Putting the ‘H’ in Herd Immunity — Humoral Immunity

An Overview of B-cells & Immunological Memory

Image Source: Brunei University London

With the pandemic getting under control and people getting vaccinated, we are building up herd immunity and getting one step closer to returning to normal. As a result, it is just as timely to talk about the amazing power of immunological memory which allows for vaccines to be successful in the first place!

To start off, humoral immunity is an aspect of the immune system where the production of antibodies/immunoglobulins is used to fight off antigens. This process is regulated by B-cells. B-cells are the components that produce large batches of antibodies when the body is fighting off an infection.

However, unlike cell-mediated immunity (which is regulated by T-cells and involves attacking infected cells), humoral immunity has an immunological memory component that allows it to mount a stronger, more potent immune response when it encounters the same pathogen again.

Image Source: F1000 Research

Before we dive into the details, I figured it would be a good idea to give you a rough sketch of B-cell development from stem cells to kickass fighters. First off, B-cells start off from stem cells and mature within the bone marrow. Once they become immature B-cells, they can either mature in the bone marrow or travel to the spleen.

As immature B-cells, they are in this ‘transitional’ state where they can either mature into follicular (B-2) b-cells or marginal zone (MZ) b-cells. The main difference between the two subtypes is that follicular b-cells are activated when there are stronger signals from b-cell receptors (sensors on the B-cell surface that detect antigens), while the marginal zone is activated when there are weaker signals.

When mature B-cells are activated, they create temporary structures called germinal centers. These germinal centers allow for the B-cell to produce the best antibody it can possibly create that has the highest antigen-binding strength. The winner of this competition then is selected to produce a multitude of plasma cells and memory b-cells to neutralize the infection.

The plasma cells secrete a large number of antibodies while memory b-cells circulate the bloodstream and neutralize pathogens. Once an infection passes, several from each group can reside in the body long-term, where it can be activated when it encounters the same pathogen. These elements allow for the formation of immunological memory.

It all starts in the bone marrow

As the name suggests, B-cells start development in the bone marrow, which is a spongy tissue inside your bones that contains stem cells. These stem cells can differentiate into red and white blood cells that can serve a variety of important functions in the body. For the purposes of this article, we’re only going to focus on white blood cells as they fight infections.

Stem cells are special cells that can develop into many different cell types whether it be muscle, tissue, or brain cells. Specifically, hematopoietic stem cells (HSCs) are stem cells that can develop into all types of blood cells, including our resident B-cell. From there, the hematopoietic stem cells can differentiate into progenitor cells, which are essentially like stem cells, but more specified into what they can differentiate into.

You can think of it in terms of the color wheel. Primary colors are considered the base colors that you can mix into varying combinations where the results of these combinations can be mixed to yield further variations. Secondary and tertiary colors are the result of continued combinations of other colors located higher on the hierarchy. In the same way, the primary colors are the ‘stem cells as they are the source of all the other colors that can be created. You can’t create primary colors and you can’t mix any of the colors located lower on the hierarchy to return to your primary colors.

Image Source: Sigma-Aldrich

Anyways, the hematopoietic stem cells can differentiate into two progenitor cells (our secondary colors): the lymphoid progenitor and the myeloid progenitor. Lymphoid progenitor cells give rise to B-cells, T-cells, and Natural Killer Cells, which can further be developed into their own subtypes. All of these cells are white blood cells or lymphocytes.

The process of a hematopoeitic stem cell developing into a mature B-cell can take up to 1–2 weeks. Before we jump into that, let’s talk about one of the most important parts of a B-cell: antibodies!

Antibodies

Antibodies, or immunoglobulins (Ig), are proteins that can identify and neutralize pathogens (i.e. infectious bacteria, fungi, viruses, etc.). If we look at the structure, there are four main parts: variable region, constant region, heavy chain, and light chain.

Image Source: MicrobiologyInfo.com

The variable region fits its name to a T: involved in antigen-binding and these locations vary between different antibody molecules which allow the body to target a wide range of pathogens in a specific manner.

On the other hand, the constant region makes up the Y base of the antibody, and in comparison, is less variable and interacts with fewer effector cells and molecules. Effector cells are cells that can respond to a stimulus and bring about a biological response. In this case, effector cells can be T cells that can elicit a response in B-cells (more on that coming later).

