Functional SARS-CoV-2-Specific Immune Memory Persists after Mild COVID-19.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus is causing a global pandemic, and cases continue to rise. Most infected individuals experience mildly symptomatic coronavirus disease 2019 (COVID-19), but it is unknown whether this can induce persistent immune memory that could contribute to immunity. We performed a longitudinal assessment of individuals recovered from mild COVID-19 to determine whether they develop and sustain multifaceted SARS-CoV-2-specific immunological memory. Recovered individuals developed SARS-CoV-2-specific immunoglobulin (IgG) antibodies, neutralizing plasma, and memory B and memory T cells that persisted for at least 3 months. Our data further reveal that SARS-CoV-2-specific IgG memory B cells increased over time. Additionally, SARS-CoV-2-specific memory lymphocytes exhibited characteristics associated with potent antiviral function: memory T cells secreted cytokines and expanded upon antigen re-encounter, whereas memory B cells expressed receptors capable of neutralizing virus when expressed as monoclonal antibodies. Therefore, mild COVID-19 elicits memory lymphocytes that persist and display functional hallmarks of antiviral immunity.

Murine red blood cells from genetically distinct donors cross-regulate when stored together.

BACKGROUND: Donor variability of red blood cell (RBC) storage has been observed in both humans and animal models. We utilized a strain of mice with RBCs known to store well (B6) and a strain known to store poorly (FVB) to test the hypothesis that RBCs affected the storage of other RBCs. STUDY DESIGN AND METHODS: Five strains of mice were used: 1) transgenic B6 mice expressing green fluorescent protein (GFP) in their RBCs (GFP.B6), 2) wild-type B6 mice, 3) wild-type FVB mice, 4) F1 crosses between GFP.B6 and FVB mice (GFP.F1), and 5) the analogous wild-type (B6xFVB) F1 cross. GFP.B6 or GFP.F1 RBCs were mixed with wild-type (non-GFP) RBCs from B6 or FVB strains before storage. Twenty-four-hour RBC recoveries were determined for stored RBCs by enumerating circulating GFP+ RBCs by flow cytometry. RESULTS: Twenty-four-hour recoveries of GFP.F1 RBCs was increased by co-storage with B6 RBCs but decreased by co-storage with FVB RBCs. This effect was dose dependent when tested with GFP.B6 RBCs; the more FVB blood added, the worse the 24-hour recoveries became. RBC cross-regulation did not occur when B6 and FVB RBCs were separated by a semipermeable membrane with a 0.4-µm size cutoff. CONCLUSION: These findings demonstrate that RBCs affect the storage of other RBCs, in both positive and negative directions, indicating not only that RBC storage is intrinsic to the RBC but that RBC-RBC communication occurs. Additional studies will be required to determine the nature of this effect and if these findings translate into human RBC storage.

Transfusion-induced alloimmunization and platelet refractoriness in a mouse model: mechanisms and interventions.

BACKGROUND: Platelet (PLT) transfusions can be an essential therapy for patients with thrombocytopenia to maintain hemostasis. However, some patients become alloimmunized to antigens on PLTs (typically HLA), which can prevent efficacy of PLT transfusion due to antibody-mediated clearance. In extreme cases, patients with alloimmunization to multiple HLAs can become "refractory" to PLT transfusion, such that insufficient compatible PLT units can be found to meet transfusion needs. MATERIALS AND METHODS: An in vivo murine model of PLT-induced alloimmunization was refined so as to include both transfusion with allogeneic leukoreduced PLTs and studies of posttransfusion PLT recoveries, allowing assessment of alloimmunization and refractoriness. Basic mechanisms of antibody-mediated PLT clearance were investigated using recipients missing either the C3 complement gene or the common gamma chain for Fc receptors. In addition, the efficacy of using costimulatory blockade as a therapeutic intervention was assessed by testing CTLA4-Ig administration before PLT transfusion. RESULTS: Fcγ receptors (but not complement C3) are required for alloantibody-mediated PLT refractoriness. In addition, levels of anti-MHC predict the extent of refractoriness in a given animal. Finally, costimulatory blockade as a therapeutic modality prevents transfusion-induced PLT refractoriness. CONCLUSIONS: Together these findings introduce new experimental methods, basic mechanistic understanding, and a potential therapeutic intervention for alloimmunization to MHC-based antigens on transfused PLTs.

Testosterone-dependent sex differences in red blood cell hemolysis in storage, stress, and disease.

