Fetal Lung Tissue Research Articles

Ultrasound-based radiomics for the evaluation of fetal rat lung maturity: A noninvasive assessment method (Ultrasound-based radiomics in fetal rat lung).

To establish a classification model for the evaluation of rat fetal lung maturity (FLM) using radiomics technology. A total of 430 high-throughput features were extracted per fetal lung image from 134 fetal lung ultrasound images (four-cardiac-chamber views) of 67 Sprague-Dawley (SD) fetal rats with a gestational age of 16-21days. The detection of fetal lung tissues included histopathological staining and the expression of surface proteins SP-A, SP-B, and SP-C. A machine learning classification model was established using a support vector machine based on histopathological results to analyze the relationship between fetal lung texture characteristics and FLM. The rat fetal lungs were divided into two groups: terminal sac period (SD1) and canalicular period (SD2). The mRNA transcription and protein expression level of SP-C protein were significantly higher in the SD1 group than in the SD2 group (p<0.05). The diagnostic performance of the rat FLM classification model was measured as follows: area under the receiver operating characteristic curve (AUC), 0.93 (training set) and 0.89 (validation set); sensitivity, 89.26% (training set) and 87.10% (validation set); specificity, 85.87% (training set) and 79.17% (validation set); and accuracy, 87.79% (training set) and 83.64% (validation set). Ultrasound-based radiomics technology can be used to evaluate the FLM of rats, which lays a foundation for further research on this technology in human fetal lungs.

Pulmonary Blastoma in a Young Adult

Pulmonary blastoma is a rare but aggressive malignancy of the lung comprising epithelial and mesenchymal elements that resemble fetal lung tissue. This report described a case of an 18-year-old male who presented with cough and weight loss for a month. Computed tomography (CT) of the thorax revealed a large mass with mixed solid and cystic lesions on the right side of chest along with pleural effusion and mediastinal lymphadenopathy. Massive debulking was performed followed by chemotherapy. A biphasic pulmonary blastoma was diagnosed on histopathology.

Hedgehog Signaling Pathway Orchestrates Human Lung Branching Morphogenesis.

The Hedgehog (HH) signaling pathway plays an essential role in mouse lung development. We hypothesize that the HH pathway is necessary for branching during human lung development and is impaired in pulmonary hypoplasia. Single-cell, bulk RNA-sequencing data, and human fetal lung tissues were analyzed to determine the spatiotemporal localization of HH pathway actors. Distal human lung segments were cultured in an air-liquid interface and treated with an SHH inhibitor (5E1) to determine the effect of HH inhibition on human lung branching, epithelial-mesenchymal markers, and associated signaling pathways in vitro. Our results showed an early and regulated expression of HH pathway components during human lung development. Inhibiting HH signaling caused a reduction in branching during development and dysregulated epithelial (SOX2, SOX9) and mesenchymal (ACTA2) progenitor markers. FGF and Wnt pathways were also disrupted upon HH inhibition. Finally, we demonstrated that HH signaling elements were downregulated in lung tissues of patients with a congenital diaphragmatic hernia (CDH). In this study, we show for the first time that HH signaling inhibition alters important genes and proteins required for proper branching of the human developing lung. Understanding the role of the HH pathway on human lung development could lead to the identification of novel therapeutic targets for childhood pulmonary diseases.

HH pathway orchestrates the epithelial‐mesenchymal crosstalk during human lung branching morphogenesis

