Curs 1

Prof.
Ileana Constantinescu MD, PhD

Immunology
Fundeni Clinical Institute

ileana.constantinescu@imunogenetica.ro
www.imunogenetica.ro

Course 1: Immunology, Prof. Ileana
Constantinescu
1
Course 1: Definition of general
aspects of immune response
Immunology
Innate immunity
Adaptive (Acquired) Immunity
Clonal selection
Humoral and Cellular Immunity
Lymphocyte migration into lymphoid tissues
Germinal centers
Follicular dendritic cells
Mucosal immune system

2 Course 1: Immunology, Prof. Ileana Constantinescu
Course 1: Immunology, Prof. Ileana Constantinescu 3
Immunology
Immunology: that branch of biomedical science concerned with the response of the
organism to antigenic chalenge, the recognition of self and not self, and all the biological
(in vivo), serological (in vitro) and physical chemical aspects of immune phenomena.
Immunology means the study of the structure and function of the immune system
(basic immunology); immunization, organ transplantation, blood banking and
immunopathology (clinical immunology); laboratory testing of cellular and humoral
immune function (laboratory immunology); and the use of antigen-antibody reactions in
other laboratory tests (serology and immunochemistry).
Immunological memory
The innate and adaptive immune systems
protect us from potentially infectious agents
(viruses, bacteria, etc.) that have gained
access to our body through the skin or the
lining of our internal organs. Such systems
have evolved to protect us from not only
intracellular (viruses, some smaller parasites,
some bacteria) and from extracellular (most
bacteria, large parasites) pathogens and
allergens (animal hair, pollen, chemicals) but
also from ourselves in potentially
uncontrolled growth such as malignancy or
autoimmune diseases.

INNATE IMMUNITY

Innate immunity is a nonspectific cellular and
humoral response that operates as the first line
of defense against pathogens. Extracellular
pathogens are immediately taken up and
degraded by neutrophils and mononuclear
phagocytes. Large parasites are killed by
eosinophils. Natural killer lymphocytes (MK) kill
some tumor and virally infected cells. Preexisting
soluble mediators of innate immunity are natural
antibodies and complement components, which
can also attach to the cell membranes of many
pathogens.

The innate and adaptive systems
Innate immunity also includes a cellular component. Pathogen
destruction in the phagolysosome.
- PRR pattern recognition receptors
The main characteristics and components of innate immunity occur in the inflammatory response.
If an organism enters the body by breaking through the barriers of the skin or mucosa lining the
inner surfaces of the body, it comes in contact with phagocytic (neutrophils, eosinophils,
monocytes/macrophages) and/or natural killer cells as well as the complement system.
Complement consists of over 20 proteins, all of which interact in a cascade fashion to attract
phagocytic cells (chemotaxis), coat bacteria (opsonize) so that phagocytic cells recognize and then
engulf them, and interact with antibody to lyse the cell membrane of the invading organism. Thus,
the main mediators of innate immunity are lysosomal enzymes and interferons as well as other
cytokines, complement proteins, and acute phase proteins (CRP MBL).
It should be noted that innate host defenses are found in all multicellular organisms, most of which
do not have an adaptive immune system. However, in multicellular organisms with an adaptive
system, many of the mediators of innate immunity can recruit adaptive immune cells.
In order to handle certain pathogens, multicellular organisms have developed a group of largely
circulating cells with very specific receptors for immunogenic peptide fragments of both
intracellular and extracellular pathogens. When these receptors are activated, the cells can secrete
various soluble mediators that attack pathogens directly (antibodies, perforin granules, etc.) or
stimulate other cells (cytokines). Antigen-presenting cells (APCs) either process or present already
processed antigenic peptides to surface immunoglobin receptors on B cells and immunoglobin-like
receptors of T cells. While surface immunoglobin on B cells can directly respond to an immunogenic
piece (epitope) of a pathogen, in order to develop into mature plasma cells, they usually require
some help in the form of cytokines from T cells.
Thus, T cells are central to an adaptive immune response. Helper T (Th) cells not only help B cells
become plasma cells, they can also help cytotoxic T cells develop as well as increase macrophage
activity. However, in contrast to B cells, the receptors on the surface of T cells can only recognize
processed antigenic epitopes presented to them in the context of surface MHC. The ability of T
cells to see only foreign antigen in the context of self MHC is called MHC restriction and is a
critical safeguard for the adaptive immune system to only eliminate pathogens and not normal
cells.

