Research in the division of pediatric endocrinology includes basic science, translational and clinical research. The division is host to a wide variety of research programs encompassing various aspects of the endocrine system of relevance to children. Topics of research include hypoglycemia, type 1 and type 2 diabetes, growth disorders, disorders of bone and mineral metabolism and obesity.
- De Leon Lab
- Grimberg Research
- Kelly Research
- Kublaoui Lab
- Levine Lab
- Levitt Katz Lab
- Lipman Research
- Magge Research
- Stanley Lab
- Willi Research
Mouse models of hyperphagic obesity, stereotaxic cannulation of mouse brain ventricles and parenchyma, isolation of hypothalamus for RNA, isolation of brain nuclei by laser capture microdissection followed by qPCR, sectioning of hypothalamus for immunostaining, and microscopy.
I am a pediatric and adult endocrinologist with an interest in obesity and the hypothalamic regulation of feeding. Our laboratory studies the paraventricular nucleus of the hypothalamus (PVN), oxytocin and the hypothalamic regulation of feeding. We have used various mouse models to study the PVN including Sim1+/- mice. We are currently using other mouse lines, to examine the importance of each neuron subtype in feeding regulation. We use various methods including surgical, pharmacological, feeding, ablation, to investigate the following:
PVN neurons in feeding regulation: Obesity is an epidemic and feeding regulation by the brain remains poorly understood. The paraventricular nucleus of the hypothalamus (PVN) functions to integrate peripheral adiposity signals such as leptin and is important in meal termination. Its ablation results in hyperphagic obesity and electrical stimulation results in a reduction in food intake. The PVN harbors the cell bodies of second order neurons that receive projections from leptin responsive Pomc and Npy/Agrp neurons in the arcuate. PVN Oxytocin (Oxt) and corticotropin releasing hormone/factor Crh/Crf neurons are leptin and Mc4r responsive and project to the Nucleus of the solitary tract (NTS) also known as the satiety center, where they interact with CCK responsive neurons. Centrally administered Oxt or Crh lead to a reduction in food intake. An Oxt antagonist delivered to the hindbrain blocks leptin-mediated reduction in food intake. We and others observed that haploinsufficiency of Sim1 results in hyperphagic obesity in humans and mice. Expression of Oxt in the PVN is reduced by 80% in Sim1+/- mice, and their hyperphagia is rescued by central Oxt administration at doses that do not affect wild-type mice. In addition they have reduced expression of Crh. Although there is ample pharmacological evidence for an important role for Oxt in feeding regulation, Oxt-/- mice are not hyperphagic. This discrepancy between neuro-anatomical and pharmacological data on one hand and mouse models on the other remains unexplained. The same discrepancy exists for Npy and Agrp whose role in feeding regulation is undisputed and pharmacological and electrophysiological evidence strongly supports the role of these neuropeptides in feeding regulation but where the Npy-/-, Agrp-/- and the Npy-/-,Agrp-/- double knockouts have no feeding phenotypes. Ablation of the Npy/Agrp neurons in adult mice but not in neonates led to starvation suggesting the presence of developmental compensatory mechanisms in knockout models. We postulate that a similar mechanism exists for Oxytocin and Crh neurons. We aim define the role of each of the above neuron subtypes in the PVN to examine the effect on feeding regulation and body weight.
Mechanism of hyperphagia in Prader-Willi Syndrome (PWS): PWS is a genetic disorder that causes neonatal feeding difficulties and failure to thrive followed by a switch to severe hyperphagia in childhood. The pathogenesis of every aspect of feeding regulation in PWS is unknown. In fact, basic elements of our understanding of the hypothalamic regulation of feeding are missing. PWS is caused by loss of several paternally inherited genes on a small locus on chromosome 15. The mouse PWS locus contains the same genes in the same order as the human locus. Like humans, mouse models of PWS also display neonatal failure to thrive with the larger deletions leading to earlier neonatal lethality. Unlike humans, PWS mice do not develop hyperphagic obesity. Many of the genes in the PWS locus are expressed in the hypothalamus and PWS is characterized by hypothalamic dysfunction. Importantly, patients with PWS have a reduced number of oxytocin neurons in the paraventricular nucleus (PVN) of the hypothalamus as well as low circulating levels of oxytocin. Oxytocin and parvocellular oxytocin neurons of the PVN have been clearly implicated in the hypothalamic regulation of feeding. We showed that Sim1 deficient mice, a mouse model of human monogenic obesity, are deficient in oxytocin and by centrally replacing oxytocin we rescued their hyperphagia. We aim to define the role of the PVN and oxytocin in feeding regulation in PWS.
