Recipient Organization
UNIV OF MARYLAND
(N/A)
COLLEGE PARK,MD 20742
Performing Department
Animal and Avian Sciences
Non Technical Summary
Copper (Cu) plays essential roles for normal growth and development. Genetically encoded diseases of metal balance, or dietary deficiency, lead to diseased states such as anemia, cognitive disorders, abnormal development, cardiovascular disease, and poor growth. With respect to animal growth and productivity it is critical to understand how animals might be genetically predisposed to abnormal metal conditions and how optimal levels of essential metals can contribute to animal health and productivity. This study will determine if Ctr1 Cu transporter insufficiency have an increased risk to cardiac failure in response to physiological and pathological stress conditions. Moreover, this investigation would provide a basis for examining whether variant forms of the Ctr1 gene in animals could be a critical factor in predisposing animals to cardiac diseases, which may arise as a result of dietary Cu deficiency, pregnancy, or other stresses, and determining whether Cu supplementation for Ctr1 insufficiency could be an effective treatment.
Animal Health Component
0%
Research Effort Categories
Basic
100%
Applied
0%
Developmental
0%
Goals / Objectives
The long-term goals of our studies are to delineate how animals regulate copper (Cu) homeostasis at the organismal level and to further elucidate the roles of Cu and the Ctr1 Cu transporter in preventing cardiovascular disease. Cu plays key catalytic and regulatory functions in biochemical reactions that are essential for normal growth, development and health. Defects in Cu metabolism cause Menkes and Wilson's disease, myeloneuropathy, cardiovascular disease and are associated with other patho-physiological states. Consequently, it is critical to understand the mechanisms by which cells control the acquisition, distribution and utilization of Cu. Although dietary Cu deficiency in animals leads to the development of cardiac hypertrophy, it was unclear whether this was due to a peripheral Cu deficiency or an intrinsic cardiac requirement for Cu. To ask this fundamental question, we previously generated mice with a cardiac-specific Ctr1 Cu importer knockout (Ctr1hrt/hrt), and these mice developed a severe cardiac hypertrophy. This investigation established a cardiac-intrinsic requirement for Cu. While Ctr1 has not previously been linked to cardiovascular disease, we outline experiments to explore whether the cardiac functions of systemic Ctr1 heterozygous mice (Ctr1+/-) are more vulnerable to stressed conditions, including pregnancy. Furthermore, we will evaluate Ctr1 SNPs as a potential risk factor for cardiomyopathy. The studies outlined in this proposal have the potential to make significant progress in our understanding of how Cu deficiency impacts human health and cardiovascular disease.Delineation of the roles of Cu and Ctr1 Cu transporter in preventing cardiovascular disease: a. Influence of Ctr1 gene dosage on pregnancy-induced cardiac hypertrophy. b. Evaluation of the Ctr1 Cu transporter as a potential risk factor for cardiomyopathy. c. Evaluation of the Ctr1P25A localization and regulation in cell culture models.
Project Methods
Influence of Ctr1 gene dosage on pregnancy-induced cardiac hypertrophy: To firmly ascertain whether Ctr1 haplo-insufficiency and the corresponding Cu deficiency predisposes mice to pregnancy-induced cardiac dysfunction, we will investigate whether multiple rounds of pregnancy can cause cardiac dysfunctions and death for the systemic Ctr1 heterozygous (Ctr1+/-) mouse, since a Ctr1+/- mouse models reflect the genetic situation of a human better than the Ctr1hrt/+ mouse. We will characterize cardiac physiology, histology and the molecular markers of Ctr1+/- mice fed a Cu-deficient diet, and under conditions of multiple pregnancies and pressure overload.Evaluation of the Ctr1 Cu transporter as a potential risk factor for cardiomyopathy:We found that the protein expressed from the Ctr1 (P25A) allele is partially defective in fostering Cu+ uptake, resulting in a relative Cu deficiency. Using tissue culture models including Ctr1-/- MEFs, we will evaluate the functional consequences (Cu levels, Cu-dependent enzyme activities) of all other known non-synonymous Ctr1 encoding SNPs (T27I, G60D, I152M,T122S), which may cause a functional consequence to Ctr1 as a Cu importer. Evaluation of the Ctr1P25A localization and regulation in cell culture models: Wild type Ctr1 localizes to the apical membrane of intestinal epithelial cells (IECs) and to intracellular vesicular compartments. We will use microscopic and biochemical approaches (cell surface biotinylation assay) in MDCK cells stably expressing wild type Ctr1 or Ctr1P25A to determine if Ctr1P25A is mislocalized.