Clinical lectures on the gallbladder and bile ducts,

"Radiology of the gallbladder, by Ernest E. Smith": p. 84-104. "Diagnostic and therapeutic duodenal drainage, by Harry M. Eberhard": p. 105-136.

ENaC activity in collecting ducts modulates NCC in cirrhotic mice.

Cirrhosis is a frequent and severe disease, complicated by renal sodium retention leading to ascites and oedema. A better understanding of the complex mechanisms responsible for renal sodium handling could improve clinical management of sodium retention. Our aim was to determine the importance of the amiloride-sensitive epithelial sodium channel (ENaC) in collecting ducts in compensate and decompensate cirrhosis. Bile duct ligation was performed in control mice (CTL) and collecting duct-specific αENaC knockout (KO) mice, and ascites development, aldosterone plasma concentration, urinary sodium/potassium ratio and sodium transporter expression were compared. Disruption of ENaC in collecting ducts (CDs) did not alter ascites development, urinary sodium/potassium ratio, plasma aldosterone concentrations or Na,K-ATPase abundance in CCDs. Total αENaC abundance in whole kidney increased in cirrhotic mice of both genotypes and cleaved forms of α and γ ENaC increased only in ascitic mice of both genotypes. The sodium chloride cotransporter (NCC) abundance was lower in non-ascitic KO, compared to non-ascitic CTL, and increased when ascites appeared. In ascitic mice, the lack of αENaC in CDs induced an upregulation of total ENaC and NCC and correlated with the cleavage of ENaC subunits. This revealed compensatory mechanisms which could also take place when treating the patients with diuretics. These compensatory mechanisms should be considered for future development of therapeutic strategies.

Histopathology findings in common marmosets (Callithrix jacchus Linnaeus, 1758) with chronic weight loss associated with bile tract obstruction by infestation with Platynosomum (Loos, 1907)

Chronic weight loss in marmosets is often associated with wasting marmoset syndrome (WMS), an important disease that occurs in callitrichid colonies around the world. Even though its etiology is very difficult to determine, particular variables, such as weight loss, diarrhea and alopecia, associated or not with infestation in the pancreatic ducts with Trichospirura leptossoma (Nematoda: Thelazioidea), seem to be linked with the syndrome. This study investigated the histopathology of the lungs, duodenum, liver, gallbladder, extrahepatic bile ducts and pancreatic ducts of six common marmosets (Callithrix jacchus) suffering from severe non-diarrheic weight loss. Three individuals died naturally and the other three were euthanized. Microscopic findings showed the presence of adult flukes (Platynosomum) in the liver. These flukes, which provoke common infection in cats, were also observed inside the gallbladder as well as in the intra and extrahepatic bile ducts in common marmosets. Portal fibrosis was observed in two animals, which developed chronic fibrosing hepatopathy (biliary pattern, grade 3). The disease progresses without diarrhea and without pancreatic lesions or infestation. With the rogression, the animals presented with ascending cholangitis, cholestasis and portal fibrosis, sometimes culminating in secondary biliary cirrhosis. Therefore, this nfirmity, associated with chronic weight loss in common marmosets, could be another tiological factor linked with WMS

Common bile duct obstruction associated with platinosoma concinum in cat

Extrahepatic obstruction in cat,can be caused by cholelithiasis, inspissated bile, pancreatic inflammation or fibrosis, duodenal inflammation, bile duct carcinoma or liver fluke infection (Amphimerus pseudofelineus e Platynosomurn concinum). A three-year-old, female neutered siames cat was present with a two-month history of progressive letargy, anorexia and emaciation. She had severe icterus of the mucous membranes, skin, and sclerae. A diagnosis of extrahepataic biliary obstruction was made based on the increased levels of conjugated bilirubin in the serum and the ultrasonography. Hepatic fluke eggs were not diagnosed in the feces because the fibrotic bile ducts were occluded and no eggs were shed into the intestine. Cholecystoduodenostomy was done to relieve posthepatic-biliary obstruction.