Interactions between the innate and adaptive immune systems
Th cells central to adaptive immune response
B cells develop with their appropriate
receptors for a cognate antigen's epitope T
and without ever seeing that antigen before,
once encountering that antigen and
generating a full immune response to it, they
will remember such a first encounter.
Thus, when these memory T and B cells
encounter that same antigen again their
response to that antigen is much faster and
more robust.

Activation of phagocytic cells through binding of pattern recognition receptors (PRRs). Binding of PRRs to pathogen
associated molecular patterns (PAMPs) on microbial cells stimulate phagocytes to become activated, increasing their size,
phagocytic activity, production of antimicrobial substances, and production of cytokines and chemokines to attract and activate
other components of the immune system.
Course 1: Immunology, Prof. Ileana Constantinescu
Natural killer (NK) cells. (A) NK cells use a variety of receptors to identify target cells to be killed. These include Fc
receptors, complement receptors, and receptors that assess the MHC I molecules present on target cells.
In addition, NK cells can influence the development of T cells in the initiation of adaptive immune responses.
(B) NK cells utilize killer activation receptors (KARs) to recognize cells undergoing stress. Once engaged, KARs
activate the NK cell to kill the target. Killer inhibitory receptors (KIRs) "examine" the target for sufficient levels of
"self" MHC I molecules. If KIRs are sufficiently engaged, the killing program is terminated. If not, killing of the
target cell proceeds.
Course 1: Immunology, Prof. Ileana Constantinescu
CLONAL SELECTION

When antigen binds to either surface
immunoglobulin on B cells or its specific
receptor on T cells, these cells are induced to
proliferate rapidly. Thus, antigen selects and
generates specific clones that bind and
respond to it.
This process is called clonal selection and is an
extremely important part of the adaptive
immune response.
Clonal selection
HUMORAL AND CELLULAR IMMUNITY

Humoral immunity is mediated by the antibody
secreted by terminally differentiated B cells, the
plasma cells. Cell-mediated immunity involves T cells
that recognize antigen in an MHC-restricted fashion
and either secrete cytokines (Th cells) or become
cytotoxic cells (Tc cells), which kill virus-infected cells or
abnormal host cells.
It should be realized that in many cases a normal
humoral response will not proceed unless there is also
a companion Th cell response; this is because Th cells
secrete various cytokines that are necessary for full B
cell maturation and antibody class switching.

Introduction to the interactions and functions of the major components of the immune system
Overview of the process involving cell-mediated immunity and antibody-mediated immunity
Induction of cell mediated immunity and antibody against a viral infection
LYMPHOCYTE MIGRATION INTO LYMPHOID TISSUES
Lymphocyte migration into lymphoid tissues involves a family of adhesion
molecules called selectins, integrins, and members of the immunoglobin
superfamily.
Both naive and memory lymphocytes continue to recirculate between blood and
secondary lymphoid tissue. However, once activated, memory cells tend to
recirculate back to the tissue where they were activated. Lymphocytes respond to
chemotactic signals from inflamed tissues or secondary lymphoid organs initially
via surface glycoproteins called selectins. E-selectin (ELAM-1) on endothelial cells
binds L-selectin (LAM-1) on lymphocytes, causing a loose association or tethering.
Firm adhesion is mediated by lymphocyte integrin proteins such as VLA-4 or LFA-1
interacting with their respective ligand VCAM or ICAM (member of the
immunoglobin superfamily) on the surface of endothelial cells.
This adhesive interaction causes an activation of the lymphocyte to increase
expression of integrin molecules, thus strengthening the adhesion. The endolhelial
cell then directs the lymphocyte toward its basal lamina while simultaneously
down-regulating ICAM expression. Migrating lymphocytes finally diapedese
between endothelial cells into an organ or tissue by up-regulating production of
proteinases.
Once a naive lymphocyte is activated, it displays a different density of adhesion
molecules specific for the tissue in which it was initially activated, thus allowing it
to repeatedly return to that tissue.

