Transmissible Gastroenteritis Virus Infection Promotes the Self-Renewal of Porcine Intestinal Stem Cells via Wnt/ß-Catenin Pathway.

Intestinal stem cells (ISCs) play an important role in tissue repair after injury. A recent report delineates the effect of transmissible gastroenteritis virus (TGEV) infection on the small intestine of recovered pigs. However, the mechanism behind the epithelium regeneration upon TGEV infection remains unclear. To address this, we established a TGEV infection model based on the porcine intestinal organoid monolayer. The results illustrated that the porcine intestinal organoid monolayer was susceptible to TGEV. In addition, the TGEV infection initiated the interferon and inflammatory responses following the loss of absorptive enterocytes and goblet cells. However, TGEV infection did not disturb epithelial integrity but induced the proliferation of ISCs. Furthermore, TGEV infection activated the Wnt/ß-catenin pathway by upregulating the accumulation and nuclear translocation of ß-catenin, as well as promoting the expression of Wnt target genes, such as C-myc, Cyclin D1, Mmp7, Lgr5, and Sox9, which were associated with the self-renewal of ISCs. Collectively, these data demonstrated that the TGEV infection activated the Wnt/ß-catenin pathway to promote the self-renewal of ISCs and resulted in intestinal epithelium regeneration. IMPORTANCE The intestinal epithelium is a physical barrier to enteric viruses and commensal bacteria. It plays an essential role in maintaining the balance between the host and intestinal microenvironment. In addition, intestinal stem cells (ISCs) are responsible for tissue repair after injury. Therefore, prompt self-renewal of intestinal epithelium will facilitate the rebuilding of the physical barrier and maintain gut health. In the manuscript, we found that the transmissible gastroenteritis virus (TGEV) infection did not disturb epithelial integrity but induced the proliferation of ISCs and facilitated epithelium regeneration. Detailed mechanism investigations revealed that the TGEV infection activated the Wnt/ß-catenin pathway to promote the self-renewal of ISCs and resulted in intestinal epithelium regeneration. These findings will contribute to understanding the mechanism of intestinal epithelial regeneration and reparation upon viral infection.

Human Type II Taste Cells Express Angiotensin-Converting Enzyme 2 and Are Infected by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2).

Chemosensory changes are well-reported symptoms of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. The virus targets cells for entry by binding of its spike protein to cell-surface angiotensin-converting enzyme 2 (ACE2). It is not known whether ACE2 is expressed on taste receptor cells (TRCs), or whether TRCs are infected directly. in situ hybridization probe and an antibody specific to ACE2 indicated presence of ACE2 on a subpopulation of TRCs (namely, type II cells in taste buds in taste papillae). Fungiform papillae of a SARS-CoV-2+ patient exhibiting symptoms of coronavirus disease 2019 (COVID-19), including taste changes, were biopsied. Presence of replicating SARS-CoV-2 in type II cells was verified by in situ hybridization. Therefore, taste type II cells provide a potential portal for viral entry that predicts vulnerabilities to SARS-CoV-2 in the oral cavity. The continuity and cell turnover of a patient's fungiform papillae taste stem cell layer were disrupted during infection and had not completely recovered 6 weeks after symptom onset. Another patient experiencing post-COVID-19 taste disturbances also had disrupted stem cells. These results demonstrate the possibility that novel and sudden taste changes, frequently reported in COVID-19, may be the result of direct infection of taste papillae by SARS-CoV-2. This may result in impaired taste receptor stem cell activity and suggest that further work is needed to understand the acute and postacute dynamics of viral kinetics in the human taste bud.

Nasal ciliated cells are primary targets for SARS-CoV-2 replication in the early stage of COVID-19.

The upper respiratory tract is compromised in the early period of COVID-19, but SARS-CoV-2 tropism at the cellular level is not fully defined. Unlike recent single-cell RNA-Seq analyses indicating uniformly low mRNA expression of SARS-CoV-2 entry-related host molecules in all nasal epithelial cells, we show that the protein levels are relatively high and that their localizations are restricted to the apical side of multiciliated epithelial cells. In addition, we provide evidence in patients with COVID-19 that SARS-CoV-2 is massively detected and replicated within the multiciliated cells. We observed these findings during the early stage of COVID-19, when infected ciliated cells were rapidly replaced by differentiating precursor cells. Moreover, our analyses revealed that SARS-CoV-2 cellular tropism was restricted to the nasal ciliated versus oral squamous epithelium. These results imply that targeting ciliated cells of the nasal epithelium during the early stage of COVID-19 could be an ideal strategy to prevent SARS-CoV-2 propagation.

