tetano
Editor, Senior Moderator
Am J Obstet Gynecol
. 2020 Oct 8;S0002-9378(20)31182-0.
doi: 10.1016/j.ajog.2020.10.010. Online ahead of print.
Reduced SARS-CoV-2 entry factors and enhanced innate immune gene expression in the nasal epithelium of pregnant rats
Arvind Palanisamy[SUP] 1 [/SUP], Tusar Giri[SUP] 2 [/SUP]
Affiliations
Abstract
Objective: An enigmatic epidemiological feature of the ongoing COVID-19 pandemic is the high rate of asymptomatic infection in pregnant women.[SUP]1[/SUP] This is puzzling because systemic immune changes predispose pregnant women to increased severity of respiratory viral infections, especially influenza A.[SUP]2[/SUP] A major roadblock in understanding this atypical clinical presentation is the poor characterization of cellular entry factors for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) - angiotensin-converting enzyme 2 (ACE2) and the androgen-sensitive transmembrane protease serine 2 (TMPRSS2) - in the respiratory tract during pregnancy. Motivated by a recent report showing estradiol-mediated downregulation of ACE2 in the airway epithelium,[SUP]3[/SUP] we hypothesized that the hormonal changes of pregnancy will decrease the expression of SARS-CoV-2 cell entry factors. Here, we compare their expression and examine the innate immune system in the nasal epithelium of term pregnant (gestational day 20) vs. non-pregnant 2-month old female rats STUDY DESIGN: All experiments were conducted after appropriate institutional approval (protocol ID:19-1071) and comply with ARRIVE guidelines. Briefly, nasal epithelia from euthanized rats (n=9 each) were dissected according to the protocol described by Dunston et al. with modifications.[SUP]4[/SUP] Collected samples were assayed for the expression of SARS-CoV-2 entry factors (Ace2, Tmprss2), innate antiviral immune genes that are highly co-expressed with Ace2 (Tnfsf10, Mx1, Nos2),[SUP]5[/SUP] and genes involved in SARS-CoV-2 detection and defense (Rig-1, Tlr7, MyD88, Irf7) with TaqMan qPCR (ThermoFisher Scientific, Inc.). In addition, we determined the expression of ACE2 (LS-c763699, LifeSpan BioSciences, Inc.; 1:1000 dilution) and TMPRSS2 (sc-515727, Santa Cruz Biotechnology, Inc.; 1:250 dilution) protein with immunoblots. Finally, we assayed ACE2 enzyme activity with a fluorometric assay (K897-100, Biovision Inc.).
Results: We observed a marked downregulation of the expression of Ace2 and Tmprss2 genes (Fig 1A) along with concomitant changes in protein expression (1B), and a significant decrease in ACE2 enzyme activity in the nasal epithelium during pregnancy (1C). Innate immune genes with antiviral function that are highly co-expressed with Ace2 (Tnfsf10, Mx1, Nos2) were significantly elevated in the nasal epithelium from pregnant rats (1D). Similarly, the expression of cytoplasmic (Rig-1) and endosomal viral sensors (Tlr7, MyD88, and Irf7) involved in the detection of SARS-CoV-2 were substantially upregulated with pregnancy (1E). Collectively, our results show a decrease in cell entry factors for SARS-CoV-2 and a surprisingly robust expression of innate immune response genes in the nasal epithelium of pregnant rats.
Conclusion: Based on our preclinical findings, we surmise that the high rate of asymptomatic infection in pregnant women is likely due to decreased SARS-CoV-2 tropism secondary to reduced expression of cell entry factors. Our observation of upregulated innate immune defense in the nasal epithelium, in contrast to the immunological indolence at the placental-fetal interface, was unexpected and novel. Considering the exquisite vulnerability of pregnant women to influenza A virus, another single-stranded RNA virus, but not SARS-CoV-2, our findings set the stage for comprehensive characterization of respiratory mucosal immunology in pregnant women to better understand host-pathogen interaction in this unique demographic subset.
