Surface vacuolar ATPase in ameloblastoma contributes to tumor invasion of the jaw bone

Abstract

Ameloblastoma is the most common benign odontogenic tumor in Japan. It is believed that it expands in the jaw bone through peritumoral activation of osteoclasts by receptor activator of nuclear factor kappa-B ligand (RANKL) released from the ameloblastoma, as in bone metastases of cancer cells. However, the clinical features of ameloblastoma, including its growth rate and patterns of invasion, are quite different from those of bone metastasis of cancer cells, suggesting that different underlying mechanisms are involved. Therefore, in the present study, we examined the possible mechanisms underlying the invasive expansion of ameloblastoma in the jaw bone. Expression levels of RANKL assessed by western blotting were markedly lower in ameloblastoma (AM-1) cells than in highly metastatic oral squamous cell carcinoma (HSC-3) cells. Experiments coculturing mouse macrophages (RAW264.7) with AM-1 demonstrated low osteoclastogenic activity, as assessed by tartrate-resistant acid phosphatase (TRAP)-positive multinuclear cell formation, probably because of low release of RANKL, whereas cocultures of RAW264.7 with HSC-3 cells exhibited very high osteoclastogenic activity. Thus, RANKL release from AM-1 appeared to be too low to generate osteoclasts. However, AM-1 cultured directly on calcium phosphate-coated plates formed resorption pits, and this was inhibited by application of bafilomycin A1. Furthermore, vacuolar-type H+-ATPase (V-ATPase) and H+/Cl- exchange transporter 7 (CLC-7) were detected on the surface of AM-1 cells by plasma membrane biotinylation and immunofluorescence analysis. Immunohistochemical analysis of clinical samples of ameloblastoma also showed plasma membrane-localized V-ATPase and CLC-7 in the epithelium of plexiform, follicular and basal cell types. The demineralization activity of AM-1 was only 1.7% of osteoclasts demineralization activity, and the growth rate was 20% of human normal skin keratinocytes and HSC-3 cells. These results suggest that the slow expansion of several typical types of ameloblastomas in jaw bone is attributable to its slow growth and low demineralization ability.

Upregulation of interleukin-8 and activin A induces osteoclastogenesis in ameloblastoma

“…Previous studies demonstrated that RANKL-positive cells were more commonly distributed throughout the stroma than in tumor cells in AM (40,47,48). The results of the present study indicated that RANKL mRNA was positively expressed in AM tissues rather than in AM cells, confirming that stromal cells are the major source of RANKL in AM.…”

“…However, the results of the present study demonstrated only a slight increase in osteoclast formation of osteoclast precursors in response to AM cells and no evidence of RANKL expression in AM cells. Yoshimoto et al (40) also failed to observe detectable expression of RANKL in AM cells. Interestingly, clear expression of IL-8 was observed in AM cells and neutralization of IL-8 prevented osteoclastogenesis induced by the CM of AM cells (data not shown), suggesting that tumor-derived IL-8 is a key factor that directly induces osteoclast differentiation in AM and that the soluble RANKL in the 1X CM of AM cells is too low to induce osteoclastogenesis.…”

Bone Resorption in Ameloblastoma and Its Underlying Mechanism

“…Besides RANKL, the progression also involved matrix metalloproteinases (MMPs). MMP-1, MMP-2, and MMP-9 were released by ameloblastoma (Yoshimoto, et al, 2016).…”

Effect of technetium‐99 conjugated with methylene diphosphonate (99Tc‐MDP) on OPG/RANKL/RANK system in vitro

“…It is associated with the surrounding bone resorption, which is a series of complex molecular biochemical reactions that includes osteoprotegerin (OPG), receptor activator of NF‐κB (RANK), and receptor activator of nuclear factor kappa‐B ligand (RANKL) mediated by osteoblasts and osteoclasts . RANKL and RANK also play an important role in jaw resorption during the development of the ameloblastomas . Previous studies in the literature showed that biphosphate drugs have a regulatory effect on the OPG/RANKL/RANK system and can significantly inhibit osteoclasts .…”

“…[1][2][3][4][5] RANKL and RANK also play an important role in jaw resorption during the development of the ameloblastomas. [1][2][3][4][5][6][7] Previous studies in the literature showed that biphosphate drugs have a regulatory effect on the OPG/RANKL/RANK system and can significantly inhibit osteoclasts. 8 99 Tc-MDP (Technetium 99 conjugated with methylene diphosphonate), as bisphosphonate drugs, possesses anti-osteoclastogenic activity against receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast formation.…”