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BMC Cancer
Published in: BMC Cancer 1/2019

Open Access 01-12-2019 | Hepatocellular Carcinoma | Research article

Induction of tolerogenic dendritic cells by activated TGF-β/Akt/Smad2 signaling in RIG-I-deficient stemness-high human liver cancer cells

Authors: Ming Zhong, Cheng Zhong, Wen Cui, Guanghui Wang, Gongpu Zheng, Li Li, Jing Zhang, Rujing Ren, Huijei Gao, Tingting Wang, Xin Li, Jiantu Che, Eiichi Gohda

Published in: BMC Cancer | Issue 1/2019

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Abstract

Background

Dendritic cells (DCs) alter their role from being immunostimulatory to immunosuppressive at advanced stages of tumor progression, but the influence of cancer stem cells (CSCs) and their secreted factors on generation and phenotypic change of DCs is unknown. Retinoic acid-inducible gene I (RIG-I) plays a role in regulation of other cellular processes including leukemic stemness besides its antiviral function.

Methods

Short hairpin RNA-mediated gene silencing was employed to generate stable RIG-I-knocked-down human hepatocellular carcinoma (HCC) cell lines. Expression levels of genes and proteins in spheres of those HCC cells were determined by quantitative real-time PCR and Western bot, respectively. Levels of secreted cytokines were measured by ELISA. The surface molecule expression levels of DCs were analyzed using flow cytometry. The ability of DCs to induce proliferation of T cells was assessed by a mixed lymphocyte reaction (MLR) assay.

Results

RIG-I-knocked-down HCC cells showed upregulated expression of stem cell marker genes, enhanced secretion of factors suppressing in vitro generation of DCs into the conditioned medium (CM), and induction of a phenotype of tumor-infiltrating DCs (TIDCs) with low levels of DC markers in their tumors in nude mice. Those DCs and TIDCs showed reduced MLR, indicating RIG-I deficiency-induced immunotolerance. The RIG-I-deficient HCC cells secreted more TGF-β1 than did reference cells. The tumors formed after injection of RIG-I-deficient HCC cells had higher TGF-β1 contents than did tumors derived from control cells. DC generation and MLR suppressed by the CM of RIG-I-deficient HCC cells were restored by an anti-TGF-β1 antibody. TGF-β1-induced phosphorylation of Smad2 and Akt was enhanced in RIG-I-deficient HCC spheres, knockdown of AKT gene expression abolishing the augmentation of TGF-β1-induced Smad2 phosphorylation. Akt and p-Akt were co-immunoprecipitated with Smad2 in cytoplasmic proteins of RIG-I-deficient spheres but not in those of control spheres, the amounts of co-immunoprecipitated Akt and p-Akt being increased by TGF-β stimulation.

Conclusions

Our results demonstrate that RIG-I deficiency in HCC cells induced their stemness, enhanced secretion and signaling of TGF-β1, tolerogenic TIDCs and less generation of DCs, and the results suggest involvement of TGF-β1 in those RIG-I deficiency-induced tolerogenic changes and involvement of CSCs in DC-mediated immunotolerance.
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Metadata
Title
Induction of tolerogenic dendritic cells by activated TGF-β/Akt/Smad2 signaling in RIG-I-deficient stemness-high human liver cancer cells
Authors
Ming Zhong
Cheng Zhong
Wen Cui
Guanghui Wang
Gongpu Zheng
Li Li
Jing Zhang
Rujing Ren
Huijei Gao
Tingting Wang
Xin Li
Jiantu Che
Eiichi Gohda
Publication date
01-12-2019
Publisher
BioMed Central
Published in
BMC Cancer / Issue 1/2019
Electronic ISSN: 1471-2407
DOI
https://doi.org/10.1186/s12885-019-5670-9

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