Immunoglobulins, immune response Martin Liška. 1. The structure of immunoglobulins

Immunoglobulins, immune response

Martin Liška

1. The structure of immunoglobulins

2. Isotypes

• (in principle) classes of antibodies distinguished on the basis of H chain structure differences

• 5 types: m (IgM), d (IgD), g (IgG), a (IgA) and e (IgE)

• in addition, we can distinguish subtypes of antibodies within some classes (IgG, IgA) based on their H chain differences

3. Domains and their biological function

• in principle: domains of V regions form a recognizing unit and domains of C regions determine secondary biological functions of antibody (i.e. biological half life, distribution in the body, binding complement, binding to cells through Fc-receptor)

4. Variable region of Ig molecule

• hypervariable loops are concentrated at the spikes of variable regions where antigen binding sites are localized

• the binding site specifity is determined by amino acid sequences and both by morphology and shape of the loop

5. The biological features of distinct Ig classes

IgG• the most abundant serum Ig • the most important Ig of secondary immune

response • the only Ig which passes through the placenta• the main opsonizing Ig• activates complement via classical pathway• biological half life 21 day

IgA

• found both in serum and seromucinous secretions

• defense of mucosa• opsonization • does not activate complement

IgM

• in pentamer form is found in serum; in monomer form is bound on membrane of B cells

• prevailing antibody of primary immune response

• high-effective agglutinant and cytolytic agent• usually isohaemagglutinins and natural

antibodies

• the best classical way complement activator

• does not bind phagocytes Fc receptor, but substantially enhances phagocytosis through complement activation

• biological half life 6 days

IgD

• free form in serum, bound on B cells membrane

• antigen receptor on B cells

IgE

• in normal conditions low amounts in serum• mainly bound on mast cells (binds through

FceR)• anti-helminth defense• immediate type allergic reactions

Ig subclasses

• differences in H chain structure and biological properties

• IgG1 a IgG3 participate in defence against viral and bacterial protein antigens

• IgG2 ensures defence against antigens which does not require help from T lymphocytes

6. Allotypic and idiotypic variations

• allotypes = allelic variants of isotypes• idiotypes = structural determinants localized in

variable region connected with the ability of antigen binding

• idiotopes = unique set of antigenic determinants (epitopes) of variable portion of an antibody (idiotype is the sum of idiotopes)

• anti-idiotypic antibodies = directed against idiotypes, in principle reflect an antigen

7. Genetic basis of Ig production

a/ L chains genes

K chain – genes located on chromosome 2 - V, J and C segments l chain – encoded in similar complex of

genes on chromosome 22

b/ genes encoding H chain

• more complicated• localized on chromosome 14• V, D, J, C segments (genes encoding

individual segments contain more regions compared with L chains)

• during completion of V/D/J exon, gene rearrangement occurs

Development of B lymphocytes• Lymphoid progenitor → pro-B cells• During maturation from pro-B cells into pre-B cells:

Ig genes of the heavy chain recombine; pre-B cells express pre-BCR

• During maturation from pre-B cells into B cells: Ig genes of the light chain recombine

• Immature B cells express membrane IgM• Mature B cells express membrane IgM and IgD =

BCR and are able to respond to antigen in peripheral lymphoid tissues

Mechanisms contributing to antibody diversity:

• chance recombinations• imprecise joining of V, D, J genes• N-region additions• extensive mutations involving variable-

region genes after antigen exposure

Isotype switching

• during the immune response, plasma cells switch from producing IgM to IgG or to another Ig class (IgA, IgE)

• the switch involves a change in the H-chain constant domains (CH)

• no changes in antigen-binding specifity ! (no alteration in the L chain or in the

variable portion of H chain)

Allelic exclusion

• once the process of rearrangement on one of chromosomes is successful, then all attempts on second chromosome are stopped

• the same rule governs both for H- and L-chains• every single B cell produces only one type of H-

and one type of L-chain

Clonal restriction

• each B cell expresses identical copies of an antibody that is specific for single epitope

• when a B cell divides, the chromosomes in its progeny cells bear the selected allelic genes, and these genes do not undergo any further V/J or V/D/J rearrangements

• immunoglobulins produced by given B cell and its progeny are identical in epitope specifity and in -k or -l chain isotype

Clonal expansion

• proliferation of lymphocytes activated by reaction with an antigen

• all lymphocytes of generated clone have the identical antigenic specifity

Monoclonal antibodies

• immunoglobulins arising from a single clone of B cells, or more precisely cells artificially created by hybridisation of B lymphocytes of specific antigenic specifity (= produced Ig have the same antigenic specifity) with tumor cell (= cells are „immortal“)

The utilization of monoclonal antibodies:

• Diagnostics (flow cytometry, ELISA, autoantibodies etc.)

