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The specific kinetics and thermodynamics of protein-protein interactions underlie the molecular mechanisms of cellular functions; hence the characterization of these interaction parameters is central to the quantitative understanding of physiological and pathological processes. Many methods have been developed to study protein-protein interactions, which differ in various features including the interaction detection principle, the sensitivity, whether the method operates in vivo, in vitro, or in silico, the temperature control, the use of labels, immobilization, the amount of sample required, the number of measurements that can be accomplished simultaneously, or the cost. Bio-Layer Interferometry (BLI) is a label-free biophysical method to measure the kinetics of protein-protein interactions. Label-free interaction assays are a broad family of methods that do not require protein modifications (other than immobilization) or labels such as fusions with fluorescent proteins or transactivating domains or chemical modifications like biotinylation or reaction with radionuclides. Besides BLI, other label-free techniques that are widely used for determining protein-protein interactions include surface plasmon resonance (SPR), thermophoresis, and isothermal titration calorimetry (ITC), among others.

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OBJECTIVE: The complement cascade as major fluid phase innate immune system is activated during progression of osteoarthritis (OA). Generated anaphylatoxins and the corresponding receptors C3aR and C5aR1 are associated with the calcification of blood vessels and involved in osteogenic differentiation. This study aims on elucidating whether complement activation products contribute to cartilage calcification of OA cartilage. METHOD: Human articular chondrocytes were osteogenically differentiated in vitro in the presence or absence of C3a, C5a, and bone morphogenetic protein (BMP) 2. Furthermore, macroscopically intact (OARSI grade ≤ 1) and highly degenerated human cartilage (OARSI grade ≥ 3) was used for C3aR and C5aR1 histochemistry. Calcification of the cartilage was assessed by Alizarin Red S and von Kossa staining. RESULTS: C3a and C5a amplified matrix mineralization during in vitro osteogenesis, while inhibition of the corresponding receptors impaired calcium deposition. Moreover, C3aR and C5aR1 expression was upregulated during osteogenic differentiation and also in degenerated cartilage. Additionally, anaphylatoxin receptor expression was positively associated with calcification of native cartilage tissue and calcium deposition during osteogenic differentiation. Finally, the pro-hypertrophic growth factor BMP2 induced the expression of C5aR1. CONCLUSIONS: Our findings indicate that anaphylatoxins and their receptors play a decisive role in cartilage calcification processes during OA progression.

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The complement system mediates diverse regulatory immunological functions. C5aR2, an enigmatic receptor for anaphylatoxin C5a, has been shown to modulate PRR-dependent pro-inflammatory cytokine secretion in human macrophages. However, the specific downstream targets and underlying molecular mechanisms are less clear. In this study, CRISPR-Cas9 was used to generate macrophage models lacking C5aR2, which were used to probe the role of C5aR2 in the context of PRR stimulation. cGAS and STING-induced IFN-ß secretion was significantly increased in C5aR2 KO THP-1 cells and C5aR2-edited primary human monocyte-derived macrophages, and STING and IRF3 expression were increased, albeit not significantly, in C5aR2 KO cell lines implicating C5aR2 as a regulator of the IFN-ß response to cGAS-STING pathway activation. Transcriptomic analysis by RNAseq revealed that nucleic acid sensing and antiviral signalling pathways were significantly up-regulated in C5aR2 KO THP-1 cells. Altogether, these data suggest a link between C5aR2 and nucleic acid sensing in human macrophages. With further characterisation, this relationship may yield therapeutic options in interferon-related pathologies.

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IMPORTANCE: Pathogenic Rickettsia species are extremely dangerous bacteria that grow within the cytoplasm of host mammalian cells. In most cases, these bacteria are able to overpower the host cell and grow within the protected environment of the cytoplasm. However, a dramatic conflict occurs when Rickettsia encounter innate immune cells; the bacteria can "win" by taking over the host, or the bacteria can "lose" if the host cell efficiently fights the infection. This manuscript examines how the immune complement system is able to detect the presence of Rickettsia and alert nearby cells. Byproducts of complement activation called anaphylatoxins are signals that "activate" innate immune cells to mount an aggressive defensive strategy. This study enhances our collective understanding of the innate immune reaction to intracellular bacteria and will contribute to future efforts at controlling these dangerous infections.

