Abstract 16446: Human Mesenchymal Stem Cells of Adult and Fetal Origin Do Not Express ACE2 and are Immune to SARS-CoV-2 Spike Pseudotyped Virus

Abstract

Background: Angiotensin-converting enzyme 2 (ACE2) expression in the lung has a protective role against injury. However, ACE2 is the main host cell receptor for the Severe Acute Respiratory Syndrome-Coronavirus 2 (SARS-CoV-2) entry. Human mesenchymal stem cells (hMSCs) are currently under investigation for the treatment of pulmonary and cardiac complications of the SARS-CoV-2 disease (COVID-19). However, it is unclear if hMSCs express ACE2 and if they possibly can secrete ACE2. Also, their predisposition to be infected by SARS-CoV-2 is unknown. Aim: To assess if hMSCs of fetal and adult origin constitutively express and secrete ACE2 and if they can be infected by SARS-CoV-2. Methods: We studied 8 hMSC lines: 4 of fetal origin isolated from human placentas (hA-MSC) and 4 of adult origin isolated from bone marrow aspirates (hBM-MSCs). The lung epithelial cancer cell line CALU-3, which expresses high levels of ACE2, was used as positive control. ACE2 expression was evaluated in both standard culture conditions and after 48 hrs of hypoxia exposure (1% O 2 ). We quantified the mRNA levels by RT-qPCR and verified protein content by western blot on both cell lysates and serum-free 48 hrs-conditioned media (MSC-CM). ACE2 levels in MSC-CM were quantified by ELISA assay. Finally, to test MSC viral susceptibility, we produced replication-defective, GFP-tagged retroviral particles bearing the SARS-CoV-2 Spike protein; the pantropic VSV glycoprotein (VSV-G) was used as positive control. Results: ACE2 mRNA level in both fetal and adult MSC was 200-fold lower than in CALU-3. ACE2 protein expression was undetectable in MSC lysates and CM, both by western blot and ELISA. Hypoxia slightly increased ACE2 mRNA level, but not protein level, which still was undetectable. Finally, CALU-3 but not MSC were infected by SARS-CoV-2 Spike pseudovirus, whereas both cell types were susceptible to VSV-G pseudovirus infection. Conclusions: Both fetal and adult MSC do not express significant levels of ACE2 both under normoxia and hypoxia, and therefore the putative therapeutic mechanism of hMSCs in COVID-19 cannot go through the release of ACE2. Importantly, MSCs are immune to SARS-CoV-2 infection and therefore their use appears safe.