Spike protein (inc vax) induced immunodeficiency & carcinogenesis megathread #3: SARS-CoV-2 Spike Protein Suppresses NK Cell Function
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Any extracts used in the following article are for non commercial research and educational purposes only and may be subject to copyright from their respective owners.
If your dendritic cells are suppressed they cannot present viral fragments to the T cells to trigger the full, early immune response. And healthy T & B cell broad spectrum memory cannot happen.
You know what else NK cells are good at? Targeting cancer cells...
"Taken together, these data indicate the ability of the SARS-CoV-2 SP1 to significantly downregulate NK cell activity in vivo via a heightened interaction between HLA-E and the inhibitory receptor CD94/NKG2A.”
"SARS-CoV-2 Spike Protein Suppresses NK Cell Function" (6/30/2020)
Much of the research thus far on the host immune response to SARS-CoV-2 infection has been geared toward the development of novel vaccines, and for this reason has focused largely on the roles of T and B cells in fighting infection. However, another class of immune cells, the so-called “natural killer” cells, are central to the response to viral infection as well. Natural killer (NK) cells are lymphocytes that are derived from the same progenitor cell line as T and B cells but are non-specific in effect and therefore part of the innate immune branch (Trinchieri, 1989). These cells play a large part in the recognition and elimination of virally infected cells and tumor cells, in which the Major Histocompatibility Complex class I (MHC I) molecules are often downregulated or mutated in an “effort” by the hijacked cell to avoid initiating an immune response. In the absence of MHC I on the surface of a host cell, the inhibition receptors on NK cells cannot be cross-linked, and the lymphocytes are subsequently activated against the pathological host cell(s) (Caligiuri, 2008; Long et al., 2013). The role of NK cells in fending against SARS-CoV-2 cannot be overstated and determining the virus’ interaction with these cells in vivo is an important next step in understanding the complete host immune response.
Bortolotti et al. investigated the interaction between the coronavirus spike proteins and NK cells in context of exhaustion of these lymphocytes. Previous work this year has demonstrated significant functional exhaustion of NK cells and CD8+ T cells in COVID-19 patients, mediated by the upregulation of the inhibitory receptor CD94/NKG2A (Zheng et al., 2020). In this study, peripheral blood natural killer cells were harvested from healthy control patients who tested negative for SARS-CoV-2, thus ensuring that these cells were entirely naïve for the spike proteins of the virus. These NK cells were cultured in the presence of either the SARS-CoV-2 spike protein domain 1 (SP1), SARS-CoV-2 spike protein domain 2 (SP2), or SARS-CoV spike protein (S), and their migration along a polycarbonate filter was measured. The authors determined that all three spike protein treatments induced significant migration compared to the control chemoattractant (CXCL12), confirming the ability of the spike protein to induce NK cell trafficking to sites of infection (Figure 1A). As a follow up, interferon-gamma (IFN-γ) expression was observed in the presence of each spike protein as a measure of NK cell activation: indeed, an increase in IFN-γ was noted in the presence of SP1, but not SP2 or S protein (Figure 1B).
Figure 1 – (A) SP1, SP2, and S protein significantly induced migration of NK cells compared to controls. (B) Interferon-gamma, a measure of NK cell activation, was markedly increased in the presence of SP1. (Bortolotti et al., 2020)
The authors then tested the effects of spike protein on the cytotoxicity of NK cells. To do so, SP1, SP2, and S protein were expressed in K562 cells, and the harvested NK cells were incubated with the K562 cells, after which expression of the molecule CD107a (a marker of NK cell degranulation) was observed. At the end of the incubation period, they noted a non-significant increase in CD107a expression for each of the three treatments compared to control K562 cells (Figure 2, top). Due to the respiratory nature of the virus, however, the experiment was repeated in the lung epithelial cell line Beas-2B. This time, the authors observed a significant decrease in CD107a in the cells expressing SP1; and a non-significant decrease in CD107a was observed in the SP2- and S-expressing cells (Figure 2, bottom). They concluded that impairment of CD107a expression indicates a reduction in the cytotoxic capacity of the NK cells due to interaction with SP1.
