• Adaptive Immunity Against Viral Threats

    Immunity Arising from Viral Infection

    Posted on Jul 18, 2020

    Innate Immunity and Adaptive Immunity

    Innate immunity is the initial immune response to infectious agents, carried out mainly by​ granulocytes and macrophages. These include a variety of responses that are genetically hard-wired to mount a quick response to families of molecules common to classes of pathogens.

    Adaptive (or acquired) immunity creates immunological memory after an initial response to a specific pathogen, leading to an enhanced response to subsequent encounters with that same pathogen. This process of acquired immunity is the basis of vaccination. 

    Summary of Adaptive Immunity

    Two groups of human leukocyte antigen (HLA) genes differentially drive immune system response to viral infection. HLA II genes are key to activating an adaptive immune response that includes antibodies against the invader produced by B cells but also involves highly specific responses by cytotoxic (killer) T cells. In addition, the initiation of B cells to produce antibodies in the first place requires stimulation by activated T helper cells. Both types of T cells, cytotoxic (killer) and helper Y cells, are activated by the binding of their T cell receptors on the surface of the cell to HLA II/viral antigen complexes. This binding includes CD8 receptors in the case of killer T cells and CD4 receptors in T helper cells. Evidence of past activations that chronicle the progress of a previous viral infection can be inferred by use of flow cytometry to follow the expression of marker genes. Flow cytometry is state of the art for this purpose, there is no practical alternative method.

    The Role of MHC I (HLA I) vs MHC II (HLA II) in Preparing for an Immune Response

    The major histocompatibility complex (MHC) is a group of proteins that has evolved to help vertebrate animals combat pathogens. Its functions are most advanced in mammals. In humans, this system of proteins is referred to as the human leukocyte antigen (HLA) complex,

    The HLA proteins are divided into two groups corresponding to MHC class I and class II. Proteins of the HLA I complex are expressed constitutively in almost all cells. In response to a viral infection,​ enzymes in infected cells chew up some of the proteins of now-internal virus particles into peptides. HLA I proteins form complexes with these peptides that then migrate to the cell surface as outward-facing receptors so that the viral peptides can be displayed to elements of the immune system. These peptides can be recognized as foreign antigens by cytotoxic (killer) T cells that then destroy the infected cell and the virus particles inside. They can also directly kill bacteria, fungi and multicellular parasites.

    The second group of HLA proteins, HLA II​, are expressed only by “professional” antigen-presenting​ cells (APCs), i.e., the dendritic cells and macrophages of the innate immune system and B cells of the adaptive immune system. The function of APCs is to encounter non-self-pathogens, process their proteins into peptides, complex these with HLA II proteins, and present them as foreign antigens on the cell surface for recognition by T cells in the first steps of mounting an adaptive immune system response to the infection.

    The Roles of Two Types of T cells in Responding to Infection

    There are two types of T cells that participate in detecting the antigens presented, and both make use of their cell-surface T cell receptor (TCR)​ proteins in recognizing and binding to the antigens on the APC. T helper (Th) cells are characterized by expression of another receptor called CD4 (in addition to TCRs), that also participates in antigen binding; Th cells are referred to as CD4​+ cells. Cytotoxic T cells instead express the CD8 receptor along with TCRs as part of APC antigen binding and are designated CD8​+ cells.

    Each of these T cell types is activated for additional immune functions by the antigen binding process, but each serves a different purpose following activation. The TCRs of activated cytotoxic CD8​+ T cells are now primed to bind foreign antigens complexed with MHC I/HLA I proteins displayed on general cells and to destroy those infected cells along with their viral contents. The activated Th CD4​+ cells can now bind and thereby activate B cells that then produce and secrete antibodies capable of binding specific epitopes of freely circulating viruses to either neutralize them by blocking entry to cells or flag them for destruction by other components of the immune system.

