key: cord-299116-1agfnjvq
authors: Bunders, Madeleine; Altfeld, Marcus
title: Implications of sex differences in immunity for SARS-CoV-2 pathogenesis and design of therapeutic interventions
date: 2020-08-17
journal: Immunity
DOI: 10.1016/j.immuni.2020.08.003
sha: 
doc_id: 299116
cord_uid: 1agfnjvq

Abstract Men present more frequently with severe manifestations of COVID-19 and are at higher risk for death. The underlying mechanisms for these differences between female and male individuals infected with SARS-CoV-2 are insufficiently understood. However, studies from other viral infections have shown that females can mount stronger immune responses against viruses than males. Emerging knowledge on the basic biological pathways that underlie differences in immune responses between women and men needs to be incorporated into research efforts on SARS-CoV-2 pathogenesis and pathology to identify targets for therapeutic interventions aimed at enhancing antiviral immune function and lung airway resilience while reducing pathogenic inflammation in COVID-19.

Our immune system defends us from pathogens. Optimal immunological homeostasis is achieved when the pathogen is removed with high efficiency whilst avoiding collateral tissue damage for the host (Holt et al., 2008) . This immunological balance is different between women and men. Increasing data demonstrates that women mount stronger immune responses against viruses and vaccines (Flanagan et al., 2017; Klein and Flanagan, 2016; Ziegler and Altfeld, 2017) , and also exhibit superior immune-mediated tissue repair capacities (Vom Steeg and Klein, 2019) . The current Severe Acute Respiratory Syndrome Corona Virus 2 (SARS-CoV-2) pandemic highlights the clinical consequences of these sex differences in antiviral immunity and tissue resilience Scully et al., 2020) , with in particular older men suffering from severe Coronavirus disease 2019 (COVID-19) and experiencing higher case mortality rates (Docherty et al., 2020; Grasselli et al., 2020; Jin et al., 2020; Salje et al., 2020) . Compiled data from Europe for example show male:female ratios of 1.5 for COVID-19 hospitalizations and of 1.7 to 1.8 for COVID-19 case fatality rates (Gebhard et al., 2020) . Although sex differences in manifestations of viral infections have been established for multiple viruses, including SARS-CoV, hepatitis C virus, ZIKA virus, respiratory syncytial virus, human immunodeficiency virus (HIV-1) and Influenza virus (Baden et al., 2014; Gabriel and Arck, 2014; Klein and Flanagan, 2016; Mazur et al., 2015; Meier et al., 2009; Robinson et al., 2011; Stanelle-Bertram et al., 2018; van Lunzen and Altfeld, 2014) , sex-specific treatment strategies are rarely applied outside the field of oncology. However, sex differences in case fatality rates seem to be higher in COVID-19 compared to these other infections (Gebhard et al., 2020) , emphasizing the need to develop treatment strategies for COVID-19 that take these differences between the sexes into account. Indeed, in response to the urgency of the COVID-19 health crisis, sex-based treatment options in COVID-19 patients, including the use of estrogen, estrogen antagonists and androgen antagonists (NCT04359329, NCT04397718, NCT04374279, NCT04389580), are currently being tested and planned. However, given the complexities underlying sex-specific differences in infectious diseases, ranging from environmental to biological factors, such as differences in sex hormones and expression of X chromosome-encoded genes, supplementing men with female hormones might not be the final answer. The rapidly expanding knowledge on the differential regulation of antiviral immunity and immunemediated tissue repair pathways between women and men needs to be harnessed for the design and testing of novel treatment strategies for SARS-CoV-2 infection aimed at enhancing antiviral immunity, preventing pathological immune activation and strengthening tissue resilience. Here we discuss biological pathways that provide women advantages over men in achieving immunological homeostasis in response to viral infections and address their potential implications for the treatment of severe COVID-19.