Levitt Katz Lab
Expertise in carbohydrate metabolism and the physiology of the insulin-like growth factor (IGF) system, insulin resistance, diabetes, and the endocrine aspects of the chromosome 22q11.2 deletion syndrome.
Dr. Levitt Katzís focus is patient-oriented research related to obesity and the metabolic syndrome, Type 2 diabetes and the prevention of Type I diabetes. She is the CHOP Principal Investigator for the Treatment Options for Type 2 Diabetes in Adolescents and Youth (TODAY) study, the NIH-sponsored multicenter study that will define the optimal treatment strategies for youth with new onset Type 2 Diabetes mellitus. She has served as a leader in the national steering committee of TODAY as well as serving on key subcommittees in planning the study, as well as substudies on cardiovascular risk. In addition, Dr. Levitt-Katz leads our efforts at CHOP in the TODAY2 follow-up study and the TODAY Genetics 2500, the multicenter study that will be the first to characterize the genetic markers of T2DM in youth. Nationally , she also serves as the CHOP PI of Type I diabetes TrialNet study that performs studies of risk assessment and prevention for youth and adults at increased risk of Type 1 Diabetes, and was the former site PI of the preceding study, The Diabetes Prevention Trial, DPT-1.
Dr. Levitt Katz received her initial training as a CAP Awardee from the NCRR to study the physiology of the insulin-like growth factor axis and carboydrate metabolism. Her curent independent research efforts include the natural history of pediatric T2DM and the pediatric metabolic syndrome. She has published results of a study of IGFBP-1 and C-peptide and other metabolic markers differentiating pediatric T1DM and T2DM as well as the first longitudinal study of glycemic control and markers of insulin secretion in youth with newly diagnosed Type 2 diabetes who were followed for 4 years duration. Collaborative work is ongoing with investigators from the Penn Weight and Eating Disorders Center as well as the CHOP Sleep Center and the Chromosome 22q11.2 Center. She served as a coinvestigator with Dr. Robert Berkowitz, former chief of Child Psychiatry, on the NIDDK funded adolescent weight loss study and is currently analyzing the metabolic and hormonal response to weight loss in this population. Dr. Levitt Katz recently published a study of sleep architecture in obese youth that defined a U-shaped relationship between sleep duration and short and long-term glycemia.
At CHOP, Dr. Katz is devoted to training fellows and junior faculty in all aspects of Patient Oriented Research. She is Associate Director of the Clinical Translational Research Center and take a leadership role in mentoring fellow and junior faculty to establish patient-oriented research studies. She previously served as a key member of the CHOP Healthy Weight Advisory Board.
HI-mouse models, isolation and culture of islets from mice, rats, islet perifusion, enzyme kinetics, isolation of ?-cells, multiplex immunoassays, CAP-certified insulin assays, EBV transformed lymphocytes. mutation detection.
I am a Pediatric Endocrinologist with a long-standing interest in translational and basic research related to disorders of insulin secretion in infants and children. My main interests have been centered on investigations into the causes of hypoglycemia in infants and children. Hypoglycemia in infants and children can lead to seizures, developmental delay, and permanent brain damage. Over the years, our work has shown that Hyperinsulinism (HI) is the most common cause of both transient and permanent disorders of hypoglycemia. HI is characterized by dysregulated insulin secretion, which results in persistent mild to severe hypoglycemia. Hyperinsulinism can also be associated with perinatal stress such as birth asphyxia, maternal toxemia, prematurity, or intrauterine growth retardation, resulting in prolonged neonatal hypoglycemia. The various forms of HI represent a group of clinically, genetically, and morphologically heterogeneous disorders.
