Induction of colitis in cftr-/- mice results in bile duct injury

It is unknown why some patients with inflammatory bowel disease develop primary sclerosing cholangitis. We have recently shown that patients with primary sclerosing cholangitis have an increased prevalence of mutations in the gene responsible for cystic fibrosis (CFTR) compared with individuals with inflammatory bowel disease alone. Our aim was to examine whether induction of colitis by oral dextran leads to bile duct injury in mice heterozygous or homozygous for mutations in CFTR. The effect of oral administration of docosahexaenoic acid to correct a fatty acid imbalance associated with cystic fibrosis was also examined to determine whether this would prevent bile duct inflammation. Wild-type mice and mice heterozygous and homozygous for CFTR mutations were given dextran orally for 14 days to induce colitis. Bile duct injury was quantitated by blinded histological scoring and measurement of serum alkaline phosphatase activity. The effect of pretreatment with docosahexaenoic acid for 7 days was examined. Treatment of mice with 100 mg dextran/day for 9 days followed by 85 mg/day for 5 days resulted in a significant increase in bile duct injury as determined by histological scoring in homozygous cystic fibrosis mice compared with wild-type mice (P = 0.005). The bile duct injury seen in cystic fibrosis mice was reflected in a threefold increase in serum alkaline phosphatase (P = 0.0006). Pretreatment with oral docosahexaenoic acid decreased both histological evidence of bile duct injury and serum alkaline phosphatase levels. In the setting of colitis, loss of CFTR function leads to bile duct injury.     

Increased expression of cystic fibrosis transmembrane conductance regulator in rat liver after common bile duct ligation

Liver disease associated with cystic fibrosis (CF) has been increasingly diagnosed and recognized as one of the major causes of death in CF during recent years. The autosomal-recessive disorder of CF results from mutation of the cystic fibrosis transmembrane conductance regulator (CFTR) that encodes the CFTR protein. Due to its existence and multifunction in biliary epithelial, over- or less-expression of CFTR in the liver may play an important role in the development of CF liver disease (CFLD). The aim of current study is to investigate the expression of CFTR in the liver of common bile duct ligated (BDL) rats. After BDL, there was an increase in the abundance of CFTR mRNA and protein. Immunohistochemical staining also demonstrated an increased intensity of CFTR staining in the liver tissue section. In conclusion, there is an increased expression of CFTR in the liver after common BDL.     

LPS signaling enhances hepatic fibrogenesis caused by experimental cholestasis in mice

Although it is clear that bile acid accumulation is the major initiator of fibrosis caused by cholestatic liver disease, endotoxemia is a common side effect. However, the depletion of hepatic macrophages with gadolinium chloride blunts hepatic fibrosis. Because endotoxin is a key activator of hepatic macrophages, this study was designed to test the hypothesis that LPS signaling through CD14 contributes to hepatic fibrosis caused by experimental cholestasis. Wild-type mice and CD14 knockout mice (CD14(-/-)) underwent sham operation or bile duct ligation and were killed 3 wk later. Measures of liver injury, such as focal necrosis, biliary cell proliferation, and inflammatory cell influx, were not significantly different among the strains 3 wk after bile duct ligation. Markers of liver fibrosis such as Sirius red staining, liver hydroxyproline, and alpha-smooth muscle actin expression were blunted in CD14(-/-) mice compared with wild-type mice after bile duct ligation. Despite no difference in lymphocyte infiltration, the macrophage/monocyte activation marker OX42 (CD11b) and the oxidative stress/lipid peroxidation marker 4-hydroxynonenal were significantly upregulated in wild-type mice after bile duct ligation but not in CD14(-/-) mice. Increased profibrogenic cytokine mRNA expression in the liver after bile duct ligation was significantly blunted in CD14(-/-) mice compared with the wild type. The hypothesis that LPS was involved in experimental cholestatic liver fibrosis was tested using mice deficient in LPS-binding protein (LBP(-/-)). LBP(-/-) mice had less liver injury and fibrosis (Siruis red staining and hydroxyproline content) compared with wild-type mice after bile duct ligation. In conclusion, these data demonstrate that endotoxin in a CD14-dependent manner exacerbates hepatic fibrogenesis and macrophage activation to produce oxidants and cytokines after bile duct ligation.     

