Overview of the Issue

 What are complex diseases?

 Major histocompatibility complex (MHC) associations with auto-immune diseases

 Identification of other genes

 Can we use this information to reduce the incidence of these diseases in dogs?

 Could there be a genetic test for complex diseases?

Summary

Auto-immune diseases are complex diseases. These are diseases that occur as a result of the influence and interaction of multiple genes, together with an environmental trigger. However, the critical feature of these diseases is that they only occur after exposure to an environmental trigger. As yet, most environmental triggers have not been identified. So whether or not someone (some dog?) develops the disease, depends on the particular combination of variants of the risk genes that they have, plus exposure to the environmental trigger.

Some genes will have a greater influence than others. Variants of 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 some variants that do not confer any increased or decreased risk of disease.

The common feature of auto-immune diseases is that the body starts to attack a particular organ or tissue. Thus, in hypothyroidism, the body starts destroying the thyroid gland, preventing the production of thyroxine. Lack of thyroxine causes the symptoms of the disease.

Within the genome there is a group of genes called the major histocompatibility complex (MHC), which plays a central role in the immune response. It is therefore likely that these genes may influence resistance and susceptibility to auto-immune diseases. In the dog the MHC genes are known as DLA (dog leucocyte 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 (every dog?) inherits one set (or haplotype) from each of their parents. Many variants have been identified for each of these genes, e.g., there are over 150 different DLA-DRB1 variants, and over 180 different three-locus haplotypes are now known. However, within any one dog breed there will only be five to seven haplotypes, of which two or three will be at high frequency. Therefore, while there is a lot of diversity within the overall dog population, within each breed there is a much less diversity.

Many studies have shown clear associations of canine auto-immune diseases with DLA. These include diabetes, immune-mediated haemolytic anaemia (IMHA), hypothyroid disease, anal furunculosis, polyarthritis, Addison's disease, systemic lupus erythematosus (SLE)-related immune-mediated rheumatic disease, symmetrical lupoid onychodystrophy, chronic inflammatory hepatitis, chronic superficial keratitis and necrotising (multiple sclerosis (MS)-like) meningoencephalitis. It is also clear that for the same disease, different breeds may have different DLA associations.

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 (interferon (IFN) gamma, interleukin (IL)-10, IL-12 beta, IL-6, insulin, protein tyrosine phosphatase, non-receptor type 22 (lymphoid) (PTPN22), IL-4, and tumour necrosis factor (TNF) alpha), whereas others were protective (IL-4, PTPN22, IL-6, insulin, insulin-like growth factor (IGF2), TNF alpha).

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 MHC Data 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.

Figure 1 shows the DLA haplotype frequencies for Dobermanns, based on a sample of 292 dogs. 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 are 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.

I would suggest that trying to change the DLA profile of this breed would 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 Dobermanns that are homozygous for haplotype 1, are at increased risk of developing chronic inflammatory hepatitis. 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.

It has been suggested, therefore, that dog breeders should try and reduce homozygosity at the MHC. I cautiously endorse this suggestion. 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, i.e., introduce other haplotypes that are not currently in the breed. The only way to do this would be to out cross with other breeds. Changing the frequency of existing haplotypes within a breed does not increase diversity.

Figure 1. DLA haplotype frequencies for Dobermanns (n = 292).

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 need 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.

Note: All samples were obtained through animal hospitals or veterinary surgeons. We do not use any experimental animals.

References

1.  Kennedy LJ, Barnes A, et al. Association of a common DLA class II haplotype with canine primary immune-mediated haemolytic anaemia. Tissue Antigens 2006;68:502–506.

2.  Kennedy LJ, Barnes A, et al. Canine DLA diversity: 3. Disease studies. Tissue Antigens 2007;69 Suppl 1:292–296.

3.  Kennedy LJ, O'Neill T, et al. Risk of anal furunculosis in German Shepherd Dogs is associated with the major histocompatibility complex. Tissue Antigens 2008;71:51–56.

4.  Short AD, Catchpole B, et al. Analysis of candidate susceptibility genes in canine diabetes. Journal of Heredity 2007;98:518–525.