Light chains and heavy chains are proteins that link together to form antibodies. Light chains are involved in the expression and secretion of functional antibodies as well as increasing antigen-binding specificity and variability. Heavy chains are involved in determining the effector function of an antibody. There are two types of light chains: lambda (λ) and kappa (κ). There is no functional difference between the two types and immunoglobulins either have one or the other, but they never have one of each. In contrast, there are 5 types of heavy chains: μ, δ, γ, α, and ε (mu, delta, gamma, alpha, and epsilon). These classes are called isotypes.

In terms of structure, immunoglobulins are made up of:

  • 2 identical heavy chains (Variable Heavy Chain [VH] and Variable Light Chain [VL])
  • 2 identical light chains (Constant Heavy Chain [CH] and Constant Light Chain [CL])

These chains are linked together by disulfide bonds. Within the chains, they contain a series of protein domains, which are distinct structural units in a protein. Light chains contain 2 domains while heavy chains contain 4.

Image Source: ABclonal.com

As if we haven’t talked enough about structure, I want to take you up to another level of antibody structure. Now that we know the component parts, we can circle back to the whole idea of constant and variable regions.

The constant region corresponds to Fc (fragment crystallizable region) while the variable region corresponds to Fab (antigen-binding fragment).

The Fc region is responsible for the biological activity of the antibody, where it can bind to specific proteins and cell receptors to ensure the appropriate immune response for a specific antigen. The Fab region is responsible for antibody specificity and antigen-binding. It also contains Fv (variable domains) regions that are the most important for binding. Essentially, Fv only represents the topmost part of the variable region that binds to the antigen while the Fab region represents the entire variable region.

If you can’t tell, we biology people love our classifications and our subtypes. However, I think understanding them and creating a recap might be useful if you get lost (honestly for my own personal reference too as I write this haha)

There are 5 different classes of immunoglobulins: IgM, IgD, IgG, IgA, IgE. Where do they get their names you ask? They correspond to the five different heavy chain types or isotypes. The main difference between them is their variable regions which are specific to different antigens. Each class can be further split up into subclasses.

Two types of light chains: lambda (λ) and kappa (κ)

5 Types of heavy chains: mu (μ), delta (δ), gamma (γ), alpha (α), and epsilon (ε)

Fab corresponds to a variable region that binds to antibodies. There are 2 Fab arms on each antibody with identical domains. The Fv regions are specifically for binding as It contains a paratope, which is the part of the antibody that binds to an antigen.

Fc corresponds to the constant region (or the base of the Y) which binds to proteins that can elicit immune responses against antigens.

B-Cell Development

Now that we have an understanding of antibody structure, what is crucial is understanding how exactly that translates into the different stages of development. When we think of how the antibodies on a B-cell are specific to only ONE antigen, it raises the question of, how exactly is our body able to respond to a diverse amount of pathogens when B-cells focus on specific targeting? Interestingly enough, as a B-cell develops, there are different mechanisms it encounters along the way that allows for the creation of functional and diverse B-cell combinations.

Image Source: https://www2.nau.edu/~fpm/immunology/Exams/Bcelldevelopment-401.html#:~:text=Immature B cells are very,naive (resting) B cells.

VDJ stands for Variable, Diversity, Joining. Each letter corresponds to different gene segments that are combined together to create unique antigen receptors that can recognize a variety of molecules. Essentially, you can think of it like a large game of Set, where you have to make find patterns within the cards placed on the table.

Just as you are taking different cards and combining them to form patterns, recombination in the biological sense deals with rearranging genetic material by joining different segments of DNA.

Set has 3 colors (red, green, purple), 3 shapes (diamonds, rounded rectangles, squiggles), and 3 patterns (open, stripes, colored). Sets must fall in line within the following patterns: 1–2–3, same shape, same color, no correlation. For instance, in a 1–2–3 set, you can have them be:

  • 1) all the same color, same pattern, but they must have different shapes
  • 2) same shape, same color, but different patterns
  • 3) all different shapes, different patterns, different colors.

Also, as you are playing the game, you want to find as many sets as quickly as possible in order to win.

Cards from Set | Image Source: Brilliant

In the same way, VDJ recombination is about finding a functional, unique arrangement of the different gene segments (V, D, J) as quickly as possible. Once a functional design is created, the cell shuts off rearrangement and expression of another allele on the homologous chromosome. While Set can be played solo or with as many people as you’d like, in this case, we would have only 2 players to represent chromosomes (we have 2 of each chromosome in our genome) playing 3 rounds.

