Tumor Biol.

DOI 10.1007/s13277-016-5215-7

ORIGINAL ARTICLE

The suppressor of cytokine signaling 2 (SOCS2) inhibits tumor

metastasis in hepatocellular carcinoma
Meiling Cui 1 & Ji Sun 2 & Jianjing Hou 1 & Tao Fang 1 & Xin Wang 2 & Chao Ge 1 &
Fangyu Zhao 1 & Taoyang Chen 3 & Haiyang Xie 4 & Ying Cui 5 & Ming Yao 1 & Jinjun Li 1 &
Hong Li 1

Received: 21 March 2016 / Accepted: 14 July 2016

# International Society of Oncology and BioMarkers (ISOBM) 2016

Abstract Hepatocellular carcinoma (HCC) is a common
cause of cancer-related death worldwide, and its incidence
continues to increase. However, the mechanism underlying
the development and progression of HCC remains unknown.
The suppressor of cytokine signaling 2 (SOCS2) is a member
of the SOCS family and influences the carcinogenesis of mul-
tiple types of tumors, but the biological roles of SOCS2 in
HCC remain unclear. In this study, we found that SOCS2
expression was reduced in HCC tissues compared with
matched noncancerous liver tissues. Moreover, decreased
SOCS2 expression was significantly associated with the pres-
ence of intrahepatic metastasis and high histological grade in
HCC patients. Colony formation assays and 3-(4,5-dimethyl-
thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays
demonstrated that overexpression of SOCS2 or knockdown of
endogenous SOCS2 did not significantly affect cell prolifera-
tion and tumorigenicity in HCC cells in vitro and in vivo.
However, SOCS2 overexpression significantly inhibited the
migration and invasion of HCC cells in vitro and inhibited
metastasis in vivo. Consistent with these findings, the knock-
down of endogenous SOCS2 enhanced migration and inva-
sion in HCC cells in vitro. Our study demonstrated that
SOCS2 inhibited human HCC metastasis, and SOCS2 might
provide a new potential therapeutic strategy for treating HCC.
Keywords SOCS2 . Metastasis . Hepatocellular carcinoma
Introduction

Hepatocellular carcinoma (HCC) is a common cause of

Taoyang Chen, Haiyang Xie and Ying Cui contributed equally to this
work.
Electronic supplementary material The online version of this article
(doi:10.1007/s13277-016-5215-7) contains supplementary material,
which is available to authorized users.
* Hong Li
hongli@shsci.org
1
State Key Laboratory of Oncogenes and Related Genes, Shanghai
Cancer Institute, Renji Hospital, Shanghai Jiaotong University
School of Medicine, Shanghai 200032, China
2
Shanghai Medical College, Fudan University, Shanghai 200032,
China
3
Qi Dong Liver Cancer Institute, Qi Dong 226200, China
4
Department of General Surgery, the First Affiliated Hospital, School
of Medicine, Zhejiang University, Hangzhou 310003, China
5
Cancer Institute of Guangxi, Nanning 530027, China
cancer-related mortality worldwide with an increasing inci-
dence [1, 2]. Among 745,500 deaths due to liver cancer oc-
curred worldwide during 2012, approximately 50 % occurred
in China [3]. HCC has become one of the five leading causes
of cancer mortality among both men and women in China [4].
The main causes of death in patients with HCC are tumor
metastasis and recurrence [5]. However, the mechanisms un-
derlying the metastasis and recurrence of HCC remain un-
known. Cancer metastasis is a complicated process, and mul-
tiple factors associated with both biology and the environment
might contribute to the incidence of the spread of tumor cells
[6]. Therefore, it is imperative to investigate the development
and progression of HCC.
The suppressor of cytokine signaling 2 (SOCS2) is a mem-
ber of the SOCS family, which includes CIS and SOCS1–7
[7]. The SOCS proteins contain three structural domains: an
N-terminal region of variable length, a cytokine-inducible Src
homology 2 domain protein (SH2) domain, and an SOCS box

