miR-155 has also been shown to contribute to alcohol-induced liver injury through induction of tumor necrosis factor production in macrophages

miR-155 has also been shown to contribute to alcohol-induced liver injury through induction of tumor necrosis factor production in macrophages.16 Interestingly, the level of miR-155 is increased in serum and plasma in patients with alcoholic and inflammatory liver injuries.17,18 These observations suggest a potential role of miR-155 in liver injury and liver diseases. by lentiviral vectors. Eight-week-old KO mice were administrated lentiviral particles containing preCmiR-155 (LV-miR-155) (in the livers harvested from KO mice injected with LV-miR-155. C: Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels 7 days after injection with lentiviral particles. Data are expressed as means SD from three independent experiments (B and C). Original magnification, 100 (A). mmc2.pdf (185K) GUID:?84FEBED8-A3E5-45C0-AA23-15F56810799C Abstract Fas-induced apoptosis is involved in diverse liver diseases. Herein, we investigated the effect of deletion on Fas-induced liver injury. Wild-type (WT) Sigma-1 receptor antagonist 2 mice and knockout (KO) mice were i.p. administered with the anti-Fas antibody (Jo2) to determine animal survival and the extent of liver injury. After Jo2 injection, the KO mice exhibited prolonged survival versus the WT mice (KO mice showed Sigma-1 receptor antagonist 2 lower alanine aminotransferase and aspartate aminotransferase levels, less liver tissue damage, fewer apoptotic hepatocytes, and lower liver tissue caspase 3/7, 8, and 9 activities compared with the WT mice, indicating that deletion prevents Fas-induced hepatocyte apoptosis and liver injury. Hepatocytes isolated from KO mice also showed resistance to Fas-induced apoptosis, KO hepatocytes compared to WT hepatocytes. A miR-155 binding site was identified in the 3-untranslated region of Mcl-1 mRNA; was identified as a direct target of miR-155 in hepatocytes. Consistently, pretreatment with a siRNA specific for reversed deletionCmediated protection against Jo2-induced liver tissue damage. Finally, restoration of expression in KO mice abolished the protection against Fas-induced hepatocyte apoptosis. Taken together, these findings demonstrate that deletion of prevents Fas-induced hepatocyte apoptosis and liver injury through the up-regulation of is up-regulated in multiple immune cell lineages on stimulation with Toll-like receptor ligands, inflammatory cytokines, and specific antigens.5C9 Subsequent studies have shown that miR-155 also mediates functions outside the hematopoietic and immune systems.10,11 In the liver, miR-155 has been shown to play a role in hepatocarcinogenesis,12C15 although its mechanism of action remains to be further defined. miR-155 has also been shown to contribute to alcohol-induced liver injury through induction of tumor necrosis factor production in macrophages.16 Interestingly, the level of miR-155 is increased in serum and plasma in patients with alcoholic and inflammatory liver injuries.17,18 These observations suggest a potential role of miR-155 in liver injury and liver diseases. However, at present, the biological functions and mechanisms of miR-155 in liver cells have not been delineated. The current Rabbit Polyclonal to TSC2 (phospho-Tyr1571) study aimed to determine the effect and mechanism of miR-155 in Fas and lipopolysaccharide (LPS)/d-galactosamine (D-GalN)Cmediated liver injury in mice. Our data show that deletion of protects against Fas-induced hepatocyte apoptosis and liver injury but not LPS/D-GalNCinduced liver injury. The role of miR-155 in hepatocytes was demonstrated by studies using hepatocytes isolated from knockout (KO) mice. Myeloid cell leukemia-1 (Mcl-1) was identified as a direct target of miR-155 in hepatocytes. Our results reveal a novel role of miR-155 in hepatocytes for regulation of and protection against Fas-induced apoptosis. Sigma-1 receptor antagonist 2 Materials and Methods Animals C57BL/6 wild-type (WT) mice and KO mice were obtained from the Jackson Laboratory (Bar Harbor, ME). The mice were maintained at 22C under a 12-hour light/dark cycle and received food and water freely at the Tulane University Health Sciences Center Animal Facility (New Orleans, LA). The experimental procedures were performed according to the guidelines of the Institutional Animal Care and Use Committee of Tulane University. Experimental Protocol Male C57BL/6 WT and KO mice were used at the age of 8 weeks. For survival experiments, the mice were injected i.p. with 0.35 g/g of body weight Jo2 anti-Fas antibody (BD Bioscience, Franklin Lakes, NJ). Jo2 was dissolved in a sterile 1?Dulbecco’s phosphate-buffered saline (PBS; Sigma-Aldrich, St. Louis, MO). The animals were observed continuously for up to 24 hours after Jo2 injection and the time of death was recorded. To assess the extent of Jo2-induced liver injury, the mice were i.p. administered 0.5 g/g of body weight Jo2?and the animals were sacrificed at specific time points. The?liver tissues were rapidly excised, and the specimens were?immediately cut into small fragments and subjected to standard formalin fixation and paraffin embedding for histological evaluation and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate digoxigenin nick-end labeling (TUNEL). The remaining liver tissues were immediately frozen in liquid nitrogen and stored at ?80C. Blood sample was collected from mouse orbital and centrifuged at 800 for 15 minutes. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities were measured with an automatic analyzer at the Department of Clinical Chemistry, Tulane University Hospital. In separate experiments, 8-week-old male C57BL/6 WT and KO mice were pretreated with 7 mg/kg siRNA (Invitrogen, Grand Island, NY) or 7 mg/kg siRNA control (Invitrogen).