Multilamellated Basement Membranes in the Capillary Network of Alveolar Capillary Dysplasia.

A minimal diffusion barrier is key to the pulmonary gas exchange. In alveolar capillary dysplasia (ACD), a rare genetically driven disease of early infancy, this crucial fibrovascular interface is compromised while the underlying pathophysiology is insufficiently understood. Recent in-depth analyses of vascular alterations in adult lung disease encouraged researchers to extend these studies to ACD and compare the changes of the microvasculature. Lung tissue samples of children with ACD (n = 12), adults with non-specific interstitial pneumonia (n = 12), and controls (n = 20) were studied using transmission electron microscopy, single-gene sequencing, immunostaining, exome sequencing, and broad transcriptome profiling. In ACD, pulmonary capillary basement membranes were hypertrophied, thickened, and multilamellated. Transcriptome profiling revealed increased CDH5, COL4A1, COL15A1, PTK2B, and FN1 and decreased VIT expression, confirmed by immunohistochemistry. In contrast, non-specific interstitial pneumonia samples showed a regular basement membrane architecture with preserved VIT expression but also increased COL15A1+ vessels. This study provides insight into the ultrastructure and pathophysiology of ACD. The lack of normally developed lung capillaries appeared to cause a replacement by COL15A1+ vessels, a mechanism recently described in interstitial lung disease. The VIT loss and FN1 overexpression might contribute to the unique appearance of basement membranes in ACD. Future studies are needed to explore the therapeutic potential of down-regulating the expression of FN1 and balancing VIT deficiency.

Pulmonary hypertension associated with lung disease: new insights into pathomechanisms, diagnosis, and management.

Patients with chronic lung diseases, particularly interstitial lung disease and chronic obstructive pulmonary disease, frequently develop pulmonary hypertension, which results in clinical deterioration, worsening of oxygen uptake, and an increased mortality risk. Pulmonary hypertension can develop and progress independently from the underlying lung disease. The pulmonary vasculopathy is distinct from that of other forms of pulmonary hypertension, with vascular ablation due to loss of small pulmonary vessels being a key feature. Long-term tobacco exposure might contribute to this type of pulmonary vascular remodelling. The distinct pathomechanisms together with the underlying lung disease might explain why treatment options for this condition remain scarce. Most drugs approved for pulmonary arterial hypertension have shown no or sometimes harmful effects in pulmonary hypertension associated with lung disease. An exception is inhaled treprostinil, which improves exercise capacity in patients with interstitial lung disease and pulmonary hypertension. There is a pressing need for safe, effective treatment options and for reliable, non-invasive diagnostic tools to detect and characterise pulmonary hypertension in patients with chronic lung disease.

Variants in FGF10 cause early onset of severe childhood interstitial lung disease: A detailed description of four affected children.

INTRODUCTION: Fibroblast growth factor 10 (FGF10) is a signaling molecule with a well-established role for lung branching morphogenesis. Rare heterozygous, deleterious variants in the FGF10 gene are known causes of the lacrimo-auriculo-dento-digital (LADD) syndrome and aplasia of lacrimal and salivary glands. Previous studies indicate that pathogenic variants in FGF10 can cause childhood Interstitial Lung Disease (chILD) due to severe diffuse developmental disorders of the lung, but detailed reports on clinical presentation and follow-up of affected children are lacking. METHODS: We describe four children with postnatal onset of chILD and heterozygous variants in FGF10, each detected by exome or whole genome sequencing. RESULTS: All children presented with postnatal respiratory failure. Two children died within the first 2 days of life, one patient died at age of 12 years due to right heart failure related to severe pulmonary hypertension (PH) and one patient is alive at age of 6 years, but still symptomatic. Histopathological analysis of lung biopsies from the two children with early postpartum demise revealed diffuse developmental disorder representing acinar dysplasia and interstitial fibrosis. Sequential biopsies of the child with survival until the age of 12 years revealed alveolar simplification and progressive interstitial fibrosis. DISCUSSION: Our report extends the phenotype of FGF10-related disorders to early onset chILD with progressive interstitial lung fibrosis and PH. Therefore, FGF10-related disorder should be considered even without previously described syndromic stigmata in children with postnatal respiratory distress, not only when leading to death in the neonatal period but also in case of persistent respiratory complaints and PH.

