Background Hepatic progenitor cells (HPCs) hold a great potential for therapeutic intervention fo... more Background Hepatic progenitor cells (HPCs) hold a great potential for therapeutic intervention for currently untreatable liver diseases. However, in human diseases molecular mechanisms involved in proliferation and differentiation of HPCs are poorly understood. Methods and results In the present study activated HPCs and their microenvironment (niche) were investigated in acute and chronic human liver disease by gene-expression analysis and immunohistochemistry/
New therapeutic concepts developed in rodent models should ideally be evaluated in large animal m... more New therapeutic concepts developed in rodent models should ideally be evaluated in large animal models prior to human clinical application. COMMD1-deficiency in dogs leads to hepatic copper accumulation and chronic hepatitis representing a Wilson's disease like phenotype. Detailed understanding of the pathogenesis and time course of this animal model is required to test its feasibility as a large animal model for chronic hepatitis. In addition to mouse models, true longitudinal studies are possible due to the size of these dogs permitting detailed analysis of the sequence of events from initial insult to final cirrhosis. Therefore, liver biopsies were taken each half year from five new born COMMD1-deficient dogs over a period of 42 months. Biopsies were used for H&E, reticulin, and rubeanic acid (copper) staining. Immunohistochemistry was performed on hepatic stellate cell (HSC) activation marker (alpha-smooth muscle actin, α-SMA), proliferation (Ki67), apoptosis (caspase-3), an...
The first aim of this study was to determine whether vitamin D supplementation influenced the eff... more The first aim of this study was to determine whether vitamin D supplementation influenced the effects of high vitamin A intake on new bone formation in adult cats. The second aim was to determine whether high vitamin A intake in cats caused liver pathology and, if so, whether the current upper limit for the dietary intake of vitamin A for healthy adult cats would be safe. Twenty-four healthy adult cats were divided into four groups that received a control diet supplemented with peanut oil (control), or peanut oil containing a 100-fold increase in vitamin A (HA), or a 100-fold increase in vitamin A and a fivefold increase in vitamin D (HAMD), or a 100-fold increase in vitamin A and a 65-fold increase in vitamin D (HAHD) over a period of 18 months. Cats did not show abnormal locomotion or clinical signs of liver failure after 18 months of supplementation but did show subtle skeletal changes and liver pathology, suggesting that the current National Research Council (2006) safe upper limit for vitamin A for cats is too high. The addition of vitamin D did not seem to influence bone pathology. While moderately elevated dietary vitamin D levels (HAMD) seemed to protect cats against the liver pathology caused by the consumption of large amounts of vitamin A, higher dietary levels of vitamin D (HAHD) did not seem to be protective.
Following chronic liver injury or when hepatocyte proliferation is impaired, ductular reactions c... more Following chronic liver injury or when hepatocyte proliferation is impaired, ductular reactions containing hepatic progenitor cells (HPCs) appear in the periportal regions and can regenerate the liver parenchyma. HPCs exist in a niche composed of myofibroblasts, macrophages and laminin matrix. Galectin-3 (Gal-3) is a β-galactoside-binding lectin that binds to laminin and is expressed in injured liver in mice and humans. We examined the role of Gal-3 in HPC activation. HPC activation was studied following dietary induced hepatocellular (choline-deficient ethionine-supplemented diet) and biliary (3,5-diethoxycarbonyl-1,4-dihydrocollidine supplemented diet) injury in wild type and Gal-3(-/-) mice. HPC proliferation was significantly reduced in Gal-3(-/-) mice. Gal-3(-/-) mice failed to form a HPC niche, with reduced laminin formation. HPCs isolated from wild type mice secrete Gal-3 which enhanced adhesion and proliferation of HPCs on laminin in an undifferentiated form. These effects were attenuated in Gal3(-/-) HPCs and in wild type HPCs treated with the Gal-3 inhibitor lactose. Gal-3(-/-) HPCs in vitro showed increased hepatocyte function and prematurely upregulated both biliary and hepatocyte differentiation markers and regulated cell cycle genes leading to arrest in G0/G1. We conclude that Gal-3 is required for the undifferentiated expansion of HPCs in their niche in injured liver.