    Detecting Antibody Evidence of Past Infection

    Detecting a former viral infection in a person by measuring antiviral antibodies, as is currently important in epidemiological tracking of the COVID-19 pandemic, is relatively straightforward. Tests are developed using specific pieces of the very few proteins viruses possess as antigens. These are the “bait” for antibodies produced by B cells against that antigen in the person’s blood. Enzyme-linked immunoassays (ELISA) are an example of this type of immunological test. ELISAs can be fast, accurate, quantitative, sensitive, and relatively inexpensive to perform.

    MarinBio has considerable expertise in the development of ELISA​s that are quantitative down to pg/ml levels with high degrees of specificity (minimizing false positives) and sensitivity (minimizing false negatives).

    Detecting T Cell Evidence of Past Infection

    In certain instances, it can be helpful to detect evidence of a former infection arising from T cell-based immunity. For example, a recent study from Germany found evidence for twice as many past infections based on T cells that was seen with antibody (B cell-based) screening. Carrying out the tests that can detect such subtle variations in T cells requires substantially more scientific competence in cell biology than is needed to perform ELISAs, and the equipment (typically a flow cytometer) used to measure cellular characteristics demands a high degree of technical expertise and significant familiarity with sophisticated instrumentation.

    MarinBio scientists have a wealth of experience with flow cytometry derived from decades of​ experience in advanced methods in cell type detection and characterization used for cell based assays.

    Recent research focusing on T cell immunity arising from COVID-19 infection has generated results that have demonstrated the importance of T cells in combating the disease and have given indications of how vaccines might be developed that exploit the strength of T cell resistance in an enhanced immune response, in addition to that from antibodies. The data also offer some hope that previous exposure to milder human-adapted coronaviruses responsible for some common colds may confer some level of immunity or at least resistance to the SARS-CoV-2 virus that causes COVID-19 disease. The research is technically demanding; there are several sub-groups to HLA II that are associated with viral infection in particular, yet careful use of sophisticated techniques in cell biology can distinguish the T cells of this type in a population.

    In this work, researchers challenge peripheral blood mononuclear cells (PBMCs; i.e., lymphocytes and monocytes) drawn from recovered subjects with pools of synthetic peptides generated from bioinformatic predictions of SARS-CoV-2 genomic sequences to see whether an immune response will be generated. The idea is that at least some of these peptides would form epitopes similar to those that T cells, involved in a previous immune response, had encountered during the course of the infection. Their reactions in this later simulation of a repeat encounter with SARS-CoV-2 can be observed at the molecular level and provide much information on the immune status and history of these T cells.

    Characterizing T Cell Subtypes Using Marker Genes in Flow Cytometry

    There are many more cell surface receptors expressed on T helper or cytotoxic T cells than just CD4 and CD8, and much is known about the expression of these other receptors that enables them to be used as “markers” to characterize the immune state of T cells in some detail. Following incubation with the SARS-CoV-2 synthetic peptides, antibodies raised against various markers that are then tagged with different fluorescent molecules were applied to the T cells before analysis on the flow cytometer. There the cells were detected using lasers that excite the fluorescent tags for detection of discrete emission spectra for multiplex analysis.

    Using these sensitive and sophisticated measures, it could be seen that 100% of recovered subjects had T helper cells reactive to SARS-CoV-2 proteins and 80% had reactive cytotoxic T cells. Responses to the spike protein (responsible for the spiky crown of coronaviruses seen in electron micrographs) were the strongest, but evidence for reactions to membrane and nuclear viral proteins was also seen. The scientists were also able to determine that 20% of their healthy controls (blood drawn before the pandemic) had some level of resistance to SARS-CoV-2, presumably from exposure to some of the four coronaviruses responsible for approximately one-quarter of human common colds.

    Contact MarinBio scientists for more information and discussion about your needs for immune-related projects or other bioassays, immunoassays, protein chemistry or molecular biology areas.​ www.marinbio.com. Phone: 415-883-8000.​