Image Source: Wikipedia

In VDJ Recombination, there are 3 steps: 1) D-J recombination 2) V-DJ recombination 3) VDJ-C recombination. This process is initiated by an enzyme complex called RAG1/RAG2. All the different gene segments contain a ‘tag’ called a recombination signal sequence (RSS) that is recognized by RAG1/RAG2. This complex creates a double-strand break (DSB) — clean cut– between the coding sequence and RSS.

Coding sequences can be combined to form a coding joint while RSS sequences can be combined to form a signal joint. The coding joints create our heavy and light chains (D-J or V-DJ for heavy chains; V-J for light chains) while the signal joints create a DNA loop that will delete unnecessary DNA present in between gene segments.

Once the different chains are created, junctional diversity acts as a mechanism that introduces even more diversity into all of the generated VDJ segments. It randomly inserts bases at junctions of V, D, and J segments via the Tdt (Terminal deoxynucleotidyl transferase) enzyme.

The final two stages of development are immature B-cell and mature B-cell. The process of generating heavy and light chains and undergoing the different recombination mechanisms take place in the previous steps of development. In the immature B-cell stage, the cell can successfully express IgM on the surface. During this stage, the cell is very sensitive to antigen-binding that they can bind self-antigen (antigens on your own cells) in the bone marrow. When this occurs, the ones that do are eliminated. Malfunctioning of this step can lead to autoimmune disorders where B-cells target healthy tissue within patients.

Finally, mature B-cells leave the bone marrow to travel to secondary lymphoid organs such as the spleen and lymph nodes. On their cellular membrane, they express both IgM and IgD. They become known as mature naive (resting) B-cells as they have not been activated yet.

B-Cell Activation

B-cells can be activated and generate two types of immune responses: T-independent and T-dependent. In T-independent responses, B-cells can directly respond to the antigen while T-dependent responses require T-cells to assist in generating a response.

Image Source: Lumen Learning

T-independent responses are when B-cell receptors bind directly to antigens that have repetitive epitopes. Epitopes are parts of an antigen molecule that an antibody binds to. When this occurs, a second signal is needed to mount an immune response, either from toll-like receptors (proteins from the innate system that activate immune responses when physical barriers were breached) or interactions with factors from the complement system (components that enhance the ability of antibodies during an immune response).

Following activation, the B-cell undergoes clonal proliferation, where it selects and reproduces one type of cell. In this case, it would be plasma cells, which are a type of B-cell that secrete antibodies in large numbers following infection. During this transition, the B-cell receptors disappear from the cell surface and the plasma cell secretes large quantities of IgM molecules.

T-independent responses are short-lived and do not produce memory B cells. As a result, there is no secondary response, since there is no immunological memory that it can work off of when it encounters that antigen.

Image Source: Lumen Learning

In comparison, T-dependent activation is more complex as it mounts a stronger immune response and generates immunological memory. B-cells bind to antigens where they are internalized (through endocytosis) and broken down. From there, the B-cell’s MHC II proteins present the processed antigens to helper T-cells. The T cell’s CD4 molecule interacts with MHC II and allows for antigen recognition. Once that occurs, the T-cell produces and secretes cytokines which serve as a GO button for B-cells to undergo clonal expansion.

Making a Clone Army aka Clonal Expansion

When you’re fighting a war, one of the best advantages you can have is strength in numbers. Similarly, clonal expansion is the process of intense cloning of a parent cell to create a multitude of daughter cells.

Image Source: Immunopaedia.org

Once the mature B-cell gets the go signal, it will either become a plasmablast or a germinal center. Plasmablasts are short-lived plasma cells that secrete a large number of antibodies (but not as many as normal plasma cells). They can divide rapidly, internalize antigens, and present them to T-cells. On the other hand, mature B-cells become blasting B-cells, which later develop into germinal centers.

Image Source: British Society for Immunology

Blasting B cells form temporary structures called germinal centers where further differentiation can occur. Germinal Centers (GCs) are structures that allow for B-cell proliferation, differentiation, and mutation of antibody genes. The main goal of the germinal center is to produce B-cells with a high-affinity receptor for the target antigen.

The germinal center has two zones, a light, and a dark. The process begins in the dark zone, where B-cells rapidly proliferate and undergo somatic hypermutation — a process to develop antibodies with stronger antigen-binding strength through single point mutations (single base pair alterations).

