Immunoglobulins

Immunoglobulin means “Immune Protein”. This is what we call the proteins produced by the B cells. When these immunoglobulins are embedded into the B cell membrane, they are called the B cell receptor (BCR). When they are secreted out of the mature B cell, they are called Immunoglobulins or antibodies. The term Immunoglobulin is abbreviated as Ig.

Antibodies are made up of four separate proteins connected by flexible disulfide bonds. They look a lot like a Y. They contain two long protein chains called the heavy chains. These heavy chains will change as the immune response progresses in a process called class switching. The other two chains are called the light chains. At the tip of each chain, there are the antigen binding sites which are called the Function Antigen Binding (FAB) region of the antibody. This is the unique part of the antibody that binds to a pathogen. The rest of the antibody structure is called the Fragment Crystallizable (Fc) region. This is the section of the antibody that interacts with other cells of the immune system. The Fc portion of the antibody is covered up when it is not bound to an antigen.

B cells are born in the bone marrow. They are first called Pro-B cells as they select to go down the development path to become fully mature B cells. The only thing they will have is the Immunoglobulin alpha and beta chains that act as the co-receptors for the B cell receptor. These Ig-a and Ig-b are the exact same for every B cell. They are the first part of the B cell receptor complex. The first thing that will occur will be the recombination of the Heavy chain. We covered the process of recombination. The heavy chain will pick one of each VDJ genes and make the heavy chain's variable region. Once the variable region of the heavy chain is created, it will be attached to the constant region. The heavy chain can use two different constant regions from IgM and IgD. The difference between the two is just splice variants. That means equal amounts of both IgM and IgD isotypes will be expressed on that B cell with the exact same antigen binding region. Then the B cell will express these heavy chains on its surface with surrogate light chains. The surrogate light chains are just placeholders to ensure the B cell can express a B cell receptor on the cell surface before it takes the time to do recombination of the actual light chains. Once that test is good, the B cell moves into the Pre-B cell stage. They will pull the receptors back in so they can do recombination with the VJ genes of the light chains. There are two possible light chains with Lambda and Kappa. The B cell will try to make one good light chain. If that one fails, then it will try to form the other light chain. You usually end up with equal amounts of B cells using light chains from both types. The 2 light chains actually have their own set of genes on different chromosomes with Kappa on chromosome 2 and Lambda on chromosome 22.

Once the B cell receptor is fully constructed, it will be expressed on the cell surface and it will become an immature B cell. All B cell receptors are tested against self antigens in the bone marrow before they leave. This prevents self reactive B cells from getting into circulation. This process is not as extensive as with T cells. There are no special cells here expressing all proteins with the AIRE. They just get checked for whatever antigens are in the bone marrow. Any that are self reactive will be allowed to try to make a new receptor. If they continue to fail, that B cell will undergo programmed cell death. This is called Central Tolerance. The immature B cell will leave the bone marrow and move into circulation. They will travel to the lymph nodes where they will assume their duties. They will act as both Antigen Presenting cells looking for antigens and Effector cells when they activate to become plasma cells.

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B Cells as APCs

B cells are the effectors of the Humoral immune response. They are capable of being antigen presenting cell and effector B cells. They can activate independently of helper T cells by engulfing pathogens with Toll Like Receptors. They need both activation of their TLR receptors and their B cell receptors to mount a T independent response.

B cells can also act as Antigen Presenting Cells for Helper T cells. They can engulf pathogens and break them down inside their lysosome. They can process antigens bound to MHC II and present them to the Helper T cells. They will bind their activated B71 and B72 receptors to the T helper cell's CD28 receptors for co-stimulation. They can even activate a third source of stimulation T helper cells need to activate and differentiate the helper T cell. They also connect with helper T cells using a unique connection of CD40 to the CD40 ligand. This is only used between B cells and Helper T cells. That makes it a unique target for autoimmune disorders. The activated B cell will in turn differentiate into Th2 or Th17 cells. They will release cytokines like IL-4 and IL-5 which will help the B cell differentiate into plasma cells and undergo clonal expansion. This will allow the B cells to undergo class switching and affinity maturation.

Without T helper support, the B cell can only mount a weak response using IgM antibodies. This is called a T independent response. The inclusion of helper T cells allows for a complete activation of the B cell and the ability to produce IgG, IgA and IgE. It allows for affinity maturation to enhance the immune response.