An isoform of Dicer protects mammalian stem cells against multiple RNA viruses.

In mammals, early resistance to viruses relies on interferons, which protect differentiated cells but not stem cells from viral replication. Many other organisms rely instead on RNA interference (RNAi) mediated by a specialized Dicer protein that cleaves viral double-stranded RNA. Whether RNAi also contributes to mammalian antiviral immunity remains controversial. We identified an isoform of Dicer, named antiviral Dicer (aviD), that protects tissue stem cells from RNA viruses-including Zika virus and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-by dicing viral double-stranded RNA to orchestrate antiviral RNAi. Our work sheds light on the molecular regulation of antiviral RNAi in mammalian innate immunity, in which different cell-intrinsic antiviral pathways can be tailored to the differentiation status of cells.

SARS-CoV-2 Infection and Disease Modelling Using Stem Cell Technology and Organoids.

In this Review, we briefly describe the basic virology and pathogenesis of SARS-CoV-2, highlighting how stem cell technology and organoids can contribute to the understanding of SARS-CoV-2 cell tropisms and the mechanism of disease in the human host, supporting and clarifying findings from clinical studies in infected individuals. We summarize here the results of studies, which used these technologies to investigate SARS-CoV-2 pathogenesis in different organs. Studies with in vitro models of lung epithelia showed that alveolar epithelial type II cells, but not differentiated lung alveolar epithelial type I cells, are key targets of SARS-CoV-2, which triggers cell apoptosis and inflammation, while impairing surfactant production. Experiments with human small intestinal organoids and colonic organoids showed that the gastrointestinal tract is another relevant target for SARS-CoV-2. The virus can infect and replicate in enterocytes and cholangiocytes, inducing cell damage and inflammation. Direct viral damage was also demonstrated in in vitro models of human cardiomyocytes and choroid plexus epithelial cells. At variance, endothelial cells and neurons are poorly susceptible to viral infection, thus supporting the hypothesis that neurological symptoms and vascular damage result from the indirect effects of systemic inflammatory and immunological hyper-responses to SARS-CoV-2 infection.

Neurotropic Effects of SARS-CoV-2 Modeled by the Human Brain Organoids.

COVID-19, caused by SARS-CoV-2, is a socioeconomic burden, which exhibits respiratory illness along with unexpected neurological complications. Concerns have been raised about whether the observed neurological symptoms are due to direct effects on CNS or associated with the virus's systemic effect. Recent SARS-CoV-2 infection studies using human brain organoids revealed that SARS-CoV-2 targets human neurons. Human brain organoids are stem cell-derived reductionist experimental systems that have highlighted the neurotropic effects of SARS-CoV-2. Here, we summarize the neurotoxic effects of SARS-CoV-2 using brain organoids and comprehensively discuss how brain organoids could further improve our understanding when they are fine-tuned.

Revealing Tissue-Specific SARS-CoV-2 Infection and Host Responses using Human Stem Cell-Derived Lung and Cerebral Organoids.

COVID-19 is a transmissible respiratory disease caused by a novel coronavirus, SARS-CoV-2, and has become a global health emergency. There is an urgent need for robust and practical in vitro model systems to investigate viral pathogenesis. Here, we generated human induced pluripotent stem cell (iPSC)-derived lung organoids (LORGs), cerebral organoids (CORGs), neural progenitor cells (NPCs), neurons, and astrocytes. LORGs containing epithelial cells, alveolar types 1 and 2, highly express ACE2 and TMPRSS2 and are permissive to SARS-CoV-2 infection. SARS-CoV-2 infection induces interferons, cytokines, and chemokines and activates critical inflammasome pathway genes. Spike protein inhibitor, EK1 peptide, and TMPRSS2 inhibitors (camostat/nafamostat) block viral entry in LORGs. Conversely, CORGs, NPCs, astrocytes, and neurons express low levels of ACE2 and TMPRSS2 and correspondingly are not highly permissive to SARS-CoV-2 infection. Infection in neuronal cells activates TLR3/7, OAS2, complement system, and apoptotic genes. These findings will aid in understanding COVID-19 pathogenesis and facilitate drug discovery.

Clinical analysis and pluripotent stem cells-based model reveal possible impacts of ACE2 and lung progenitor cells on infants vulnerable to COVID-19.

