Background
Giant cell tumor of bone (GCTB), a primary tumor of bone, frequently occurs in metaphysis of long bones, especially in the distal femur and proximal tibia [
1,
2]. It has a high recurrence rate after surgery with involvement of joints and their surrounding soft tissues and even causes serious dysfunction. Sometimes, it also incurs lung metastasis. At present, Campanacci imaging classification was an accepted classification standard of GCTBs.
GCTB is a low-grade malignant tumor with invasive, expansive, and destructive growth, and its biological behavior and prognosis are uncertain. At present, the relation between its particular gene expression and biological behavior is a hot research topic.
Ezrin is abnormally expressed in many tumor tissues and affects cellular motility, proliferation, apoptosis, and cycle and plays an important role in invasion and metastasis of tumors. In recent years, it has attracted more attention of researchers with more researches [
3]. CD44 is a kind of cell adhesion factors, and changes in its expression are related to the progression, invasion, and metastasis of tumors and tumor-free survival rate of patients. VEGF is an important predisposing factor for tumor angiogenesis, promotes tumor cell proliferation, hydrolyzes basement membrane, and enhances vascular permeability through its specific binding with the receptor. Therefore, ezrin, CD44, and VEGF were used as the research targets in this study.
In this study, expression status of ezrin, CD44, and VEGF were measured with immunohistochemical technique in GCTB tissue samples, association of ezrin, CD44, and VEGF with clinical pathological characteristics and prognosis of GCTB patients, and their role in invasion, metastasis, and recurrence of GCTB was analyzed in order to provide the theoretical basis for the treatment of GCTB.
Discussion
Invasion, metastasis, and recurrence are major biological and clinical characteristics of GCTB and the most important factors affecting the prognosis of GCTB patients. Invasion, metastasis, and recurrence of tumor are a complex, changeable, and multistep process, which can be affected by many factors. In this study, the role of ezrin, CD44, and VEGF in biological behaviors of GCTB and their application was studied.
Ezrin, a link protein of cell membrane and cytoskeleton, can anchor actin in specific membrane, maintain cell polarity, take part in cell shape regulation and cell movement, interact with a variety of cell surface molecules (CD44, Met, etc.) [
6], regulate cell-cell adhesion with cell-extracellular matrix, and promote metastasis and proliferation of tumor cells. Ezrin can regulate cell signal transduction by Rho factor, tyrosine kinase receptor, and Ras-to-MAPK pathway and plays an important role in cell movement, proliferation, apoptosis, cell cycle, invasion, and metastasis of tumor cells [
7-
9]. In addition, ezrin is related with micro-RNA and c-Myc [
10,
11] where ezrin affects invasion and metastasis of tumors.
It was reported that high ezrin expression levels play a role in promoting invasion and metastasis of esophageal carcinoma, breast cancer, ovarian cancer, liver cancer, osteosarcoma, and other common tumors [
7,
8,
12-
15]. High ezrin expression status can induce conversion of a variety of cell lines and abnormal proliferation [
16]. Bruce et al. [
5] analyzed the ezrin expression in more than 5,000 tumor tissue specimens, including breast cancer, lung cancer, prostate cancer, colon cancer, sarcoma, and their adjacent normal tissue samples with microarray immunohistochemical method, showing that high ezrin expression status is closely related with the pathological grade of sarcoma and recurrence of breast cancer.
Our study showed that the high ezrin expression rate was significantly higher in GCTB tissue than in its adjacent normal tissue (58.8% vs 11.2%, P < 0.05). The high ezrin expression rate gradually increased in Campanacci stages I to III of GCTB and was significantly different between Campanacci stage III and stages I to II of GCTB. The ezrin expression status was significantly higher in GCTB patients with recurrence and metastasis than in those without recurrence and metastasis, indicating that ezrin expression status is higher in GCTB patients and may influence the biological behaviors of GCTB, such as invasion, recurrence, and metastasis.
Survival analysis of the follow-up data showed that ezrin induced the GCTB biological behaviors, suggesting that recurrence of GCTB is not related with the age and gender of GCTB patients and tumor-free survival rate is significantly lower for ezrin (+ +) GCTB patients than for ezrin (−, +) GCTB patients. The median recurrence and metastasis time of GCTB patients with a high ezrin expression status was 32 months. The median recurrence and metastasis of GCTB patients with a low ezrin expression was longer than that (90 months) in our study (P < 0.05). Kaplan-Meier analysis and log-rank comparison showed that high ezrin expression status in GCTB patients was closely related with invasion, metastasis, and recurrence of GCTB after operation.
CD44, a receptor of hyaluronic acid and a marker of tumor stem cells, is a kind of cell adhesion factors. Changes in its expression are related to the progression, invasion, and metastasis of GCTB and the tumor-free survival rate of GCTB patients. CD44 and its ligand hyaluronan are widely distributed in human body, and its binding to hyaluronic acid caused by abnormal expression of CD44 may influence the occurrence and development of tumors [
17]. CD44 is involved in cell adhesion and migration, lymphocyte activation, extracellular matrix accumulation, and metastasis of tumor cells [
18]. Elliott et al. [
11] showed that ezrin may play an important role in tumorigenesis by regulating cell adhesion molecules. Ezrin can directly interact with the cytoplasm of CD44 molecules, influence conformation of cytoskeletal protein and its distribution, change apoptosis of tumor cells, regulate cell-cell adhesion with extracellular matrix, and promote metastasis and proliferation of tumors. In this study, CD44 was abnormally expressed in GCTB patients. The higher the Campanacci stage was, the higher the positive CD44 expression rate was. Moreover, survival analysis showed that CD44 could affect the recurrence of GCTB, and its expression was positively related to that of ezrin, indicating that the expression of CD44 is related to the occurrence, development, and infiltration of GCTB.
Kumta et al. [
19] detected the expression of VEGF in 14 GCTB patients by semiquantitative RT-PCR and immunohistochemistry, respectively, showing that multiple subtypes of VEGF, especially 121 subtypes, are highly expressed in GCTB patients and the expression of VEGF is related to the osteolytic destruction and local recurrence of GCTB. In this study, the high expression rate of VEGF was significantly higher in GCTB tissues than in its adjacent normal tissues (61.3%
vs 26.2%). In addition, the VEGF expression status was higher in GCTB patients at Campanacci stage III with recurrence and metastasis than in those at Campanacci stages I and II with no recurrence and metastasis. The results of our study agreed with those of Kumta [
19]. However, the mechanism of VEGF is not fully understood. Knowles et al. [
20] showed that hypoxia can induce the expression of hypoxia-inducible factor (HIF) and regulate the expression of VEGF by autocrine and paracrine which indirectly affect the formation of osteoclasts. Matsumoto et al. [
21] reported that the VEGF-Flt-1-FAK pathways consisting of VEGF and its receptor are involved in aggregation and proliferation of osteoclast precursor cells and the bone destruction. Furthermore, the expression of ezrin and VEGF in GCTB patients was positively correlated with the correlation coefficient (
r = 0.741,
P < 0.001) in this study. Youn [
22] showed that ezrin can enhance VEGF-induced nitric oxide production by activating calpain, thereby regulating the function of endothelial cells and vascular generation, suggesting that VEGF may play a role in the recurrence and metastasis of GCTB, and interact with ezrin.
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Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
ZZY, JZ, and JD were responsible for the study concept and design. XM and JLZ conducted the data acquisition while ML and YC conducted the immunohistochemistry. JZ, XM, YYD, KL, and ZPZ analyzed and interpreted the data. JZ prepared the manuscript while ZZY reviewed the manuscript. All authors have read and approved the final manuscript.