Cholic Acid Induces a Cftr Dependent Biliary Secretion and Liver Growth Response in Mice

The cause of Cystic fibrosis liver disease (CFLD), is unknown. It is well recognized that hepatic exposure to hydrophobic bile salts is associated with the development of liver disease. For this reason, we hypothesize that, CFTR dependent variations, in the hepatic handling of hydrophobic bile salts, are related to the development CFLD. To test our hypothesis we studied, in Cftr^-/- and control mice, bile production, bile composition and liver pathology, in normal feeding condition and during cholate exposure, either acute (intravenous) or chronic (three weeks via the diet). In Cftr^-/- and control mice the basal bile production was comparable. Intravenous taurocholate increased bile production to the same extent in Cftr^-/- and control mice. However, chronic cholate exposure increased the bile flow significantly less in Cftr^-/- mice than in controls, together with significantly higher biliary bile salt concentration in Cftr^-/- mice. Prolonged cholate exposure, however, did not induce CFLD like pathology in Cftr^-/- mice. Chronic cholate exposure did induce a significant increase in liver mass in controls that was absent in Cftr^-/- mice. Chronic cholate administration induces a cystic fibrosis-specific hepatobiliary phenotype, including changes in bile composition. These changes could not be associated with CFLD like pathological changes in CF mouse livers. However, chronic cholate administration induces liver growth in controls that is absent in Cftr^-/- mice. Our findings point to an impaired adaptive homeotrophic liver response to prolonged hydrophobic bile salt exposure in CF conditions.     

Cystic fibrosis mice carrying the missense mutation G551D replicate human genotype-phenotype correlations.

We have generated a mouse carrying the human G551D mutation in the cystic fibrosis transmembrane conductance regulator gene (CFTR) by a one-step gene targeting procedure. These mutant mice show cystic fibrosis pathology but have a reduced risk of fatal intestinal blockage compared with 'null' mutants, in keeping with the reduced incidence of meconium ileus in G551D patients. The G551D mutant mice show greatly reduced CFTR-related chloride transport, displaying activity intermediate between that of cftr(mlUNC) replacement ('null') and cftr(mlHGU) insertional (residual activity) mutants and equivalent to approximately 4% of wild-type CFTR activity. The long-term survival of these animals should provide an excellent model with which to study cystic fibrosis, and they illustrate the value of mouse models carrying relevant mutations for examining genotype-phenotype correlations.     

Drug-induced bile duct injury

Drug-induced liver injury includes a spectrum of pathologies, some related to the mode of injury, some to the cell type primarily damaged. Among these, drug-induced bile duct injury is characterized by the destruction of the biliary epithelium following exposure to a drug. Most of the drugs associated with bile duct injury cause immune-mediated lesions to the epithelium of interlobular ducts. These share common histopathological features with primary biliary cholangitis, such as inflammation and necrosis at the expense of cholangiocytes and, if the insult persists, bile duct loss and biliary cirrhosis. Some drugs selectively target larger ducts. Such injury is often dose-dependent and thought to be the result of intrinsic drug toxicity. The histological changes resemble those seen in primary sclerosing cholangitis. This overview focuses on the clinical and pathological features of bile duct injury associated with drug treatment and on the immunological and biochemical effects that drugs exert on the biliary epithelium. This article is part of a Special Issue entitled: Cholangiocytes in Health and Disease edited by Jesus Banales, Marco Marzioni, Nicholas LaRusso and Peter Jansen.     