Early in my research career, I described methods for diagnosis and treatment of infants with hyperinsulinism and was actively involved in the discovery of many of the known genetic disorders of ketogenesis and fatty acid oxidation. Over the past 15 years, I have contributed to the identification of the four major genetic loci for hyperinsulinism, GLUD-1, GCK, SUR-1, and KIR6.2. More recently, I have been characterizing defects in other genes that lead to HI, including SCHAD and MCT1. SUR-1 and Kir6.2 combine to form the ?-cell plasma membrane KATP channel. The channel is a heterooctameric complex comprising 4 Kir6.2 subunits which form the ion pore, coupled to 4 SUR-1 regulatory subunits. Inactivating mutations in the KATP channel result in constitutive closure of the channel allowing membrane depolarization and calcium influx into the ?-cell, resulting in constitutive insulin secretion from the ?-cell. These mutations cause KATP channel hyperinsulinism (KATP-HI), the most common and severe form of HI. More than 100 mutations of SUR-1 and 20 mutations of KIR6.2 have been found. Most of the mutations are recessive, but a few dominantly expressed mutations have been reported. There are two distinct histological forms of KATPHI, diffuse HI and focal HI. The preoperative differentiation of these two forms is very important because the surgical management is radically different. The focal form of the disease can be cured if the focal lesion can be localized accurately and completely resected with surgery. One of my ongoing research interests has been to assess the reliability of [18F]DOPA PET scans in its accuracy in localizing focal lesions to aid in their surgical excision.
My lab discovered glutamate dehydrogenase HI (GLUD-1), the second most common form of HI. It is also known as the hyperinsulinism and hyperammonemia (HI/HA) syndrome. It is caused by activating mutations in GDH, a mitochondrial enzyme (9 ), and a key regulator of amino acid and ammonia metabolism in ?-cells, liver, and brain. A mutation in HADH, the gene encoding the mitochondrial enzyme SCHAD, is also associated with HI. SCHAD catalyzes the third of 4 steps in the mitochondrial fatty acid oxidation pathway by catalyzing the oxidation of short-chain substrates. My laboratory has recently shown that dysregulation of insulin secretion associated with SCHAD deficiency is due to an activation of GDH enzyme activity, reflecting the loss of an inhibitory protein-protein interaction of SCHAD upon GDH, which has not previously been recognized.
More recently, my laboratory has begun to focus on HI in children that involves both novel molecular defects of known loci, as well as, previously unrecognized new genetic loci. Using linkage analysis and NexGen sequencing techniques, we have characterized a family with HI that formed the basis for the identification of idiopathic hypoglycemia of infancy in 1954. We have found no mutations in the known HI loci, however, our analysis has identified a novel 8.2Mb locus in chromosome 10 containing 49 genes. Gene capture and next-gen sequencing techniques are being used to identify disease-causing mutations in this region.
I also serve as the Medical Director of the Translational Core Laboratory (TCL), which is part of the Clinical and Translational Research Center (formerly the General Clinical Research Center) here at the Childrenís Hospital of Philadelphia (CHOP). The Clinical and Translational Research Center is part of a integrated strategy to support clinical and translational research and education by the University of Pennsylvania, (CHOP), the Wistar Institute, and University of the Sciences in Philadelphia. The functions of the CHOP-TCL are grouped into four sub-cores: (1) Specimen Collection, Processing and Shipping Core, (2) Biochemistry Core, (3) DNA Isolation-Cell Culture Core, and (4) Molecular Biology-Genetics Core. The TCL provides CTRC investigators with access to a wide range of unique assays and services for patient-oriented research. This includes help with protocol design, specimen collection, processing, tracking, and storage, immunoassays (single and multi-plex), DNA isolation and re-sequencing, cell culture services, PCR-based assays (SNP, real-time PCR gene expression), and mutation detection. The CHOP TCL is focused on performing these clinical research-quality assays in a cost effective manner.