22 Course 1 - Immunology - Prof. Dr. Ileana Constantinescu
GERMINAL CENTERS

Germinal centers (GCs) develop within primary follicles of lymph nodes and spleen during T cell-
dependent immune responses. GCs are the sites of centroblast clonal expansion and V
H
region-
directed somatic hypermutation, selection of high affinity antibody-producing centrocytes, deletion
of low affinity centrocytes, Th1- and Th2-directed antibody class isotype production and eventual
generation of plasmablasts or memory cells.
B cells in lymph nodes and spleen first encounter Ag in the T cell-rich areas of the paracortex and
periarterial lymphoid sheath (PALS), respectively. Ag is presented to them by dendritic cells (DCs),
which express high levels of class II major histocompatibility complex (MHC), adhesion, and co-
stimulatory molecules. After activation, and within 2 days, these B blasts produce low affinity
unmutated lg, which is capable of forming Ag-Ig complexes on follicular dendritic cells within
primary follicles. Many of these cells die within 10 to 12 clays. On the average, three B blasts
colonize each primary follicle and are then called centroblasts. Centroblasts, which are found in the
base or dark zone of the follicle, down-regulate their surface lg and undergo a clonal expansion
phase, dividing every 6 to 7 h. During proliferation, they activate a site-specific hypermutation
mechanism that introduces random point mutations into their lg V
H
region genes.
Centrocytes are the progeny of centroblasts that migrate apically into the basal light zone, where
they up-regulate their somatically mutated surface lg (receptors). Here, they interact with follicular
dendritic cells (FDCs), which can contain unprocessed Ag in the form of immune (Ag-Ab) complexes
on their surface for months. The centrocytes that express somatically mutated surface lg receptors
with high ( ) affinity for FDC-bound Ag up-regulate bcl-2 expression and are positively selected
(which is opposite to positive selection of thymocytes). These centrocytes interact with Th
2
cells
and can become either plasmablasts or memory B cells, depending on the co-stimulatory signals
they receive from FDC and Th1 or Th2 cells. However, they also have a high propensity to recycle
back into centroblasts for further proliferation and even higher affinity maturation. Centrocytes
with low affinity somatically mutated surface lg (receptors) for FDC-Ag do not up-regulate bcl-2 and
die via apoptosis. Thus, there is extensive fine tuning of centrocytes before they are allowed to
produce antibody for export.

Germinal centers (GC) zones
The recombination-activating genes RAG1 and RAG2, which drive genomic V(D)J rearrangements
are also induced to be reexpressed in preapoptotic centrocytes of the basal light zone. This
induction of an immature state in GC centrocytes may be necessary for further light chain receptor
editing in an effort to save these cells.
This editing of VL genes, along with the bcl-2-regulated apoptosis, provides an additional
mechanism for removing potentially autoreactive antibody-producing cells. Thus, in GC, there is an
opportunity for antigen-dependent secondary V(D)J rearrangements to occur in order to fine tune
the specificity of the peripheral antibody repertoire.
Once centrocytes with high affinity surface Ig and high expression of bcl-2 have passed the
discriminatory environment of the basal light zone, they migrate to the apical light zone. Here Ab
class switching and maturation into plasmablast or memory cells occur. Plasmablasts migrate into
the medullary cords of lymphnodes and red pulp cord of the spleen, where they terminally
differentiate intomature plasma cells and secrete their antibodies into the circulation .
Some activated B cells or plasmablasts can leave the secondary lymphoid organs, return to the
circulation, and reappear in the bone marrow and the gut. Memory B cells are found in the areas
surrounding GCs (follicular mantles) a well as the marginal zone of the spleen.
Initial proliferation of centroblasts gives rise to the first foci in primary folicles within 2 days, and
the development of a full dark and light zone GC take 14 days. The development of GCs requires
cluster formation between LFA-1 on B cells and ICAM-1 on FDCs as well as VLA-4 on activated B
cells and VCAP on FDCs.
In addition, interleukin-4, -5, and -10 must be elicited from Th1 or Th2 cells in order for antibody
class switching and full centrocyte development to plasmablasts or memory cells to occur. GCs in
spleen and lymph nodes peakat 2 weeks and begin to wane after 3 weeks; however, GCs remain
constitutively in the gut.

