Bile Duct Ligation Induces ATZ Globule Clearance in a Mouse Model of α-1 Antitrypsin Deficiency.
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ABSTRACT:
SUBMITTER: Khan Z
PROVIDER: S-EPMC5296240 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Cholestatic liver diseases can be caused by genetic defects, drug toxicities, hepatobiliary malignancies or obstruction of the biliary tract. Cholestasis leads to accumulation of bile acids (BAs) in hepatocytes. Direct toxicity of BAs is currently the most accepted hypothesis for cholestatic liver injury. However, information on which bile acids are actually accumulating during cholestasis is limited.To assess the BA composition in liver and serum after bile duct ligation (BDL) in male C57Bl/6 mice between 6 h and 14 days and evaluate toxicity of the most abundant BAs.Bile acid concentrations increased in liver (27-fold) and serum (1400-fold) within 6 h after surgery and remained elevated up to 14 days. BAs in livers of BDL mice became more hydrophilic than sham controls, mainly because of increased 6?-hydroxylation and taurine conjugation. Among the eight unconjugated and 16 conjugated BAs identified in serum and liver, only taurocholic acid (TCA), ?-muricholic acid (?MCA) and T?MCA were substantially elevated representing >95% of these BAs over the entire time course. Although glycochenodeoxycholic acid and other conjugated BAs increased in BDL animals, the changes were several orders of magnitude lower compared with TCA, ?MCA and T?MCA. A mixture of these BAs did not cause apoptosis or necrosis, but induced inflammatory gene expression in cultured murine hepatocytes.The concentrations of cytotoxic BAs are insufficient to cause hepatocellular injury. In contrast, TCA, ?MCA and T?MCA are able to induce pro-inflammatory mediators in hepatocytes. Thus, BAs act as inflammagens and not as cytotoxic mediators after BDL in mice.
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Project description:Portal vein ligation (PVL) induces atrophy/hypertrophy complex (AHC). We hypothesised that simultaneous bile duct and portal vein ligation (BPL) might induce proper bile acid (BA) retention to enhance AHC by activating BA-mediated FXR signalling in the intact liver and promoting apoptosis in the ligated liver. We established rat models of 90% BPL and 90% PVL and found that BPL was well-tolerated and significantly accelerated AHC. The enhanced BA retention in the intact liver promoted hepatocyte proliferation by promoting the activation of FXR signalling, while that in the ligated liver intensified caspase3-mediated apoptosis. Decreasing the BA pools in the rats that underwent BPL could compromise these effects, whereas increasing the bile acid pools of rats that underwent PVL could induce similar effects. Second-stage resection of posterior-caudate-lobe-spearing hepatectomy was performed 5 days after BPL (B-Hx), PVL (V-Hx) or sham (S-SHx), as well as whole-caudate-lobe-spearing hepatectomy 5 days after sham (S-Hx). The B-Hx group had the most favourable survival rate (93.3%, the S-SHx group 0%, the S-Hx group 26.7%, the V-Hx group 56.7%, P < 0.01) and the most sustained regeneration. We conclude that BPL is a safe and effective method, and the acceleration of AHC was bile acid-dependent.
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Project description:Copper levels are elevated in a variety of liver fibrosis conditions. Lowering copper to a certain level protects against fibrosis. However, whether severe copper deficiency is protective against liver fibrosis is not known. The purpose of the present study is to evaluate this question by inducing severe copper deficiency using the copper chelator, tetrathiomolybdate (TM), in a bile duct ligation (BDL) rat model. Male Sprague-Dawley rats were divided into four groups: sham, sham plus TM, BDL, and BDL plus TM. TM was given in a daily dose of 10 mg/kg by body weight by means of intragastric gavage, beginning 5 days after BDL. All animals were killed 2 weeks after surgery. Severe copper deficiency was induced by TM overdose in either sham or BDL rats, as shown by decreased plasma ceruloplasmin activity. Liver injury and fibrosis were exacerbated in BDL rats with TM treatment, as illustrated by robustly increased plasma aspartate aminotransferase and hepatic collagen accumulation. Iron stores, as measured by plasma ferritin, were significantly increased in copper-deficient BDL rats. Moreover, hepatic heme oxygenase-1 expression was markedly down-regulated by copper deficiency in BDL rats. In addition, hepatic gene expression involving mitochondrial biogenesis and ?-oxidation was significantly up-regulated in BDL rats, and this increase was abolished by copper deficiency. In summary, severe copper deficiency exacerbates BDL-induced liver injury and liver fibrosis, probably caused by increased iron overload and decreased antioxidant defenses and mitochondrial dysfunction.
