Intrahepatic cholangiocarcinoma (ICC) is a highly fatal malignancy characterized by a vast amount of intra-tumoral fibroblasts. These fibroblasts are potentially implicated in maintaining the high aggressiveness of ICC, whereas its pro-cancer mechanisms remain scarcely reported. Here, by establishing co-culture models of ICC cells and hepatic stellate cells (HSCs), we identified that HSCs triggered the expression of nuclear receptor family 4 subgroup A member 2 (NR4A2), a transcription factor previously reported as a molecular switch between inflammation and cancer, in ICC cells. Functionally, NR4A2 promotes tumor proliferation, metastatic potentiality and represents an independent prognostic indicator for overall survival in ICC patients. Mechanistically, NR4A2 upregulates osteopontin (OPN) expression through transcriptional activation and thereby augments the activity of Wnt/β-catenin signaling. Intriguingly, in the context of co-culture, vascular endothelial growth factor (VEGF), a previously proved NR4A2 stimulus, not only enhances NR4A2 expression, but also can be blunted by the interference of the NR4A2-OPN axis. Altogether, this study suggests the NR4A2/OPN/Wnt signaling axis to be a pivotal executor of HSC-instigated cancer-promoting roles in ICC, and the NR4A2/OPN/VEGF positive feedback loop may help to reinforce the effect.
Subscribe to Journal
Get full journal access for 1 year
only $7.98 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Tax calculation will be finalised during checkout.
Rent or Buy article
Get time limited or full article access on ReadCube.
All prices are NET prices.
Kalluri R. The biology and function of fibroblasts in cancer. Nat Rev Cancer. 2016;16:582–98.
Smyth MJ, Ngiow SF, Ribas A, Teng MW. Combination cancer immunotherapies tailored to the tumour microenvironment. Nat Rev Clin Oncol. 2016;13:143–58.
Sanmamed MF, Chen L. A paradigm shift in cancer immunotherapy: from enhancement to normalization. Cell. 2018;175:313–26.
Chang CH, Qiu J, O’Sullivan D, Buck MD, Noguchi T, Curtis JD, et al. Metabolic competition in the tumor microenvironment is a driver of cancer progression. Cell. 2015;162:1229–41.
Cuiffo BG, Campagne A, Bell GW, Lembo A, Orso F, Lien EC, et al. MSC-regulated microRNAs converge on the transcription factor FOXP2 and promote breast cancer metastasis. Cell Stem Cell. 2014;15:762–74.
Bertuccio P, Malvezzi M, Carioli G, Hashim D, Boffetta P, El-Serag HB, et al. Global trends in mortality from intrahepatic and extrahepatic cholangiocarcinoma. J Hepatol. 2019;71:104–14.
Saha SK, Zhu AX, Fuchs CS, Brooks GA. Forty-year trends in cholangiocarcinoma incidence in the u.s.: intrahepatic disease on the rise. oncologist. 2016;21:594–9.
Bridgewater J, Galle PR, Khan SA, Llovet JM, Park JW, Patel T, et al. Guidelines for the diagnosis and management of intrahepatic cholangiocarcinoma. J Hepatol. 2014;60:1268–89.
Primrose JN, Fox RP, Palmer DH, Malik HZ, Prasad R, Mirza D. Capecitabine compared with observation in resected biliary tract cancer(BILCAP): a randomised, controlled, multicentre, phase 3 study. The Lancet Oncology. 2019;20:663–73.
Capecitabine Extends Survival for Biliary Tract Cancer. Cancer Discov. 2017;7:OF1.
Paule B, Herelle MO, Rage E, Ducreux M, Adam R, Guettier C, et al. Cetuximab plus gemcitabine-oxaliplatin (GEMOX) in patients with refractory advanced intrahepatic cholangiocarcinomas. Oncology. 2007;72:105–10.
Sirica AE, Gores GJ, Groopman JD, Selaru FM, Strazzabosco M, Wei Wang X, et al. Intrahepatic cholangiocarcinoma: continuing challenges and translational advances. Hepatology. 2019;69:1803–15.
Coulouarn C, Clement B. Stellate cells and the development of liver cancer: therapeutic potential of targeting the stroma. J Hepatol. 2014;60:1306–9.
Li L, Piontek K, Ishida M, Fausther M, Dranoff JA, Fu R. et al. Extracellular vesicles carry microRNA-195 to intrahepatic cholangiocarcinoma and improve survival in a rat model. Hepatology. 2017;65:501–14.
Jing CY, Fu YP, Huang JL, Zhang MX, Yi Y, Gan W, et al. Prognostic nomogram based on histological characteristics of fibrotic tumor stroma in patients who underwent curative resection for intrahepatic cholangiocarcinoma. oncologist. 2018;23:1482–93.
