Involvement of Mast Cells in the Pathology of COVID-19: Clinical and Laboratory Parallels.

Recent studies suggested the potential role of mast cells (MCs) in the pathology of coronavirus disease 2019 (COVID-19). However, the precise description of the MCs' activation and the engagement of their proteases is still missing. The objective of this study was to further reveal the importance of MCs and their proteases (chymase, tryptase, and carboxypeptidase A3 (CPA3)) in the development of lung damage in patients with COVID-19. This study included 55 patients who died from COVID-19 and 30 controls who died from external causes. A histological analysis of the lung parenchyma was carried out to assess the protease profiles and degranulation activity of MCs. In addition, we have analyzed the general blood test, coagulogram, and C-reactive protein. The content of tryptase-positive MCs (Try-MCs) in the lungs of patients with COVID-19 was higher than in controls, but their degranulation activity was lower. The indicators of chymase-positive MCs (Chy-MCs) were significantly lower than in the controls, while the content of CPA3-positive MCs (CPA3-MCs) and their degranulation activity were higher in patients with COVID-19. In addition, we have demonstrated the existence of correlations (positive/negative) between the content of Try-MCs, Chy-MCs, and CPA3-MCs at different states of their degranulation and presence (co-adjacent/single) and the levels of various immune cells (neutrophils, eosinophils, basophils, and monocytes) and other important markers (blood hemoglobin, activated partial thromboplastin time (aPTT), international normalized ratio (INR), and fibrinogen). Thus, the identified patterns suggest the numerous and diverse mechanisms of the participation of MCs and their proteases in the pathogenesis of COVID-19, and their impact on the inflammatory process and coagulation status. At the same time, the issue requires further study in larger cohorts of patients, which will open up the possibility of using drugs acting on this link of pathogenesis to treat lung damage in patients with COVID-19.

Wnt/ß-Catenin-Signaling Modulates Megakaryopoiesis at the Megakaryocyte-Erythrocyte Progenitor Stage in the Hematopoietic System.

The bone marrow (BM) hematopoietic system (HS) gives rise to blood cells originating from hematopoietic stem cells (HSCs), including megakaryocytes (MKs) and red blood cells (erythrocytes; RBCs). Many steps of the cell-fate decision remain to be elucidated, being important for cancer treatment. To explore the role of Wnt/ß-catenin for MK and RBC differentiation, we activated ß-catenin signaling in platelet-derived growth factor b (Pdgfb)-expressing cells of the HS using a Cre-lox approach (Ctnnb1BM-GOF). FACS analysis revealed that Pdgfb is mainly expressed by megakaryocytic progenitors (MKPs), MKs and platelets. Recombination resulted in a lethal phenotype in mutants (Ctnnb1BM-GOFwt/fl, Ctnnb1BM-GOFfl/fl) 3 weeks after tamoxifen injection, showing an increase in MKs in the BM and spleen, but no pronounced anemia despite reduced erythrocyte counts. BM transplantation (BMT) of Ctnnb1BM-GOF BM into lethally irradiated wildtype recipients (BMT-Ctnnb1BM-GOF) confirmed the megakaryocytic, but not the lethal phenotype. CFU-MK assays in vitro with BM cells of Ctnnb1BM-GOF mice supported MK skewing at the expense of erythroid colonies. Molecularly, the runt-related transcription factor 1 (RUNX1) mRNA, known to suppress erythropoiesis, was upregulated in Ctnnb1BM-GOF BM cells. In conclusion, ß-catenin activation plays a key role in cell-fate decision favoring MK development at the expense of erythroid production.

Fibroblast growth factor 10 reverses cigarette smoke- and elastase-induced emphysema and pulmonary hypertension in mice.