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Project description:Adherens junctions, consisting of cadherins and catenins, are a group of cell-to-cell junctions that mediate mechanistic linkage between neighboring cells. By doing so, adherens junctions ensure direct intercellular contact and play an indispensable role in maintaining tissue architecture. Considering these critical functions, it is not surprising that adherens junctions are frequently involved in disease. In the present study, the effects of bile duct ligation-a surgical procedure to experimentally induce cholestatic and fibrotic liver pathology-on hepatic adherens junctions were investigated in mice. In essence, it was found that liver mRNA and protein levels of E-cadherin, ?-catenin and ?-catenin drastically increase following bile duct ligation. These results could suggest a cytoprotective role for hepatic adherens junctions following bile duct ligation.
| S-EPMC6843439 | biostudies-literature
Project description:Male Sprague-Dawley rats weighing 250-300 g were purchased from Japan SLC Co. (Shizuoka, Japan). The animals were housed under a daily controlled 12-h light and 12-h dark cycle at 23 °C with free access to rat chow (Japan SLC Co.) and water for 1 week prior to the experiments. Rats were laparotomized under ether anesthesia in each experiment, and the common bile duct of each rat was ligated between the liver hilus and small intestine. Control animals underwent a sham operation with exposure but without ligation of the common bile duct. After the CBDL or sham operation, animals were allowed access to food and water ad libitum. Groups of three rats were sacrificed under ether anesthesia at 1, 3 and 12 hr after the CBDL or sham operation. At the time of sacrifice, the right lateral liver lobe was removed and flash-frozen in liquid nitrogen and stored at -80 °C. Total RNA isolated from frozen livers using TRIzol reagent (Invitrogen Co., Carlsbad, CA) was mixed to minimize variation among animals. Poly(A) RNA was purified using Oligotex-dT30 (Takara Shuzo Co., Ltd., Kyoto, Japan) in accordance with manufacturer’s instructions. Fluorescence-labeled probes were prepared by reverse transcription using a superscript II reverse transcriptase (Invitrogen Co.) and cyanine-3- and cyanine-5-dUTP (Perkin-Elmer Inc., Wellesley, MA). Poly(A) RNAs derived from rats that has undergone CBDL and from those that has undergone a sham operation were labeled with cyanine-3 and cyanine-5, respectively, and vise varsa. Fluorescence-labeled probes were purified using a MinElute PCR Products Purification Kit (Quiagen GmbH, Hilden, Germany) in accordance with manufacturer’s instructions. Each purified probe was suspended in hybridization buffer containing 1.6 mg/mL poly(A) (Roche Diagnostics, Basel, Switzerland) and yeast tRNA (Roche Diagnostics), 0.67 mg/mL herring sperm, 16% 20 x SSC and 0.3% SDS and applied to a cDNA microarray containing 1,800 rat genes on a slide glass (Asahi Technoglass Co., Tokyo, Japan). Three house-keeping genes (GAPDH, HPRT and b-actin) and rat unrelated traits (Lambda DNA) were also spotted on the slide as internal positive and negative controls, respectively. Hybridization was carried out twice to eliminate any dye bias. In one experiment, duplicate slides were hybridized with probes derived from CBDL and control rats that had been labeld with cyanine-3 and cyanine-5, respectively. In a replicate experiment, other duplicate slides were hybridized with probes derived from CBDL rats and control rats that had been labeled with cyanine-5 and cyanine-3, respectively (color swap). Then, the slides were cover-slipped and incubated in a sealed chamber (Asahi Technoglass Co.) for 16 hrs under a humidified (65 °C) condition. After being washed in low-stringency buffer (2 x SSC and 0.1% SDS), high-stringency buffer (0.2 x SSC and 0.1% SDS) and 0.2 x SSC and then rinsing with 99.5% ethanol, the slide was dried by centrifugation at low speed and used for scanning. Fluorescence was scanned by using a ScanArray 4000 (Packard BioChip Technologies, Billerica, MA) and quantified by using QuantArray Software (Packard BioChip Technologies, Billerica, MA). Keywords: time-course
2003-12-13 | GSE898 | GEO
Project description:Dynamic changes in growth factor expression is observed after bile duct ligation at different time points compared to sham-treated mice, corresponding to increase in biliary cell proliferation and repair