Figure 2 –(Top) CD107a was slightly increased, though not significantly, when NK cells were incubated with K562 cells transfected with SP1, SP2, or S protein. (Bottom) A significant decrease in CD107a expression was observed when NK cells were incubated with lung epithelial Beas-2B cells transfected with SP1, SP2, or S protein. (Bortolotti et al., 2020)
Finally, to determine the exact mechanism of the reduced cytotoxicity, the authors expressed SP1, SP2, and S in Beas-2B cells and stained them with anti-classical Human Leukocyte Antigen I antibodies (anti-HLA-A, anti-HLA-B, and anti-HLA-C). They observed a significant decrease in HLA I cell surface expression in Beas-2B cells that were transfected with SP1, indicating that SP1 has a large role in inhibiting the presentation of viral antigens by the host cells (Figure 3, top; see updates 6/15/2020 for discussion of MHC [HLA] I presentation). HLA-E is a “non-classical” HLA I molecule that is expressed on the surfaces of many cell types and interacts specifically with the CD94/NKG2A inhibitory receptors on NK cells. When the researchers stained the transfected Beas-2B cells with anti-HLA-E antibody, they noted a significant increase in HLA-E surface expression in the presence of SP1, but not SP2 or S (Figure 3, middle). Lastly, when NK cells were incubated with SP1-transfected Beas-2B cells, there was a significant increase in CD94/NKG2A expression on the NK cell surfaces (Figure 3, bottom). Taken together, these data indicate the ability of the SARS-CoV-2 SP1 to significantly downregulate NK cell activity in vivo via a heightened interaction between HLA-E and the inhibitory receptor CD94/NKG2A.
One limitation of this study is the fact that the subunits of the SARS-CoV-2 spike protein were only tested individually. It is not clear why the SP1 subunit should exert the observed effects on NK cells when the SP2 subunit does not. Further characterization of the function(s) of each subunit may shed light on the reason behind this disparity. It is not likely that the SP1 subunit is viable to initiate and sustain infection on its own, and it is likely not the case that the subunits of the spike protein would be encountered individually anyway. Future work should focus on determining the effects of the entire SARS-CoV-2 spike protein on NK cell cytotoxicity and exhaustion.
Figure 3 – (Top) In the presence of SP1, classical HLA I molecules were significantly downregulated on Beas-2B cells. (Middle) In the presence of SP1, non-classical HLA-E was significantly upregulated on Beas-2B cells. (Bottom) In the presence of SP1, the inhibitory receptor CD94/NKG2A was significantly upregulated on NK cells. (Bortolotti et al., 2020)
Spike protein (inc vax) induced immunodeficiency & carcinogenesis megathread #3: SARS-CoV-2 Spike Protein Suppresses NK Cell Function
Spike protein (inc vax) induced immunodeficiency & carcinogenesis megathread #3: SARS-CoV-2 Spike Protein Suppresses NK Cell Function
Spike protein (inc vax) induced immunodeficiency & carcinogenesis megathread #3: SARS-CoV-2 Spike Protein Suppresses NK Cell Function
Any extracts used in the following article are for non commercial research and educational purposes only and may be subject to copyright from their respective owners.
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If your dendritic cells are suppressed they cannot present viral fragments to the T cells to trigger the full, early immune response. And healthy T & B cell broad spectrum memory cannot happen.
You know what else NK cells are good at? Targeting cancer cells...
"Taken together, these data indicate the ability of the SARS-CoV-2 SP1 to significantly downregulate NK cell activity in vivo via a heightened interaction between HLA-E and the inhibitory receptor CD94/NKG2A.”
"SARS-CoV-2 Spike Protein Suppresses NK Cell Function" (6/30/2020)
Much of the research thus far on the host immune response to SARS-CoV-2 infection has been geared toward the development of novel vaccines, and for this reason has focused largely on the roles of T and B cells in fighting infection. However, another class of immune cells, the so-called “natural killer” cells, are central to the response to viral infection as well. Natural killer (NK) cells are lymphocytes that are derived from the same progenitor cell line as T and B cells but are non-specific in effect and therefore part of the innate immune branch (Trinchieri, 1989). These cells play a large part in the recognition and elimination of virally infected cells and tumor cells, in which the Major Histocompatibility Complex class I (MHC I) molecules are often downregulated or mutated in an “effort” by the hijacked cell to avoid initiating an immune response. In the absence of MHC I on the surface of a host cell, the inhibition receptors on NK cells cannot be cross-linked, and the lymphocytes are subsequently activated against the pathological host cell(s) (Caligiuri, 2008; Long et al., 2013). The role of NK cells in fending against SARS-CoV-2 cannot be overstated and determining the virus’ interaction with these cells in vivo is an important next step in understanding the complete host immune response.
