1. Immunoglobulins, immune response
Martin Liška

2. 1. The structure of immunoglobulins

4. 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

5. 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)

6. 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 specificity is determined by amino acid sequences and both by morphology and shape of the loop

7. 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

8. IgA found both in serum and seromucinous secretions defense of mucosa
opsonization
does not activate complement

9. 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

10. the best classical way complement activator
does not bind phagocytes Fc receptor, but substantially enhances phagocytosis through complement activation
biological half life 6 days

11. IgD free form in serum, bound on B cells membrane
antigen receptor on B cells

12. IgE in normal conditions low amounts in serum
mainly bound on mast cells (binds through FceR)
anti-helminth defense
immediate type allergic reactions

13. 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

14. 6. Allotypic and idiotypic variations
allotypes = allelic variants of isotypes
idiotypes = structural determinants localized in variable region having connection with the ability of antigen binding
idiotopes = epitopes in variable region (idiotype is the sum of idiotopes)
anti-idiotypic antibodies = in principle reflect the antigen

15. 7. Genetic basis of Ig production
a/ L chains genes
K chain – genes located on chromosome 2
- V, J and C segments
chain – encoded in similar complex of
genes on chromosome 22

17. 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

18. 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

19. 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 change in antigen-binding specificity !
(no alteration in the L chain or in the variable portion of H chain)

20. 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

21. 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 specificity and in k- or l-chain isotype

22. Clonal expansion
proliferation of lymphocytes activated by reaction with an antigen
all lymphocytes of generated clone have the identical antigenic specificity

23. 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 specificity (= produced Ig have the same antigenic specificity) with tumor cell (= cells are „immortal“)

24. The utilization of monoclonal antibodies:
Diagnostics (flow cytometry, ELISA, autoantibodies etc.)
Treatment (anti-IgE, anti-TNF-a, anti-CD3)

25. 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

26. 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)

27. Phases of humoral immune responses

28. 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; typicky polysaccharides, lipids

29. 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

31. 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

32. Ontogenesis of immune response
a/ prenatal

33. 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

34. 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

35. B lymphocytes Precursors form 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

36. Monocytes-macrophages
Macrophages can be detected by week 3-4
Mature monocytes appear by month 5 in fetal circulation

37. Neutrophils
Mature cells are detectable from week 12-14

38. 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 yr.
Humoral response to polysaccharide antigen arises by the age of 2 yr.

39. 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

40. 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)

41. 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