MCB 321

Cells of the immune system

Immunity is the ability of an organism to resist infection. The human immune system employs a
two-pronged defense against invading pathogens. Innate immunity or natural immunity
(nonspecific immune response), the first of these interconnected defensive mechanisms, is the
built-in capacity of the immune system of multicellular organisms to target common pathogens
regardless of their identity. Innate immunity does not involve recognition of a specific microbe
i.e. lack immunological memory. Innate immunity first-line defenses include skin and mucous
membranes, and the second-line defenses include natural killer cells, phagocytes, inflammation,
fever, and antimicrobial substances. Innate immune responses represent immunity’s early-
warning system and are designed to prevent microbes from gaining access into the body and to
help eliminate those that do gain access. By contrast, adaptive or specific immunity (specific
immune response) is based on a specific response to a specific microbe once a microbe has
breached the innate immunity defenses. Moreover, the effectiveness of specific immune
responses increases on repeated exposure to foreign agents such as viruses, bacteria, or toxins;
that is to say, specific responses have “memory. Adaptive immunity involves lymphocytes (a
type of white blood cell) called T cells (T lymphocytes) and B cells (B lymphocytes).
The immune response results from the activities of a wide variety of specialized cells that
circulate throughout the body, primarily through the blood and lymph, a fluid similar to blood
but which lacks red blood cells. Blood and lymph interact directly or indirectly with every major
organ system.
Blood consists of fluid, called plasma, and formed elements— that is, cells and cell fragments
suspended in plasma. The formed elements include erythrocytes, or red blood cells (RBCs);
leukocytes, or white blood cells (WBCs); and platelets. Platelets, which are actually fragments
arising from large cells called megakaryocytes, are important for blood clotting. White blood
cells, or leukocytes, are important in all host defenses. The average adult has approximately
7,400 leukocytes per mm3 of blood. This average value shifts substantially during infectious and
allergic responses of the host. Formed elements are created in red bone marrow by stem cells in
a process called hematopoiesis (he¯m-a-to¯-POY-e¯-sis). Hematopoietic cells are capable of
long-term self-renewal, meaning they can divide repeatedly. Hematopoietic stem cells are
induced to develop into the various types of blood cells by a group of proteins called colony-
stimulating factors (CSFs). It begins when a cell called a pluripotent stem cell develops into two
other types of cells called myeloid stem cells and lymphoid stem cells. From these two stem
cells, all of the formed elements develop. Leukocytes are divided into two main categories based
on their appearance under a light microscope: granulocytes and agranulocytes. Granulocytes owe
their name to the presence of large granules in their cytoplasm that can be seen with a light
microscope after staining. They are differentiated into three types of cells based on how the
granules stain; neutrophils, basophils, and eosinophils.
Neutrophils stain pale lilac with a mixture of acidic and basic dyes. Neutrophils are also
commonly called polymorphonuclear leucocytes (PMNs), or polymorphs. The term
polymorphonuclear refers to the fact that the nuclei of neutrophils contain two to five lobes.
They normally account for over half of the circulating white blood cells, and their numbers
increase during most bacterial infections. Neutrophils, which are highly phagocytic and motile,
are active in the initial stages of an infection. They have the ability to leave the blood, enter an
infected tissue, and destroy microbes and foreign particles.
Basophils stain blue-purple with the basic dye methylene blue. Basophils (and mast cells)
possess high affinity receptors for one type of antibody, known as immunoglobulin E (IgE), that
is associated with allergic responses. When these cells become coated with IgE antibodies,
binding of antigen to the IgE can trigger the secretion of histamine. Histamine are important in
inflammation and allergic responses. Mast cells are similar in appearance and function to
basophils but are found in tissues rather than blood. They do not come from the same precursor
cells as basophils. Mast cells are important in the inflammatory response and are responsible for
many allergic reactions.
Eosinophils stain red or orange with the acidic dye eosin. Eosinophils are somewhat phagocytic
and also have the ability to leave the blood. Their major function is to kill certain parasites, such
as helminths or protozoan. Although eosinophils are physically too small to ingest and destroy
helminths, they can attach to the outer surface of the parasites and discharge peroxide ions that
destroy them. Their number increases significantly during certain parasitic worm infections and
hypersensitivity (allergy) reactions.
Agranulocytes also have granules in their cytoplasm, but the granules are not visible under the
light microscope after staining. There are three different types of agranulocytes: monocytes,
dendritic cells, and lymphocytes.
Monocytes: Monocytes [Greek monos, single, and cyte, cell] are mononuclear leukocytes with
