In infants and children, many of the diseases that affect the liver and bile ducts, such as cysts of the biliary tree and congenital hepatic fibrosis, represent true birth defects. Other diseases are caused by immaturity of hepatic structure and function, which influences how the liver reacts to injury from viruses and drugs. Knowledge of the biochemical and molecular events that affect liver development is critical to understanding how developmental defects of the liver and bile ducts occur and why infants are more susceptible to the development of cholestasis in association with bacterial infection, intravenous feeding, or malfunction of other organs, such as the heart. Injury to the liver during critical periods of development may adversely affect its growth and capacity to perform vital functions, such as processing nutrients, providing energy, and excreting wastes through biliary secretion.
AREAS OF INTEREST
- Much recent information indicates that hepatocytes and cholangiocytes may derive from a common precursor, or stem cell. Specific genes expressed in each cell type eventually produce the final differentiated cell (1,2). The mechanisms by which these specific genes are activated or suppressed have been defined only partially. Understanding differential gene expression in these two types of epithelial cells is critical to understanding how the liver and biliary tree develop and how certain functions, particularly related to bile formation, are restricted either to hepatocytes or to cholangiocytes.
- A number of disorders unique to childhood, such as intrahepatic bile duct paucity and biliary atresia, result in progressive injury and destruction of the intrahepatic and extrahepatic bile ducts. To understand how these disorders affect the liver in infants and children, it will be essential to define the normal and abnormal development of the intrahepatic and extrahepatic bile duct systems (3).
- The intrahepatic bile ducts derive from an initial primitive streak of endodermal cells destined to become the liver. During early fetal development, these growing cells express genes that direct their development into cells found exclusively in the liver (hepatocytes) or the bile ducts (cholangiocytes). The factors that cause this differentiation have not been defined completely, but this knowledge is essential to understanding a number of pediatric disorders, such as bile duct paucity, some cases of biliary atresia, and cystic diseases of the biliary tree.
- The development of the extrahepatic bile ducts and the gallbladder is even more poorly understood. These structures may grow out of an adjacent portion of the intestine. During early development of the human embryo, the primitive gallbladder and common bile ducts consist of a solid cord of cells without an open channel. The process that controls remodeling of these structures to form open channels has not been defined. Some forms of biliary atresia, particularly those that occur in association with birth defects of the intestine and other organs, probably relate to this developmental process. Basic research must focus on the developmental biology of these structures.
- The healthy infant undergoes a period of physiologic cholestasis caused by immature pathways for the formation of bile. This immaturity of liver function may make the infant more susceptible to cholestatic liver disease during episodes of infection or during administration of drugs or parenteral nutrition. Understanding has progressed considerably of the transport mechanisms that contribute to bile formation at the level of the hepatocyte and cholangiocyte (4). However, much remains to be discovered about how the specific transporters develop and are affected by disease in the infant.
- Recent studies of cholestatic disorders in children have demonstrated inherited defects in a number of transport mechanisms located at the canalicular membrane (5,6). Acquired dysfunction of these transporters probably occurs in other liver diseases, and these abnormalities may be associated with significant morbidity and even mortality. The mechanism by which these inherited defects progress to liver injury is understood only partially. Understanding this mechanism potentially will provide therapeutic interventions to decrease injury caused by cholestasis.
- The intrahepatic and extrahepatic bile ducts are frequent sites of involvement in many inherited and acquired liver diseases of childhood. Recent progress has been made in our ability to study the functional properties of cholangiocytes in the adult liver (7). However, little understanding exists of how these cells function during development and how they contribute to bile formation (8). We know little about the differences between the genes that are expressed in cholangiocytes of the developing liver and those of the adult liver.
- The role of the liver in fetal growth and development has been partially clarified. The liver is a key source of amino acids necessary for fetal growth. Transport of nutrients across the hepatocyte is a key regulatory step. Clarifying the role of the unique developmental aspects of hepatic metabolism at the organ, zonal, and cellular level in healthy and abnormal fetal and neonatal growth and development is a critical area of study that may affect the care of preterm and newborn infants.
- To define the array of genes that are activated or suppressed during development to produce fully differentiated hepatocytes and cholangiocytes
- To understand the molecular mechanisms underlying the structural development of the liver and biliary tract and their relationship to birth defects, such as some forms of biliary atresia and fibrocystic diseases of the liver
- To determine the normal development of liver transport mechanisms that contribute to bile formation and how cholestasis affects transporters of organic and inorganic solutes
- To define the functional properties of cholangiocytes and their contribution to bile formation during early life
- To define the regulation of developmental changes in fetal hepatic amino acid, carbohydrate, and lipid metabolism
- To define the array of genes activated or suppressed during development to produce fully differentiated hepatocytes and cholangiocytes: The fully differentiated hepatocyte and cholangiocyte are derived from a common precursor stem cell. An array of genes, many of which are transcription factors, are activated or suppressed during development to produce the mature cell. This process is poorly understood and requires intensive study. Some important genes may be identified through differential screening methods and further tested by targeted inactivation (knockout). For example, recent research indicated that when a gene for a growth factor, transcription factor, or signaling molecule was inactivated in the mouse, abnormal liver development occurred, or the liver failed to develop at all. It was often unclear how these molecules influenced liver development.
