Osteopontin promotes a cancer stem cell-like phenotype in hepatocellular carcinoma cells via an integrin–NF-κB–HIF-1α pathway

// Lei Cao 1, 2, 5, * , Xiaoyu Fan 2, * , Wei Jing 3 , Yingchao Liang 2 , Rui Chen 2 , Yingying Liu 2, 3 , Minhui Zhu 3 , Rongjie Jia 2 , Hao Wang 2 , Xueguang Zhang 5 , Yanyun Zhang 1 , Xuyu Zhou 3 , Jian Zhao 2, 4, 6 , Yajun Guo 2, 4, 6 1 School of Medicine, Shanghai Jiao Tong University, Shanghai, 200025, People's Republic of China 2 International Joint Cancer Institute, The Second Military Medical University, Shanghai, 200433, People's Republic of China 3 Changhai Hospital, The Second Military Medical University, Shanghai, 200433, People's Republic of China 4 PLA General Hospital Cancer Center, PLA Postgraduate School of Medicine, Beijing, 100853, People's Republic of China 5 Clinical Immunology Laboratory, The First Affiliated Hospital of Suzhou University, Suzhou, 215007, Jiangsu, People's Republic of China 6 Beijing Key Laboratory of Cell Engineering & Antibody, Beijing, 100853, People's Republic of China * These authors have contributed equally to this work Correspondence to: Jian Zhao, e-mail: zhaojian@smmu.edu.cn Xuyu Zhou, e-mail: xuyu2006@medmail.com.cn Keywords: Cancer stem cell, hepatocellular carcinoma, osteopontin, stemness Received: November 19, 2014      Accepted: January 08, 2015      Published: February 25, 2015 ABSTRACT There is increasing evidence to suggest that hepatocellular carcinomas (HCCs) are sustained by a distinct subpopulation of self-renewing cells known as cancer stem cells. However, the precise signals required for maintenance of stemness-like properties of these cells are yet to be elucidated. Here, we demonstrated that the level of oncoprotein osteopontin (OPN) in tumor cells of the edge of bulk tumors was significantly correlated with the clinical prognosis of patients with HCC. OPN was highly expressed in side population fractions of HCC cell lines, as well as in dormant cells, spheroids and chemo-resistant cancer cells, all of which are considered as having stemness-like cellular features. Depletion of OPN in HCC cell lines resulted in a reduction in the proportion of side population fractions, formation of hepato-spheroids, expression of stem-cell-associated genes and decreased tumorigenecity in immunodeficient mice. Mechanistically, OPN was demonstrated to bind to integrin α v β 3 and activate the transcription factor NF-κB, which resulted in upregulation of HIF-1α transcription and its downstream gene, BMI1 , to mediate maintenance of the stemness-like phenotype. Suppression of the α v β 3 –NF-κB–HIF-1α pathway decreased OPN-mediated self-renewal capabilities. Levels of OPN protein expression were significantly correlated with HIF-1α protein levels in HCC tumor tissue samples. OPN might promote a cancer stem cell-like phenotype via the α v β 3 –NF-κB–HIF-1α pathway. Our findings offer strong support for OPN requirement in maintaining stem-like properties in HCC cells.