BACKGROUND: COPD is an incurable disease and a leading cause of death worldwide. In mice, fibroblast growth factor (FGF)10 is essential for lung morphogenesis, and in humans, polymorphisms in the human FGF10 gene correlate with an increased susceptibility to develop COPD. METHODS: We analysed FGF10 signalling in human lung sections and isolated cells from healthy donor, smoker and COPD lungs. The development of emphysema and PH was investigated in Fgf10+/- and Fgfr2b+/- (FGF receptor 2b) mice upon chronic exposure to cigarette smoke. In addition, we overexpressed FGF10 in mice following elastase- or cigarette smoke-induced emphysema and pulmonary hypertension (PH). RESULTS: We found impaired FGF10 expression in human lung alveolar walls and in primary interstitial COPD lung fibroblasts. In contrast, FGF10 expression was increased in large pulmonary vessels in COPD lungs. Consequently, we identified impaired FGF10 signalling in alveolar walls as an integral part of the pathomechanism that leads to emphysema and PH development: mice with impaired FGF10 signalling (Fgf10+/- and Fgfr2b+/- ) spontaneously developed lung emphysema, PH and other typical pathomechanistic features that generally arise in response to cigarette smoke exposure. CONCLUSION: In a therapeutic approach, FGF10 overexpression successfully restored lung alveolar and vascular structure in mice with established cigarette smoke- and elastase-induced emphysema and PH. FGF10 treatment triggered an initial increase in the number of alveolar type 2 cells that gradually returned to the basal level when the FGF10-mediated repair process progressed. Therefore, the application of recombinant FGF10 or stimulation of the downstream signalling cascade might represent a novel therapeutic strategy in the future.

Combined Pulmonary Fibrosis and Emphysema: When Scylla and Charybdis Ally.

Combined pulmonary fibrosis and emphysema (CPFE) is a recently recognized syndrome that, as its name indicates, involves the existence of both interstitial lung fibrosis and emphysema in one individual, and is often accompanied by pulmonary hypertension. This debilitating, progressive condition is most often encountered in males with an extensive smoking history, and is presented by dyspnea, preserved lung volumes, and contrastingly impaired gas exchange capacity. The diagnosis of the disease is based on computed tomography imaging, demonstrating the coexistence of emphysema and interstitial fibrosis in the lungs, which might be of various types and extents, in different areas of the lung and several relative positions to each other. CPFE bears high mortality and to date, specific and efficient treatment options do not exist. In this review, we will summarize current knowledge about the clinical attributes and manifestations of CPFE. Moreover, we will focus on pathophysiological and pathohistological lung phenomena and suspected etiological factors of this disease. Finally, since there is a paucity of preclinical research performed for this particular lung pathology, we will review existing animal studies and provide suggestions for the development of additional in vivo models of CPFE syndrome.

Nicotine promotes e-cigarette vapour-induced lung inflammation and structural alterations.

BACKGROUND: Electronic cigarette (e-cigarette) vapour is gaining popularity as an alternative to tobacco smoking and can induce acute lung injury. However, the specific role of nicotine in e-cigarette vapour and its long-term effects on the airways, lung parenchyma and vasculature remain unclear. RESULTS: In vitro exposure to nicotine-containing e-cigarette vapour extract (ECVE) or to nicotine-free e-cigarette vapour extract (NF ECVE) induced changes in gene expression of epithelial cells and pulmonary arterial smooth muscle cells (PASMCs), but ECVE in particular caused functional alterations (e.g. a decrease in human and mouse PASMC proliferation by 29.3±5.3% and 44.3±8.4%, respectively). Additionally, acute inhalation of nicotine-containing e-cigarette vapour (ECV) but not nicotine-free e-cigarette vapour (NF ECV) increased pulmonary endothelial permeability in isolated lungs. Long-term in vivo exposure of mice to ECV for 8 months significantly increased the number of inflammatory cells, in particular lymphocytes, compared to control and NF ECV in the bronchoalveolar fluid (BALF) (ECV: 853.4±150.8 cells·mL-1; control: 37.0±21.1 cells·mL-1; NF ECV: 198.6±94.9 cells·mL-1) and in lung tissue (ECV: 25.7±3.3 cells·mm-3; control: 4.8±1.1 cells·mm-3; NF ECV: 14.1±2.2 cells·mm-3). BALF cytokines were predominantly increased by ECV. Moreover, ECV caused significant changes in lung structure and function (e.g. increase in airspace by 17.5±1.4% compared to control), similar to mild tobacco smoke-induced alterations, which also could be detected in the NF ECV group, albeit to a lesser degree. In contrast, the pulmonary vasculature was not significantly affected by ECV or NF ECV. CONCLUSIONS: NF ECV components induce cell type-specific effects and mild pulmonary alterations, while inclusion of nicotine induces significant endothelial damage, inflammation and parenchymal alterations.

