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miR-150 Deficiency Protects against FAS-Induced Acute Liver Injury in Mice through Regulation of AKT.

Chen W, Han C, Zhang J, Song K, Wang Y, Wu T - PLoS ONE (2015)

Bottom Line: Although miR-150 is implicated in the regulation of immune cell differentiation and activation, it remains unknown whether miR-150 is involved in liver biology and disease.Luciferase reporter assays in hepatocytes transfected with the Akt1 or Akt2 3'-UTR reporter constructs (with or without mutation of miR-150 binding site) established Akt1 and Akt2 as direct targets of miR-150.Tail vein injection of lentiviral particles containing pre-miR-150 enhanced Jo2-induced liver injury in miR-150 KO mice.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, Tulane University School of Medicine,1430 Tulane Avenue SL-79, New Orleans, Louisiana, United States of America.

ABSTRACT
Although miR-150 is implicated in the regulation of immune cell differentiation and activation, it remains unknown whether miR-150 is involved in liver biology and disease. This study was performed to explore the potential role of miR-150 in LPS/D-GalN and Fas-induced liver injuries by using wild type and miR-150 knockout (KO) mice. Whereas knockout of miR-150 did not significantly alter LPS/D-GalN-induced animal death and liver injury, it protected against Fas-induced liver injury and mortality. The Jo2-induced increase in serum transaminases, apoptotic hepatocytes, PARP cleavage, as well as caspase-3/7, caspase-8, and caspase-9 activities were significantly attenuated in miR-150 KO mice. The liver tissues from Jo2-treated miR-150 KO mice expressed higher levels of Akt1, Akt2, total Akt, as well as p-Akt(Ser473) compared to the wild type livers. Pretreatment with the Akt inhibitor V reversed Jo2-induced liver injury in miR-150 KO mice. The primary hepatocytes isolated from miR-150 KO mice also showed protection against Fas-induced apoptosis in vitro (characterized by less prominent PARP cleavage, less nuclear fragmentation and less caspase activation) in comparison to hepatocytes from wild type mice. Luciferase reporter assays in hepatocytes transfected with the Akt1 or Akt2 3'-UTR reporter constructs (with or without mutation of miR-150 binding site) established Akt1 and Akt2 as direct targets of miR-150. Tail vein injection of lentiviral particles containing pre-miR-150 enhanced Jo2-induced liver injury in miR-150 KO mice. These findings demonstrate that miR-150 deficiency prevents Fas-induced hepatocyte apoptosis and liver injury through regulation of the Akt pathway.

No MeSH data available.


Related in: MedlinePlus

Effect of Akt inhibitor V on Fas-induced liver injury and hepatocyte apoptosis.(A) WT and miR-150 KO mice were injected intraperitoneally with vehicle or Akt inhibitor V (1mg/kg of body weight) 30 minutes before PBS administration (n = 3 each group). Mice were sacrificed 3 hours after PBS administration. Western blot showed that Akt inhibitor V treatment inhibited the phosphorylation of Akt. (B) WT and miR-150 KO mice were injected intraperitoneally with vehicle or Akt inhibitor V (1mg/kg of body weight) 30 minutes before Jo2 (0.5μg/g body weight) administration (n = 6 each group). Mice were sacrificed 3 hours after Jo2 administration. H&E staining (100×, scale bar 20μm) of formalin-fixed, paraffin-embedded liver tissues. (C) Serum levels of ALT and AST in mice with or without Akt inhibitor V pretreatment (data are expressed as mean ± SD **p<0.01). (D) TUNEL staining of the liver tissues from WT and miR-150 KO mice 3 hours after Jo2 injection (with or without Akt inhibitor V pretreatment). Representative photographs (100×, scale bar 20μm) are shown at the left panel; quantification of TUNEL-positive cells is shown at the right panel (the data are expressed as mean ± SD; *p<0.05, **p<0.01). (E) Immunostaining for cleaved caspase-3 in liver tissues from mice with or without Akt inhibitor V pretreatment (100×, scale bar 20μm). Quantification of cleaved caspase-3 positive cells is shown at the right panel (the data are expressed as mean ± SD; *p<0.05, **p<0.01). (F) Caspase-3/7, caspase-8, and caspase-9 activities in liver tissue from mice with or without Akt inhibitor V pretreatment (the data are expressed as mean ± SD; *p<0.05, **p<0.01).
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pone.0132734.g005: Effect of Akt inhibitor V on Fas-induced liver injury and hepatocyte apoptosis.(A) WT and miR-150 KO mice were injected intraperitoneally with vehicle or Akt inhibitor V (1mg/kg of body weight) 30 minutes before PBS administration (n = 3 each group). Mice were sacrificed 3 hours after PBS administration. Western blot showed that Akt inhibitor V treatment inhibited the phosphorylation of Akt. (B) WT and miR-150 KO mice were injected intraperitoneally with vehicle or Akt inhibitor V (1mg/kg of body weight) 30 minutes before Jo2 (0.5μg/g body weight) administration (n = 6 each group). Mice were sacrificed 3 hours after Jo2 administration. H&E staining (100×, scale bar 20μm) of formalin-fixed, paraffin-embedded liver tissues. (C) Serum levels of ALT and AST in mice with or without Akt inhibitor V pretreatment (data are expressed as mean ± SD **p<0.01). (D) TUNEL staining of the liver tissues from WT and miR-150 KO mice 3 hours after Jo2 injection (with or without Akt inhibitor V pretreatment). Representative photographs (100×, scale bar 20μm) are shown at the left panel; quantification of TUNEL-positive cells is shown at the right panel (the data are expressed as mean ± SD; *p<0.05, **p<0.01). (E) Immunostaining for cleaved caspase-3 in liver tissues from mice with or without Akt inhibitor V pretreatment (100×, scale bar 20μm). Quantification of cleaved caspase-3 positive cells is shown at the right panel (the data are expressed as mean ± SD; *p<0.05, **p<0.01). (F) Caspase-3/7, caspase-8, and caspase-9 activities in liver tissue from mice with or without Akt inhibitor V pretreatment (the data are expressed as mean ± SD; *p<0.05, **p<0.01).

