POSSIBLE IMMUNOGENETIC BASIS FOR AUTISM

Roger A. Burger* and Reed P. WarrenUtah State University, Logan, Utah

Autism results from several different etiologies or combina-tion of pathological mechanisms. Mounting evidence indicates thatimmune dysfunction along with an environmental pathogen may befactors contributing to the development of some cases of autism.One of the immune deficiencies observed in autism is abnormalT-cell mediated immunity. Another is altered levels of certainclasses of antibodies (immunoglobulins), including decreased levels ofimmunoglobulin A and deficient complement activity, basedon the inheritance of a null allele of the C4B gene. In additionto the C4B gene, other genes on chromosome 6 also appear to beassociated with autism. In the developing child, genetically deter-mined immune deficiencies might increase the risk for autism in twoways: 1) A pathogen or its toxins might damage the brain, 2) thepathogen might trigger an autoimmune mechanism that wouldinterfere with brain functioning. In the mother, immune deficiencymight allow a pathogen to persist in utero, damaging the fetal braindirectly or triggering a maternal immune response that createspathogenesis in the fetal brain. r 1998 Wiley-Liss, Inc.MRDD Research Reviews 1998;4:137–141.

Key Words: autism; immune system

Immunologic disorders often have a genetic basis. The bestevidence for this in man comes from family studies, especiallystudies of twins. The incidence of a particular disease in

monozygotic and dizygotic twins is compared. If a disease showsa high concordance in all twins (both monozygotic or dizygoticpairs), this would point to shared genetic or environmentalfactors. This is because both monozygotic and dizygotic twinstend to be brought up in similarly shared environmentalconditions. If high concordance is restricted to monozygoticrather than dizygotic twins, however, then genetic factors arelikely to be more important than environmental factors. Studieswith twins have been undertaken for several human disorders inwhich autoimmunity plays an important role. These includeinsulin-dependent diabetes mellitus, rheumatoid arthritis, mul-tiple sclerosis, and systemic lupus erythematosus. In each case,around 20% of pairs of monozygotic twins show diseaseconcordance, compared with less than 5% of dizygotic twins.Thus, both inherited and environmental factors may play a rolein inducing autoimmune disease.

Folstein and Rutter [1977] found a higher concordancerate of autism in monozygotic twins than that seen in same-sexdizygotic twins. Also, other family studies reveal that about 3–7%of probands with autism have affected siblings [Bailey et al.,1996], a rate exceeding by 50 to 100 times that expected bychance. Thus, genetic factors are strongly implicated in thisdisorder. Notably, certain genes located on chromosome 6 areassociated with autoimmune disorders, and, as discussed below,appear to be associated with autism.

Most immunologic disorders, especially autoimmunedisorders, are associated with infections. More than 20 reportshave been published on prenatal and postnatal viral infections inautism [Gillberg and Coleman, 1992]. Other studies [Deykin andMacMahon, 1979; Stubbs, 1978] have implicated infection byother viruses with onset of autistic symptoms. Four studies foundan excess of births of children with autism in the month of March[Gillberg, 1990], thus providing circumstantial evidence for viralinfection in autism. It is conceivable that a pathogen involved inautism would be more operative or epidemic during the earlywinter (the second trimester for March babies).

THE IMMUNE SYSTEM AND IMMUNEABNORMALITIES IN AUTISM

The Immune SystemBlood cells (red cells, platelets, and white cells) arise in the

bone marrow and are derived from a common precursor cell, thehematopoietic stem cell. Upon stimulation by unknown factors,stem cells divide repeatedly. Some daughter cells develop intored blood cells, others into platelets, and still others into the twomajor types of white blood cells, the myeloid cells and thelymphocytes. The myeloid cells include the monocytes andgranulocytes. Monocytes mature into macrophages, which areone of two types of phagocytic immune cells. Macrophages,widely distributed in body tissues, play a critical role in natural(nonimmune) resistance to infections by engulfing and destroy-ing many pathogens. The macrophages also play a major role inimmune activation by presenting antigens (typically foreignsubstances not produced by one’s own system, but see below) tolymphocytes.