Human lung virtual histology by multi-scale x-ray phase-contrast computed tomography.

Objectives.As the central organ of the respiratory system, the human lung is responsible for supplying oxygen to the blood, which reaches the erythrocytes by diffusion through the alveolar walls and is then distributed throughout the body. By exploiting the difference in electron density detected by a phase shift in soft tissue, high-resolution x-ray phase-contrast computed tomography (XPCT) can resolve biological structures in a sub-µm range, shedding new light on the three-dimensional structure of the lungs, physiological functions and pathological mechanisms.Approach.This work presents both synchrotron and laboratory XPCT results of postmortem tissue from autopsies and biopsies embedded with various preparation protocols such as precision-cut lung slices, cryogenically fixed lung tissue, as well as paraffin and alcohol fixed tissue. The selection of pathological abnormalities includes channel of Lambert, bronchus-associated lymphoid tissue and alveolar capillary dysplasia with misalignment of pulmonary veins. Subsequently, quantification and visualization approaches are presented.Main results.The overall high image quality even of in-house XPCT scans for the case of FFPE biopsies can be exploited for a wide range of pulmonary pathologies and translated to dedicated and optimized instrumentation which could be operated in clinical setting. By using synchrotron radiation, contrast can be further increased to resolve sub-µm sized features down to the sub-cellular level. The results demonstrate that a wide range of preparation protocols including sample mounting in liquids can be used.Significance.With XPCT, poorly understood 3D structures can be identified in larger volume overview and subsequently studied in more detail at higher resolution. With the full 3D structure, the respective physiological functions of airways or vascular networks, and the different pathophysiologic mechanisms can be elucidated or at least underpinned with structural data. Moreover, synchrotron data can be used to validate laboratory protocols and provide ground truth for standardizing the method.

Fibrosis-Related Gene Profiling in Liver Biopsies of PiZZ α1-Antitrypsin Children with Different Clinical Courses.

PiZZ (Glu342Lys) α1-antitrypsin deficiency (AATD) is characterized by intrahepatic AAT polymerization and is a risk factor for liver disease development in children. The majority of PiZZ children are disease free, hence this mutation alone is not sufficient to cause the disease. We investigated Z-AAT polymers and the expression of fibrosis-related genes in liver tissues of PiZZ children with different clinical courses. Liver biopsies obtained during 1979-2010 at the Department of Paediatrics, Karolinska University Hospital, Sweden, were subjected to histological re-evaluation, immunohistochemistry and NanoString-based transcriptome profiling using a panel of 760 fibrosis plus 8 bile acid-related genes. Subjects were divided into three groups based on clinical outcomes: NCH (neonatal cholestasis, favourable outcome, n = 5), NCC (neonatal cholestasis, early cirrhosis and liver transplantation, n = 4), and NNCH (no neonatal cholestasis, favourable outcome, n = 5, six biopsies). Hepatocytes containing Z-AAT polymers were abundant in all groups whereas NCC showed higher expression of genes related to liver fibrosis/cirrhosis and lower expression of genes related to lipid, aldehyde/ketone, and bile acid metabolism. Z-AAT accumulation per se cannot explain the clinical outcomes of PiZZ children; however, changes in the expression of specific genes and pathways involved in lipid, fatty acid, and steroid metabolism appear to reflect the degree of liver injury.

The fatal trajectory of pulmonary COVID-19 is driven by lobular ischemia and fibrotic remodelling.

