Specific Aims

The Tulane University Hypertension and Renal Center of Excellence (THRCE) has received over $11 million grant from National Center for Research Resources (NCRR) of the National Institutes of Health (NIH) to continue its research in the field of high blood pressure and its effect on the kidneys as well as the role of kidneys in the development of high blood pressure, through 2012. The center was first established by the NIH as one of its Centers of Biomedical Research Excellence (COBRE) with a $10.8 million grant in 2002.

In spite of Katrina, which led to serious disruption in its program, the first period of support was highly successful in fulfilling all the major goals originally established in 2002. Under COBRE, the Hypertension Center continued to grow and develop an outstanding program including support of ten junior faculty investigators, development of a state-of-the-art molecular and analytical core facilities, sponsorship of local and regional meetings on Hypertension, and public education programs to increase awareness of the dangers of hypertension. In addition to support of the ten junior faculty investigators, the COBRE project provides support for post-doctorial fellows, graduate students, and summer research fellowships for medical students. As a mark of success, five original junior faculty investigator received substantive NIH and other funding.

For the renewal period, the objectives are to provide an enriched mentoring environment to junior faculty investigators so that they can achieve nationally competitive status and to augment and strengthen the biomedical research capacity and infrastructure for Tulane investigators in hypertensive renal and cardiovascular diseases. The new COBRE grant will support junior faculty investigators from the Departments of Physiology, Medicine, and Pediatrics in the School of Medicine , and from the Department of Epidemiology in the School of Public Health and Tropical Medicine.

Projects & Investigators

Transcriptional Control of Ureteric Bud Growth and Branching
	Zubaida Saifudeen, Ph.D., Assistant Professor, Department of Pediatrics, Section of Nephrology, Tulane University Health Sciences Center. Dr. Saifudeen joined the Section of Pediatric Nephrology at Tulane University School of Medicine in July 1999 as a Postdoctoral fellow. After completing a NRSA fellowship (2002-2005) in Dr. Samir El-Dahr's lab, Zubaida stayed on as Research Assistant Professor at Tulane Pediatrics. Early in her post-doctoral fellowship, Zubaida discovered that the bradykinin B2-receptor (B2R), a G-protein coupled seven transmembrane receptor, is a p53-target gene (JBC, 2000), a novel finding which had many important scientific implications.
The goal of Dr. Saifudeen's project is to understand the role of p53 protein in early kidney development, as aberrations during the early stages of development can lead to congenital defects. Dr. Saifudeen has found that loss of p53 in mice embryos causes kidney structure and growth abnormalities (kidney/ureter duplication, obstruction of ureter). Infants born with duplex kidneys may suffer from unilateral or bilateral vesicoureteral reflux, hydronephrosis, and cystic-dysplasia. A subset of p53-null/deficient mice exhibit profound defects in early renal development including duplex ureters/kidneys, hypoplasia and hydronephrosis.  The etiology of these abnormalities is currently unknown.  The objective of this proposal is to elucidate the role of p53 in early events of renal development with specific emphasis on UB outgrowth and patterning.  The overall hypothesis is that p53 expression in the metanephric anlagen is required to restrict UB induction to one specific site along the nephric duct.  In Specific Aim 1, the nephric cell lineage in which p53 expression is required to restrict UB induction to a specific site on the nephric duct will be determined by A) in situ hybridization, and B) Conditional deletion of the p53 ORF from either the UB or the MM using cre-lox transgenic mice. In Specific Aim 2, we propose to investigate the mechanisms by which p53 controls UB growth and branching.  Identifying how p53 works in early kidney development will be an important step to formulating new therapies against congenital kidney diseases

