RNA and viral spike glycoprotein expression. Increasing concentrations of viral RNA are detected in supernatants of infected cardiomyocytes, which induced infections in Caco-2 cell lines, documenting productive infections. SARS-CoV-2 infection and induced cytotoxic and proapoptotic effects associated with it abolished cardiomyocyte beating. RNA sequencing confirmed a transcriptional response to viral infection as demonstrated by the up-regulation of genes associated with pathways related to viral response and interferon signalling, apoptosis, and reactive oxygen stress. SARS-CoV-2 infection and cardiotoxicity was confirmed in a 3D cardiosphere tissue model. Importantly, viral spike protein and viral particles were detected in living human heart slices after infection with SARS-CoV-2. Coronavirus particles were further observed in cardiomyocytes of a patient with coronavirus disease 2019. Infection of induced pluripotent stem cell-derived cardiomyocytes was dependent on cathepsins and angiotensin-converting enzyme 2, and was blocked by remdesivir. CONCLUSION: This study demonstrates that SARS-CoV-2 infects cardiomyocytes in vitro in an angiotensin-converting enzyme 2- and cathepsin-dependent manner. SARS-CoV-2 infection of cardiomyocytes is inhibited by the antiviral drug remdesivir.

BACKGROUND: Since 2009, IPF patients across Europe are recruited into the eurIPFreg, providing epidemiological data and biomaterials for translational research. METHODS: The registry data are based on patient and physician baseline and follow-up questionnaires, comprising 1700 parameters. The mid- to long-term objectives of the registry are to provide clues for a better understanding of IPF phenotype sub-clusters, triggering factors and aggravating conditions, regional and environmental characteristics, and of disease behavior and management. RESULTS: This paper describes baseline data of 525 IPF subjects recruited from 11/2009 until 10/2016. IPF patients had a mean age of 68.1 years, and seeked medical advice due to insidious dyspnea (90.1%), fatigue (69.2%), and dry coughing (53.2%). A surgical lung biopsy was performed in 32% in 2009, but in only 8% of the cases in 2016, possibly due to increased numbers of cryobiopsy. At the time of inclusion in the eurIPFreg, FVC was 68.4% ± 22.6% of predicted value, DLco ranged at 42.1% ± 17.8% of predicted value (mean value ± SD). Signs of pulmonary hypertension were found in 16.8%. Steroids, immunosuppressants and N-Acetylcysteine declined since 2009, and were replaced by antifibrotics, under which patients showed improved survival (p = 0.001). CONCLUSIONS: Our data provide important insights into baseline characteristics, diagnostic and management changes as well as outcome data in European IPF patients over time. TRIAL REGISTRATION: The eurIPFreg and eurIPFbank are listed in ClinicalTrials.gov( NCT02951416 ).

Abstract Coronavirus disease 2019 (COVID-19) can damage cerebral small vessels and cause neurological symptoms. Here we describe structural changes in cerebral small vessels of patients with COVID-19 and elucidate potential mechanisms underlying the vascular pathology. In brains of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected individuals and animal models, we found an increased number of empty basement membrane tubes, so-called string vessels representing remnants of lost capillaries. We obtained evidence that brain endothelial cells are infected and that the main protease of SARS-CoV-2 (M pro ) cleaves NEMO, the essential modulator of nuclear factor-κB. By ablating NEMO, M pro induces the death of human brain endothelial cells and the occurrence of string vessels in mice. Deletion of receptor-interacting protein kinase (RIPK) 3, a mediator of regulated cell death, blocks the vessel rarefaction and disruption of the blood–brain barrier due to NEMO ablation. Importantly, a pharmacological inhibitor of RIPK signaling prevented the M pro -induced microvascular pathology. Our data suggest RIPK as a potential therapeutic target to treat the neuropathology of COVID-19.

