studies

HEPATOLOGY, VOLUME 62, NUMBER 1 (SUPPL) AASLD ABSTRACTS 305A
190
Knockout of the substance P receptor, neurokinin-1,
reduces liver fibrosis in cholestatic bile duct ligated (BDL)
mice
Ying Wan 2 , Nan Wu 3 , Julie Venter 3 , Yuyan Han 3 , Holly A.
Standeford 4 , Haibo Bai 2 , Shanika Avila 3 , Heather L. Francis 1 ,
Lindsey Kennedy 4 , Micheleine Guerrier 3 , Tina Kyritsi 3 , Tami Annable
4 , Shannon S. Glaser 1 , Gianfranco Alpini 1 , Fanyin Meng 1 ;
1 Research, S&W DDRC and Medicine, Central Texas Veterans
Health Care System and Scott & White, Temple, TX; 2 Operational
Funds, BaylorScott&White, Temple, TX; 3 Medicine, Texas A&M
University HSC, Temple, TX; 4 Research, Central Texas Veterans
Health Care System, Temple, TX
During the progression of cholestatic liver diseases, proliferating
cholangiocytes acquire neuroendocrine features and
secrete sensory neurotransmitters such as α-calcitonin gene
related peptide (α-CGRP) and substance P (SP). We have previously
shown that: (i) SP, a proteolytic product of tachykinin
(Tac1) that is deactivated by membrane metalloendopeptidase
(MME), stimulates cholangiocarcinoma growth by interacting
with its receptor, NK1R; and (ii) knockdown of NK1R decreases
biliary hyperplasia in cholestatic bile duct ligated (BDL) mice.
Thus, we aim to determine the autocrine/paracrine role of the
SP/NK1R axis on liver fibrosis in normal and cholestatic mice.
Methods: In normal and BDL WT and NK1RKO mice, we measured
serum SP levels and the mRNA expression of Tac1 and
MME in total liver samples and purified small and large cholangiocytes.
We measured: (i) liver fibrosis by Sirius Red staining
in liver sections; and (ii) the mRNA expression of the fibrosis
genes, α-SMA and fibronectin, in total liver tissues from: (i)
normal wild-type (WT) mice treated for 1 wk with saline or
SP (2.5 mmol/Kg BW by IP implanted minipumps); and (ii)
normal and BDL WT and NK1R KO mice. In vitro, murine
cholangiocyte lines were treated with BSA (basal) or SP (100
nM) in the absence/presence of L-733, 060 (25 μM, NK1R
antagonist) for 6-72 hr before evaluating the expression of the
aforementioned fibrotic genes. Results: There was an increase
in SP serum levels in normal WT mice treated with SP and in
BDL WT compared to normal WT mice. The expression of Tac1
increased in BDL WT mice, whereas MME mRNA expression
decreased in BDL WT compared to normal mice; an opposite
pattern was observed in BDL NK1RKO mice compared to
BDL WT mice. There was enhanced fibrosis and fibrosis gene
expression in normal WT mice treated with SP and in BDL WT
mice compared to normal WT mice. A significant decrease
in liver fibrosis and the expression of α-SMA and fibronectin
was observed in BDL NK1RKO mice compared to BDL WT
mice. Small cholangiocytes isolated from WT BDL mice and
NK1RKO mouse livers displayed a less fibrotic phenotype
when compared to large cholangiocytes. In addition, large
cholangiocytes derived from NK1RKO mice showed a significant
reduction in fibrotic markers relative to BDL WT mice.
Treatment of large murine cholangiocytes with SP increased the
expression of fibrosis genes, whereas L-733, 060 reduced the
SP-induced expression of α-SMA and fibronectin. Conclusion:
We propose that modulation of the synthesis of neurotransmitters
modulated by sensory innervation may be important in the
modulation of liver fibrosis during the progression of cholestatic
disorders.
Disclosures:
The following authors have nothing to disclose: Ying Wan, Nan Wu, Julie Venter,
Yuyan Han, Holly A. Standeford, Haibo Bai, Shanika Avila, Heather L. Francis,
Lindsey Kennedy, Micheleine Guerrier, Tina Kyritsi, Tami Annable, Shannon S.
Glaser, Gianfranco Alpini, Fanyin Meng
191
Loss of Cytoglobin Exacerbates Liver Fibrosis and Cancer
Development in Steatohepatitis by Activating the
Oxidative Stress Pathway
Thuy T. Le 2 , Yoshinari Matsumoto 2 , Tuong Thi Van Thuy 2 , Hoang
Hai 2 , Katsutoshi Yoshizato 1 , Norifumi Kawada 2 ; 1 Academic Advisor’s
Office, PhoenixBio Co.Ltd., Hiroshima, Japan; 2 Department
of Hepatology, Graduate School of Medicine, Osaka City University,
Osaka, Japan
Objective: To clarify the role of cytoglobin (CYGB) in the
development of liver fibrosis and cancer in humans and in a
mouse model of non-alcoholic steatohepatitis (NASH). Methods:
CYGB expression was assessed in patients with NASH or
hepatocellular carcinoma (HCC). Cygb-deficient mice (Cygb / )
and primary hepatic stellate cells (HSCs) isolated from Cygb /
or wild-type (WT) mice were characterized. A mouse NASH
model was generated in Cygb / and WT mice by feeding a
CDAA define diet for 8-32 weeks. A subset of mice was treated
with the antioxidant N-acetylcysteine (NAC). Histopathology
and gene expression were analyzed. Results: CYGB was
expressed in HSCs of intact human livers, and its expression
was reduced in NASH and HCC patients. In Cygb / mice, 67%
of those aged 1 to 2 years spontaneously exhibited abnormalities
and cancer development in multiple organs, including
the liver, lung, lymph nodes and heart, compared with 4.7%
in WT mice (p