. 2020 Oct 8;S0002-9378(20)31182-0.
doi: 10.1016/j.ajog.2020.10.010. Online ahead of print.
Reduced SARS-CoV-2 entry factors and enhanced innate immune gene expression in the nasal epithelium of pregnant rats
Arvind Palanisamy[SUP] 1 [/SUP], Tusar Giri[SUP] 2 [/SUP]
Affiliations
- PMID: 33039392
- PMCID: PMC7544630
- DOI: 10.1016/j.ajog.2020.10.010
Abstract
Objective: An enigmatic epidemiological feature of the ongoing COVID-19 pandemic is the high rate of asymptomatic infection in pregnant women.[SUP]1[/SUP] This is puzzling because systemic immune changes predispose pregnant women to increased severity of respiratory viral infections, especially influenza A.[SUP]2[/SUP] A major roadblock in understanding this atypical clinical presentation is the poor characterization of cellular entry factors for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) - angiotensin-converting enzyme 2 (ACE2) and the androgen-sensitive transmembrane protease serine 2 (TMPRSS2) - in the respiratory tract during pregnancy. Motivated by a recent report showing estradiol-mediated downregulation of ACE2 in the airway epithelium,[SUP]3[/SUP] we hypothesized that the hormonal changes of pregnancy will decrease the expression of SARS-CoV-2 cell entry factors. Here, we compare their expression and examine the innate immune system in the nasal epithelium of term pregnant (gestational day 20) vs. non-pregnant 2-month old female rats STUDY DESIGN: All experiments were conducted after appropriate institutional approval (protocol ID:19-1071) and comply with ARRIVE guidelines. Briefly, nasal epithelia from euthanized rats (n=9 each) were dissected according to the protocol described by Dunston et al. with modifications.[SUP]4[/SUP] Collected samples were assayed for the expression of SARS-CoV-2 entry factors (Ace2, Tmprss2), innate antiviral immune genes that are highly co-expressed with Ace2 (Tnfsf10, Mx1, Nos2),[SUP]5[/SUP] and genes involved in SARS-CoV-2 detection and defense (Rig-1, Tlr7, MyD88, Irf7) with TaqMan qPCR (ThermoFisher Scientific, Inc.). In addition, we determined the expression of ACE2 (LS-c763699, LifeSpan BioSciences, Inc.; 1:1000 dilution) and TMPRSS2 (sc-515727, Santa Cruz Biotechnology, Inc.; 1:250 dilution) protein with immunoblots. Finally, we assayed ACE2 enzyme activity with a fluorometric assay (K897-100, Biovision Inc.).
Results: We observed a marked downregulation of the expression of Ace2 and Tmprss2 genes (Fig 1A) along with concomitant changes in protein expression (1B), and a significant decrease in ACE2 enzyme activity in the nasal epithelium during pregnancy (1C). Innate immune genes with antiviral function that are highly co-expressed with Ace2 (Tnfsf10, Mx1, Nos2) were significantly elevated in the nasal epithelium from pregnant rats (1D). Similarly, the expression of cytoplasmic (Rig-1) and endosomal viral sensors (Tlr7, MyD88, and Irf7) involved in the detection of SARS-CoV-2 were substantially upregulated with pregnancy (1E). Collectively, our results show a decrease in cell entry factors for SARS-CoV-2 and a surprisingly robust expression of innate immune response genes in the nasal epithelium of pregnant rats.
Conclusion: Based on our preclinical findings, we surmise that the high rate of asymptomatic infection in pregnant women is likely due to decreased SARS-CoV-2 tropism secondary to reduced expression of cell entry factors. Our observation of upregulated innate immune defense in the nasal epithelium, in contrast to the immunological indolence at the placental-fetal interface, was unexpected and novel. Considering the exquisite vulnerability of pregnant women to influenza A virus, another single-stranded RNA virus, but not SARS-CoV-2, our findings set the stage for comprehensive characterization of respiratory mucosal immunology in pregnant women to better understand host-pathogen interaction in this unique demographic subset.