an ovoid- or kidney-shaped nucleus and granules in the cytoplasm that stain gray-blue with basic
dyes. They are produced in the bone marrow and enter the blood, circulate for about eight hours,
enlarge, migrate to the tissues, and mature into macrophages or dendritic cells. Because
macrophages [Greek macros, large, and phagein, to eat] are derived from monocytes, they are
also classified as mononuclear phagocytic leukocytes. However, they are larger than monocytes,
contain more organelles that are critical for phagocytosis, and have a plasma membrane covered
with microvilli.
Macrophages have surface molecules that function as receptors to nonspecifically recognize
common components of pathogens. These receptors include mannose and fucose receptors and a
special class of molecules called toll-like receptors which bind lipopolysaccharide (LPS),
peptidoglycan, fungal cell wall components called zymosan, viral nucleic acids, and foreign
DNA. In addition, macrophages possess specialized scavenger receptors including a surface
protein called CD14, which also binds LPS. Macrophages also have receptors for antibodies and
serum glycoproteins known as complement. Both antibody and complement proteins can coat
microorganisms or foreign material and enhance their phagocytosis. This enhancement is termed
opsonization.
Dendritic cells are also believed to be derived from the same precursor cells as monocytes. They
have long extensions that resemble the dendrites of nerve cells, thus their name. Dendritic cells
are especially abundant in the epidermis of the skin, mucous membranes, the thymus, and lymph
nodes. Dendritic cells destroy microbes by phagocytosis and initiate adaptive immune responses
by collecting antigens from the tissues and then bring them to lymphocytes that gather in the
secondary lymphoid organs (e.g., lymph nodes, spleen, appendix, tonsils). This is a process
known as antigen presentation. Antigen presentation triggers the activation of T cells, which are
critical for the initiation and regulation of an effective specific immune response. Thus not only
do dendritic cells destroy invading pathogens as part of the innate response, but they also help
trigger specific immune responses.
Lymphocytes: Lymphocytes are specialized leukocytes derived from lymphoid precursor cells.
There are three types of lymphocytes: B cells (B lymphocytes), T cells (T lymphocytes), and
natural killer cells (innate lymphoid cells (ILCs) i.e. lack specificity in their mechanism of
antigen recognition). Mature B and T cells circulate through the blood and lymph system but are
concentrated in the lymph nodes and spleen where they interact with antigens. T cells and B cells
are not usually phagocytic but play a key role in adaptive immunity.
B cells originate and mature in the bone marrow and fetal liver. In birds, B cells are
differentiated in the bursa of Fabricius. Like dendritic cells and macrophages, B cells are
specialized APCs, but in addition to this, they are the precursors of antibody producing plasma
cells. Antibodies (immunoglobulins) are soluble proteins that interact with specific antigens and
are produced by B cells and plasma cells (a mature activated B cell).
T cells, which interact with antigens presented by APCs, begin their development in the bone
marrow but migrate to the thymus to mature. Unlike B cells, however, T cells do not secrete
antibodies. Activated T cells produce and secrete proteins called cytokines. Cytokines can have
various effects on other cells including other T cells, B cells, granulocytes, and other somatic
cells. In some cases, the cytokines stimulate cells to mature and differentiate, produce new
effector products, and even cause some cells to die. Because B and T cells must be activated by
specific antigens, they are included in the adaptive or specific immune system.
Natural killer (NK) cells: Natural killer (NK) cells are a small population of large, non-
phagocytic granular lymphocytes that play an important role in innate immunity. They are found
in blood and in the spleen, lymph nodes, and red bone marrow. Natural killer cells cells have the
ability to kill a wide variety of infected body cells and certain tumor cells. Natural Killer cells
attack any body cells that display abnormal or unusual plasma membrane proteins. The binding
of NK cells to a target cell, such as an infected human cell, causes the release of toxic substances
from lytic granules in NK cells. Lytic granules are a secretory organelle unique to NK cells.
Some granules contain a protein called perforin, which inserts into the plasma membrane of the
target cell and creates channels (perforations) in the membrane. As a result, extracellular fluid
flows into the target cell and the cell bursts, a process called cytolysis (s¯-TOL-i-sis; I cyto- =
cell; -lysis = loosening). Other granules of NK cells release granzymes, which are protein-
digesting enzymes that induce the target cell to undergo apoptosis, or self-destruction. This type
of attack kills infected cells but not the microbes inside the cells; the released microbes, which
may or may not be intact, can be destroyed by phagocytes.
In mammals, the bone marrow and thymus are called primary lymphoid organs because they are
the sites where lymphoid stem cells mature into functional, antigen-reactive lymphocytes

