key: cord-0844475-7h4t4xhv
authors: de Miguel-Gómez, Lucía; Romeu, Mónica; Castells-Ballester, Joan; Pellicer, Nuria; Faus, Amparo; Mullor, José Luis; Pellicer, Antonio; Cervelló, Irene
title: Undetectable viral RNA from SARS-CoV-2 in endometrial biopsies from women with COVID-19: a preliminary study
date: 2021-10-21
journal: Am J Obstet Gynecol
DOI: 10.1016/j.ajog.2021.10.019
sha: 2183b5276209cb20e5c924852a1966efc53a04ca
doc_id: 844475
cord_uid: 7h4t4xhv

nan

From its first appearance, the novelty of severe acute respiratory syndrome 2 coronavirus 2 (SARS-CoV-2), responsible for coronavirus disease 2019 (COVID-3 19), has generated many important questions about its effects on human health. 4 SARS-CoV-2 is a single-stranded enveloped RNA virus that incorporates three 5 types of transmembrane proteins [spike (S), envelope, and membrane] and a 6 nucleocapsid (N) structural protein. 1 For effective cell entry and consequent 7 infection, the S protein needs to specifically bind to cell-surface receptors such 8 as angiotensin-converting enzyme 2 (ACE2), which is thought to be a crucial 9 receptor in this process. S protein also requires priming by host cell serine 10 proteases such as transmembrane protease serine 2 (TMPRSS2). 2 11

Interestingly the gene encoding ACE2 is expressed in female reproductive 12 organs and cells, including the oocytes, ovaries, placenta, vagina, and uterus. 3 In 13

2009, expression of the ACE2 gene in the uterine mucous layer (endometrium) 14 was shown to be higher in epithelial cells (versus stromal cells) and during the 15 secretory phase (compared to the proliferative phase) of the menstrual cycle. 4 16 Protein expression analysis reaffirmed increased expression of ACE2 in the 17 secretory phase (when decidualization, which is essential for achieving 18 pregnancy, occurs); furthermore, stromal cells during this phase showed higher 19 expression of ACE2 than the epithelial compartments. 5 20

These studies underscore the importance of determining whether there is 21 sufficient ACE2 expression in the human endometrium to allow SARS-CoV-2 22

infection of this tissue. Because the endometrium is critical for reproduction, viral 23 infection could influence reproductive outcomes and potentially even impact 1 future fertility treatments. 2

The research ethics committee of Hospital Universitari i Politècnic La Fe 4 approved this study (registration number: 2020-268-1) on May 12, 2020. All study 5

participants received written and oral information on the study characteristics and 6 consented verbally in the presence of at least one witness. 7

Human tissue samples were obtained by endometrial aspiration (Cornier Pipelle) For statistical significance, one-tailed t-test was applied to log2−ΔΔCT. 23

We enrolled 15 women [mean age = 35.80 years, standard deviation (SD) = 7.49] 2 in different phases of the menstrual cycle, with a mean of 5 days (SD = 4.44) 3 between confirmed diagnosis of COVID-19 (positive nasopharyngeal RT-PCR; 4 data not available) and collection of endometrial biopsies (Table 1) . 5

All endometrial samples (except for patient seven, due to insufficient total RNA 6 available) tested negative for the virus (neither the N1 nor the N2 gene was 7 detected) ( Figure 1A ). ACE2 expression was detected in 10 of 14 endometrial 8 samples (excluding patient seven) ( Figure 1A ). 9

Comparison of ACE2 endometrial expression values to those detected in the 10

confirmed the receptor's low-level expression in the endometrium, and this 12 difference was statistically significant in 8 samples ( Figure 1B) . 13

Little is known about the impact of SARS-CoV-2 on the endometrium, and to our 15 knowledge, no study has assessed this tissue from patients with COVID-19. The 16 present study is the first to analyze such samples. Our findings indicate that, 17

despite the presence of the receptor ACE2 (proposed to be key for SARS-CoV-18 2 cell entry) 2 in the endometrium, its mRNA expression is insufficient to allow viral 19 infection (none of the analyzed endometrial biopsies revealed the presence of 20 viral mRNA). Previous findings in the human endometrium indicated differential 21 ACE2 expression across the menstrual cycle and in different endometrial cell 22 types. 4,5 In agreement with these reports, we could appreciate lower levels of 23 ACE2 expression in the proliferative versus secretory phase (with a notable trend 1 of high expression in the early secretory endometrium). In line with this, an in 2 silico study indicated that the endometrium is likely safe from SARS-CoV-2 3 infection based on a lack of several key molecular effectors, including ACE2 and 4 TMRPSS2. 6 These results align with another recently published in silico work 5 that describes the apparent low risk of infection of the endometrium due to the 6 low percentage of cells expressing ACE2. 7 Interestingly, the percentage of cells 7 co-expressing ACE2 and TMRPSS2 was even lower during the window of 8 implantation (secretory phase). 7 Together with our pilot results, these findings 9

indicate that the human endometrium seems safe (negative diagnostic test However, we note several limitations. Since samples were not easy to collect or 20 process, our population size was small. Additionally, due to health measures and 21 the infectious condition of the patients, we collected biopsies only from 22 symptomatic hospitalized patients (excluding asymptomatic women). We 

Structures and 2 distributions of SARS-CoV-2 spike proteins on intact virions

SARS-CoV-2 cell entry depends on ACE2 and TMPRSS2 and is 6 blocked by a clinically proven protease inhibitor

Potential influence of COVID-19/ACE2 on the female reproductive system

The vasoactive peptide angiotensin-(1-7), its receptor Mas 12 and the angiotensin-converting enzyme type 2 are expressed in the 13 human endometrium

Angiotensin converting enzyme 2 (ACE2) is required for human 16

SARS-CoV-2 infection risk assessment in the endometrium: 19 Viral infection-related gene expression across the menstrual cycle