studies

884A AASLD ABSTRACTS HEPATOLOGY, October, 2015
by the same parameters as above. We conclude that simultaneous
targeting of peripheral CB 1
R and iNOS by novel hybrid
inhibitors offers improved efficacy and safety in the pharmacotherapy
of liver fibrosis.
Disclosures:
Resat Cinar - Employment: National Institutes of Health; Patent Held/Filed:
National Institutes of Health
George Kunos - Patent Held/Filed: US Patent
The following authors have nothing to disclose: Malliga Iyer, Ziyi Liu, Tony Jourdan,
Zongxian Cao, Katalin Erdelyi, Grzegorz Godlewski, Gergo Szanda, Jie
Liu, Pal Pacher, Kenner Rice
1375
Proof of liver regeneration after resolution of fibrosis by
siRNA against HSP47 encapsulated in vitamin A-coupled
liposome employing CCl 4
mice engineered with
Sox9-Cre ERT2 /ROSA-YFP genes
Naoki Fujitani 1 , Akihiro Yoneda 2 , Yuya Murakami 1 , Miyuki
Nishimura 1 , Miyono Miyazaki 3 , Keiko Kajiwara 3 , Kenjiro
Minomi 3 , Yasuaki Tamura 2 , Yoshiro Niitsu 1 ; 1 School of Medicine,
Department of Molecular Exploration, Sapporo Medical University,
Sapporo, Japan; 2 Department of Molecular Therapeutics, Center
for Food and Medical Innovation, Institute for the Promotion of
Business-Regional Collaboration, Sapporo, Japan; 3 Hokkaido Laboratory,
Nitto Denko Corporation, Sapporo, Japan
Background and Aim We have previously shown that hepatic
fibrosis could be resolved by the administration of siRNA
against HSP47 encapsulated in vitamin A-coupled liposome
(VA-lip-siRNA HSP47) in various liver cirrhosis rodent models
(Sato et al. Nat. Biotechnol. 2008). However, a real clinical
benefit of such anti-fibrosis modality would be a recovery of
impaired liver function with tissue regeneration. We therefore,
in the present study intended to demonstrate that fibrotic liver
is capable to regenerate after resolution of fibrosis, employing
bile duct ligation (BDL) model rats, dimthylnitrosoamine (DMN)
model rats and carbon tetrachloride (CCl 4
) model of transgenic
mice carrying Sox9-Cre ERT2 /ROSA-YFP genes. Methods BDL
and DMN models of rats were established as reported earlier
(Sato et al. Nat. Biotechnol. 2008). Transgenic mice with
Sox9-Cre ERT2 /ROSA-YFP genes were produced by repeated
crossing of Sox9-Cre ERT2 mice onto ROSA-YFP mice obtained
from The Jackson Laboratory. Fibrosis was induced with the
intraperitoneal administration of CCl 4
for 10 weeks. VA-lipsiRNA
HSP47 was formulated as described earlier (Sato et al.
Nat. Biotechnol. 2008). Immuno-stainings for Sox9, αSMA,
EpCAM, YFP, HNF4α, BrdU and PCNA were carried out using
each specific antibodies. Results and discussion In the liver of
BDL rats, EpCAM positive; progenitors were localized at portal
area and were surrounded by deposited collagen and αSMA
positive cells, activated stellate cells (aHSC) as revealed by
immune-staining. In the liver of DMN rats which were administrated
with VA-lip-siRNA HSP47 for 4 times, both aHSC and
progenitors markedly diminished, and PCNA staining onto
HNF4α positive hepatocytes was significantly increased as
compared with that of control rats, suggesting a concept that
in the cirrhotic liver progenitors surrounded by aHSCs forming
niche like structure underwent differentiation into hepatocytes
upon destruction of niche structure with administration
of VA-lip-siRNA HSP47. To verify this concept, fate-tracing of
progenitor after resolution of fibrosis was conducted with Sox9-
Cre ERT2 /ROSA-YFP mice. When the CCl 4
mice were injected
with tamoxifen, followed by the treatment of VA-lip-siRNA
HSP47 for 10 times, initiating at 10 th week of CCl 4
administration,
and YFP expression was examined at 13 th week of
CCl 4
administration, HNF4α positive hepatocytes were clearly
immune-stained for YFP. Thus, present study demonstrates that
cirrhotic liver is capable of undergoing regeneration with our
anti-fibrosis treatment even under the condition of continuous
exposure to hepatotoxic agent.
