Location: Houston, Texas2011 Annual Report
1a. Objectives (from AD-416)
Objective 1: Elucidate the role of vitamin A in vascular development and hematopoiesis using mouse embryos, in vitro assays, and complementary techniques. Sub-objective 1.A. Determine the intracellular mechanism(s) by which endothelial cell migration is regulated during vascular remodeling, and quantify the effects of retinoic acid deficiency on this process. Sub-objective 1.B. Investigate the role(s) of retinoic acid in the specification of yolk sac primitive endothelium to hemogenic endothelium and its differentiation to blood cell lineages. Sub-objective 1.C. Identify and characterize hemogenic endothelium within the aorta-gonad-mesonephros (AGM) region of the embryo, and determine whether retinoic acid regulates its fate and function. Objective 2: Identify target genes downstream of retinoic acid signaling that are required for blood and blood vessel development. Sub-objective 2.A. Determine which endoderm-derived soluble factors, downstream of retinoic acid, regulate endothelial cell migration and enable vascular remodeling. Sub-objective 2.B. Test whether genes downregulated in hemogenic endothelial cells in the absence of retinoic acid signaling mediate the specification of hemogenic endothelium. Objective 3: Gain knowledge regarding the impact of nutrition on the Hedgehog signaling pathway.
1b. Approach (from AD-416)
Children's Nutrition Research Center scientists will further investigate, on a cellular and molecular level, the role of retinoid signaling in the regulation of blood vessel formation and hemaotpoiesis. These studies will a employ transgenic mouse models to define the contribution of retinoids to these processes in vivo, and well-controlled cell culture systems to aid in elucidating their cellular and molecular roles in vitro. We will also employ RT-PCR and Microarray approaches to discover genes downstream of retinoic acid signaling that directly regulate blood and blood vessel formation. Information gained from these studies will further the understanding of normal development and the role of nutrients, such as retinoids, therein. Examine mutant animal models with adipose tissue-specific inactivation of Hedgehog pathway components.
3. Progress Report
In Obj. 1-2, we determined the characteristics of endothelial cells within the aorta that can make blood during embryonic development. We demonstrated their blood-forming ability in vitro and within embryos using real time imaging of transgenic mice. We also started studying the factors that regulate the development of these special endothelial cells, and are working toward understanding the role of Vitamin A in this process. In Obj. 3-4, to establish the hedgehog (Hh) pathway as an important regulator of adipose tissue formation, we examined the gene expression levels and cellular localization of the Hh pathway components in several previously published preadipocyte cell lines. We found that the core components of the Hh pathway are expressed in these cell lines and localize to the primary cilium, the cellular compartment responsible for mediating activation of the Hh signaling pathway. More importantly, we found that differentiation of these cell lines into mature adipocytes is reduced by Hh treatment, suggesting that the Hh pathway plays an inhibitory role in adipocyte maturation. Finally, we completed the setup of our mouse colonies for the proposed dietary studies to examine the expression levels of the Hh pathway components in the adipose tissue under different nutritional conditions. The ADODR monitors project activities by visits, review of purchases of equipment, review of ARS-funded foreign travel, and review of ARS funds provided through the SCA.
1. Characterization of hemogenic endothelial cells within the embryonic aorta (heart). Children's Nutrition Research Center researchers previously knew that blood was derived from aortic endothelial cells (a thin layer of cells that lines the interior surface of blood vessels) during development, but did not know the specific characteristics of these cells. These scientists, based in Houston, TX, discovered the cells' physical characteristics, and demonstrated their function in vitro and in vivo on a single cell level. These studies will allow researchers to prospectively isolate such cells and/or derive them from human stem cells so that we can use them to generate blood cells in vitro for hematopoietic (blood making) cell therapy. These findings provide possible technologies to address health concerns in the future.