RationaleHedgehog (HH) pathway plays an essential role in lung branching morphogenesis in rodents. However, little is known about the role of this pathway in human lung development. We hypothesize that HH pathway is necessary for branching during human lung development and hence in pulmonary hypoplasia.MethodsExpression of HH pathway components was analyzed in single cell and bulk RNA‐sequencing data from publicly available lung datasets. Fresh human fetal lung tissues between gestational ages 10‐18 weeks were collected. RT‐qPCR, fluorescent in situ hybridization (FISH) and immunofluorescence staining (IF) were performed to determine spatiotemporal localization of HH pathway actors. In addition, distal lung segments were cultured in air‐liquid interface for up to 48 hours in the presence or absence of HH inhibitor (5E1). RT‐qPCR, western blot and IF were performed to investigate the effect of HH pathway inhibition on epithelial‐mesenchymal markers and associated signaling pathways.ResultsBulk and scRNA‐seq analysis showed an early expression of HH pathway components during human lung development. We noted an inverse expression between activators and inhibitors receptors (PTCH1/HHIP) over time. PTCH1 had a higher expression at 20 weeks compared to 14 weeks while the opposite was observed for HHIP. HHIP FISH staining was restricted to ACTA2+ airway smooth muscle cells while PTCH1 was primarily localized in mesenchymal populations adjacent to the tip of airway epithelial buds where SHH ligand is produced. Inhibiting HH signaling ex vivo reduced branching during development (2.5‐fold vs control, p&lt;0.01, n=8). Consequently, SOX9expression, an epithelial progenitor marker, was increased whereas ACTA2 was decreased (respectively 1.5‐ and 2‐fold vs control, p&lt;0.05, n=8). These results were confirmed by western blot and IF staining. No change was observed for SOX2 gene expression while a significant increase in protein expression was observed. FGF10 increased following HH inhibition (1.5‐fold vs control, p&lt;0.01, n=8), but the expression of ETV4and 5 decreased (1.5‐fold vs control, p&lt;0.05, n=8). Finally, we demonstrated that HH signaling elements (including SHH, PTCH1, GLI2) were downregulated in lung tissues from congenital diaphragmatic hernia patients.ConclusionWe have determined for the first‐time gene expression and localization of HH pathway during human lung morphogenesis. Moreover, we showed that HH signaling inhibition alters important genes and proteins expression, such as epithelial‐mesenchymal progenitors and associated pathways, required for proper branching processes. Understanding the role of HH pathway on human lung development could lead to the identification of novel therapeutic targets for childhood pulmonary diseases.

Sex-Specific Differences in MicroRNA Expression During Human Fetal Lung Development.

Background: Sex-specific differences in fetal lung maturation have been well described; however, little is known about the sex-specific differences in microRNA (miRNA) expression during human fetal lung development. Interestingly, many adult chronic lung diseases also demonstrate sex-specific differences in prevalence. The developmental origins of health and disease hypothesis suggests that these sex-specific differences in fetal lung development may influence disease susceptibility later in life. In this study, we performed miRNA sequencing on human fetal lung tissue samples to investigate differential expression of miRNAs between males and females in the pseudoglandular stage of lung development. We hypothesized that differences in miRNA expression are present between sexes in early human lung development and may contribute to the sex-specific differences seen in pulmonary diseases later in life. Methods: RNA was isolated from human fetal lung tissue samples for miRNA sequencing. The count of each miRNA was modeled by sex using negative binomial regression models in DESeq2, adjusting for post-conception age, age2, smoke exposure, batch, and RUV factors. We tested for differential expression of miRNAs by sex, and for the presence of sex-by-age interactions to determine if miRNA expression levels by age were distinct between males and females. Results: miRNA expression profiles were generated on 298 samples (166 males and 132 females). Of the 809 miRNAs expressed in human fetal lung tissue during the pseudoglandular stage of lung development, we identified 93 autosomal miRNAs that were significantly differentially expressed by sex and 129 miRNAs with a sex-specific pattern of miRNA expression across the course of the pseudoglandular period. Conclusion: Our study demonstrates differential expression of numerous autosomal miRNAs between the male and female developing human lung. Additionally, the expression of some miRNAs are modified by age across the pseudoglandular stage in a sex-specific way. Some of these differences in miRNA expression may impact susceptibility to pulmonary disease later in life. Our results suggest that sex-specific miRNA expression during human lung development may be a potential mechanism to explain sex-specific differences in lung development and may impact subsequent disease susceptibility.

An Uncommon Cause for Respiratory Failure in a Preterm Neonate.

An Uncommon Cause for Respiratory Failure in a Preterm Neonate.

Investigation of the Possible Toxicological Effects of Paracetamol on Lung in the Trimesters; An Experimental Study

