SARS-CoV-2 Cell Entry Factors ACE2 and TMPRSS2 Are Expressed in the Microvasculature and Ducts of Human Pancreas but Are Not Enriched in ? Cells
ABSTRACT: Isolated reports of new-onset diabetes in individuals with COVID-19 have led to the hypothesis that SARS-CoV-2 is directly cytotoxic to pancreatic islet ? cells. This would require binding and entry of SARS-CoV-2 into ? cells via co-expression of its canonical cell entry factors, angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2); however, their expression in human pancreas has not been clearly defined. We analyzed six transcriptional datasets of primary human islet cells and found that ACE2 and TMPRSS2 were not co-expressed in single ? cells. In pancreatic sections, ACE2 and TMPRSS2 protein was not detected in ? cells from donors with and without diabetes. Instead, ACE2 protein was expressed in islet and exocrine tissue microvasculature and in a subset of pancreatic ducts, whereas TMPRSS2 protein was restricted to ductal cells. These findings reduce the likelihood that SARS-CoV-2 directly infects ? cells in vivo through ACE2 and TMPRSS2. Graphical Abstract Highlights • ACE2 and TMPRSS2 mRNA are expressed in the human pancreas• ACE2 protein is expressed in some islet and exocrine capillaries and pericytes• ACE2 and TMPRSS2 proteins are co-expressed in some pancreatic ducts• ACE2 and TMPRSS2 protein is not detected in ? cells of the human pancreas Coate et al. examined expression of canonical SARS-CoV-2 entry proteins ACE2 and TMPRSS2 in the human pancreas and report ACE2 expression in the microvasculature, including islet pericytes, whereas both ACE2 and TMPRSS2 are expressed in some ducts. Conversely, neither protein is detected in ? cells, arguing against direct ? cell viral infection in vivo.
Project description:Diabetes is associated with increased mortality from severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Given literature suggesting a potential association between SARS-CoV-2 infection and diabetes induction, we examined pancreatic expression of angiotensin-converting enzyme 2 (ACE2), the key entry factor for SARS-CoV-2 infection. Specifically, we analyzed five public scRNA-seq pancreas datasets and performed fluorescence in situ hybridization, western blotting, and immunolocalization for ACE2 with extensive reagent validation on normal human pancreatic tissues across the lifespan, as well as those from coronavirus disease 2019 (COVID-19) cases. These in silico and ex vivo analyses demonstrated prominent expression of ACE2 in pancreatic ductal epithelium and microvasculature, but we found rare endocrine cell expression at the mRNA level. Pancreata from individuals with COVID-19 demonstrated multiple thrombotic lesions with SARS-CoV-2 nucleocapsid protein expression that was primarily limited to ducts. These results suggest SARS-CoV-2 infection of pancreatic endocrine cells, via ACE2, is an unlikely central pathogenic feature of COVID-19-related diabetes. Graphical Abstract Highlights • ACE2 mRNA and protein are expressed in human pancreatic ducts and microvasculature• ACE2 mRNA was rarely detected and at low levels in human pancreatic endocrine cells• Pancreatic ACE2 protein expression changes across the lifespan and correlates with BMI• SARS-CoV-2 NP was detected in ducts, but not endocrine cells, of COVID-19 pancreata Kusmartseva et al. demonstrate preferential ACE2 expression in pancreatic microvascular and ductal structures, suggesting these constitute a more likely target than islet endocrine cells in SARS-CoV-2 infection. This notion was supported by detection of SARS-CoV-2 nucleocapsid protein in ductal epithelium, but not endocrine cells, of pancreata from individuals with COVID-19.
Project description:The SARS-CoV-2 virus has already caused over a million COVID-19 cases and over fifty-thousand deaths globally. There is an urgent need to create novel models to study SARS-CoV-2 virus using human disease-relevant cells and tissues to understand key features of virus biology. We present a platform comprised of nine different cell and organoid derivatives from human pluripotent stem cells (hPSCs) representing all three primary germ layers, including lung progenitors and alveolar type II (AT2) cells, pancreatic endocrine cells, liver organoids, endothelial cells, cardiomyocytes, macrophages, microglia, and both cortical and dopaminergic neurons. We systematically probed which cell types are permissive to SARS-CoV-2 infection. Human pancreatic beta cells and hepatocytes were strikingly permissive to SARS-CoV-2 infection, further validated using adult primary human islets and liver organoids. Both in vitro and in a humanized mouse model, human lung progenitors and AT2 cells express the ACE2 viral receptor and were highly permissive to SARS-CoV-2 infection. Transcriptomic analysis following SARS-CoV-2 infection of hPSC-derived pancreatic and lung organoids revealed upregulation of chemokines but not type I/III interferon signaling, similar to what was seen in primary human COVID-19 pulmonary infection. Therefore, hPSC-derived cells phenocopy human COVID-19 disease and provide a valuable resource to understand SARS-CoV-2 biology and search for novel therapeutics. Overall design: Human islets were dissociated into single cells and analyzed using 10X scRNA-seq to determine the expression of ACE2 and TMPRSS2 expression in human islet alpha and beta cells.
