obeticholic-acid and Reperfusion-Injury

We have previously shown that obeticholic acid (OCA) upregulates the biliary excretion of asymmetric dimethylarginine (ADMA), an inhibitor of iNOS regulating the activity of matrix metalloproteinases (MMPs). Here, the effects of OCA on MMP-2 and MMP-9 activity in liver, bile and serum were evaluated after hepatic ischemia/reperfusion (I/R) injury.. Male Wistar rats (n = 20) were orally administered 10 mg/kg/day of OCA (5 days) and subjected to a 60-min ischemia and 60-min reperfusion. Bile, serum and tissue were collected for MMP-2 and MMP-9 activity quantification. The MMP regulator tissue reversion-inducing cysteine rich protein with Kazal motifs (RECK), tissue inhibitor of metalloproteinases (TIMPs), iNOS and biliary levels of LDH, γGT, glucose and ADMA were quantified.. In the I/R group, OCA administration reduced MMP-2 and MMP-9 in liver, bile and serum. A downregulation of tissue RECK and TIMPs, observed under I/R, were recovered by OCA. Immunohistochemical staining of hepatic tissue demonstrated that RECK expression is mainly localized in both cholangiocytes and hepatocytes. Hepatic iNOS positively correlated with tissue MMP-2 and MMP-9 activity. Biliary levels of LDH, γGT and glucose were lower in I/R rats treated with OCA; in bile, MMP levels positively correlated with LDH and γGT.. Thus, OCA administration confers protection to cholangiocytes via downregulation of biliary MMPs in livers submitted to I/R. This event is associated with hepatic RECK- and TIMP-mediated MMP decrease.

Topics: Animals; Biliary Tract; Chenodeoxycholic Acid; Liver; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Rats, Wistar; Reperfusion Injury

The farnesoid X receptor (FXR) plays an important role in bile acid metabolism, intestinal homeostasis, and intestinal ischemia-reperfusion (I/R) injury. We aimed to clarify the potential effects of FXR on intestinal epithelial cell tolerance to intestinal I/R injury and reveal the underlying mechanisms. An intestinal I/R injury model was established by the occlusion of the superior mesenteric artery for ischemia for 1 h, followed by reperfusion for 4 h in C57BL/6 (wild type [WT]) and FXR mice. The small intestine injury was assessed by histological analysis. Diamine oxidase and TNF-α levels in the serum were measured. Expressions of Bcl-2, Bax, caspase-3, and cystathionine-γ-lyase (CSE) were determined by immunohostochemical staining. Oxygen-glucose deprivation/reperfusion (OGD/R) was used to make injury in cultured Caco-2 cells pretreated with FXR agonist (INT-747) or DL-propargylglycine (PAG) for 24 h. Cell viability and the expressions of NF-κB, TNF-α, and IL-6 were assessed. Compared with WT I/R mice, FXR knockout mice exacerbated intestinal I/R injury, intestinal epithelial apoptosis, and inflammatory response. The I/R injury in WT mice was alleviated with INT-747 pretreatment. CSE expression increased after intestinal I/R injury in WT but not in FXR mice. INT-747 enhanced Caco-2 cell viability and inhibited inflammatory response by blocking the NF-κB pathway after OGD/R injury, which was diminished by a CSE-specific inhibitor (PAG). Thus, we demonstrated that FXR activation enhances intestinal epithelial cell tolerance to I/R by suppressing the inflammatory response and NF-κB pathway via CSE mediation.

Topics: Animals; Caco-2 Cells; Chenodeoxycholic Acid; Humans; Intestinal Diseases; Intestine, Small; Male; Mice; Mice, Knockout; NF-kappa B; Receptors, Cytoplasmic and Nuclear; Reperfusion Injury; Signal Transduction