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BACKGROUND AND OBJECTIVE: Cytokine storm (CS) is a major contributor to the fatal outcome of severe infectious diseases, including Covid-19. Treatment with the complement (C) C5 inhibitor eculizumab was beneficial in end-stage Covid-19, however, the mechanism of this effect is unknown. To clarify this, we analyzed the relationship between C activation and production of pro-inflammatory cytokines in a PBMC model. METHODS: Human PBMC with or without 20 % autologous serum was incubated with C3a, C5a, zymosan or zymosan-pre-activated serum (ZAS) for 24 h with or without eculizumab or the C5a receptor antagonist, DF2593A. C activation (sC5b-9) and 9 inflammatory cytokines were measured by ELISA. RESULTS: In serum-free unstimulated PBMC only IL-8 release could be measured during incubation. Addition of C5a increased IL-8 secretion only, ZAS induced both IL-2 and IL-8, while zymosan led to significant production of all cytokines, most abundantly IL-8. In the presence of serum the above effects were greatly enhanced, and the zymosan-induced rises of IL-1α, IL-1ß IFN-γ and IL-2 were significantly attenuated by eculizumab but not by DF2593a. CONCLUSIONS: These data highlight the complexity of interrelationships between C activation and cytokine secretion under different experimental conditions. The clinically relevant findings include the abundant formation of the chemokine IL-8, which was stimulated by C5a, and the suppression of numerous inflammatory cytokines by eculizumab, which explains its therapeutic efficacy in severe Covid-19. These data strengthen the clinical relevance of the applied PBMC model for drug screening against CS, enabling the separation of complex innate immune cross-talks.

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Leptospirosis is a neglected worldwide zoonosis involving farm animals and domestic pets caused by the Gram-negative spirochete Leptospira interrogans. This bacterium deploys a variety of immune evasive mechanisms, some of them targeted at the complement system of the host's innate immunity. In this work, we have solved the X-ray crystallographic structure of L. interrogans glyceraldehyde-3-phosphate dehydrogenase (GAPDH) to 2.37-Å resolution, a glycolytic enzyme that has been shown to exhibit moonlighting functions that potentiate infectivity and immune evasion in various pathogenic organisms. Besides, we have characterized the enzyme's kinetic parameters toward the cognate substrates and have proven that the two natural products anacardic acid and curcumin are able to inhibit L. interrogans GAPDH at micromolar concentration through a noncompetitive inhibition modality. Furthermore, we have established that L. interrogans GAPDH can interact with the anaphylatoxin C5a of human innate immunity in vitro using bio-layer interferometry and a short-range cross-linking reagent that tethers free thiol groups in protein complexes. To shed light into the interaction between L. interrogans GAPDH and C5a, we have also carried out cross-link guided protein-protein docking. These results suggest that L. interrogans could be placed in the growing list of bacterial pathogens that exploit glycolytic enzymes as extracellular immune evasive factors. Analysis of the docking results indicates a low affinity interaction that is consistent with previous evidence, including known binding modes of other α-helical proteins with GAPDH. These findings allow us to propose L. interrogans GAPDH as a potential immune evasive factor targeting the complement system.

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Allergic conditions are associated with canonical and noncanonical activation of the complement system leading to the release of several bioactive mediators with inflammatory and immunoregulatory properties that regulate the immune response in response to allergens during the sensitization and/or the effector phase of allergic diseases. Further, immune sensors of complement and regulator proteins of the cascade impact on the development of allergies. These bioactive mediators comprise the small and large cleavage fragments of C3 and C5. Here, we provide an update on the multiple roles of immune sensors, regulators, and bioactive mediators of complement in allergic airway diseases, food allergies, and anaphylaxis. A particular emphasis is on the anaphylatoxins C3a and C5a and their receptors, which are expressed on many of the effector cells in allergy such as mast cells, eosinophils, basophils, macrophages, and neutrophils. Also, we will discuss the multiple pathways, by which the anaphylatoxins initiate and control the development of maladaptive type 2 immunity including their impact on innate lymphoid cell recruitment and activation. Finally, we briefly comment on the potential to therapeutically target the complement system in different allergic conditions.