B cells and T cells are the two major types of lymphocytes.B cells become activated through exposure to an antigen by amacrophage. They are further stimulated by peptide messengerscalled lymphokines, which are produced by activated T cells.The B cells differentiate into plasma cells that secrete antibodies,including immunoglobulin A (IgA) (see Fig. 1). The moleculesof IgA are found in mucus-coated body surfaces such as therespiratory, digestive, and reproductive tracts, where theyneutralize infectious agents. Mother’s milk delivers them to themucus lining of her newborn’s gut. Quantitatively, IgA is themost important of immunoglobulins when both secretory andserum IgA are taken into account. Its rate of synthesis exceedsthat of all other immunoglobulins combined. Selective IgA

*Correspondence to: Roger A. Burger, Immunology, Utah State University,Logan, UT 84322–6895. E-mail: Roger@cpd2.usu.edu

MENTAL RETARDATION AND DEVELOPMENTAL DISABILITIESRESEARCH REVIEWS 4: 137–141 (1998)

r 1998 Wiley-Liss, Inc.

deficiency is the most common immuno-deficiency disorder in man [reviewed byAmmann, 1991], having a prevalenceestimated at 1:800. Although its cause isnot yet known, it may be related to anarrest in the development of B cells. AnIgA deficiency predisposes one to avariety of diseases, including allergies andautoimmune disorders.

There are two main types of Tcells. One is the cytotoxic T cell (alsocalled CD8 cell or ‘‘killer T cell’’), whichdestroys virally infected cells and tumorcells. The second type of T cell, the CD4cell, functions mainly as an activator ofother cells, including B cells and macro-phages. Both T and B cells have on theircell surface protein receptors that bind toantigens, and a mature lymphocyte has areceptor specific for only one particularantigen. The T cell antigen receptor(TCR) is very important. In order for Tcells to become activated, the receptormust not only bind to a foreign antigen,but also to an HLA (histocompatibility

locus antigen) molecule that is com-plexed (bound) to that antigen. ThisHLA-antigen complex is expressed onthe surface of an antigen-presenting cell(APC) such as a macrophage or B cell (seeFig. 2).

Immune System Abnormalities inAutism

Studies have documented numer-ous abnormalities [see Gillberg and Cole-man, 1992, for a recent review]: 1) Ourlaboratory has reported decreased serumIgA levels in 8 of 40 individuals withautism [Warren et al., 1997], a numberthat is far greater than might be expectedby chance. 2) Stubbs [1976] observed thatsome children failed to produce antibod-ies following rubella vaccination, suggest-ing that they lacked specific reactivitywith rubella. 3) Warren and colleagues[1986] found that individuals with autismhad decreased immune responses toT-cell mitogens [confirming an earlierfinding by Stubbs et al., 1977]. Also

reported are significantly reduced num-bers of CD41 T cells [Warren et al.,1986, 1990; Yonk et al., 1990], a findingconfirmed by two other laboratories[Wright et al., 1990; Denny et al., 1996].4) Children with autism have shownpositive reactions to skin tests with severalallergens and abnormally high antibodyresponses to tetanus and diphtheria anti-gens after booster injections [Tsaltas,1986]. 5) Warren and colleagues [1987]reported decreased activity of naturalkiller cells—a first line of defense againstviral infections and malignant cells. 6)Plioplys and colleagues [1994a] observedthat a majority of their participants withautism had increased numbers of DR1(activated) T cells, thus suggesting im-mune system activation. This finding wasconfirmed subsequently by Warren andcolleagues [1995]. 7) Individuals withautism have also shown aberrant cell-mediated and antibody responses tomyelin basic protein [Weizman et al.,1982; Singh et al., 1993] and neurofila-ment proteins [Plioplys et al., 1994b].