[8, 9]. Members of the SOCS family are considered important
factors in the development and progression of carcinoma.
Recent studies have demonstrated that SOCS1, SOCS3, and
SOCS6 are involved in the tumorigenesis and progression of
HCC [10–16]. SOCS1 was identified as a tumor suppressor in
HCC. Methylation of the SOCS1 promoter was inversely cor-
related with HB(s) Ag status and strongly associated with
HCC stemming from liver cirrhosis and with tumor size [11,
13–15]. The loss of SOCS3 might promote cell growth, mi-
gration, and invasion by enhancing the JAK/STAT and FAK
signaling pathways in human HCC [12, 16]. Qiu et al. evalu-
ated SOCS2 expression in 106 pairs of HCC tissues and
matched noncancerous liver tissues using qRT-PCR and
Western blot analysis and found that SOCS2 downregulation
was an independent prognostic indicator of overall and
disease-free survival in HCC. In addition, similar results were
observed with SOCS6 [10]. However, the functional role of
SOCS2 in HCC remains unclear.
In the present study, we found that SOCS2 expression was
significantly downregulated in HCC tissues and low levels of
SOCS2 were significantly associated with the presence of
intrahepatic metastasis and high histological grade in HCC
patients. Moreover, SOCS2 inhibited migration and invasion
in HCC cell lines.
Materials and methods
Clinical specimens
A total of 236 human HCC tissue samples were obtained from
patients who underwent surgical treatment at the Guangxi
Cancer Institute (Nanning, China), the Qidong Liver Cancer
Institute (Qidong, China), or the First Affiliated Hospital of
Zhejiang University (Hangzhou, China). The 236 HCC pa-
tients included 190 males and 46 females (mean age
50.9 years, ranging from 21 to 83 years). No patient received
preoperative chemotherapy or radiotherapy. All procedures
were performed under consensus agreements and in accor-
dance with the China Ethical Review Committee.
Immunohistochemistry
All tissue samples were fixed in 4 % phosphate-buffered neu-
tral formalin for at least 72 h and routinely embedded in par-
affin. Tissue microarrays (TMAs) were constructed and im-
munohistochemical staining of TMAs were performed as de-
scribed previously [5, 17]. Immunohistochemical analyses
were performed with the anti-SOCS2 polyclonal antibody
(SAB4300809) which was purchased from Sigma-Aldrich
(Louis, MO, USA).
Tumor Biol.
Evaluation of tissue microarray
All the slides were observed and photographed using an
Axioskop 2 microscope (Carl Zeiss, Oberkochen, Germany).
Considered both staining intensity (0, no color; 1, light yel-
low; 2, brownish yellow; 3, brownish black) and the percent-
age of positive cells (0, <10 %; 1, 10–25 %; 2, 25–50 %; 3,
50–75 %; 4, >75 %), immunohistochemical results of SOCS2
were scored 0 to 4. When the score of cancer tissue was higher
than the matched noncancerous liver tissues, we considered
that SOCS2 was overexpressed; when the score of cancer
tissue was lower than the matched noncancerous liver tissues,
we considered that SOCS2 was underexpressed; when the
score of cancer tissue was same with the matched noncancer-
ous liver tissues, we considered that SOCS2 was unchanged.
TCGA cohort
A total of 373 liver cancer patients who underwent a minimum
follow-up of 3 months from The Cancer Genome Atlas
(TCGA, https://tcga-data.nci.nih.gov/tcga/, updated
December 31, 2014) database (hereinafter referred to as the
TCGA cohort) were enrolled in this study for the survival and
clinicopathological features analyses. SOCS2 levels were
evaluated in 50 patients from the TCGA for whom SOCS2
messenger RNA (mRNA) expression data in both cancer tis-
sues and matched adjacent noncancerous tissues were
available.
Cell lines and cell culture