Journal of Trace Elements in Medicine and Biology, 2015
COMMD1-deficient dogs progressively develop copper-induced chronic hepatitis. Since high copper l... more COMMD1-deficient dogs progressively develop copper-induced chronic hepatitis. Since high copper leads to oxidative damage, we measured copper metabolism and oxidative stress related gene products during development of the disease. Five COMMD1-deficient dogs were studied from 6 months of age over a period of five years. Every 6 months blood was analysed and liver biopsies were taken for routine histological evaluation (grading of hepatitis), rubeanic acid copper staining and quantitative copper analysis. Expression of genes involved in copper metabolism (COX17, CCS, ATOX1, MT1A, CP, ATP7A, ATP7B, ) and oxidative stress (SOD1, catalase, GPX1 ) was measured by qPCR. Due to a sudden death of two animals, the remaining three dogs were treated with d-penicillamine from 43 months of age till the end of the study. Presented data for time points 48, 54, and 60 months was descriptive only. A progressive trend from slight to marked hepatitis was observed at histology, which was clearly preceded by an increase in semi-quantitative copper levels starting at 12 months until 42 months of age. During the progression of hepatitis most gene products measured were transiently increased. Most prominent was the rapid increase in the copper binding gene product MT1A mRNA levels. This was followed by a transient increase in ATP7A and ATP7B mRNA levels. In the sequence of events, copper accumulation induced progressive hepatitis followed by a transient increase in gene products associated with intracellular copper trafficking and temporal activation of anti-oxidative stress mechanisms.
The liver progenitor cell compartment in the normal canine liver and in spontaneous canine acute ... more The liver progenitor cell compartment in the normal canine liver and in spontaneous canine acute (AH) and chronic hepatitis (CH) was morphologically characterised and compared to its human equivalents. Immunohistochemistry was performed for cytokeratin-7 (CK7), human hepatocyte marker (Hep Par 1), multidrug resistance-associated protein-2 (MRP2), and breast cancer resistance protein (BCRP) on paraffin and frozen sections from canine and human tissues. Normal liver showed similar morphology and immunohistochemical reaction of the progenitor cell compartment/canal of Hering in man and dog. In addition, a ductular reaction, comparable in terms of severity, location and immunohistochemical characteristics, was observed in canine and human AH and CH. CK7 was a good marker for canine progenitor cells, including intermediate cells, which were positively identified in cases of AH and CH. In both species, BCRP was expressed in both hepatocytes and bile ducts of the normal liver, and in ductular reaction in AH and CH. MRP2 detected bile canalicular membranes in man and dog. These findings underline the similarities between canine and human liver reaction patterns and may offer mutual advantage for comparative research in human and canine spontaneous liver diseases.
Naturally occurring liver disease in dogs resemble human liver disease in great detail; including... more Naturally occurring liver disease in dogs resemble human liver disease in great detail; including the activation of liver progenitor cells (LPC) in acute and chronic liver disease. The aim of the present study was to isolate, culture, and characterize progenitor cells derived from healthy mature dog livers. A nonparenchymal cell fraction enriched with small hepatocytes was isolated and cultured in Hepatozyme-serum-free media (SFM) to stimulate the growth of colony-forming small epithelial cells. After 2 weeks of culturing, clonal expansion of keratin 7 (K7) immunopositive small cells with a large nucleus/cytoplasm ratio emerged in the hepatocyte monolayer. These colonies expressed genes of several hepatocyte (CYP1A1, ALB, and KRT18), cholangiocyte/LPC (KRT7 and KRT19), and progenitor cell markers (alpha-fetoprotein, CD44, prominin1, KIT, THY1, and neural cell adhesion molecule 1), indicating their immature and bipotential nature. Gene-expression profiles indicated a more pronounced hepatic differentiation in Hepatozyme-SFM compared to William's Medium E (WME). Furthermore, colony-forming cells differentiated toward intermediate hepatocyte-like cells with a more pronounced membranous K7 immunostaining. In conclusion, colony-forming small epithelial cells in long-term canine liver cell cultures express LPC markers and have differentiating capacities. These cells may therefore be considered as progenitor cells of the liver.