FOLLICULAR DENDRITIC CELLS

FDCs have long processes and Fc, C, TNF receptors, as well as ICAM-1, and VCAM-1
but no MHC-II. They contain Ag-Ab complexes on their surface for months to years.
Centrocytes that continue to maintain high levels of bcl-2 expression can receive
membrane-bound immune complexes (iccosomes) from FDC and induce the
expression of T cell-reactive surface molecules such as B7.2. This resultant
interaction determines whether there is differentiation into a mature GC
centrocyte or centroblast recycling. However, the formation of FDC clusters is
critical dependent on TNF and LT, since these cytokines are necessary for FDC
cluster formation and consequent primary as well as secondary follicles indicating
that they have TNF-R1s.
The fact that FDC can contain Ag for prolonged periods of time is the reason why
we do not need to be frequently immunized with certain antigens, since FDCs
provide a continuous depot of Ag even without an existing GC. It has recently been
recognized that the human immunodeficiency virus, upon disappearing from the
blood, can be sequestered on the dendritic arms of FDC for a number of years.
Eventually, the FDC can no longer hold the virus in check, the virus is released into
peripheral blood, and the patient enters the lethal stages of AIDS.

MUCOSAL IMMUNE SYSTEM

The mucosal immune system (MIS) consists of lymphoid tissues within and directly beneath the epithelial
lining of the respiratory, genitourinary, and gastrointestinal tracts as well as beneath the ductal system of
the salivary, lacrimal, and mammary glands. The surface area of mucosal surfaces is over 100 times greater
than that of skin, and the MIS contains up to 75% of all the B cells of the body. The primary product of the
MIS is IgA.
Within the gastrointestinal tract, soluble antigen is taken up by villus epithelium and particulate antigens are
primarily taken up in the ileal portion of the small intestine by specialized surface lining microfold (M) cells.
M cells internalize Ag and transport it to underlying lamina propria macrophages, which process it and
present it to surrounding collections of lymphoid cells forming ileal Peyer's patches (PPs).
PPs contain follicles, similar to lymph nodes and spleen, with high endothelial venules and significant
collections of T cells in between. T and B blasts activated here go to the nearest mesenteric lymph node for
further maturation, alpha H chain class switching, or J chain formation or deletion. They then enter the
thoracic duct and bloodstream, homing back to the same or distant mucosal sites. Thus, even though much
of the processing and reactivity to antigen occurs in the ileum, protection through IgA occurs at many
mucosal surfaces.
Serum IgA is monmeric and represents only 11% of all serum immunoglobulin. Secretory IgA represents over
95% of all Ig found in secretions and is primarily dimeric with two monomerlic units covalently joined by a J
chain. Dimeric IgA binds to a polymeric immunoglobulin receptor (pIGR) on the basal surface of mucosal
epithelial cells. This IgA-pIGR complex is endocytosed and transported to the apical (luminal) surface of the
epithelial cell.
During this transport process, a small piece of the pIGR is cleaved with the remaining component now called
the secretory component. Thus, IgA is secreted as dimeric IgA bound to a secretory component.
Secretory IgA does not activate the complement system but coats bacteria and some viruses (polio,
coxsackie, rota, and herpes), thus preventing their adherence to mucosal lining epithelium. Also, some
viruses within surface epithelia can be neutralized by pIGR-internalized IgA. However, it should be noted that
oral immunization with soluble Ag does not always generate an immune response and can instead generate
unresponsiveness.

27
Course 1 - Immunology - Prof. Dr. Ileana Constantinescu
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