Project description:Background:The new coronavirus (SARS-CoV-2), which has been responsible for the recent coronavirus disease 2019 (COVID-19) pandemic, uses the cell receptor angiotensin converting enzyme-2 (ACE2) for entry and the serine protease TMPRSS2 for spike (S) protein priming. Meanwhile, the presence of B0AT1 (SLC6A19) may prevent TMPRSS2 from accessing the cutting position on ACE2. Identifying the expression of these cell receptor-related genes of SARS-CoV-2 is critical for understanding the pathogenesis of SARS-CoV-2 in various tissues, especially in the kidney. Methods:The single-cell transcription datasets of the human cell landscape (HCL) and other relevant single-cell transcription databases were used to analyze the expression of ACE2, TMPRSS2, and SLC6A19 in various organs and tissues, but mainly in the kidney. Results:ACE2 was significantly expressed in the S1, S2, and S3 segments of proximal tubule (PT) cells. TMPRSS2 was widely expressed in several renal tubule populations extending from the PT cells to the collection system cell type, of which intercalated cells and the distal convoluted tubule cells showed more significant expression than PT cells. Unexpectedly, although expressed on various renal tubule populations, SLC6A19 was mainly enriched in PT cells, in line with ACE2 expression. Although alveolar-type (AT) 2 cells of the lung and intestinal epithelial cells expressed ACE2 as well as PT cells, AT 2 cells significantly expressed TMPRSS2 but not SLC6A19, while all 3 genes were significantly expressed in intestinal epithelial cells. Conclusions:ACE2 was widely expressed in specific cell subgroups of various human tissues, especially in intestinal epithelial cells, kidney PT cells, and also AT 2 cells of the lung. These 3 types of cells demonstrated significant differences in the distribution of the cell receptor-related genes of SARS-CoV-2, which may indicate the diversity of cell surface structures and differences in the affinity between SARS-CoV-2 and cells.
Project description:Infection by the Severe Acute Respiratory Syndrome-Coronavirus-2 (SARS-CoV-2) results in the novel coronavirus disease COVID-19, which has posed a serious threat globally. Infection of SARS-CoV-2 during pregnancy is associated with complications such as preterm labor and premature rupture of membranes, and a proportion of neonates born to infected mothers are also positive for the virus. During pregnancy, the placental barrier protects the fetus from pathogens and ensures healthy development. To predict if the placenta is permissive to SARS-CoV-2, we utilized publicly available single-cell RNA-seq data to identify if the placental cells express the necessary factors required for infection. SARS-CoV-2 binding receptor ACE2 and the S protein priming protease TMPRSS2 are co-expressed by a subset of syncytiotrophoblasts (STB) in the first trimester and extravillous trophoblasts (EVT) in the second trimester human placenta. In addition, the non-canonical receptor BSG/CD147 and other proteases (CTSL, CTSB, and FURIN) are detected in most of the placental cells. Other coronavirus family receptors (ANPEP and DPP4) were also expressed in the first and second trimester placental cells. Additionally, the term placenta of multiple species including humans expressed ACE2, DPP4, and ANPEP along with the viral S protein proteases. The ACE2- and TMPRSS2-positive (ACE2 + TMPRSS2 +) placental subsets expressed mRNA for proteins involved in viral budding and replication. These cells also had the mRNA for proteins that physically interact with SARS-CoV-2 in host cells. Further, we discovered unique signatures of genes in ACE2 + TMPRSS2 + STBs and EVTs. The ACE2 + TMPRSS2 + STBs are highly differentiated cells and express genes involving mitochondrial metabolism and glucose transport. The second trimester ACE2 + TMPRSS2 + EVTs are enriched for markers of endovascular trophoblasts. Both these subtypes abundantly expressed genes in the Toll-like receptor pathway. The second trimester EVTs are also enriched for components of the JAK-STAT pathway that drives inflammation. We carried out a systematic review and identified that in 12% of pregnant women with COVID-19, the placenta was infected with SARS-CoV-2, and the virus was detected in STBs. To conclude, herein we have uncovered the cellular targets for SARS-CoV-2 entry and have shown that these cells can potentially drive viremia in the developing human placenta. Our results provide a basic framework toward understanding the paraphernalia involved in SARS-CoV-2 infections in pregnancy.
