SARS-CoV-2 Variants of Concern Hijack IFITM2 for Efficient Replication in Human Lung Cells.

It has recently been shown that an early SARS-CoV-2 isolate (NL-02-2020) hijacks interferon-induced transmembrane proteins (IFITMs) for efficient replication in human lung cells, cardiomyocytes, and gut organoids. To date, several "variants of concern" (VOCs) showing increased infectivity and resistance to neutralization have emerged and globally replaced the early viral strains. Here, we determined whether the five current SARS-CoV-2 VOCs (Alpha, Beta, Gamma, Delta, and Omicron) maintained the dependency on IFITM proteins for efficient replication. We found that depletion of IFITM2 strongly reduces viral RNA production by all VOCs in the human epithelial lung cancer cell line Calu-3. Silencing of IFITM1 had modest effects, while knockdown of IFITM3 resulted in an intermediate phenotype. Strikingly, depletion of IFITM2 generally reduced infectious virus production by more than 4 orders of magnitude. In addition, an antibody directed against the N terminus of IFITM2 inhibited SARS-CoV-2 VOC replication in induced pluripotent stem cell (iPSC)-derived alveolar epithelial type II cells, thought to represent major viral target cells in the lung. In conclusion, endogenously expressed IFITM proteins (especially IFITM2) are critical cofactors for efficient replication of genuine SARS-CoV-2 VOCs, including the currently dominant Omicron variant. IMPORTANCE Recent data indicate that SARS-CoV-2 requires endogenously expressed IFITM proteins for efficient infection. However, the results were obtained with an early SARS-CoV-2 isolate. Thus, it remained to be determined whether IFITMs are also important cofactors for infection of emerging SARS-CoV-2 VOCs that outcompeted the original strains in the meantime. This includes the Omicron VOC, which currently dominates the pandemic. Here, we show that depletion of endogenous IFITM2 expression almost entirely prevents productive infection of Alpha, Beta, Gamma, Delta, and Omicron SARS-CoV-2 VOCs in human lung cells. In addition, an antibody targeting the N terminus of IFITM2 inhibited SARS-CoV-2 VOC replication in iPSC-derived alveolar epithelial type II cells. Our results show that SARS-CoV-2 VOCs, including the currently dominant Omicron variant, are strongly dependent on IFITM2 for efficient replication, suggesting a key proviral role of IFITMs in viral transmission and pathogenicity.

Function of epithelial stem cell in the repair of alveolar injury.

Alveoli are the functional units of blood-gas exchange in the lung and thus are constantly exposed to outside environments and frequently encounter pathogens, particles and other harmful substances. For example, the alveolar epithelium is one of the primary targets of the SARS-CoV-2 virus that causes COVID-19 lung disease. Therefore, it is essential to understand the cellular and molecular mechanisms by which the integrity of alveoli epithelial barrier is maintained. Alveolar epithelium comprises two cell types: alveolar type I cells (AT1) and alveolar type II cells (AT2). AT2s have been shown to function as tissue stem cells that repair the injured alveoli epithelium. Recent studies indicate that AT1s and subgroups of proximal airway epithelial cells can also participate alveolar repair process through their intrinsic plasticity. This review discussed the potential mechanisms that drive the reparative behaviors of AT2, AT1 and some proximal cells in responses to injury and how an abnormal repair contributes to some pathological conditions.

Surfactant-based therapy against COVID-19: A review

The coronavirus disease 2019 (COVID-19) has led to serious health and economic damage to all over the world, and it still remains unstoppable. The SARS-CoV-2, by using its S-glycoprotein, binds with an angiotensin-converting enzyme 2 re-ceptor, mostly present in alveolar epithelial type II cells. Eventually pulmonary surfactant depletion occurs. The pulmonary surfactant is necessary for maintaining the natural immunity as well as the surface tension reduction within the lung alveoli during the ex-piration. Its insufficiency results in the reduction of blood oxyge-nation, poor pulmonary regeneration, lung fibrosis, and finally the respiratory system collapses. Exogenous surfactants have pre-viously shown great promise in the treatment of infant respiratory distress syndrome, and they may also aid in the healing of dam-aged alveolar cells and the prevention of respiratory failure. Sur-factant based therapy has been advised for the prevention of COVID-19, and the trials have begun around the world. Further-more, greater research on the timing, dose, and the distribution of surfactant to the COVID-19 patients is required before this tech-nique can be implemented in clinical practice.