When hepatocyte replication during liver disease is insufficient for regeneration, liver progenit... more When hepatocyte replication during liver disease is insufficient for regeneration, liver progenitor cells (LPCs) are activated. The cells and stroma in the immediate environment of LPCs, together termed the LPC niche, are thought to play an important role in this activation. Among these cells are the hepatic stellate cells (HSCs)/myofibroblasts (MFs). We assessed the activation of HSC/MFs and LPCs in relation to the histological location and extent of liver disease in immunohistochemically (double) stained serial sections. Markers of HSC/MFs [alpha-smooth muscle actin, glial fibrillary acidic protein (GFAP), neurotrophin 3 and neural-cell adhesion molecule], markers of LPCs (keratin 7 and keratin 19) and a proliferation marker (Ki67) were used. A very relevant spontaneous model to evaluate LPC niche activation in a translational approach seems to be the dog. Therefore, both human and canine liver diseases with different degree of fibrosis and disease activity were included. In human and canine liver disease, type and extent of LPC niche activation depended on type and severity of disease (P<0.05) and corresponded to the main location of disease. Activated HSCs surrounded the activated LPCs. In chronic hepatitis and non-alcoholic steatohepatitis lobular-type HSCs were activated, while during biliary disease portal/septal MFs were mainly activated. In canine liver, GFAP further presented as an early marker of HSC activation. Activation of the LPCs correlated with disease location and severity (P<0.01), and was inversely related to hepatocyte proliferation, as was previously shown in man. A shared involvement of HSC/MFs, LPCs and disease severity during hepatic disease processes is shown, which is highly similar in man and dog.
American Cocker Spaniels are predisposed to chronic hepatitis. To describe the clinical and histo... more American Cocker Spaniels are predisposed to chronic hepatitis. To describe the clinical and histological features of chronic hepatitis in Japanese American Cocker Spaniels. Thirteen cases examined from 2003 to 2009. Retrospective study. Medical records were searched for American Cocker Spaniels with chronic liver diseases. History, physical examination, clinicopathologic features, hepatic ultrasonographic findings, hepatic histopathology, and immunohistochemistry were evaluated. The median age was 4.6 (1.9-10.7) years. Clinical signs included inappetence (11/13), ascites (11/13), lethargy (9/13), diarrhea (7/13), and melena (2/13). Only 1/13 dogs was jaundiced. Clinicopathological abnormalities were increased liver enzymes (gamma-glutamyl transpeptidase: 9/12, aspartate aminotransferase: 7/10, alanine aminotransferase: 6/13, alkaline phosphatase: 6/13), increased total serum bile acid concentrations (10/12), and hypoalbuminemia (10/13). The liver had an irregular surface in all dogs and acquired portosystemic collaterals were verified in 11/13 dogs by abdominal ultrasound (2), laparoscopy (4), or both (5). Liver histology revealed severe fibrosis and cirrhosis in all cases, subdivided in lobular dissecting hepatitis (7), periportal fibrosis (1), micronodular cirrhosis (3), and macronocular cirrhosis (2). Inflammatory activity was low to mild. Immunohistochemical stains showed ductular proliferation. The median survival time was 913 (range: 63-1981) days. Hepatitis in Japanese American Cocker Spaniels is clinically silent until an advanced stage and is associated with severe hepatic fibrosis leading to cirrhosis, extensive ductular/putative hepatic progenitor cell proliferation, portal hypertension, and acquired portosystemic collateral shunting, but relatively long survival times. Lobular dissecting hepatitis seems more prevalent than in previously reported cases from other countries.