The human HCC lines PLC/PRF/5, Hep3B, and SK-Hep1
were obtained from the American Type Culture Collection
(Manassas, VA, USA). The Huh7 cell lines were purchased
from Riken Cell Bank (Tsukuba, Japan). MHCC-97L,
MHCC-97H, and MHCC-LM3 cell lines were provided by
the Liver Cancer Institute, Zhongshan Hospital of Fudan
University (Shanghai, China). The SMMC-7721 and Li-7 cell
lines were obtained from the cell bank of the Institute of
Biochemistry and Cell Biology of the Chinese Academy of
Sciences (Shanghai, China). The HCC-LY5 and HCC-LY10
cell lines were established in our laboratory. All cell lines were
cultivated in Dulbecco’s modified Eagle’s medium (DMEM)
(Sigma-Aldrich, St. Louis, MO, USA) with 10 % fetal bovine
serum (FBS) (Hyclone, Logan, UT, USA) at 37 °C in 5 %
CO2.
Quantitative real-time polymerase chain reaction
(qRT-PCR)
Total RNA was isolated from cells using TRIzol reagent
(Invitrogen, Carlsbad, CA, USA). Reverse transcription was
performed using the PrimeScript RT Reagent Kit (Perfect Real
Tumor Biol.
Time) (TaKaRa Biotechnology, Dalian, China) according to
the manufacturer’s instructions. Real-time PCR analysis was
performed using the following socs2 gene-specific primers:
forward, 5′-ATGACCCTGCGGTGCCT-3′, and reverse, 5′-
AAAGTTCCTTCTGGTGCCTCT-3′. The expression level
of human glyceraldehyde-3-phosphate dehydrogenase
(GAPDH) was used as an internal control: forward, 5′-
AGAAGGCTGGGGCTCATTTG-3′, and reverse, 5′-AGGG
GCCATCCACAGTCTTC-3′.
Western blotting
Total proteins were extracted from cell lysates and separated
using 10 or 15 % sodium dodecyl sulfate-polyacrylamide gel
electrophoresis (SDS-PAGE). The proteins were transferred to
polyvinylidene difluoride (PVDF) or nitrocellulose mem-
branes according to the manufacturer’s supplied protocol
(Sigma-Aldrich). The membrane was subsequently blocked
with 5 % nonfat milk and incubated with the specific primary
antibody against SOCS2 (Cell Signaling Technology,
Danvers, MA). β-Actin (Sigma-Aldrich) levels were used as
an internal control.
MTT assays
Approximately 2000 to 5000 cells per well were cultured in
triplicate in 96-well plates for 7 days. The cells were incubated
with 100 μl of MTT reagent (5 mg/ml, Sigma-Aldrich) for 4 h
every day. The crystals were dissolved in dimethylsulfoxide
(DMSO). The optical density (OD) value was measured at two
wavelengths (570 and 630 nm).
Colony formation assays
Approximately 2000 to 5000 cells per well were incubated in
triplicate in six-well plates for 2 weeks and fixed with 10 %
formaldehyde for 30 min at room temperature. The cells were
subsequently stained with Giemsa (Sigma-Aldrich) for
30 min, and the cell colonies were quantified.
RNA interference and overexpression of SOCS2
The lentiviral vectors expressing shSOCS2-1, shSOCS2-2,
and shNC were respectively constructed by inserting annealed
oligos (shSOCS2-1, 5′-GAAGGAACTTTCTTGATTA-3′;
shSOCS2-2, 5′-GGACCAACTAATCTTCGAA-3′; shNC,
5′-TTCTCCGAACGTGTCACGT-3′) into the lentiviral vec-
tor pLVTHM (Addgene, Cambridge, MA) as described on the
Addgene website. The lentiviral vector pWPXL (Addgene,
Cambridge, MA) was used to construct the overexpression
plasmid of SOCS2. Western blot and real-time PCR analyses
were used to confirm the efficiency of SOCS2 gene silencing
or overexpression.
In vitro migration and invasion assays
Equal numbers of cells were added to the upper chamber of a
transwell (BD Biosciences, Franklin Lakes, NJ, USA) in
serum-free media. DMEM containing 10 % FBS was added
to the bottom chambers, and the cells were incubated at 37 °C
for 24 or 48 h. Cells that had migrated from the upper chamber
to the lower chamber were fixed with paraformaldehyde and
stained with Giemsa solution or crystal violet.