Mentions: To further investigate the role of Akt for protection against Fas-induced liver injury, we assessed the extent of Jo2-induced liver injury in WT and miR-150 KO mice pretreated with Akt inhibitor V (Triciribine). The efficacy of Akt inhibitor V was confirmed by the observation that Akt inhibitor V treatment inhibited the phosphorylation of Akt in mice without Jo2 treatment (Fig 5A). To evaluate the effect of Akt inhibitor V on Jo2-induced liver injury, WT and miR-150 KO mice were pretreated with vehicle or Akt inhibitor V (1 mg/kg) 30 minutes prior to the administration of Jo2 (0.5 μg/g body weight) or PBS, and the animals were sacrificed 3 hours after Jo2 injection. We observed that pretreatment with Akt inhibitor V reversed Jo2-induced liver injury in miR-150 KO mice, as documented by histopathological examination of the liver tissues (Fig 5B), analysis for serum transaminases (Fig 5C), TUNEL assay (Fig 5D), immunostain for cleaved caspase-3 (Fig 5E), as well as assessment for caspase-3/7, caspase-8 and caspase-9 activities (Fig 5F). These findings suggest an important role of Akt for protection against Fas-induced liver injury in miR-150 KO mice.


miR-150 Deficiency Protects against FAS-Induced Acute Liver Injury in Mice through Regulation of AKT.

Chen W, Han C, Zhang J, Song K, Wang Y, Wu T - PLoS ONE (2015)

