Restriction fragment length polymorphism of the T-cell receptor beta-chain gene in dogs

Abstract Since T-cells and the T-cell receptor (TCR) play a pivotal role in the response of the immune system, they are a target for pathogenesis studies in immune mediated diseases and have been used to generate markers for T-cell dependent diseases in humans and dogs. TCR rearrangement is generated at the genomic DNA level and can be analyzed by Southern blotting techniques. In the present study this method to detect rearrangement of the TCR beta chain in the dog was critically examined. To search for restriction fragment length differences due to either inherited polymorphism or in diseases with suspected superantigen influence (X-linked severe combined immune deficiency and canine juvenile polyarteriitis syndrome) 13 dog families of three different breeds were examined. In addition primary spleen cell cultures, stimulated with either phytohemagglutinin A (PHA) or staphylococcus enterotoxin A (SEA) and B (SEB) were studied. The germline digest pattern of the enzymes Pst I, Sst I, Bgl II, Eco RI and Eco RV were identical in all dogs examined with the exception of one dog with canine juvenile polyarteriitis syndrome. In this dog an additional band was found in the Bgl II and Eco RV digestion suggestive of specific TCR rearrangement. Bam HI digestion revealed restriction fragment length polymorphisms (RFLPs) showing Mendelian inheritance. After digestion of the genomic DNA extracted from PHA, SEA or SEB stimulated spleen cells and Southern blot analysis, no differences in fragment patterns between the unstimulated cells and the stimulated cells could be detected. An important point to consider before a specific pattern variation between dogs is classified to be a marker for a specific disease or is used in pathogenesis studies, is the possibility of an inherited RFLP, especially after Bam HI digestion. In such studies the combined examination of the parents and the offspring must be recommended.