Objectives of the Presentation

 What are complex diseases?

 Canine MHC associations with auto-immune diseases

 Identification of other genes associated with auto-immune diseases

 How should breeders use this information?

 Will there be genetic tests for these diseases?

Overview of the Issue

Auto-immune diseases are complex diseases. These are diseases that occur as a result of the influence and interaction of multiple genes. However, the critical feature of these diseases is that they only occur after exposure to an environmental trigger.

So whether or not someone (or some dog) will develop the disease, depends on the particular combination of variants of the risk genes that they have, plus exposure to the environmental trigger.

Can we identify any of the gene variants that cause complex diseases?

Can we use that information to reduce the incidence of those diseases in dog breeds?

Could there be a genetic test for complex diseases?

Hypothesis

Many canine auto-immune diseases have been shown to have MHC associations.

Can we use this information to reduce the disease incidences?

Should we do so?

Additional Details

Auto-immune diseases are complex diseases. These are diseases that occur as a result of the influence/interaction of multiple genes, together with an environmental trigger. As yet, most environmental triggers have not been identified.

Some genes will have a greater influence than others. Variants at some genes will affect susceptibility to the disease, while different variants of the same gene, or variants of other genes, may be protective. Some variants may affect the severity of the disease. Each gene will have one variant that does not confer any increased or decreased risk of disease. This is known as the normal or wild type.

The common feature of auto-immune diseases is that the body starts to attack a particular organ or tissue, because it has lost the ability to distinguish "self" from "non-self" for that area. Thus, in hypothyroidism, the body starts destroying the thyroid gland, preventing the production of thyroxin. Lack of thyroxin causes the symptoms of the disease. Fortuitously, oral supplements of thyroxin can counteract these effects.

Within the genome there is a group of genes called the Major Histocompatibility Complex (MHC). These genes are central to the way in which an animal can distinguish self and non-self. It is therefore reasonable to suppose that these genes may have an influence on resistance and susceptibility to auto-immune diseases. In the dog the MHC genes are known as DLA (for Dog Leukocyte Antigen). We study three MHC genes, called DLA-DRB1, DQA1 and DQB1. These genes lie close together on chromosome 12, and are inherited in sets, which are called haplotypes. Everyone (and every dog) inherits one set (or haplotype) from each of their parents.

Many studies have shown clear associations of canine auto-immune diseases with DLA. These include diabetes (Catchpole et al. 2005; Kennedy et al. 2007; Kennedy et al. 2006b), immune mediated haemolytic anaemia (IMHA) (Kennedy et al. 2006a), hypothyroid disease (Kennedy et al. 2006c; Kennedy et al. 2006d; Wilbe et al. 2010a), anal furunculosis (Barnes et al. 2009; Kennedy et al. 2008), polyarthritis (Ollier et al. 2001), Addison's disease (Hughes et al. 2010), SLE related immune-mediated rheumatic disease (Wilbe et al. 2009), symmetrical lupoid onychodystrophy (Wilbe et al. 2010b), chronic inflammatory hepatitis (Dyggve et al. 2011), chronic superficial keratitis (Jokinen et al. 2010) and necrotising (MS-like) meningoencephalitis (Greer et al. 2010). It is also clear that for the same disease, different breeds may have different DLA associations (e.g., diabetes).

Candidate gene studies have been carried out for some of these diseases. These studies screen several genes that have been selected as potentially involved in the disease process. For diabetes we selected genes that had been shown to be associated with Type I diabetes in humans. We identified a complex pattern of associations, and different gene variants were associated with susceptibility in different breeds. Thus some associations were with increased susceptibility to the disease (IFN gamma, IL-10, IL-12 beta, IL-6, insulin, PTPN22, IL-4, and TNF alpha), whereas others were protective (IL-4, PTPN22, IL-6, insulin, IGF2, TNF alpha) (Short et al. 2007).

Currently Genome-wide Association Studies (GWAS) are being performed for many canine auto-immune diseases, in order to identify other regions of the genome that are associated with each disease.

Can We Use DLA Information to Reduce Disease Susceptibility?

There has been a suggestion that if a DLA allele or haplotype has been associated with a specific disease in a breed, then we should use this MHC information in mate selection to reduce the frequency of that haplotype.

I believe very strongly that we should not do this.

There may be a reason why a haplotype is at low frequency in a breed. Perhaps it is associated with another disease that is currently rare in the breed.