This research bridges the gap between clinical research and patient care. Participants for studies are often recruited from Endocrine clinic. Also, we work with many different parts of the CTRC, including the Vascular Core for noninvasive imaging studies, the Bionutrition Core, and the nutritionists.
Dr. Sheela N. Magge pursues patient-oriented research in the fields of pediatric obesity, insulin resistance, dyslipidemia, and type 2 diabetes. With the increased prevalence of obesity among children and adolescents, physicians are seeing more insulin resistance, dyslipidemia, metabolic syndrome, and type 2 diabetes in children. During her subspecialty training, Dr. Magge obtained a Masters in Clinical Epidemiology and Biostatistics from the UPenn School of Medicine, in order to gain formal training in clinical research methodology. Her Masterís thesis work resulted in a publication demonstrating that obese siblings of children diagnosed with type 2 diabetes had a prevalence of abnormal glucose tolerance of 40%, compared to only 14% in obese children without a sibling with diabetes. She has also been involved in studies of the metabolic syndrome in adolescents, and in the natural history of type 2 diabetes in children.
As a consequence of T2DM developing during childhood (instead of almost exclusively during adulthood), comorbidities of T2DM such as cardiovascular disease (CVD) may present decades earlier in these children, than in previous generations. This will present a tremendous human toll, and could result in a public health crisis. It is important to delineate which children are at highest CVD risk, in order to guide screening, and target lifestyle and/or pharmacologic interventions. Dr. Magge was awarded a K23 Career Development Award from the NIH to study dyslipidemia and CVD risk factors in pediatric obesity and type 2 diabetes, and this study is currently underway. The study compares metabolic dyslipidemia and cardiovascular risk factors in four groups of adolescents: lean controls, obese insulin-sensitive, obese insulin-resistant, and obese with type 2 diabetes. Measured outcomes include carotid intima-media thickness, brachial reactivity, insulin resistance measured through the frequently sampled intravenous glucose tolerance test, adipocytokines such as adiponectin, and lipoprotein subclass analysis. The study is being conducted through the Upenn Clinical and Translational Research Center (CTRC). By identifying which children are at the greatest CVD risk, Dr. Magge would like to then pursue a prospective intervention trial in at risk adolescents, to help prevent or decrease longterm cardiovascular morbidity.
Dr. Magge is also a co-PI of a pilot study looking at CVD risk factors in obese, vitamin D deficient, African American adolescents before and after vitamin D3 treatment. Lower 25-OH vitamin D levels have been associated with obesity and insulin resistance, as well as increased CVD mortality in adults. This pilot study in adolescents will be used to gather preliminary data for a future randomized control trial in this population.
Finally, Dr. Magge is also interested in genetic syndromes of insulin resistance. She has collaborated with investigators in England to learn more about this spectrum of diseases, and how to care for patients with these rare disorders.
De Leon Lab
Mouse model of KATP hyperinsulinism, isolation and functional evaluation of mouse and human pancreatic islets, immunoassays. Clinical and laboratory protocols for the evaluation of the entero-insular axis in mice and humans.
I am a pediatric endocrinologist with an interest in congenital and acquired forms of hyperinsulinemic hypoglycemia. I am particularly interested in the role that alterations of the entero-insular axis may play in the pathophysiology of these disorders. The importance of the entero-insular axis on the regulation of glucose metabolism is well established. Impairments on the axis, resulting in decreased incretin response, have been linked to the pathophysiology of type 2 diabetes. Thus, agents that target the entero-insular axis provide a new therapeutic approach to treat diabetes. Less is known about the potential role of incretin hormones, particularly GLP-1, in disorders of insulin regulation resulting in hypoglycemia.