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The complement system plays a crucial role in cancer development. Our study investigated the role of C3a anaphylatoxin on the tumor microenvironment. Our models consisted of mesenchymal stem cells (MSC-like, 3T3-L1), macrophages (Raw 264.7 Blue, (RB)) and tumor cells (melanoma B16/F0). Recombinant mouse (Mo) C3a (rC3a) was produced in CHO cells transfected with a Mo-IL10-signal peptide-Mo C3a plasmid construct. The effects of rC3a, IFN-γ, TGF-ß1, and LPS were tested on the expression of C3, C3aR, PI3K, cytokines, chemokines, transcription factors, antioxidant defense mechanisms, angiogenesis and macrophage polarization (M1/M2). 3T3-L1 expressed the highest levels of C3, while C3aR was expressed more by RB. Interestingly, expression of C3/3T3-L1 and C3aR/RB was markedly upregulated by IFN-γ. rC3a was found to upregulate the expression of anti-inflammatory cytokines (IL-10) on 3T3-L1 and TGF-ß1 on RB. rC3a also upregulated the expression of pro-inflammatory cytokines in RB. The expression of CCL-5 increased in 3T3-L1 in response to rC3a. On RB, rC3a did not alter M1/M2 polarization but upregulated the expression of antioxidant defense genes, HO-1, and VEGF. C3/C3a produced mainly by MSC may play a critical role in TME remodeling by stimulating both anti-inflammatory and proangiogenic activities of tumor stromal cells.

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Mitochondrial dynamics is a morphological balance between fragmented and elongated shapes, reflecting mitochondrial metabolic status, cellular damage, and mitochondrial dysfunction. The anaphylatoxin C5a derived from complement component 5 cleavage, enhances cellular responses involved in pathological stimulation, innate immune responses, and host defense. However, the specific response of C5a and its receptor, C5a receptor (C5aR), in mitochondria is unclear. Here, we tested whether the C5a/C5aR signaling axis affects mitochondrial morphology in human-derived retinal pigment epithelial cell monolayers (ARPE-19). C5aR activation with the C5a polypeptide induced mitochondrial elongation. In contrast, oxidatively stressed cells (H2O2) responded to C5a with an enhancement of mitochondrial fragmentation and an increase in the number of pyknotic nuclei. C5a/C5aR signaling increased the expression of mitochondrial fusion-related protein, mitofusin-1 (MFN1) and - 2 (MFN2), as well as enhanced optic atrophy-1 (Opa1) cleavage, which are required for mitochondrial fusion events, whereas the mitochondrial fission protein, dynamin-related protein-1 (Drp1), and mitogen-activated protein kinase (MAPK)-dependent extracellular signal-regulated protein kinase (Erk1/2) phosphorylation were not affected. Moreover, C5aR activation increased the frequency of endoplasmic reticulum (ER)-mitochondria contacts. Finally, oxidative stress induced in a single cell within an RPE monolayer (488 nm blue laser spot stimulation) induced a bystander effect of mitochondrial fragmentation in adjacent surrounding cells only in C5a-treated monolayers. These results suggest that C5a/C5aR signaling produced an intermediate state, characterized by increased mitochondrial fusion and ER-mitochondrial contacts, that sensitizes cells to oxidative stress, leading to mitochondrial fragmentation and cell death.

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Anaphylaxis is a serious allergic or hypersensitivity reaction with a sudden onset that can be life-threatening or fatal. Previous studies have highlighted two pathways of anaphylaxis in mice. One is the classical immunoglobulin E (IgE)-mediated pathway that involves mast cells and histamine. The other is an alternative IgG-mediated pathway that involves basophils, monocytes/macrophages, neutrophils, and the platelet-activating factor (PAF). However, little is known about the mechanism by which complement anaphylatoxins contribute to the induction of anaphylaxis. Infection is a cofactor that potentially amplifies the risk of anaphylaxis. Here, we showed that priming with a lipopolysaccharide (LPS), which mimics bacterial infection, exacerbates anaphylatoxin C5a-induced anaphylaxis in mice. LPS plus C5a-induced anaphylaxis was mediated by histamine and lipid mediators, especially PAF. Cell depletion experiments demonstrated that LPS plus C5a-induced anaphylaxis depended on monocytes/macrophages, basophils, and neutrophils. These results suggest that C5a is a potent inducer of anaphylaxis in bacterial infections. Remarkably, the molecular and cellular mediators of LPS plus C5a-induced anaphylaxis are mostly shared with IgE- and IgG-mediated anaphylaxis. Therefore, combined inhibition of histamine and PAF may be beneficial as a second-line treatment for severe anaphylaxis.

