IMMUNOLOGY TODAY

ow the im

efore the germ theory of dis-

ease was introduced by Louis

Pasteur in 1870 (Ref. 11, it was

not known that infectious dis-

eases are caused by germs - microorganisms

that are visible only under a microscope.

Pasteur’s correct theory revolutionized

medicine. The causes of about 20 major dis-

eases were identified between 1880 and

1900, owing to the availability of suitable

microscopes and stains for bacteria2. The

germ theory was applied to surgery by

Joseph Listerl, and thus surgeons began to

wash their hands before operations and ap-

ply asepsis. This freed surgery of inevitable

wound infection and allowed major opera-

tions to be performed successfully.

Furthermore, awareness of microbial dis-

ease led to the discovery of the immune sys-

tem, which provides defense against infec-

tious disease. Antibodies were discovered,

and used for passive immunization against

diphtheria and tetanus. Researchers noted

the power with which antibodies destroy

bacteria via complement-mediated lysis,

and wondered if the immune system ever

malfunctions and causes diseases by react-

ing with a host antigen instead of a micro-

organism. Unfortunately, World War I inter-

vened and, after that terrible disruption, it

was wrongly taught that Paul Ehrlich’s

dictum ‘horror autotoxicus’4 was based on

proof that autoimmunity never occurs.

Hence, euphemisms, such as ‘so-called

autoimmunity’ and ‘autoallergy’5, were

used. Ehrlich would have been horrified at

this mistake, which delayed recognition of

autoimmune disease by 40 year&‘.

Meanwhile, in New Zealand, research into

goitre by Charles Hercuss and Dick Purves9

led to successful abolition of the endemic by

the addition of one part of potassium iodide

to every 20000 parts of sodium chloride in

domestic salt. This research continued with

the study of the pit4tary gland by Walter

wncan s

The imzune system has evolmi to

defenci the host against itlfrctiozts

disease. Here, Dmcat~ Aclnms

outlines fhe hisforic developmet~ts

thnf hazle led to our crirrerzf

uizdeusfdiug of 11070 tile inzmfne

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mrtoinimune &senses nnd hc?iCl

these could be clrren or preuenfed.

Griesbach’O and the introduction of radio

active iodine by Purves for thyroid research.

While seeking the cause of Graves’ disease,

Duncan Adams and Purves developed a

bioassay for measuring levels of thyroid-

stimulating hormone (TSI-0, which is pro-

duced by the pituitary gland. This assay

demonstrated the presence of autoantibodies,

known as long-acting thyroid stimulator

(LATS), in the blood of patients with Graves’

disease; these autoantibodies react with the

TSH receptor of the thyroid gland to cause

excessive secretion of thyroid hormone and

hence thyrotoxicosis’. In 1957, Deborah

Doniach and Ivan Roitt” in the UK, and

Ernst Witebsky and Noel RoseI in the USA,

discovered autoantibodies to thyroglobulin

and the disease autoimmune thyroiditis.

Thus, the existence of autoimmune disease

was at last established.

Why do autoimmune diseases occur? To

understand this, we need to knoti how the

immune system works (Box 1). Niels Jerne13

received the Nobel Prize for showing that

antibodies are not formed on an antigen

template, but are preformed in myriad di-

versity, awaiting selection by a comple-

mentary antigen. Macfarlane Burnet14 be-

came aware that antibodies are produced by

lymphocytes and that each’ lymphocyte

makes only one type of antibody. He real-

ized that the unit of specificity in the im-

munity system is the lymphocyte clone, a

subset of lymphocytes with identical recep-

tors for antigen. Each clone comprises mil-

lions of lymphocytes and an individual has

millions of different clones, thus providing

the possibility of having a clone available

for reaction with any possible pathogenic

microbe that might infect humans.

But-net’s forbidden-clone theory As well as being trained in medicine, Burnet

was a bacteriologist, accustomed to grow-

ing bacteria on agar culture plates contain-

ing blood and counting their mutation rates

as they underwent millions nf cell divisions.

Bumet knew that lymphocytes multiply in

our bodies in response to antigenic stimu-

lation from microbial infection. He realized

that somatic mutations (DNA-copying errors)

must occur in population5 of multiplying

lymphocytes, just as in bacterial populations.

In fetuses, clones reactive with host antigens

are destroyed as they arise, so that we are all

born free of autoimmune disease. However,

as we grow older, we are all at risk of devel-

oping autoimmune diseases. Bumet proposed

that this is because random somatic mutations

in the variable (VI-region genes of multiply-

ing lymphocytes give rise to new clones.