My translational research program focuses on examining the role of the insulinotropic hormone glucagon-like peptide-1 in the pathophysiology of congenital and acquired forms of hyperinsulinemic hypoglycemia. Hyperinsulinemic hypoglycemia is the most common cause of hypoglycemia in children and adults. Most commonly in children, hyperinsulinemic hypoglycemia is the result of genetic defects affecting regulation of insulin secretion, particularly KATP channel mutations, also known as KATP hyperinsulinism (KATPHI). Surgical procedures affecting nutrient delivery to the gastrointestinal tract, particularly Nissen fundoplication and gastric bypass surgery, can also result in hyperinsulinemic hypoglycemia. We have estimated that approximately 24% of children undergoing a Nissen fundoplication develop hypoglycemia, which is frequently unrecognized.
Using an animal model of KATP hyperinsulinsm (SUR1-/- mouse) we have shown that antagonism of the GLP-1 receptor suppresses the dysregulated insulin secretion and corrects fasting hypoglycemia. These findings suggest that GLP-1 and its receptor may play a role in the pathophysiology of KATPHI and that the GLP-1 receptor may be a viable therapeutic target for this disease.
In children with hyperinsulinemic hypoglycemia (late dumping syndrome) after Nissen fundoplication, we have shown that endogenous secretion of GLP-1 in response to a meal is significantly increased compared to normal children. We postulate that this increased secretion of GLP-1 is responsible for the exaggerated insulin response and subsequent hypoglycemia observed.
Currently, we are conducting proof-of-concept studies to examine the effects of the GLP-1 receptor antagonist, exendin-(9-39), on glucose metabolism and pancreatic islet function in children with hyperinsulinemic hypoglycemia. In addition, we are conducting studies in mouse and human islets to understand the mechanisms by which exendin-(9-39) inhibits KATP-independent insulin secretion. Please visit www.clinicaltrials.gov for a complete list of our clinical trials.
The aim of my research program is to improve our understanding of mechanisms responsible for hyperinsulinemic hypoglycemia with the goal of developing effective and innovative therapies for this condition and to further our understanding of the pathophysiologic interactions between the gut and the pancreatic islet.
Dr. Lipman's major areas of research have been the epidemiology of diabetes and assessment of growth of children. She has maintained the Philadelphia Pediatric Diabetes Registry since 1990. It is longest ongoing pediatric diabetes registry in the US. She was the principal investigator of a multicenter randomized, controlled trail evaluating growth assessment in 900 children in primary care practices in 8 US cities. Dr. Lipman's current research includes racial disparities in children with endocrine disorders, the effect of the economic insecurity on health and food choices in adolescents, and abdominal adiposity as a predictor of metabolic and cardiovascular risk factors. Dr. Lipman also focuses on community based participatory research, collaborating with an urban high school to screen diabetes risk factors in children in the community.
Expertise in the molecular biology of the growth hormone (GH)/insulin-like growth factor (IGF) system. Expertise in molecular biology and clinical research.
Trained as a translational physician-scientist, my primary research focuses on the growth hormone (GH)/insulin-like growth factor (IGF)-I axis and clinical issues related to child growth. I was appointed Scientific Director of the Diagnostic and Research Growth Center at CHOP, where approximately 80 new growth patients are referred for evaluation per month and approximately 2000 children per year are followed for growth faltering. I was also appointed Chair of the taskforce charged with drafting the new guidelines for rhGH and rhIGF-I use in children and adolescents for the Pediatric Endocrine Society.
I was trained, with support from a KO8 award from NIDDK, in the molecular biology of the GH/IGF axis and how it interacts with the p53 tumor suppressor pathway in cancer development. I have acquired expertise in the modulation of IGF bioactivity in the context of intersecting cell signaling pathways that together, determine cell fate. For example, I discovered that IGF binding protein-2 is a novel target of p53 and more recently, in collaboration, that IGF-I enriches colon cancer stem cells. On-going collaborative work focuses on the regulation of IGF secretion. While pursuing my molecular biology research, I was inspired by patients I had seen in clinic to embark on studies investigating clinical issues related to growth, such as gender and racial disparities in the evaluation and management of children with short stature, optimizing the electronic health records (EHR) system as a tool for evaluating growth in the primary care setting, and growth problems associated with specific diseases like Alagille syndrome. My research is funded by NIH and foundation grants.
