Tumor Necrosis Factor-α Produced by Osteoclasts Might Induce Intractable Pain in a Rat Spinal Metastasis Model of Breast Cancer

Abstract

Introduction Causes of pain due to spinal metastases have been insufficiently investigated. Tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were the focus of this study. Both are known as proinflammatory cytokines associated with the pathophysiology of pain syndromes 1 ) . It is well known that cancer cells produce these cytokines, but whether osteoclasts produce them as well remains unclear. We hypothesize that osteoclasts produce these cytokines; in other words, pain from spinal metastasis is stronger than pain from the primary tumor. Methods We made a rat spinal metastasis model of breast cancer (metastasis group) and models with a hole in the vertebrae (puncture group) and resected the vertebrae. Tartrate-resistant acid phosphatase (TRAP) staining was performed to reconfirm that osteoclasts increase in vertebrae with spinal metastasis. We then evaluated TNF-α and IL-6 expression using immunohistochemistry and real-time polymerase chain reaction (PCR). Results The results of TRAP staining showed that osteoclasts increase in metastatic vertebrae. The osteoclasts in the puncture models were TNF-α negative but were TNF-α positive in the metastasis model. The osteoclasts in the puncture models and metastasis model were both IL-6 positive. According to the real-time PCR results, TNF-α in vertebrae increased in the metastasis model, but IL-6 did not increase in the metastasis model compared with in the puncture model. Conclusions The number of osteoclasts is higher in the metastasis model. While TNF in the osteoclasts increased in the spinal metastasis model, IL-6 did not. This probably means that breast cancer affects TNF production in osteoclasts. This increase of TNF-α may lead to pain from spinal metastasis.

High Expression of Parathyroid Hormone-related Protein and Tumor Necrosis Factor-α in Cancer Cells as Risk Factors for Hypercalcemia in Bone Metastases Lytic Lesions

“…Mazaki et al (2019) performed a real-time PCR test to samples taken from mice vertebrae with breast cancer metastases, proving an increase in TNF-α gene expression compared to IL-6, and this was in line with the results of immunohistochemical expression. It is evident that TNF-α affects lytic lesions in bone metastases [35]. Takahashi et al (2003) studied 29 cancer patients with hypercalcemia and found that TNF-α levels in serum increased, though not significantly [36].…”

“…Correspondingly, serum TNF-α levels were proven to have a positive correlation with high invasion of cancer cells to bone and poor patient prognosis [34]. Mazaki et al (2019) performed a real-time PCR test to samples taken from mice vertebrae with breast cancer metastases, proving an increase in TNF-α gene expression compared to IL-6, and this was in line with the results of immunohistochemical expression. It is evident that TNF-α affects lytic lesions in bone metastases [35].…”

What Is the Role of Interleukins in Breast Cancer Bone Metastases? A Systematic Review of Preclinical and Clinical Evidence

“…However, in order to evaluate the main contributors involved in IL-6 expression during breast cancer bone metastases, numerous in vitro and in vivo studies analyzed tumor cells, bone cells, immune cells, and the organic bone matrix. Several preclinical studies showed that metastatic breast cancer cells, directly [54][55][56][57][58][59] or through Jagged1-expressing tumor cells [58], induced osteoblasts to express high levels of IL-6 [54][55][56][57][58][59][60][61], while they seemed to not affect IL-6 production by osteoclasts [62,63]. The high expression of IL-6 by osteoblasts, particularly activated throughout the bone marrow [60], completely suppresses osteoblast functions [55] and stimulates osteoclastogenesis in the presence or absence of the receptor activator of nuclear factor kappa-B (RANK)/RANK ligand (RANKL) pathway [56,59].…”