Transcript Slide 1

Sri Ram .P
Date: 11/25/03
Course: Scientific Discovery
Instructor: Dr. A. Vankley
Cell Mediated Immunity
“Discoveries concerning the specificities of
Cell Mediated Immune defenses and their
implications ”
Immunity?
Foreign Invaders
Self Markers
Markers of Non-Self
Organs and tissues of the
immune system

Lymphatic vessels
form a circulatory
system that
operates in close
partnership with
blood circulation.
Lymph Node
Cells of the Immune System

B cells become
plasma cells, which
produce antibodies
when a foreign
antigen triggers the
immune response.
Antibody

Antibodies
produced by cells of
the immune system
recognize foreign
antigens and mark
them for
destruction.

Activation of B cells to
make antibody

T lymphocytes
become CD4+ or
helper T cells, or
they can become
CD8+ cells, which in
turn can become
killer T cells, also
called cytotoxic T
cells.

Activation of helper T
cells

Activation of cytotoxic
T cells
Cytokines
Complement
Natural Killer cells , Phagocytes and
Granulocytes
asa
Immunity and Cancer
Human Tissue Typing for Organ
Transplants
Rolf M. Zinkernagel
Rolf M. Zinkernagel
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Born: January 6, 1944, Basel, Switzerland
Primary and Secondary Education in and around
Basel.
1962-68:University of Basel, Faculty of
Medicine
1969- Began his life as Surgeon in Basel but
soon realized this was not his field.
1969-70:Postdoctoral Fellow, Laboratory for
Electron Microscopy, Institute of Anatomy,
University of Basel
Rolf M. Zinkernagel

1971-1973 Postdoctoral Fellow, Institute of
Biochemistry, University of Lausanne,
Switzerland
 He learnt his immunology here.
 He also familiarized himself with the 51-Cr.
Release assay to study the immune mechanism
destruction of host cells.
 His work with infectious agents and immunity
studies motivated him for further study .
Rolf M. Zinkernagel
 1973-75:Visiting
Fellow, Department of
Microbiology, The John Curtin School of
Medical Research, Australian National
University, Canberra, Australia
 1976-79:Associate (Assistant Professor),
Department of Immunopathology,
Research Institute of Scripps Clinic, La
Jolla, California
Rolf M. Zinkernagel
 1979-88:Associate
Professor, Department
of Pathology, University of Zurich,
University Hospital, Zurich
 1988-92-Full professor in same place
 1992-Head, Institute of Experimental
Immunology, Zurich
Peter C. Doherty
Peter C. Doherty

Born: October 15, 1940, Australia
 1962:BVSc University of Queensland, Australia
 1966:MVSc University of Queensland, Australia
 1967-71:Scientific Officer, Senior Scientific
Officer, Department of Experimental Pathology,
Moredun Research Institute, Edinburgh,
Scotland
Peter C. Doherty