Effects of selective bile duct ligation on liver parenchyma in young animals: histologic and molecular evaluations

Background/Purpose: The mechanisms of increased collagen production and liver parenchyma fibrosis are poorly understood. These phenomena are observed mainly in children with biliary obstruction (BO), and in a great number of patients, the evolution to biliary cirrhosis and hepatic failure leads to the need for liver transplantation before adolescence. However, pediatric liver transplantation presents with biliary complications in 20% to 30% of cases in the postoperative period. Intra-or extrahepatic stenosis of bile ducts is frequent and may lead to secondary biliary cirrhosis and the need for retransplantation. It is unknown whether biliary stenosis involving isolated segments or lobes may affect the adjacent nonobstructed lobes by paracrine or endocrine means, leading to fibrosis in this parenchyma. Therefore, the present study aimed to create an experimental model of selective biliary duct ligation in young animals with a subsequent evaluation of the histologic and molecular alterations in liver parenchyma of the obstructed and nonobstructed lobes. Methods: After a pilot study to standardize the surgical procedures, weaning rats underwent ligation of the bile ducts of the median, left lateral, and caudate liver lobes. The bile duct of the right lateral lobe was kept intact. To avoid intrahepatic biliary duct collaterals neoformation, the parenchymal connection between the right lateral and median lobes was clamped. The animals were divided into groups according to the time of death: 1, 2, 3, 4, and 8 weeks after surgical procedure. After death, the median and left lateral lobes (with BO) and the right lateral lobe (without BO [NBO]) were harvested separately. A group of 8 healthy nonoperated on animals served as controls. Liver tissues were subjected to histologic evaluation and quantification of the ductular proliferation and of the portal fibrosis. The expressions of smooth muscle alpha-actin (alpha-SMA), desmin, and transforming growth factor beta 1 genes were studied by molecular analyses (semiquantitative reverse transcriptase-polymerase chain reaction and real-time polymerase chain reaction, a quantitative method). Results: Histologic analyses revealed the occurrence of ductular proliferation and collagen formation in the portal spaces of both BO and NBO lobes. These phenomena were observed later in NBO than BO. Bile duct density significantly increased 1 week after duct ligation; it decreased after 2 and 3 weeks and then increased again after 4 and 8 weeks in both BO and NBO lobes. The portal space collagen area increased after 2 weeks in both BO and NBO lobes. After 3 weeks, collagen deposition in BO was even higher, and in NBO, the collagen area started decreasing after 2 weeks. Molecular analyses revealed increased expression of the alpha-SMA gene in both BO and NBO lobes. The semiquantitative and quantitative methods showed concordant results. Conclusions: The ligation of a duct responsible for biliary drainage of the liver lobe promoted alterations in the parenchyma and in the adjacent nonobstructed parenchyma by paracrine and/or endocrine means. This was supported by histologic findings and increased expression of alpha-SMA, a protein related to hepatic fibrogenesis. (C) 2012 Elsevier Inc. All rights reserved.

Alternative splicing of the rat sodium/bile acid transporter changes its cellular localization and transport properties

Bile secretion involves the structural and functional interplay of hepatocytes and cholangiocytes, the cells lining the intrahepatic bile ducts. Hepatocytes actively secrete bile acids into the canalicular space and cholangiocytes then transport bile acids in a vectorial manner across their apical and basolateral plasma membranes. The initial step in the transepithelial transport of bile acids across rat cholangiocytes is apical uptake by a Na+-dependent bile acid transporter (ASBT). To date, the molecular basis of the obligate efflux mechanism for extrusion of bile acids across the cholangiocyte basolateral membrane remains unknown. We have identified an exon-2 skipped, alternatively spliced form of ASBT, designated t-ASBT, expressed in rat cholangiocytes, ileum, and kidney. Alternative splicing causes a frameshift that produces a 154-aa protein. Antipeptide antibodies detected the ≈19 kDa t-ASBT polypeptide in rat cholangiocytes, ileum, and kidney. The t-ASBT was specifically localized to the basolateral domain of cholangiocytes. Transport studies in Xenopus oocytes revealed that t-ASBT can function as a bile acid efflux protein. Thus, alternative splicing changes the cellular targeting of ASBT, alters its functional properties, and provides a mechanism for rat cholangiocytes and other bile acid-transporting epithelia to extrude bile acids. Our work represents an example in which a single gene appears to encode via alternative splicing both uptake and obligate efflux carriers in a bile acid-transporting epithelial cell.