BACKGROUND: COVID-19 is characterized by a heterogeneous clinical presentation, ranging from mild symptoms to severe courses of disease. 9-20% of hospitalized patients with severe lung disease die from COVID-19 and a substantial number of survivors develop long-COVID. Our objective was to provide comprehensive insights into the pathophysiology of severe COVID-19 and to identify liquid biomarkers for disease severity and therapy response. METHODS: We studied a total of 85 lungs (n = 31 COVID autopsy samples; n = 7 influenza A autopsy samples; n = 18 interstitial lung disease explants; n = 24 healthy controls) using the highest resolution Synchrotron radiation-based hierarchical phase-contrast tomography, scanning electron microscopy of microvascular corrosion casts, immunohistochemistry, matrix-assisted laser desorption ionization mass spectrometry imaging, and analysis of mRNA expression and biological pathways. Plasma samples from all disease groups were used for liquid biomarker determination using ELISA. The anatomic/molecular data were analyzed as a function of patients' hospitalization time. FINDINGS: The observed patchy/mosaic appearance of COVID-19 in conventional lung imaging resulted from microvascular occlusion and secondary lobular ischemia. The length of hospitalization was associated with increased intussusceptive angiogenesis. This was associated with enhanced angiogenic, and fibrotic gene expression demonstrated by molecular profiling and metabolomic analysis. Increased plasma fibrosis markers correlated with their pulmonary tissue transcript levels and predicted disease severity. Plasma analysis confirmed distinct fibrosis biomarkers (TSP2, GDF15, IGFBP7, Pro-C3) that predicted the fatal trajectory in COVID-19. INTERPRETATION: Pulmonary severe COVID-19 is a consequence of secondary lobular microischemia and fibrotic remodelling, resulting in a distinctive form of fibrotic interstitial lung disease that contributes to long-COVID. FUNDING: This project was made possible by a number of funders. The full list can be found within the Declaration of interests / Acknowledgements section at the end of the manuscript.

A Morphomolecular Approach to Alveolar Capillary Dysplasia.

Alveolar capillary dysplasia (ACD) is a rare lung developmental disorder leading to persistent pulmonary arterial hypertension and fatal outcomes in newborns. The current study analyzed the microvascular morphology and the underlying molecular background of ACD. One ACD group (n = 7), one pulmonary arterial hypertension group (n = 20), and one healthy con1trol group (n = 16) were generated. Samples of histologically confirmed ACD were examined by exome sequencing and array-based comparative genomic hybridization. Vascular morphology was analyzed using scanning electron microscopy of microvascular corrosion casts. Gene expression and biological pathways were analyzed using two panels on inflammation/kinase-specific genes and a comparison analysis tool. Compartment-specific protein expression was analyzed using immunostaining. In ACD, there was an altered capillary network, a high prevalence of intussusceptive angiogenesis, and increased activity of C-X-C motif chemokine receptor 4 (CXCR4), hypoxia-inducible factor 1α (HIF1A), and angiopoietin signaling pathways compared with pulmonary arterial hypertension/healthy controls. Histologically, there was a markedly increased prevalence of endothelial tyrosine kinase receptor (TEK/TIE2)+ macrophages in ACD, compared with the other groups, whereas the CXCR4 ligand CXCL12 and HIF1A showed high expression in all groups. ACD is characterized by dysfunctional capillaries and a high prevalence of intussusceptive angiogenesis. The results indicate that endothelial CXCR4, HIF1A, and angiopoietin signaling as well as TIE2+ macrophages are crucial for the induction of intussusceptive angiogenesis and vascular remodeling. Future studies should address the use of anti-angiogenic agents in ACD, where TIE2 appears as a promising target.

Comparative Analysis of Gene Expression in Fibroblastic Foci in Patients with Idiopathic Pulmonary Fibrosis and Pulmonary Sarcoidosis.