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C3 is the central effector molecule of the complement system, mediating its multiple functions through different binding sites and their corresponding receptors. We will introduce the C3 forms (native C3, C3 [H2 O], and intracellular C3), the C3 fragments C3a, C3b, iC3b, and C3dg/C3d, and the C3 expression sites. To highlight the important role that C3 plays in human biological processes, we will give an overview of the diseases linked to C3 deficiency and to uncontrolled C3 activation. Next, we will present a structural description of C3 activation and of the C3 fragments generated by complement regulation. We will proceed by describing the C3a interaction with the anaphylatoxin receptor, followed by the interactions of opsonins (C3b, iC3b, and C3dg/C3d) with complement receptors, divided into two groups: receptors bearing complement regulatory functions and the effector receptors without complement regulatory activity. We outline the molecular architecture of the receptors, their binding sites on the C3 activation fragments, the cells expressing them, the diversity of their functions, and recent advances. With this review, we aim to give an up-to-date analysis of the processes triggered by C3 activation fragments on different cell types in health and disease contexts.

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Hidradenitis suppurativa (HS) is a chronic auto-inflammatory skin disease with a complex and multifactorial pathogenesis involving both the innate and adaptive immune system. Despite limited evidence for local complement activation, conflicting results have been published on the role of systemic complement activation in HS. It was hypothesized that complement was consumed in highly inflamed HS skin, trapping complement from the circulation. Therefore, the aim of this study was to evaluate this local complement deposition in HS skin lesions using routine and commonly used complement antibodies.Direct immunofluorescence for C1q, C3c, C4d, C5b-9, and properdin was performed on frozen tissue sections of 19 HS patients and 6 controls. C5a receptor 1 (C5aR1) was visualized using immunohistochemistry. Overall, we found no significant local complement deposition in HS patients versus controls regarding C1q, C3c, C4d, C5b-9, or properdin on either vessels or immune cells. C5aR1 expression was exclusively found on immune cells, predominantly neutrophilic granulocytes, but not significantly different relatively to the total infiltrate in HS lesions compared with controls. In conclusion, despite not being able to confirm local complement depositions of C1q, C3c, C4d, or properdin using highly sensitive and widely accepted techniques, the increased presence of C5aR1 positive immune cells in HS suggests the importance of complement in the pathogenesis of HS and supports emerging therapies targeting this pathway.

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Numerous publications have underlined the link between complement C5a and the clinical course of COVID-19. We previously reported that levels of C5a remain high in the group of severely ill patients up to 90 days after hospital discharge. We have now evaluated which complement pathway fuels the elevated levels of C5a during hospitalization and follow-up. The alternative pathway (AP) activation marker C3bBbP and the soluble fraction of C4d, a footprint of the classical/lectin (CP/LP) pathway, were assessed by immunoenzymatic assay in a total of 188 serial samples from 49 patients infected with SARS-CoV-2. Unlike C5a, neither C3bBbP nor C4d readouts rose proportionally to the severity of the disease. Detailed correlation analyses in hospitalization and follow-up samples collected from patients of different disease severity showed significant positive correlations of AP and CP/LP markers with C5a in certain groups, except for the follow-up samples of the patients who suffered from highly severe COVID-19 and presented the highest C5a readouts. In conclusion, there is not a clear link between persistently high levels of C5a after hospital discharge and markers of upstream complement activation, suggesting the existence of a non-canonical source of C5a in patients with a severe course of COVID-19.

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Osteoarthritis (OA) affects more than 500 million people worldwide and is among the five diseases in Germany causing the highest suffering of the patients and cost for the society. The quality of life of OA patients is severely compromised, and adequate therapy is lacking owing to a knowledge gap that acts as a major barrier to finding safe and effective solutions. Chronic, low-grade inflammation plays a central role in OA pathogenesis and is associated with both OA pain and disease progression. Innate immune pathways, such as the complement- and pattern-recognition receptor pathways, are pivotal to the inflammation in OA and key components of the innate immune system implicated in OA include DAMP-TLR signaling, the complement system, carboxypeptidase B (CPB), and mononuclear cells. Anaphylatoxins C3a and C5a are small polypeptides (77 and 74 amino acids, respectively) which are released by proteolytic cleavage of the complement components C3 and C5. The alternative complement pathway seems to play a crucial role in OA pathogenesis as these complement components, mostly C3 and its activation peptide C3a, were detected at high levels in osteoarthritic cartilage, synovial membrane, and cultured chondrocytes. Targeting the complement system by using anti-complement drugs as a therapeutic option bears the risk of major side effects such as increasing the risk of infection, interfering with cell regeneration and metabolism, and suppressing the clearance of immune complexes. Despite those adverse effects, several synthetic complement peptide antagonists show promising effects in ameliorating inflammatory cell responses also in joint tissues.

