| Immune-mediated hemolytic anemia (IMHA) results in regenerative anemia and is the most well studied immune-mediated anemia in dogs. However, the underlying cause of IMHA remains unknown, and mortality remains a problem. In a different condition, dogs with suspected precursor-targeted immune-mediated anemia (PIMA) present with nonregenerative anemia and ineffective erythropoiesis, which have occasionally been associated with phagocytosis of erythroid precursors (rubriphagocytosis) or myelofibrosis. The pathogenesis of PIMA has not yet been determined, but roles for immunoglobulin and/or complement have been suspected. Additional involvement of apoptosis-like mechanisms with phosphatidylserine (PS) exposure is possible in both canine IMHA and PIMA based on previous studies in people and dogs. Our central hypothesis is that PIMA is part of a spectrum of immune-mediated anemias including IMHA, in which IgG, with or without the contribution of phosphatidylserine (PS), target and promote phagocytosis of different stages of erythroid cells, simultaneously or independently. The rationale was that better characterizing the pathogenesis of PIMA and its association with IMHA will ultimately help establish diagnostic criteria, identify appropriate therapeutic strategies based on knowledge of the pathogenesis of the diseases, and raise veterinary awareness of PIMA. This dissertation first describes a retrospective study of dogs with PIMA with the main goal of helping characterize canine PIMA and facilitate its recognition and diagnosis. Then it describes the development of flow cytometric assays for RBC and erythroid precursor IgG, including a Percoll gradient separation for erythroid populations. These methods were used for isolation of erythroid populations and their assessment for IgG and PS in IMHA and PIMA dogs, healthy dogs, and sick dogs with no evidence of IMHA or PIMA. We found that IMHA dogs had significantly higher IgG and PS when compared to healthy and non-IMHA dogs. Additionally, we showed that a subset of PIMA dogs had increased IgG-positive erythroid precursors when compared to healthy and non-IMHA dogs, and erythroid precursors from most tested PIMA dogs had more PS-positive erythroid precursors than healthy dogs; however, no sick dogs without PIMA were tested for comparison. These findings suggest a role for IgG in canine PIMA and for PS in canine IMHA and PIMA. Finally, we demonstrated the expression of DEA1.1 on canine erythroid precursors from rubriblasts through metarubricytes. |