Mechanisms of cholangiocyte responses to injury

Cholangiocytes, epithelial cells that line the biliary epithelium, are the primary target cells for cholangiopathies including primary sclerosing cholangitis and primary biliary cholangitis. Quiescent cholangiocytes respond to biliary damage and acquire an activated neuroendocrine phenotype to maintain the homeostasis of the liver. The typical response of cholangiocytes is proliferation leading to bile duct hyperplasia, which is a characteristic of cholestatic liver diseases. Current studies have identified various signaling pathways that are associated with cholangiocyte proliferation/loss and liver fibrosis in cholangiopathies using human samples and rodent models. Although recent studies have demonstrated that extracellular vesicles and microRNAs could be mediators that regulate these messenger/receptor axes, further studies are required to confirm their roles. This review summarizes current studies of biliary response and cholangiocyte proliferation during cholestatic liver injury with particular emphasis on the secretin/secretin receptor axis. This article is part of a Special Issue entitled: Cholangiocytes in Health and Diseaseedited by Jesus Banales, Marco Marzioni, Nicholas LaRusso and Peter Jansen.     

Pathogenesis of biliary fibrosis

Chronic cholestatic liver diseases such as primary biliary cholangitis (PBC) and primary sclerosing cholangitis (PSC) are associated with active hepatic fibrogenesis, and, ultimately, to the development of cirrhosis. However, the precise relationship between cholestasis, in its broad meaning, and liver tissue fibrosis is still poorly defined. Fibrogenesis is currently viewed as a dynamic process that appears strictly related to the extent and duration of parenchymal injury. This relationship is clearly evident in the presence of reiterative hepatocellular necrosis due to viral infection or alcohol abuse. It appears that “pure” intralobular intrahepatic cholestasis secondary to biliary secretory failure of the hepatocyte, in absence of hepatocellular damage, lobular inflammation and bile duct damage and/or proliferation, is not associated with marked and/or progressive liver tissue fibrosis. In contrast, marked and progressive liver tissue fibrosis always follows liver diseases characterized by chronic inflammatory bile duct damage as seen in PBC and PSC or chronic mechanical obstruction of the biliary tree. Overall, the fibrogenic process in these clinical conditions appears to be related to a more complex interaction between immune/inflammatory mechanisms, cytokine networks and the derangement of the homeostasis between epithelial and mesenchymal cells. The elucidation of these mechanisms is indeed crucial for the identification of potential diagnostic and therapeutic targets.     

Small duct autoimmune sclerosing cholangitis and Crohn colitis in a 10-year-old child. A case report and review of the literature

ABSTRACT: Autoimmune sclerosing cholangitis is an overlap syndrome characterized by features of both autoimmune hepatitis and primary sclerosing cholangitis, the latter usually involving the large bile ducts. Autoimmune sclerosing cholangitis occurs more often in children than in adults and is frequently associated with inflammatory bowel disease, predominantly ulcerative colitis. We report a unique case of a 10-year-old Danish boy with severe small duct autoimmune sclerosing cholangitis and synchronic Crohn colitis. He was referred with a history of weight loss, abdominal pain, vomiting and diarrhea. Biochemical anomalies included elevated alanine aminotransferase, gamma-glutamyl transferase and immunoglobulin G levels and the presence of smooth muscle antibodies and perinuclear antineutrophil cytoplasmic antibodies but normal alkaline phosphatase. Liver biopsy specimen revealed features of both autoimmune hepatitis and sclerosing cholangitis, the latter characterized by acute, hyperplastic and destructive inflammation - granulocytic epithelial lesion - of the small ducts. Magnetic resonance cholangiography was normal. Colonoscopic biopsies showed chronic inflammatory changes of the caecum and the ascending and transverse colon compatible with Crohn disease. Ursodeoxycholic acid and immunosuppressive treatment was initiated and within four weeks of treatment the general condition improved. Normalization of aminotransferase was seen at 21 weeks and gamma-glutamyl transferase at 72 weeks after first admittance, while immunoglobulin G remained slightly increased. Virtual slides The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1418596609736470.     

