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Cytokine Storms and Immune Response
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Injury to multiple organs has been observed in severe cases of COVID-19. Brain inflammation, seizures, and strokes have been reported along with loss of sense of smell. In addition to blood clot formation, heart attacks and cardiac inflammation can occur in response to SARS-CoV-2. The liver and kidney have also shown signs of severe damage and failure. The lower gastrointestinal tract is rich in ACE2 receptors and symptoms of COVID-19 can include diarrhea. Despite a growing list of organ systems affected by COVID-19, there remains much to be investigated. It is not well understood whether the injury observed in these organ systems is due to direct viral damage or are indirect effects of hyperactivity of the immune system (cytokine storms), clotting abnormalities, or side effects of drug administration.
The complement pathway includes over 30 proteins and is an important component of innate immunity and host defense systems. Products of the complement pathway are stimulated in the lung at the alveolar-capillary interface following endothelial damage caused by viral infections and cytokine signaling. The complement pathway activates tissue factor expression and the secretion of von Willebrand factor (VWF) and platelet factor V, which are important components of the coagulation pathway. Over-activation of the complement pathway is known to promote hyper-inflammation, endothelial cell damage, thrombosis, thrombophilia, and ultimately multiple organ system failure.
There are many proposed explanations for how the complement system contributes to the clotting abnormalities observed in COVID-19, such as damaging feedback loops between NET formation, complement, and coagulation pathways. It is likely that the combined effects of endothelial injury, complement activation, dysregulated neutrophils, and hyper-coagulation together exacerbate the severity of COVID-19. Despite many hypotheses regarding the crosstalk between the complement pathway, coagulation cascades, and inflammation, it is not well understood how they contribute to thrombosis in the realm of COVID-19 pathogenesis.
Cytokine storms cause endothelial cell damage and cell death in blood vessels. As a result, several events occur that contribute to the development of thrombosis and other COVID-19 complications:
- Coagulation and Complement Activation: The complement system is activated following endothelial cell damage, which triggers the coagulation cascade. The coagulation pathway is essential for repairing the damaged vascular tissue and minimizing blood loss. However, when the complement pathway is not regulated properly, a hypercoagulable state can occur that drives endothelial damage and increases the risk of thrombosis.
- Capillary Leak Syndrome: Capillary leak syndrome develops when plasma drains from capillaries into neighboring tissues leading to hypotension, edema, organ damage, and acute respiratory failure.
- Erythrophagocytosis: Eythrophagocytosis is the destruction of red blood cells, which could lead to anemia. This is caused by high levels of cytokine-mediated activation of macrophages.
- Emergency Granulopoiesis: Elevated systemic cytokines are associated with low platelet numbers and the rapid production of neutrophils, eosinophils, and basophils. These granulocytes and monocytes are quickly mobilized to migrate to sites of inflammation and infection to function in defensive roles. This can also lead to the increased production of pro- inflammatory cytokines from natural killer and T cells.
- Immuno-thrombosis: Neutrophils release neutrophil extracellular traps (NETs) to capture pathogens and form thrombi, called immuno-thrombosis. Immuno-thrombosis further amplifies cytokine production which could lead to widespread thrombi formation.
- Disseminated Intravascular Coagulation: Disruption to vascular system hemostasis can occur following thrombosis, which could drive disseminated intravascular coagulation (DIC). DIC is the formation of blood clots that block small blood vessels throughout the body which can cause shortness of breath, low blood pressure, multiple organ failure, and death.
A positive SARS-CoV-2 antibody result suggests that an individual has been exposed to SARS-CoV-2 and has generated an immune response to it. It does not indicate an active SARS-CoV-2 infection, but rather suggests that exposure has occurred. Tracking SARS-CoV-2 antibody levels will be crucial for population studies to discern the spread of the virus within a population, how antibody levels change over time, and how the data correlate to immunity. Serology studies will also be informative on the effectiveness of SARS-CoV-2 vaccines. The goal of vaccines is to facilitate the body to manufacture antibodies specific to the SARS-CoV-2 virus. Being able to measure antibody levels in response to vaccine delivery may be indicative of the efficacy of the vaccine. It is important to keep in mind that antibodies can be developed against different antigens of the SARS-CoV-2 virus, such as against the S protein or the N protein. Detection of different viral proteins will help to determine mechanisms of immunity and viral function. Anti-SARS-CoV-2 antibody levels correlate with COVID-19 disease severity, which suggests that immune response is elevated in those with severe symptoms.