During lung development, parabronchial SMC (PSMC) progenitors in the distal mesenchyme secrete fibroblast growth factor 10 (Fgf10), which acts on distal epithelial progenitors to promote their proliferation. β-catenin signaling within PSMC progenitors is essential for their maintenance, proliferation, and expression of Fgf10. Here, we report that this Wnt/Fgf10 embryonic signaling cascade is reactivated in mature PSMCs after naphthalene-induced injury to airway epithelium. Furthermore, we found that this paracrine Fgf10 action was essential for activating surviving variant Clara cells (the cells in the airway epithelium from which replacement epithelial cells originate) located at the bronchoalveolar duct junctions and adjacent to neuroendocrine bodies. After naphthalene injury, PSMCs secreted Fgf10 to activate Notch signaling and induce Snai1 expression in surviving variant Clara cells, which subsequently underwent a transient epithelial to mesenchymal transition to initiate the repair process. Epithelial Snai1 expression was important for regeneration after injury. We have therefore identified PSMCs as a stem cell niche for the variant Clara cells in the lung and established that paracrine Fgf10 signaling from the niche is critical for epithelial repair after naphthalene injury. These findings also have implications for understanding the misregulation of lung repair in asthma and cancer.

Reactive oxygen species (ROS) are highly reactive oxygen derivatives that include free radicals such as superoxide anion radical (O2•−) and hydroxyl radical (HO•), as well as non-radical molecules hydrogen peroxide (H2O2), peroxynitrite (ONOO−), and hypochlorous acid (HOCl) [...]

Pulmonary hypertension (PH) is characterized by pulmonary artery remodeling that can subsequently culminate in right heart failure and premature death. Emerging evidence suggests that hypoxia-inducible factor (HIF) signaling plays a fundamental and pivotal role in the pathogenesis of PH. This Review summarizes the regulation of HIF isoforms and their impact in various PH subtypes, as well as the elaborate conditional and cell-specific knockout mouse studies that brought the role of this pathway to light. We also discuss the current preclinical status of pan- and isoform-selective HIF inhibitors, and propose new research areas that may facilitate HIF isoform-specific inhibition as a novel therapeutic strategy for PH and right heart failure.

BACKGROUND: The prevention of hypertension with the angiotensin-converting enzyme inhibitor ramipril in patients with high-normal blood pressure study addresses the issue of whether progression to manifest hypertension in patients with high-normal blood pressure can be prevented with treatment. METHODS: A total of 1008 participants with high-normal office blood pressure were randomized to ramipril treatment group (n = 505) and a control group (n = 503). The patients were followed up for 3 years. Primary endpoint was to prevent or delay the progression to manifest hypertension. Secondary endpoints were reduction in the incidence of cerebrovascular and cardiovascular events, as well as the development of hypertension as defined by ambulatory blood pressure monitoring. FINDINGS: One hundred and fifty-five patients (30.7%) in the ramipril group, and 216 (42.9%) in the control group reached the primary endpoint (relative risk reduction 34.4%, P = 0.0001). Ramipril also proved to be more effective in reducing the incidence of manifest office hypertension in patients with baseline ambulatory blood pressure monitoring high-normal blood pressure. The incidence of cerebrovascular and cardiovascular events showed no statistically significant differences between the two groups. Cough was more frequent in the ramipril group (4.8 vs. 0.4%). INTERPRETATION: There is now good clinical evidence that patients with high-normal blood pressure (prehypertension) are more likely to progress to manifest hypertension than patients with optimal or normal blood pressure. Additional ambulatory blood pressure monitoring seems to be essential to achieve correct diagnosis. Treatment of patients with high-normal office blood pressure with the angiotensin-converting enzyme inhibitor was well tolerated, and significantly reduced the risk of progression to manifest hypertension.