Table 1 shows the DLA haplotype frequencies for Dobermann Pinschers, based on a sample of 292. There are five haplotypes, one at a high frequency, 71%, one at a frequency of 14%, three at a frequency of about 4% plus a few others found in single dogs. This is a typical profile for a purebred dog breed. In the last column I have listed the diseases that are associated with each haplotype. It is clear why chronic inflammatory hepatitis is so common in this breed, since that disease is associated with the most frequent haplotype (no. 1). Similarly hypothyroid disease is associated with the second most frequent haplotype (no. 2). However, while the other three haplotypes have not been specifically associated with diseases in the Dobermann, two of them have been associated with either diabetes or IMHA in other breeds.

Table 1: Dobermann Pinschers (n=292).

I would suggest that trying to change the DLA profile of this breed may not be such a good idea. A similar situation is likely to exist in many other breeds.

Several of the studies on canine auto-immune diseases have shown that dogs who are homozygous for DLA may be at increased risk of developing these diseases. Certainly it appears that Dobermans that are homozygous for haplotype 1, are at increased risk of developing chronic inflammatory hepatitis (Dyggve et al. 2011). Many MHC studies in wild populations of different species, have suggested that heterozygosity confers a survival advantage. There are also data from human studies suggesting that spontaneous abortions are more frequent when the parents share MHC haplotypes.

Therefore it has been suggested that dog breeders should try to reduce homozygosity at the MHC.

I cautiously endorse this suggestion.

So my suggested use for MHC in mate selection is as follows:

 Choose several sires for your bitch, based on your usual criteria.

 If your bitch is homozygous for an MHC haplotype, then you could use MHC information to avoid selecting a sire that is homozygous for the same haplotype.
However, if all the sires have the same MHC, then it does not matter which one you choose.

 If your bitch is not homozygous, then you do not need to use MHC in your mate selection.

 It is much more important to avoid mating two dogs that are either both affected with the same disease or both have first degree relatives with the same disease.

 There is no reason to exclude homozygous dogs from mating, and it is perfectly acceptable to sell homozygous dogs.
Homozygous dogs may be slightly less able to respond to specific immune challenges, but since other genes also contribute to these processes, lack of variety at the MHC is not enough of a reason to exclude these dogs from the gene pool.

 The gene pool in any dog breed is fairly limited, so excluding dogs from the gene pool has to be very carefully considered.

It could be argued that the best solution would be to increase MHC diversity in dog breeds, but the only way to do this would be to out cross with other breeds.
I do not think this would be a popular solution!!

Could There Be Genetic Tests For Complex Diseases?

Monogenic diseases are caused by a defect in a single gene, which may be dominant or recessive. There are generally no environmental triggers, so that if a genetic test is performed, it is easy to predict whether an animal will develop the disease.

This is not the case for complex diseases. While we are beginning to identify some of the gene variants that are associated with each disease, it is clear that just having a single "bad" gene variant cannot be used to predict whether the disease will develop. Once we have identified more of the genes, we will then have to screen large cohorts of cases and controls for each breed and each disease, to estimate the risk each gene confers.

A genetic test for a complex disease will give an estimate of the risk that is conferred by the combination of all the genes that have been tested. However, if the environmental trigger is not encountered, even an animal with a high risk may still not develop the disease.

Summary

Auto-immune diseases are complex, and will have multiple risk and protective gene associations.

The MHC is just one of many such genes.

Each gene will confer a different level of risk.

Manipulating MHC haplotype frequencies may not reduce disease risk, and could cause more problems.

MHC information should be used with great caution in mate selection.

We may never have definitive yes/no genetic tests for complex diseases, or at least, not until we can identify the environmental triggers.

It is likely that such tests will give a measure of risk.

References/Suggested Reading

1.  Barnes A, O'Neill T, Kennedy LJ, Short AD, Catchpole B, House A, Binns M, Fretwell N, Day MJ, Ollier WE. Association of canine anal furunculosis with TNFA is secondary to linkage disequilibrium with DLA-DRB1*. Tissue Antigens 2009;73:218–224.

2.  Catchpole B, Ristic JM, Fleeman LM, Davison LJ. Canine diabetes mellitus: can old dogs teach us new tricks? Diabetologia 2005;48: 1948–1956.