We previously showed that increased asymmetric dimethylarginine (ADMA) biliary excretion occurs during hepatic ischemia/reperfusion (I/R), prompting us to study the effects of the farnesoid X receptor (FXR) agonist obeticholic acid (OCA) on bile, serum and tissue levels of ADMA after I/R.. Male Wistar rats were orally administered 10mg/kg/day of OCA or vehicle for 5 days and were subjected to 60 min partial hepatic ischemia or sham-operated. After a 60 min reperfusion, serum, tissue and bile ADMA levels, liver mRNA and protein expression of ADMA transporters (CAT-1, CAT-2A, CAT-2B, OCT-1, MATE-1), and enzymes involved in ADMA synthesis (protein-arginine-N-methyltransferase-1, PRMT-1) and metabolism (dimethylarginine-dimethylaminohydrolase-1, DDAH-1) were measured.. OCA administration induced a further increase in biliary ADMA levels both in sham and I/R groups, with no significant changes in hepatic ADMA content. A reduction in CAT-1, CAT-2A or CAT-2B transcripts was found in OCA-treated sham-operated rats compared with vehicle. Conversely, OCA administration did not change CAT-1, CAT-2A or CAT-2B expression, already reduced by I/R. However, a marked decrease in OCT-1 and increase in MATE-1 expression was observed. A similar trend occurred with protein expression.. The reduced mRNA expression of hepatic CAT transporters suggests that the increase in serum ADMA levels is probably due to decreased liver uptake of ADMA from the systemic circulation. Conversely, the mechanism involved in further increasing biliary ADMA levels in sham and I/R groups treated with OCA appears to be MATE-1-dependent.

Topics: Animals; Antiporters; Arginine; Biliary Tract; Blotting, Western; Carrier Proteins; Chenodeoxycholic Acid; Liver Diseases; Male; Nitric Oxide Synthase; Organic Cation Transport Proteins; Protein-Arginine N-Methyltransferases; Rats; Rats, Wistar; Real-Time Polymerase Chain Reaction; Receptors, Cytoplasmic and Nuclear; Reperfusion Injury; RNA, Messenger; Up-Regulation

The farnesoid X receptor (FXR) is abundantly expressed in the ileum, where it exerts an enteroprotective role as a key regulator of intestinal innate immunity and homeostasis, as shown in pre-clinical models of inflammatory bowel disease. Since intestinal ischemia reperfusion injury (IRI) is characterized by hyperpermeability, bacterial translocation and inflammation, we aimed to investigate, for the first time, if the FXR-agonist obeticholic acid (OCA) could attenuate intestinal ischemia reperfusion injury.. In a validated rat model of intestinal IRI (laparotomy + temporary mesenteric artery clamping), 3 conditions were tested (n = 16/group): laparotomy only (sham group); ischemia 60min+ reperfusion 60min + vehicle pretreatment (IR group); ischemia 60min + reperfusion 60min + OCA pretreatment (IR+OCA group). Vehicle or OCA (INT-747, 2*30mg/kg) was administered by gavage 24h and 4h prior to IRI. The following end-points were analyzed: 7-day survival; biomarkers of enterocyte viability (L-lactate, I-FABP); histology (morphologic injury to villi/crypts and villus length); intestinal permeability (Ussing chamber); endotoxin translocation (Lipopolysaccharide assay); cytokines (IL-6, IL-1-β, TNFα, IFN-γ IL-10, IL-13); apoptosis (cleaved caspase-3); and autophagy (LC3, p62).. It was found that intestinal IRI was associated with high mortality (90%); loss of intestinal integrity (structurally and functionally); increased endotoxin translocation and pro-inflammatory cytokine production; and inhibition of autophagy. Conversely, OCA-pretreatment improved 7-day survival up to 50% which was associated with prevention of epithelial injury, preserved intestinal architecture and permeability. Additionally, FXR-agonism led to decreased pro-inflammatory cytokine release and alleviated autophagy inhibition.. Pretreatment with OCA, an FXR-agonist, improves survival in a rodent model of intestinal IRI, preserves the gut barrier function and suppresses inflammation. These results turn FXR into a promising target for various conditions associated with intestinal ischemia.

Topics: Animals; Apoptosis; Autophagy; Biomarkers; Chenodeoxycholic Acid; Disease Models, Animal; Endotoxins; Ileum; Inflammation Mediators; Intestinal Mucosa; Intestines; Male; Permeability; Rats; Receptors, Cytoplasmic and Nuclear; Reperfusion Injury; Signal Transduction