Antigens
Substances that induce production of antibodies are called antigens (short for antibody
generators). Most antigens are either proteins or large polysaccharides. Lipids and nucleic acids
are usually antigenic only when combined with proteins and polysaccharides. Pathogens can
have multiple antigenic sites. Components of invading microbes—such as capsules, cell walls,
flagella, fimbriae, bacterial toxins, and viral coats—all tend to be antigenic. However, a
compound doesn’t have to be part of an invading pathogen to be deemed antigenic by the
immune system. Non-microbial antigens may include pollen, egg white, blood cell surface
molecules, serum proteins from other individuals or species, and surface molecules of
transplanted tissues and organs. Detection of an antigen provokes production of highly specific,
corresponding antibodies. Antigens that elicit an immune response are often known
as immunogens. Generally speaking, antibodies react with specific regions on antigens called
epitopes or antigenic determinants. The nature of the interaction depends on the size, shape, and
chemical structure of the binding site on the antibody molecule. Similarly, epitopes can be
displayed by antigen presenting cells (such as when macrophages and dendritic cells present
them to T cells). A bacterium or virus may have several epitopes that cause the production of
different antibodies. Most antigens have a molecular mass of 10,000 da or higher. A foreign
substance that has a low molecular mass is often not antigenic unless it is attached to a carrier
molecule. These low molecular-mass compounds are called haptens. Allergic reactions are a type
of hypersensitivity reaction that occurs when the immune system reacts to something that is
normally non-pathogenic, such as pollen. In these people, when penicillin combines with host
proteins, the resulting combined molecule initiates an immune response.

Antibodies
Antibodies are immunoglobulins (Igi) which can react specifically with the antigen which
stimulate their production. These are large, Y-shaped blood proteins produced by plasma cells or
B lymphocyte. B cells are lymphocytes that specialize in antibody production and contain
antibodies (B cell receptors, BCRs) on their surfaces; each B cell has an estimated 100,000
BCRs of identical antigen specificity. They are found in the serum and in other body fluid, and
tissues, including urine, spinal fluid, lymph nodes, spleen, etc. Immunoglobulins comprise about
20% of total plasma proteins. They bind to foreign particles and invade them. These particles are
foreign bodies that get attacked by Antibody. Antigens are foreign pathogens that invade the
body and have the capability to give rise to a response from our immunity system either by
grouping up with a larger molecule or alone after binding with antibodies for a particular
immune response. Hence, antigens stimulate the production of antibodies by the immune system.

Antibody Structure
An antibody has a Y-shaped structure, made up of four polypeptide subunits. Each subunit has
two identical light and heavy chains. Each light chain has about 220 amino acids, and each heavy
chain has about 440 amino acids. Both light (L) and heavy (H) chains contain two different
regions (constant and variable). Constant (C) regions (CL and CH) have amino acid sequences
that do not vary significantly between antibodies of the same class. The variable (V) regions (VL
and VH) from different antibodies have different amino acid sequences. It is the variable regions
(VL and VH) that, when folded together, form the antigen-binding sites. The four chains are
covalently linked by disulfide bonds. This breakage produces two identical Fab fragments, which
carry the antigen-binding sites, and one Fc fragment (crystallizable fragment), which is involved
in placental transfer, complement fixation, attachment to various cells, and other biologic
activities. The number of antigen-binding sites on an antibody is called the valence of that
antbody. For example, most human antibodies have two binding sites; therefore, they are
bivalent. Because a bivalent antibody has the simplest molecular structure, it is called a
monomer. An L chain consists of one variable domain (VL) and one constant domain (CL). The
L chains can be either κ (kappa) or λ (lambda) and their classification is made based on amino
acid differences in their constant regions. Although, an individual immunoglobulin molecule has
either k or λ chain, not both. Most H chains consist of one variable domain (VH) and three or
more constant domains (CH). L and H chains are subdivided into variable regions and constant
regions. The regions are composed of three dimensionally folded, repeating segments called
domain, each domain is approximately 110 amino acids long. Variable regions are responsible
for antigen binding; constant regions are responsible for the biologic functions which is involved
in placental transfer, complement fixation, attachment to various cells, and other biologic
activities.