Dr. Kellyís patient-oriented research interests focus on endocrine co-morbidities in chronic childhood disease with a special interest in cystic fibrosis. I work closely with the Childrenís Hospital of Philadelphia Cystic Fibrosis Center and Hospital of University of Pennsylvania Adult Cystic Fibrosis Center to study cystic fibrosis related diabetes (CFRD). This work aims 1) to understand the link between hyperglycemia and worse nutritional status and pulmonary health, 2) the mechanisms responsible for development of insulin secretion and glucose abnormalities in cystic fibrosis, and 3) CFRD screening.
Dr. Kelly also works with the Division of Gastroenterology and Nutrition to study body composition and bone health in cystic fibrosis using dual x-ray absorptiometry and quantitative computerized tomography measures. Bone deficits in a relatively healthy population of children and adolescents with CF are largely explained by short stature and decreases in lean body mass. Additional work strives to understand the impact of intramuscular fat on these relationships.
Work funded by my K-23 revolved around the development of insulin resistance and cardiometabolic risk in children with obstructive sleep apnea. This work also identified a relationship between vitamin D and insulin resistance. Thus, bridging this work and her interest in body composition, Dr. Kelly is also studying the impact of vitamin D supplementation upon vitamin D levels, muscle strength, and cardiometabolic risk in obese African-American adolescents with vitamin D deficiency.
During my fellowship and early training, I performed basic and clinical research in the field of congenital hyperinsulinism. This research primarily focused on amino acid stimulated insulin secretion and the role of the enzyme glutamate dehydrogenase (GDH) in the second most common form of congenital hyperinsulinism, the hyperinsulinism-hyperammonemia syndrome. This research interest remains, and I continue to study the central nervous system manifestations of GDH-hyperinsulinism.
Steven M. Willi, M.D.
Dr. Williís major research focus is in the areas of diabetes and obesity, including intensive management of, and novel therapies for, type 1 diabetes. Notable previous efforts include the use of a very low calorie diet (VLCD) to treat morbid obesity and T2DM in adolescence and optimal insulin pump therapy in type 1 diabetes. He currently serves on the Safety and Monitoring Committee for a multicenter, national pediatric type 2 diabetes treatment trial (STOPP-T2Dís TODAY study), and was on the Publications and Presentations Committee for a multicenter, national pediatric type 2 diabetes prevention trial (STOPP-T2Dís "Healthy" study). He also serves as chair of the Data Safety Monitoring Board for the ďTake ControlĒ Study which is designed to improve diabetes outcomes in children with type 1 diabetes.
Dr. Willi's ongoing research trials include three that are designed to delay the progression of type 1 diabetes in new onset patients. He is also involved in two trials utilizing new developments in type 1 diabetes therapy; including studies utilizing continuous glucose sensors, and novel insulin products. His previous work with the sensor augmented insulin pump has recently been published in the New England Journal of Medicine and Diabetes Care. This technology represents a tangible step toward an ďartificial pancreasĒ which has the potential to revolutionize diabetes care in the coming decade.
During his tenure as Director of the Diabetes Center for Children, Dr. Willi has spearheaded the development of an ADA certified Diabetes Education "Center of Excellence" at the Children's Hospital of Philadelphia. He has furthermore initiated several evidence-based practice initiatives within the Center. This effort has led to the formation of numerous teams of physicians, nurses, nutritionists and social workers. These teams collaborate to develop practice guidelines, which are then assessed through clinical data analysis and enhanced to improve outcomes.
Fellows involved in Dr. Williís research group will be able to become involved in patient oriented research projects exploring new approaches to treatment of type 1 and type 2 diabetes. Dr. Williís group is currently conducting over 10 clinical trials which include several examining the safety and efficacy of therapies to preserve beta cell function in type 1 diabetes, 2 phase 1 PK/PD trails using novel type 2 diabetes therapies, the examination of blood glucose profiles in preschool age children with type 1 diabetes using glucose sensor technology, etc. His group is also participating in a multicenter type 1 diabetes consortium which will allow the examination of research questions through analysis of prospectively collected clinical and biochemical data. Dr. Willís participation in this consortium affords the fellow an opportunity to frame unique research questions and propose methods for data collection to be conducted at CHOP, or even disseminated across a number of collaborating pediatric and adult diabetes centers.