Paracetamol (PAR) is an analgesic and antipyretic drug that is frequently used during pregnancy in many countries of the world. Although the fetal and maternal effects of toxic or long-term therapeutic doses used during pregnancy are known, the data on fetal effects during trimesters are insufficient. In this study, the possible effects of PAR exposure on both maternal and fetal tissues were investigated during pregnancy and in different trimester periods. Pregnant rats were exposed to different doses of PAR (low dose: 50 mg/kg and high dose: 500 mg/kg) in first trimester (1-7th days), second trimester (8-14th days) and third trimester (15-21st days) of pregnancy. On the 21st day, fetuses were removed by cesarean section under anesthesia. Maternal and fetal lung tissues samples were taken for biochemical analysis. PAR exposure decreased antioxidants levels and increased oxidative stress parameters levels in maternal and infant lung tissues. TBARS and TOS values increased compared to the control group, and it was statistically significant in all groups except the 1LD group (p &lt;0.05). SOD, CAT, GPX, TAS, and GSH parameters decreased compared to the control group, which was statistically significant in maternal lungs (p &lt;0.05). In infant lungs, it was also statistically significant for all groups in SOD, CAT, and TAS parameters. This experimental study demonstrates that exposure to PAR during pregnancy caused toxic damage to both maternal and fetal lung organs, especially in long-term usage during pregnancy, and in the third trimester.

Fetal lung adenocarcinoma (low-grade) – A case report of local recurrence

Low-grade fetal lung adenocarcinoma (L-FLAC) is a rare tumour most often affecting young females in their third to fourth decade. The histology resembles fetal lung tissue in the pseudoglandular stage of development.1 It is clinicopathologically distinct from its high-grade counterpart (H-FLAC), with differing morphology, immunohistochemical profile, molecular features and prognosis.2 L-FLAC accounts for 0.3% of all pulmonary adenocarcinomas, with H-FLAC having an incidence between 0.5–1.4%.1

Antenatal Mesenchymal Stromal Cell Extracellular Vesicle Therapy Prevents Preeclamptic Lung Injury in Mice.

In preeclamptic pregnancies, a variety of intrauterine alterations lead to abnormal placentation, release of inflammatory and/or antiangiogenic factors, and subsequent fetal growth restriction with significant potential to cause a primary insult to the developing fetal lung. Thus, modulation of the maternal intrauterine environment may be a key therapeutic avenue to prevent preeclampsia-associated developmental lung injury. Abiologic therapy of interest is mesenchymal stromal cell-derived extracellular vesicles (MEx), which we have previously shown to ameliorate preeclamptic physiology through intrauterine immunomodulation. To evaluate the therapeutic potential of MEx to improve developmental lung injury in experimental preeclampsia, using the heme oxygenase-1-null mouse (Hmox1-/-) model, preeclamptic pregnant dams were administered intravenous antenatal MEx treatment during each week of pregnancy followed by analysis of fetal and postnatal lung tissues, amniotic fluid protein profiles, and lung explant and amniotic fluid cocultures in comparison with controland untreated preeclamptic pregnancies. We first identified that a preeclamptic intrauterine environment had a significant adverse impact on fetal lung development, including alterations in fetal lung developmental gene profiles in addition to postnatal alveolar and bronchial changes. Amniotic fluid proteomic analysis and fetal lung explant and amniotic fluid cocultures further demonstrated that maternally administered MEx altered the expression of multiple inflammatory mediators in the preeclamptic intrauterine compartment, resulting in the normalization of fetal lung branching morphogenesis and developmental gene expression. Our evaluation of fetal and postnatal parameters overall suggests that antenatal MEx treatment may provide a highly valuable preventative therapeutic modality for amelioration of lung development in preeclamptic disease.

Development and Functional Characterization of Fetal Lung Organoids.

Preterm infants frequently suffer from pulmonary complications due to a physiological and structural lung immaturity resulting in significant morbidity and mortality. Novel in vitro and in vivo models are required to study the underlying mechanisms of late lung maturation and to facilitate the development of new therapeutic strategies. Organoids recapitulate essential aspects of structural organization and possibly organ function, and can be used to model developmental and disease processes. We aimed at generating fetal lung organoids (LOs) and to functionally characterize this in vitro model in comparison to primary lung epithelial cells and lung explants ex vivo. LOs were generated with alveolar and endothelial cells from fetal rat lung tissue, using a Matrigel-gradient and air-liquid-interface culture conditions. Immunocytochemical analysis showed that the LOs consisted of polarized epithelial cell adhesion molecule (EpCAM)-positive cells with the apical membrane compartment facing the organoid lumen. Expression of the alveolar type 2 cell marker, RT2-70, and the Club cell marker, CC-10, were observed. Na+ transporter and surfactant protein mRNA expression were detected in the LOs. First time patch clamp analyses demonstrated the presence of several ion channels with specific electrophysiological properties, comparable to vital lung slices. Furthermore, the responsiveness of LOs to glucocorticoids was demonstrated. Finally, maturation of LOs induced by mesenchymal stem cells confirmed the convenience of the model to test and establish novel therapeutic strategies. The results showed that fetal LOs replicate key biological lung functions essential for lung maturation and therefore constitute a suitable in vitro model system to study lung development and related diseases.