In vivo metastasis assays
Approximately 6- to 8-week-old BALB/C-nu/nu nude mice
were randomly divided into two groups. Li-7 cells stably over-
expressing SOCS2 or expressing pWPXL were injected into
the left hepatic lobe, and 1 × 106 cells were used for each
mouse. After 6 weeks, the mice were euthanized. The lungs
and livers were resected and fixed in 10 % neutral phosphate-
buffered formalin. The tissues were embedded in paraffin and
stained with hematoxylin and eosin (HE) staining. All the
animal experiments were performed according to the require-
ments approved by the Shanghai Medical Experimental
Animal Care Commission.
Statistical analysis
Statistical analyses were conducted using Statistical Product
and Service Solutions (SPSS) 13.0 software. All results are
presented as the mean ± standard deviation (SD), and compar-
isons were evaluated using Student’s t test. Survival was ana-
lyzed using the Kaplan–Meier method. Statistical computa-
tions were performed by GraphPad Prism version 5.0.
p < 0.05 was considered statistically significant: *p < 0.05;
**p < 0.01; *** p < 0.001.
Results
SOCS2 is decreased and negatively correlated
with malignancies in HCC
SOCS2 protein expression in 236 pairs of HCC tissues and
matched noncancerous liver tissues was analyzed using im-
munohistochemistry (IHC) staining to explore the associa-
tions between SOCS2 expression and the clinicopathological
features of HCC patients. Compared with matched noncancer-
ous liver tissue, SOCS2 was significantly downregulated in
172/236 (72.88 %) and upregulated in 10/236 (4.24 %) of the
HCC tissue samples. No difference in SOCS2 was observed in
54/236 (22.88 %) of the HCC specimens (Fig. 1a, b). We also
evaluated the levels of SOCS2 mRNA in 29 pairs of HCC
tissues and matched noncancerous liver tissues using qRT-
PCR. These data demonstrated that SOCS2 was significantly
Tumor Biol.
 Tumor Biol.
ƒFig. 1 Decreased SOCS2 expression is associated with HCC
malignancy and progression. a The expression of SOCS2 in 236 pairs
of HCC tissues and matched noncancerous tissues was evaluated using
IHC (magnification ×40 and ×400). b Statistical analysis of SOCS2
expression in HCC tissues and matched noncancerous tissues. c qRT-
PCR was performed to measure the expression levels of the SOCS2 gene
in 29 pairs of HCC tissues and matched noncancerous tissues. d
Statistical analysis of SOCS2 mRNA levels in 50 pairs of HCC tissues
and matched noncancerous tissues in the TCGA cohort. e SOCS2 mRNA
was expressed at lower levels in HCC tissues derived from patients with
advanced stage (III–IV) HCC compared with patients with earlier disease
stages (I–II) in the TCGA cohort. f SOCS2 mRNA levels were inversely
correlated with pathological stage in the TCGA cohort. g Kaplan–Meier
analysis of the correlation between SOCS2 mRNA levels and overall
survival in the TCGA cohort
downregulated in HCC tissues (Fig. 1c), and similar results
were observed in the 50 pairs of HCC cases from the TCGA
cohort (Fig. 1d).
In addition, the downregulation of SOCS2 expression was
significantly associated with histological grade (p = 0.037)
and intrahepatic metastasis (p = 0.026) in the 236 pairs of
HCC tissue samples was evaluated. However, no correlation
was noted between SOCS2 expression and the other clinico-
pathological factors evaluated, including age, gender, tumor
size, serum alpha-fetoprotein (AFP), hepatitis B surface anti-
gen (HBsAg) levels, and the presence of cirrhosis (Table 1). In
the TCGA cohort, SOCS2 expression levels were inversely
correlated with serum AFP levels (p = 0.0003)
(Supplementary Fig. 1a), histological grade (p = 0.0052)
(Fig. 1e), pathological stage (p = 0.0291) (Fig. 1f), and vas-
