Location: Children's Nutrition Research Center2009 Annual Report
1a. Objectives (from AD-416)
It has now been convincingly demonstrated by human epidemiological investigations that the origins of many of the major chronic diseases that are manifest in adult life have their origins during development. Additionally, molecular, cellular and animal studies have now shown conclusively that interference with, or alteration of, developmental pathways during critical windows of development can provide the pathophysiological basis for the events that take place later in adult life. Our research attempts to answer the following: 1) differentiate the effects of fetal versus postnatal maternal dietary protein restriction on satellite cell accretion and skeletal muscle mass; 2) determine if impaired catch-up growth upon nutritional rehabilitation is due to aberrant epigenetic mechanisms intrinsic to the satellite cell and/or an absence of the extracellular cues necessary to sufficiently accelerate satellite cell division; 3) develop novel techniques to study amino acid metabolism in conscious mouse models, with special emphasis on hepatic and enteral metabolism; 4)determine the role of urea cycle intermediates in maintaining nitric oxide and ureagenesis during different physiological and pathophysiological conditions; 5) determine gene expression in human lactating mammary epithelium; 6) characterize inbred mouse strains for lactation performance, gene expression, and weight gain among offspring in lean and obese animals, making use of a cross-fostering design where appropriate; 7) study the effect of nutrients on mammary gland development and function in mouse models; define the critical window for effects on mammary gland development and function; 8) elucidate the role of vitamin A in vascular development and hematopoiesis using mouse embryos, in vitro assays, and complementary techniques; and 9) identify target genes downstream of retinoic acid signaling that are required for blood and blood vessel development.
1b. Approach (from AD-416)
These research studies will use various techniques to accomplish the research to be undertaken. Establishing the critical window of development during which an inadequate nutrient supply permanently compromises the growth of the skeletal muscle and to understand the responsible mechanisms is of great importance. Research studies will focus on the satellite cell in animal models in order to understand how the nutrition of the fetus and infant has lifelong consequences for the health of the individual. Studies will take place on the role of amino acids in the urea cycle (crucial for nitric oxide synthesis), which is in demand for physiological (growth, pregnancy) and pathophysiological (trauma, sepsis) conditions. Lactation research will be conducted in mouse models to obtain an improved understanding of the genomic factors regulating mammary gland function that is central to providing therapeutic interventions that can aid women to establish and maintain a productive lactation. Since the abnormal formation of blood and blood vessels in adults is central to the progression of prevalent pathologies, including atherosclerosis, tumor angiogenesis, and anemia, researchers will develop an understanding of the cellular and molecular regulation of blood and blood vessel formation. Scientists will study retinoids in their role in the formation and maturation of blood vessels, as well as hematopoiesis, during embryonic development. Additionally, CNRC researchers will investigate, on a cellular and molecular level, the role of retinoid signaling in the regulation of these processes.
3. Progress Report
Skeletal muscles were collected from 21-d-old pups suckled on control and PR dams have been collected and subjected IHF analysis. We have performed manual morphometric analysis on a portion of the samples and evaluated the number of fibers that need to be analyzed to establish with confidence the various parameters of interest. This exercise identified the necessity of developing software to automate morphometric analysis of images so as to reduce the time required for analysis. Various versions of macros are being currently tested and validated. Additionally, we developed surgical techniques to be used later on to accomplish the goals of the project. We acquired transgenic mice and expanded our mice colony. We titrated the LPS response in control mice to determine the LPS dose in future studies. (Project 1) IRB applications for objective 1 have been submitted for approval. Once we have received permission we will initiate pilot studies to determine on which post partum day we can detect mRNA. This will be essential in the design of the subsequent studies. Additionally we have imported 32 strains of inbred mice and have established mating colonies of these strains to generate quantitative phenotypic data on traits linked to lactation performance. We have conducted a preliminary study comparing lactation capacity in an obese strain of mice, the Avy mouse, with that of a lean mouse, C57BL6/J. Also, we have started breeding mice to generate experimental animals for objective 3. (Project 2) We have determined the signaling hierarchy that mediates endodermal induction of mesoderm differentiation toward vascular endothelium, and migration of endothelial cells during vascular remodeling (Boles et al.; and another ms submitted). We have also identified candidate molecules that mediate RA regulation of embryonic hematopoiesis (Goldie et al. 2008a and 2008b), and are investigating the function of these in vitro (Kelly et al. 2008) and in vivo (Kienstra et al. 2008). (Project 3)
1. Intragastric Delivery of Nutrients: To conduct metabolic studies utilizing stable isotopes in mouse models it is essential that a steady metabolic state is attained. Children's Nutrition Research Center researchers have developed a surgical protocol for the gastric catheterization of mice that permits the infusion of nutrients (and tracers) directly into the stomach. This protocol will enable the study of mice in the fed state, as well as the delivery of tracers intragastrically. This technique is important for science and will let researchers study first pass metabolism of nutrients and intestinal metabolism in intact conscious mice. (Project 1: Organ-Specific Metabolism and Growth under Varying Nutritional Conditions during Development)
2. Maternal Obesity and Lactation: The purpose of this study was to establish the usefullness of a mouse model of obesity for studies to determine how maternal obesity impairs lactation. Children's Nutrition Research Center researchers demonstrated that obese Avy mice have decreased milk production during early lactation in comparison to lean mice. This was accomplished by comparing the rate of litter growth among the two strains (obese vs. lean mice) during the first 8 days of lactation. This finding supports the conclusion that the Avy mice will be a good model for use in understanding biologically, how maternal obesity impairs lactation. (Project 2: Functional Genomics of Lactation: Effects of Genetics, Hormones and Substrates)