studies

902A AASLD ABSTRACTS HEPATOLOGY, October, 2015
1419
Serine Protease Omi/HtrA2 Reduces Oxidative Stress
and Preserves Mitochondrial Function in Liver Fibrosis
Seung Kew Yoon 1,2 , Wonhee Hur 1 , Joon Ho Lee 1 , Sung Woo
Hong 1 , Jung-Hee Kim 1 , Jung Eun Choi 1 , Sung Min Kim 1 , Eun Byul
Lee 1 ; 1 The Catholic University Liver Research Center & WHO Collaborating
Center of Viral Hepatitis, Seoul, Korea (the Republic
of); 2 Department of Internal Medicine, St. Mary’s Hospital, The
Catholic University of Korea, Seoul, Korea (the Republic of)
Background: Liver fibrosis is one of chronic liver diseases
caused by various causes. A mitochondrial serine protease
Omi/high temperature requirement protein A2 (HtrA2) is
involved in several disease, but has been poorly understood
in relation to liver fibrosis. The aim of this study is to investigate
the role of Omi/HtrA2 in the liver fibrosis. Methods: Protein
expression of Omi/HtrA2 was analyzed with liver tissues
from CCl - 4
induced mice and liver cirrhosis patients. Reactive
oxygen species (ROS) damage was measured by H 2
DCF-DA
and mito-Sox staining in isolated hepatocytes from mnd2 mice
which have the S276C mutation in the protease domain of
Omi/HtrA2. The Omi/HtrA2 gene was delivered by hydrodynamic
injection in the CCl 4
induced-liver fibrosis mice model
which had lower Omi/HtrA2 expression. The change of collagen
accumulation was measured by Masson’s trichrome (MT)
staining and hydorxyproline assay. Morphological alteration
of mitochondria was analyzed by using transmission electron
microscopy (TEM). Result: Protein expression of Omi/HtrA2
was decreased in liver tissues from CCl 4
induced mice and
liver cirrhosis patients. In mnd2 mice, oxidative stress increased
compared with wild type mice and abnormal morphology of
mitochondria was observed under TEM. Mitochondrial damage
induced with CCl 4
was protected by hydrodynamic based
gene delivery of Omi/HtrA2. In addition, the overexpression
of Omi/HtrA2 decreased collagen accumulation in the liver
tissue. Conclusion: Our findings suggest that Omi/HtrA2
overexpression modulates mitochondrial ROS generation in
liver fibrosis. Omi/HtrA2 gene delivery in liver fibrosis mice
model showed anti-oxidative effect through preserving liver
mitochondrial functions. Therefore, the modulation of Omi/
HtrA2 through the mitochondrial antioxidant defense mechanism
might be promising in anti-fibrotic therapeutic approach.
Disclosures:
The following authors have nothing to disclose: Seung Kew Yoon, Wonhee Hur,
Joon Ho Lee, Sung Woo Hong, Jung-Hee Kim, Jung Eun Choi, Sung Min Kim,
Eun Byul Lee
1420
Cancer associated fibroblasts in intrahepatic cholangiocarcinoma
is derived from portal fibroblasts, but
not hepatic stellate cells; immunohistochemical study in
human tissues
Naoki Uyama 1 , Rei A. Ito 1 , Ami Kurimoto 1 , Tomohiro Okamoto 1 ,
Akito Yada 1 , Hideaki Sueoka 1 , Seikan Hai 1 , Hisashi Kosaka 1 ,
Yuichi Kondo 1 , Ikuo Nakamura 1 , Kazuhiro Suzumura 1 , Yasukane
Asano 1 , Toshihiro Okada 1 , Tadamichi Hirano 1 , Junichi
Yamanaka 1 , Norifumi Kawada 2 , Jiro Fujimoto 1 ; 1 Hepato-biliary-pancreas
surgery, Hyogo College of Medicine, Nishinomiya,
Japan; 2 Department of Hepatology, Osaka City University Graduate
School of Medicine,, Osaka, Japan
(Background and aim) Cancer associated fibroblasts (CAFs),
a key player in the extracellular matrix production of cancer
tissue, is known to be involved in the regulation of cancer
behavior. Although origin of CAFs in intrahepatic cholangiocarcinoma
(ICC) is suspected to be composed of hepatic
stellate cells (HSCs), portal fibroblasts (PF), or bone marrow
derived cells such as fibrocytes, detailed analysis has lacked
so far. Until now, it has been reported that fascin is a specific
marker for HSC and fibrin-2 and Thy-1 are specific markers
for PFs, while α-SMA and PDGFRβ are expressed by both
cells. However, there has been no report on the expression
of PDGFRα in hepatic mesenchymal cells. In this study, we
performed immunohistochemistry in order to characterize
CAFs in human ICC. (Materials and Methods) Ten pairs of ICC
cancerous and noncancerous tissues were prepared. Tissue
sections were Zinc-fixed and paraffin-embedded. Immunohistochemistry
of α-SMA, PDGFRα, PDGFRβ, fascin, fibrin-2 and
Thy-1 in cancerous and noncancerous area was performed.
