Leukemia | 2021
An evidence that SARS-Cov-2/COVID-19 spike protein (SP) damages hematopoietic stem/progenitor cells in the mechanism of pyroptosis in Nlrp3 inflammasome-dependent manner
Abstract
TO THE EDITOR: Mounting evidence accumulates that hematopoietic stem/progenitor cells (HSPCs) and endothelial progenitor cells (EPCs) are damaged during severe SARS-Cov-2/COVID-19 infection [1, 2]. It has been reported that patient infected with COVID-19 are frequently presented with anemia, lymphopenia, and thrombocytopenia [1–3]. This negative effect of the virus on human hematopoiesis and endothelium has been reported in infected patients and demonstrated in vitro after exposure of cells to SARS-Cov-2/COVID19 spike protein (SP) [1, 3, 4]. It is known that virus may enter cells and, directly in case of productive infection, lead to their irreversible damage. On the other hand, the interaction of viral SP with some of the receptors expressed on the cell surface may lead to their damage as well [1–3]. We have proposed that interaction of SP with the target cell surface receptors induces intracellular hyperactivation of Nlrp3 inflammasome which may lead to cell death by pyroptosis [5]. It is known that pyroptosis is characterized by the creation in a caspase-1 dependent manner of N-gasdermin pores in the cell membrane, which leads to the release of cytosol components to extracellular space and final cell lysis [6]. As reported, SARS-CoV-2/COVID-19 enters human cells after binding to the angiotensin-converting enzyme 2 (ACE2) receptor utilizing SP for attachment and subsequent internalization. Moreover, transmembrane protease 2 (TMPRSS2) cleavage of SP may augment viral entry [7]. This facilitates its interaction with ACE2 and the subsequent fusion of viral and cellular membranes. The other receptor postulated to be involved in virus entry is toll-like receptor-4 (TLR4) [8]. Thus, this virus is using cell surface receptors that play a physiological role in the conversion of angiotensin II to angiotensin (1–7) (Ang [1–7]) as it is a case of ACE2 and TLR4 which belongs to pattern recognition cell surface receptor family and is responsible for inflammatory cytokine production and activation of the innate immunity responses. As demonstrated, both ACE2 and TLR4 are highly expressed on HSPCs and EPCs [3, 4]. In addition to these two receptors, SARS-CoV-2/COVID-19 may also interact with the extracellular matrix metalloproteinase inducer basigin, known as cluster of differentiation 147 (CD147) [9] as well as recently proposed with C-type lectin receptor and Tweety family member 2 [10]. In our recent perspective paper in Leukemia, we proposed that the adverse effects of infection on stem cell compartments result from the uncontrolled hyperactivation of the Nlrp3 inflammasome [5]. To support this, we noticed that exposure of human UCBpurified CD34linCD45 HSCs to recombinant SP for 16 h lead to upregulation of mRNA expression for Nlrp3 inflammasome [4]. As determined by ELISA, we also detected elevated levels of IL-1β in the conditioned media (CM) from cells exposed to SP [4]. Release from cells of IL-1β in a caspase-1-dependent manner is an important indicator of Nlrp3 inflammasome activation. This Letter to Editor reports that human CD34 cells enriched for HSPCs and human CD34CD133CD31CD144 cells enriched for EPCs activate Nlrp3 inflammasome after exposure to SP as evidenced by measuring the cytosolic activity of caspase-1 by employing functional sensitive bioluminescent glow assay. Fig. 1A shows that caspase-1 becomes activated in HSPCs in response to SP, and Fig. 1B shows that the same phenomenon occurs in EPCs. Moreover, this activation has been inhibited after preincubation of SP with human recombinant ACE2 protein (rhACE2). Interestingly, activation of Nlrp3 inflammasome was higher in EPCs as compared to HSCs. Next, we exposed human CD34 cells to SP in the absence or presence of Nlrp3 inflammasome-specific inhibitor MCC950 (Fig. 1C, D) and plated these cells in methylcellulose cultures to grow CFU-Mix and CFU-GM colonies. As demonstrated, exposure to SP decreased clonogeneic growth of CFU-Mix and CFU-GM, and inhibition of Nlrp3 inflammasome by MCC950 reversed this effect. In the next set of experiments, we become interested in which receptors responsible for SP binding are accountable for the