Types of Antibodies

The amino acid sequence of heavy-chain constant domains determines antibody class, the heavy
chain called gamma (g) defines the IgG class. Likewise, alpha (a) defines IgA; mu (μ) defines
IgM; delta (d) defines IgD; and epsilon (e) defines IgE.

IgG
Each IgG molecule consists of two L chains and two H chains (molecular formula H2 L2).
Because it has two identical antigen-binding sites, it is divalent. It circulates in the blood and
lymph and constitutes 50-80% of total antibodies present in serum. Therefore, IgG is the
predominant antibody in secondary immune responses and provides the longest-term protection
of any antibody class. It has the molecular weight of 180,000 daltons. It is the only antibody able
to cross the placenta and provide passive immunity to the foetus. IgG is divided into four
subclasses- IgG1, IgG2, IgG3, and IgG4. Among these, only IgG3 and IgG4 possess the ability
to cross the placenta. These can also pass the walls of blood vessels and enter in tissue fluid.
These can bind to bacteria and viruses, and also can neutralize the toxins secreted by them. After
binding the antigen, these enhance the effectiveness of phagocytic cells to engulf and ingest
them. It is also one of the two immunoglobulin classes that activate complement by the classical
pathway. The normal IgG level in adult serum is 1000-1500mg/dL.

IgM
IgM is the first antibody produced in response to a microbial attack by B cells. It is composed of
five H2 L2 units (each similar to one IgG unit) and one molecule of J (joining) chain. The
pentamer (MW 900,000 daltons) has a total of 10 identical antigen-binding sites and thus a
valence of 10. It has a large number of antigenic sites on its surface and therefore facilitates
efficient activation of the immune system. It circulates in the blood and lymph and constitutes 5-
10% of the total antibody content in the serum. IgM are especially effective at cross linking
particulate antigens and causing their aggregration because of its numerous antigen binding site.
It is the most efficient immunoglobulin in agglutination (clumping), complement fixation, and
other antigen–antibody reactions and is important also in defense against bacteria and viruses. It
can increase the digestion of target cells by the phagocytic cells as IgG. Since IgM does not cross
the placenta, IgM antibodies in the newborn are thus considered a sign of intrauterine infection.
The normal adult serum level is 60-180mg/dL and this level is reached 6-9 months after birth.

IgA

IgA is mainly produced by the plasma cells that reside in the mucosa-associated lymphoid tissues
(MALT). Recall that plasma cells are the antibody-secreting form of B cells. IgA, when
transported from the mucus-associated lymphoid tissue to mucosal surfaces, acquires a protein
called the secretory component. It constitutes 15 % of the total antibody content in the serum
and is divided into 2 sub-classes- IgA1 and IgA2. Among these, IgA1 is highly found in the
secretions and is also called the secretory immunoglobulin. It exists in both monomeric as well
as dimeric forms. The dimeric forms consist of two H2 L2 units and one molecule each of J
chain peptide and secretory component that aids in transport of IgA across membranes. They are
found in body secretions e.g. saliva, sweat, tears, serum and secretion from respiratory,
genitourinary tracts, gastrointestinal tract and colostrum (breast milk) as well. IgA in breast milk
protects an infant’s gastrointestinal tract from infection. It provides the first line of defense
against the pathogens and limits inflammation. Secretory IgA can neutralize viruses and can
inhibit attachment of bacteria to epithelial cells. It has the molecular weight of 360,000 daltons.
The normal adult serum level is 100-400mg/dL.