COVID-19, caused by SARS-CoV-2, was first reported from China in December 2019 (Zavascki and Falci, 2020; Zhang, 2020) . Although Middle East respiratory syndrome coronavirus (MERS-CoV) and SARS-CoV infections have higher mortality rates than SARS-CoV-2, the more successful person-to-person spread of SARS-CoV-2 has resulted in high numbers of worldwide infections and deaths. Initial manifestations of viral infections depend on the tissue distribution of the receptors that allow for viral binding and entry into host cells. Angiotensin converting enzyme 2 (ACE2) serves as the principal entry receptor for SARS-COV-2 (Hoffmann et al., 2020; Zhou et al., 2020a) and is expressed in numerous tissues, including nasal, respiratory and intestinal epithelial cells, kidney and blood vessels (Lamers et al., 2020; Puelles et al., 2020; Sungnak et al., 2020; Teuwen et al., 2020; Tukiainen et al., 2017) . The principal mode of SARS-CoV-2 transmission appears to be the respiratory route with initial infection of the upper respiratory tract (Sungnak et al., 2020) , and infected individuals suffer from symptoms similar to those of other upper respiratory tract infections. Control of further viral dissemination at this early stage prevents the development of more severe COVID-19 symptoms; however SARS-CoV-2 can spread through aerosols to new hosts. It has been suggested that SARS-CoV-2 is able to spread efficiently amongst the human population due to a higher affinity for ACE2-binding compared to SARS-CoV Wrapp et al., 2020) and efficient cell fusion facilitated by TMPRSS2 (Hoffmann et al., 2020; Zang et al., 2020) . Failure to prevent viral dissemination beyond the upper respiratory tract results in infection of the lower respiratory airways, apoptosis of pneumocytes and decreased capacity for gas exchange, leading to reduced oxygenation and eventually requirement for respiratory support. Attempts of the immune response to control SARS-CoV-2 infection in the lower respiratory tract can aggravate oxygenation by causing collateral lung tissue damage, resulting in pulmonary edema and further occlusion of small airways and eventually leading to acute respiratory distress syndrome (ARDS). The broad tissue expression of ACE2 furthermore enables SARS-CoV-2 to spread to other tissues and to infect the vascular endothelium, resulting in multi-organ failure accompanied by vasoocclusion and enhanced coagulation, and impairing vital functions (Puelles et al., 2020; Wichmann et al., 2020) . An additional characteristic of severe COVID-19 is a massive cytokine release syndrome believed to be triggered by SARS-CoV-2-induced cytokine production by macrophages and dendritic cells that further contributes to pathogenesis (Hirano and Murakami, 2020; Mangalmurti and Hunter, 2020; Merad and Martin, 2020; Moore and June, 2020) . In the following, we will address the different stages of SARS-CoV-2 pathogenesis, including viral entry and sensing, induction of antiviral immune responses and inflammation, and immune-mediated tissue-repair, in the context of critical differences in immune responses that exist between the sexes and contribute to the male-bias in development of severe COVID-19.