2022-05-24 | GSE182049 | GEO
Project description:BACKGROUND:Alpha-1-antitrypsin (AAT) deficiency (AATD) of Z, Mmalton, Siiyama type is associated with liver storage of the mutant proteins and liver disease. The Z variant can be diagnosed on isoelectric focusing (IEF) while Mmalton and Siiyama may be missed or misdiagnosed with this technique. Therefore, molecular analysis is mandatory for their characterization. In particular, that holds true for the Mmalton variant as on IEF profile it resembles the wild M2 subtype. METHODS:This is a retrospective analysis involving review of medical records and of liver biopsy specimens from a series of Mmalton, Z and Siiyama Alpha-1-antitrypsin deficiency patients. The review has been implemented by additional histological stains, electron microscopic observations and 3-D modeling studies of the sites of the mutations. RESULTS:Z, Mmalton and Siiyama liver specimen contained characteristic intrahepatocytic PAS-D globules. The globules differed in the three variants as only Mmalton cases showed dark basophilic precipitates within the AAT inclusions. The precipitates were visualized in haematoxylin-eosin (H.E.) stained preparations and corresponded to calcium precipitates as demonstrated by von Kossa staining. On immunohistochemistry, ZAAT inclusions were stained by polyclonal as well as monoclonal noncommercial anti-AAT antibody (AZT11), whilst Mmalton and Siiyama inclusion bodies remained negative with the monoclonal anti-Z antibody. 3-D protein analysis allowed to predict more severe misfolding of the Mmalton molecule as compared to Z and Siiyama that could trigger anomalous interaction with endoplasmic reticulum chaperon proteins, namely calcium binding proteins. CONCLUSIONS:Mmalton AAT inclusion bodies contain calcium precipitates inside them that allow the differential diagnosis with Siiyama and ZAAT inclusions in routine histological sections. The study has confirmed the specificity of the monoclonal AZT11 for the Z mutant. Thus, the combination of these two features is crucial for the distinction between the three variants and for predicting the genotype, whose confirmation would definitely require molecular analysis. Our study provides new data on the pathomorphogenesis of Mmalton inclusion bodies whose mineralization could play a central role in disease pathogenesis of Mmalton that is distinct from the Z and Siiyama variants. Calcium is known to be a major effector of cell death either via the increased intracellular concentration or the alteration of homeostasis.
| S-EPMC5956786 | biostudies-literature
Project description:UnlabelledHuman chronic cholestatic liver diseases are characterized by cholangiocyte proliferation, hepatocyte injury, and fibrosis. Yes-associated protein (YAP), the effector of the Hippo tumor-suppressor pathway, has been shown to play a critical role in promoting cholangiocyte and hepatocyte proliferation and survival during embryonic liver development and hepatocellular carcinogenesis. Therefore, the aim of this study was to examine whether YAP participates in the regenerative response after cholestatic injury. First, we examined human liver tissue from patients with chronic cholestasis. We found more-active nuclear YAP in the bile ductular reactions of primary sclerosing cholangitis and primary biliary cirrhosis patient liver samples. Next, we used the murine bile duct ligation (BDL) model to induce cholestatic liver injury. We found significant changes in YAP activity after BDL in wild-type mice. The function of YAP in the hepatic response after BDL was further evaluated with liver-specific Yap conditional deletion in mice. Ablating Yap in the mouse liver not only compromised bile duct proliferation, but also enhanced hepatocyte necrosis and suppressed hepatocyte proliferation after BDL. Furthermore, primary hepatocytes and cholangiocytes isolated from Yap-deficient livers showed reduced proliferation in response to epidermal growth factor in vitro. Finally, we demonstrated that YAP likely mediates its biological effects through the modulation of Survivin expression.ConclusionOur data suggest that YAP promotes cholangiocyte and hepatocyte proliferation and prevents parenchymal damage after cholestatic injury in mice and thus may mediate the response to cholestasis-induced human liver disease.
