The Regulatory Role Of ANGPTL4 In Giant Cell Tumor Of Bone

Giant cell tumor(GCT) is a common primary bone tumor which typically occurs in the metaphyseal regions of long bones and less arises in sacrum, pelvis and spine. It is composed of osteoclast-like multinucleated giant cells, spindle-shaped stromal cells and mononuclear precursor cells. Spindle-shaped stromal cells of GCT(GCTSCs) originating from mesenchymal stem cells in bone marrow play a neoplastic role in GCT. GCT is classified as a benign tumor with a potentially aggressive behavior, malignant transformation and metastasis are not common. Surgical resection in the treatment of GCT is recommended, but the overall recurrence rate from 18% to 60% remains high. Osteolysis formation is an essential feature of GCT, however, the mechanism of which is still not fully understood. Thus further understanding the biology of GCT is critically important.Angiopoietin-like 4 Protein(ANGPTL4), a member of ANGPTLs’ family, is a secreted protein closely related to angiogenesis, lipid and glucose metabolism. ANGPTL4 is mainly expressed in the liver, adipose tissue, and placenta as well as in ischemic tissues, and is upregulated after fasting and hypoxia. It was identified as a paracrine or endocrine regulator of lipid metabolism in a previous search and a target of peroxisome proliferators-activated receptors(PPARs). In recent years emerging studies have implicated ANGPTL4’s role in the aggressiveness and metastasis of tumors due to its function in angiogenesis, inflammation and hypoxia. The role of ANGPTL4 in lipid metabolism has been well investigated, however, the relevance of ANGPTL4 in tumor biology remains largely undefined. ANGPTL4 expression is upregulated in clear cell renal-cell carcinoma and oral tongue squamous cell carcinoma(SCC). In addition, tumor-derived ANGPTL4 has been shown to promote breast tumors for lung metastasis. However, to the best of our knowledge, the expression and functional role of ANGPTL4 in GCT has not yet been reported.The transforming growth factor β(TGF-β) signaling pathway is a key player in nearly all cell types and with prominent roles during embryo development and tissue homeostasis. The regulatory cytokine TGF-β has a multifunctional function and regulates the production of microenvironment sensors and modulators, including cytokines, extracellular matrix components, and cell-surface receptors. TGF-β has an effect that tumor cells must avoid from for malignant evolution. However, paradoxically, TGF-β also modulates processes such as cell invasion, immune regulation, and microenvironment modification that tumor cells may exploit to their advantage. In an established research, TGF-β acted on its target gene ANGPTL4, which primes breast tumors for lung metastasis due to the disruption of lung capillary endothelial junctions. However, the TGF-β-ANGPTL4 signaling pathway has been not investigated in other kinds of tumors and the mechanism of this signaling pathway in tumor progress and invasion has not been fully elucidated.In this study, ANGPTL4 had a significantly increased expression in GCTSCs by western blot analysis, qRT-PCR analysis, immunolocalization and H&E staining, then we found that there was a significant enhancement of TGF-β in GCTSCs and a high level of TGF-β2 could directly increase ANGPTL4 expression in GCTSCs. Furthermore, we elaborated that ANGPTL4 could mediate osteoclast differentiation, osteolysis and angiogenesis on a cellular level through building a ANGPTL4-/-Knockout strain. We established a CAM model for GCT and observed its effects on GCT and angiogenesis in vivo.We verified that an up-regulation of ANGPTL4 was detected in GCTSCs and initially elaborated that ANGPTL4 significantly promoted GCTSCs’ proliferation, osteolytic lesion and angiogenesis in GCT by:(a) ANGPTL4 had a significantly increased expression in GCTSCs and a high level of TGF-β2 could directly increase ANGPTL4 expression in GCTSCs;(b) ANGPTL4 could mediate osteoclast differentiation, osteolysis and angiogenesis on a cellular level through building a ANGPTL4-/-Knockout strain;(c) ANGPTL4-Ab had an inhibitory effect on GCT and inhibit the tumor angiogenesis by establishing a CAM model. Initially, our finding indicated that ANGPTL4 could been positively regulated through TGF-β2 ligand and further control the biological behavior of GCT. In addition, we also assumed that this may offers a new target for the diagnosis and treatment of GCT.