Project description:Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) has rapidly caused a global pandemic associated with a novel respiratory infection: coronavirus disease-19 (COVID-19). Angiotensin-converting enzyme-2 (ACE2) is necessary to facilitate SARS-CoV-2 infection, but-owing to its essential metabolic roles-it may be difficult to target it in therapies. Transmembrane protease serine 2 (TMPRSS2), which interacts with ACE2, may be a better candidate for targeted therapies. Using publicly available expression data, we show that both ACE2 and TMPRSS2 are expressed in many host tissues, including lung. The highest expression of ACE2 is found in the testes, whereas the prostate displays the highest expression of TMPRSS2. Given the increased severity of disease among older men with SARS-CoV-2 infection, we address the potential roles of ACE2 and TMPRSS2 in their contribution to the sex differences in severity of disease. We show that expression levels of ACE2 and TMPRSS2 are overall comparable between men and women in multiple tissues, suggesting that differences in the expression levels of TMPRSS2 and ACE2 in the lung and other non-sex-specific tissues may not explain the gender disparities in severity of SARS CoV-2. However, given their instrumental roles for SARS-CoV-2 infection and their pleiotropic expression, targeting the activity and expression levels of TMPRSS2 is a rational approach to treat COVID-19.
Project description:The SARS-CoV-2 pandemic affecting the human respiratory system severely challenges public health and urgently demands for increasing our understanding of COVID-19 pathogenesis, especially host factors facilitating virus infection and replication. SARS-CoV-2 was reported to enter cells via binding to ACE2, followed by its priming by TMPRSS2. Here, we investigate ACE2 and TMPRSS2 expression levels and their distribution across cell types in lung tissue (twelve donors, 39,778 cells) and in cells derived from subsegmental bronchial branches (four donors, 17,521 cells) by single nuclei and single cell RNA sequencing, respectively. While TMPRSS2 is strongly expressed in both tissues, in the subsegmental bronchial branches ACE2 is predominantly expressed in a transient secretory cell type. Interestingly, these transiently differentiating cells show an enrichment for pathways related to RHO GTPase function and viral processes suggesting increased vulnerability for SARS-CoV-2 infection. Our data provide a rich resource for future investigations of COVID-19 infection and pathogenesis.
Project description:Background:Recent evidence demonstrated that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) propagates in intestinal epithelial cells expressing Angiotensin-Converting Enzyme 2 (ACE2), implying that these cells represent an important entry site for the viral infection. Nicotinic receptors (nAChRs) have been put forward as potential regulators of inflammation and of ACE2 expression. As vagus nerve stimulation (VNS) activates nAChRs, we aimed to investigate whether VNS can be instrumental in affecting intestinal epithelial ACE2 expression. Methods:By using publicly available datasets we qualified epithelial ACE2 expression in human intestine, and assessed gene co-expression of ACE2 and SARS-CoV-2 priming Transmembrane Serine Protease 2 (TMPRSS2) with nAChRs in intestinal epithelial cells. Next, we investigated mouse and human ACE2 expression in intestinal tissues after chronic VNS via implanted devices. Results:We show co-expression of ACE2 and TMPRSS2 with nAChRs and ?7 nAChR in particular in intestinal stem cells, goblet cells, and enterocytes. However, VNS did not affect ACE2 expression in murine or human intestinal tissue, albeit in colitis setting. Conclusions:ACE2 and TMPRSS2 are specifically expressed in epithelial cells of human intestine, and both are co-expressed with nAChRs. However, no evidence for regulation of ACE2 expression through VNS could be found. Hence, a therapeutic value of VNS with respect to SARS-CoV-2 infection risk through ACE2 receptor modulation in intestinal epithelia could not be established.