Hepatic progenitor cells (HPCs) hold a great potential for therapeutic intervention for currently... more Hepatic progenitor cells (HPCs) hold a great potential for therapeutic intervention for currently untreatable liver diseases. However, in human diseases molecular mechanisms involved in proliferation and differentiation of HPCs are poorly understood. In the present study activated HPCs and their microenvironment (niche) were investigated in acute and chronic human liver disease by gene-expression analysis and immunohistochemistry/immunofluorescence. Cryopreserved liver tissues were used from patients with parenchymal versus biliary diseases: acute necrotising hepatitis (AH), cirrhosis after hepatitis C infection, and primary biliary cirrhosis in order to study differentiation of HPCs towards hepatocytic versus biliary lineage. Keratin 7 positive HPCs/reactive ductules were captured by means of laser capture microdissection and gene-expression profiles were obtained by using a customized PCR array. Gene expression results were confirmed by immunohistochemistry and immunofluorescence double staining. In all disease groups, microdissected HPCs expressed progenitor cell markers such as KRT7, KRT19, NCAM, ABCG2, LIF, KIT, OCT4, CD44 and TERT. In AH, HPCs were most activated and showed a high expression of prominin-1 (CD133) and alpha-fetoprotein, and a strong activation of the Wnt pathway. In contrast to parenchymal diseases, HPCs in primary biliary cirrhosis (biliary differentiation) showed a high activation of Notch signalling. A distinct pattern of HPC surface markers was found between acute and chronic liver diseases. Similar to what is known from animal experiments, strong evidence has been found signifying the role of Wnt signalling in proliferation of human HPCs whereas Notch signalling is involved in biliary differentiation. These pathways can be targeted in future therapies.
BackgroundThe liver has a large regenerative capacity. Hepatocytes can replicate and regenerate a... more BackgroundThe liver has a large regenerative capacity. Hepatocytes can replicate and regenerate a diseased liver. However, as is the case in severe liver diseases, this replication may become insufficient or exhausted and hepatic progenitor cells (HPCs) can be activated in an attempt to restore liver function. Due to their bi-potent differentiation capacity, these HPCs have great potential for regenerative approaches yet over-activation does pose potential health risks. Therefore the mechanisms leading to activation must be elucidated prior to safe implementation in the veterinary clinic. Wnt/ß-catenin and Notch signalling have been implicated in the activation of HPCs in mouse models and in humans. Here we assessed the involvement in canine HPC activation. Gene-expression profiles were derived from laser microdissected HPCs niches from lobular dissecting hepatitis (LDH) and normal liver tissue, with a focus on Wnt/ß-catenin and Notch signalling. Immunohistochemical and immunofluore...
Background Hepatic progenitor cells (HPCs) hold a great potential for therapeutic intervention fo... more Background Hepatic progenitor cells (HPCs) hold a great potential for therapeutic intervention for currently untreatable liver diseases. However, in human diseases molecular mechanisms involved in proliferation and differentiation of HPCs are poorly understood. Methods and results In the present study activated HPCs and their microenvironment (niche) were investigated in acute and chronic human liver disease by gene-expression analysis and immunohistochemistry/
New therapeutic concepts developed in rodent models should ideally be evaluated in large animal m... more New therapeutic concepts developed in rodent models should ideally be evaluated in large animal models prior to human clinical application. COMMD1-deficiency in dogs leads to hepatic copper accumulation and chronic hepatitis representing a Wilson's disease like phenotype. Detailed understanding of the pathogenesis and time course of this animal model is required to test its feasibility as a large animal model for chronic hepatitis. In addition to mouse models, true longitudinal studies are possible due to the size of these dogs permitting detailed analysis of the sequence of events from initial insult to final cirrhosis. Therefore, liver biopsies were taken each half year from five new born COMMD1-deficient dogs over a period of 42 months. Biopsies were used for H&E, reticulin, and rubeanic acid (copper) staining. Immunohistochemistry was performed on hepatic stellate cell (HSC) activation marker (alpha-smooth muscle actin, α-SMA), proliferation (Ki67), apoptosis (caspase-3), an...
The first aim of this study was to determine whether vitamin D supplementation influenced the eff... more The first aim of this study was to determine whether vitamin D supplementation influenced the effects of high vitamin A intake on new bone formation in adult cats. The second aim was to determine whether high vitamin A intake in cats caused liver pathology and, if so, whether the current upper limit for the dietary intake of vitamin A for healthy adult cats would be safe. Twenty-four healthy adult cats were divided into four groups that received a control diet supplemented with peanut oil (control), or peanut oil containing a 100-fold increase in vitamin A (HA), or a 100-fold increase in vitamin A and a fivefold increase in vitamin D (HAMD), or a 100-fold increase in vitamin A and a 65-fold increase in vitamin D (HAHD) over a period of 18 months. Cats did not show abnormal locomotion or clinical signs of liver failure after 18 months of supplementation but did show subtle skeletal changes and liver pathology, suggesting that the current National Research Council (2006) safe upper limit for vitamin A for cats is too high. The addition of vitamin D did not seem to influence bone pathology. While moderately elevated dietary vitamin D levels (HAMD) seemed to protect cats against the liver pathology caused by the consumption of large amounts of vitamin A, higher dietary levels of vitamin D (HAHD) did not seem to be protective.