BACKGROUND: Red blood cell (RBC) hemolysis represents an intrinsic mechanism for human vascular disease. Intravascular hemolysis releases hemoglobin and other metabolites that inhibit nitric oxide signaling and drive oxidative and inflammatory stress. Although these pathways are important in disease pathogenesis, genetic and population modifiers of hemolysis, including sex, have not been established. STUDY DESIGN AND METHODS: We studied sex differences in storage or stress-induced hemolysis in RBC units from the United States and Canada in 22 inbred mouse strains and in patients with sickle cell disease (SCD) using measures of hemolysis in 315 patients who had homozygous SS hemoglobin from the Walk-PHASST cohort. A mouse model also was used to evaluate posttransfusion recovery of stored RBCs, and gonadectomy was used to determine the mechanisms related to sex hormones. RESULTS: An analysis of predisposition to hemolysis based on sex revealed that male RBCs consistently exhibit increased susceptibility to hemolysis compared with females in response to routine cold storage, under osmotic or oxidative stress, after transfusion in mice, and in patients with SCD. The sex difference is intrinsic to the RBC and is not mediated by plasmatic factors or female sex hormones. Importantly, orchiectomy in mice improves RBC storage stability and posttransfusion recovery, whereas testosterone repletion therapy exacerbates hemolytic response to osmotic or oxidative stress. CONCLUSION: Our findings suggest that testosterone increases susceptibility to hemolysis across human diseases, suggesting that male sex may modulate clinical outcomes in blood storage and SCD and establishing a role for donor genetic variables in the viability of stored RBCs and in human hemolytic diseases.

Lupus IgA1 autoantibodies synergize with IgG to enhance pDC responses to RNA-containing immune complexes.

Autoantibodies to nuclear antigens are hallmarks of the autoimmune disease systemic lupus erythematosus (SLE) where they contribute to pathogenesis. However, there remains a gap in our knowledge regarding how different isotypes of autoantibodies contribute to disease, including the production of the critical type I interferon (IFN) cytokines by plasmacytoid dendritic cells (pDCs) in response to immune complexes (ICs). We focused on IgA, which is the second most prevalent isotype in serum, and along with IgG is deposited in glomeruli in lupus nephritis. Here, we show that individuals with SLE have IgA autoantibodies against most nuclear antigens, correlating with IgG against the same antigen. We investigated whether IgA autoantibodies against a major SLE autoantigen, Smith ribonucleoproteins (Sm/RNPs), play a role in IC activation of pDCs. We found that pDCs express the IgA-specific Fc receptor, FcαR, and there was a striking ability of IgA1 autoantibodies to synergize with IgG in RNA-containing ICs to generate robust pDC IFNα responses. pDC responses to these ICs required both FcαR and FcγRIIa, showing a potent synergy between these Fc receptors. Sm/RNP IC binding to and internalization by pDCs were greater when ICs contained both IgA1 and IgG. pDCs from individuals with SLE had higher binding of IgA1-containing ICs and higher expression of FcαR than pDCs from healthy control individuals. Whereas pDC FcαR expression correlated with blood ISG signature in SLE, TLR7 agonists, but not IFNα, upregulated pDC FcαR expression in vitro. Together, we show a new mechanism by which IgA1 autoantibodies contribute to SLE pathogenesis.

Lupus IgA1 autoantibodies synergize with IgG to enhance plasmacytoid dendritic cell responses to RNA-containing immune complexes.

Autoantibodies to nuclear antigens are hallmarks of systemic lupus erythematosus (SLE) where they contribute to pathogenesis. However, there remains a gap in our knowledge regarding how different isotypes of autoantibodies contribute to this autoimmune disease, including the production of the critical type I interferon (IFN) cytokines by plasmacytoid dendritic cells (pDCs) in response to immune complexes (ICs). We focused on IgA, which is the second-most prevalent isotype in serum and, along with IgG, is deposited in glomeruli in individuals with lupus nephritis. We show that individuals with SLE have serum IgA autoantibodies against most nuclear antigens, correlating with IgG against the same antigen. We investigated whether IgA autoantibodies against a major SLE autoantigen, Smith ribonucleoprotein (Sm/RNP), played a role in IC activation of pDCs. We found that pDCs expressed the IgA-specific Fc receptor, FcαR, and IgA1 autoantibodies synergized with IgG in RNA-containing ICs to generate robust primary blood pDC IFN-α responses in vitro. pDC responses to these ICs required both FcαR and FcγRIIa, showing synergy between these Fc receptors. Sm/RNP IC binding to and internalization by pDCs were greater when ICs contained both IgA1 and IgG. Circulating pDCs from individuals with SLE had higher binding of IgA1-containing ICs and higher expression of FcαR than pDCs from healthy control individuals. Although pDC FcαR expression correlated with the blood IFN-stimulated gene signature in SLE, Toll-like receptor 7 agonists, but not IFN-α, up-regulated pDC FcαR expression in vitro. Together, we show a mechanism by which IgA1 autoantibodies contribute to SLE pathogenesis.

