Deletion of osteopontin in non-small cell lung cancer cells affects bone metabolism by regulating miR-34c/Notch1 axis: a clue to bone metastasis

Abstract

Lung cancer is prone to bone metastasis, and osteopontin (OPN) has an important significance in maintaining bone homeostasis. The goal of this study was to explore the impact of OPN level on bone metabolism and the molecular mechanism of inhibiting bone metastasis in non-small cell lung cancer (NSCLC). The expression of OPN in NSCLC was ascertained by Western blot and immunohistochemistry, and the correlation between the expression level of OPN and survival of patients was analyzed. Then the shRNA technology was applied to reduce the expression of OPN in NSCLC cells, and CCK-8 assay was carried out to investigate the effect of low expression of OPN on the proliferation of NSCLC cells. In addition, the effects of low expression of OPN on osteoclast differentiation, osteoblast generation and mineralization were studied using osteoclast precursor RAW264.7 and human osteoblast SaOS-2 cells, and whether OPN could regulate miR-34c/ Notch pathway to affect bone metabolism was further explored. The findings showed that the high level of OPN in NSCLC was closely related to the poor prognosis of patients and the abnormal proliferation of NSCLC cell lines. The suppression of OPN was beneficial to inhibit the differentiation of osteoclasts and promote the mineralization of osteoblasts. Besides, this study confirmed that the deletion of OPN can regulate bone metabolism through the regulation of miR-34c/Notch1 pathway, which will contribute to inhibiting the occurrence of osteolytic bone metastasis in NSCLC.

Immunoregulatory Roles of Osteopontin in Diseases

“…The binding of OPN with integrins initiates the activation of several downstream signaling effectors, such as phosphatidylinositol 3 kinase (PI3K) /protein kinase B (AKT), focal adhesion kinase (FAK)/AKT and nuclear factor kappa-B (NF-κB), leading to cell proliferation, migration, epithelial–mesenchymal transition (EMT), inflammation, neurotoxic microglial phenotype, tumor growth, migration and invasion, as well as angiogenesis within the chronic subdural hematoma (CSDH) outer membrane [ 15 , 16 , 17 , 18 , 19 , 20 ]. In addition, OPN regulates other signal pathways or signal molecules, such as Janus kinase (JAK)/signal transducer and activator of transcription (STAT) [ 21 , 22 , 23 , 24 ], PI3K/AKT [ 15 , 25 , 26 , 27 ], NOTCH [ 28 , 29 ], extracellular regulated protein kinase1/2 (ERK1/2) [ 30 ], the ubiquitin C -terminal hydrolase L1 (UCHL1)–ubiquitin–proteasome system (UPS) axis [ 31 ] and transforming growth factor β (TGF-β) [ 32 ], influencing cellular physiological processes and disease progression. OPN also acts as a ligand for CD44 that results in attracting mesenchymal stem cells (MSCs) to the tumor microenvironment, promoting EMT and tumor budding (TB), inducing macrophage migration and activation, stimulating intestinal growth, differentiation and maturation, cell growth, proliferation, migration and cell-cycle activity and promoting oxidative stress [ 15 , 17 , 23 , 33 , 34 , 35 , 36 ].…”