GENETIC FACTORS: THEMAJOR HISTOCOMPATIBILITYCOMPLEX OF GENES

BackgroundHLA molecules bind peptide frag-

ments derived from pathogens and dis-play them on the cell surface of APCs forrecognition by the appropriate T cells.The consequences of such presentationleads to destruction of virus-infectedcells, activation of macrophages to kill thebacteria that have been engulfed, andantibody production by B cells. Twodifferent characteristics of HLA mol-ecules make it difficult for pathogens toevade immune responses: 1) severaldifferent HLA class I and HLA class IIgenes encode proteins with differentranges of abilities to bind pathogenicpeptides; and 2) the HLA is highlypolymorphic, that is, there are multipleforms (alleles) of an HLA gene. This isanalogous to the polymorphisms seenwith the gene for eye color, in whichdifferent alleles lead to blue eyes, browneyes, and so on. In fact, the HLA genesare the most polymorphic genes knownin the human genome.

HLA covers a distance of morethan 2 centimorgans of DNA, or about4 3 10 basepairs, and contain more than100 genes. For simplicity, only thosegenes that have been studied in individu-als with autism will be included in thisreview. These include genes for the class I

Fig. 1. The production of IgA antibodies begins with the presentation of antigen by anAPC such as a monocyte. Both the B cell and T cell contact the APC and are activated bythe same specific antigen. The T cell produces lymphokines which induce a conversion ofthe activated B cell to a plasma cell. The plasma cell then secretes large numbers of IgAmolecules that can bind and neutralize the specific antigen.

138 MRDD RESEARCH REVIEWS • POSSIBLE IMMUNOGENETIC BASIS FOR AUTISM • BURGER & WARREN

molecules, called HLA-B, genes for twoHLA class II molecules, called HLA-DRand -DQ, and genes for three of thecomplement components. The particularcombination of HLA alleles found of thevarious HLA genes on an individualchromosome is known as an HLAhaplotype (Fig. 3).

The number of different HLAalleles affects antigen recognition indi-rectly by controlling peptide binding.The products of individual HLA genescan differ from one another by up to 20amino acids, making each allele quitedistinct. Most of these differences are onthe surface of the HLA molecule con-fined to the site that binds peptides(peptide-binding groove). Thus, theamino acids lining the peptide-bindinggroove determine the peptide-bindingproperties of the different HLA mol-ecules. The T cells responding to a givenprotein antigen presented by severaldifferent HLA molecules will usuallyhave to recognize different peptides. Inrare cases, a protein will have no peptidescapable of binding to any of the HLAmolecules expressed on the APCs of anindividual. When this happens, the indi-vidual fails to respond to the antigen.

The Role of HLA in AutoimmunityAn adaptive immune response di-

rected at self-antigens (those produced byone’s own body) is called an autoimmuneresponse or autoimmunity. The resultingautoimmune diseases are characterized bytissue damage. Well-known examplesinclude rheumatoid arthritis, which causesdestruction of synovial or joint-associatedtissue, and multiple sclerosis, whichinvolves damage to myelin tissue in thebrain. It is clear that autoimmunity isinitiated by responses involving T cells.Cytotoxic T-cell responses and inappro-priate activation of macrophages cancause extensive tissue damage, and inap-propriate T-cell help can initiate aharmful antibody response to self-antigens. Autoimmune responses are anatural consequence of both B-cell andT-cell receptors being able to recognizeany peptide, since such receptors will alsoinclude those reactive to self-antigens. Itis not known what triggers autoimmunitybut both environmental and geneticfactors, especially HLA genotype, areclearly important.

It was realized early in the study ofimmunity that the powerful effectormechanisms used to defend the individualfrom disease could, if turned against theindividual, lead to severe tissue damage.Typically, individuals do not mountsustained T-cell responses to their own

antigens and, although transient responsesto damaged tissues do occur, these rarelycause further tissue damage. However,while a lack of immune response to one’sown tissue is the general rule, sustainedimmune responses to self-tissues dooccur.

The association of HLA alleles withautoimmune disease is not surprising,since all autoimmune responses involve Tcells, and the ability of T cells to respondto a particular antigen depends on HLAgenotype. However, the way that HLAalleles determine susceptibility to autoim-mune disease is not known.