Cheng JT, Deng YN, Yi HM, Wang GY, Fu BS, Chen WJ, et al. Hepatic carcinoma-associated fibroblasts induce IDO-producing regulatory dendritic cells through IL-6-mediated STAT3 activation. Oncogenesis. 2016;5:e198.
Ji J, Eggert T, Budhu A, Forgues M, Takai A, Dang H, et al. Hepatic stellate cell and monocyte interaction contributes to poor prognosis in hepatocellular carcinoma. Hepatology. 2015;62:481–95.
Cadamuro M, Nardo G, Indraccolo S, Dall’olmo L, Sambado L, Moserle L, et al. Platelet-derived growth factor-D and Rho GTPases regulate recruitment of cancer-associated fibroblasts in cholangiocarcinoma. Hepatology. 2013;58:1042–53.
Gentilini A, Rombouts K, Galastri S, Caligiuri A, Mingarelli E, Mello T, et al. Role of the stromal-derived factor-1 (SDF-1)-CXCR4 axis in the interaction between hepatic stellate cells and cholangiocarcinoma. J Hepatol. 2012;57:813–20.
Fingas CD, Bronk SF, Werneburg NW, Mott JL, Guicciardi ME, Cazanave SC, et al. Myofibroblast-derived PDGF-BB promotes Hedgehog survival signaling in cholangiocarcinoma cells. Hepatology. 2011;54:2076–88.
Okabe H, Beppu T, Hayashi H, Horino K, Masuda T, Komori H, et al. Hepatic stellate cells may relate to progression of intrahepatic cholangiocarcinoma. Ann surgical Oncol. 2009;16:2555–64.
Han Y, Cai H, Ma L, Ding Y, Tan X, Liu Y, et al. Nuclear orphan receptor NR4A2 confers chemoresistance and predicts unfavorable prognosis of colorectal carcinoma patients who received postoperative chemotherapy. Eur J Cancer. 2013;49:3420–30.
McEvoy C, de Gaetano M, Giffney HE, Bahar B, Cummins EP, Brennan EP, et al. NR4A receptors differentially regulate NF-kappaB signaling in myeloid cells. Front Immunol. 2017;8:7.
Zhao D, Desai S, Zeng H. VEGF stimulates PKD-mediated CREB-dependent orphan nuclear receptor Nurr1 expression: role in VEGF-induced angiogenesis. International journal of cancer. J Int du Cancer. 2011;128:2602–12.
Crean D, Cummins EP, Bahar B, Mohan H, McMorrow JP, Murphy EP. Adenosine modulates NR4A orphan nuclear receptors to attenuate hyperinflammatory responses in monocytic cells. J Immunol. 2015;195:1436–48.
Sekiya T, Kashiwagi I, Inoue N, Morita R, Hori S, Waldmann H, et al. The nuclear orphan receptor Nr4a2 induces Foxp3 and regulates differentiation of CD4+ T cells. Nat Commun. 2011;2:269.
Han YF, Cao GW. Role of nuclear receptor NR4A2 in gastrointestinal inflammation and cancers. World J Gastroenterol. 2012;18:6865–73.
Mohan HM, Aherne CM, Rogers AC, Baird AW, Winter DC, Murphy EP. Molecular pathways: the role of NR4A orphan nuclear receptors in cancer. Clin Cancer Res. 2012;18:3223–8.
Karki K, Li X, Jin UH, Mohankumar K, Zarei M, Michelhaugh SK, et al. Nuclear receptor 4A2 (NR4A2) is a druggable target for glioblastomas. J Neurooncol. 2020;146:25–39.
Shigeishi H, Higashikawa K, Hatano H, Okui G, Tanaka F, Tran TT, et al. PGE(2) targets squamous cell carcinoma cell with the activated epidermal growth factor receptor family for survival against 5-fluorouracil through NR4A2 induction. Cancer Lett. 2011;307:227–36.
Han Y, Cai H, Ma L, Ding Y, Tan X, Chang W, et al. Expression of orphan nuclear receptor NR4A2 in gastric cancer cells confers chemoresistance and predicts an unfavorable postoperative survival of gastric cancer patients with chemotherapy. Cancer. 2013;119:3436–45.
Zagani R, Hamzaoui N, Cacheux W, de Reynies A, Terris B, Chaussade S, et al. Cyclooxygenase-2 inhibitors down-regulate osteopontin and Nr4A2-new therapeutic targets for colorectal cancers. Gastroenterology. 2009;137:1358–66 e1351-1353.
Zheng Y, Zhou C, Yu XX, Wu C, Jia HL, Gao XM, et al. Osteopontin promotes metastasis of intrahepatic cholangiocarcinoma through recruiting MAPK1 and mediating Ser675 phosphorylation of beta-Catenin. Cell Death Dis. 2018;9:179.
Wu L, Amarachintha S, Xu J, Oley F Jr., Du W. Mesenchymal COX2-PG secretome engages NR4A-WNT signalling axis in haematopoietic progenitors to suppress anti-leukaemia immunity. Br J Haematol. 2018;183:445–56.