1972-75:Research Fellow, Department of
Microbiology, The John Curtin School
of Medical Research, Australian National
University, Canberra, Australia
 1975-82: Associate Professor/Professor, The
Wistar Institute, Philadelphia, PA
 1982-88:Professor and Head, Department of
Experimental Pathology, The John Curtin
School of Medical Research, Australian National
University, Canberra
Peter C. Doherty
 1988:Chairman,
Department of
Immunology, St. Jude Children's Research
Hospital,
Memphis, TN
 1992:Adjunct Professor, Departments of
Pathology and Pediatrics, University of
Tennessee, College of Medicine,
Memphis, TN
Paired Up
 Peter
Doherty first studied the
pathogenesis of Semliki Forest virus
infection in the mouse, then switched to
the lymphocytic choriomeningitis virus
(LCMV) model which was a much more
powerful tool for immunological analysis.
Paired Up
 Zinkernagel
wanted to work with R.
Blanden on cell-mediated immunity
against Salmonella and Listeria to learn
more about the role of cell-mediated
versus antibody-dependent immune
effector mechanisms in these infectious
disease models . But lack of space in the
lab paired both of them.
Background
In 1960-70s immunology was
attempted to be understood in terms of
infectious diseases. It was then Largely
pre-occupied with antibody and T-cell
responses against foreign protein
antigens or chemically defined small
molecules called haptens.
Background
 Mechanism
of foreign-organ graft rejection
was intensively studied, although the
biological function of MHC was largely
unclear.
 Only few people studied immunity against
infectious agents.
Background
 Antibacterial
and antiviral T-cell mediated
immunity and the capacity of immunized
cytotoxic CD8+ T cells to destroy either
virus infected or allogeneic target cells in
vitro- was the work in progress at JCSMR.
Techniques and Study
 Doherty
and Zinkernagel jointly begin
work on CMI in LCMV (Lymphocytic
choriomeningitis virus).
 51 cr. Release assays as cytotoxicity
assay was used by Zinkernagel.
 Doherty was efficient in cannulation and
could draw few ml of CSF from cisterna
magna of the mice.
Techniques and Study
 Whether
inflammatory cells in the CSF of
mice infected intra cerebrally with LCMV
were cytolytic in vitro and whether there
was any correlation between cytotoxic Tcell activity and severity of
choriomeningitis .
Observations
 Cytotoxic
T cells specifically destroying
LCMV infected target cells could be found
in CSF of normal mice but not in nude
mice lacking thymus and T-cells.
 T-cells probably also destroyed infected
meningeal and ependymal cells in vivo
and this was the pathogenic mechanism
causing choriomeningitis.
Observations
 The
findings were published in the
Journal of Experimental Medicine in March
1973.
 Same journal had a paper showing mice
with different major histocompatibility gene
complexes differed in susceptibility to
LCMV after cerebral infection.
 This prompted to experiment further on
this.
Experiment
 6-8
mice of inbred and cross-bred strains
were infected intra cerebrally with LCMV.
 2 of each were sacrificed on day 7 after
infection when first mouse became sick. –
to test antiviral cytotoxic T –cell activities
in spleens.
 Remaining mice were monitored for lethal
disease during next 10days .
Experiment
 All
mice died in course of time.
 But only some generated virus-specific
cytotoxic activity that was measurable in
vitro.
 Result- Either cytotoxic T-cells have
nothing to do with choriomeningitis or the
test was inadequate.
Reassessment
 Mouse
L-929 cells (fibroblast cell lines)
were used as target cells to assess
cytotoxic T-cell activity.
 Fortunately the mouse CBA strain and the
L-cells derived from mouse strain C3H
were closely related.
 Both possessed the same MHC-molecules
(H-2k).
Reassessment

Studying further, LCMV -immune spleen cells
from all mice that possessed H-2k haplotype (as
do CBA mice) including the cross breeds with H2k lysed L-929 cells infected with virus.
 But did not lyse uninfected targets or those
infected with third-party virus.
 All spleen cells derived from immunized mice
that were not of H-2k type failed to do so.
Further Studies
 Two
additional experiments showed that
LCMV immune lymphocytes from non-H2k
strains of mice were able to lyse LCMV
infected target cells of same MHC –type.
 LCMV did not infect these cell lines.
 Used macrophages from the peritoneum
of the mice as target cells. Adhered to
plastic, readily infectable and labeled with
Cr 51.
Further Studies
 Criss-cross
experiments showed that
LCMV immune T-cells from H-2b mice lyse
LCMV-infected macrophages of H-2b
origin but not those of other H-2 types and
vice versa.
 These findings were reported in December
to Nature and were published in April,1974
Similar Finding
 TNP-specific
cytotoxic T cells lysed
syngeneic TNP-lated targets more
efficiently than allogeneic TNP-lated
targets.
 European journal of immunology –same
time . But independent.
How to Interpret
 Biological
function of MHC and TA was
unknown in early 1970s.
 TA-Gorer and Snell
 HLA-Dausset and Van Rood
 Many patients were typed and disease
susceptibilities were linked to TA
 Mice MHC was mapped due to availability
of well-bred strains.
How to interpret
 MHC
polymorphism- to prevent mutual
parasitism or transmission of tumor cells
or to prevent viruses or pathogens
mimicking TA and eliminate the species.
 TA were though to act as enzymes or
generators of antibody diversity.
The best proposal
 H.S
Lawrence proposed infectious agents
complexed with TA and formed a (self+x)
complex.
 All
this was foundation to reveal the
essential role of MHC and T-cell
recognition.
Crucial findings
 Finding
of double specifity by Tcells– for
virus and MHC from intial experiements.
 Findings that H-2 incompatible T-helper
cells transfused to T-cell deficient nude
mice were not able to help mice nude B
cells make antibodies
 Histo-incompatible B cells and T cells were
not interacting successfully to produce a
good IgM to IgG switch.
Crucial findings and Analysis