Following chronic liver injury or when hepatocyte proliferation is impaired, ductular reactions c... more Following chronic liver injury or when hepatocyte proliferation is impaired, ductular reactions containing hepatic progenitor cells (HPCs) appear in the periportal regions and can regenerate the liver parenchyma. HPCs exist in a niche composed of myofibroblasts, macrophages and laminin matrix. Galectin-3 (Gal-3) is a β-galactoside-binding lectin that binds to laminin and is expressed in injured liver in mice and humans. We examined the role of Gal-3 in HPC activation. HPC activation was studied following dietary induced hepatocellular (choline-deficient ethionine-supplemented diet) and biliary (3,5-diethoxycarbonyl-1,4-dihydrocollidine supplemented diet) injury in wild type and Gal-3(-/-) mice. HPC proliferation was significantly reduced in Gal-3(-/-) mice. Gal-3(-/-) mice failed to form a HPC niche, with reduced laminin formation. HPCs isolated from wild type mice secrete Gal-3 which enhanced adhesion and proliferation of HPCs on laminin in an undifferentiated form. These effects were attenuated in Gal3(-/-) HPCs and in wild type HPCs treated with the Gal-3 inhibitor lactose. Gal-3(-/-) HPCs in vitro showed increased hepatocyte function and prematurely upregulated both biliary and hepatocyte differentiation markers and regulated cell cycle genes leading to arrest in G0/G1. We conclude that Gal-3 is required for the undifferentiated expansion of HPCs in their niche in injured liver.
Journal of Trace Elements in Medicine and Biology, 2015
COMMD1-deficient dogs progressively develop copper-induced chronic hepatitis. Since high copper l... more COMMD1-deficient dogs progressively develop copper-induced chronic hepatitis. Since high copper leads to oxidative damage, we measured copper metabolism and oxidative stress related gene products during development of the disease. Five COMMD1-deficient dogs were studied from 6 months of age over a period of five years. Every 6 months blood was analysed and liver biopsies were taken for routine histological evaluation (grading of hepatitis), rubeanic acid copper staining and quantitative copper analysis. Expression of genes involved in copper metabolism (COX17, CCS, ATOX1, MT1A, CP, ATP7A, ATP7B, ) and oxidative stress (SOD1, catalase, GPX1 ) was measured by qPCR. Due to a sudden death of two animals, the remaining three dogs were treated with d-penicillamine from 43 months of age till the end of the study. Presented data for time points 48, 54, and 60 months was descriptive only. A progressive trend from slight to marked hepatitis was observed at histology, which was clearly preceded by an increase in semi-quantitative copper levels starting at 12 months until 42 months of age. During the progression of hepatitis most gene products measured were transiently increased. Most prominent was the rapid increase in the copper binding gene product MT1A mRNA levels. This was followed by a transient increase in ATP7A and ATP7B mRNA levels. In the sequence of events, copper accumulation induced progressive hepatitis followed by a transient increase in gene products associated with intracellular copper trafficking and temporal activation of anti-oxidative stress mechanisms.
