The goal of this project is to delineate the role of oxidant signaling as the pivotal step leading to flow-induced dilation within the newborn intestinal circulation. Earlier work from this laboratory has determined that nitric oxide (NO) plays a substantial role in the regulation of the intestinal circulation during newborn life, much more so than during adult life. We contend that oxidantmediated activation of the endothelial isoform of nitric oxide synthase (eNOS) in response to shear stress is the principal means by which NO is generated in the newborn intestinal circulation. We have gathered preliminary data which demonstrate that: i) the dilatory response of in situ intestinal loops and in vitro terminal mesenteric arteries to a flow stimulus is significantly greater in 3d than in 40d old subjects and the flow-induced dilation in 3d arterioles is eliminated by the antioxidant N-acetylcysteine, ii) the rate of NO production by mesenteric arterial arcades closely correlates to flow rate and vascular resistance in 3d, but not 40d old subjects, iii) oxidant and NO are generated by shear stimulus applied to ld-MAECs, but not 180d MAECs; moreover, the shear-induced generation of oxidants and NO are blocked by the antioxidant diphenyliodonium, and iv) phosphorylation of Akt occurs in ld-MAECs in response to shear exposure. Based on these data, as well as recently published reports regarding the means by which shear stress activates eNOS, we propose the following novel hypothesis: the mechanostimulus of shear stress activates a signal transduction pathway leading to phosphorylation, and hence activation of eNOS that is unique to newborn mesenteric artery endothelial cells, and that the trigger event is an oxidant burst generated by NADPH oxidase within mesenteric endothelial cells. To test this hypothesis, we will: i) demonstrate that shear-induced oxidant production results in eNOS phosphorylation, and subsequent flow induced dilation; this work will be carried out in 1 degree cultures of mesenteric artery endothelial cells (MAECs) and in terminal mesenteric arterioles, ii) demonstrate that the shear-induced increase in NO production in ld-MAECs is mediated by a signal transduction pathway which includes Rac, PI-3K and Akt; this work will be carried out in MAECs and will involve the use of transient transfection with dominant-negative constructs of Rac and Akt, and iii) demonstrate that the principal factor which distinguishes ld from 180d old MAECs is the inability of the latter to produce oxidants in response to a shear stimulus; here. we will compare the effects of shear and non- shear stimuli on CNOS activity and also use a constitutively-active Rac construct to increase oxidant production in 180d-MAECs. The proposed work is relevant to the pathogenesis of necrotizing entercolitis, the most common gastrointestinal disease afflicting newborn infants. Vascular pathology plays a pivotal role in the etiology of this disuse. The novel mechanism proposed in this application may target agents which trigger the onset of NEC.
| Reber, Kristina M; Nankervis, Craig A (2004) Necrotizing enterocolitis: preventative strategies. Clin Perinatol 31:157-67 |
| Su, Baogen Y; Reber, Kristina M; Nankervis, Craig A (2004) Developmental expression of endothelin receptors in postnatal swine mesenteric artery. Pediatr Res 56:359-65 |
| Su, Baogen Y; Reber, Kristina M; Nankervis, Craig A et al. (2003) Development of the myogenic response in postnatal intestine: role of PKC. Am J Physiol Gastrointest Liver Physiol 284:G445-52 |
| Reber, Kristina M; Su, Baogen Y; Clark, K Reed et al. (2002) Developmental expression of eNOS in postnatal swine mesenteric artery. Am J Physiol Gastrointest Liver Physiol 283:G1328-35 |
| Reber, Kristina M; Nankervis, Craig A; Nowicki, Philip T (2002) Newborn intestinal circulation. Physiology and pathophysiology. Clin Perinatol 29:23-39 |
| Nankervis, C A; Reber, K M; Nowicki, P T (2001) Age-dependent changes in the postnatal intestinal microcirculation. Microcirculation 8:377-87 |