Isolation of small polarized bile duct units.

Fragments of small interlobular bile ducts averaging 20 microns in diameter can be isolated from rat liver. These isolated bile duct units form luminal spaces that are impermeant to dextran-40 and expand in size when cultured in 10 microM forskolin for 24-48 hr. Secretion is Cl- and HCO3- dependent and is stimulated by forskolin > dibutyryl cAMP > secretion but not by dideoxyforskolin, as assessed by video imaging techniques. Secretin stimulates Cl-/HCO3- exchange activity, and intraluminal pH increases after forskolin administration. These studies establish that small polarized physiologically intact interlobular bile ducts can be isolated from rat liver. These isolated bile duct units should be useful preparations for assessing the transport properties of small bile duct segments, which are the primary site of injury in cholestatic liver disorders, known as "vanishing bile duct syndromes."

Histopathology findings in common marmosets (Callithrix jacchus Linnaeus, 1758) with chronic weight loss associated with bile tract obstruction by infestation with Platynosomum (Loos, 1907)

Chronic weight loss in marmosets is often associated with wasting marmoset syndrome (WMS), an important disease that occurs in callitrichid colonies around the world. Even though its etiology is very difficult to determine, particular variables, such as weight loss, diarrhea and alopecia, associated or not with infestation in the pancreatic ducts with Trichospirura leptossoma (Nematoda: Thelazioidea), seem to be linked with the syndrome. This study investigated the histopathology of the lungs, duodenum, liver, gallbladder, extrahepatic bile ducts and pancreatic ducts of six common marmosets (Callithrix jacchus) suffering from severe non-diarrheic weight loss. Three individuals died naturally and the other three were euthanized. Microscopic findings showed the presence of adult flukes (Platynosomum) in the liver. These flukes, which provoke common infection in cats, were also observed inside the gallbladder as well as in the intra and extrahepatic bile ducts in common marmosets. Portal fibrosis was observed in two animals, which developed chronic fibrosing hepatopathy (biliary pattern, grade 3). The disease progresses without diarrhea and without pancreatic lesions or infestation. With the rogression, the animals presented with ascending cholangitis, cholestasis and portal fibrosis, sometimes culminating in secondary biliary cirrhosis. Therefore, this nfirmity, associated with chronic weight loss in common marmosets, could be another tiological factor linked with WMS

Histopathology findings in common marmosets (Callithrix jacchus Linnaeus, 1758) with chronic weight loss associated with bile tract obstruction by infestation with Platynosomum (Loos, 1907)

Chronic weight loss in marmosets is often associated with wasting marmoset syndrome (WMS), an important disease that occurs in callitrichid colonies around the world. Even though its etiology is very difficult to determine, particular variables, such as weight loss, diarrhea and alopecia, associated or not with infestation in the pancreatic ducts with Trichospirura leptossoma (Nematoda: Thelazioidea), seem to be linked with the syndrome. This study investigated the histopathology of the lungs, duodenum, liver, gallbladder, extrahepatic bile ducts and pancreatic ducts of six common marmosets (Callithrix jacchus) suffering from severe non-diarrheic weight loss. Three individuals died naturally and the other three were euthanized. Microscopic findings showed the presence of adult flukes (Platynosomum) in the liver. These flukes, which provoke common infection in cats, were also observed inside the gallbladder as well as in the intra and extrahepatic bile ducts in common marmosets. Portal fibrosis was observed in two animals, which developed chronic fibrosing hepatopathy (biliary pattern, grade 3). The disease progresses without diarrhea and without pancreatic lesions or infestation. With the rogression, the animals presented with ascending cholangitis, cholestasis and portal fibrosis, sometimes culminating in secondary biliary cirrhosis. Therefore, this nfirmity, associated with chronic weight loss in common marmosets, could be another tiological factor linked with WMS

Fine mapping and replication of genetic risk Loci in primary sclerosing cholangitis