Identification and characterization of pulmonary mesenchymal stem cells derived from rat fetal lung tissue

Pulmonary mesenchymal stem cells (PMSCs) have great potential in lung tissue repair and regeneration, which have been isolated from some mammalian species, including mice, bovine and pig. However, the isolation, characteristics and differentiation potential of rat PMSCs have not been reported. In this study, we successfully isolated PMSCs from Sprague-Dawley rat fetal lung tissue in vitro for the first time and attempted to evaluate its multilineage differentiation potentials. The cultured PMSCs showed typical spindle-shaped morphology and high proliferative potential, and could be passaged for at least 13 passages and maintained high hereditary stability with more than 93.6 % of cells were diploid (2n = 42) by G-banding analysis. Furthermore, the PMSCs could express mesenchymal markers Sca-1, CD29, CD44, CD73 and CD90, but not hematopoietic markers CD34 and CD45. Besides, the expression of cell markers of AT2 (SFTPC), AT1 (PDPN) and macrophage (CD11b) were also negative. Cell cycle examination revealed majority of the PMSCs were in G0/G1 phase, which are similar with previously reported pig PMSCs. In addition, the PMSCs were multipotent and could differentiated into osteocytes, adipocytes, hepatocytes and neurons in vitro. Together, the present study demonstrated the stemness and multi-differentiation potentials of rat PMSCs, which conferred a potential regenerative cell resource for cell regenerative therapy of lung injury.

Bioaccumulation of legacy organic contaminants in pregnant Indo-Pacific humpback dolphins (Sousa chinensis): Unique features on the transplacental transfer

The placental transfer and congener composition of organic contaminants (OCs) in Indo-Pacific humpback dolphins have been little studied. In the present study, 16 polycyclic aromatic hydrocarbons (PAHs), 18 organochlorine pesticides (OCPs), and 28 polychlorinated biphenyl (PCB) congeners were analyzed in muscle, lung, liver, kidney, and blubber tissues from three mother–fetus pairs of this species stranded along the Pearl River Estuary, China. For PCBs, tetra-, penta-, hexa-, and hepta-chlorobenzene congeners were dominant in all the analyzed samples. Among them, hexachlorobenzene congeners showed the highest level in most dolphin mother-fetus tissue samples. The concentrations of PCBs and OCPs in adult females were higher in the detoxification organs (liver and kidneys) than in the muscles and lungs, whereas muscle tissues in fetuses generally exhibited higher PCBs and OCPs levels than the livers and kidneys. The most abundant PAHs in the four tissues were those with lower molecular weights, which were also the most water-soluble and bioavailable. Negative correlations between the octanol/water partition coefficients (log KOW) and the fetus/mother ratios for PCB congeners revealed that the transfer of PCBs may be determined by their lipid solubility. OCPs and PCBs with low molecular weights and low log KOW values would be more likely to accumulate in the dolphin liver, lung, kidney and muscle tissues. Furthermore, OCs with low molecular weights and low log KOW values were more concentrated in the fetal blubber, lung, and liver tissues than in their respective mothers. The ubiquitous existence of OCs in the mother–fetus pairs strongly suggested that these contaminants could pass through the placenta and partition in fetal tissue. The high transfer efficiency of PAHs and PCBs indicated that the placenta might not be an efficient barrier for these pollutants. PCBs levels in both mother and fetus dolphins could cause immunosuppression.

Molecular regulation of lung maturation in near-term fetal sheep by maternal daily vitamin C treatment in late gestation