Sex differences in SARS-CoV-2 entry ACE2 has been described as the principal receptor enabling SARS-CoV-2 to enter human cells through binding of the spike (S) glycoprotein ( Figure 1 ) (Hoffmann et al., 2020; Zhou et al., 2020a) , and blocking these interactions using antibodies against ACE2 or soluble human ACE2 can prevent infection of human cells (Monteil et al., 2020; Wrapp et al., 2020) . Differences in expression of ACE2 in the nasal epithelium of small children and adults have been linked to a reduced risk of children to present with severe COVID-19 (Bunyavanich et al., 2020) , suggesting that lower ACE2 expression levels might be protective. ACE2 is encoded by the X chromosome, but studies have demonstrated that ACE2 expression is lower in lung tissues of women compared to men (Tukiainen et al., 2017) , and it has been suggested that estrogen downregulates the expression of ACE2 (Liu et al., 2010) . Furthermore, circulating plasma concentrations of ACE2 are higher in men with heart failure than in women (Sama et al., 2020) and in sputum cells of men compared to women (Peters et al., 2020) . These data suggest that differences in ACE2 amounts between women and men can translate into differences in the development of severe COVID-19, emphasizing the clinical promise of interventions that reduce binding of SARS-CoV-2 to ACE2, including antibodies directed against the ACE-2-binding site. More efficient entry of SARS-CoV-2 into cells from men might be furthermore facilitated by higher expression of the mucosa-specific serine protease TMPRSS2 that promotes virus entry into human host cells (Hoffmann et al., 2020; Zang et al., 2020) . TMPRSS2 expression has been suggested to be enhanced by androgens (Stopsack et al., 2020; ; and enhanced expression in male sputum cells has been described (Peters et al., 2020) . Furthermore, androgen-deprivation therapies for prostate cancer have been suggested to partially protect patients from SARS-CoV-2 infection in a population-based study from Italy (Montopoli et al., 2020) , although data on TMPRSS2 expression were not included in that study. These data suggest that sex differences in the expression of molecules involved in SARS-CoV-2 entry might facilitate the spread of SARS-CoV-2 in men. However, the role of ACE2 in SARS-CoV-2 infection is likely more complex (Table 1) . While being high jacked by SARS-CoV-2 as an entry receptor, ACE2 has important anti-inflammatory properties by catalyzing angiotensin II, resulting in the production of Ang1-7 that can ameliorate ARDS (Oudit and Pfeffer, 2020) . ACE2 mRNA expression is upregulated by type I IFNs, and ACE2 has been suggested to serve as an interferon-stimulated gene (ISG) during respiratory infections that promotes lung airway function (Ziegler et al., 2020) . Future studies are needed to determine whether ACE2 expression in lung tissues differs between women and men during SARS-CoV-2 infection, and whether potentially higher ACE2 expression in infected women, resulting from a stronger ISG-response (Chang et al., 2013; Meier et al., 2009; Ziegler et al., 2020) , might translate into beneficial anti-inflammatory effects mediated through Ang1-7 in COVID19. Sex differences in ACE2 expression might therefore have different effects between the sexes at different stages of SARS-CoV-2 infection (Table 1) . While lower estrogen-mediated baseline expression of ACE2 might reduce spreading of SARS-CoV-2 beyond the initial infection of the upper respiratory tract, higher interferoninduced ACE2 expression in women during progressive SARS-CoV-2 infection of the lower respiratory tract might protect from ARDS. The potential beneficial effect of clinical interventions currently tested in patients with severe COVID-19, such as the administration of estrogens to men or the addition of IFNβ to antiviral interventions, will therefore critically depend on the stage of the disease, and also might result in different effects between the sexes that need to be considered in the design of these studies, including sample size calculations.

In general, women develop stronger innate and adaptive immune responses against most viral infections and vaccines (Flanagan et al., 2017; Klein and Flanagan, 2016) , and significant sex differences in the induction of type I IFN responses, B cell functions and T cell functions have been described. Type I IFN responses that are critical for restriction of viral replication but also induction of antiviral immune responses, are differentially regulated between men and women ( Figure 1 ) (Ziegler and Altfeld, 2017) . Infections by RNA viruses, such as SARS-CoV-2, are sensed by Toll-like receptors (TLRs), including TLR7, resulting in the induction of IFNβ and IFNα production by pDCs (Heil et al., 2004) . TLR7 is encoded by the X chromosome, and studies have shown that the gene encoding for TLR7 can escape inactivation of the second X chromosome in women, resulting in higher TLR7 expression in immune cells of women compared to men (Souyris et al., 2018) . Several studies have demonstrated that plasmacytoid dendritic cells (pDCs) from women produce more IFNα/β in response to TLR7 ligands, including viral RNAs, resulting in higher induction of ISG in women (Berghofer et al., 2006; Chang et al., 2013; Meier et al., 2009; Seillet et al., 2012) . Both X chromosomal factors and sex hormones have been implicated in these sex differences in type I IFN responses (Griesbeck et al., 2015; Laffont et al., 2014; Scheuplein et al., 2015; Seillet et al., 2012) . Coronaviruses, including MERS-CoV and SARS-CoV, can induce type I IFN responses through TLR7 (Karnam et al., 2012; Li et al., 2016; Li et al., 2013; Scheuplein et al., 2015) , and SARS-CoV and SARS-CoV-2 have been shown to be sensitive to viral restriction by ISGs in vivo and in vitro (Haagmans et al., 2004; Mantlo et al., 2020) . The first studies assessing type I IFN responses induced by SARS-CoV-2 infections suggest induction of ISGs by SARS-CoV-2 (Blanco- Melo et al., 2020; Chu et al., 2020; Zhou et al., 2020b ), but to a lower level than observed in SARS-CoV infection (Blanco-Melo et al., 2020; Chu et al., 2020) , indicating a potential immune evasion mechanism. Remarkably, the presence of genetic loss-of-function variants of the X-chromosomal TLR7 was observed in 4 men with severe COVID-19 (van der Made et al., 2020), further highlighting the critical role of TLR7 and type I IFNs in SARS-CoV-2 pathogenesis. These studies have provided rationale for the treatment of COVID-19 patients with IFNs (O'Brien et al., 2020; Park and Iwasaki, 2020; Sallard et al., 2020) . Results from a retrospective study suggest that therapeutic interventions using IFNα early in SARS-CoV-2 infection can reduce mortality, while IFNα administration during late-stage sever COVID-19 was associated with increased mortality . However, significantly fewer women were included in the groups of patients receiving IFNα , representing a potential bias in the interpretation of the clinical outcomes. Some studies have furthermore reported that SARS-CoV-2 might be detected for longer in nasal swabs of men compared to women (Xu et al., 2020; Zheng et al., 2020) , suggesting a reduced ability to restrict viral replication and enhanced risk for transmitting the virus in men. Enhancing type I IFN-mediated restriction of viral replication, for example by subcutaneous administration of IFNβ, might therefore represent an early intervention with particular benefit for men that needs to be evaluated in randomized controlled clinical studies that take the sex differences in type I IFN responses into account.