IgD

It accounts for only 0.2% of total antibodies of serum. It has the molecular weight of 180,000
daltons. They are abundant in combination with IgM on the surface of B cells and thus are part
of the B cell receptor complex. Therefore their function is to signal the B cell to start antibody
production upon antigen binding. Like others, they cannot pass across the placenta and help the
new baby to initiate the immune response as their population remains very high on the surface of
B-cells. It is involved in the production of the antibody by B cells. It is present as a monomer and
weighs around 1,80,000 daltons. It acts as a receptor on B cell surface and participates in B cell
activation and differentiation. Normal adult serum level is about 3-5mg/dL.

IgE
IgE is present in the least amounts, around 0.002% of the antibody content in the serum. It has
the molecular weight of 200,000 daltons. They are a little larger than IgG. This is found as a
monomer in the body. These are present in the linings of the respiratory and intestinal tracts and
respond to allergic reactions. IgE also binds to tissue mast cells, basophils and eosinophils,
where subsequent binding of antigen to antigen-binding sites of IgE causes degranulation of the
mast cells. The ensuing release of chemical mediators, such as histamine and serotonin, triggers
immediate-type hypersensitivities (a type of allergic response). In a highly allergic person,
abnormally a high concentration of IgE is found. Unfortunately for allergy sufferers, basophils
and mast cells also release their chemicals when IgE binds to normally harmless materials such
as foods, dusts, and pollens, leading to immediate reactions such as coughing, sneezing, and
tissue swelling. Normal adult serum level is about 0.03mg/dL.

Functions of Antibody
Following are some of the key functions of antibody:
(i) Binds to pathogens.

(ii) Activates the immune system in case of bacterial pathogens.

(iii) Directly attacks viral pathogens.

(iv) Assists in phagocytosis.

(v) Antibody provides long-term protection against pathogens because it persists for years after
the presence of the antigen.

(vi) It neutralizes the bacterial toxins and binds the antigen to enhance its efficiency.

(vii) They also act as the first line of defense for mucosal surfaces.

(viii) They ingest cells by phagocytosis.

(ix) Few antibodies bind the antigen present on the pathogens. These aggregates the pathogen
and
they remain in secretions. When the secretion is expelled out, the antigen is also expelled.

The antibody serves to coat and identify the non-self agent as a target for immunological attack
and to activate nonspecific immune responses that can destroy the target. Phagocytes have Fc
receptors for immunoglobulin on their surface, so bacteria that are covered with antibodies are
better targets for phagocytosis by neutrophils and macrophages. This is termed opsonization.

Production of Antibodies

Antibody production may or may not depends on T antigens.

Antibody production against T-dependent antigens: Antibody production by B-cells is also

cooperated by the other cells such as T-cells and macrophages. For example, B-cells produce
antibodies in cooperation with T-cells against a type of antigen known as T-dependent antigen.
The T-dependent antigen consists of bacteria, some of proteins, RBCs etc. The macrophages or
dendritic cells (highly branched cells) play a significant role. The B-cells need the help of
antigen presenting cells (APC) (i.e the macrophages and the dendritic cells that attack the T-
dependent antigen and partially digest them and specialized helper T cells to produce antibodies
against T-dependent antigens. The process is accomplished in the following steps:

The APC attaches the T-dependent antigens and partially digest them. The APC usually takes up
the polypeptide fragments of the antigens and keep them on the cell surface. The specific T-cells
which are known as helper T-cells interact with the APC. The antigen fragments and certain self-
antigens are recognized by helper T-cells and carry them on the surface of APC. The self-
antigens are actually the cell surface proteins that are the components of the major
histocompability complex. They serve as the self- antigens, the signals by which immune system
distinguishes the self from non-self. The foreign antigen is recognized by the helper T-cells only
when it gets combined with a self -antigen on the surface of APC. This results in stimulation of
suitable B-cells. For that the possession of self antigens by B-cells is very necessary. Helper T-
cells are required by all immune responses that induce IgG, IgM and IgE. However a single
complex of the antigen and the self- antigen present on the cell surface are recognized by T-cell
receptors.