The liver progenitor cell compartment in the normal canine liver and in spontaneous canine acute ... more The liver progenitor cell compartment in the normal canine liver and in spontaneous canine acute (AH) and chronic hepatitis (CH) was morphologically characterised and compared to its human equivalents. Immunohistochemistry was performed for cytokeratin-7 (CK7), human hepatocyte marker (Hep Par 1), multidrug resistance-associated protein-2 (MRP2), and breast cancer resistance protein (BCRP) on paraffin and frozen sections from canine and human tissues. Normal liver showed similar morphology and immunohistochemical reaction of the progenitor cell compartment/canal of Hering in man and dog. In addition, a ductular reaction, comparable in terms of severity, location and immunohistochemical characteristics, was observed in canine and human AH and CH. CK7 was a good marker for canine progenitor cells, including intermediate cells, which were positively identified in cases of AH and CH. In both species, BCRP was expressed in both hepatocytes and bile ducts of the normal liver, and in ductular reaction in AH and CH. MRP2 detected bile canalicular membranes in man and dog. These findings underline the similarities between canine and human liver reaction patterns and may offer mutual advantage for comparative research in human and canine spontaneous liver diseases.
Naturally occurring liver disease in dogs resemble human liver disease in great detail; including... more Naturally occurring liver disease in dogs resemble human liver disease in great detail; including the activation of liver progenitor cells (LPC) in acute and chronic liver disease. The aim of the present study was to isolate, culture, and characterize progenitor cells derived from healthy mature dog livers. A nonparenchymal cell fraction enriched with small hepatocytes was isolated and cultured in Hepatozyme-serum-free media (SFM) to stimulate the growth of colony-forming small epithelial cells. After 2 weeks of culturing, clonal expansion of keratin 7 (K7) immunopositive small cells with a large nucleus/cytoplasm ratio emerged in the hepatocyte monolayer. These colonies expressed genes of several hepatocyte (CYP1A1, ALB, and KRT18), cholangiocyte/LPC (KRT7 and KRT19), and progenitor cell markers (alpha-fetoprotein, CD44, prominin1, KIT, THY1, and neural cell adhesion molecule 1), indicating their immature and bipotential nature. Gene-expression profiles indicated a more pronounced hepatic differentiation in Hepatozyme-SFM compared to William's Medium E (WME). Furthermore, colony-forming cells differentiated toward intermediate hepatocyte-like cells with a more pronounced membranous K7 immunostaining. In conclusion, colony-forming small epithelial cells in long-term canine liver cell cultures express LPC markers and have differentiating capacities. These cells may therefore be considered as progenitor cells of the liver.
When hepatocyte replication during liver disease is insufficient for regeneration, liver progenit... more When hepatocyte replication during liver disease is insufficient for regeneration, liver progenitor cells (LPCs) are activated. The cells and stroma in the immediate environment of LPCs, together termed the LPC niche, are thought to play an important role in this activation. Among these cells are the hepatic stellate cells (HSCs)/myofibroblasts (MFs). We assessed the activation of HSC/MFs and LPCs in relation to the histological location and extent of liver disease in immunohistochemically (double) stained serial sections. Markers of HSC/MFs [alpha-smooth muscle actin, glial fibrillary acidic protein (GFAP), neurotrophin 3 and neural-cell adhesion molecule], markers of LPCs (keratin 7 and keratin 19) and a proliferation marker (Ki67) were used. A very relevant spontaneous model to evaluate LPC niche activation in a translational approach seems to be the dog. Therefore, both human and canine liver diseases with different degree of fibrosis and disease activity were included. In human and canine liver disease, type and extent of LPC niche activation depended on type and severity of disease (P<0.05) and corresponded to the main location of disease. Activated HSCs surrounded the activated LPCs. In chronic hepatitis and non-alcoholic steatohepatitis lobular-type HSCs were activated, while during biliary disease portal/septal MFs were mainly activated. In canine liver, GFAP further presented as an early marker of HSC activation. Activation of the LPCs correlated with disease location and severity (P<0.01), and was inversely related to hepatocyte proliferation, as was previously shown in man. A shared involvement of HSC/MFs, LPCs and disease severity during hepatic disease processes is shown, which is highly similar in man and dog.
