Section 5 -

Terminology Changes Regarding "Ductular Reactions" Neil Theise, M.D. Romil Saxena, M.D.

Case 5A 21 year old woman commits suicide with acetaminophen overdose. Liver tissue obtained at autopsy. Diagnosis: Massive hepatic necrosis secondary to acetaminophen overdose. Case 5B 42 year old woman, complaining of fatigue presents with elevated alkaline phosphatase and anti-mitochondrial antibody positive 1:2560. Liver biopsy for diagnosis and staging Diagnosis: Primary biliary cirrhosis, stage 2/4 (diagnostic biopsy). Case 5C 54 year old man transplanted for hepatitis C cirrhosis. Diagnosis: Chronic hepatitis, mildly active with cirrhosis (modified Ishak stage 4/4), compatible with hepatitis C. Comments excerpted from: "Nomenclature of the Finer Branches of the Biliary Tree: Canals, Ductules and Ductular Reactions in Human Livers" Roskams T, Theise ND, et al. Hepatology. 2004 Jun;39(6):1739-45 Structure of Normal Biliary Tree The fine detail of normal liver microanatomy is not well-understood. This is true whether discussing hepatic vasculature, bile ducts, stroma and matrix, innervation, or lymphatics. Some points are known, but gaps remain. The distal branches of the biliary tree are reasonably well defined: the common bile duct arises from confluence of the right and left hepatic ducts, which arise from segmental ducts, which arise from septal ducts arising from interlobular duc. It is known that these interlobular ducts arise from still smaller cholangiocyte-lined structures and that the lumina of these in turn are in structural continuity with the lumen of hepatocellular bile canaliculi. But the terms used for these smallest, most proximal structures have been confusing. The structure that came to be known as the canal of Hering was first incompletely described by Ewald Hering, reporting dye excretion studies identifying a link between the hepatocyte canalicular system and bile ducts in 1867. Within this publication he included a drawing of bile channels that were partly lined by hepatocytes and partly by cholangiocytes. This structure led in turn to a channel completely lined by cholangiocytes. Descriptions and terms for these structures varied widely over the course of the next century and a half. The ductular-hepatocellular junction, lined partly by hepatocytes, partly by cholangiocytes (canal of Hering), apparently varies in length. Recent work using immunohistochemical staining of the biliary tree and three dimensional analyses of serial sections of normal liver definitively established that the canal of Hering often does not stop at the limiting plate of hepatocytes surrounding the portal tract, but, rather, extends through it, linking to the hepatocyte canalicular system within the lobule, usually in the periportal region. In two dimensional sections, immunohistochemical staining for biliary markers reveals cholangiocytes, singly or in small clusters, at variable distances from the portal tract. But, by and large, these cells prove not to be isolated; instead, they often represent cross-sections of these most proximal branches of the biliary tree. Immunophenotyping studies further support this description. The recent interest in these seemingly isolated cells - usually representing cross-sections of the smallest biliary branches - particularly arises from their identification as an intra-organ stem or progenitor cell compartment of the liver. Such identification guarantees that they will be an increasing focus of research in the coming years. It thus behooves us to come to consensus on terminology so that such investigations can be performed, reviewed, and published in an expedient and generally comprehensible fashion. Terminology of Normal Structures The terminology for these small structures has been highly variable and confusing. Here is presented an updated view of these structures and names for the component parts. Well-known terms have been retained to avoid confusion through unnecessary change, but are defined to reflect current understanding of anatomy (Table 1): The canal of Hering is a channel partly lined by hepatocytes and partly by cholangiocytes. It represents the anatomic and physiological link between the intralobular canalicular system and the biliary tree. Cells of morphology and immunophenotype intermediate between hepatocytes and cholangiocytes ("intermediate cells", see below) are not recognized in normal tissue. A corollary is that the true interface of hepatocytes and biliary tree does not, as has been assumed, at the "limiting plate", but rather along an array of sites that project star-like from the portal tracts, along the canals of Hering. The canal of Hering continues into a channel lined entirely by cholangiocytes, which is termed