The development of antibodies against SARS-CoV-2, including neutralizing antibodies, has been shown in SARS-CoV-2-infected persons and rhesus macaques Kreer et al., 2020; Ni et al., 2020; Robbiani et al., 2020; Rogers et al., 2020; Wolfel et al., 2020; Zhao et al., 2020) . SARS-CoV-2-specific, but also SARS-CoV-specific antibodies Pinto et al., 2020) , can block infection and prevent disease manifestations in rhesus macaques Yu et al., 2020) , and rhesus macaques receiving experimental vaccines that induce antibodies against SARS-CoV-2 infection are protected from disease . Furthermore, potential clinical benefits in COVID-19 patients receiving adoptive transfer of antibodies from convalescent plasma of SARS-CoV-2-infected individuals have been described (Shen et al., 2020a) , indicating that antibodies against SARS-CoV-2 might provide a clinical benefit in severe COVID-19 and prevent infection in vaccinated individuals (Figure 1 ). Multiple studies have demonstrated that women develop more rapid and stronger antibody responses to infections and vaccinations, and have implicated sex hormones and X chromosomal factors into these sex differences in antibody responses against influenza virus (Flanagan et al., 2017; Klein and Flanagan, 2016) . Studies in mice have demonstrated that estrogens promote and testosterone can suppress the development of antibodies (Fink et al., 2018; Flanagan et al., 2017; Klein and Flanagan, 2016) , and studies in humans described lower immune responses to influenza vaccination in men compared to women, and in particular in men with high levels of testosterone at the time of vaccination (Furman et al., 2014) . The results from these studies demonstrate an important role of sex hormones in mediating sex differences in antibody responses. More recent studies have however also implicated X chromosomal factors, including higher expression of TLR7 in B cells from females due to escape from X chromosome inactivation (XCI) that resulted in the enhanced propensity to IgG class switch in females (Souyris et al., 2018) . Other genes located on the X chromosome encoding for proteins that play an important role in the regulation of antibody responses, including CD40L and BTK, can also escape XCI (Tukiainen et al., 2017) , and might contribute to better induction and maintenance of antibody responses in women. This is further supported by studies demonstrating sex differences in T follicular helper (Tfh) cells that support B cell maturation, including higher IL-21 and IL-27 expression in females (Dimitrijevic et al., 2020) , and an increase in circulating Tfh cells at the time of induction of plasma cells was recently described in a case study of a woman with non-severe COVID-19 (Thevarajan et al., 2020) . Finally, the ability to induce or maintain antibody responses further decreases with age, in particular in men (Marquez et al., 2020) . These data strongly suggest that women have an immunological advantage in developing antibody responses against SARS-CoV-2. These sex differences might translate into clinical benefits for women, as early antibody signatures have been associated with COVID-19 outcome (Atyeo et al., 2020) (Shen et al., 2020b) , suggesting that they can curb the initial infection and prevent further spreading of the virus. This female bias has to be considered when clinical studies aimed at inducing SARS-CoV-2specific antibodies by vaccination are designed, both in the evaluation of potential adverse events as well as immunological and clinical endpoints (Flanagan et al., 2017). Given the described relatively short persistence of antibodies directed against SARS-CoV (Cao et al., 2007; Liu et al., 2006) , and a significantly more rapid decline of these antibody titers in men (Liu et al., 2006) , differences between women and men need to be in particular accounted for in studies investigating the persistence of antibody responses directed against SARS-CoV-2, and their ability to prevent infection or re-infection. (Polanczyk et al., 2004; Tai et al., 2008) . Furthermore, the gene encoding for FoxP3, the transcription factor that determines Treg cell differentiation, is expressed on the X chromosome, as is the gene encoding for CD40L that is involved in CD4+ T cell-mediated help for B cells (Tukiainen et al., 2017) . Enhanced ability of T cells from women to reduce excessive immune activation and to promote B cell differentiation might therefore contribute to the less frequent development of severe COVID-19 in women (Figure 1) . However, recent studies have shown that males have higher Treg numbers than females (Vasanthakumar et al., 2020) , and additional studies assessing the implication of Treg cells in SARS-CoV-2 pathogenesis are required. Furthermore, data regarding the role of T cells in SARS-CoV-2 pathogenesis are very limited. CD4+ and CD8+ T cell populations tend to be activated and reduced in the peripheral blood of patients with COVID-19 (Chen et al., 2020; Kuri-Cervantes et al., 2020; Mathew et al., 2020; Qin et al., 2020) , potentially due to recruitment into affected tissues, and the degree of T cell reduction has been associated with the severity of COVID-19 (Chen et al., 2020; Kuri-Cervantes et al., 2020; Mathew et al., 2020; Qin et al., 2020) . In addition, a reduction in Treg cells has been described in severe COVID-19 cases (Chen et al., 2020; Qin et al., 2020) . SARS-CoV-2-specific CD4+ and CD8+ T cells have been detected in COVID-19 patients Le Bert et al., 2020; Weiskopf et al., 2020) , but little is known regarding their antiviral activity, and no data on differences between women and men have been included in these first reports. Furthermore, neither CD4+ nor CD8+ T cell responses directed against SARS-CoV-2 showed a correlation with protection from disease in vaccinated or re-convalescent rhesus macaques Yu et al., 2020) . The sparsity of data emphasizes the need for additional studies to determine the role of T cell-mediated immunity in SARS-CoV-2 infection, and such studies need to take differences in T cell responses between the sexes into account, and report data in the context of the sex of the study subjects.