| S-EPMC3431197 | biostudies-literature
Project description:Hepatic stellate cells (HSCs) play pivotal roles in hepatic fibrosis as they synthesize glial fibrillary acidic protein (GFAP), which is increased in activated HSCs. GFAP-expressing HSCs and myofibroblasts accumulate in and around hepatic fibrosis lesions. Peptidylarginine deiminase 2 (PAD2) is responsible for the citrullination of GFAP (cit-GFAP). However, the involvement of PAD2 and cit-GFAP in hepatic fibrosis remains unclear. To determine the expression of PAD2 and cit-GFAP in hepatic fibrosis, C57BL/6 mice underwent bile duct ligation (BDL) or a sham operation. In BDL livers, the expression of PAD2 and its enzyme activity were significantly increased compared with controls. In addition, PAD2-postitive cells were rarely observed in only the portal vein and the small bile duct in sham-operated livers, whereas an increased number of PAD2-positive cells were detected in the bile duct and Glisson's sheath in BDL livers. Interestingly, PAD2 was colocalized with ?-SMA-positive cells and CK19-positive cells in BDL livers, indicating upregulated PAD2 in activated HSCs and portal fibroblasts of the livers of BDL mice. We also found that citrullinated proteins were highly accumulated in the livers of BDL mice compared with controls. Moreover, the expression level of GFAP and the amount of cit-GFAP were higher in BDL livers than in control livers. In correlation with PAD2 localization, cit-GFAP was observed in ?-SMA-positive and CK19-positive cells in the livers of BDL mice. These results suggest that the increased expression and activation of PAD2 along with increased citrullinated proteins, specifically cit-GFAP, may play important roles in the pathogenesis of hepatic fibrosis.
| S-EPMC6072123 | biostudies-literature
Project description:Tetrathiomolybdate (TM), a potent copper-chelating drug, was initially developed for the treatment of Wilson's disease. Our working hypothesis is that the fibrotic pathway is copper-dependent. Because biliary excretion is the major pathway for copper elimination, a bile duct ligation (BDL) mouse model was used to test the potential protective effects of TM. TM was given in a daily dose of 0.9 mg/mouse by means of intragastric gavage 5 days before BDL. All the animals were killed 5 days after surgery. Plasma liver enzymes and total bilirubin were markedly decreased in TM-treated BDL mice. TM also inhibited the increase in plasma levels of tumor necrosis factor (TNF)-alpha and transforming growth factor (TGF)-beta1 seen in BDL mice. Cholestatic liver injury was markedly attenuated by TM treatment as shown by histology. Hepatic collagen deposition was significantly decreased, and it was paralleled by a significant suppression of hepatic smooth muscle alpha-actin and fibrogenic gene expression in TM-treated BDL mice. Although the endogenous antioxidant ability was enhanced, oxidative stress as shown by malondialdehyde and 4-hydroxyalkenals, hepatic glutathione/oxidized glutathione ratio, was not attenuated by TM treatment, suggesting the protective mechanism of TM may be independent of oxidative stress. In summary, TM attenuated BDL-induced cholestatic liver injury and fibrosis in mice, in part by inhibiting TNF-alpha and TGF-beta1 secretion. The protective mechanism seems to be independent of oxidative stress. Our data provide further evidence that TM might be a potential therapy for hepatic fibrosis.
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