cular invasion (p = 0.0057) (Supplementary Fig. 1b). Kaplan–
Meier survival analysis of the TCGA cohort revealed that
higher levels of SOCS2 were associated with more favorable
outcomes (Fig. 1g). Together, these findings indicate that low-
er levels of SOCS2 expression are associated with the malig-
nant progression of HCC.
SOCS2 had no significant effect on tumorigenicity of HCC
cells
To explore the function of SOCS2 in HCC progression, we
first examined the expression of SOCS2 in HCC lines using
qRT-PCR. We found that SOCS2 mRNA levels were relative-
ly low in Li-7, MHCC-97L, and Huh7 cells and higher in SK-
Hep1 and HCC-LY5 cells (Supplementary Fig. 1c). To eval-
uate the function of SOCS2 in HCC, we stably overexpressed
SOCS2 in Li-7, MHCC-97L, and Huh7 cells and knocked
down the endogenous expression of SOCS2 in SK-Hep1
and HCC-LY5 cells with two specific shRNAs using lentiviral
infection. The overexpression and knockdown efficiencies
were verified by qRT-PCR and Western blot analysis
(Fig. 2a, b and Supplementary Fig. 1d). We performed MTT
assays and colony formation assays to assess the effect of
SOCS2 on HCC cell proliferation. The results of these assays
demonstrated that SOCS2 did not significantly affect HCC
cell growth in vitro (Fig. 2c, d). Consistent with these findings,
overexpression of SOCS2 did not affect the tumorigenicity of
Li-7 cells in a xenograft mouse model (Fig. 2e).
SOCS2 significantly inhibits HCC cell migration
and invasion in vitro
Based on the observation that SOCS2 expression levels were
inversely correlated with the presence of intrahepatic metasta-
ses in HCC, we hypothesized that SOCS2 might contribute to
HCC metastasis. Therefore, we explored the effects of SOCS2
on the migratory and invasive abilities of HCC cells using
transwell migration and invasion assays. SOCS2 overexpres-
sion in the Li-7, MHCC-97L, and Huh7 HCC cell lines mark-
edly inhibited migration and invasive efficiencies of HCC
cells in vitro compared with the control empty vector-
transfected cells (Fig. 3a, b), whereas the knockdown of en-
dogenous SOCS2 in SK-Hep1 and HCC-LY5 cells signifi-
cantly enhanced migration and invasion (Fig. 3c, d). These
results suggested that SOCS2 significantly inhibits invasive-
ness of HCC cells in vitro.
SOCS2 remarkable suppressed metastasis in vivo
To investigate whether SOCS2 affects HCC metastasis
in vivo, 1 × 106 Li-7 cells stably overexpressing SOCS2 were
injected into the left hepatic lobe of nude mice. Li-7-pWPXL
cells were used as a control. After 6 weeks, we observed
metastatic tumors in the liver and lung of mice in both the
control and experimental groups. Five of the eight mice in
the control group that developed primary tumors (one mouse
did not develop a primary tumor) developed intrahepatic me-
tastases (71.43 %), and three mice in the control group devel-
oped lung metastases (42.86 %). However, only four of the
nine nude mice in the experimental group developed
intrahepatic metastasis (44.44 %) and one mouse developed
lung metastasis (11.11 %). Further analysis indicated that the
mice in the experimental group exhibited fewer metastatic
nodules compared with the control mice (Fig. 4a). The effi-
ciency of SOCS2 overexpression in the xenografts was veri-
fied by qRT-PCR and Western blot (Fig. 4b, c). Therefore,
these findings demonstrated that SOCS2 overexpression
markedly suppressed HCC metastasis in vivo.
Discussion
SOCS2, as other members of this family, is thought to be
involved in the regulation of cytokine signaling via the
Janus kinase (JAK)/signal transducer and activators of tran-
scription (STAT) signal transduction pathway [18]. Previous
study demonstrated that SOCS2 negatively regulates cell
 Tumor Biol.