Potential long-term effects of SARS-CoV-2 infection on the pulmonary vasculature: Multilayered cross-talks in the setting of coinfections and comorbidities.

The Coronavirus Disease 2019 (COVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) and its sublineages pose a new challenge to healthcare systems worldwide due to its ability to efficiently spread in immunized populations and its resistance to currently available therapies. COVID-19, although targeting primarily the respiratory system, is also now well established that later affects every organ in the body. Most importantly, despite the available therapy and vaccine-elicited protection, the long-term consequences of viral infection in breakthrough and asymptomatic individuals are areas of concern. In the past two years, investigators accumulated evidence on how the virus triggers our immune system and the molecular signals involved in the cross-talk between immune cells and structural cells in the pulmonary vasculature to drive pathological lung complications such as endothelial dysfunction and thrombosis. In the review, we emphasize recent updates on the pathophysiological inflammatory and immune responses associated with SARS-CoV-2 infection and their potential long-term consequences that may consequently lead to the development of pulmonary vascular diseases.

Role of mast cells in the pathogenesis of severe lung damage in COVID-19 patients.

BACKGROUND: There is still insufficient knowledge with regard to the potential involvement of mast cells (MCs) and their mediators in the pathology of coronavirus disease-2019 (COVID-19). Therefore, our study aimed to investigate the role of MCs, their activation and protease profiles in the pathogenesis of early and late lung damage in COVID-19 patients. METHODS: Formalin-fixed and paraffin embedded lung specimens from 30 patients who died from COVID-19 and 9 controls were used for histological detection of MCs and their proteases (tryptase, chymase) followed by morphometric quantification. RESULTS: Our results demonstrated increased numbers of MCs at early stage and further augmentation of MCs number during the late stage of alveolar damage in COVID-19 patients, as compared to the control group. Importantly, the percentage of degranulated (activated) MCs was higher during both stages of alveolar lesions in comparison to the controls. While there was no prominent alteration in the profile of tryptase-positive MCs, our data revealed a significant elevation in the number of chymase-positive MCs in the lungs of COVID-19 patients, compared to the controls. CONCLUSIONS: MCs are characterized by dysregulated accumulation and increased activation in the lungs of patients suffering from COVID-19. However, future profound studies are needed for precise analysis of the role of these immune cells in the context of novel coronavirus disease.

Circulating Microparticles Are Differentially Increased in Lowlanders and Highlanders with High Altitude Induced Pulmonary Hypertension during the Cold Season.

The role of microparticles (MPs) and cold in high altitude pulmonary hypertension (HAPH) remains unexplored. We investigated the impact of long-term cold exposure on the pulmonary circulation in lowlanders and high-altitude natives and the role of MPs. Pulmonary hemodynamics were evaluated using Doppler echocardiography at the end of the colder and warmer seasons. We further examined the miRNA content of MPs isolated from the study participants and studied their effects on human pulmonary artery smooth muscle (hPASMCs) and endothelial cells (hPAECs). Long-term exposure to cold environment was associated with an enhanced pulmonary artery pressure in highlanders. Plasma levels of CD62E-positive and CD68-positive MPs increased in response to cold in lowlanders and HAPH highlanders. The miRNA-210 expression contained in MPs differentially changed in response to cold in lowlanders and highlanders. MPs isolated from lowlanders and highlanders increased proliferation and reduced apoptosis of hPASMCs. Further, MPs isolated from warm-exposed HAPH highlanders and cold-exposed highlanders exerted the most pronounced effects on VEGF expression in hPAECs. We demonstrated that prolonged exposure to cold is associated with elevated pulmonary artery pressures, which are most pronounced in high-altitude residents. Further, the numbers of circulating MPs are differentially increased in lowlanders and HAPH highlanders during the colder season.

HIV and Schistosoma Co-Exposure Leads to Exacerbated Pulmonary Endothelial Remodeling and Dysfunction Associated with Altered Cytokine Landscape.