The respective antitumoral and protumoral roles of M1 and M2 tumor-associated macrophages (TAM) typify the complexity of macrophage function in cancer. In lung cancer, density and topology of distinct TAM phenotypes at the tumor center (TC) versus the invasive margin (IM) are largely unknown. Here, we investigated TAM subtype density and distribution between TC and IM in human lung cancer and TAM associations with overall survival. Macrophages isolated from adjacent nontumor tissue (NM), the TC (TC-TAM), and the IM (IM-TAM) were analyzed with RNA-sequencing (RNA-seq). Lung tumor tissue microarrays from 104 patient samples were constructed. M1 and M2 TAMs were identified using multiplex immunofluorescence staining and a tumor cell-TAM proximity analysis was performed. RNA-seq identified marked differences among NM, TC-TAM, and IM-TAM. On the basis of a panel of five selected markers (CD68, IL12, CCR7, CD163, and ALOX15), M2 predominance over M1 and M2 proximity to tumor cells was observed, especially at IM. Tumor cell proximity to TAM was linked with tumor cell survival and hypoxia was associated with accumulation of M2 TAM. Notably, lower density of M1 TC-TAM and higher proximity of tumor cells to M2 IM-TAM or lower proximity to M1 IM-TAM were linked with poor survival. In addition, three novel molecules (UBXN4, MFSD12, and ACTR6) from RNA-seq served as potential prognostic markers for lung cancer, and M2 predominance and juxtaposition of M2 TAM near tumor cells were associated with poor survival. Together, our results reveal the marked heterogeneity of TAM populations in different tumor regions, with M2 TAM predominance, particularly at IM. SIGNIFICANCE: This study underlines the significance of the density, spatial distribution, and gene expression of TAM phenotypes as prognostic factors for overall survival in lung cancer. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/20/4414/F1.large.jpg.

Studies from the USA have reported that sleep apnoea is common in congestive heart failure (CHF), with Cheyne-Stokes respiration (CSR) being the most frequent type of sleep-disordered breathing (SDB) in these patients. Within the present study, the authors sought to assess the prevalence and type of SDB among CHF patients in Germany. A total of 203 CHF patients participated in this prospective multicentre study. All patients were stable in New York Heart Association classes II and III and had a left ventricular ejection fraction (LVEF)<40%. The patients were investigated by polygraphy and all data were centrally analysed. Patient enrolment was irrespective of sleep-related symptoms. The majority of patients were male with a mean age of 65 yrs and hospitalised. Of the 203 patients, 145 (71%) had an apnoea/hypopnoea index>10.h(-1), obstructive sleep apnoea (OSA) occurred in 43% (n=88) and CSR in 28% (n=57) of patients. The prevalence of sleep-disordered breathing is high in patients with stable severe congestive heart failure from a European population. As sleep-disordered breathing may have a negative impact on the prognosis of congestive heart failure, a sleep study should be performed in every patient with congestive heart failure and a left ventricular ejection fraction of <40%. This diagnostic approach should probably be adopted for all of these patients irrespective of the presence of sleep-related symptoms.

Portopulmonary hypertension (POPH) is regarded as a subtype of pulmonary arterial hypertension (PAH); however, established PAH therapies have not been evaluated for this condition. The current authors treated 14 patients (four male, 10 female; mean (range) age 55 (39-75) yrs) with moderate (n = 1) or severe (n = 13) POPH caused by alcoholic liver disease (n = 7), chronic viral hepatitis (n = 3), autoimmune hepatitis (n = 3), and hepatic manifestation of hereditary haemorrhagic teleangiectasia (n = 1) with oral sildenafil. Eight patients were newly started on pulmonary vasoactive treatment, while six patients were already on treatment with inhaled prostanoids (iloprost, n = 5; treprostinil, n = 1). During treatment with sildenafil, mean +/- sd 6-min walk distance increased from 312 +/- 111 m to 397 +/- 99 m after 3 months, and 407 +/- 97 m after 12 months. Mean +/- sd pro-brain natriuretic peptide levels decreased from 582 +/- 315 ng x mL(-1) to 230 +/- 278 ng x mL(-1), and to 189 +/- 274 ng x mL(-1) after 3 and 12 months, respectively. Two patients died after 1 and 2 months from liver failure and cardiac failure, respectively. There was a similar response to sildenafil treatment after 3 and 12 months in patients on monotherapy and those on combination therapy. In conclusion, sildenafil might be effective in monotherapy and in combination therapy with inhaled prostanoids in portopulmonary hypertension, leading to significant improvement by 3 months and sustained response over 12 months.