Introduction: An increasing number of children with severe coronavirus disease 2019 (COVID-19) is being reported, yet the spectrum of disease severity and expression patterns of angiotensin-converting enzyme 2 (ACE2) in children at different developmental stages are largely unknow. Methods: We analysed clinical features in a cohort of 173 children with COVID-19 (0-15 yrs.-old) between January 22, 2020 and March 15, 2020. We systematically examined the expression and distribution of ACE2 in different developmental stages of children by using a combination of children's lung biopsies, pluripotent stem cell-derived lung cells, RNA-sequencing profiles, and ex vivo SARS-CoV-2 pseudoviral infections. Results: It revealed that infants (< 1yrs.-old), with a weaker potency of immune response, are more vulnerable to develop pneumonia whereas older children (> 1 yrs.-old) are more resistant to lung injury. The expression levels of ACE2 however do not vary by age in children's lung. ACE2 is notably expressed not only in Alveolar Type II (AT II) cells, but also in SOX9 positive lung progenitor cells detected in both pluripotent stem cell derivatives and infants' lungs. The ACE2+SOX9+ cells are readily infected by SARS-CoV-2 pseudovirus and the numbers of the double positive cells are significantly decreased in older children. Conclusions: Infants (< 1 yrs.-old) with SARS-CoV-2 infection are more vulnerable to lung injuries. ACE2 expression in multiple types of lung cells including SOX9 positive progenitor cells, in cooperation with an unestablished immune system, could be risk factors contributing to vulnerability of infants with COVID-19. There is a need to continue monitoring lung development in young children who have recovered from SARS-CoV-2 infection.

Direct Exposure to SARS-CoV-2 and Cigarette Smoke Increases Infection Severity and Alters the Stem Cell-Derived Airway Repair Response.

Current smoking is associated with increased risk of severe COVID-19, but it is not clear how cigarette smoke (CS) exposure affects SARS-CoV-2 airway cell infection. We directly exposed air-liquid interface (ALI) cultures derived from primary human nonsmoker airway basal stem cells (ABSCs) to short term CS and then infected them with SARS-CoV-2. We found an increase in the number of infected airway cells after CS exposure with a lack of ABSC proliferation. Single-cell profiling of the cultures showed that the normal interferon response was reduced after CS exposure with infection. Treatment of CS-exposed ALI cultures with interferon ß-1 abrogated the viral infection, suggesting one potential mechanism for more severe viral infection. Our data show that acute CS exposure allows for more severe airway epithelial disease from SARS-CoV-2 by reducing the innate immune response and ABSC proliferation and has implications for disease spread and severity in people exposed to CS.

Single-cell analysis identified lung progenitor cells in COVID-19 patients.

OBJECTIVES: The high mortality of severe 2019 novel coronavirus disease (COVID-19) cases is mainly caused by acute respiratory distress syndrome (ARDS), which is characterized by increased permeability of the alveolar epithelial barriers, pulmonary oedema and consequently inflammatory tissue damage. Some but not all patients showed full functional recovery after the devastating lung damage, and so far there is little knowledge about the lung repair process. We focused on crucial roles of lung progenitor cells in alveolar cell regeneration and epithelial barrier re-establishment and aimed to uncover a possible mechanism of lung repair after severe SARS-CoV-2 infection. MATERIALS AND METHODS: Bronchoalveolar lavage fluid (BALF) of COVID-19 patients was analysed by single-cell RNA-sequencing (scRNA-seq). Transplantation of a single KRT5+ cell-derived cell population into damaged mouse lung and time-course scRNA-seq analysis was performed. RESULTS: In severe (or critical) COVID-19 patients, there is a remarkable expansion of TM4SF1+ and KRT5+ lung progenitor cells. The two distinct populations of progenitor cells could play crucial roles in alveolar cell regeneration and epithelial barrier re-establishment, respectively. The transplanted KRT5+ progenitors could long-term engraft into host lung and differentiate into HOPX+ OCLN+ alveolar barrier cell which restored the epithelial barrier and efficiently prevented inflammatory cell infiltration. CONCLUSIONS: This work uncovered the mechanism by which various lung progenitor cells work in concert to prevent and replenish alveoli loss post-severe SARS-CoV-2 infection.

Three-Dimensional Human Alveolar Stem Cell Culture Models Reveal Infection Response to SARS-CoV-2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is the cause of a present pandemic, infects human lung alveolar type 2 (hAT2) cells. Characterizing pathogenesis is crucial for developing vaccines and therapeutics. However, the lack of models mirroring the cellular physiology and pathology of hAT2 cells limits the study. Here, we develop a feeder-free, long-term, three-dimensional (3D) culture technique for hAT2 cells derived from primary human lung tissue and investigate infection response to SARS-CoV-2. By imaging-based analysis and single-cell transcriptome profiling, we reveal rapid viral replication and the increased expression of interferon-associated genes and proinflammatory genes in infected hAT2 cells, indicating a robust endogenous innate immune response. Further tracing of viral mutations acquired during transmission identifies full infection of individual cells effectively from a single viral entry. Our study provides deep insights into the pathogenesis of SARS-CoV-2 and the application of defined 3D hAT2 cultures as models for respiratory diseases.