3.  Dyggve H, Kennedy LJ, Meri S, Spillmann T, Lohi H, Speeti M. Association of Doberman hepatitis to canine major histocompatibility complex II. Tissue Antigens 2011;77:30–35.

4.  Greer KA, Wong AK, Liu H, Famula TR, Pedersen NC, Ruhe A, Wallace M, Neff MW. Necrotizing meningoencephalitis of Pug dogs associates with dog leukocyte antigen class II and resembles acute variant forms of multiple sclerosis. Tissue Antigens 2010;76:110–118.

5.  Hughes AM, Jokinen P, Bannasch DL, Lohi H, Oberbauer AM. Association of a dog leukocyte antigen class II haplotype with hypoadrenocorticism in Nova Scotia Duck Tolling Retrievers. Tissue Antigens 2010;75: 684–690.

6.  Jokinen P, Rusanen EM, Kennedy LJ, Lohi H. MHC class II risk haplotype associated with canine chronic superficial keratitis in German Shepherd dogs. Vet Immunol Immunopathol 2011;140(1–2):37–41.

7.  Kennedy LJ, Barnes A, Ollier WER, Day MJ. Association of a common DLA class II haplotype with canine primary immune-mediated haemolytic anaemia. Tissue Antigens 2006a;68:502–506.

8.  Kennedy LJ, Barnes A, Short AD, Brown JJ, Seddon JM, Fleeman LM, Brkljacic M, Happ GM, Catchpole B, Ollier WER. Canine DLA diversity: 3. Disease studies. Tissue Antigens 2007;69(Suppl 1):292–296.

9.  Kennedy LJ, Davison LJ, Barnes A, Short AD, Fretwell N, Jones CA, Lee AC, Ollier WER, Catchpole B. Identification of susceptibility and protective Major Histocompatibility Complex (MHC) haplotypes in canine diabetes mellitus. Tissue Antigens 2006b;68:467–476.

10. Kennedy LJ, Huson HJ, Leonard J, Angles JM, Fox LE, Wojciechowski JW, Yuncker C, Happ GM. Association of Hypothyroid disease in Doberman Pinscher dogs with a rare Major Histocompatibility Complex DLA class II haplotype. Tissue Antigens 2006c;67:53–56.

11. Kennedy LJ, O'Neill T, House A, Barnes A, Kyostila K, Innes J, Fretwell N, Day MJ, Catchpole B, Lohi H, Ollier WE. Risk of anal furunculosis in German Shepherd dogs is associated with the major histocompatibility complex. Tissue Antigens 2008;71:51–56.

12. Kennedy LJ, Quarmby S, Happ GM, Barnes A, Ramsey IK, Dixon RM, Catchpole B, Rusbridge C, Graham PA, Roethel C, Dodds WJ, Carmichael N, Ollier WER. Association of canine hypothyroidism with a common Major Histocompatibility Complex DLA class II allele. Tissue Antigens 2006d;68:82–86.

13. Ollier WER, Kennedy LJ, Thomson W, Barnes AN, Bell SC, Bennett D, Angles JM, Innes JF, Carter SD. Dog MHC alleles containing the "human RA shared epitope" confer susceptibility to canine rheumatoid arthritis. Immunogenetics 2001;53: 669–673.

14. Short AD, Catchpole B, Kennedy LJ, Barnes A, Fretwell N, Jones C, Thomson W, Ollier WER. Analysis of candidate susceptibility genes in canine diabetes. J Hered 2007;98:518–525.

15. Wilbe M, Jokinen P, Hermanrud C, Kennedy LJ, Strandberg E, Hansson-Hamlin H, Lohi H, Andersson G. MHC class II polymorphism is associated with a canine SLE-related disease complex. Immunogenetics 2009;61:557–564.

16. Wilbe M, Sundberg K, Hansen IR, Strandberg E, Nachreiner RF, Hedhammar A, Kennedy LJ, Andersson G, Bjornerfeldt S. Increased genetic risk or protection for canine autoimmune lymphocytic thyroiditis in Giant Schnauzers depends on DLA class II genotype. Tissue Antigens 2010a;75(6):712–719.

17. Wilbe M, Ziener ML, Aronsson A, Harlos C, Sundberg K, Norberg E, Andersson L, Lindblad-Toh K, Hedhammar A, Andersson G, Lingaas F. DLA class II alleles are associated with risk for canine symmetrical lupoid onychodystrophy [corrected](SLO). PLoS One 2010b;5:e12332.