IgG Subclass Determines Suppression Versus Enhancement of Humoral Alloimmunity to Kell RBC Antigens in Mice.

It has long been appreciated that immunoglobulins are not just the effector endpoint of humoral immunity, but rather have a complex role in regulating antibody responses themselves. Donor derived anti-RhD IgG has been used for over 50 years as an immunoprophylactic to prevent maternal alloimmunization to RhD. Although anti-RhD has dramatically decreased rates of hemolytic disease of the fetus and newborn (for the RhD alloantigen), anti-RhD also fails in some cases, and can even paradoxically enhance immune responses in some circumstances. Attempts to generate a monoclonal anti-RhD have largely failed, with some monoclonals suppressing less than donor derived anti-RhD and others enhancing immunity. These difficulties likely result, in part, because the mechanism of anti-RhD remains unclear. However, substantial evidence exists to reject the common explanations of simple clearance of RhD + RBCs or masking of antigen. Donor derived anti-RhD is a mixture of 4 different IgG subtypes. To the best of our knowledge an analysis of the role different IgG subtypes play in immunoregulation has not been carried out; and, only IgG1 and IgG3 have been tested as monoclonals. Multiple attempts to elicit alloimmune responses to human RhD epitopes in mice have failed. To circumvent this limitation, we utilize a tractable animal model of RBC alloimmunization using the human Kell glycoprotein as an antigen to test the effect of IgG subtype on immunoregulation by antibodies to RBC alloantigens. We report that the ability of an anti-RBC IgG to enhance, suppress (at the level of IgM responses), or have no effect is a function of the IgG subclass in this model system.

Functional SARS-CoV-2-specific immune memory persists after mild COVID-19.

The recently emerged SARS-CoV-2 virus is currently causing a global pandemic and cases continue to rise. The majority of infected individuals experience mildly symptomatic coronavirus disease 2019 (COVID-19), but it is unknown whether this can induce persistent immune memory that might contribute to herd immunity. Thus, we performed a longitudinal assessment of individuals recovered from mildly symptomatic COVID-19 to determine if they develop and sustain immunological memory against the virus. We found that recovered individuals developed SARS-CoV-2-specific IgG antibody and neutralizing plasma, as well as virus-specific memory B and T cells that not only persisted, but in some cases increased numerically over three months following symptom onset. Furthermore, the SARS-CoV-2-specific memory lymphocytes exhibited characteristics associated with potent antiviral immunity: memory T cells secreted IFN-γ and expanded upon antigen re-encounter, while memory B cells expressed receptors capable of neutralizing virus when expressed as antibodies. These findings demonstrate that mild COVID-19 elicits memory lymphocytes that persist and display functional hallmarks associated with antiviral protective immunity.

Functional SARS-CoV-2-specific immune memory persists after mild COVID-19.

The recently emerged SARS-CoV-2 virus is currently causing a global pandemic and cases continue to rise. The majority of infected individuals experience mildly symptomatic coronavirus disease 2019 (COVID-19), but it is unknown whether this can induce persistent immune memory that might contribute to herd immunity. Thus, we performed a longitudinal assessment of individuals recovered from mildly symptomatic COVID-19 to determine if they develop and sustain immunological memory against the virus. We found that recovered individuals developed SARS-CoV-2-specific IgG antibody and neutralizing plasma, as well as virus-specific memory B and T cells that not only persisted, but in some cases increased numerically over three months following symptom onset. Furthermore, the SARS-CoV-2-specific memory lymphocytes exhibited characteristics associated with potent antiviral immunity: memory T cells secreted IFN-γ and expanded upon antigen re-encounter, while memory B cells expressed receptors capable of neutralizing virus when expressed as antibodies. These findings demonstrate that mild COVID-19 elicits memory lymphocytes that persist and display functional hallmarks associated with antiviral protective immunity.