Citrate transport in the proximal tubule
	Junior Faculty Investigator: Kathleen S. Hering-Smith, Ph.D., Research Assistant Professor, Department of Medicine, Tulane University School of Medicine. Dr. Hering-Smith has extensive experience in studies of sodium transport and acid-base transport including that of citrate; these studies have primarily utilized isolated perfused tubules, cell culture, and a variety of cell and molecular techniques.  Her career as an independent investigator began after she received her PhD from Tulane University in 2004.
Urinary citrate is one of the most important inhibitors of calcium nephrolithiasis Various studies estimate that 19-63% of individuals with calcium containing kidney stones have hypocitraturia as a contributing cause.  Understanding the mechanisms of the regulation of citrate transport will hopefully lead to improved diagnosis of causes of hypocitraturia. Urinary citrate is an important inhibitor of calcium nephrolithiasis and is primarily determined by fractional reabsorption in the proximal tubule. The dicarboxylate transporter (NaDC1) is presumably the main mechanism of apical uptake of filtered citrate along the nephron. The most important physiologic regulator of urinary citrate excretion is acid-base status. Also urinary citrate increases as urinary calcium increases. The proposed studies will address the acute regulation of citrate transport by calcium, and chronic regulation of citrate transport by acid-base perturbations and hypokalemia. Using a newly characterized in vitro model of citrate transport, OK cells studied under particular conditions, citrate and dicarboxylate uptake are sensitive to extracellular calcium. These studies indicate that the OK cell citrate transport system is likely a novel citrate transporter.  Recently another cell line of dicarboxylate transport was developed.  Human retinal pigmented epithelial cells stably transfected with human NaDC1 (CUBS cells) are responsive to acid-base conditions in vitro and will therefore represent a powerful new model. Two hypotheses will be examined: 1. Calcium acutely inhibits a novel citrate transport process in mammalian proximal tubule cells.  2. Chronic regulation of proximal tubule transport of citrate is accomplished by redundant mechanisms including changes in NaDC1 protein production and insertion of pre-existing NaDC1 protein into the apical membrane from sub-apical vesicles.  The specific aims are:  1.  To delineate the calcium sensitive citrate transport process by: demonstrating that the calcium sensitive citrate transport process is a novel transporter, not NaDC1, and determining the cellular mechanisms whereby extracellular calcium alters this citrate transport process. 2.  To delineate the mechanisms of chronic regulation of citrate transport by acid-base perturbations and hypokalemia.   To achieve this aim three modes of regulation will be examined: transcriptional (or mRNA stability) regulation, regulation at the protein level, and regulation by trafficking of NaDC1 into and out of the apical membrane from sub-apical vesicles under conditions of metabolic acidosis and hypokalemia.

Heme Oxygenase in Angiotensin II Hypertension
	Fady T. Botros, Ph.D., Instructor, Department of Physiology, Tulane University School of Medicine. Dr. Botros received his Ph.D. in the Department of Pharmacology at New York Medical College in 2004.  He completed his postdoctoral fellowship with us in the Department of Physiology and was appointed as Instructor in January 2007.  His research interests include cardiovascular and renal physiology with emphasis on the role of the heme-heme oxygenase (HO) system in normal regulation of vascular function and how dysfunction of this system may contribute to the development of hypertension.
Hypertension affects about 33 % of the population; if not controlled, it may cause end organ damage including renal failure. The angiotensin II-infused rat is a model for angiotensin II-dependent hypertension, in these rats both kidneys are exposed to elevated levels of angiotensin II and elevated perfusion pressure. Previous studies have shown that heme oxygenase (HO), an enzyme that catalyzes the conversion of heme to biliverdin, free iron and carbon monoxide (CO), is upregulated in kidneys from angiotensin II-infused rats indicating a possible protective role of heme oxygenase in angiotensin II-mediated hypertension. We hypothesize that heme oxygenase is upregulated in kidneys of angiotensin II-dependent hypertensive rats, and that HO-derived metabolites (CO and/or bilirubin) regulate renal hemodynamics by counteracting the effects of elevated angiotensin II levels and dilating the renal afferent and efferent arterioles, thus improving renal function. The aims of this study are: 1) To determine the effect of chronic angiotensin II infusion on renal arteriolar and tubular HO expression using immunohistochemical and western blot analyses. 2) To determine the effect of HO induction or inhibition on afferent and efferent arteriolar vasoactivity and responsiveness to angiotensin II, and on afferent arteriolar autoregulatory responses in normotensive and angiotensin II-hypertensive rats. Afferent and efferent arteriolar responses will be evaluated using the blood-perfused juxtamedullary nephron preparation. 3) To determine the effect of acute HO inhibition on renal hemodynamics, autoregulation, and pressure-natriuresis in angiotensin II-infused rats, using in vivo renal clearance studies.