Methods for Establishing theAssociation Between the HLAGenotype and Disease

One determines such an associationby comparing the frequency of differentHLA alleles in clinical populations withthe frequency in the normal population.In insulin-dependent diabetes mellitus,for example, this approach has demon-

strated an association between diseasesusceptibility and the HLA class II allelesHLA-DR3 and HLA-DR4. Anotherway of determining whether HLA genesare important in autoimmune disease is tostudy the families of affected individuals.In family studies, it has been shown thattwo siblings affected with the sameautoimmune disease are far more likelythan expected to share the same HLAhaplotypes. However, HLA genotypealone does not determine whether aperson develops disease. Identical twins,sharing all of their genes, are far morelikely to develop the same autoimmunedisease than HLA-identical siblings, dem-onstrating that other genetic factors alsoaffect disease susceptibility. Recent stud-ies of the genetics of autoimmune diabe-tes in humans and mice have shown thatthere are several independently segregat-ing disease susceptibility loci in additionto the HLA.

There is evidence that several otherfamilies of genes may play an important

Fig. 2. Foreign substances (antigens) are engulfed and processed by APC and presentedto T cells by the major histocompatibility complex (MHC). The MHC-antigen complexinteracts with the T-cell receptor (TCR) initiating activation through a complex series ofreactions within the T cell. The hypervariability of the TCR allows for the recognition ofthousands of different antigens, giving the immune system the adaptability required toprotect the individual from infection.

Fig. 3. The human extended MHC on the short arm of chromosome 6 consisting of: theHLA-DQ region and the DR regions (including the DRb1 gene); C4B, C4A, Bf, C2; the genesfor TNF a and b, and the HLA-B region. Spacing of genes in the illustration approximatelycorresponds to the spacing of the genes on chromosome 6, except that the distancebetween DR and C4B and between the TNF genes and HLA-B are greater than shown.

MRDD RESEARCH REVIEWS • POSSIBLE IMMUNOGENETIC BASIS FOR AUTISM • BURGER & WARREN 139

role in increasing susceptibility to autoim-mune disease. In humans, normal plasmacontains nine (C1–C9) heat-labile pro-teins of the complement system. Theseproteins augment the antibacterial activ-ity of antibodies through a cascade ofinterdependent reactions called the classi-cal complement pathway. Inherited totalor partial deficiency of the early proteins(C4 or C2) of the classical pathway ofcomplement is very strongly associatedwith the development of systemic lupuserythematosus (SLE). These genes arelocated between the genes for HLA-Band HLA-DR (see Fig. 3). The mecha-nism of this association is unknown butmay involve the inability to clear certaininfections or the abnormal processing ofimmune complexes in the absence of afunctional classical pathway of comple-ment fixation.

Association of HLA Alleles WithAutism

Our laboratory reported that theallelic products of certain HLA genes areassociated with autism [Warren et al.,1991, 1992; Daniels et al., 1995] includ-ing the null allele of the C4B gene. A nullallele is a deficient gene from which noproduct is produced. An inherited C4Bnull allele would result in a deficiency ofC4B protein (a subset of the C4 comple-ment protein) and a breakdown in theclassical complement pathway. We foundthat the gene frequency of the C4B nullallele was significantly increased in indi-viduals with autism as compared to that ofunaffected individuals. Moreover, themothers of the individuals with autismhad an elevated C4B null gene frequency.The paternal C4B frequency was notsignificantly elevated.

Studying HLA extended haplo-types is another way of determiningwhether a given disorder is associatedwith HLA. Extended haplotypes areobserved when certain HLA haplotypessegregate together far more often thanexpected from the relative distancesbetween their respective genes. Someunknown mechanism prevents crossovers(gene exchanges between chromosomes)from occurring within this portion of theHLA, resulting in a similarity of DNAsequences [Alper et al., 1989; Degli-Esposti et al., 1992]. As Table 1 shows,about a dozen or so extended haplotypeswith a gene frequency of 0.0043 orgreater have been identified. One ormore of these extended haplotypes havebeen associated with specific disorders,most of which are known or suspected tobe autoimmune in nature [Degli-Espostiet al., 1992; Fraser et al., 1990].