Acetylcholine (ACh), a classical transmitter of parasympathetic nerve fibres in the airways, is also synthesized by a large number of non-neuronal cells, including airway surface epithelial cells. Strongest expression of cholinergic traits is observed in neuroendocrine and brush cells but other epithelial cell types--ciliated, basal and secretory--are cholinergic as well. There is cell type-specific expression of the molecular pathways of ACh release, including both the vesicular storage and exocytotic release known from neurons, and transmembrane release from the cytosol via organic cation transporters. The subcellular distribution of the ACh release machineries suggests luminal release from ciliated and secretory cells, and basolateral release from neuroendocrine cells. The scenario as known so far strongly suggests a local auto-/paracrine role of epithelial ACh in regulating various aspects on the innate mucosal defence mechanisms, including mucociliary clearance, regulation of macrophage function and modulation of sensory nerve fibre activity. The proliferative effects of ACh gain importance in recently identified ACh receptor disorders conferring susceptibility to lung cancer. The cell type-specific molecular diversity of the epithelial ACh synthesis and release machinery implies that it is differently regulated than neuronal ACh release and can be specifically targeted by appropriate drugs.

Alveolar macrophages (AMs) are the sentinel cells of the alveolar space, maintaining homeostasis, fending off pathogens, and controlling lung inflammation. During acute lung injury, AMs orchestrate the initiation and resolution of inflammation in order to ultimately restore homeostasis. This central role in acute lung inflammation makes AMs attractive targets for therapeutic interventions. Single-cell RNA-Seq and spatial omics approaches, together with methodological advances such as the generation of human macrophages from pluripotent stem cells, have increased understanding of the ontogeny, function, and plasticity of AMs during infectious and sterile lung inflammation, which could move the field closer to clinical application. However, proresolution phenotypes might conflict with proinflammatory and antibacterial responses. Therefore, therapeutic targeting of AMs at vulnerable time points over the course of infectious lung injury might harbor the risk of serious side effects, such as loss of antibacterial host defense capacity. Thus, the identification of key signaling hubs that determine functional fate decisions in AMs is of the utmost importance to harness their therapeutic potential.

Skin wounds are characterized by injury to the skin due to trauma, tearing, cuts, or contusions. As such injuries are common to all human groups, they may at times represent a serious socioeconomic burden. Currently, increasing numbers of studies have focused on the role of mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) in skin wound repair. As a cell-free therapy, MSC-derived EVs have shown significant application potential in the field of wound repair as a more stable and safer option than conventional cell therapy. Treatment based on MSC-derived EVs can significantly promote the repair of damaged substructures, including the regeneration of vessels, nerves, and hair follicles. In addition, MSC-derived EVs can inhibit scar formation by affecting angiogenesis-related and antifibrotic pathways in promoting macrophage polarization, wound angiogenesis, cell proliferation, and cell migration, and by inhibiting excessive extracellular matrix production. Additionally, these structures can serve as a scaffold for components used in wound repair, and they can be developed into bioengineered EVs to support trauma repair. Through the formulation of standardized culture, isolation, purification, and drug delivery strategies, exploration of the detailed mechanism of EVs will allow them to be used as clinical treatments for wound repair. In conclusion, MSC-derived EVs-based therapies have important application prospects in wound repair. Here we provide a comprehensive overview of their current status, application potential, and associated drawbacks.