Gene expression profiling of alpha‐radiation‐induced rat osteosarcomas: Identification of dysregulated genes involved in radiation‐induced tumorigenesis of bone

Abstract

To better understand the molecular basis of radiation-induced osteosarcoma (OS), we performed global gene expression profiling of rat OS tumors induced by the bone-seeking alpha emitter 238 Pu, and the expression profiles were compared with those of normal osteoblasts (OB). The expressions of 72 genes were significantly differentially expressed in the tumors related to OB. These included genes involved in the cell adhesion (e.g., Podxl, Col18a1, Cd93, Emcn and Vcl), differentiation, developmental processes (e.g., Hhex, Gata2, P2ry6, P2rx5, Cited2, Osmr and Igsf10), tumorsuppressor function (e.g., Nme3, Blcap and Rrm1), Src tyrosine kinase signaling (e.g., Hck, Shf, Arhgap29, Cttn and Akap12), and Wnt/b-catenin signaling (e.g., Fzd6, Lzic, Dkk3 and Ctnna1) pathways. Expression changes of several genes were validated by quantitative real-time RT-PCR analysis. Notably, all of the identified genes involved in the Wnt/b-catenin signaling pathway were known or proposed to be negative regulators of this pathway and were downregulated in the tumors, suggesting the activation of bcatenin in radiation-induced OS. By using immunohistochemical and immunoblot analyses, constitutive activation of the Wnt/b-catenin signaling pathway in the tumors was confirmed by observing nuclear and/or cytoplasmic localization of b-catenin and a decrease in its inactive (phosphorylated) form. Furthermore, we found a significant reduction in the levels of glycogen synthase kinase 3b (GSK-3b) protein in the tumors relative to OB. Taken together, these findings provide new insights into the molecular basis of radiation-induced OS. ' UICCKey words: osteosarcoma; osteoblast; ionizing radiation; carcinogenesis; transcriptome Osteosarcoma (OS) is the most common primary tumor of bone in children and adolescents. The peak incidence of OS occurs in the second decade of life, with an additional smaller peak in the elderly population. This tumor is highly aggressive and is thought to arise primarily from osteoblasts (OB), boneforming cells. An increased risk for developing OS is known to be associated with some genetic disorders, such as hereditary retinoblastoma and Li-Fraumeni syndrome with germline mutations in the retinoblastoma (RB1) and TP53 genes, respectively. 1,2 In agreement with this, abnormalities of genes involved in the RB1 and TP53 tumor-suppressor pathways are often found in OS. 3,4 On the other hand, the association between ionizing radiation and the subsequent development of OS has been well documented in prior studies. For example, OS is known as one of the most frequent secondary malignant neoplasms occurring within the radiation field in patients, especially with retinoblastoma, treated with radiation therapy. 1,5 The risk of OS has been reported to be increased following the internal exposure to bone-seeking radioisotopes from occupational or medicinal use. 6-8 Interestingly, some studies have revealed the genetic and cytogenetic changes in radiation-induced OS and suggested the presence of additional tumor-associated genes. ...

The genetic basis for inactivation of Wnt pathway in human osteosarcoma

“…In contrast to these results, most previous studies have suggested that active Wnt signaling contributes to osteosarcoma development [6,7]. Furthermore, it is reported that the Wnt pathway is transcriptionally active in radiation-induced rat osteosarcomas [25], and that the Wnt/β-catenin pathway antagonists, curcumin and PKF118-310, demonstrate anti-tumor activity against human osteosarcoma cells [26]. These seemingly conflicting results suggest that the complexity of this important signaling pathway is still poorly understood in human osteosarcoma [5].…”

Modeling distinct osteosarcoma subtypes in vivo using Cre:lox and lineage-restricted transgenic shRNA

“…Of these, syntenic sections of 10, 11 and 16 are also lost in cases of human OS [43,44]. Candidate genes of interest on chromosome 7 include the Wnt pathway members DKK3 [45] and Wnt11 [46] which have roles in promoting osteoblast maturation. Chromosome 15 in mouse shows homology with human chromosomes 5, 8, 12 and 22.…”

Influence of β-catenin small interfering RNA on human osteosarcoma cells

“…More recently, it was reported that the Wnt pathway is transcriptionally active in radiation-induced rat OS [22] and Wnt/β-catenin pathway antagonists curcumin and PKF118-310 possess anti-tumour activity against human OS cells [23] . In addition, the Wnt inhibitory factor 1 (Wif1) is silenced in human OS and accelerates radiation-induced OS formation when it is deleted in mice [24] .…”