Down-regulation of the anti-inflammatory protein annexin A1 in cystic fibrosis knock-out mice and patients

Cystic fibrosis is a fatal human genetic disease caused by mutations in the CFTR gene encoding a cAMP-activated chloride channel. It is characterized by abnormal fluid transport across secretory epithelia and chronic inflammation in lung, pancreas, and intestine. Because cystic fibrosis (CF) pathophysiology cannot be explained solely by dysfunction of cystic fibrosis transmembrane conductance regulator (CFTR), we applied a proteomic approach (bidimensional electrophoresis and mass spectrometry) to search for differentially expressed proteins between mice lacking cftr (cftr(tm1Unc), cftr-/-) and controls using colonic crypts from young animals, i.e. prior to the development of intestinal inflammation. By analyzing total proteins separated in the range of pH 6-11, we detected 24 differentially expressed proteins (>2-fold). In this work, we focused on one of these proteins that was absent in two-dimensional gels from cftr-/- mice. This protein spot (molecular mass, 37 kDa; pI 7) was identified by mass spectrometry as annexin A1, an anti-inflammatory protein. Interestingly, annexin A1 was also undetectable in lungs and pancreas of cftr-/- mice, tissues known to express CFTR. Absence of this inhibitory mediator of the host inflammatory response was associated with colonic up-regulation of the proinflammatory cytosolic phospholipase A2. More importantly, annexin A1 was down-regulated in nasal epithelial cells from CF patients bearing homozygous nonsense mutations in the CFTR gene (Y122X, 489delC) and differentially expressed in F508del patients. These results suggest that annexin A1 may be a key protein involved in CF pathogenesis especially in relation to the not well defined field of inflammation in CF. We suggest that decreased expression of annexin A1 contributes to the worsening of the CF phenotype.     

Maternal obesogenic diet enhances cholestatic liver disease in offspring

Human and animal model data show that maternal obesity promotes nonalcoholic fatty liver disease in offspring and alters bile acid (BA) homeostasis. Here we investigated whether offspring exposed to maternal obesogenic diets exhibited greater cholestatic injury. We fed female C57Bl6 mice conventional chow (CON) or high fat/high sucrose (HF/HS) diet and then bred them with lean males. Offspring were fed 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) for 2 weeks to induce cholestasis, and a subgroup was then fed CON for an additional 10 days. Additionally, to evaluate the role of the gut microbiome, we fed antibiotic-treated mice cecal contents from CON or HF/HS offspring, followed by DDC for 2 weeks. We found that HF/HS offspring fed DDC exhibited increased fine branching of the bile duct (ductular reaction) and fibrosis but did not differ in BA pool size or intrahepatic BA profile compared to offspring of mice fed CON. We also found that after 10 days recovery, HF/HS offspring exhibited sustained ductular reaction and periportal fibrosis, while lesions in CON offspring were resolved. In addition, cecal microbiome transplant from HF/HS offspring donors worsened ductular reaction, inflammation, and fibrosis in mice fed DDC. Finally, transfer of the microbiome from HF/HS offspring replicated the cholestatic liver injury phenotype. Taken together, we conclude that maternal HF/HS diet predisposes offspring to increased cholestatic injury after DDC feeding and delays recovery after returning to CON diets. These findings highlight the impact of maternal obesogenic diet on hepatobiliary injury and repair pathways during experimental cholestasis.     

Animal models for cystic fibrosis liver disease (CFLD)

Liver disease is a severe complication in patients with Cystic Fibrosis (CF), a genetic disease caused by mutations in the gene encoding for cystic fibrosis transmembrane conductance regulator (CFTR) channel. The sequence of events leading to CFLD is still unclear and has limited the development of more specific treatments other than the bile acid UDCA. However, in the last twenty years, several gaps have been filled, which have mainly been possible due to the availability of different animal models that mimic CF. CF mice, although they lack a spontaneous liver manifestation, have been essential to better understand the multiple functions of CFTR expression on the apical membrane of cholangiocytes, from chloride channel to regulator of epithelial innate immunity. Additionally, we have learned that the gut microbiota might be a pathogenetic factor for the development of liver disease. The recent creation of novel CF animal models (i.e. pig and ferret) that better reproduce the human disease, will allow for comparative studies with species that spontaneously develop the liver disease and will hopefully lead to novel therapeutic treatments. In this review, we have compared and summarized the main features of the current available CF animal models and their applicability for the study of the liver phenotype.     