Antigen specifc proliferative T-cell responses
found only when primed T cells and antigen
presenting cells were with same MHC type.
 All in vitro and in vivo tests confirmed the HLA
restriction for T cells.
 Virus infection somehow caused alterations of
TA on the cell surface by forming a complex of
viral antigen with MHC molecules .
 These alterations were recognized by the T-cell
receptors
Intimacy Model

Foreign TA altered forms of self-TA
 Lymphocytes and target cells interact mutually
via TA. H-2k interacts best with H-2k in a
symmetrical like-like complementarity.
 This initmacy model was soon excluded by the
F1-experiment showing virus specific cytotoxic T
lymphocytes from heterozygote F1 mice
consisted of at least 2 subpopulations-each
being specific for infected H-2k and other for H2b targets.
Codominant Expression
 Since
MHC expressed codominantly some
T-cell receptors of one population were
probably specific for H-2k plus virus and
other sub-population was specific for H-2b
plus virus.
 Working
with Blanden’s experiments
showed that H-2d and H-2k regions coding
for class1MHC molecule were involved in
virus-specific cytotoxic t-cell recognition.
 This seperated MHC restricted recognition
by virus-specific cytotoxic T-cells from
MHC class 2
Conclusions
In 2nd letter to the Nature -T-cells might function
to survey the integrity of TA. Recognition of cell
surface alteration due to virus infection ,chemical
modification or genetic differences may be
accommodated in the same model.
 General hypothesis formulated in Lancet was
that function of MHC is to signal modifications of
self-MHC to the immune system.

Conclusions
 Tried
to extend the explanations to helper
T cells-they might recognize antigeninduced modifications of 1a(as the MHC
class 2 molecules) on B cells and
macrophages.
 Explained the extensive polymorphisms of
MHC molecules minimizing failure of some
pathogens to cause immuno-modification
and risk general unresponsiveness.
Role of Peptides

Not know then , MHC molecules are recognized
as complex with antigenic peptide.
 By works of Unanue, Grey and others on class 2
antigens and Townsted works showed- class 1
molecules of the virus infected cells present
peptides ,9-10 amino acids long to virus specific
cytotoxic t cells. These peptides were later
successfully eluted
Role of Peptides

In 1987 x-ray crystallography revealed a peptide
binding cleft.
 X-ray structure of the complete complex of the T
cell receptor-MHC class1 plus the bound peptide
in same year of Nobel prize.
 Still unclear which part of the TCR and whether
always corresponding parts of the TCR
recognized the peptide and the MHC molecule
in the same general position.
Implications
 The
above findings and conclusions gave
immense scope for further understanding
of the immune system and its clinical
implications in the field of medicine.
Role of Thymus