BackgroundIn the fetus, the appropriate balance of prooxidants and antioxidants is essential to negate the detrimental effects of oxidative stress on lung maturation. Antioxidants improve respiratory function in postnatal life and adulthood. However, the outcomes and biological mechanisms of antioxidant action in the fetal lung are unknown.MethodsWe investigated the effect of maternal daily vitamin C treatment (200 mg/kg, intravenously) for a month in late gestation (105–138 days gestation, term ~145 days) on molecular regulation of fetal lung maturation in sheep. Expression of genes and proteins regulating lung development was quantified in fetal lung tissue. The number of surfactant-producing cells was determined by immunohistochemistry.ResultsMaternal vitamin C treatment increased fetal lung gene expression of the antioxidant enzyme SOD-1, hypoxia signaling genes (HIF-2α, HIF-3α, ADM, and EGLN-3), genes regulating sodium movement (SCNN1-A, SCNN1-B, ATP1-A1, and ATP1-B1), surfactant maturation (SFTP-B and ABCA3), and airway remodeling (ELN). There was no effect of maternal vitamin C treatment on the expression of protein markers evaluated or on the number of surfactant protein-producing cells in fetal lung tissue.ConclusionsMaternal vitamin C treatment in the last third of pregnancy in sheep acts at the molecular level to increase the expression of genes that are important for fetal lung maturation in a healthy pregnancy.ImpactMaternal daily vitamin C treatment for a month in late gestation in sheep increases the expression of gene-regulating pathways that are essential for normal fetal lung development.Following late gestation vitamin C exposure in a healthy pregnancy, an increase in lung gene but not protein expression may act as a mechanism to aid in the preparation for exposure to the air-breathing environment after birth.In the future, the availability/development of compounds with greater antioxidant properties than vitamin C or more specific targets at the site of oxidative stress in vivo may translate clinically to improve respiratory outcomes in complicated pregnancies at birth.

Distinct phenotypic expression levels of macrophages in neonatal lungs.

Alveolar macrophages are the front-line defense against environmental pathogens. However, to the best of our knowledge, differences in function and phenotypic expression levels of macrophages between neonatal and adult lungs have not previously been determined. The present study investigated lung tissues and analyzed blood samples to find cell markers of M1 and M2 macrophages in neonatal and adult rats. Pulmonary sepsis was induced by intrapleural instillation of lipopolysaccharide (LPS; 20 mg/kg) and survival time after administration of LPS was measured. In certain neonates, a selective inducible nitric oxide synthase (iNOS) inhibitor, 1400w, was administered prior to induction of pulmonary sepsis. Compared with adults, fetal and neonatal lung tissues had significantly higher levels of iNOS and CD86 (M1 markers), whereas the expression levels of CD206 and arginase-1 (M2 markers) were lower in the neonatal lung. The circulating cells that co-expressed CD68 (monocytes and macrophages) and CD86 in the blood were also significantly higher in neonates than in adults (25.9±6.6 vs. 11.6±2.2%; P=0.007. At basal unstimulated conditions, lung tissue concentrations of nitrite and nitrate (NOx) were significantly lower in the neonates than in adults (112.1±55.9 vs. 340.9±124.9 µM/g; P<0.001). However, NOx was increased following administration of LPS. Administration of 1400w suppressed lung tissue levels of NOx and improved the survival time in neonatal rats treated with LPS. The present study demonstrated that M1 is the primary macrophage phenotype in the neonatal lung and that higher iNOS expression levels do not have a protective effect against pulmonary endotoxins in neonates. Overproduction of NO by iNOS in neonatal alveolar macrophages may result in detrimental effects during pulmonary inflammation.

Intrauterine Gardnerella vaginalis Infection Results in Fetal Growth Restriction and Alveolar Septal Hypertrophy in a Rabbit Model.

Background: Gardnerella vaginalis (GV) is most frequently associated with bacterial vaginosis and is the second most common etiology causing intrauterine infection after Ureaplasma urealyticum. Intrauterine GV infection adversely affects pregnancy outcomes, resulting in preterm birth, fetal growth restriction, and neonatal pneumonia. The knowledge of how GV exerts its effects is limited. We developed an in vivo animal model to study its effects on fetal development.Materials and Methods: A survival mini-laparotomy was conducted on New Zealand rabbits on gestational day 21 (28 weeks of human pregnancy). In each dam, fetuses in the right uterine horn received intra-amniotic 0.5 × 102 colony-forming units of GV injections each, while their littermate controls in the left horn received sterile saline injections. A second laparotomy was performed seven days later. Assessment of the fetal pups, histopathology of the placenta and histomorphometric examination of the fetal lung tissues was done.Results: Three dams with a combined total of 12 fetuses were exposed to intra-amniotic GV, and 9 fetuses were unexposed. The weights of fetuses, placenta, and fetal lung were significantly lower in the GV group than the saline-inoculated control group [mean gross weight, GV (19.8 ± 3.8 g) vs. control (27.9 ± 1.7 g), p < 0.001; mean placenta weight, GV (5.5 ± 1.0 g) vs. control (6.5 ± 0.7 g), p = 0.027; mean fetal lung weight, GV (0.59 ± 0.11 g) vs. control (0.91 ± 0.08 g), p = 0.002. There was a two-fold increase in the multinucleated syncytiotrophoblasts in the placenta of the GV group than their littermate controls (82.9 ± 14.9 vs. 41.6 ± 13.4, p < 0.001). The mean alveolar septae of GV fetuses was significantly thicker than the control (14.8 ± 2.8 μm vs. 12.4 ± 3.8 μm, p = 0.007). Correspondingly, the proliferative index in the interalveolar septum was 1.8-fold higher in the GV group than controls (24.9 ± 6.6% vs. 14.2 ± 2.9%, p = 0.011). The number of alveoli and alveolar surface area did not vary between groups.Discussion: Low-dose intra-amniotic GV injection induces fetal growth restriction, increased placental multinucleated syncytiotrophoblasts and fetal lung re-modeling characterized by alveolar septal hypertrophy with cellular proliferative changes.Conclusion: This intra-amniotic model could be utilized in future studies to elucidate the acute and chronic effects of GV intrauterine infections.