Ursodeoxycholate modulates bile flow and bile salt pool independently from the cystic fibrosis transmembrane regulator (Cftr) in mice

Cystic fibrosis liver disease (CFLD) is treated with ursodeoxycholate (UDCA). Our aim was to evaluate, in cystic fibrosis transmembrane regulator knockout (Cftr(-/-)) mice and wild-type controls, whether the supposed therapeutic action of UDCA is mediated via choleretic activity or effects on bile salt metabolism. Cftr(-/-) mice and controls, under general anesthesia, were intravenously infused with tauroursodeoxycholate (TUDCA) in increasing dosage or were fed either standard or UDCA-enriched chow (0.5% wt/wt) for 3 wk. Bile flow and bile composition were characterized. In chow-fed mice, we analyzed bile salt synthesis and pool size of cholate (CA). In both Cftr(-/-) and controls intravenous TUDCA stimulated bile flow by ～250% and dietary UDCA by ～500%, compared with untreated animals (P < 0.05). In non-UDCA-treated Cftr(-/-) mice, the proportion of CA in bile was higher compared with that in controls (61 ± 4 vs. 46 ± 4%; P < 0.05), accompanied by an increased CA synthesis [16 ± 1 vs. 10 ± 2 μmol·h(-1)·100 g body wt (BW)(-1); P < 0.05] and CA pool size (28 ± 3 vs. 19 ± 1 μmol/100 g BW; P < 0.05). In both Cftr(-/-) and controls, UDCA treatment drastically reduced the proportion of CA in bile below 5% and diminished CA synthesis (2.3 ± 0.3 vs. 2.2 ± 0.4 μmol·day(-1)·100 g BW(-1); nonsignificant) and CA pool size (3.6 ± 0.6 vs. 1.5 ± 0.3 μmol/100 g BW; P < 0.05). Acute TUDCA infusion and chronic UDCA treatment both stimulate bile flow in cystic fibrosis conditions independently from Cftr function. Chronic UDCA treatment reduces the hydrophobicity of the bile salt pool in Cftr(-/-) mice. These results support a potential beneficial effect of UDCA on bile flow and bile salt metabolism in cystic fibrosis conditions.     

Increased prevalence of CFTR mutations and variants and decreased chloride secretion in primary sclerosing cholangitis

Primary sclerosing cholangitis (PSC) and cystic fibrosis (CF) are both slowly progressive cholestatic liver diseases characterized by fibro-obliterative inflammation of the biliary tract. We hypothesized that dysfunction of the CF gene product, cystic fibrosis transmembrane conductance regulator (CFTR), may explain why a subset of patients with inflammatory bowel disease develop PSC. We prospectively evaluated CFTR genotype and phenotype in patients with PSC ( n=19) compared with patients with inflammatory bowel disease and no liver disease ( n=18), primary biliary cirrhosis ( n=17), CF ( n=81), and healthy controls ( n=51). Genetic analysis of the CFTR gene in PSC patients compared with disease controls (primary biliary cirrhosis and inflammatory bowel disease) demonstrated a significantly increased number of mutations/variants in the PSC group (37% vs 8.6% of disease controls, P=0.02). None of the PSC patients carried two mutations/variants. Of PSC patients, 89% carried the 1540G-variant-containing genotypes (resulting in decreased functional CFTR) compared with 57% of disease controls ( P=0.03). Only one of 19 PSC patients had neither a CFTR mutation nor the 1540G variant. CFTR chloride channel function assessed by nasal potential difference testing demonstrated a reduced median isoproterenol response of 14 mV in PSC patients compared with 19 mV in disease controls ( P=0.04) and 21 mV in healthy controls ( P=0.003). These data indicate that there is an increased prevalence of CFTR abnormalities in PSC as demonstrated by molecular and functional analyses and that these abnormalities may contribute to the development of PSC in a subset of patients with inflammatory bowel disease.     