Bortolotti et al. investigated the interaction between the coronavirus spike proteins and NK cells in context of exhaustion of these lymphocytes. Previous work this year has demonstrated significant functional exhaustion of NK cells and CD8+ T cells in COVID-19 patients, mediated by the upregulation of the inhibitory receptor CD94/NKG2A (Zheng et al., 2020). In this study, peripheral blood natural killer cells were harvested from healthy control patients who tested negative for SARS-CoV-2, thus ensuring that these cells were entirely naïve for the spike proteins of the virus. These NK cells were cultured in the presence of either the SARS-CoV-2 spike protein domain 1 (SP1), SARS-CoV-2 spike protein domain 2 (SP2), or SARS-CoV spike protein (S), and their migration along a polycarbonate filter was measured. The authors determined that all three spike protein treatments induced significant migration compared to the control chemoattractant (CXCL12), confirming the ability of the spike protein to induce NK cell trafficking to sites of infection (Figure 1A). As a follow up, interferon-gamma (IFN-γ) expression was observed in the presence of each spike protein as a measure of NK cell activation: indeed, an increase in IFN-γ was noted in the presence of SP1, but not SP2 or S protein (Figure 1B).
Figure 1 – (A) SP1, SP2, and S protein significantly induced migration of NK cells compared to controls. (B) Interferon-gamma, a measure of NK cell activation, was markedly increased in the presence of SP1. (Bortolotti et al., 2020)
The authors then tested the effects of spike protein on the cytotoxicity of NK cells. To do so, SP1, SP2, and S protein were expressed in K562 cells, and the harvested NK cells were incubated with the K562 cells, after which expression of the molecule CD107a (a marker of NK cell degranulation) was observed. At the end of the incubation period, they noted a non-significant increase in CD107a expression for each of the three treatments compared to control K562 cells (Figure 2, top). Due to the respiratory nature of the virus, however, the experiment was repeated in the lung epithelial cell line Beas-2B. This time, the authors observed a significant decrease in CD107a in the cells expressing SP1; and a non-significant decrease in CD107a was observed in the SP2- and S-expressing cells (Figure 2, bottom). They concluded that impairment of CD107a expression indicates a reduction in the cytotoxic capacity of the NK cells due to interaction with SP1.
Figure 2 –(Top) CD107a was slightly increased, though not significantly, when NK cells were incubated with K562 cells transfected with SP1, SP2, or S protein. (Bottom) A significant decrease in CD107a expression was observed when NK cells were incubated with lung epithelial Beas-2B cells transfected with SP1, SP2, or S protein. (Bortolotti et al., 2020)
Finally, to determine the exact mechanism of the reduced cytotoxicity, the authors expressed SP1, SP2, and S in Beas-2B cells and stained them with anti-classical Human Leukocyte Antigen I antibodies (anti-HLA-A, anti-HLA-B, and anti-HLA-C). They observed a significant decrease in HLA I cell surface expression in Beas-2B cells that were transfected with SP1, indicating that SP1 has a large role in inhibiting the presentation of viral antigens by the host cells (Figure 3, top; see updates 6/15/2020 for discussion of MHC [HLA] I presentation). HLA-E is a “non-classical” HLA I molecule that is expressed on the surfaces of many cell types and interacts specifically with the CD94/NKG2A inhibitory receptors on NK cells. When the researchers stained the transfected Beas-2B cells with anti-HLA-E antibody, they noted a significant increase in HLA-E surface expression in the presence of SP1, but not SP2 or S (Figure 3, middle). Lastly, when NK cells were incubated with SP1-transfected Beas-2B cells, there was a significant increase in CD94/NKG2A expression on the NK cell surfaces (Figure 3, bottom). Taken together, these data indicate the ability of the SARS-CoV-2 SP1 to significantly downregulate NK cell activity in vivo via a heightened interaction between HLA-E and the inhibitory receptor CD94/NKG2A.
One limitation of this study is the fact that the subunits of the SARS-CoV-2 spike protein were only tested individually. It is not clear why the SP1 subunit should exert the observed effects on NK cells when the SP2 subunit does not. Further characterization of the function(s) of each subunit may shed light on the reason behind this disparity. It is not likely that the SP1 subunit is viable to initiate and sustain infection on its own, and it is likely not the case that the subunits of the spike protein would be encountered individually anyway. Future work should focus on determining the effects of the entire SARS-CoV-2 spike protein on NK cell cytotoxicity and exhaustion.
Figure 3 – (Top) In the presence of SP1, classical HLA I molecules were significantly downregulated on Beas-2B cells. (Middle) In the presence of SP1, non-classical HLA-E was significantly upregulated on Beas-2B cells. (Bottom) In the presence of SP1, the inhibitory receptor CD94/NKG2A was significantly upregulated on NK cells. (Bortolotti et al., 2020)
Full paper:
http://sites.utexas.edu/melamed-lab/2020/06/30/6-30-20/
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