studies

HEPATOLOGY, VOLUME 62, NUMBER 1 (SUPPL) AASLD ABSTRACTS 701A
factor for HCV infection in Vero cells. We also supplemented
Vero cells with the human Apolipoprotein E (ApoE), one of
the host factors important for virus production, and found that
ApoE supplementation enabled them to produce infectious
viruses. Finally, we established Vero cells expressing all these
factors, miR-122, human SR-B1 and ApoE. In this cell line, HCV
replication and infectious virus production could be observed
after HCVcc infection, indicating the reconstitution of the entire
life cycle of HCV. In conclusion, we demonstrate that miR-122,
SR-B1 and ApoE are necessary and sufficient for the reconstitution
of the complete HCV life cycle in non-human non-hepatic
Vero cells. These identified factors may be key determinants for
species and organ tropism of HCV infection. The established
novel HCV cell culture system with Vero cells will be useful to
understand the species and organ specific restriction factors of
HCV and to establish the mass culture system of viral particles
in non-cancer cells for HCV vaccine.
Disclosures:
The following authors have nothing to disclose: Asako Murayama, Nao Sugiyama,
Takaji Wakita, Takanobu Kato
1004
Tim-3 Inhibits Monocyte Functions via T-bet during Hepatitis
C Virus Infection
Ying Zhang 1 , Wenjing Yi 1 , Peixin Zhang 1 , Yan Liang 1 , Zhi Q.
Yao 2 , Zhansheng Jia 1 ; 1 Department of Infectious Diseases, Tangdu
Hospital, Fourth Military Medical University, Xi’an, China; 2 Department
of Internal Medicine, Division of Infectious Diseases, James
H. Quillen College of Medicine, East Tennessee State University,
Johnson City, TN
Hepatitis C virus (HCV) dysregulates innate immune responses
and induces persistent viral infection. We have previously
demonstrated that T-cell immunoglobulin and mucin domain
protein-3 (Tim-3) plays a pivotal role in negative regulation
of Toll-like receptor (TLR)-mediated innate immune responses.
While it is clear that Tim-3 is up-regulated on monocyte/macrophages
(M/M) during HCV infection, little is known about the
transcription factor that controls its expression in these cells.
Recent studies have revealed that Tim-3 expression is controlled
by the transcription factor T-box expressed in T cells (T-bet).
In this study, we further investigated the regulatory effects of
T-bet on Tim-3 transcription/translation in M/M during HCV
infection. Our results demonstrate that T-bet is constitutively
expressed on resting peripheral blood CD14+ M/M. M/M
from chronically HCV-infected subjects at baseline exhibit
significantly increased T-bet expression that is positively correlated
with the Tim-3 expression. Up-regulation of T-bet is also
observed in CD14+ M/M incubated with HCV+ Huh7.5 cell in
a time-dependent manner. In addition, HCV core protein can
induce T-bet expression in primary M/M or monocytic THP-1
cells, which is reversible by blocking the HCV core/gC1qR
interactions. Moreover, the HCV core-induced up-regulation of
T-bet and Tim-3 expression in CD14+ monocytes can be abrogated
by incubating the cells with SP600125 - an inhibitor for
JNK signaling pathway. Notably, silencing T-bet gene expression
decreases Tim-3 expression and enhances IL-12 secretion
as well as STAT-1 phosphorylation. These data suggest that
T-bet, induced by HCV core/gC1qR through JNK pathway,
enhances Tim-3 expression, leading to dampened M/M function
during chronic HCV infection. These findings provide new
information regarding Tim-3 transcriptional/translational regulation
via T-bet during HCV infection and novel immunotherapy
targets to combat this global epidemic viral infection.
Disclosures:
The following authors have nothing to disclose: Ying Zhang, Wenjing Yi, Peixin
Zhang, Yan Liang, Zhi Q. Yao, Zhansheng Jia
1005
Downregulation of let-7 family miRs is strongly associated
with increasing stages of hepatic fibrosis in chronic
hepatitis C (HCV) Genotype 3a
Manish C. Choudhary 1 , Sadaf Dar 1 , Ekta Gupta 2 , Jaswinder S.
Maras 1 , Syed N. Kazim 3 , Gayatri Ramakrishna 1 , Shiv K. Sarin 1 ;
1 Research, Institute of Liver and Biliary Science, New Delhi, India;
2 Virology, Institute of Liver and Biliary Sciences, New Delhi, India;
3 Centre for Interdisciplinary Research in Basic Sciences, Jamia
Milia Islamia, New Delhi, India
Background: Chronic hepatitis C (CHC) infection is a major
cause of liver fibrosis and end stage liver disease. Circulating
miRNAs have evolved as a reliable biomarker for various
pathological conditions. However, there is limited data on systematic
miRNA based biomarker study on hepatic fibrogenesis
in HCV genotype 3a (G3a). Aims: To profile circulating repertoire
of miRNAs in the plasma of HCV G3a infected patients
with different stages of hepatic fibrosis.Patients and Methods:
47 subjects with CHC and histological assessment by two independent
hepato-pathologists were categorized based on stage
of hepatic fibrosis: F0-1 (n=32), F3-4 (n=15) were compared
with healthy controls (n=28). Differentially expressed miRNAs
in plasma of CHC F0,1 (n=4) and F3,4 (n=4) were studied
and compared with controls using miRCURY LNA TM Serum/
Plasma Focus panel. These miRNAs were analysed in context
of fibrosis progression by making comparison between CHC
F0,1 and F3,4 patients. Subsequent validation of array data
was done by stem-loop RT-PCR. miRNA target gene prediction
was done by multiMiR R analysis. Results: 185 miRNAs were
screened and 31 miRNAs were commonly identified in all the
groups. 4 miRNA were significantly down regulated (p