Usually this is beneficial, tightening the

affinity of clones for pathogenic microbes

and so enabling us to recover from Infectious

disease. However, because of the random

element in the somatic mutations, some-

times a clone will arise that can react with

one of our self-antigens instead of a microbial

one. Burnet called such self-antigen-reactive

clones ‘forbidden clones’ and proposed that

they are the cause of the various autoimmune

diseases’5,‘h. It has since been shown that the

thyroid-stimulating autoantibodies in indi-

vidual patients with Graves’ disease contain

only one type of the two possible im-

munoglobulin light chains, K or A, but never

both”, thus demonstrating that the forbid-

den clones that make these pathogenic

autoantibodies arise from single lymphocytes

by somatic mutation. Hence, Burnet’s theory

he immune system

The purpose of the immune system is to defend the host from con- stantly changing microbial pathogens. Autoimmune diseases are caused by failure to distinguish between host and microbiaI anti- gens. The major components of the immune system are listed below. V genes Variable O-region genes mutate, and code for antigen receptors on lymphocytes. GermIine V genes provide the initial repertoire of lymphocyte clones, which is expanded by random V-gene changes in lymphocytes multiplying under antigenic stimulus. The random changes tighten affinity for pathogens, enabling recovery from infectious diseases, but also produce forbidden clones (see below). H genes Histocompatibility @I! genes are constant, and code for H anti-

gens on all nucleated cells. These antigens delete any nascent lymphocyte clones that have complementary antigen receptors.

They police the constantly changing immune repertoire to reduce risk of autoimmune disease. Apparent predisposition to auto- immune diseases is actually reduced protection.

Lymphocytes Lymphocytes bear antigen receptors and mediate immune speci- ficity as follows: @ B cells secrete antibodies for defense against infectious disease 0 Cytotoxic T lymphocytes k_iJl virus-infebed cells (virus factories) @ Helper T cells, when active, produce cytokines, that promote

immune response at infection sites and, when inactive, induce tolerance to host antigens centrally

Macrophages Macrophages collect soluble antigens for presentation to lymphocytes. Clones

The unit of immunological specificity is the lymphocyte clone. Each clone comprises millions of lymphocytes with identical antigen receptors. Forbidden clones Forbidden clones react with a host antigen rather than a microbial antigen. Most are harmless responses to tissue damage, but some cause autoimmune disease by: 0) stimulating a cell receptor (e.g. Graves’ disease); (2) blocking a cell receptor (e.g. myasthenia gtavis); (3) destroying specific cells (e.g. insulin-dependent diabetes mellitus, myxedema and multiple sclerosis); (4) complement-mediated cytolysis (e.g. Goodpasture’s syndrome and systemic lupus erythematosis); (5) perverting a functional molecule (e.g. C3 nephritic syndrome and lupus anticoagulant); or (6) unknown means (e.g. rheuma- toid arthritis). Police clones Police clones occur naturally and forestall or destroy forbidden clones. ArtifSally produced police clones could cure auto-

immune diseases.

has been proved correct for Graves’ disease. Futhermore, Burnet’s theory is probably true for all autoimmune diseases, including those caused by T-cell forbidden clones’“.

The H-gene theory Autoimmune diseases show familial aggre- gation, and the familial aggregation shows

disease specificity19,20. When V genes were discovered, they were proposed to be re- sponsible for the genetic predisposition to

autoimmune diseases as they could account for the disease specificity of the familial ag- gregationlR. However, although germline V genes have some influence on the risk of autoimmune diseaseIs, the histocompatibility

(H) genes [major, minor and H-Y (the male sex gene)] are involved to the greatest ex- tentIE. This is demonstrated by the altered risk of several autoimmune diseases im- posed by various major histocompatibility

complex (MHC) antigens?‘, minor H antigens

and H-Y (Refs 19,20). The H-gene theorylgv2” proposes that

germline V genes for the antigen receptors of B and T cells provide the initial repertoire

of immunological clones. The H genes,

which remain the same throughout life, im- mediately delete from the virgin repertoire any clones that are complementary to them, leaving the others to react with invading microbes. In the course of our recurrent sub- clinical and clinical infections, we con-

stantly add to our immune repertoires since somatic mutations and DNA rearrange- ments in the V genes of our lymphocytes result in the production of new clones with altered receptors. Those new clones whose

receptors have highest affinity for available antigens are selectively enlarged. Any new clone that reacts with an H antigen is dehted. Hence, an H antigen is never the target of autoimmunity, which shows that the H antigens impose on the immune repertoire perfect, unbreakable tolerance to themselves. This is nature’s device for mini- mizing autoimmune disease, while retain- ing maximal possible flexibility for finding

reactive clones for the constantly changing microbial pathogens that plague us”.