HIV and Schistosoma infections have been individually associated with pulmonary vascular disease. Co-infection with these pathogens is very common in tropical areas, with an estimate of six million people co-infected worldwide. However, the effects of HIV and Schistosoma co-exposure on the pulmonary vasculature and its impact on the development of pulmonary vascular disease are largely unknown. Here, we have approached these questions by using a non-infectious animal model based on lung embolization of Schistosoma mansoni eggs in HIV-1 transgenic (HIV) mice. Schistosome-exposed HIV mice but not wild-type (Wt) counterparts showed augmented pulmonary arterial pressure associated with markedly suppressed endothelial-dependent vasodilation, increased endothelial remodeling and vessel obliterations, formation of plexiform-like lesions and a higher degree of perivascular fibrosis. In contrast, medial wall muscularization was similarly increased in both types of mice. Moreover, HIV mice displayed an impaired immune response to parasite eggs in the lung, as suggested by decreased pulmonary leukocyte infiltration, small-sized granulomas, and augmented residual egg burden. Notably, vascular changes in co-exposed mice were associated with increased expression of proinflammatory and profibrotic cytokines, including IFN-γ and IL-17A in CD4+ and γδ T cells and IL-13 in myeloid cells. Collectively, our study shows for the first time that combined pulmonary persistence of HIV proteins and Schistosoma eggs, as it may occur in co-infected people, alters the cytokine landscape and targets the vascular endothelium for aggravated pulmonary vascular pathology. Furthermore, it provides an experimental model for the understanding of pulmonary vascular disease associated with HIV and Schistosoma co-morbidity.

Quantitative Lipidomic Analysis of Takotsubo Syndrome Patients' Serum.

Takotsubo syndrome (TTS), also known as the transient left ventricular apical ballooning syndrome, is in contemporary times known as novel acute cardiac syndrome. It is characterized by transient left ventricular apical akinesis and hyperkinesis of the basal left ventricular portions. Although the precise etiology of TTS is unknown, events like the sudden release of stress hormones, such as the catecholamines and the increased inflammatory status might be plausible causes leading to the cardiovascular pathologies. Recent studies have highlighted that an imbalance in lipid accumulation might promote a deviant immune response as observed in TTS. However, there is no information on comprehensive profiling of serum lipids of TTS patients. Therefore, we investigated a detailed quantitative lipid analysis of TTS patients using ES-MSI. Our results showed significant differences in the majority of lipid species composition in the TTS patients compared to the control group. Furthermore, the computational analyses presented was able to link the altered lipids to the pro-inflammatory cytokines and disseminate possible mechanistic pathways involving TNFα and IL-6. Taken together, our study provides an extensive quantitative lipidome of TTS patients, which may provide a valuable Pre-diagnostic tool. This would facilitate the elucidation of the underlying mechanisms of the disease and to prevent the development of TTS in the future.

SPARC, a Novel Regulator of Vascular Cell Function in Pulmonary Hypertension.

BACKGROUND: Pulmonary hypertension (PH) is a life-threatening disease, characterized by excessive pulmonary vascular remodeling, leading to elevated pulmonary arterial pressure and right heart hypertrophy. PH can be caused by chronic hypoxia, leading to hyper-proliferation of pulmonary arterial smooth muscle cells (PASMCs) and apoptosis-resistant pulmonary microvascular endothelial cells (PMVECs). On reexposure to normoxia, chronic hypoxia-induced PH in mice is reversible. In this study, the authors aim to identify novel candidate genes involved in pulmonary vascular remodeling specifically in the pulmonary vasculature. METHODS: After microarray analysis, the authors assessed the role of SPARC (secreted protein acidic and rich in cysteine) in PH using lung tissue from idiopathic pulmonary arterial hypertension (IPAH) patients, as well as from chronically hypoxic mice. In vitro studies were conducted in primary human PASMCs and PMVECs. In vivo function of SPARC was proven in chronic hypoxia-induced PH in mice by using an adeno-associated virus-mediated Sparc knockdown approach. RESULTS: C57BL/6J mice were exposed to normoxia, chronic hypoxia, or chronic hypoxia with subsequent reexposure to normoxia for different time points. Microarray analysis of the pulmonary vascular compartment after laser microdissection identified Sparc as one of the genes downregulated at all reoxygenation time points investigated. Intriguingly, SPARC was vice versa upregulated in lungs during development of hypoxia-induced PH in mice as well as in IPAH, although SPARC plasma levels were not elevated in PH. TGF-ß1 (transforming growth factor ß1) or HIF2A (hypoxia-inducible factor 2A) signaling pathways induced SPARC expression in human PASMCs. In loss of function studies, SPARC silencing enhanced apoptosis and reduced proliferation. In gain of function studies, elevated SPARC levels induced PASMCs, but not PMVECs, proliferation. Coculture and conditioned medium experiments revealed that PMVECs-secreted SPARC acts as a paracrine factor triggering PASMCs proliferation. Contrary to the authors' expectations, in vivo congenital Sparc knockout mice were not protected from hypoxia-induced PH, most probably because of counter-regulatory proproliferative signaling. However, adeno-associated virus-mediated Sparc knockdown in adult mice significantly improved hemodynamic and cardiac function in PH mice. CONCLUSIONS: This study provides evidence for the involvement of SPARC in the pathogenesis of human PH and chronic hypoxia-induced PH in mice, most likely by affecting vascular cell function.