Elevated plasma IL-6 associates with increased risk of advanced fibrosis and cholangiocarcinoma in individuals infected by Opisthorchis viverrini

Opisthorchis viverrini is considered among the most important of the food-borne trematodes due to its strong association with advanced periductal fibrosis and bile duct cancer (cholangiocarcinoma). We investigated the relationship between plasma levels of Interleukin (IL)-6 and the risk of developing advanced fibrosis and bile duct cancer from chronic Opisthorchis infection. We show that IL-6 circulates in plasma at concentrations 58 times higher in individuals with advanced fibrosis than age, sex, and nearest-neighbor matched controls and 221 times higher in individuals with bile duct cancer than controls. We also observed a dose-response relationship between increasing levels of plasma IL-6 and increasing risk of advanced fibrosis and bile duct cancer; for example, in age and sex adjusted analyses, individuals with the highest quartiles of plasma IL-6 had a 19 times greater risk of developing advanced periductal fibrosis and a 150 times greater risk of developing of bile duct cancer than individuals with no detectable level of plasma IL-6. Finally, we show that a single plasma IL-6 measurement has excellent positive predictive value for the detection of both advanced bile duct fibrosis and bile duct cancer in regions with high O. viverrini transmission. These data support our hypothesis that common mechanisms drive bile duct fibrosis and bile duct tumorogenesis from chronic O. viverrini infection. Our study also adds a unique aspect to the literature on circulating levels of IL-6 as an immune marker of hepatobiliary pathology by showing that high levels of circulating IL-6 in plasma are not related to infection with O. viverrini, but to the development of the advanced and often lethal pathologies resulting from chronic O. viverrini infection.     

TNFalpha is required for cholestasis-induced liver fibrosis in the mouse

TNFα, a mediator of hepatotoxicity in several animal models, is elevated in acute and chronic liver diseases. Therefore, we investigated whether hepatic injury and fibrosis due to bile duct ligation (BDL) would be reduced in TNFα knockout mice (TNFα−/−). Survival after BDL was 60% in wild-type mice (TNFα+/+) and 90% in TNFα−/− mice. Body weight loss and liver to body weight ratios were reduced in TNFα−/− mice compared to TNFα+/+ mice. Following BDL, serum alanine transaminases (ALT) levels were elevated in TNFα+/+ mice (268.6 ± 28.2 U/L) compared to TNFα−/− mice (105.9 U/L ± 24.4). TNFα−/− mice revealed lower hepatic collagen expression and less liver fibrosis in the histology. Further, α-smooth muscle actin, an indicator for activated myofibroblasts, and TGF-β mRNA, a profibrogenic cytokine, were markedly reduced in TNFα−/− mice compared to TNFα+/+ mice. Thus, our data indicate that TNFα induces hepatotoxicity and promotes fibrogenesis in the BDL model.     

No indications for altered essential fatty acid metabolism in two murine models for cystic fibrosis

A deficiency of essential fatty acids (EFA) is frequently described in cystic fibrosis (CF), but whether this is a primary consequence of altered EFA metabolism or a secondary phenomenon is unclear. It was suggested that defective long-chain polyunsaturated fatty acid (LCPUFA) synthesis contributes to the CF phenotype. To establish whether cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction affects LCPUFA synthesis, we quantified EFA metabolism in cftr-/-CAM and cftr+/+CAM mice. Effects of intestinal phenotype, diet, age, and genetic background on EFA status were evaluated in cftr-/-CAM mice, DeltaF508/DeltaF508 mice, and littermate controls. EFA metabolism was measured by 13C stable isotope methodology in vivo. EFA status was determined by gas chromatography in tissues of cftr-/-CAM mice, DeltaF508/DeltaF508 mice, littermate controls, and C57Bl/6 wild types fed chow or liquid diet. After enteral administration of [13C]EFA, arachidonic acid (AA) and docosahexaenoic acid (DHA) were equally 13C-enriched in cftr-/-CAM and cftr+/+CAM mice, indicating similar EFA elongation/desaturation rates. LA, ALA, AA, and DHA concentrations were equal in pancreas, lung, and jejunum of chow-fed cftr-/-CAM and DeltaF508/DeltaF508 mice and controls. LCPUFA levels were also equal in liquid diet-weaned cftr-/-CAM mice and littermate controls, but consistently higher than in age- and diet-matched C57Bl/6 wild types. We conclude that cftr-/-CAM mice adequately absorb and metabolize EFA, indicating that CFTR dysfunction does not impair LCPUFA synthesis. A membrane EFA imbalance is not inextricably linked to the CF genotype. EFA status in murine CF models is strongly determined by genetic background.     