Expression of SARS-CoV-2 entry factors in lung epithelial stem cells and its potential implications for COVID-19.

SARS-CoV-2 can infiltrate the lower respiratory tract, resulting in severe respiratory failure and a high death rate. Normally, the airway and alveolar epithelium can be rapidly reconstituted by multipotent stem cells after episodes of infection. Here, we analyzed published RNA-seq datasets and demonstrated that cells of four different lung epithelial stem cell types express SARS-CoV-2 entry factors, including Ace2. Thus, stem cells can be potentially infected by SARS-CoV-2, which may lead to defects in regeneration capacity partially accounting for the severity of SARS-CoV-2 infection and its consequences.

Bargain with the tooth fairy - The savings accounts for dental stem cells.

Despite the hard times COVID-19 has imposed on us, the Biomedical Journal strives to provide fresh and compelling reading material - to be enjoyed safely from home. In this issue, we glance behind the scenes of dental stem cell preservation for potential therapeutic use, and discover that cancer cells hijack podoplanin expression to induce thrombosis. Moreover, we learn how the helicase DDX17 promotes tumour stemness, how genetic defects in meiosis and DNA repair cause premature ovarian insufficiency, and that the brain-derived neurotrophic factor is associated with several psychiatric diseases. Further accounts relate the role of miR-95-3p in colorectal cancer, the protective power of eggplants against mercury poisoning, and the predictive value of inhibin A for premature delivery. Finally, the very rare case of adenoid cystic carcinoma in the external auditory canal receives some attention, and we get to read up on how 3D imaging and modelling combines functional and aesthetic repair of cleft lip and palate cases.

SARS-CoV-2 infection and stem cells: Interaction and intervention.

The emergence of the novel severe acute respiratory coronavirus 2 (SARS-CoV-2) in China and its rapid national and international spread have created a global health emergency. The resemblance with SARS-CoV in spike protein suggests that SARS-CoV-2 employs spike-driven entry into angiotensin-converting enzyme 2 (ACE2)-expressing cells. From a stem cell perspective, this review focuses on the possible involvement of ACE2+ stem/progenitor cells from both the upper and lower respiratory tracts in coronavirus infection. Viral infection-associated acute respiratory distress syndrome and acute lung injury occur because of dysregulation of the immune response. Mesenchymal stem cells appear to be a promising cell therapy given that they favorably modulate the immune response to reduce lung injury. The use of exogenous stem cells may lead to lung repair. Therefore, intervention by transplantation of exogenous stem cells may be required to replace, repair, remodel, and regenerate lung tissue in survivors infected with coronavirus. Ultimately, vaccines, natural killer cells and induced-pluripotent stem cell-derived virus-specific cytotoxic T lymphocytes may offer off-the-shelf therapeutics for preventing coronavirus reemergence.

CD147 as a Target for COVID-19 Treatment: Suggested Effects of Azithromycin and Stem Cell Engagement.

The expressive number of deaths and confirmed cases of SARS-CoV-2 call for an urgent demand of effective and available drugs for COVID-19 treatment. CD147, a receptor on host cells, is a novel route for SARS-CoV-2 invasion. Thus, drugs that interfere in the spike protein/CD147 interaction or CD147 expression may inhibit viral invasion and dissemination among other cells, including in progenitor/stem cells. Studies suggest beneficial effects of azithromycin in reducing viral load of hospitalized patients, possibly interfering with ligand/CD147 receptor interactions; however, its possible effects on SARS-CoV-2 invasion has not yet been evaluated. In addition to the possible effect in invasion, azithromycin decreases the expression of some metalloproteinases (downstream to CD147), induces anti-viral responses in primary human bronchial epithelial infected with rhinovirus, decreasing viral replication and release. Moreover, resident lung progenitor/stem are extensively differentiated into myofibroblasts during pulmonary fibrosis, a complication observed in COVID-19 patients. This process, and the possible direct viral invasion of progenitor/stem cells via CD147 or ACE2, could result in the decline of these cellular stocks and failing lung repair. Clinical tests with allogeneic MSCs from healthy individuals are underway to enhance endogenous lung repair and suppress inflammation.