the ductule. Ductules may or may not traverse the limiting plate, and thus may have an intralobular segment in addition to their intra-portal course. The ductules in turn link to the smallest interlobular bile ducts. Both these structures may branch at any point along their continuum, though their names still depend on whether they are partially or completely lined by cholangiocytes. It should be recognized that the cholangiocytes are a heterogeneous group of cells. They perform a variety of functional and physiological roles: they are a conduction system for bile; they process and modify bile as it flows by, they serve as an intra-organ facultative progenitor cell compartment, and they may be involved in maintenance of matrix as well as in fibrogenesis. They may well have other functions not yet identified. It remains unclear how many of these functions can be performed simultaneously. When specialized studies (e.g. immunohistochemical staining or electron microscopy) identify isolated cells in sections of typical thickness (<6 microns), most of these structures will be sampled in cross section. These cross sections will have the appearance of individual cells, small cell clusters, or "cuboidal strings." Such cells may be referred to by the markers which highlight them (usually biliary-type markers or others, such as NCAM-1/CD56) and/or by reference to the predominant activity being studied (e.g. "cholangiocytes", "progenitor cells"). The diagnostic pathologist and the investigator should maintain keen awareness of two points: 1) that the appearance as isolated cells is usually an artifact of sectioning and visualization in a near two-dimensional plane; 2) that these cells are often engaged in multiple physiological tasks, not simply the one under investigation in a given study. Terminology of Reactive Lesions In disease states, a commonly used term for the expanded population of epithelial cells at the interface of the biliary tree and the hepatocytes is "ductular proliferation." This pair of words is problematic because the reactive lesions may not simply arise from proliferation of pre-existing bile ductular cells, as they may also originate from activated and differentiated progenitor cells, they may originate from cells which entered from the circulation and differentiate towards liver cells, or, more rarely, arise from biliary metaplasia of hepatocytes. The term ductular reaction was coined by Popper et al in 1957 and has subsequently been used in the hepatology literature. We prefer this to "ductular proliferation" for the reasons given in the preceding paragraph. "Ductular reaction" implies a reaction of ductular phenotype, possibly but not necessarily of ductular origin This is in keeping with current practice in oncologic pathology, where the names of neoplasms reflect phenotype, as the supposed cell of origin is not always known. As noted, the epithelial component of the ductular reaction may actually derive from several sources: not only from the proximal branches of the biliary tree, but also from the circulation (often if not always from bone marrow), and from biliary metaplasia of hepatocytes. "Reaction" encompasses the complex of stroma, inflammatory cells, and other structures of diverse systems all of which participate in the reactive lesion. It was in the epithelial components of the ductular reaction that investigators first began to see features that suggested a correspondence with the "oval cells" of rodent models of carcinogenesis and stem/progenitor lineages. Thus, investigators studying human tissues have often been tempted or required to use corresponding terminology: oval cells or, worse yet, "oval-like cells." While similarities exist between the progenitor cell compartments of human and rodent livers, the different rodent models are not entirely comparable with the human situation, and use of the same term has created confusion as to what characteristics may be expected in the human ductular reaction. Therefore, it is suggested that "oval cell" and "oval-like cell" be no longer used in descriptions of human tissue. Currently, the progenitor functioning of the ductular reaction attracts much attention. In particular, cells of intermediate morphology and intermediate immunophenotyping are of interest. It is suggested that these cells be referred to as just that: intermediate cells, defined as larger than 6 microns in diameter (the approximate size of the normal canal of Hering cell, i.e. the smallest cholangiocytes), but less than 40 microns (the typical size of an hepatocyte), with other features suggesting dual characteristics of both hepatocytes and cholangiocytes. These include, but are not limited to: simultaneous expression of biliary antigens (e.g. cytokeratins 19, 7, OV-6) and hepatocyte antigens (e.g. HepPar1, albumin, alpha-1-antitrypsin, biliary glycoprotein-1 detected by canalicular staining with polyclonal anti-CEA, and, occasionally, alpha-fetoprotein), other markers such as NCAM-1/CD56, and structural features such as basement membrane formation typical of cholangiocytes and canalicular membranes typical of hepatocytes. If the context of usage does not make clear that the term "intermediate cell" refers to hepatobiliary cells, as opposed to hepatic stellate cells or other cell populations, then one may use the phrase "intermediate hepatobiliary cells" for increased clarity. This may particularly be useful in titles or figure legends where context is absent. "Typical" and "atypical ductular proliferation" are problematic terms that perhaps are best avoided. These terms arose from attempts to histologically differentiate between extrahepatic and intrahepatic cholestasis in man. Typical ductules allegedly have a recognizable lumen lined by cuboidal cells, whereas atypical ductules are described as thin, elongated structures that extend irregularly into the lobules, are lined by flattened cells and lack easily discernible lumina. However, distinction between them is not easy, as emphasized already many years ago. Furthermore, the advent of better imaging of the liver and biliary tree in the 1970's decreased the need for histological differentiation between intra-and extrahepatic cholestasis, whereas early endoscopic removal of impacted gall stones and stenting of obstructed bile ducts have rendered the histological observation of complete extrahepatic obstruction in man an uncommon occurrence. On the other hand, in incomplete extrahepatic obstruction and in vanishing bile duct diseases, the histological appearance is complicated by both hepatocellular and cholangiocytic damage, resulting in not only biliary obstructive lesions but also aspects of regeneration of cells which differentiate towards the most damaged cell type. Since the term 'atypical' is further burdened by the (pre)-malignant connotation of this term in diagnostic histopathology in general, it is suggested that these terms are best avoided, awaiting more reliable markers of differentiation between reactions of ductules to obstructive and non-obstructive triggers. The ductular reaction is not to be confused with ductal plate malformations. Ductal plate malformations are abnormal bile duct-like structures: malformations, not reactive lesions. The ductal plate malformation is observed in fibro-cystic liver diseases and in a subgroup of patients with extrahepatic bile duct atresia. Table 1: Suggested terms, with definitions, and terms to be discontinued   Suggested terminology Definition Discouraged Terms Normal Liver Canal of Hering Physiologic link between hepatocyte canaliculi and the biliary tree. Partially lined by hepatocytes and by cholangiocytes (not by cells of intermediate morphology, which are not identified in normal livers).     Bile Ductule Link between canals of Hering and the interlobular bile ducts. Lined entirely by cholangiocytes, may begin at the edge of portal tract stroma, or may traverse the limiting plate, in which case it will have an "intralobular" as well as an "intraportal" segment.     "Isolated" cholangiocytes or progenitor cells in 2-dimensional tissue sections. These cells are cross sections of canals of Hering and intralobular bile ductules and are not all, therefore, truly isolated. They may be referred to by the immunomarkers used to define them (e.g. CK19+, CK7+, NCAM+) and by the function which is under investigation (e.g. "cholangiocytes", "progenitor cells"), always recognizing, however that they have multiple functions. Oval cells Oval-like cells Stem cells Diseased Liver Ductular Reaction A reaction of ductular phenotype, possibly but not necessarily of ductular origin, in acute and chronic liver disease. May arise from: 1) proliferation of pre-existing cholangiocytes; 2) progenitor cells (local and/or circulating cells probably bone marrow-derived); 3) rarely, biliary metaplasia of hepatocytes. Ductular proliferation, "typical" or "atypical" Biliary piecemeal necrosis Oval cells Oval-like cells   Intermediate Cells or Intermediate Hepatobiliary Cells Cells in ductular reactions that are morphologically or phenotypically intermediate between hepatocytes and cholangiocytes: > 6 microns, <40 microns, dual phenotyping by immunostaining or by electron microscopy. NOTE: If context does not make clear that the term refers to parenchymal cells of the liver, such as in titles or figure legends, then "hepatobiliary" may be inserted for clarity. Ductular proliferation Biliary piecemeal necrosis Oval cells Oval-like cells