studies

HEPATOLOGY, VOLUME 62, NUMBER 1 (SUPPL) AASLD ABSTRACTS 1241A
these mice, the anti-HCV effect and development of human
mononuclear cell chimerism by human PBMCs and IFN-α treatment
was completely negated, suggesting that IFN-γ produced
by NKT cells is essential not only for its anti-viral effect, but also
for the development of human lymphocyte chimerism in human
PBMCs and IFN-α treated mice. Conclusion: IFN-α stimulates
IFN-γ expression in type I NKT cells and enhances the inhibition
of HCV replication. Type I NKT cells might represent a new
therapeutic target for chronic hepatitis C patients.
Disclosures:
Kazuaki Chayama - Consulting: AbbVie; Grant/Research Support: Ajinomoto,
Astellas, Torii, Tsumura, Aska, Bayer, Zeria, Daiichi Sankyo, Dainippon Sumitomo,
Eisai, Eli Lily, Janssen, Kowa, Mitsubishi Tanabe, MSD, Nippon Kayaku,
Nippon Shinyaku, Otsuka, Roche, Takeda, Toray; Speaking and Teaching:
Ajinomoto, AbbVie, Abott, Astellas, AstraZeneca, Aska, Bayer, BMS, Chugai,
Daiichi Sankyo, Dainippon Sumitomo, Eisai, J & J, Jimro, Miyarisan, MSD, Nihon
Kayaku, Olympus
The following authors have nothing to disclose: Eisuke Miyaki, Michio Imamura,
Nobuhiko Hiraga, Takuro Uchida, Hiromi Kan, Masataka Tsuge, Hiromi Abe, C.
Nelson Hayes, Hiroshi Aikata, Chise Tateno
2121
Small specific-sized Hyaluronic Acid normalizes
TLR4-mediated signaling in Kupffer cells after chronic
ethanol exposure associated with M2 polarization
Paramananda Saikia 1 , Katherine A. Pollard 1 , Megan R. McMullen
1 , Laura E. Nagy 1,2 ; 1 Center For Liver Disease Research, Pathobiology,
Lerner Research Institute, Cleveland Clinic, Cleveland,
OH; 2 Molecular Medicine, Case Western Reserve University,
Cleveland, OH
Hyaluronan (HA) is the major glycosaminoglycan in the extracellular
matrix and ubiquitously present in many tissues. While
HA has been used as a biomarker for liver injury for decades,
the contribution of HA to alcoholic liver disease is not well
understood. During acute and chronic inflammation or tissue
injury, reactive oxygen species and matrix metalloproteinases
increase HA turnover, resulting in local and systemic accumulation
of HA fragments of different molecular weights. HA
is recognized as an important damage associated pattern
molecule (DAMP) that regulates innate immunity. However,
depending on their size, HA fragments can have either pro-inflammatory
or anti-inflammatory functions. Chronic alcohol
consumption is associated with an increase in the sensitivity of
hepatic macrophages to signaling via TLR4 associated with an
M1 polarization and enhanced expression of pro-inflammatory
cytokines. Here we investigated the impact of specific-sized
small HA on TLR4 mediated signaling in Kupffer cells after
chronic ethanol exposure to rats. Primary cultures of rat Kupffer
cells were isolated from rats chronically exposed to ethanol
via the Lieber-DeCarli diet or pair-fed control diets. Kupffer
cells from ethanol-fed rats were more sensitive to stimulation
with LPS, resulting in increased expression of mRNA for the
pro-inflammatory cytokine, TNFα. When Kupffer cells were
pre-treated with HA fragments for 5 hr prior to LPS challenge
for 60 min, specific small-sized HA normalized TNFα mRNA
expression in Kupffer cells from ethanol fed rats. In contrast,
higher molecular weight HA fragments had no effect on TNF-α
production of Kupffer cells. Normalization of TLR4 signaling
after treatment with small-sized HA was associated with an
increase expression of the M2 phenotypic gene arginase-1
(arg-1) and a decrease expression of the M1 associated gene
iNOS. In summary, these data demonstrate for the first time
that specific small-sized HA fragments promote polarization
of Kupffer cells to M2/anti-inflammatory phenotype to normalize
chronic ethanol induced sensitization of TLR4 signaling in
Kupffer cells.
Disclosures:
The following authors have nothing to disclose: Paramananda Saikia, Katherine
A. Pollard, Megan R. McMullen, Laura E. Nagy
2122
Novel in vivo and in vitro models of Hepatitis E virus
genotype 3 infectivity for chronic human HEV infection.
Martijn D. van de Garde 1 , Suzan D. Pas 3 , Guido van der Net 3 ,
Bart L. Haagmans 3 , Andre Boonstra 1 , Thomas Vanwolleghem 1,2 ;
1 Department of Gastroenterology and Hepatology, Erasmus MC
University Hospital, Rotterdam, Netherlands; 2 Gastroenterology
and Hepatology, University Hospital Antwerp, Antwerp, Belgium;
3 Viroscience, Erasmus Medical Center, Rotterdam, Netherlands
Hepatitis E virus (HEV) genotype 3 (gt3) infections are emerging
in western countries. The pathogenesis of HEV infection
with associated liver pathology and clinical disease is poorly
understood due to a lack of suitable model systems. Here we
apply a novel in vivo and in vitro model system to characterize
the infectivity of different HEV RNA containing human clinical
samples. Human-liver chimeric mice (uPA +/+ Nod-SCID-IL2Rγ -/- )
and human lung adenocarcinoma A549 cells were challenged
with HEV gt3 obtained from human plasma, feces or liver-biopsy
of 3 patients. Detection and quantification of HEV RNA
was performed in mouse serum, feces, bile and liver or in
A549 culture supernatant using RT-qPCR. High HEV RNA levels
(up to 8 log IU HEV RNA/gr) were consistently detected in
100% of chimeric mouse livers (n=10) from week 2-14 post
inoculation with human feces-derived HEV (6-8 logs IU). Feces
of these mice was positive for HEV RNA at least once during
follow-up, while HEV viremia was inconsistently detectable,
with maximum viral loads of 3.6 log IU/ml. HEV derived from
a cryopreserved liver biopsy (5.5 log IU) similarly resulted in
moderate to high HEV RNA levels in mouse feces, bile and
liver (n=2). In contrast, anti-HEV IgG/IgM negative, HEV RNA
positive plasma (6 log IU) was not infectious in any of the
inoculated chimeric animals (n=8). In these animals human
albumin plasma levels were stable and comparable to HEV-infected
chimeric mice, indicating a functional hepatocyte graft.
Infection of A549 cells with HEV RNA positive clinical samples
showed rapid and increasing HEV RNA titers in culture
supernatant within 7 days and could be maintained for more
than 7 passages. Consistent in vitro HEV infectivity was seen
with feces samples of all 3 patients, while only 2 of 3 human
serum samples could be cultured in vitro, showing less efficient
propagation. In conclusion, infectivity of feces derived human
HEV is higher compared to serum derived HEV both in vitro
and in vivo. The sustained HEV gt 3 liver infections induced in
chimeric mice, show preferential viral shedding towards mouse
bile and feces, mimicking the course of infection in humans.
Besides antiviral efficacy studies, both models will guide future
investigations into the biology of HEV infectivity with implications
towards transfusion transmitted HEV in humans.
Disclosures:
Andre Boonstra - Grant/Research Support: BMS, Janssen Pharmaceutics, Merck,
Roche, Gilead
The following authors have nothing to disclose: Martijn D. van de Garde, Suzan
D. Pas, Guido van der Net, Bart L. Haagmans, Thomas Vanwolleghem