Reconstitution of lethally radiated H-2b recipient
mice with bone-marrow stem cells of (H-2k*H2b)F-1 origin resulted in bone marrow chimeras
tolerant to H-2k and H-2b .
 When immunized these reacted against only H2b+minor HC antigens or H-2b+virus only.
 So, MHC restricted T cells were specifically
selected during T cell maturation according to
the MHC expressed in the thymus.
Role of Thymus
 MHC
specificity studied in mice that lacked
thymus did not have mature t-cells.
 Introducing H-2k thymus into F-1 mice
gave T–cells recognizing virus infected H2k but not infected H-2b target cells.
implications – It is not only
necessary to deplete T cells to avoid lethal
graft versus host disease.
 But also, host, transplanted bone marrow
and hosts own or transplanted thymus
grafts must share MHC molecules.
 Other wise no proper immune reaction can
be mounted in such reconstituted hosts.
 Clinical
New Vaccines
As peptides from viruses, bacteria and
parasites are presented to the MHC class
1 or 2 molecules it was suggested that
instead of live potentially harmful peptides
could possibly be used as vaccines to
induce T- cell responses.
New vaccines
 Peptide
life was short lived therefore
adjuvants are required to guarantee slow
and long term release of the peptides
triggering T cells over a prolonged period.
Vaccinations
 Positive
vaccination- increase the T cell
precursor frequency to enhance
protection.
 Negative vaccination- to reduce or delete
Tcells by excess peptide . To exhaust or
delete immuno patholgical disease
causing T-cells. Tried in diabetes.
Mutant viruses
T
cell epitope escapes mutant viruses Mutation of the 9-10 a.a peptides such that
its presentation by MHC molecules or
recognition by t-cells is no longer possible.
this helps viruses escape immune
surveillance
Mutant Viruses
 Infected
mice in the footpad gave 2 peaks
of immune reactions.
 Similar mutant virus are seen in HIV and
HBV infections.
MHC Associations
 Linkage
between some disease
susceptibilities and certain HLA-types –
important role of MHC molecules in
immunity.
 These autoimmune or immunopathological
diseases are linked often to HLA class1
rather than class 2 molecules.
MHC Associations
 As
antigenicity and immunogenicity are
linked to MHC and correlate with different
strengths of the T cell response, shows
that different MHC molecules directly
determine and regulate resistance to
diseases.
HLA associated diseasesAnkylosing Spondylitis
Autoimmune Diseases
 Non
cytopathologial viruses are not
directly responsible for disease by
themselves ,instead by the damaging
effect of the protective T –cell responses.
 Differences in MHC may influence the
severity of the disease depending on the
immunopathological response of T-cells.
 These are the basis for autoimmune or
immunopathological diseases.
Autoimmune Diseases
 Multiple
sclerosis
 Diabetes mellitus
 Rheumatoid Arthritis
 SLE
 Psoriasis
 Hashimotos thyroiditis,etc
Autoimmune Diseases
Immunological memory
 The
concept of immunological memory
has been exploited in vaccinations.
 The B and T cells were found to exist as
memory cells and hence maintained
certain precursor levels to fight infections
better.
 The use of immune memory were
explained for the mother and fetus.
Other Related Diseases
 1-
Immune complex diseases. Ex.
Glomerulonephritis
 2-Immune Deficiency diseases. Ex. AIDS
Team Work
Thus the combined efforts of doctors,
immunologists, geneticists, virologists,
microbiologists, etc helped us reveal the
recognition of viral infected cells by T cells.
Giving insight into immunological specificity
and memory and help understand
immunological disease pathogenesis.

In recognition of these
contributions Peter
Doherty and Rolf
Zinkernagel were
awarded the Nobel
Peace Prize in
Medicine in year
1996.
References
1. Zinkernagel RM, Doherty PC. Restriction of in vitro T cell-mediated
cytotoxicity in lymphocytic choriomeningitis within a syngenic and
semiallogeneic system. Nature 248, 701- 702, 1974.
2. Zinkernagel RM, Doherty PC. Immunological surveillance against
altered self components by sensitised T lymphocytes in lymphocytic
choriomeningitis. Nature 251, 547-548, 1974.
3. Doherty PC, Zinkernagel RM. A biological role for the major
histocompatibility antigens. Lancet, 1406-1409, 1975.
4. Zinkernagel RM, Doherty PC. MHC restricted cytotoxic T cells:
Studies on the biological role of polymorphic major transplantation
antigens determining T cell restriction specificity. Advances in
Immunology 27, 51-177, 1979.
References
 5-Press
Release: The 1996 Nobel Prize in
Physiology or Medicine
 6-Nobel Lecture, December 8, 1996
Cellular Immune Recognition and the
Biological Role of Major
Transplantation Antigens
 7-Nobel Lecture, December 8, 1996 Cell
Mediated Immunity in Virus Infections
References
 8-www.nobel.se
 9http://press2.nci.nih.gov/sciencebehind/i
mmune/immune17.htm
 10www.niaid.nih.gov/publications/autoimm
une/graphics/Synovium.gif