Project description:ObjectiveSince December 2019, a newly identified coronavirus (severe acute respiratory syndrome coronavirus (SARS-CoV-2)) has caused outbreaks of pneumonia in Wuhan, China. SARS-CoV-2 enters host cells via cell receptor ACE II (ACE2) and the transmembrane serine protease 2 (TMPRSS2). In order to identify possible prime target cells of SARS-CoV-2 by comprehensive dissection of ACE2 and TMPRSS2 coexpression pattern in different cell types, five datasets with single-cell transcriptomes of lung, oesophagus, gastric mucosa, ileum and colon were analysed.DesignFive datasets were searched, separately integrated and analysed. Violin plot was used to show the distribution of differentially expressed genes for different clusters. The ACE2-expressing and TMPRRSS2-expressing cells were highlighted and dissected to characterise the composition and proportion.ResultsCell types in each dataset were identified by known markers. ACE2 and TMPRSS2 were not only coexpressed in lung AT2 cells and oesophageal upper epithelial and gland cells but also highly expressed in absorptive enterocytes from the ileum and colon. Additionally, among all the coexpressing cells in the normal digestive system and lung, the expression of ACE2 was relatively highly expressed in the ileum and colon.ConclusionThis study provides the evidence of the potential route of SARS-CoV-2 in the digestive system along with the respiratory tract based on single-cell transcriptomic analysis. This finding may have a significant impact on health policy setting regarding the prevention of SARS-CoV-2 infection. Our study also demonstrates a novel method to identify the prime cell types of a virus by the coexpression pattern analysis of single-cell sequencing data.
Project description:AIMS:Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) directly binds to ACE2 (angiotensin-converting enzyme 2) to facilitate cellular entry. Compared with the lung or respiratory tract, the human heart exhibits greater ACE2 expression. However, little substantial damage was found in the heart tissue, and no viral particles were observed in the cardiac myocytes. This study aims to analyse ACE2 and SARS-CoV-2 spike (S) protein proteases at the single-cell level, to explore the cardiac involvement in COVID-19 and improve our understanding of the potential cardiovascular implications of COVID-19. METHODS AND RESULTS:With meta-analysis, the prevalence of cardiac injury in COVID-19 patients varies from 2% [95% confidence interval (CI) 0-5%, I2 = 0%] in non-ICU patients to 59% (95% CI 48-71%, I2 = 85%) in non-survivors. With public single-cell sequence data analysis, ACE2 expression in the adult human heart is higher than that in the lung (adjusted P < 0.0001). Inversely, the most important S protein cleavage protease TMPRSS2 (transmembrane protease serine protease-2) in the heart exhibits an extremely lower expression than that in the lung (adjusted P < 0.0001), which may restrict entry of SARS-CoV-2 into cardiac cells. Furthermore, we discovered that other S protein proteases, CTSL (cathepsin L) and FURIN (furin, paired basic amino acid cleaving enzyme), were expressed in the adult heart at a similar level to that in the lung, which may compensate for TMPRSS2, mediating cardiac involvement in COVID-19. CONCLUSION:Compared with the lung, ACE2 is relatively more highly expressed in the human heart, while the key S protein priming protease, TMPRSS2, is rarely expressed. The low percentage of ACE2+/TMPRSS2+ cells reduced heart vulnerability to SARS-CoV-2 to some degree. CTSL and FURIN may compensate for S protein priming to mediate SARS-CoV-2 infection of the heart.
Project description:PURPOSE:To determine the expressions of SARS-CoV-2 receptor angiotensin-converting enzyme 2 (ACE2) and type II transmembrane serine protease (TMPRSS2) genes in human and mouse ocular cells and comparison to other tissue cells. METHODS:Human conjunctiva and primary pterygium tissues were collected from pterygium patients who underwent surgery. The expression of ACE2 and TMPRSS2 genes was determined in human primary conjunctival and pterygium cells, human ocular and other tissue cell lines, mesenchymal stem cells as well as mouse ocular and other tissues by reverse transcription-polymerase chain reaction (RT-PCR) and SYBR green PCR. RESULTS:RT-PCR analysis showed consistent expression by 2 ACE2 gene primers in 2 out of 3 human conjunctival cells and pterygium cell lines. Expression by 2 TMPRSS2 gene primers could only be found in 1 out of 3 pterygium cell lines, but not in any conjunctival cells. Compared with the lung A549 cells, similar expression was noted in conjunctival and pterygium cells. In addition, mouse cornea had comparable expression of Tmprss2 gene and lower but prominent Ace2 gene expression compared with the lung tissue. CONCLUSION:Considering the necessity of both ACE2 and TMPRSS2 for SARS-CoV-2 infection, our results suggest that conjunctiva would be less likely to be infected by SARS-CoV-2, whereas pterygium possesses some possibility of SARS-CoV-2 infection. With high and consistent expression of Ace2 and Tmprss2 in cornea, cornea rather than conjunctiva has higher potential to be infected by SARS-CoV-2. Precaution is necessary to prevent possible SARS-CoV-2 infection through ocular surface in clinical practice.