Background and aims. Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease characterized by progressive inflammation and fibrosis of the bile ducts eventually leading to biliary cirrhosis. Recent genetic studies in PSC have identified associations at 2q13, 2q35, 3p21, 4q27, 13q31 and suggestive association at 10p15. The aim of this study was to further characterize and refine the genetic architecture of PSC. Methods. We analyzed previously reported associated SNPs at four of these non-HLA loci and 59 SNPs tagging the IL-2/IL-21 (4q27) and IL2RA (10p15) loci in 992 UK PSC cases and 5162 healthy UK controls. Results. The most associated SNPs identified were rs3197999 (3p21 (MST1), p = 1.9 × 10 -6, OR A vs G = 1.28, 95% CI (1.16-1.42)); rs4147359 (10p15 (IL2RA), p = 2.6 × 10 -4, OR A vs G = 1.20, 95% CI (1.09-1.33)) and rs12511287 (4q27 (IL-2/IL-21), p = 3.0 × 10 -4, OR A vs T = 1.21, 95% CI (1.09-1.35)). In addition, we performed a meta-analysis for selected SNPs using published summary statistics from recent studies. We observed genome-wide significance for rs3197999 (3p21 (MST1), P combined = 3.8 × 10 -12) and rs4147359 (10p15 (IL2RA), P combined = 1.5 × 10 -8). Conclusion. We have for the first time confirmed the association of PSC with genetic variants at 10p15 (IL2RA) locus at genome-wide significance and replicated the associations at MST1 and IL-2/IL-21 loci in a large homogeneous UK population. These results strongly implicate the role of IL-2/IL2RA pathway in PSC and provide further confirmation of MST1 association. © Informa Healthcare.

Epidemiology and Treatment Options of Primary Biliary Cirrhosis

Primary biliary cirrhosis (PBC) is caused by an autoimmune inflammation of the small bile ducts. It results to destruction of bile ducts, accumulation of the bile in the liver, and cirrhosis. The prevalence and incidence of PBC is increasing in the Western world. The prevalence is highest in the USA (402 per million) and incidence in Scotland (49/million/year). Our aim was to assess the epidemiology of PBC in Finland. Patients for the epidemiological study were searched from the hospital discharge records from year 1988 to 1999.The prevalence rose from 103 to 180/million from 1988 to 1999, an annual increase of 5.1%. The incidence rose from 12 to 17 /million/year, an annual increase of 3.5%. The age at death increased markedly from 65 to 76 years. The risk of liver related deaths diminished over time. The treatment of PBC is based on Ursodeoxycholic acid (UDCA). During 20 years 50% of patients end up with cirrhosis. Our treatment option was to combine budesonide, a potent corticosteroid with a high first pass metabolism in the liver, to UDCA and evaluate the liver effects and systemic effects such as bone mass density (BMD) changes. Our aim was to find out if combination of laboratory tests would serve as a surrogate marker for PBC and help reducing the need for liver biopsy. Non-cirrhotic PBC patients were randomized to receive budesonide 6 mg/day combined to UDCA 15 mg /kg/day or UDCA alone for three years. The combination therapy with UDCA and budesonide was effective: stage improved 22%, fibrosis 25%, and inflammation 32%. In the UDCA group the changes were: 20% deterioriation in stage and 70% in fibrosis, but a 10% improvement in inflammation. BMD in femoral neck decreased by 3.6% in the combination group and by 1.9% in the UDCA group. The reductions in lumbar spine were 2.8% and 0.7%. Pharmacokinetics did not differ between the stages of PBC. HA, PIIINP, bile acids, and AST were significantly different within stages I-III and could differentiate the mild fibrosis (F0F1) from the moderate (F2F3). The combination of these individual markers (PBC-score) further improved the accuracy. The area under the ROC of the PBC score, using a cut of value 66, had a sensitivity of 81.4% and a specificity of 65.2% to classify the stage of PBC. The prevalence of PBC in Finland increases, which results from increasing incidence and improved survival. The combination of budesonide and UDCA improves liver histology compared to UDCA alone in non-cirrhotic stages of PBC. The treatment may reduce BMD. Hyaluronic acid, PIIINP, AST, and bile acids may serve as tools to monitor the treatment response in the early stages of PBC. The budesonide and UDCA combination therapy is an option for those patients who do not receive full response from UDCA and are still at the non-cirrhotic stage of PBC.