In addition to their direct antiviral activities, cells of the immune system can also shape local host defense mechanisms exerted by epithelial cells (Vivier et al., 2018) . Key to the prevention of severe COVID-19 is the containment of SARS-CoV-2 to the upper respiratory tract. Mucosal tissues employ the so called "weep and sweep" response to contain and remove pathogens (Gause et al., 2020) . Respiratory epithelial cells can secrete fluid that contains antimicrobial products and sweep infected cells or pathogens caught in the mucus up to the oral cavity where it can be removed. This "weep and sweep" response is enhanced by cytokines such as IL-4, IL-5 and IL-13, which are secreted by CD4+ Th2 cells and innate lymphocytes (ILCs) type 2, referred to as type 2 immunity (Gause et al., 2020) (figure 1). Although reduced levels of IL-13 have been detected in COVID-19 patients compared to healthy controls other studies suggest IL-13 to be increased in severe COVID-19 (Lucas et al., 2020; Schultheiss et al., 2020) . Type 2 immunity is more pronounced in women (Laffont et al., 2017a) who also suffer more frequently from diseases linked to dysregulated type 2 immunity, including allergies and asthma (Holgate et al., 2015) . An analyses of multiple studies showed an overall low prevalence of pre-existing respiratory diseases in COVID-19 patients (de Lusignan et al., 2020; Guan et al., 2020; Lupia et al., 2020) , which is in line with reports from previous coronavirus epidemics ( Moni and Lio, 2014) . However, in a recent study, patients with severe COVID-19 had suffered from a recent asthma exacerbation requiring treatment, suggesting that preexisting respiratory illnesses can predispose to severe COVID-19 (Bhatraju et al., 2020) . Upon damage signals, cytokines secreted by Th2 and ILC2s promote epithelial cell turnover in general and in particular that of secretory cells such as goblet cells (Gause et al., 2020) , which are increased in the female respiratory tract and patients with asthma (Hayashi and Huber, 1977; Kuperman et al., 2005) . Sex hormones can impact ILC2 numbers, as androgens reduce ILC2 numbers and cytokine production in male mice (Laffont et al., 2017b; Ricardo-Gonzalez et al., 2018) . In addition, the receptor for IL-13 (IL-13RA1) is encoded by the X chromosome, providing women a gene dosage advantage upon escape from XCI (Tukiainen et al., 2017) , potentially resulting in enhanced IL-13-signaling in epithelial cells (figure 1). A recent study showed that IL-13 decreased the expression of ACE2 in epithelial cells in vitro, and respiratory epithelial cells of asthma patients with enhanced type 2 signaling exhibited reduced ACE2-expression compared to healthy controls (Peters et al., 2020) . Together these data provide potential mechanisms by which sex differences in ILC2 and Th2 cells and the IL-13-IL-13R axis mediate a stronger airway "weep and sweep" response in women that can impact early stages of SARS-CoV-2 infection.