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INTRODUCTION: Both increased activity of the complement system (CS) and the role of the pituitary hormone prolactin (PRL) are implicated in osteoarthritis (OA) pathogenesis. Besides, Cathepsin D (CatD) activity is increased in the context of OA and can exert not only proteolytic but also non-proteolytic effects on cells. For the first time, possible crosstalk between two separate humoral systems: the CS and the PRL hormone systems in chondrocytes are examined together. METHODS: Primary human articular chondrocytes (hAC) were stimulated with complement protein C5 (10 µg /mL), PRL (25 ng/mL), CatD (100 ng/mL), or anaphylatoxin C5a (25 ng/mL) for 24 h or 72 h, while unstimulated cells served as controls. In addition, co-stimulations of C5 or PRL with CatD were carried out under the same conditions. The influence of the stimulants on cell viability, cell proliferation, and metabolic activity of hAC, the chondrosarcoma cell line OUMS-27, and endothelial cells of the human umbilical cord vein (HUVEC) was investigated. Gene expression analysis of C5a receptor (C5aR1), C5, complement regulatory protein CD59, PRL, PRL receptor (PRLR), CatD, and matrix metal-loproteinases (MMP)-13 were performed using real-time PCR. Also, collagen type (Col) I, Col II, C5aR1, CD59, and PRL were detected on protein level using immunofluorescence labeling. RESULTS: The stimulation of the hAC showed no significant impairment of the cell viability. C5, C5a, and PRL induced cell growth in OUMS-27 and HUVEC, but not in chondrocytes. CatD, as well as C5, significantly reduced the gene expression of CatD, C5aR1, C5, and CD59. PRLR gene expression was likewise impaired by C5, C5a, and PRL+CatD stimulation. On the protein level, CatD, as well as C5a, decreased Col II as well as C5aR1 synthesis. CONCLUSIONS: The significant suppression of the C5 gene expression under the influence of PRL+CatD and that of CD59 via PRL+/-CatD and conversely a suppression of the PRLR gene expression via C5 alone or C5a stimulation indicates an interrelation between the two mentioned systems. In addition, CatD and C5, in contrast to PRL, directly mediate possible negative feedback of their own gene expression.

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Emerging evidence suggests that signaling through the C3a anaphylatoxin receptor (C3aR) protects against various inflammation-related diseases. However, the role of C3aR in psoriasis remains unknown. The purpose of this study was to investigate the possible protective role of C3aR in psoriasis and to explore the underlying molecular mechanisms. We initially found that the psoriatic epidermis exhibited significantly decreased C3aR expression. C3aR showed protective roles in mouse models of imiquimod (IMQ)- and interleukin-23-induced psoriasis. Furthermore, increased epidermal thickness and keratin 6 (K6), K16, and K17 expression occurred in the ears and backs of C3aR-/- mice. Pharmacological treatment with a C3aR agonist ameliorated IMQ-induced psoriasiform lesions in mice and decreased the expression of K6, K16, and K17. Additionally, the signal transducer and activator of transcription 3 (STAT3) pathway participated in the protective function of C3aR. More importantly, the expression levels of K6, K16, and K17 in keratinocytes were all restored in HaCaT cells transfected with a C3aR-overexpression plasmid after treating them with colivelin (a STAT3 activator). Our findings demonstrate that C3aR protects against the development of psoriasis and suggest that C3aR confers protection by negatively regulating K6, K16, and K17 expression in a STAT3-dependent manner, thus inhibiting keratinocyte proliferation and helping reverse the pathogenesis of psoriasis.