Regulatory interaction between the cystic fibrosis transmembrane conductance regulator and HCO3- salvage mechanisms in model systems and the mouse pancreatic duct

The pancreatic duct expresses cystic fibrosis transmembrane conductance regulator (CFTR) and HCO3- secretory and salvage mechanisms in the luminal membrane. Although CFTR plays a prominent role in HCO3- secretion, the role of CFTR in HCO3- salvage is not known. In the present work, we used molecular, biochemical, and functional approaches to study the regulatory interaction between CFTR and the HCO3- salvage mechanism Na+/H+ exchanger isoform 3 (NHE3) in heterologous expression systems and in the native pancreatic duct. We found that CFTR regulates NHE3 activity by both acute and chronic mechanisms. In the pancreatic duct, CFTR increases expression of NHE3 in the luminal membrane. Thus, luminal expression of NHE3 was reduced by 53% in ducts of homozygote DeltaF508 mice. Accordingly, luminal Na+-dependent and HOE694- sensitive recovery from an acid load was reduced by 60% in ducts of DeltaF508 mice. CFTR and NHE3 were co-immunoprecipitated from PS120 cells expressing both proteins and the pancreatic duct of wild type mice but not from PS120 cells lacking CFTR or the pancreas of DeltaF508 mice. The interaction between CFTR and NHE3 required the COOH-terminal PDZ binding motif of CFTR, and mutant CFTR proteins lacking the C terminus were not co-immunoprecipitated with NHE3. Furthermore, when expressed in PS120 cells, wild type CFTR, but not CFTR mutants lacking the C-terminal PDZ binding motif, augmented cAMP-dependent inhibition of NHE3 activity by 31%. These findings reveal that CFTR controls overall HCO3- homeostasis by regulating both pancreatic ductal HCO3- secretory and salvage mechanisms.     

Protective role of melatonin in early‐stage and end‐stage liver cirrhosis

The liver is composed of hepatocytes, cholangiocytes, Kupffer cells, sinusoidal endothelial cells, hepatic stellate cells (HSCs) and dendritic cells; all these functional and interstitial cells contribute to the synthesis and secretion functions of liver tissue. However, various hepatotoxic factors including infection, chemicals, high‐fat diet consumption, surgical procedures and genetic mutations, as well as biliary tract diseases such as sclerosing cholangitis and bile duct ligation, ultimately progress into liver cirrhosis after activation of fibrogenesis. Melatonin (MT), a special hormone isolated from the pineal gland, participates in regulating multiple physiological functions including sleep promotion, circadian rhythms and neuroendocrine processes. Current evidence shows that MT protects against liver injury by inhibiting oxidation, inflammation, HSC proliferation and hepatocyte apoptosis, thereby inhibiting the progression of liver cirrhosis. In this review, we summarize the circadian rhythm of liver cirrhosis and its potential mechanisms as well as the therapeutic effects of MT on liver cirrhosis and earlier‐stage liver diseases including liver steatosis, nonalcoholic fatty liver disease and liver fibrosis. Given that MT is an antioxidative and anti‐inflammatory agent that is effective in eliminating liver injury, it is a potential agent with which to reverse liver cirrhosis in its early stage.