Hashemi Goradel N, Najafi M, Salehi E, Farhood B, Mortezaee K. Cyclooxygenase-2 in cancer: a review. J Cell Physiol. 2019;234:5683–99.
Li P, Shan JX, Chen XH, Zhang D, Su LP, Huang XY, et al. Epigenetic silencing of microRNA-149 in cancer-associated fibroblasts mediates prostaglandin E2/interleukin-6 signaling in the tumor microenvironment. Cell Res. 2015;25:588–603.
Akeson A, Herman A, Wiginton D, Greenberg J. Endothelial cell activation in a VEGF-A gradient: relevance to cell fate decisions. Microvasc Res. 2010;80:65–74.
Raja R, Kale S, Thorat D, Soundararajan G, Lohite K, Mane A, et al. Hypoxia-driven osteopontin contributes to breast tumor growth through modulation of HIF1alpha-mediated VEGF-dependent angiogenesis. Oncogene. 2014;33:2053–64.
Ramchandani D, Weber GF. Interactions between osteopontin and vascular endothelial growth factor: Implications for cancer. Biochim Biophys Acta. 2015;1855:202–22.
Chakraborty G, Jain S, Kundu GC. Osteopontin promotes vascular endothelial growth factor-dependent breast tumor growth and angiogenesis via autocrine and paracrine mechanisms. Cancer Res. 2008;68:152–61.
Ahmed M, Sottnik JL, Dancik GM, Sahu D, Hansel DE, Theodorescu D, et al. An osteopontin/CD44 axis in RhoGDI2-mediated metastasis suppression. Cancer Cell. 2016;30:432–43.
Dai J, Peng L, Fan K, Wang H, Wei R, Ji G, et al. Osteopontin induces angiogenesis through activation of PI3K/AKT and ERK1/2 in endothelial cells. Oncogene. 2009;28:3412–22.
Urtasun R, Lopategi A, George J, Leung TM, Lu Y, Wang X, et al. Osteopontin, an oxidant stress sensitive cytokine, up-regulates collagen-I via integrin alpha(V)beta(3) engagement and PI3K/pAkt/NFkappaB signaling. Hepatology. 2012;55:594–608.
Kothari AN, Arffa ML, Chang V, Blackwell RH, Syn WK, Zhang J, Mi Z, Kuo PC. Osteopontin-A Master Regulator of Epithelial-Mesenchymal Transition. J Clin Med. 2016;5:39. https://doi.org/10.3390/jcm5040039.
Weber CE, Kothari AN, Wai PY, Li NY, Driver J, Zapf MA, et al. Osteopontin mediates an MZF1-TGF-beta1-dependent transformation of mesenchymal stem cells into cancer-associated fibroblasts in breast cancer. Oncogene. 2015;34:4821–33.
Zhu Y, Yang J, Xu D, Gao XM, Zhang Z, Hsu JL, et al. Disruption of tumour-associated macrophage trafficking by the osteopontin-induced colony-stimulating factor-1 signalling sensitises hepatocellular carcinoma to anti-PD-L1 blockade. Gut. 2019;68:1653–66.
Sulpice L, Rayar M, Desille M, Turlin B, Fautrel A, Boucher E, et al. Molecular profiling of stroma identifies osteopontin as an independent predictor of poor prognosis in intrahepatic cholangiocarcinoma. Hepatology. 2013;58:1992–2000.
Lammi J, Huppunen J, Aarnisalo P. Regulation of the osteopontin gene by the orphan nuclear receptor NURR1 in osteoblasts. Mol Endocrinol.2004;18:1546–57.
This project is supported by the National Natural Science Foundation of China (NO.81772510; NO.81902419; NO.81902993; NO. 82072677, NO. 82072672); National Key Research and Development Program of China (NO.2017YFC0908101 and NO.2017YFC0908102); Research Programs of Science and Technology Commission Foundation of Shanghai (NO.18XD1401100); Medical and Health Key Programs of Science and Technology Plan of Xiamen (NO.3502Z20191105); 2020 Youth Talent Program of Obstetrics & Gynecology Hospital of Fudan University obtained by Dr. Yi-Peng Fu and Research and Development Program of Zhongshan Hospital, Fudan University (NO. 2020ZSFZ13, NO. 2019ZSFZ13, NO. 2018ZSFZ055, NO. ZSLC62, NO. ZHZS17, NO. 2019-275, NO. 2020-312, NO. 2021-353).
Conflict of interest
The authors declare no competing interests.
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
About this article
Cite this article
Jing, CY., Fu, YP., Zhou, C. et al. Hepatic stellate cells promote intrahepatic cholangiocarcinoma progression via NR4A2/osteopontin/Wnt signaling axis. Oncogene 40, 2910–2922 (2021). https://doi.org/10.1038/s41388-021-01705-9