Antibodies function by neutralizing or destroying the virus in the blood or mucosa before it invades a cell. Therefore, measuring the levels of antibodies in the blood can be an indicator of immune protection. Recent studies have suggested that the amount of antibodies against SARs-CoV-2 decline rapidly in the blood over time. It is not yet known whether antibody protection occurs when levels are low or undetectable, or if antibodies against SARS-CoV-2 will provide protection against re-infection. More long-term surveillance and immunology studies are needed.
However, T cell immunity is promising to provide long-term immune protection. T cells function by targeting and killing cells that have been infected by the virus. The formation of memory T cells following exposure to the virus will convey virus-specific immune response abilities over time. There is a need for more research that investigates the long-term immunological responses to SARS-CoV-2.
Immunoglobulins are glycoproteins produced by plasma cells and are important for antibodies to recognize and bind antigens. Each isotype has distinct effector functions. Antibodies specific to SARS- CoV-2 are primarily found in the immunoglobulin classes of IgM, IgA, and IgG.
- IgM are the primary response antibodies that are first detected following SARS-CoV-2 infection.
- IgA is secreted into tears, mucus, and saliva of the respiratory epithelium and gastrointestinal tract. SARS-CoV-2 specific IgA can also be seroconverted for detection in serum and have effector functions in a first line of defense against the SARS-CoV-2 respiratory pathogen. IgA can also be detected in early stages with IgM and may show higher diagnostic sensitivity than IgM.
- IgG antibodies are detected at later time periods post-infection, but remain in the serum for the longest period of time.
The correlation of antibody response, especially IgA, to COVID-19 disease progression is striking and needs further investigation.
B cells are the antibody-producing cells of the body that provide humoral immune responses. B cells encounter antigens through contact with dendritic and macrophage immune cells. B cells are formed in germinal centers (GC) where clonal expansion occurs via proliferation and B cells undergo class switch recombination. Immunoglobulin class switching is important to diversify and enhance the cell’s ability to eliminate the pathogen. Activation by T cell cytokines induces B cells to undergo clonal selection to enrich for B cells with the highest antigen-binding affinities. These selected B cells will ultimately differentiate into memory B cells and plasma cells that rapidly produce SARs-CoV-2-specific antibodies. There is potential for these memory B cells and plasma cells to be long-lived and provide lasting protection against the SARS-CoV-2 pathogen, but more investigation is required to determine protective immunity over time.
Despite having a bad reputation to form cytokine storms, cytokines are functionally important during B cell development, survival, differentiation, and proliferation. B cells produce neutralizing antibodies during immune responses to viral invasion. During infection, B cells are recruited to the pathogen by chemokine interferons. After arrival to the virus, B cell receptors bind antigens on the cell membrane with high affinity. Further B cell stimulation occurs by cytokines released from T helper cells. B cells also produce cytokines that are important for adaptive immune system functions. In the context of COVID- 19, cytokine storms may be dangerous, but cytokines also serve important functions during protective immunity development.
Yes. Cytokines normally function in a protective capacity in response to pathogenic invasion. However, when cytokine levels are elevated and rapidly circulating throughout the body, the protective functions are lost. There are regulatory mechanisms in place to resolve cytokine immune responses. For example, cytokines typically have short half-lives to contain the response to the local site of infection. At levels above threshold, these regulatory safeguards are overcome and cytokines become pathological, driving collateral tissue damage throughout the body.
Cytokines function as indicators of inflammation and can also be contributors to the COVID-19 disease progression. A cytokine storm initially presents as a persistent fever and constitutional symptoms (headache, fatigue, etc). As it progresses, systemic inflammation can occur that manifests as hypotension, high fever, vasodilation, and ultimately organ failure. Important areas of research that are attracting much-needed attention are surrounding the mechanisms and risk-factors driving cytokine storms, and how to best therapeutically treat them before they become deadly.
Elevated levels of circulating cytokines are called a cytokine storm. Cytokines function to mobilize neutrophils and monocytes to sites of inflammation in response to pathogens. Cytokine signaling is a rapid response that normally dissipates after the pathogen and infected cells have been eliminated. In the context of SARS-CoV-2, the release of cytokines IL-12, IFN-a, MHC Class 1 expression, and NK cell activation are collectively needed to resist viral replication and eradicate SARS-CoV-2 infected cells. However, these signaling pathways may also lead to the rapid production of proinflammatory cytokines and chemokines in a faulty positive-feedback loop during a cytokine storm. The list of pro-inflammatory cytokines that are activated following SARS-CoV-2 infection is extensive and includes IFN-a, IFN-g, IL-1, IL-6, IL-17, CXCL10, TNF-a, and TGFb, along with others. The scientific community has identified key participants of cytokine storms, but more research about the mechanisms of how they develop and how to stop them is critically needed.