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B Cells as Plasma Cells

When the B cell is immature, it has a very small cytoplasm. Then it becomes activated and ramps up the antibody building factories. It grows its cytoplasm significantly. The antibodies receptors are exactly the same as the B cell receptor, but have one small change to the tail. Instead of being stuck in the membrane as a receptor, it is now secreted. The average plasma cell can make and secret thousands of these antibodies per second. These antibodies can travel through the blood, pass into the tissue and seek out anything that matches their receptor. They will bind to that pathogen and coat it in a process called opsonization. Once the antibody is bound to a pathogen, the Fc portion will change its shape to become active. The Fc part of the antibody is the real business end of this amazing immune protein. It has several functions it can use to interact with many of the other immune cells.

The first role of the antibody is the neutralization of the pathogen. The antibodies can coat a pathogen and make them incapable from infecting cells. This is done by blocking their ability to make entry into cells. This slows down the pathogen and infection, and it gives other immune cells a chance to clear the opsonized pathogens. The second is the Fc region of the antibody can bind C1 of the complement system. We went over this in the complement section. This will trigger the Classical complement pathway that will lead to inflammation and cell lysis. This process can also be activated by autoantibodies in autoimmune disorders. Those antibodies can bind to a cell surface antigen and trigger the classical complement pathway. This will lead to inflammation, cell damage and/or cell death. The third function is that the Fc region can bind to the Fc receptors on phagocytes and NK cells. The phagocytes are the neutrophils and macrophages. They have receptors on them for the Fc part of the antibody.

The antibodies will coat pathogens and flag them for phagocytes to come along and ingest them. Then the phagocytes will break them down using their enzymes or oxygen burst to destroy them. This process is called Antibody Dependent Phagocytosis. This can also occur with cells that the antibody binds to by binding to cell surface antigens on the cells and calling in the macrophages with their Fc receptor. The macrophage can ingest the entire cell and break it down. This is called Antibody Dependent Cellular Phagocytosis (ADCP). This process can occur in autoimmune disorders, but it also is used in therapies using antibodies to target and destroy cancer cells.

The final option is that the Fc region can bind to the Fc receptors on Natural Killer (NK) cells. This is known as the CD16 receptor. The NK cell can bind to antibodies and release its perforins and granzymes to kill the cell. This is called Antibody Dependent Cellular Cytotoxicity (ADCC). It is another key function used in clearing infections that can also be used to target cancer cells.

When an antibody binds with its matching antigen, this is called the antibody/antigen complex or Immune Complex. This is key when dealing with the antibody agglutination. Agglutination is the clumping up of the pathogens into clumps so the effector cells can deal with them quicker. Agglutination can become pathological when these Immune Complexes become deposited into the tissues. This can cause inflammation through complement activation. It can also cause tissue damage from the recruitment of and degranulation of neutrophils. Antigen/Antibody complexes drive several autoimmune disorders.

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B Cells as Memory Cells

As the B cells differentiate into plasma cells to make antibodies. Some of them will become long lived memory cells. The first response of the B cell is to produce only IgM antibodies. As the immune response progresses, the B cells will begin to class switch to IgG antibodies. These are more specific to the pathogen and have higher affinity. This becomes the indicator of a Memory B cell. Immature B cells will only ever have IgM and IgD, but a memory B cell will also display IgG on its surface. These memory B cells will often migrate to the Bone Marrow. There, they can live for months or even many years secreting low levels of protective antibodies incase of reinfection. They do undergo mitosis in the bone marrow to stay healthy. This means they can sometimes mutate and become cancerous. When this happens with the plasma cells in the Bone Marrow, we call it Multiple Myeloma. The purpose of the memory B cell is to give long term protection, but in rare cases, it can go wrong. There are about 32,000 cases of Multiple Myeloma in the US each year. Most of them are in patients over 70.

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Class Switching

All B cells have the immunoglobulins IgM and IgD on their cell surface. The DNA contains both the information to create these two classes of immunoglobulins. They will be produced in equal amounts during the B cell development. Once a B cell is activated toward a pathogen, it will begin to secrete immunoglobulins (antibodies) in the form of the IgM. This is the only thing it is able to create and secret at this point. As the infection progresses, the helper T cells will release IL-4 and IL-5 which will help in the B cells and plasma cells with switching classes. Why do they need to switch classes? Some classes are specialized toward different kinds of infections and work in different areas of the body. By class switching, the antibodies become better at doing their job. The process of class switching is done to the DNA just like with recombination. In class switching, the receptor stays completely the same, but only the constant region of the heavy chain changes. This just swaps out the Fc region with something that has more specialized functions. There are three additional classes of antibodies that an antibody can switch to with IgG, IgA and IgE. These three additional antibodies will have other different functions from the IgM that is first secreted. The Fc regions of these classes of antibodies are critical in their response.