Immune cells furthermore have a critical role beyond host defense in regulating tissue regeneration (Karin and Clevers, 2016; Vivier et al., 2018) , ranging from ILCs, Th17 and Th22 cells maintaining mucosal integrity (Chung et al., 2012) to NK cells and macrophages shaping the architecture of the placenta (Vento-Tormo et al., 2018) . Frequencies of IL-22-J o u r n a l P r e -p r o o f and IL-17-producing RORɣt+ CD4+ T cells are enhanced in women compared to men, and have been shown to be regulated by sex hormones (Fuseini et al., 2019; Fuseini et al., 2018; Newcomb et al., 2015; Sankaran-Walters et al., 2013) . IL-22 promotes epithelial stem cell proliferation via STAT3 signaling (Lindemans et al., 2015) , contributing to tissue resilience upon damage. Protective effects of IL-22 have also been demonstrated in several viral infections including influenza, where IL-22 reduces inflammation, protects against bacterial superinfections and promotes lung epithelial repair (Pociask et al., 2013) . Amphiregulin produced by ILC2s, is a member of the epidermal growth factor family and promotes tissue repair (figure 1), resulting in improved barrier integrity and lung function in influenza-infected mice (Monticelli et al., 2011) . Amphiregulin is also produced by Treg cells and together with IL-10 and transforming growth factor-β allows Treg cells to promote tissue repair (Forbes and Rosenthal, 2014; Zaiss et al., 2006) Treg cells have been described to ameliorate severe complications in influenza infection in mice (Egarnes and Gosselin, 2018; Moser et al., 2014; Oliphant et al., 2015) . These data suggest that immune cell profiles in women are skewed towards tissue repair responses compared to men (figure 1), potentially contributing to reduced COVID-19 associated morbidity. However amphiregulin is also produced by epithelial cells and expressed at higher levels in male mice in an animal model with enhanced morbidity in influenza-infected female mice (Vermillion et al., 2018) . Furthermore, cell intrinsic non-immune-mediated tissue regeneration has been described to be better in men (Grishina et al., 2014; Joseph and Dyson, 1965; Kadel and Kovats, 2018) , indicating that sex-biased tissue repair pathways both in epithelial and immune cells can contribute to the different outcomes of respiratory infections between the sexes, and need to be included in models assessing SARS-CoV-2 pathogenesis.