• Treatment (anti-IgE, anti-TNF-a, anti-CD3)

Humoral immune response

• The recognition of antigen by specific Ig on the surface of naive B lymphocyte

• The binding of antigen cross-links Ig receptors of specific B cells and then activation signals are delivered inside the B cell; the necessary second signal is provided by a breakdown product of the complement protein C3

• Clonal expansion of B cell and secretion of low levels of IgM

Humoral immune response• Protein antigens activate antigen-specific T helper cells which

stimulate B cell; antigen presentation of these antigens to T helper cells is required

• T helper cells exprime CD40L on their surface and secrete cytokines → proliferation and differentiation of antigen-specific B cells, isotype switching

• Affinity maturation = affinity of antibodies for protein antigens increases with prolonged or repeated exposure to the antigens (B cells migrate into follicles and form germinal centers → proliferate rapidly and their Ig V genes undergo extensive somatic mutations; at the same time, the antigen complexed with secreted antibody is displayed by FDC → B cells that recognize the antigen with high affinity are selected to survive)

Phases of humoral immune responses

Primary immune response

• First antigen exposure• The amounts of antibody produced is smaller

2 types of antigens:• T-dependent – help from antigen-specific T

helper cells is required; protein antigens• T-independent – antibody production is

induced directly, without the involvement of T helper cells; typically polysaccharides, lipids

Secondary immune response

• Subsequent antigen exposure• Higher amount of antibodies is produced• With protein antigens, secondary responses

show increased isotype switching and affinity maturation (= production of antibodies with increased affinity to antigen)

• Memory cells involvement

Affinity and avidity of antibodies

• affinity = the strength of the binding between a single binding site of a molecule (e.g.antibody) and a ligand

• avidity = expresses the strength of interaction of polyvalent antibody with a polyvalent antigen

Effector functions of antibodies

• Neutralization of microbes and their toxins• Opsonization of microbes (binding to phagocytes

through Fc-receptors, stimulation of their microbicidal activity)

• ADCC (Antibody-dependent cell-mediated cytotoxicity) – microbe, which is opsonized by IgG, is killed by NK-cell after binding of immunocomplex to Fc-receptor

• Complement system activation (classical pathway)

Ontogenesis of immune response

a/ prenatal

Hematopoiesis

• Mesoblast – from 2nd (3rd) week of gestation

• Liver – from 6th (8th) week of gestation, in liver hematopoiesis persists whole prenatal period

• Bone marrow – from 10th (12th) week of g., from 20th week the main organ of hematopoiesis

T lymphocytes

• Precursors from week 7, from week 8-9 lymphocytes move into thyme, where they differentiate

• TCR gene segments rearrangement, expression of TCR on the surface of T lymphocytes

• Selection

B lymphocytes

• Precursors from day 8• Fetal B lymphocytes express IgM on their

surface• Synthesis of specific antibodies start at

week 20-24, but IgA+M levels are in fact undetectable, IgG production starts after birth

Monocytes-macrophages

• Macrophages can be detected by week 3-4

• Mature monocytes appear by month 5 in fetal circulation

Neutrophils

• Mature cells are detectable from week 12-14

Postnatal

B lymphocytes • relative counts decrease after the birth• respond to immunization presumably by IgM

production, switching to other isotypes is slower• slow increase of child’s own IgG connected with

decrease of maternal IgG levels (by month 3-6)• IgM reaches levels common i adults at the age of

1-3 yr., IgG+A between the age of 10-15 yr. • Humoral response to polysaccharide antigen arises

by the age of 2 yr.

T lymphocytes

• More than 90% are naive, but their numbers decrease in adult age

• Proliferation under mitogen stimulation similar to adults X response to specific antigens only after contact with them

• Lower cytotoxic activity of T lymphocytes

Innate imunity

• Newborns´phagocytes have generally decreased functional ability, activity of NK-cells is decreased

• Decreased total complement activity (concentration of its compounds is of 35-70% of adults)

c/ Old age

• decreased cytotoxicity of NK-cells and macrophages

• decreased resistance against viral infections, decreased anti-tumour immunity

• switching from Th1 to Th2• weaker humoral response under new stimuli• increased production of autoantibodies