The immune response to SARS-CoV-2 is mediated through cytokines. There is an increase in the release of proinflammatory cytokines IP-10, MCP-1, MIP-1A, and TNF-a following SARS-CoV-2 infection that correlates with COVID-19 severity and mortality. Antigen presenting cells (APCs), such as dendritic cells and macrophages, are triggered after SARS-CoV-2 gains entry into host cells. APCs recognize structural components of the virus through pattern recognition receptors. This recognition induces a signaling cascade to activate immune effector cells, such as B cells, and prompt the production of proinflammatory proteins IFN-a, NF-kB, MAPK, and TNF-b in an initial line of defense. This is important because the expression of IFN-stimulated genes functions to suppress viral replication. However, this initial immune response is delicate because excessive expression of proinflammatory genes can lead to hyperinflammation which drives acute respiratory distress syndrome, which can be lethal in COVID-19. Despite the many research studies that have teased out the mechanisms of immune responses to infection, not everybody reacts to pathogens the same way. This is especially seen in COVID-19 where there is a variety of immune responses, ranging from undetectable to dangerous cytokine storms that can be fatal.
After gaining entry into target cells via the ACE2 receptor, SARS-CoV-2 releases viral genetic material into the cytoplasm of the host cell. The viral genetic material is in the form of mRNA that is ready for translation into protein. The nucleocapsid, or N protein, is bound to the viral mRNA and functions to localize the genetic material to the endoplasmic reticulum-Golgi host machinery. Nonstructural proteins are translated and form replicase-transcriptase complexes that function to transcribe endogenous viral genetic material. The translation and assembly of accessory and structural proteins, such as S protein, occur through the host endoplasmic reticulum followed by the endoplasmic reticulum-Golgi intermediate compartment (ERGIC). The N proteins join the newly replicated mRNA and together move into the ERGIC to join with other structural proteins, ultimately forming new SARS-CoV-2 viruses. Small vesicles export the newly formed viruses out of the host cell via exocytosis. As the virus leaves the host cell, it is suggested that proteases on the cell surface prime the S protein to enable efficient attack on other cells. The intricate mechanisms of the SARS-CoV-2 viral life cycle have more to be understood.
SARS-CoV-2 primarily enters the body through respiratory cells in the airway and lungs. A common method used to predict the cell types susceptible to SARS-CoV-2 infection is to analyze the expression of entry-associated genes, such as ACE2 and TMPRSS2. ACE2 is the receptor SARS-CoV-2 binds to gain entry into target cells and TMPRSS2 is necessary for S protein priming. Cells that co-express ACE2 and TMPRSS2 include lung alveolar type II epithelial cells, ileal enterocytes, nasal goblet cells, and cells of the esophagus and colon. Interestingly, ACE2 is expressed at low levels throughout the airway epithelium and exhibits the highest levels of expression in nasal secretory cells. In airway cells, TMPRSS2 is expressed in a small subset of ACE2+ cells, suggesting that SARS-CoV-2 may be utilizing additional host proteases. For example, SARS-CoV-2 could be using the lysosomal protease cathepsin B, which is co-expressed in 70-90% of ACE2+ cells. Another hypothesis that requires more investigation is that ACE2+ TMPRSS2+ nasal goblet cells could be the initial location of infection and function in a dual capacity as a viral reservoir. Much more research is needed to identify the cell types that are being directly infected by SARS-CoV-2.
Understanding how SARS-CoV-2 infects cells is a critical step for understanding its pathogenicity. SARS- CoV-2 gains entry into cells through the ACE2 cell surface receptor with the assistance of host protease activators. The spike protein (S Protein) from SARS-CoV-2 binds to ACE2 on host cells with high affinity through the receptor-binding domain (RBD). Host proteases, such as TMPRSS2, furin, and lysosomal proteases, are required to proteolytically activate the viral S protein. It has been suggested that protease activation confers a structural change to the S protein that enhances RBD binding to ACE2 and promotes membrane fusion. The functional mechanisms of how host proteases promote SARS-CoV-2 binding and entry into cells requires further investigation.
According to the World Health Organization, SARS-CoV-2 transmission occurs through close contact with respiratory droplets and saliva secretions from infected carriers. SARS-CoV-2 transmission can even be airborne during aerosol-generating medical procedures. However, it is unclear if SARS-CoV-2 is transmitted by aerosol routes during non-aerosol generating procedures. Contaminated surfaces can harbor viable SARS-CoV-2 viruses, although there is no consistent evidence suggesting that SARS-CoV-2 can be transmitted after contact with contaminated surfaces. SARS-CoV-2 RNA has been detected in feces, urine, plasma, and serum, but it is not known if transmission of the virus can occur through these routes. The transmission and subsequent spread of SARS-CoV-2 infection have more to be understood.