The History and Mystery of Alveolar Epithelial Type II Cells: Focus on Their Physiologic and Pathologic Role in Lung.

Alveolar type II (ATII) cells are a key structure of the distal lung epithelium, where they exert their innate immune response and serve as progenitors of alveolar type I (ATI) cells, contributing to alveolar epithelial repair and regeneration. In the healthy lung, ATII cells coordinate the host defense mechanisms, not only generating a restrictive alveolar epithelial barrier, but also orchestrating host defense mechanisms and secreting surfactant proteins, which are important in lung protection against pathogen exposure. Moreover, surfactant proteins help to maintain homeostasis in the distal lung and reduce surface tension at the pulmonary air-liquid interface, thereby preventing atelectasis and reducing the work of breathing. ATII cells may also contribute to the fibroproliferative reaction by secreting growth factors and proinflammatory molecules after damage. Indeed, various acute and chronic diseases are associated with intensive inflammation. These include oedema, acute respiratory distress syndrome, fibrosis and numerous interstitial lung diseases, and are characterized by hyperplastic ATII cells which are considered an essential part of the epithelialization process and, consequently, wound healing. The aim of this review is that of revising the physiologic and pathologic role ATII cells play in pulmonary diseases, as, despite what has been learnt in the last few decades of research, the origin, phenotypic regulation and crosstalk of these cells still remain, in part, a mystery.

Identification of AAV serotypes for lung gene therapy in human embryonic stem cell-derived lung organoids.

Gene therapy is being investigated for a range of serious lung diseases, such as cystic fibrosis and emphysema. Recombinant adeno-associated virus (rAAV) is a well-established, safe, viral vector for gene delivery with multiple naturally occurring and artificial serotypes available displaying alternate cell, tissue, and species-specific tropisms. Efficient AAV serotypes for the transduction of the conducting airways have been identified for several species; however, efficient serotypes for human lung parenchyma have not yet been identified. Here, we screened the ability of multiple AAV serotypes to transduce lung bud organoids (LBOs)-a model of human lung parenchyma generated from human embryonic stem cells. Microinjection of LBOs allowed us to model transduction from the luminal surface, similar to dosing via vector inhalation. We identified the naturally occurring rAAV2 and rAAV6 serotypes, along with synthetic rAAV6 variants, as having tropism for the human lung parenchyma. Positive staining of LBOs for surfactant proteins B and C confirmed distal lung identity and suggested the suitability of these vectors for the transduction of alveolar type II cells. Our findings establish LBOs as a new model for pulmonary gene therapy and stress the relevance of LBOs as a viral infection model of the lung parenchyma as relevant in SARS-CoV-2 research.

RAAS, ACE2 and COVID-19; a mechanistic review.

The use of angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin receptor blockers (ARBs) in coronavirus disease 2019 (COVID-19) patients has been claimed as associated with the risk of COVID-19 infection and its subsequent morbidities and mortalities. These claims were resulting from the possibility of upregulating the expression of angiotensin-converting enzyme 2 (ACE2), facilitation of SARS-CoV-2 entry, and increasing the susceptibility of infection in such treated cardiovascular patients. ACE2 and renin-angiotensin-aldosterone system (RAAS) products have a critical function in controlling the severity of lung injury, fibrosis, and failure following the initiation of the disease. This review is to clarify the mechanisms beyond the possible deleterious effects of angiotensin II (Ang II), and the potential protective role of angiotensin 1-7 (Ang 1-7) against pulmonary fibrosis, with a subsequent discussion of the latest updates on ACEIs/ARBs use and COVID-19 susceptibility in the light of these mechanisms and biochemical explanation.

Expression of SARS-CoV-2 Entry Factors in Human Alveolar Type II Cells in Aging and Emphysema.

Alveolar type II (ATII) cells proliferate and restore the injured epithelium. It has been described that SARS-CoV-2 infection causes diffuse alveolar damage in the lungs. However, host factors facilitating virus infection in ATII cells are not well known. We determined the SARS-CoV-2-related genes and protein expression using RT-PCR and Western blotting, respectively, in ATII cells isolated from young and elderly non-smokers, smokers, and ex-smokers. Cells were also obtained from lung transplants of emphysema patients. ACE2 has been identified as the receptor for SARS-CoV-2, and we found significantly increased levels in young and elderly smokers and emphysema patients. The viral entry depends on TMPRSS2 protease activity, and a higher expression was detected in elderly smokers and ex-smokers and emphysema patients. Both ACE2 and TMPRSS2 mRNA levels were higher in this disease in comparison with non-smokers. CD209L serves as a receptor for SARS-CoV-2, and we found increased levels in ATII cells obtained from smokers and in emphysema patients. Also, our data suggest CD209L regulation by miR142. Endoplasmic reticulum stress was detected in ATII cells in this disease. Our results suggest that upregulation of SARS-CoV-2 entry factors in ATII cells in aging, smokers, and emphysema patients may facilitate infection.