The dual role of immune cells in host defense and tissue repair have shaped dimorphic immune responses between the sexes to maximize reproductive success in females, providing women with a superior capacity for immune-mediated tissue repair. An enhanced utilization of these immune-mediated tissue repair pathways might therefore provide women with better resilience upon SARS-CoV-2 infection and protection from development of severe COVID-19. These observations provide a rationale to test interventions that promote tissue resilience in clinical studies. Treatment strategies are available to antagonize androgenmediated effects, which can promote protective ILC2 and Th22 cell responses when initiated early during SARS-CoV-2 infection. Recombinant amphiregulin has shown beneficial effects in influenza-infected mice, and IL-13 has been proposed as a therapeutic intervention to reduce inflammation and promote tissue repair in preclinical models of stroke (Kolosowska et al., 2019) . However, given the pro-inflammatory role of cytokines in immune-mediated diseases such as Crohns disease and rheumatoid arthritis, as well as enhanced levels detected in COVID-19 patients (Lucas et al., 2020; Mateen et al., 2019; Shah et al., 2018) , the timing of initiation of immune-therapies in SARS-CoV-2 infection will be critical to promote viral containment and tissue resilience without contributing to the cytokine release syndrome in the advanced stages of COVID-19.

The female immune system has evolved to optimize antiviral immune responses protecting the unborn or newborn infant and to enable cyclic promotion of tissue development and regeneration required for reproduction. This has resulted in differences in immune functions between women and men, which are manifested by stronger immune responses in women against pathogens and vaccines, but also in higher susceptibility to autoimmune diseases. The emerging understanding of the mechanisms underlying these sex differences in immune functions provide important insights into protective as well as disease-promoting pathways during SARS-CoV-2 infection, which can be targeted therapeutically. Interventions aimed at enhancing antiviral mechanisms, reducing excessive inflammation and promoting tissueprotective immune functions during COVID-19 have to take these sex specific differences into account to optimize treatment outcomes for women and men. Furthermore, sex has to J o u r n a l P r e -p r o o f be taken into account as a critical variable in the analysis of immunological and clinical data from COVID-19 patients to better understand the biology of the disease.

J o u r n a l P r e -p r o o f Sex-specific differences in immune-mediated antiviral mechanisms are described on the left and in tissue repair mechanisms on the right. Left: SARS-CoV-2 binds to ACE2 receptors to gain entrance into cells. SARS-CoV-2 can be sensed by TLR7 and TLR8, and induce the production of type I IFNs by pDCs. Tfh cells provide help through cytokines and CD40/CD40L to B cells that produce neutralizing antibodies against SARS-CoV-2. These antiviral mechanisms are stronger in females, and might be partially regulated by higher expression of genes encoded by the X chromosome, including TLR7, TLR8 and CD40L. Right: Th22 cells and ILC2s produce cytokines (IL-22, IL-5, IL-13and AREG) that promote lung airway epithelial cell growth and function. Stronger Treg and ILC2 function as well as higher expression of the IL-13R in women might promote lung airway tissue resilience and function in SARS-CoV-2-infected women. Molecules encoded by genes located on the X chromosome are shown in red, including ACE2, TLR7, TLR8, CD40L and IL-13R. Wedges on the bottom of the figure represent overall higher immune-mediated antiviral and tissue repair mechanisms in women compared to men. genes in HIV-1-infected females after adjusting for the level of viral replication. The Journal of infectious diseases 208, 830-838. Chen, G., Wu, D., Guo, W., Cao, Y., Huang, D., Wang, H., Wang, T., Zhang, X., Chen, H., Yu, H., et al. (2020) Gause, W.C., Rothlin, C., and Loke, P. (2020). Heterogeneity in the initiation, development and function of type 2 immunity. Nature reviews. Immunology.