Macronutrient Composition of Diet and Risk Factors for Cardiovascular Disease
	Lydia A. Bazzano, M.D., Ph.D., M.PH., Assistant Professor of Epidemiology, Department of Medicine, Section of Epidemiology, Tulane University School of Public Health and Tropical Medicine. Dr. Bazzano received both her MD and Ph.D. from Tulane University , earning her Ph.D from the Department of Epidemiology in 2000 and MD in 2002.  She completed her Medical Internship and Residency at Harvard Medical School , and in 2005, was appointed as Assistant Professor of Epidemiology in  the Tulane University School of Public Health and Tropical Medicine.  Her proposed study addresses important clinical and public health issues, which will contribute to our knowledge regarding the efficacy of low-carbohydrate diets on traditional and novel cardiovascular disease risk factors.
Cardiovascular diseases (CVD) remain the leading cause of death globally as well as here in the United States . Manipulations of the macronutrient (protein, carbohydrate and fat) contents of diet have been used extensively for weight loss and weight control in the past several decades. Low carbohydrate diets, in particular, have gained popularity for weight loss. However, few studies have examined the effects of a diet low in carbohydrates on traditional and novel cardiovascular risk factors in the long term, particularly in contrast to the current dietary recommendations for decreased fat intake to reduce risk of CVD. In this proposal, we plan to conduct a 12-month, parallel-arm, randomized controlled trial of a diet low in carbohydrates versus the currently recommended diet low in fat diet to reduce CVD risk factors among obese adults. The objective of this trial is to examine the long-term effects of a diet low in carbohydrates, as compared to one low in fat, on CVD risk factors, including blood pressure (BP), body weight and composition, serum lipids, plasma glucose, insulin, adipocytokines (adiponectin, leptin, resistin), and C-reactive protein (CRP) among obese adults. In order to accomplish these objectives we will randomize 130 eligible participants (n=65 in each group) to consume either a diet low in carbohydrates (<40 g/d) or a diet low in fat (<7% saturated fat, <35% total fat). Neither of the diets will be energy-restricted. Participants will meet with a dietitian for one-on-one counseling session weekly for the first 4 weeks, then bi-weekly in small group sessions for the next 5 months, and monthly in larger group sessions for the final 6 months of the intervention. Data on both traditional and novel CVD risk factors will be collected at baseline, 3, 6, and 12 months. We hypothesize that a diet low in carbohydrates as compared to a diet low in fat will lower systolic and diastolic BP, body weight, total percent body fat, waist circumference, serum levels of triglycerides, and plasma levels of insulin, glucose, leptin, resistin, and CRP, and increase serum levels of HDL-cholesterol and adiponectin. Because CVD is the most common cause of death here in the U.S. and world-wide, this study has important public health implications. It will provide new information on the potential long-term effects of diets low in carbohydrates on both the traditional risk factors for CVD as well as novel risk factors and inflammatory factors.  The results from this study will help to determine if a diet low in carbohydrates as compared to the currently recommended low fat diet can decrease the risk of CVD among obese adults. In addition, this trial will provide the junior investigator, Dr. Bazzano, an opportunity to acquire research skills and experience necessary to launch a successful, sustainable, and independent research career in the investigation and dietary and lifestyle approaches to reduce risk factors for hypertension and CVD.