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Anaphylatoxin C3a is a small signaling polypeptide that is generated during complement activation. C3a is involved in the regulation of various innate and adaptive immune system processes; however, the role of C3a in macrophage differentiation and polarization is poorly elucidated. Here we showed that C3a impairs alternative M2 polarization of human macrophages and suppressed CD206, IL1Ra and CCL22 expression. C3a leads to a decrease of nuclear receptor PPARγ expression via the ERK1/2 signaling pathway, resulting in repressed PPARγ-dependent activation of CD36, FABP4 and LXRα genes and blunted response to an LXR ligand TO901317. Using small interfering RNA and agonist/antagonist approaches we showed that C3a decreases CD206, IL1Ra and CCL22 transcription at least partly in a PPARγ-dependent manner in M2 macrophages. Moreover, C3a impairs efferocytosis by M2 macrophages and inhibits their migratory activity. By contrast, macrophages treated with C3a during differentiation show blunted response to lipopolysaccharide stimulation owing to downregulation of TLR4 and lipid raft content. At the same time, differentiation of macrophages with C3a does not change M1 polarization in interferon gamma (IFNγ) and IFNγ + lipopolysaccharide-treated macrophages. These data provide a novel role of complement system and C3a in the regulation of M2 macrophage polarizations and suggest crosstalk between C3a, TLR4, PPARγ and LXR signaling pathways.

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Coronavirus disease-2019 (COVID-19) was declared as a pandemic by WHO in March 2020. SARS-CoV-2 causes a wide range of illness from asymptomatic to life-threatening. There is an essential need to identify biomarkers to predict disease severity and mortality during the earlier stages of the disease, aiding treatment and allocation of resources to improve survival. The aim of this study was to identify at the time of SARS-COV-2 infection patients at high risk of developing severe disease associated with low survival using blood parameters, including inflammation and coagulation mediators, vital signs, and pre-existing comorbidities. This cohort included 89 multi-ethnic COVID-19 patients recruited between July 14th and October 20th 2020 in Doha, Qatar. According to clinical severity, patients were grouped into severe (n=33), mild (n=33) and asymptomatic (n=23). Common routine tests such as complete blood count (CBC), glucose, electrolytes, liver and kidney function parameters and markers of inflammation, thrombosis and endothelial dysfunction including complement component split product C5a, Interleukin-6, ferritin and C-reactive protein were measured at the time COVID-19 infection was confirmed. Correlation tests suggest that C5a is a predictive marker of disease severity and mortality, in addition to 40 biological and physiological parameters that were found statistically significant between survivors and non-survivors. Survival analysis showed that high C5a levels, hypoalbuminemia, lymphopenia, elevated procalcitonin, neutrophilic leukocytosis, acute anemia along with increased acute kidney and hepatocellular injury markers were associated with a higher risk of death in COVID-19 patients. Altogether, we created a prognostic classification model, the CAL model (C5a, Albumin, and Lymphocyte count) to predict severity with significant accuracy. Stratification of patients using the CAL model could help in the identification of patients likely to develop severe symptoms in advance so that treatments can be targeted accordingly.

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T cells play a fundamental role in the early control and clearance of many viral infections of the respiratory system. In SARS-CoV-2-infected individuals, lymphopenia with drastically reduced CD4+ and CD8+ T cells correlates with Coronavirus disease 2019 (COVID-19)-associated disease severity and mortality. In this study, we characterized cellular and humoral immune responses induced in patients with mild, severe and critical COVID-19. Peripheral blood mononuclear cells of 37 patients with mild, severe and critical COVID-19 and 10 healthy individuals were analyzed by IFNγ ELISpot and multi-color flow cytometry upon stimulation with peptide pools covering complete immunodominant SARS-CoV-2 matrix, nucleocapsid and spike proteins. In addition SARS-CoV-2 antibody levels, neutralization abilities and anaphylatoxin levels were evaluated by various commercially available ELISA platforms. Our data clearly demonstrates a significantly stronger induction of SARS-CoV-2 specific CD8+ T lymphocytes and higher IFNγ production in patients with mild compared to patients with severe or critical COVID-19. In all patients SARS-CoV-2-specific antibodies with similar neutralizing activity were detected, but highest titers of total IgGs were observed in critical patients. Finally, elevated anaphylatoxin C3a and C5a levels were identified in severe and critical COVID-19 patients probably caused by aberrant immune complex formation due to elevated antibody titers in these patients. Crucially, we provide a full picture of cellular and humoral immune responses of COVID-19 patients and prove that robust polyfunctional CD8+ T cell responses concomitant with low anaphylatoxin levels correlate with mild infections. In addition, our data indicates that high SARS-CoV-2 antibody titers are associated with severe disease progression.

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