Our laboratory has studied ex-tended haplotypes [Warren et al., 1992;Daniels et al., 1995] in autism. We havediscovered that one such haplotype,B44-S30-DR4, was increased more thansixfold. This particular haplotype consistsof the 44 allele of the HLA-B region, theS allele of the BF gene, the 3 allele ofC4A, the 0 or null allele of C4B, and theDR4 allele (see Fig. 3). Further, as Table1 shows the three extended haplotypescontaining DR7 occurred about twice asfrequently in individuals with autism as incontrols.

The relevance of HLA to pathoge-nicity and/or autoimmune mechanisms isnot fully understood. However, it isknown that HLA molecules are vital torecognition of foreign substances, such aspathogens, since T cells react to foreignantigens only as they are presented by

self-HLA molecules on APCs. Anothervery interesting finding was that some ofthe mothers of children with autism hadB44-S30-DR4 on one of their chromo-somes that did not pass to the child. Onits face, this finding would seem to argueagainst an association between B44-S30-DR4 and autism. However, if thisextended haplotype is associated with animmune abnormality/deficiency, its pres-ence in the mother may be sufficient toaffect the child. As noted earlier, maternalimmune deficiency itself may have al-lowed damage to the developing fetalbrain. Interestingly, in mothers withB44-S30-DR4 indications of autism werenoted very early, at birth or within thefirst year of life.

SUMMARYAutism likely results from several

different etiologies or a combination ofpathological mechanisms. As this briefreview suggests, evidence suggests thatsome cases of autism are associated withimmune abnormalities, pathogen-autoim-mune processes, and/or the HLA. Re-searchers in several centers in the UnitedStates and Europe continue to investigatethe link between the immune system andautism. Extensive pedigree analysis ofindividuals with autism and their rela-tives, further exploration of immuneabnormalities, and attempts to identifylinkages with autoimmunity may offer away of identifying families at increasedrisk for autism. j

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Table 1. List of HLA Extended Haplotypes and Their ReportedDisease Associations [Alper et al., 1989; Degli-Esposti et al., 1992]

Class I Class III Class II Known Disease

B BF C4A C4B DR DR_1 Abbreviation Associationsa

44 S 3 0 4 0401 B44-S30-DR4 RA (child onset), FS44 F 3 1 7 0701 B44-F31-DR7 CD, IgA Def57 S 6 1 7 0701 B57-S61-DR7 IgA Def, Psoriasis13 S 3 1 7 0701 B13-S31-DR7 None reported7 S 3 1 15 0501 B7-S31-DR15 MS, CD8 S 0 1 3 0301 B8-S01-DR3 GMG, IgA Def, SLE

18 F 3 0 3 0301 B18-F130-DR3 IDDM35 F 3,2b 0 1 0101 B35-F(3,2)0-DR1 HIV (rapid progression)35 S 3 0 1 0101 B35-S30-DR1 HIV (rapid progression)35 S 3 1 4 0404 B35-S31-DR4 None reported62 S 3 3 4 0401 B62-S33-DR4 IDDM, RA65 S 2 1 1 2 1 0101 B65-S2(1,2)-DR1 IgA Def

aRA, rheumatoid arthritis, and childhood onset; FS, Feltys syndrome; CD, Celiac disease; IgA Def, immunoglobulin A deficiency; MS,Multiple sclerosis; SLE, systemic lupus erythematosus; GMG, myasthenia gravis; IDDM, insulin-dependent diabetes mellitus; HIV, humanimmunodeficiency virus.bThis designation denotes a fairly rare duplication of the C4A gene; one allele is C4A3 and the other allele is C4A2.

140 MRDD RESEARCH REVIEWS • POSSIBLE IMMUNOGENETIC BASIS FOR AUTISM • BURGER & WARREN

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