Functions of the MHC An important function of the MHC class I antigens is to put surface tags on ail nucleated

cells for defense against virus infection”. This is necessary because viruses replicate with explosive speed”. For example, a singIe influenza virus infecting a cell produces 1000 progeny in ten hours, each capable of infecting another cell and producing 1000

progeny in each; this would cause death in

two or three days. For~~ately, we are usually

saved by having large numbers of cytotoxic

T lymphocyte (CTL) clones that recognize

MHC class I antigens bearing viral peptides. Without this arrangement, the billions of free virus particles would ‘muffle’ the T-cell

receptors of the CTLs, and thus prevent

reaction with the virus particles on host cell

surfaces. Hence, large numbers of pre-

formed CTLs are required to kill the virus

factories before their products kill us3.

These clones are also responsible for the rejection of grafted kidneys and hearts,

through their reactivity with the unmodified

MHC antigens of the donor tissue’“.

The MHC class II ar.tigens are present

in myriad diversity on lymphocytes and macrophages. Their prime function is to

fine-tune the immune repertoire by deleting

all new clones that react with them’“.

As well as subtracting clones from the

1996

Box 2. A suggested course cyection for research towards immunotherapy and prophylaxis of autoimmune diseases

Ideas 1 N&e &at the autoimmune diseases are potentially curable by selective destruction of

the pathogenic ‘f~r&kkn’ (i.e. self-antigen-reactive) clones and are preventtible by judi-

cious va&nation against the triggering microbes.

2 Acknowledge the forbidden clone and histocompatibiity (H)gene theories.

3 Note that the H genes act predominantly by deleting nascent complementary clones,

not by altering specificity of antigen presentation16.

4 Note that B-cell clones reactive with host antigens occur commonly, often as passive re-

actions to tissue damage; they are not selected against by reproductive disadvantage.

Thus, it would be best to msetve the term forbidden clone for those B cells that cause disease.

5 Do not focus only on antigen-presenting cells and the major histocompatibiity complex

(MHC) restriction since cytotoxic T lymphocytes (CTLs) react with anything that fits,

including unmodified MHC antigens of other individuals, as shown by graft rejectton”.

6 Do not confuse tolerance and the activity of ‘police’ clones (see Box 1)‘s.

7 Note that nonspecific T-cell help probably suffices for the activity of B-cell forbidden

clones that cause autoantibody-mediated diseases.

8 Assume that sex-related differences of possible autoimmune diseases are based on

H-Y-antigen-mediated tolerances imposed on the immune repertoire.

Actions 1 Seek the forbidden clones causing suspected autoimmune disease by testing patients’

inummoglobulins or lymphocytes for reactivity with the likely autoantigens.

2 Use family studies and simple sequence repeat DNA markers to find the variable (V)

and major and minor H genes.predisposing to the various autoimmune diseases, then

determine sequences to gain specificity information.

3 Try to make artificial police clones - for instance, for curing Graves’ disease, where the

autoantigen has been clonedz4 and the B-cell forbidden clones produce readily measur-

able autoantibodies that are agonist competitors with thyroid-stimulating hormone,

Knowledge of the amino acid sequences of the antigen receptors on the forbidden clones

(which will vary somewhat from patient to patient because of the random element in

their generation) should enable creation of therapeutic, anti-idiotypic, CTL police clones

that lack attackable MHC antigens.

4 Seek the microbiil triggers of the autoimmune diseases and &move them with vac-

c& that lack the host-crossreactive antigens, as is being $oneered for rheumatic feve9.

repertoire, new clones are added: because of

competition of clones for microbial anti-

genie stimuli, deletion of a high-affinity

clone for a microbial antigen will cause

increased development of lower-affinity

clones for related microbial antigens. In this

way, tolerance imposed by, for example, the

antigen HLA-DW could increase the risk of

insulin-dependent diabetes mellitus (IDDM)

by fostering development of the forbidden

clones that cause R-cell destruction.

Accord with Darwin’s law of evolution Natural selection of reproductive advan-

tage, in accord with Charles Darwin’s law

of evolution, can be seen to act on the im-

mune system at two levels. First, on muiti-

plying lymphocytes for selection of higher-

affinity clones for reaction with pathogenic

microbes. Second, on multiplying people

for selection of the germline H and V genes

that are most able to provide maximal de-

fense against infectious disease coupled

with minimal risk of autoimmune disease.

Concluding remarks What has history taught us as to how we

should progress in the search for efficacious

strategies for treatment or prevention of auto-

immune diseases? Box 2 summarizes a sug-

gested sequence of ideas and actions that

stem from a consideration of the underlying

principles.

The author is indebted to D. Stewart, R. Irvine

and the Vivian Tod Bequest Fund.

Duncan Adarns (duncanQofago.ac.rIzl is af the

Faculfy of Medicine at the University of Otago

Medical School, Box 913, Dunedin, New Zealand.

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