Antibody production against T-independent antigen: The T-independent antigens without the
interference of T-cells can also induce a response from B-cells. The T-independent antigens
contain the repeating subunits of polysaccharide or protein. For example bacterial flagella
(composed of proteins) and lipopolysaccharide layer of Gram-negative bacteria. Multiple bonds
are formed with B-cells by these antigens. But the immune response by these antigens is weaker
as compared to T-dependent antigens. They bind to B-cells which produce antibodies only of the
class IgM.

Complement system
The complement system consists of over 30 proteins produced by the liver that circulate in blood
serum and within tissues throughout the body. The complement system, or simply complement,
is a group of sequentially interacting proteins, many with enzymatic activity, that functions to
boost the efficiency of both innate and adaptive immune responses for the destruction of
pathogens. This is another example of the cooperation between the innate and adaptive immune
systems. Together, proteins of the complement system destroy microbes by cytolysis,
opsonization, and inflammation and they also prevent excessive damage to host tissues.
Some of the complement proteins are opsonins in that they bind to microbial cells, coating them
for recognition by phagocytes. Additionally, other complement proteins are strong chemotactic
signals that recruit phagocytes to the site of their activation. Still other complement proteins
puncture cell membranes to cause lysis. Interestingly, one of the several triggers that can activate
the complement process is the recognition of specific antibody on a target cell. Complement
proteins are produced in an inactive form; they become active following enzymatic cleavage.
Activated fragments (products) carry out the destructive actions. Complement proteins are
usually designated by an uppercase letter C and are numbered C1 through C9, named for the
order in which they were discovered. Activated fragments are indicated by lowercase letters a
and b. For example, inactive complement protein C3 is split into activated fragments, C3a and
C3b. Complement proteins act in a cascade, where one reaction triggers another, which in turn
triggers another. More product is formed with each succeeding reaction in the cascade,
amplifying the effects. The cascade of complement proteins that occurs during an infection is
called complement activation. It may occur in three pathways that end in the activation of C3.
The Classical Pathway
The classical pathway was the first discovered. It is initiated when antibodies bind to antigens.
(1) Antibodies attach to antigens (for example, proteins or large polysaccharides on the surface
of a bacterium or other cell), forming antigen–antibody complexes. The antigen–antibody
complexes bind to and activate C1.
(2) Next, activated C1 activates C2 and C4 by splitting each of them. Then C2 splits into
fragments C2a and C2b, and C4 is split into C4a and C4b.
(3) C2a and C4b combine and together activate C3 by splitting it into C3a and C3b fragments.
C3a participates in inflammation, and C3b functions in cytolysis and opsonization.

The Alternative Pathway

The alternative pathway is so named because it was discovered after the classical pathway.
Unlike the classical pathway, it does not involve antibodies. The alternative pathway is activated
by contact between certain complement proteins and a pathogen.
(1) C3, constantly present in the blood, combines with complement proteins called factor B,
factor D, and factor P (properdin) on the microbe’s surface. The complement proteins are
attracted to microbial cell surface material (mostly lipid–carbohydrate complexes of certain
bacteria and fungi).
(2) Once the complement proteins combine and interact, C3 splits into fragments C3a and C3b.
As in the classical pathway, C3a participates in inflammation, and C3b functions in cytolysis
and opsonization.

The Lectin Pathway

The lectin pathway is the most recently discovered mechanism for complement activation.
When macrophages ingest bacteria, viruses, and other foreign matter by phagocytosis, they
release cytokines that stimulate the liver to produce lectins, proteins that bind to carbohydrates.
(1) Mannose-binding lectin (MBL) binds to the carbohydrate mannose. MBL binds to many
pathogens because MBL molecules recognize a distinctive pattern of carbohydrates that
includes mannose, which is found in bacterial cell walls and on some viruses.
(2) As a result of binding, MBL functions as an opsonin to enhance phagocytosis and activates
C2 and C4.
(3) C2a and C4b activate C3. As with the other two b 5 6 7 8 9 mechanisms, C3 splits into
fragments C3a, which participates in inflammation; and C3b, which functions in cytolysis
and opsonization.