Table 1 Relationship between

SOCS2 protein expression and Clinicopathological Number of SOCS2 immunostaining
clinicopathological features in features cases
HCC tissues Low expression, N High expression, N p value
(%) (%)

Age (years)
≤60 159 69 (68.32) 90 (67.16) 0.852
>60 76 32 (31.68) 44 (32.84)
Gender
Male 190 77 (76.24) 113 (83.70) 0.152
Female 46 24 (23.76) 22 (16.30)
Tumor size
≤5 cm 113 50 (50.51) 63 (48.46) 0.759
>5 cm 116 49 (49.49) 67 (51.54)
AFP (ng/ml)
≤20 79 28 (28.28) 51 (38.35) 0.110
>20 153 71 (71.72) 82 (61.65)
HBV infection
Negative 42 22 (22.22) 20 (15.38) 0.185
Positive 187 77 (77.78) 110 (84.62)
Cirrhosis
Absent 38 17 (16.83) 21 (15.56) 0.792
Present 198 84 (83.17) 114 (4.44)
Histological grade
I, II 119 43 (42.57) 76 (56.30) 0.037*
III, IV 117 58 (57.43) 59 (43.70)
Intrahepatic metastasis
Absent 161 61 (60.40) 100 (74.07) 0.026*
Present 75 40 (39.60) 35 (25.93)

p Value represents the probability from a chi-squared test for different expression of SOCS2 in HCC tissues (the
immunohistochemical score of 0–1 was considered as low expression; the score of 2–4 was considered as high
expression)
AFP alpha-fetoprotein, N number of cases
*p < 0.05