BACKGROUND: Fibroblastic foci (FF) are characteristic features of usual interstitial pneumonia (UIP)/idiopathic pulmonary fibrosis (IPF) and one cardinal feature thought to represent a key mechanism of pathogenesis. Hence, FF have a high impact on UIP/IPF diagnosis in current guidelines. However, although less frequent, these histomorphological hallmarks also occur in other fibrotic pulmonary diseases. Currently, there is therefore a gap in knowledge regarding the underlying molecular similarities and differences of FF in different disease entities. METHODS: In this work, we analyzed the compartment-specific gene expression profiles of FF in IPF and sarcoidosis in order to elucidate similarities and differences as well as shared pathomechanisms. For this purpose, we used laser capture microdissection, mRNA and protein expression analysis. Biological pathway analysis was performed using two different gene expression databases. As control samples, we used healthy lung tissue that was donated but not used for lung transplantation. RESULTS: Based on Holm Bonferroni corrected expression data, mRNA expression analysis revealed a significantly altered expression signature for 136 out of 760 genes compared to healthy controls while half of these showed a similar regulation in both groups. Immunostaining of selected markers from each group corroborated these results. However, when comparing all differentially expressed genes with the fdr-based expression data, only 2 of these genes were differentially expressed between sarcoidosis and IPF compared to controls, i.e., calcium transport protein 1 (CAT1) and SMAD specific E3 ubiquitin protein ligase 1 (SMURF1), both in the sarcoidosis group. Direct comparison of sarcoidosis and IPF did not show any differentially regulated genes independent from the statistical methodology. Biological pathway analysis revealed a number of fibrosis-related pathways pronounced in IPF without differences in the regulatory direction. CONCLUSIONS: These results demonstrate that FF of end-stage IPF and sarcoidosis lungs, although different in initiation, are similar in gene and protein expression, encouraging further studies on the use of antifibrotic agents in sarcoidosis.

Periprocedural safety and outcome after pump implantation for intravenous treprostinil administration in patients with pulmonary arterial hypertension.

METHODS: In this retrospective observational study, we analyzed all patients with pulmonary arterial hypertension undergoing LenusPro® pump implantation between November 2013 and October 2019 at our center. Periprocedural safety was assessed by describing all complications that occurred within 28 days after surgery; complications that occurred later were described to assess long-term safety. Clinical outcomes were measured by comparison of clinical parameters and echocardiographic measurements of right ventricular function from baseline to 6-months-follow-up. RESULTS: Fifty-four patients underwent LenusPro® pump implantation for intravenous treprostinil treatment during the investigation period. Periprocedural complications occurred in 5 patients; the only anesthesia-related complication (right heart failure with recovery after prolonged intensive care and death in the further course) occurred in the only patient who underwent general anesthesia. All other patients underwent local anesthesia with or without short-acting (analgo-) sedation. Eighteen long-term complications occurred in 15 patients, most notably pump pocket or catheter related problems. Transplant-free survival rates at 1, 2, and 3 years were 77 %, 56 %, and 48 %, respectively. CONCLUSIONS: Subcutaneous pump implantation under local anesthesia and conscious analgosedation while avoiding intubation and mechanical ventilation is feasible in patients with advanced PAH. Controlled studies are needed to determine the safest anesthetic approach for this procedure. BACKGROUND/OBJECTIVES: Intravenous treprostinil treatment via a fully implantable pump is a treatment option for patients with advanced pulmonary arterial hypertension. However, there is no consensus on the preferred anesthetic approach for the implantation procedure. Primary objective was to assess periprocedural safety with particular attention to feasibility of local anesthesia and conscious analgosedation instead of general anesthesia. Long-term safety and clinical outcomes were secondary endpoints.

Pharmacokinetics of Meropenem in People with Cystic Fibrosis-A Proof of Concept Clinical Trial.

Anti-infective treatment of pulmonary exacerbations is a major issue in people with cystic fibrosis (CF). Individualized dosing strategies and adaptation of infusion times are important concepts to optimize anti-infective therapy. In this prospective non-randomized controlled open-label trial, we compared pharmacokinetics of meropenem in 12 people with CF experiencing a pulmonary exacerbation, of whom six received parenteral meropenem 2 g tid as short infusion over 30 min and six extended infusion over 120 min. We measured blood concentrations of meropenem at five predetermined time points over 240 min and calculated differences in the percentages of the time above the minimal inhibitory concentration (fT > MIC) for meropenem concentrations >16 and >32 mg/L, respectively. Mean percentages of fT > 16 and fT > 32 mg/L were higher in the extended compared to the short infusion group (83 and 56% vs. 59% and 34%), with a statistically significant prolongation of the fT > 32 mg/L (mean 134 vs. 82 min; p = 0.037). Our results demonstrate that, in people with CF, longer fT > MIC can be achieved with a simple modification of meropenem dosing. Further studies are needed to clarify if this may translate into improved microbiological and clinical outcomes, in particular in adults with difficult-to-treat chronic infection by carbapenem-resistant Pseudomonas aeruginosa.