Affinity chromatography based phosphoproteome research on lung cancer cells and its application

磷酸化是蛋白质翻译后修饰的重要形式之一,其异常往往会导致细胞内信号通路的紊乱和疾病的发生。固定化金属离子亲和色谱(IMAC)是磷酸化肽段的高效富集技术,在磷酸化蛋白质组研究方面应用广泛。该研究以金属钛离子(Ti4+)螯合IMAC材料(Ti4+-IMAC)为载体,进行磷酸化肽段富集。比较了10 μm Ti4+-IMAC通过振荡法和固相萃取法(SPE)富集磷酸肽的效果,发现振荡法可以富集到更多的磷酸肽;对比了两种尺寸(10 μm和30 μm)Ti4+-IMAC在磷酸化肽段富集中的差异,发现小尺寸材料富集效果更佳。进一步采用优化的策略比较了不同转移能力肺癌细胞的磷酸化蛋白质组,免标记定量蛋白质组学结果表明,优化的Ti4+-IMAC方法可以从正常的肺成纤维细胞MRC5、低转移肺癌细胞95C和高转移肺癌细胞95D中分别鉴定到510、863和1108种磷酸化蛋白质,其中317种为3组所共有。该研究共鉴定到1268种磷酸化蛋白质上的7560个磷酸化位点,其中1130个为差异磷酸化位点,文献报道显示部分异常表达的激酶与癌症转移密切相关。通过生信对比分析发现,异常表达的磷酸化蛋白质主要与细胞侵袭、迁移和死亡等细胞迁移方面的功能有关。通过优化磷酸化肽富集策略,初步阐明了磷酸化蛋白质网络的异常与肺癌转移之间的相关性,该方法有望用于肺癌进展相关的磷酸化位点、磷酸化蛋白质及其信号通路研究。

A SCID mouse-human lung xenograft model of SARS-CoV-2 infection.

SARS-CoV-2 infection, which is responsible for the current COVID-19 pandemic, can cause life-threatening pneumonia, respiratory failure and even death. Characterizing SARS-CoV-2 pathogenesis in primary human target cells and tissues is crucial for developing vaccines and therapeutics. However, given the limited access to clinical samples from COVID-19 patients, there is a pressing need for in vitro/in vivo models to investigate authentic SARS-CoV-2 infection in primary human lung cells or tissues with mature structures. The present study was designed to evaluate a humanized mouse model carrying human lung xenografts for SARS-CoV-2 infection in vivo.Methods: Human fetal lung tissue surgically grafted under the dorsal skin of SCID mice were assessed for growth and development after 8 weeks. Following SARS-CoV-2 inoculation into the differentiated lung xenografts, viral replication, cell-type tropism and histopathology of SARS-CoV-2 infection, and local cytokine/chemokine expression were determined over a 6-day period. The effect of IFN-α treatment against SARS-CoV-2 infection was tested in the lung xenografts.Results: Human lung xenografts expanded and developed mature structures closely resembling normal human lung. SARS-CoV-2 replicated and spread efficiently in the lung xenografts with the epithelial cells as the main target, caused severe lung damage, and induced a robust pro-inflammatory response. IFN-α treatment effectively inhibited SARS-CoV-2 replication in the lung xenografts.Conclusions: These data support the human lung xenograft mouse model as a useful and biological relevant tool that should facilitate studies on the pathogenesis of SARS-CoV-2 lung infection and the evaluation of potential antiviral therapies.