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Tissue signals imprint ILC2 identity with anticipatory function

Convergent antibody responses to SARS-CoV-2 in convalescent individuals

Elevated 17beta-estradiol protects females from influenza A virus pathogenesis by suppressing inflammatory responses

Isolation of potent SARS-CoV-2 neutralizing antibodies and protection from disease in a small animal model

Estimating the burden of SARS-CoV-2 in France

Circulating plasma concentrations of angiotensin-converting enzyme 2 in men and women with heart failure and effects of renin-angiotensin-aldosterone inhibitors

Sex differences matter in the gut: effect on mucosal immune activation and inflammation

High secretion of interferons by human plasmacytoid dendritic cells upon recognition of Middle East respiratory syndrome coronavirus

Next-Generation Sequencing of T and B Cell Receptor Repertoires from COVID-19 Patients Showed Signatures Associated with Severity of Disease

Considering how biological sex impacts immune responses and COVID-19 outcomes

The TLR-mediated response of plasmacytoid dendritic cells is positively regulated by estradiol in vivo through cell-intrinsic estrogen receptor alpha signaling

Sex-Based Differences in Incidence of Inflammatory Bowel Diseases-Pooled Analysis of Population-Based Studies From Western Countries

Treatment of 5 Critically Ill Patients With COVID-19 With Convalescent Plasma

Delayed specific IgM antibody responses observed among COVID-19 patients with severe progression

TLR7 escapes X chromosome inactivation in immune cells

Male offspring born to mildly ZIKV-infected mice are at risk of developing neurocognitive disorders in adulthood

TMPRSS2 and COVID-19: Serendipity or Opportunity for Intervention? Cancer discovery

SARS-CoV-2 entry factors are highly expressed in nasal epithelial cells together with innate immune genes

Induction of regulatory T cells by physiological level estrogen

COVID-19: the vasculature unleashed

Breadth of concomitant immune responses prior to patient recovery: a case report of non-severe COVID-19

Landscape of X chromosome inactivation across human tissues

Presence of Genetic Variants Among Young Men With Severe COVID-19

Sex-specific adipose tissue imprinting of regulatory T cells

Single-cell reconstruction of the early maternal-fetal interface in humans

Production of amphiregulin and recovery from influenza is greater in males than females

Innate Lymphoid Cells: 10 Years On

Sex and sex steroids impact influenza pathogenesis across the life course

Function, and Antigenicity of the SARS-CoV-2 Spike Glycoprotein

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is likely to be androgen mediated

Androgen sensitivity gateway to COVID-19 disease severity

Retrospective Multicenter Cohort Study Shows Early Interferon Therapy Is Associated with Favorable Clinical Responses in COVID-19 Patients

Phenotype and kinetics of SARS-CoV-2-specific T cells in COVID-19 patients with acute respiratory distress syndrome

Autopsy Findings and Venous Thromboembolism in Patients With COVID-19

Virological assessment of hospitalized patients with COVID-2019

Cryo-EM structure of the 2019-nCoV spike in the prefusion conformation

Factors associated with prolonged viral RNA shedding in patients with COVID-19

DNA vaccine protection against SARS-CoV-2 in rhesus macaques

TMPRSS2 and TMPRSS4 promote SARS-CoV-2 infection of human small intestinal enterocytes

Clinical Characteristics of Covid-19 in China

Epidemiology of Covid-19. The New England journal of medicine 382

Antibody responses to SARS-CoV-2 in patients of novel coronavirus disease 2019

Viral load dynamics and disease severity in patients infected with SARS-CoV-2 in Zhejiang province, China

Heightened Innate Immune Responses in the Respiratory Tract of COVID-19 Patients

SARS-CoV-2 Receptor ACE2 Is an Interferon-Stimulated Gene in Human Airway Epithelial Cells and Is Detected in Specific Cell Subsets across Tissues

Human Immunodeficiency Virus 1 and Type I Interferons-Where Sex Makes a Difference

J o u r n a l P r e -p r o o f • Highlights:• Men present more frequently with severe COVID-19 • Sex-differences in immunity contribute to better control of SARS-CoV-2 in women • Immune-mediated tissue repair mechanisms differ between the sexes

Clinical manifestations of SARS-CoV-2 infection differ between women and men, with men presenting more frequently with severe COVID-19. We review the impact of sex differences in immunity for these differences in COVID-19, and discuss the implications for clinical interventions.