The IgM class has a very large Fc region with three domains. It is the largest of all antibodies and not capable of leaving the blood due to its large size. They are the first antibodies released by plasma cells. They have very low affinity, but they are capable of binding their Fc regions together into a pentamer. That is five IgM bound together to form a snowflake-like structure. They are great at binding a pathogen and clumping it up to neutralize them. The main effect of IgM is neutralization. The IgM Fc region is capable of binding C1 complement and triggering the classical complement pathway which leads to inflammation. The first response trait of IgM makes it an indicator of early primary infections.

IgG is the most common of all antibodies, it is the smallest in size. It is made to cross out of the blood and into the tissue where many pathogens enter the body. It is the only antibody small enough to pass through the umbilical cord from mother to fetus. The IgG makes up 70% of the antibodies in our bodies. It has a very high affinity for its antigen. Their Fc region is able to bind C1 complement and trigger the classical complement pathway for inflammation. All of the phagocytes have IgG receptors. Phagocytes can ingest coated pathogens and destroy them. The NK cells have an IgG receptor to bind and kill infected cells. The IgG has four different subtypes in IgG1, IgG2, IgG3 and IgG4. There are only slight differences between them and their activity.

IgA is the second most abundant antibody. They are designed to work in the mucus that lines our respiratory tract and GI tracts. It is present in the saliva, tears, and even sweat. It can be passed through breast milk from mother to infant. They are capable of binding into dimers where two IgA bind their Fc regions together. This helps in neutralization. Many epithelial cells that line the mucosal surfaces can display IgA as a first line of defense for pathogens. The IgA is capable of being taken into the epithelial cells and carried across the cell and to the outside. This can protect the linings in the respiratory tract and GI tract. This process of carrying IgA across the epithelial cells is called transcytosis. The IgA comes in two subtypes of IgA1 and IgA2.

IgE is the lowest concentration in circulation. They have the larger Fc regions with three domains and bind to the Mast Cells. IgE plays a major role in protection against parasites. The IgE will sit onto the IgE receptor on the mast cell. When it binds antigen, it will cause mast cell degranulation. This mechanism that is designed to protect from parasites also harms people in allergic reactions. The formation of IgE against harmless substances like pollen leads to allergies. The activation of mast cells through activation of IgE created toward an allergen causes mast cells to degranulate and release histamine. The level of allergy can vary widely from mild to life threatening like Anaphylactic Stock.

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Affinity Maturation

Somatic Hypermutation is commonly called Affinity Maturation. This is a process by which B cells purposefully mutate their receptors. This is a process by which B cells test and mutate their receptor against a pathogen to make them better at fighting that antigen. They do this by mutating their receptors and testing them against a pathogen to see if it works better or worse. The receptors that bind better will go on while any that do worse will die through programmed cell death.

The process of affinity maturation is governed by Activation Induced Cytosine Deaminase (AID). This enzyme will randomly bind to a cytosine nucleotide in the DNA for the B cell receptor. The AID enzyme will remove the amino group from the cytosine turning it into a Uracil. The Uracil is not normal to DNA so the repair enzymes will come in and erase the bad nucleotide and randomly replace it with another. The four possible nucleotides it can replace it with are Cytosine (C), Guanine (G), Adenine (A) or Thymine (T). If it replaces the old cytosine with another cytosine, then nothing happens. There is a 75% chance it replaces the old cytosine with another nucleotide like A, G, or T. Then the repair mechanism will fix the DNA. This creates a Single Nucleotide Polymorphism (SNP). The new receptor can encode different amino acids from the old. This will allow it to bind antigens differently. Since the process is random, the outcome could end up being better or worse. There are Follicular Dendritic cells inside the lymph nodes. They will constantly present the antigens for a pathogen on their surface. The B cells will test these new receptors on the antigens. Any B cell receptors that have a stronger binding will get a signal to survive. Any that bind weaker will get a signal to undergo programmed cell death.

This process of embracing mutation in B cells toward affinity maturation leads to many mutations. Sometimes these mutations can lead to B cell cancers. It is a necessary risk by evolution that is designed to keep up with mutating pathogens, but sometimes goes awry and leads to cancer in a small percent of the population. The need to protect us from pathogens outweighs the risk of possible cancer.

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