Recognition of Dr. Willi's clinical expertise is evident from his inclusion in "Best Doctors in America", Marquis Who's Who and the Consumer Research Council of America's Guide to America's Top Pediatrician's. He has been recognized as a clinical researcher by being chosen for three NIH/NIDDK Special Emphasis Panels as well as a Special Advisory Panel to the United States Office of Human Research Protection (OHRP).
Dr. Michael A. Levine
Our laboratory investigates the molecular pathophysiology of genetic disorders of bone and mineral metabolism. Our approach has been translational, and is based on research questions that address clinical disorders that affect the growing skeleton. Our studies focus on three areas of investigation:
1. Elucidation of the pre- and post-natal roles of the transcription factor Gcm2, an essential molecular switch that acts during vertebrate development to control very early differentiation steps in the initiation of parathyroid morphogenesis. The parathyroid glands secrete parathyroid hormone (PTH), the principle hormone that regulates serum levels of calcium and phosphate. Our laboratory has identified mutations in the GCM2 gene that account for hypoparathyroidism, a condition in which children fail to develop parathyroid glands. We are currently using both transgenic and cellular approaches to identify the molecular and protein targets of Gcm2 action. Our current model is a mouse line that we have created in which we can disrupt Gcm2 alleles in a time and spatially controlled manner.
2. A second area of investigation focuses on the study of the GTP-binding proteins that couple receptors for PTH and other hormones to stimulation of signal effector proteins. We have identified and characterized molecular defects in the GNAS gene, an imprinted gene that encodes the alpha chain of the GTP-binding protein Gs (Gs ), in patients with two metabolic syndromes, Albright hereditary osteodystrophy (AHO) and McCune-Albright syndrome (MAS). Germline GNAS mutations that impair expression or function of Gs protein are present in most patients with AHO, and account for generalized hormone resistance in these patients. In addition, other mutations that are cis to GNAS disrupt imprinting of Gs? and impair expression of the maternal GNAS allele in imprinted tissues. By contrast, somatic cell gene mutations that activate Gs are present in subjects with MAS. The activated Gs protein leads to constitutive signaling and accounts for the autonomous proliferation and excess function of endocrine, skin, and bone tissues. Based on human studies and experiments in transgenic mice that we have developed, we have learned that obesity in patients (and mice) with mutations in GNAS reflects disordered hormonal signaling in the brain. Current work is focused on determining the basis for obesity in these patients through a combination of both animal and human studies.
3. The third focus of our research is the elucidation of novel genetic mechanisms that regulate vitamin D metabolism. Nutritional rickets most commonly results from vitamin D deficiency and is typically associated with inadequate sunlight. Thus, the existence of rickets in many tropical countries, where sunlight is abundant, is unexpected, and has suggested that genetic, hormonal, and other nutritional factors may cause rickets in susceptible children. We previously identified CYP21 as the enzyme required for the first step of vitamin D activation. We have now discovered missense mutations in the CYP2R1 genes of affected members of two large Nigerian families with inherited rickets. To assess the functional consequences of these mutations, we administered 50,000 IU of ergocalciferol (D2) or cholecalciferol (D3) on separate occasions to normal and affected subjects and determined serum levels of 25(OH)D2 and 25(OH)D3 by isotope-dilution liquid chromatography-tandem mass spectrometry. Homozygous and heterozygous patients with rickets showed approximately 90% and 50% less 25(OH)D production, respectively, than normal subjects. These studies provide evidence of a new form of vitamin D dependent rickets that appears to be inherited in a semidominant manner. Moreover, our results suggest that genetic polymorphisms in CYP2R1 might provide a novel explanation for variation in the vitamin D requirements of different children and adults.