Fetal Lung Tissue Engineering.

Lung transplantation may be considered as a final treatment option for diseases such as chronic lung disease, pulmonary hypertension, bronchopulmonary dysplasia, pulmonary fibrosis, and end-stage lung disease. The five-year survival rate of lung transplants is nearly 50%. Unfortunately, many patients will die before a suitable lung donor can be found. Importantly, the shortage of donor organs has been a significant problem in lung transplantation. The tissue engineering approach uses de- and recellularization of lung tissue to create functional lung substitutes to overcome donor lung limitations. Decellularization is hope for generating an intact ECM in the development of the engineered lung. The goal of decellularizationis to prepare a suitable scaffold of lung tissue that contains an appropriate framework for the functionality of regenerated lung tissue. In this chapter, we aim to describe the decellularization protocols for lung tissue regenerative purposes.

Functional Complexes of Angiotensin-Converting Enzyme 2 and Renin-Angiotensin System Receptors: Expression in Adult but Not Fetal Lung Tissue.

Angiotensin-converting enzyme 2 (ACE2) is a membrane peptidase and a component of the renin-angiotensin system (RAS) that has been found in cells of all organs, including the lungs. While ACE2 has been identified as the receptor for severe acute respiratory syndrome (SARS) coronaviruses, the mechanism underlying cell entry remains unknown. Human immunodeficiency virus infects target cells via CXC chemokine receptor 4 (CXCR4)-mediated endocytosis. Furthermore, CXCR4 interacts with dipeptidyl peptidase-4 (CD26/DPPIV), an enzyme that cleaves CXCL12/SDF-1, which is the chemokine that activates this receptor. By analogy, we hypothesized that ACE2 might also be capable of interactions with RAS-associated G-protein coupled receptors. Using resonance energy transfer and cAMP and mitogen-activated protein kinase signaling assays, we found that human ACE2 interacts with RAS-related receptors, namely the angiotensin II type 1 receptor (AT1R), the angiotensin II type 2 receptor (AT2R), and the MAS1 oncogene receptor (MasR). Although these interactions led to various alterations of signal transduction, but, more importantly, ligand binding to AT1R resulted in the downregulation of ACE2 cell surface expression, while ligand binding to AT2R, but not to MasR, resulted in upregulation of ACE2 cell surface expression. Proximity ligation assays performed in situ revealed macromolecular complexes containing ACE2 and AT1R, AT2R or MasR in adult but not fetal mouse lung tissue. These findings highlight the relevance of RAS in SARS-CoV-2 infection and the role of ACE2-containing complexes as potential therapeutic targets.

Effect of Intrauterine Smoke Exposure on microRNA-15a Expression in Human Lung Development and Subsequent Asthma Risk.

Background: In utero smoke (IUS) exposure is associated with asthma susceptibility. Objective: We sought to test the hypothesis that changes in miRNA expression by IUS exposure during human lung development is associated with asthma susceptibility. Methods: Gene expression was profiled from 53 IUS unexposed and 51 IUS exposed human fetal lung tissues. We tested for the differential expression of miRNAs across post-conception age and by IUS using linear models with covariate adjustment. We tested the IUS-associated miRNAs for association with their gene expression targets using pair-wise inverse correlation. Using our mouse model, we investigated the persistence of the IUS-associated miRNA signature using RT-PCR from the lungs of mouse pups with and without IUS at postnatal day 14. MiRNAs were then tested for association with asthma and exacerbations using whole blood gene expression profiles from Asthma BRIDGE. Results: Five miRNAs were differentially expressed across post-conception age (adjusted p < 0.0002) including two that were differentially expressed by IUS exposure in human fetal lung (p < 0.05). MiR-15a was differentially expressed by post-conception age (p = 0.00002), IUS exposure in human fetal lung (p = 0.005), and in the post-natal mouse lung (p = 0.01). MiR-15a was also associated with the in utero expression of GSDMB (adjusted p = 0.0002), a known childhood asthma gene and with asthma exacerbations (p = 0.0009) in Asthma BRIDGE. Thus, miR-15a is expressed during human lung development, is impacted by IUS exposure, regulates the intrauterine expression of asthma genes, and is associated with asthma severity. Conclusions: These results provide evidence for the role of miR-15a in the fetal origin of asthma.