As for noncancerous area, peri-portal area and sinusoidal
area were intensively observed. To investigate the expression
of individual markers in PFs, HSCs and CAF, double-labeled
immunofluorescent staining with α-SMA was performed under
confocal microscope. (RESULTS) In immunohistochemistry of
noncancerous area of ICC, PDGFRα expression was detected
at periportal area, but not in sinusoidal area. Double immunofluorescent
staining for PDGFRα and α-SMA revealed that both
protein expressions were overlapped in PFs, but not in HSCs,
indicating PDGFRα as a novel marker for PFs. Further immunohistochemical
analyses showed PFs are characterized as
α-SMA + , PDGFRβ + , fascin - , fibrin-2 + and Thy-1 + and HSCs are
as α-SMA + , PDGFRβ + , fascin + , fibrin-2 - and Thy-1 - . In cancerous
area, immunohistochemistry revealed that cancer stroma
was positive for all of α-SMA, PDGFRα, PDGFRβ, fibrin- and
Thy-1. As for fascin, 3 samples were positive, but the others
were negative. α-SMA expression was overlapped with any of
PDGFRα, PDGFRβ, fibrin-2 or Thy-1 in CAFs. (Conclusion) Our
human study revealed that PDGFRα is a specific marker for
PFs. HSCs are α-SMA + , PDGFRβ + , fascin + , fibrin-2 - , Thy-1 - and
PDGFRα - , while PFs are α-SMA + , PDGFRβ + , fascin - , fibrin-2 + ,
Thy-1 + and PDGFRα + . CAFs associated with ICC are positive
for PF markers (fibrin-2 + , Thy-1 + and PDGFRα + ) in addition to
α-SMA and PDGFRβ. Taken together, it is plausible that CAFs
are derived from PFs rather than HSCs.
Disclosures:
Norifumi Kawada - Grant/Research Support: BMS, Chugai, Kowa; Speaking
and Teaching: MSD, Janssen
The following authors have nothing to disclose: Naoki Uyama, Rei A. Ito, Ami
Kurimoto, Tomohiro Okamoto, Akito Yada, Hideaki Sueoka, Seikan Hai, Hisashi
Kosaka, Yuichi Kondo, Ikuo Nakamura, Kazuhiro Suzumura, Yasukane Asano,
Toshihiro Okada, Tadamichi Hirano, Junichi Yamanaka, Jiro Fujimoto
1421
Human neutrophil peptide-1 enhances alcohol-induced
hepatocyte apoptosis in vivo and in vitro.
Rie Ibusuki 2,1 , Hirofumi Uto 2,3 , Masaya Kozono 2 , Kohei Oda 2 ,
Akihiko Oshige 2 , Kotaro Kumagai 2 , Seiichi Mawatari 2 , Tsutomu
Tamai 2 , Akihiro Moriuchi 2 , Hirohito Tsubouchi 4,5 , Takezaki
Toshiro 1 , Akio Ido 2,4 ; 1 Department of International Island and
Community Medicine, Kagoshima University Graduate School of
Medical and Dental Sciences, Kagoshima, Japan; 2 Digestive and
Lifestyle Diseases, Department of Human and Environmental Science,
Kagoshima University Graduate School of Medical and Dental
Sciences, Kagoshima, Japan; 3 Center for Digestive and Liver
Diseases, Miyazaki Medical Center Hospital,, Miyazaki, Japan;
4 Department of HGF Tissue Repair and Regenerative Medicine,
Kagoshima University Graduate School of Medical and Dental Sciences,
Kagoshima, Kagoshima, Japan; 5 Kagoshima City Hospital,,
Kagoshima, Japan
Background Neutrophil infiltration of the liver is a typical
feature of alcoholic liver injury and nonalcoholic steatohepatitis.
Human neutrophil peptide (HNP)-1 is an antimicrobial
peptide secreted by neutrophils. We previously reported that