The ACE2 Receptor for COVID-19 Entry Is Expressed on the Surface of Hematopoietic Stem/Progenitor Cells and Endothelial Progenitors As Well As Their Precursor Cells and Becomes Activated in Nlrp3 Inflammasome-Dependent Manner By Virus Spike Protein - a Potential Pathway Leading to a "Cytokine Storm"

Abstract

Background . The SARS-CoV-2 pandemic, with its high mortality, has become an urgent clinical problem. It is well established that SARS-CoV-2 enters human cells after binding to the angiotensin-converting enzyme 2 (ACE2) receptor and utilizes a spike protein (S) for attachment and entry into the cells, leading to their lysis or damage. This infection damages several organs, including lungs, heart, blood vessels, kidneys, and intestines and may lead to a fatal complication known as a "cytokine storm", which is the result of uncontrolled hyperactivation of the innate immunity-initiated response and the release of several pro-inflammatory cytokines. An important potential aspect of SARS-CoV-2 infection is damage to the stem cell compartment, which may lead to severe complications from the infection. The ACE2 receptor has been described as being expressed on the surfaces of certain types of stem cells, including specified hematopoietic stem cells (HSCs) and endothelial progenitor cells (EPCs). These cells could be potentially damaged and lysed after virus entry or could undergo pyroptosis due to hyperactivation of Nlrp3 inflammasomes. Supporting this possibility, we recently reported that all Nlrp3 inflammasome components are expressed in HSCs (Adamiak, M et al. Nlrp3 Inflammasome Signaling Regulates the Homing and Engraftment of Hematopoietic Stem Cells (HSPCs) by Enhancing Incorporation of CXCR4 Receptor into Membrane Lipid Rafts. Stem Cell Rev and Rep (2020).https://doi.org/10.1007/s12015-020-10005-w). Hypothesis. We hypothesized that one of the triggers of a cytokine storm could be interaction of the ACE2 receptor with the SARS-CoV-2 spike protein, leading to hyperactivation of Nlrp3 inflammsomes in target cells including population of stem cells.Materials and Methods. Experiments were performed on human stem cells at different levels of specification, including very small CD34+CD133+lin-CD45- cells, which may become specified into HSCs and EPCs, as well as human CD34+Lin-CD45+ HSCs and CD34+ CD133+ KDR+ CD31+ EPCs. These cells were phenotyped for expression of ACE2 and the SARS-CoV-2 entry-facilitating transmembrane protease TMPRSS2 at the mRNA level and by FACS at the protein level. Next, we exposed these cells to the NCP-CoV (2019-nCoV) spike protein (S1+S2 ECD, expressed with a His-tag; Sino Biological) at a concentration of 10 nM. After 16 h of incubation, the cells were lysed, and total RNA was isolated for qRT-PCR analysis of Nlrp3 and essential Nlrp3 inflammasome components, including ASC, caspase 1, IL1b, and IL18. In some experiments, UCB-derived HSCs were plated into 96-well plates and stimulated with NCP-CoV (2019-nCoV) spike protein, as described above, alone or together with angiotensin 1-7 or the anti-inflammatory heme oxygenase 1 (HO-1) activator CoPP. Results. First, we observed that the ACE2 receptor and SARS-CoV-2 entry-facilitating transmembrane protease TMPRSS2 are expressed by all types of stem cells evaluated in our studies. Moreover, we detected activation of Nlrp3 inflammasomes in response to viral spike protein. This activation was inhibited by exposure of the stimulated cells to angiotensin 1-7 or CoPP. Conclusions. We envision that, in addition to directly infecting target cells, virus can hyperactivate the Nlrp3 inflammasome in stem cells, which may trigger their pyroptosis. Therefore, since we still do not have an effective SARS-CoV-2 vaccine in hand, the results presented in our current work suggest that inhibition of the Nlrp3 inflammasome by the small-molecule inhibitor MCC950 or application of Nlrp3 inflammasome inhibitors, such as Ang (1-7) or heme oxygenase 1 activators, could find potential clinical application to prevent the onset of a cytokine storm and cell pyroptosis. Disclosures No relevant conflicts of interest to declare.

Stem Cells as Potential Therapeutics and Targets for Infection by COVID19 – Special Issue on COVID19 in Stem Cell Reviews and Reports

“…Therefore, more studies are needed to address if, in addition to ACE2, spike protein may also activate Nlrp3 inflammasome in TLR4 or CD147-dependent manner. A recent report supports this possibility in the case of TLR4 [7]. In conclusion let's hope that next months will bring answers to all these outstanding yet questions.…”

Hematopoietic responses to SARS-CoV-2 infection

“…The authors of this study have concluded that human bone marrow cells could be prone to SARS-CoV-2 infection. In support of this hypothesis, they have reported that different human stem cell lineages, such as CD34 + CD133 + lin − CD45 − cells, which may differentiate to HSCs and endothelial progenitor cells (EPCs), CD34 + Lin − CD45 + HSCs and CD34 + CD133 + KDR + CD31 + EPCs, express ACE2 and TMPRSS2 at the gene and protein levels [ 88 ]. In addition, it was observed that the viral spike protein activates NLrp3 inflammasomes in HSCs, which likely triggers their pyroptosis [ 88 ].…”

“…In support of this hypothesis, they have reported that different human stem cell lineages, such as CD34 + CD133 + lin − CD45 − cells, which may differentiate to HSCs and endothelial progenitor cells (EPCs), CD34 + Lin − CD45 + HSCs and CD34 + CD133 + KDR + CD31 + EPCs, express ACE2 and TMPRSS2 at the gene and protein levels [ 88 ]. In addition, it was observed that the viral spike protein activates NLrp3 inflammasomes in HSCs, which likely triggers their pyroptosis [ 88 ]. Post-mortem examination of COVID-19 patients has revealed evidence of direct viral infection of endothelial cells with accumulation of inflammatory cells.…”

“…Another possible mechanism of thrombocytopenia could be related to the invasion of HSCs by SARS-CoV-2. Because of the expression of ACE2 and TMPRSS2 by HSCs and their susceptibility to SARS-CoV-2 infection [ 88 , 90 ], it is possible to speculate that viral invasion and apoptosis of HSCs may suppress their differentiation into megakaryocyte progenitors. Moreover, cytokines and stem cell factors can influence the bone marrow niches thereby resulting in thrombocytopenia.…”

Therapeutic Targeting of Innate Immune Receptors Against SARS-CoV-2 Infection

“…The spike protein of SARS-CoV-2 has also been found to activate the NLRP3 inflammasome ( Kucia et al, 2020 ). In another research article, it is reported that in cultured murine alveolar macrophages, ultramicronized palmitoylethanolamide inhibits NLRP3 inflammasome expression and the pro-inflammatory response activated by the SARS-CoV-2 Spike protein ( Del Re et al, 2021 ).…”