Fasciola hepatica: Comparative effects of host resistance and parasite intra-specific interactions on size and reproductive histology in flukes from rats infected with isolates differing in triclabendazole sensitivity

The efficacies of putative fasciolicides and vaccines against Fasciola hepatica are frequently monitored in clinical and field trials by determination of fluke egg output in host faeces and by worm counts in the host liver at autopsy. Less often used are parameters based on fluke size and histology, yet these can provide important indications of specific effects on the development of particular germ-line or somatic tissues, especially in relation to the timing and profligacy of egg production. In this study. F. hepatica metacercariae of two distinct isolates, the triclabendazole (TCBZ)-sensitive Cullompton isolate and the TCBZ-resistant Oberon isolate, were administered to rats as single-isolate or mixed-isolate infections. At autopsy 16 weeks later individual adult flukes were counted, measured and the reproductive organs were examined histologically. The degree of development of the testis tubules in each fluke was represented by a numerical score, based on the proportion of the histological section profiles occupied by testis tissue. The level of anti-F. hepatica antibody in the serum of each rat was determined by ELISA. It was found that Cullompton flukes were significantly larger than Oberon flukes, and that significantly more Cullompton metacercariae developed to adults than Oberon metacercariae. The Cullompton flukes showed histological evidence of aspermy and spermatogenic arrest, which was reflected in quantitatively reduced testicular development, as compared with the Oberon isolate. In Cullompton flukes, parthenogenetic egg development is implied. The size of Cullompton and Oberon flukes was significantly related to the number of adult flukes recovered, to the number of metacercariae administered, and to the percentage success of infection. The testis development score in both isolates was significantly related to the number of adult flukes recovered but not to the number of metacercariae administered, or to the percentage success of infection. Fluke size was positively related to testis score for both isolates, and a significant negative relationship was found between percentage success of infection and metacercarial dose. The results are interpreted in terms of differing interactions between various numbers of young flukes and host immunity during invasion of and migration in the hepatic parenchyma, and of fluke intra-specific (possibly pheromonal) stimulatory effects in the final stages of development, within the host bile ducts. No significant relationships were found between host antibody levels and fluke size or testis score. False positive serological reactions were found in some rats that had been infected, but found to harbour no flukes at autopsy. Clearly the act of eliminating the flukes involved generation of an immune response. (C) 2011 Elsevier B.V. All rights reserved.

Secreted cysteine proteases of the carcinogenic liver fluke, Opisthorchis viverrini: Regulation of cathepsin F activation by autocatalysis and trans-processing by cathepsin B

Opisthorchis viverrini is an important helminth pathogen of humans that is endemic in Thailand and Laos. Adult flukes reside within host bile ducts and feed on epithelial tissue and blood cells. Chronic opisthorchiasis is associated with severe hepatobiliary diseases such as cholangiocarcinoma. Here we report that adult O. viverrini secrete two major cysteine proteases: cathepsin F (Ov-CF-1) and cathepsin B1 (Ov-CB-1). Ov-CF-1 is secreted as an inactive zymogen that autocatalytically processes and activates to a mature enzyme at pH 4.5 via an intermolecular cleavage at the prosegment-mature domain junction. Ov-CB-1 is also secreted as a zymogen but, in contrast to Ov-CF-1, is fully active against peptide and macromolecular substrates despite retaining the N-terminal prosegment. The active Ov-CB-1 zymogen was capable of trans-activating Ov-CF-1 by proteolytic removal of its prosegment at pH 5.5, a pH at which the Ov-CF-1 zymogen cannot autocatalytically activate. Both cathepsins hydrolyse human haemoglobin but their combined action more efficiently degrades haemoglobin to smaller peptides than each enzyme alone. Ov-CF-1 degraded extracellular matrix proteins more effectively than Ov-CB-1 at physiological pH. We propose that Ov-CB-1 regulates Ov-CF-1 activity and that both enzymes work together to degrade host tissue contributing to the development of liver fluke-associated cholangiocarcinoma.