TY - JOUR
T1 - Effects of modulating M3 muscarinic receptor activity on azoxymethane-induced liver injury in mice
AU - Khurana, Sandeep
AU - Jadeja, Ravirajsinh
AU - Twaddell, William
AU - Cheng, Kunrong
AU - Rachakonda, Vikrant
AU - Saxena, Neeraj
AU - Raufman, Jean Pierre
N1 - Funding Information:
We thank Dr. Jürgen Wess, Molecular Signaling Section, Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, for kindly providing mice used in this work. We thank Grazyna Zaidel and Esther Lucas for technical assistance with immunostaining, and Xue-Min Gao for assistance with animal handling and mini-pump insertion. Sandeep Khurana is supported by NIH grant K08 DK081479 and the Baltimore Research and Education Foundation . Jean-Pierre Raufman is supported by NIH grants R01 CA107345 and R01 CA120407 . Neeraj Saxena is supported by NIH grants K01 DK077137 and R03 DK089130 . Vikrant Rachakonda was supported by T32 DK067872 .
PY - 2013
Y1 - 2013
N2 - Previously, we reported that azoxymethane (AOM)-induced liver injury is robustly exacerbated in M3 muscarinic receptor (M3R)-deficient mice. We used the same mouse model to test the hypothesis that selective pharmacological modulation of M3R activity regulates the liver injury response. Initial experiments confirmed that giving a selective M3R antagonist, darifenacin, to AOM-treated mice mimicked M3R gene ablation. Compared to vehicle controls, mice treated with the M3R antagonist had reduced survival and increased liver nodularity and fibrosis. We next assessed AOM-induced liver injury in mice treated with a selective M3R agonist, pilocarpine. After pilocarpine treatment, stimulation of post-M3R signaling in the liver was evidenced by ERK and AKT activation. In contrast to the damaging effects of the M3R antagonist, administering pilocarpine to AOM-treated mice significantly attenuated hepatic stellate cell activation, collagen deposition, bile ductule proliferation, and liver fibrosis and nodularity. As anticipated from these findings, livers from pilocarpine-treated mice exhibited reduced expression of key players in fibrosis (a1 collagen, a-smooth muscle actin, TGF-b1, PGDF, TGF-b1R, PGDFR) and decreased mRNA levels for molecules that regulate extracellular matrix formation (TIMP-1, TIMP-2, MMP-2, MMP-13). Cleaved caspase-3, nitrotyrosine and BrdU immunostaining provided evidence that pilocarpine treatment reduced hepatocyte apoptosis and oxidative stress, while increasing hepatocyte proliferation. Collectively, these findings identify several downstream mechanisms whereby M3R activation ameliorates toxic liver injury. These novel observations provide a proof-of-principle that selectively stimulating M3R activation to prevent or diminish liver injury is a therapeutic strategy worthy of further investigation.
AB - Previously, we reported that azoxymethane (AOM)-induced liver injury is robustly exacerbated in M3 muscarinic receptor (M3R)-deficient mice. We used the same mouse model to test the hypothesis that selective pharmacological modulation of M3R activity regulates the liver injury response. Initial experiments confirmed that giving a selective M3R antagonist, darifenacin, to AOM-treated mice mimicked M3R gene ablation. Compared to vehicle controls, mice treated with the M3R antagonist had reduced survival and increased liver nodularity and fibrosis. We next assessed AOM-induced liver injury in mice treated with a selective M3R agonist, pilocarpine. After pilocarpine treatment, stimulation of post-M3R signaling in the liver was evidenced by ERK and AKT activation. In contrast to the damaging effects of the M3R antagonist, administering pilocarpine to AOM-treated mice significantly attenuated hepatic stellate cell activation, collagen deposition, bile ductule proliferation, and liver fibrosis and nodularity. As anticipated from these findings, livers from pilocarpine-treated mice exhibited reduced expression of key players in fibrosis (a1 collagen, a-smooth muscle actin, TGF-b1, PGDF, TGF-b1R, PGDFR) and decreased mRNA levels for molecules that regulate extracellular matrix formation (TIMP-1, TIMP-2, MMP-2, MMP-13). Cleaved caspase-3, nitrotyrosine and BrdU immunostaining provided evidence that pilocarpine treatment reduced hepatocyte apoptosis and oxidative stress, while increasing hepatocyte proliferation. Collectively, these findings identify several downstream mechanisms whereby M3R activation ameliorates toxic liver injury. These novel observations provide a proof-of-principle that selectively stimulating M3R activation to prevent or diminish liver injury is a therapeutic strategy worthy of further investigation.
KW - Azoxymethane
KW - Darifenacin
KW - Liver injury
KW - Muscarinic receptors
KW - Pilocarpine
UR - http://www.scopus.com/inward/record.url?scp=84884907991&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84884907991&partnerID=8YFLogxK
U2 - 10.1016/j.bcp.2013.05.010
DO - 10.1016/j.bcp.2013.05.010
M3 - Article
C2 - 23707755
AN - SCOPUS:84884907991
SN - 0006-2952
VL - 86
SP - 329
EP - 338
JO - Biochemical Pharmacology
JF - Biochemical Pharmacology
IS - 2
ER -