American Cocker Spaniels are predisposed to chronic hepatitis. To describe the clinical and histo... more American Cocker Spaniels are predisposed to chronic hepatitis. To describe the clinical and histological features of chronic hepatitis in Japanese American Cocker Spaniels. Thirteen cases examined from 2003 to 2009. Retrospective study. Medical records were searched for American Cocker Spaniels with chronic liver diseases. History, physical examination, clinicopathologic features, hepatic ultrasonographic findings, hepatic histopathology, and immunohistochemistry were evaluated. The median age was 4.6 (1.9-10.7) years. Clinical signs included inappetence (11/13), ascites (11/13), lethargy (9/13), diarrhea (7/13), and melena (2/13). Only 1/13 dogs was jaundiced. Clinicopathological abnormalities were increased liver enzymes (gamma-glutamyl transpeptidase: 9/12, aspartate aminotransferase: 7/10, alanine aminotransferase: 6/13, alkaline phosphatase: 6/13), increased total serum bile acid concentrations (10/12), and hypoalbuminemia (10/13). The liver had an irregular surface in all dogs and acquired portosystemic collaterals were verified in 11/13 dogs by abdominal ultrasound (2), laparoscopy (4), or both (5). Liver histology revealed severe fibrosis and cirrhosis in all cases, subdivided in lobular dissecting hepatitis (7), periportal fibrosis (1), micronodular cirrhosis (3), and macronocular cirrhosis (2). Inflammatory activity was low to mild. Immunohistochemical stains showed ductular proliferation. The median survival time was 913 (range: 63-1981) days. Hepatitis in Japanese American Cocker Spaniels is clinically silent until an advanced stage and is associated with severe hepatic fibrosis leading to cirrhosis, extensive ductular/putative hepatic progenitor cell proliferation, portal hypertension, and acquired portosystemic collateral shunting, but relatively long survival times. Lobular dissecting hepatitis seems more prevalent than in previously reported cases from other countries.
Hepatic progenitor cells (HPCs) hold a great potential for therapeutic intervention for currently... more Hepatic progenitor cells (HPCs) hold a great potential for therapeutic intervention for currently untreatable liver diseases. However, in human diseases molecular mechanisms involved in proliferation and differentiation of HPCs are poorly understood. In the present study activated HPCs and their microenvironment (niche) were investigated in acute and chronic human liver disease by gene-expression analysis and immunohistochemistry/immunofluorescence. Cryopreserved liver tissues were used from patients with parenchymal versus biliary diseases: acute necrotising hepatitis (AH), cirrhosis after hepatitis C infection, and primary biliary cirrhosis in order to study differentiation of HPCs towards hepatocytic versus biliary lineage. Keratin 7 positive HPCs/reactive ductules were captured by means of laser capture microdissection and gene-expression profiles were obtained by using a customized PCR array. Gene expression results were confirmed by immunohistochemistry and immunofluorescence double staining. In all disease groups, microdissected HPCs expressed progenitor cell markers such as KRT7, KRT19, NCAM, ABCG2, LIF, KIT, OCT4, CD44 and TERT. In AH, HPCs were most activated and showed a high expression of prominin-1 (CD133) and alpha-fetoprotein, and a strong activation of the Wnt pathway. In contrast to parenchymal diseases, HPCs in primary biliary cirrhosis (biliary differentiation) showed a high activation of Notch signalling. A distinct pattern of HPC surface markers was found between acute and chronic liver diseases. Similar to what is known from animal experiments, strong evidence has been found signifying the role of Wnt signalling in proliferation of human HPCs whereas Notch signalling is involved in biliary differentiation. These pathways can be targeted in future therapies.
BackgroundThe liver has a large regenerative capacity. Hepatocytes can replicate and regenerate a... more BackgroundThe liver has a large regenerative capacity. Hepatocytes can replicate and regenerate a diseased liver. However, as is the case in severe liver diseases, this replication may become insufficient or exhausted and hepatic progenitor cells (HPCs) can be activated in an attempt to restore liver function. Due to their bi-potent differentiation capacity, these HPCs have great potential for regenerative approaches yet over-activation does pose potential health risks. Therefore the mechanisms leading to activation must be elucidated prior to safe implementation in the veterinary clinic. Wnt/ß-catenin and Notch signalling have been implicated in the activation of HPCs in mouse models and in humans. Here we assessed the involvement in canine HPC activation. Gene-expression profiles were derived from laser microdissected HPCs niches from lobular dissecting hepatitis (LDH) and normal liver tissue, with a focus on Wnt/ß-catenin and Notch signalling. Immunohistochemical and immunofluore...
Uploads
Papers by Baukje Schotanus