Heterogeneous expression of the SARS-Coronavirus-2 receptor ACE2 in the human respiratory tract.

BACKGROUND: Zoonotically transmitted coronaviruses are responsible for three disease outbreaks since 2002, including the current COVID-19 pandemic, caused by SARS-CoV-2. Its efficient transmission and range of disease severity raise questions regarding the contributions of virus-receptor interactions. ACE2 is a host ectopeptidase and the receptor for SARS-CoV-2. Numerous reports describe ACE2 mRNA abundance and tissue distribution; however, mRNA abundance is not always representative of protein levels. Currently, there is limited data evaluating ACE2 protein and its correlation with other SARS-CoV-2 susceptibility factors. MATERIALS AND METHODS: We systematically examined the human upper and lower respiratory tract using single-cell RNA sequencing and immunohistochemistry to determine receptor expression and evaluated its association with risk factors for severe COVID-19. FINDINGS: Our results reveal that ACE2 protein is highest within regions of the sinonasal cavity and pulmonary alveoli, sites of presumptive viral transmission and severe disease development, respectively. In the lung parenchyma, ACE2 protein was found on the apical surface of a small subset of alveolar type II cells and colocalized with TMPRSS2, a cofactor for SARS-CoV2 entry. ACE2 protein was not increased by pulmonary risk factors for severe COVID-19. Additionally, ACE2 protein was not reduced in children, a demographic with a lower incidence of severe COVID-19. INTERPRETATION: These results offer new insights into ACE2 protein localization in the human respiratory tract and its relationship with susceptibility factors to COVID-19.

Heterogeneous groups of alveolar type II cells in lung homeostasis and repair.

Alveoli are the gas-exchanging units of the lung, and the alveolar barrier is often a key battleground where pathogens, allergens, and other insults from the environment are encountered. This is seen in the current coronavirus disease 2019 (COVID-19) pandemic, as alveolar epithelium is one of the major targets of SARS-COV-2, the virus that causes COVID-19. Thus, it is essential to understand the mechanisms in order to maintain the integrity of alveoli epithelium. Alveolar type II (AT2) cells behave as tissue stem cells that repair alveoli epithelium during steady-state replacement and after injury. However, not all AT2 cells are equal in their ability for self-renewal or differentiation. Through marker gene identification, lineage tracing, and single-cell RNA-sequencing (scRNA-seq), distinct subpopulations of AT2 cells have been identified that play the progenitor role in a different context. The revelation of AT2 heterogeneity has brought new insights into the role of AT2 cells in various lung disease settings and potentiates the finding of more therapeutics targets. In this mini review, we discuss the recently identified subpopulations of AT2 cells and their functions under steady-state, postinjury, and pathological conditions.

Thoughts on the alveolar phase of COVID-19.

COVID-19 can be divided into three clinical stages, and one can speculate that these stages correlate with where the infection resides. For the asymptomatic phase, the infection mostly resides in the nose, where it elicits a minimal innate immune response. For the mildly symptomatic phase, the infection is mostly in the pseudostratified epithelium of the larger airways and is accompanied by a more vigorous innate immune response. In the conducting airways, the epithelium can recover from the infection, because the keratin 5 basal cells are spared and they are the progenitor cells for the bronchial epithelium. There may be more severe disease in the bronchioles, where the club cells are likely infected. The devastating third phase is in the gas exchange units of the lung, where ACE2-expressing alveolar type II cells and perhaps type I cells are infected. The loss of type II cells results in respiratory insufficiency due to the loss of pulmonary surfactant, alveolar flooding, and possible loss of normal repair, since type II cells are the progenitors of type I cells. The loss of type I and type II cells will also block normal active resorption of alveolar fluid. Subsequent endothelial damage leads to transudation of plasma proteins, formation of hyaline membranes, and an inflammatory exudate, characteristic of ARDS. Repair might be normal, but if the type II cells are severely damaged alternative pathways for epithelial repair may be activated, which would result in some residual lung disease.