Endothelial Dysfunction, Adipocytokins, Inflammation & Chronic Kidney Disease
	Jing Chen, M.D., M.Sc., Assistant Professor of Medicine and Epidemiology, Department of Medicine, Tulane University School of Public Health and Tropical Medicine. Dr. Chen received her MD at Jiangxi Medical College , PR China, in 1984 and MSc in Biotechnology from Johns Hopkins University   in 1995. She was appointed Assistant Professor in the department of Medicine, section of Nephrology in 2003. Her research interest is primarily related to the role of endiothelial dysfunction in contributing to chronic kidney disease and related cardiovascular disease.
Chronic kidney disease (CKD) has become an important public health challenge in the United States . CKD is a major risk factor for end-stage renal disease (ESRD), cardiovascular disease (CVD), and premature death. Understanding novel risk factors for CKD may provide effective approaches for early intervention in order to reduce the morbidity and mortality related to CKD. Endothelial dysfunction, adipocytokines and inflammation have been associated with ESRD and CVD in small clinical studies and animal experiements. However their role in the etiology of CKD has not been established.  The overall objectives of this proposed study are to examine the effects of endothelial dysfunction, adipocytokines, and inflammation on the risk of CKD. The specific aims of the proposed study are:  (1) to examine the association between biomarkers of endothelial dysfunction (plasma levels of asymmetric dimethylarginine, endothelin-1, intercellular adhesion molecule-1, vascular cell adhesion molecule 1, E-selectin, L-arginine and NO2/NO3 (NOx), and urinary excretion of NO2/NO3 (NOx)) as well as endothelial function assessed by brachial artery reactivity using high-resolution ultrasound and risk of CKD; (2) to examine the association between adipocytokines (leptin, resistin, and adiponectin) and risk of CKD; (3) to examine the association between inflammation (C-reactive protein, interleukin-6, and tumor necrosis factor-a) and risk of CKD; and (4) to examine the correlation between biochemical markers of endothelial dysfunction and endothelial function assessed by brachial artery reactivity using high-resolution ultrasound. This study has important clinical and public health implications. Understanding the nature of endothelial dysfunction, adipocytokines and inflammation in patients with CKD will provide insight into developing tailored intervention strategies including normalizing endothelial dysfunction, targeting adipocytokines and inflammation for the prevention and treatment of CKD and related CVD. In addition, the proposed study, if funded, will provide important preliminary data to conduct a prospective cohort study to examine the longitudinal association of endothelial dysfunction, adipocytokines and inflammation with the progression of CKD and related CVD.

Mechanism of Resistance Artery Structural Remodeling in Hypertension
	Khalid Matrougui, Ph.D., Research Assistant Professor, Department of Physiology, Tulane University School of Public Health and Tropical Medicine. Dr. Khalid Matrougui graduated from the University of Paris VI, France, earning his MS in 1995, and his PhD in 1998. His post-doctorial trainings were at the Aarhus University, Denmark, the University Paris VII, France, and the University of Alabama at Birmingham, USA. Dr. Matrougui continued as Faculty Instructor at UAB's department of Biophysics & Physiology until his appointment as faculty member to Louisiana State University Health Science Center. Dr. Matrougui joined Tulane School of Medicine in 2007, as Research Assistant Professor to the Department of Physiology, bringing with his appointment a new dimensions of expertise related to the growing efforts in Vascular Biology and Pathophysiology of Hypertension.
Hypertension, the most important risk factor for cardiovascular disease, is at epidemic levels in the UNITED STATES being responsible for increased prevalence of vascular complications, and morbidity and mortality. Many studies have shown mophological changes in resistance arteries (RA) from hypertensive patients and animal models; however, in vivo signaling pathways contributing to vascular structural remodeling in hypertension are poorly understood. Preliminary data show increased collagen type 1 content, artery stiffness and eutrophic structural remodeling induction of RA from angiotensin II (ANG II)-dependent hypertensive mice. Those RA morphological changes are associated with increased oxidative stress, inhibitory IkappaB proteins (IKB) phosphorylation, p50/p65 Nuclear Factor kappa B (NFkB) phosphorylation and translocation to the nucleus, avß3-integrin shedding, and TGFß1 expression. Based on these findings, Dr. Matrougui hypothesizes that elevated oxidative stress activates NFKB pathway leading to increase of avß3-integrin shedding and/or TGFß1 expression and bio-activity, which induces abnormal accumulation of collagen type 1 responsible for structural wall remodeling and increased stiffness of RA from ANG II-dependent hypertensive mice. The goals of this project are: 1) To demonstrate that increased collagen type 1 content, stiffness, and eutrophic remodeling induction are dictated by enhanced oxidative stress-dependent NFKB pathway activation of RA from ANG II-dependent hypertensive mice. In this aim we will determine that oxidative stress is upstream signaling that activates NFkB pathway responsible for morphological changes of RA from ANG II-dependent hypertensive mice. 2) To delineate the role of enhanced avß3-integrin shedding and TGFß1 expression on increased collagen type 1 content and stiffness, and eutrophic remodeling induction of RA from ANG II-dependent hypertensive mice. In this aim we will establish the role of avß3-integrin and TGFß1, down stream signaling to oxidative stress and NFkB, regulating collagen type 1 turnover, structural remodeling and stiffness of resistance arteries from ANG II-dependent hypertensive mice.