Additionally, class switch recombination can occur, which focuses on changing the Y base section of the antibody. by doing so, this mechanism changes what class of immunoglobulin the B-cell is producing (i.e. from production IgM to IgG). Unlike somatic hypermutation, this process focuses on changing out the constant region, which causes the effector function of the antibody to change. That means that antigen-binding remains the same, but now, the B-cell’s Y-base can interact with other proteins that it previously couldn’t in its original configuration.

During this process, several rounds of somatic hypermutation and class switch recombination occur where the B-cells migrate back and forth between the zones.

Image Source: British Society for Immunology

Next, the B-cells enter the light zone and compete with each other for antigen. Whichever B-cell clone has the highest binding affinity will survive and exit the germinal center as either plasma cells or memory B cells.

The plasma cells will secrete antibodies several weeks after activation. Afterward, it will migrate to the bone marrow and reside indefinitely where it will be activated if it encounters the same pathogen again. In contrast, memory B-cells will circulate throughout the body looking for antigens and stopping infection. Once things are stable, the memory B–cell will reside in different lymphoid organs such as the bone marrow, tonsil, and spleen or continue to circulate the bloodstream until activated.

Contrary to what we typically think, antibodies aren’t used to directly kill pathogens. They merely bind and target pathogens, while signaling to other destructive immune cell types to annihilate the enemy.

This process is called opsonization, where antibodies mark pathogens to recruit cells that will destroy target cells. This is done using one of the following mechanisms: Antibody-Dependent Cellular Phagocytosis (ADCP), Antibody-Dependent Cellular Cytotoxicity (ADCC), and Complement-Dependent Cytotoxicity (CDC). It’s kind of like an SOS signal where ships launch fireworks into the air hoping that other ships nearby will be able to see it and respond to save the sinking ship.

Image Source: Creative Biolabs

Both ADCP and ADCC are dependent on Fc receptors of effector cells binding to the Fc region (Y-base/constant region) of the antibody.

In ADCC, once binding occurs, non-specific cytotoxic T-cells — a type of T-cell that kills infected or damaged cells through apoptosis — effector cells are activated and substances (i.e. perforin, granzyme, TNF) that cause cell death are released.

For ADCP, the antibody binds to the target cell and induces phagocytosis. Phagocytosis is a cellular process that removes pathogens and dead/dying cells. Cells that perform phagocytosis, phagocytes (i.e. monocytes, macrophages, neutrophils, dendritic cells), link their Fc receptor to the antibody’s Fc domain and begin destroying them.

The last process, CDC is a mechanism where antibody-coated target cells activate the complement system which leads to cell lysis. Lysis is the destruction of a cell through rupturing the cell wall or membrane. The complement system is a part of the immune system that enhances the ability of antibodies and phagocytic cells to get rid of microbes and damaged cells.

When antibodies bind to an antigen, they form an immunocomplex. When IgG or IgM binds to the antigen, a protein complex from the complement system, C1q can bind to the immunocomplex. This interaction gives rise to the formation of a Membrane Attack Complex (MAC), which is an effector of the immune system that causes cell lysis.

Closing Thoughts

If I’m being honest here, part of the reason that I wrote this is for a school presentation more than my genuine interest in the topic. And I figured that this would be a good way to practice explaining all these complex components while also actually posting an article on my Medium since it’s been a while (killing 2 birds with one stone 😉 ).

So it was kind of a surprise that I enjoyed this far more than I expected. Yes, I’m into bio, but I was never really interested in medicine or immunology. I also did some surface-level research into B-cell technology, and it’s interesting how scientists are working on developing treatments such as cancer vaccines using B-cells or creating a B-cell depletion therapy, which aims to eliminate cancerous B-cells. Not going into that because I rambled way too long in this article, but if anyone is interested, I’ll consider writing about it.

Also, even though things are opening up and we’re all getting vaccinated, I still hope that everyone can stay safe and remain cautious as we go back to normal. Please take care of your health! Make things a smidge easier for your humoral immune system.

Hi! My name is Maggie and I am an ambitious 16-year-old looking to impact the world through emerging biotech. At the moment, I’m exploring topics such as biocomputing, philosophy, ethics, and climate change.

If you got to the end, thank you for reading my article! Feel free to connect with me on Linkedin or sign up for my personal newsletter if you would like to receive monthly updates on my biocomputing journey!

A 17-year-old who knows less about life than she thought she did. Fascinated by biology, specifically biocomputing, synthetic biology, and bioinformatics.