- To understand the molecular mechanisms that underlie structural development of the liver and biliary tract and their association to birth defects, such as some forms of biliary atresia and fibrocystic diseases of the liver: The liver in the embryo derives from a primitive streak of endodermal cells in the foregut that form hepatocytes and cholangiocytes. The molecular mechanisms underlying formation of the liver are understood only partially. In particular, the factors leading to the formation and remodeling of the so-called ductal plate to produce the intrahepatic biliary tree must be defined, including the genes that determine cell fate, proliferation, and apoptosis (programmed cell death). Although the gene responsible for autosomal dominant polycystic kidney disease is expressed in the liver and probably contributes to cystic abnormalities of the biliary tree, the role of this gene and its protein product polycystin in the morphogenesis of bile ducts has not been defined. It is also unknown how the defective gene in Alagille syndrome, the JAG1 gene, leads to the loss of interlobular bile ducts and to the small size of the extrahepatic bile ducts.
- To determine the normal development of liver transport mechanisms that contribute to bile formation and how transporters of organic and inorganic solutes are affected by cholestasis: Cholestasis occurs more frequently and earlier in the course of liver disease in infancy than at any other time of life. It has recently been established that inherited defects in several adenosine triphosphate–dependent transporters localized to the bile canalicular membrane, including transporters for bile acids and phospholipids, lead to progressive cholestatic liver disease. Little is known about how these transport mechanisms develop and are regulated in the fetus and neonate. Moreover, research must define how the function of these transporters is altered in acquired cholestasis as a result of bile duct obstruction and intrahepatic disease. It is feasible to study the properties of these transport systems in animal models as well as in human models. Overexpression of transport proteins in transgenic animals and targeted deletion studies will help define the importance of each system to the process of bile formation and will contribute significantly to our understanding of the pathophysiology of cholestasis.
- To define the functional properties of cholangiocytes and their contribution to bile formation during early life: The bile ducts are a frequent site of injury in pediatric liver disease. However, we lack knowledge about the functional properties of cholangiocytes during development.
- Recent studies have demonstrated the ability to isolate and culture cholangiocytes and small segments of the biliary tree (isolated bile duct units). However, such research has not been performed in developing animals. The functional properties of cholangiocytes must be determined, including the localization and expression of transporters and whether, as in the adult biliary system, functional heterogeneity between large and small cholangiocytes exists. It is also unknown how these cells respond to hormonal agonists such as secretin. Determining the genes that are specifically expressed in developing cholangiocytes, compared with mature cholangiocytes, is essential to understanding the biology of these cells. Strategies to target genes specifically to the biliary tree will be important in developing gene replacement therapies for biliary disorders, such as cystic fibrosis.
- To define the regulation of developmental changes in fetal hepatic amino acid, carbohydrate, and lipid metabolism: The liver is a key site in the coordinated regulation of fetal and neonatal metabolism. Recent studies have demonstrated unique features of fetal and neonatal hepatic glycogen, serine, and glutamine metabolism. However, the underlying regulation of these processes must be defined. The next step in this research will be to determine how abnormalities in this regulation affect fetal development. Determination of the genes that are specifically expressed in the developing liver and that differ from those expressed in the mature liver will be essential to understanding the role of the liver in fetal metabolism. This research also will require the development of models for testing hypotheses generated by cellular work. These models will allow the development of methods (such as stable isotope tracing) for use in human studies.
PROJECTED TIMETABLE AND FUNDING REQUIREMENTS
Investigations into the proposed areas of emphasis will require approximately 2 to 4 additional RO1-type grants in each of the five areas, for a total of 10 to 20 grants. The direct costs for each year would total approximately $1.5 million to $3 million. Program project grants should be encouraged so that basic science departments and research hepatologists could collaborate to study the problems detailed in this report; the direct costs would total approximately $2 million per year. Success in this arena will require a group of well-trained basic investigators from clinical and basic science departments. To this end, three new training grants that focus on pediatric hepatology research and that total approximately $450,000 in direct costs per year should be initiated. Funding an additional digestive disease center that would focus on clinical and basic investigations in pediatric hepatology also would be desirable. The direct cost per year for this center would be approximately $750,000.
HEALTH AND ECONOMIC OUTCOMES
Currently, the charges for the care of children with extrahepatic biliary atresia alone may exceed $200 million per year. The cost of care for patients with more common metabolic liver diseases, such as glycogen storage disease, α-1-antitrypsin deficiency, and Wilson disease, is more than $150 million per year. Moreover, liver transplantation, when a child is fortunate enough to undergo the procedure, costs $200,000 to $300,000 for the first year of care. Lifelong immunosuppression adds to the burden of healthcare costs, particularly considering the prolonged survival of pediatric transplant patients.
Prevention and treatment strategies targeted to this age group probably will be especially cost-effective because when chronic liver disease is prevented or successfully cured in a child, a healthy, productive citizen is returned to the workforce for many decades. However, basic insight into the development of liver structure and function is essential to our efforts to treat many of the hepatic disorders that affect infants and children.
Substantial progress has been made in identifying the unique range of hepatobiliary diseases that affect infants and children and in saving the lives of some patients with end-stage liver disease through liver transplantation. Chronic and progressive liver disease, whether caused by a virus, an inborn error of metabolism, or congenital malformation, may be fatal or may significantly impair normal physical and mental development. Better understanding of liver development and the mechanisms that underlie pediatric hepatobiliary disease is necessary so that effective treatment and strategies for prevention can be devised. Progress will come only from substantial and sustained investment in basic and clinical research. The basis of many treatment and preventive strategies will come from further insight into the development of liver structure and function (1–8).
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