Effect of Akt inhibitor V on Fas-induced liver injury and hepatocyte apoptosis.(A) WT and miR-150 KO mice were injected intraperitoneally with vehicle or Akt inhibitor V (1mg/kg of body weight) 30 minutes before PBS administration (n = 3 each group). Mice were sacrificed 3 hours after PBS administration. Western blot showed that Akt inhibitor V treatment inhibited the phosphorylation of Akt. (B) WT and miR-150 KO mice were injected intraperitoneally with vehicle or Akt inhibitor V (1mg/kg of body weight) 30 minutes before Jo2 (0.5μg/g body weight) administration (n = 6 each group). Mice were sacrificed 3 hours after Jo2 administration. H&E staining (100×, scale bar 20μm) of formalin-fixed, paraffin-embedded liver tissues. (C) Serum levels of ALT and AST in mice with or without Akt inhibitor V pretreatment (data are expressed as mean ± SD **p<0.01). (D) TUNEL staining of the liver tissues from WT and miR-150 KO mice 3 hours after Jo2 injection (with or without Akt inhibitor V pretreatment). Representative photographs (100×, scale bar 20μm) are shown at the left panel; quantification of TUNEL-positive cells is shown at the right panel (the data are expressed as mean ± SD; *p<0.05, **p<0.01). (E) Immunostaining for cleaved caspase-3 in liver tissues from mice with or without Akt inhibitor V pretreatment (100×, scale bar 20μm). Quantification of cleaved caspase-3 positive cells is shown at the right panel (the data are expressed as mean ± SD; *p<0.05, **p<0.01). (F) Caspase-3/7, caspase-8, and caspase-9 activities in liver tissue from mice with or without Akt inhibitor V pretreatment (the data are expressed as mean ± SD; *p<0.05, **p<0.01).
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pone.0132734.g005: Effect of Akt inhibitor V on Fas-induced liver injury and hepatocyte apoptosis.(A) WT and miR-150 KO mice were injected intraperitoneally with vehicle or Akt inhibitor V (1mg/kg of body weight) 30 minutes before PBS administration (n = 3 each group). Mice were sacrificed 3 hours after PBS administration. Western blot showed that Akt inhibitor V treatment inhibited the phosphorylation of Akt. (B) WT and miR-150 KO mice were injected intraperitoneally with vehicle or Akt inhibitor V (1mg/kg of body weight) 30 minutes before Jo2 (0.5μg/g body weight) administration (n = 6 each group). Mice were sacrificed 3 hours after Jo2 administration. H&E staining (100×, scale bar 20μm) of formalin-fixed, paraffin-embedded liver tissues. (C) Serum levels of ALT and AST in mice with or without Akt inhibitor V pretreatment (data are expressed as mean ± SD **p<0.01). (D) TUNEL staining of the liver tissues from WT and miR-150 KO mice 3 hours after Jo2 injection (with or without Akt inhibitor V pretreatment). Representative photographs (100×, scale bar 20μm) are shown at the left panel; quantification of TUNEL-positive cells is shown at the right panel (the data are expressed as mean ± SD; *p<0.05, **p<0.01). (E) Immunostaining for cleaved caspase-3 in liver tissues from mice with or without Akt inhibitor V pretreatment (100×, scale bar 20μm). Quantification of cleaved caspase-3 positive cells is shown at the right panel (the data are expressed as mean ± SD; *p<0.05, **p<0.01). (F) Caspase-3/7, caspase-8, and caspase-9 activities in liver tissue from mice with or without Akt inhibitor V pretreatment (the data are expressed as mean ± SD; *p<0.05, **p<0.01).
Mentions: To further investigate the role of Akt for protection against Fas-induced liver injury, we assessed the extent of Jo2-induced liver injury in WT and miR-150 KO mice pretreated with Akt inhibitor V (Triciribine). The efficacy of Akt inhibitor V was confirmed by the observation that Akt inhibitor V treatment inhibited the phosphorylation of Akt in mice without Jo2 treatment (Fig 5A). To evaluate the effect of Akt inhibitor V on Jo2-induced liver injury, WT and miR-150 KO mice were pretreated with vehicle or Akt inhibitor V (1 mg/kg) 30 minutes prior to the administration of Jo2 (0.5 μg/g body weight) or PBS, and the animals were sacrificed 3 hours after Jo2 injection. We observed that pretreatment with Akt inhibitor V reversed Jo2-induced liver injury in miR-150 KO mice, as documented by histopathological examination of the liver tissues (Fig 5B), analysis for serum transaminases (Fig 5C), TUNEL assay (Fig 5D), immunostain for cleaved caspase-3 (Fig 5E), as well as assessment for caspase-3/7, caspase-8 and caspase-9 activities (Fig 5F). These findings suggest an important role of Akt for protection against Fas-induced liver injury in miR-150 KO mice.

Bottom Line: Although miR-150 is implicated in the regulation of immune cell differentiation and activation, it remains unknown whether miR-150 is involved in liver biology and disease.Luciferase reporter assays in hepatocytes transfected with the Akt1 or Akt2 3'-UTR reporter constructs (with or without mutation of miR-150 binding site) established Akt1 and Akt2 as direct targets of miR-150.Tail vein injection of lentiviral particles containing pre-miR-150 enhanced Jo2-induced liver injury in miR-150 KO mice.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, Tulane University School of Medicine,1430 Tulane Avenue SL-79, New Orleans, Louisiana, United States of America.

ABSTRACT
Although miR-150 is implicated in the regulation of immune cell differentiation and activation, it remains unknown whether miR-150 is involved in liver biology and disease. This study was performed to explore the potential role of miR-150 in LPS/D-GalN and Fas-induced liver injuries by using wild type and miR-150 knockout (KO) mice. Whereas knockout of miR-150 did not significantly alter LPS/D-GalN-induced animal death and liver injury, it protected against Fas-induced liver injury and mortality. The Jo2-induced increase in serum transaminases, apoptotic hepatocytes, PARP cleavage, as well as caspase-3/7, caspase-8, and caspase-9 activities were significantly attenuated in miR-150 KO mice. The liver tissues from Jo2-treated miR-150 KO mice expressed higher levels of Akt1, Akt2, total Akt, as well as p-Akt(Ser473) compared to the wild type livers. Pretreatment with the Akt inhibitor V reversed Jo2-induced liver injury in miR-150 KO mice. The primary hepatocytes isolated from miR-150 KO mice also showed protection against Fas-induced apoptosis in vitro (characterized by less prominent PARP cleavage, less nuclear fragmentation and less caspase activation) in comparison to hepatocytes from wild type mice. Luciferase reporter assays in hepatocytes transfected with the Akt1 or Akt2 3'-UTR reporter constructs (with or without mutation of miR-150 binding site) established Akt1 and Akt2 as direct targets of miR-150. Tail vein injection of lentiviral particles containing pre-miR-150 enhanced Jo2-induced liver injury in miR-150 KO mice. These findings demonstrate that miR-150 deficiency prevents Fas-induced hepatocyte apoptosis and liver injury through regulation of the Akt pathway.

No MeSH data available.


Related in: MedlinePlus