Novel Therapeutic Targets for the Treatment of Right Ventricular Remodeling: Insights from the Pulmonary Artery Banding Model.

Right ventricular (RV) function is the main determinant of the outcome of patients with pulmonary hypertension (PH). RV dysfunction develops gradually and worsens progressively over the course of PH, resulting in RV failure and premature death. Currently, approved therapies for the treatment of left ventricular failure are not established for the RV. Furthermore, the direct effects of specific vasoactive drugs for treatment of pulmonary arterial hypertension (PAH, Group 1 of PH) on RV are not fully investigated. Pulmonary artery banding (PAB) allows to study the pathogenesis of RV failure solely, thereby testing potential therapies independently of pulmonary vascular changes. This review aims to discuss recent studies of the mechanisms of RV remodeling and RV-directed therapies based on the PAB model.

Beneficial effects of inhaled surfactant in patients with COVID-19-associated acute respiratory distress syndrome.

BACKGROUND: We have investigated the use of nebulized surfactant as a potential therapeutic option for the patients with coronavirus disease 2019 (COVID-19)-associated acute respiratory distress syndrome (ARDS) undergoing non-invasive ventilation. METHODS: The patients were divided into 2 groups: surfactant (n = 33) and control (n = 32). The subjects in the surfactant group received the inhaled surfactant at daily dose of 150-300 mg. The oxygenation parameters and several clinical outcomes were analyzed. RESULTS: On the 5 day of therapy, PaO2/FiO2 improved significantly in the surfactant group compared to the control group (184 (155-212) mmHg vs 150 (91-173) mmHg, p = 0.02). The inhaled surfactant significantly reduced the need for transfer of patients to intensive care units (24.2% vs 46.9%, p = 0.05) and invasive mechanical ventilation (18.2% vs 40.6%, p = 0.04). Even more, the nebulized surfactant shortened the length of non-invasive ventilation (7 (3-13) days vs 11 (5-22) days, p = 0.02) and time spent in hospital (18 (16-27) days vs 26 (21-31) days, p = 0.003) in patients suffering from COVID-19-linked ARDS. CONCLUSIONS: Our preliminary data provided indications that inhaled surfactant therapy may represent a promising option for patients with COVID-19-associated ARDS. However, larger clinical trials are crucially needed.

An Exaggerated Rise in Pulmonary Artery Pressure in a High-Altitude Dweller during the Cold Season.

Chronic hypoxia-induced sustained pulmonary vasoconstriction and vascular remodeling lead to mild-to-moderate elevation of pulmonary artery pressure in high-altitude residents. However, in some of them, severe pulmonary hypertension may develop. Besides hypoxia, high-altitude residents also face other environmental challenges such as low ambient temperatures. We describe a case of a 49-year-old woman of Kyrgyz ethnicity with abnormally increased pulmonary artery pressure, revealed by Doppler echocardiography. Significantly elevated pulmonary artery pressure was detected in late winter and this was not associated with right ventricular hypertrophy or right ventricular dysfunction. Repeat echocardiography performed in late summer disclosed a significant attenuation of pulmonary artery pressure elevation, with no changes in right ventricular performance parameters. This case illustrates that, in susceptible individuals, long-term cold exposure could induce an abnormal pulmonary artery pressure rise, which can be reversed during warm seasons as in our patient. In certain circumstances, however, additional factors could contribute to a sustained pulmonary artery pressure increase and the development of persistent pulmonary hypertension, which often leads to right heart failure and premature death.