Disclosures:
Miyuki Nishimura - Grant/Research Support: Nitto Denko
The following authors have nothing to disclose: Naoki Fujitani, Akihiro Yoneda,
Yuya Murakami, Miyono Miyazaki, Keiko Kajiwara, Kenjiro Minomi, Yasuaki
Tamura, Yoshiro Niitsu
1376
Oral hepatotropic DPP4 inhibitors attenuate NASH and
bilary fibrosis in part through macrophage modulation
Xiaoyu Wang 1 , Shih-Yen Weng 1 , Yong Ook Kim 1 , Thomas Klein 3 ,
Detlef Schuppan 1,2 ; 1 Institue of Translational Immunology, Mainz,
Germany; 2 Division of Gastroenterology, Boston, MA; 3 Boehringer
Ingelheim Pharma, Biberach an der Riss, Germany
Background and aims: Non-alcoholic steatohepatitis (NASH) is
characterized by steatosis, lobular inflammation and progressive
parenchymal fibrosis. Glucagon like peptide 1 (GLP-1)
analogues are proven drugs to treat type 2 diabetes and insulin
resistance. GLP-1 levels are increased by agents that inhibit cell
surface dipeptidyl peptidase-4 (DPP-4) which is ubiquitously
expressed and rapidly inactivates secreted GLP-1. Here, we
studied the effect of two hepatotropic DPP4-inhibitors on liver
inflammation and fibrosis in models of NASH and biliary fibrosis.
Methods: Linagliptin and Sitagliptin were administered
daily by oral gavage to Mdr2KO mice and to C57BL/6mice
fed a methionine and choline deficient (MCD) diet for 6 weeks.
Hepatic fibrosis was assessed by morphometric analysis of
Sirius red stained collagen and measurement of hydroxyproline
content. Hepatic inflammation and fibrosis were assessed by
semiquantitative immunohistochemistry. Fibrosis and inflammation
related transcript levels were measured by quantitative
real-time polymerase chain reaction (qPCR). Ex vivo analysis
of hepatic inflammatory cells was done by FACS. Results: In
the MCD model, both Linagliptin and Sitagliptin lead to a significantly
attenuated hepatic fat accumulation a reduction of
transcript levels of SREBP-1c, FAS and LPL, lowered serum ALT,
AST, ALP and LDH and significantly suppressed fibrosis and
inflammation related transcripts (aSMA, procollagen α1(I),
TIMP-1, TGFβ1, MMP-13, CD68, CCL3 and TNFα) compared
to vehicle-treated controls. This was accompanied by a significant
decrease of collagen area (Sirius red), αSMA, procollagen
type III, CD68, F4/80, Caspase 3, as determined by semiquantitive
immunohistochemistry. Boih DPPIV-inhibitors significantly
decreased hepatic CD11b + Ly6C + high (proinflammatory monocytes/macrophages)
and total F4/80 + cells (macrophages,
Kupffer cells) compared to mice on the MCD diet without treatment.
In nonsteatotic, spontaneously fibrotic Mdr2KO mice 10
mg/kg/day of Linagliptin significantly decreased procollagen
α1(I), TGFβ1, TIMP-1, MMP-8 transcript levels, and increased
putatively anti-fibrotic MMP-9 and -13. Conclusion: In mice fed
the MCD diet for 6 weeks, the DPPIV-inhibitors significantly
decreased inflammation and apoptosis. In both the MCD and
Mdr2KO model Linagliptin and Sitagliptin mildly decreased
liver injury and fibrosis through inhibition of inflammatory and
apoptosis pathways and via a decrease of proinflammatory
and profibrotic monocytes-macrophages.
Disclosures:
Thomas Klein - Employment: Boehringer Ingelheim
Detlef Schuppan - Consulting: Boehringer Ingelheim, Conatus, GLG, Merck,
Mitsubishi-Tanabe, Takeda, Silence, Glenmark, Isis; Grant/Research Support:
Boehringer-Ingelheim
The following authors have nothing to disclose: Xiaoyu Wang, Shih-Yen Weng,
Yong Ook Kim