growth via several mechanisms, including binding to the insulin-
like growth factor 1 (IGF-1) receptor [19, 20] or STAT5b [21].
The involvements of SOCS2 in some malignancies have been
reported [22–27]. The expression of SOCS2 is low in pulmonary
adenocarcinoma [27], breast cancer, and ovarian cancers [22, 26]
and is associated with a low proliferation rate and high differen-
tiation in breast cancer [22]. SOCS2 deletion promotes the spon-
taneous development of intestinal tumors driven by mutations in
the adenomatous polyposis coli/beta-catenin pathway and acti-
vates AP-1 [25]. Interestingly, the function of SOCS2 in prostate
cancer is controversial [23, 24]. Iglesias-Gato et al. found that
SOCS2 protein levels are reduced in castration-resistant prostate
cancer (CRPC) and that SOCS2 suppresses proliferation and
invasion in prostate cancer cells [24]. However, Hoefer et al.
found that SOCS2 levels were markedly increased in prostate
cancer tissues and that SOCS2 knockdown significantly
inhibited tumor growth in vivo and in vitro [23].
In the present study, we analyzed 236 pairs of HCC tissues
and matched noncancerous liver tissues using IHC to explore
Fig. 2 SOCS2 did not significantly affect the tumorigenicity of HCC„
cells. a Western blot analysis was performed to assess the
overexpression efficiency of SOCS2 in Li-7, MHCC-97L, and Huh7.
β-Actin was used as a loading control. b qRT-PCR was conducted to
evaluate the knockdown efficiency of SOCS2 in HCC-LY5 and SK-
Hep1 cells stably expressing shSOCS2. c MTT assays were conducted
to evaluate the proliferation of Li-7, MHCC-97L, Huh7, HCC-LY5, and
SK-Hep1 cells stably transfected with SOC2, shSOCS2, or the control. d
Colony formation assays were performed in Li-7, MHCC-97L, Huh7,
HCC-LY5, and SK-Hep1 cell lines stably transfected with SOC2,
shSOCS2, or the control. e Li-7 cells infected with pWPXL-SOCS2
were orthotopically injected into nude mice. Mice injected with the
empty vector were used as a control. Tumors were resected from the
nude mice approximately 6 weeks after injection. Representative
images of mouse livers bearing tumors and the weight of these livers
are presented
Tumor Biol.
Tumor Biol.
 Tumor Biol.
ƒFig. 3 SOCS2 inhibited HCC cell migration and invasion in vitro. a Cell
migration in Li-7, MHCC-97L, and Huh7 cells stably transfected with
SOCS2 was evaluated using transwell assays in vitro. Cells transfected
with the empty vector were used as a control. b Cell invasion capability in
Li-7, MHCC-97L, and Huh7 cells stably transfected with SOCS2 was
assessed using transwell assays in vitro. Cells transfected with the empty
vector were used as a control. c Cell migration in HCC-LY5 and SK-
Hep1 cells stably transfected with shSOCS2 was assessed using transwell
assays in vitro. Cells transfected with shNC were used as a control. d Cell
invasion in HCC-LY5 and SK-Hep1 cells stably transfected with
shSOCS2 was evaluated using transwell assays in vitro. Cells
transfected with shNC were used as a control. *p < 0.05; **p < 0.01;
***p < 0.001
Fig. 4 SOCS2 markedly inhibited metastasis in vivo. a The effect of
ectopic SOCS2 expression on intrahepatic and lung metastasis in Li-7
cell-derived liver tumors. Representative images of liver and lung meta-
static nodules derived from Li-7 cells stably transfected with SOCS2 or
the control are presented. The numbers of intrahepatic and lung metastasis
the association between clinicopathological features of HCC
and SOCS2. Consistent with a previous study [10], SOCS2
protein levels were significantly downregulated in HCC tis-
sues and we observed a statistically significant decrease in the
expression of SOCS2 in HCC with advanced (stages III and
IV) stages or with intrahepatic metastasis. Moreover, patients
in the TCGA cohort with higher levels of SOCS2 expression
exhibited a significantly greater overall survival rate.
We established Li-7, MHCC-97L, and Huh7 cell lines sta-
bly overexpressing SOCS2 and SK-Hep1 and HCC-LY5 cells
stably overexpressing shRNA targeting SOCS2 to evaluate
nodules are presented in the right panels. (magnification 1, 2, 5, and 6:
×40; 3, 4, 7, and 8: ×200). b qRT-PCR was used to evaluate the effect of
SOCS2 overexpression in mouse liver tumors. c Western blot analysis
was performed to verify the effect of SOCS2 overexpression on mouse
liver tumors. *p < 0.05; **p < 0.01; ***p < 0.001

the function of SOCS2 in HCC. Similar to other members of
SOCS family, SOCS2 inhibited HCC cell migration and inva-
sion in vitro and inhibited metastasis in vivo. However,
SOCS2 did not affect HCC cell growth in our study.
Masuzaki et al. reported that SOCS2 plays a dual role in mod-
ulating the rate of hepatocyte proliferation by regulating
growth hormone (GH) signaling [28]. A recent study demon-
strates that SOCS2 decreased FLT3-ITD-mediated cell prolif-
eration [29]. SOCS2 is induced by various cytokines that also
activate STAT5, and SOCS2 regulates multiple signaling
pathways induced by GH, PRL, LIF, IL-2, IL-3, IL-6, EGF,
and IGF-1 [19, 30–32]. The mechanism by which SOCS2
affects HCC metastasis merits further investigation.
In conclusion, our study demonstrates that SOCS2 plays an
important role in HCC progression and that SOCS2 sup-
presses the metastatic potential of HCC cells in vivo and
in vitro. SOCS2 might represent a novel therapeutic strategy
in the treatment and prevention of HCC metastasis.
Acknowledgments This work was supported in part by grants from the
National KeySci-Tech Special Project of China (2013ZX10002-011),
National Natural Science Foundation of China (81201623, 81372192),
Innovation Program of Shanghai Municipal Education Commission
(13ZZ082), Key Discipline and Specialty Foundation of Shanghai
Municipal Commission of Health and Family Planning, and the
SKLORG Research foundation (91-12-04, 91-13-02).
Compliance with ethical standards
Conflicts of interest None
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