## 4 Discussion
The production of IL-6 by osteoblasts is significant, as IL-6 has been reported to be important to osteoclast development and bone resorption, both directly and indirectly, via the upregulation of RANKL [12]. Stimulated osteoblasts also produced increased levels of IL-8, which was significant in the platform-matched groups, achieving results up to four-fold higher than the control after 3 days of exposure. The release of IL-8 from osteoblasts in response to implant materials was also confirmed by the work of [PERSON] [39]. The authors illustrated the distinctively high expression of IL-8 from osteoblasts after cultivation onto a wide range of commercially available implant materials, regardless of the material's surface roughness. On the other hand, [PERSON] et al., using actual dental implants, have demonstrated that titanium implants led to a more pronounced increase in IL-8 expression when compared with zirconia implants after incubation in human blood [40]. However, the magnitude of IL-8 expression in the work of [PERSON] et al. was far higher (>65-fold) than the levels expressed in the current study. This apparent difference could be attributed to either the different cells types used, the use of osteoblasts versus whole blood or the different incubation times [40]. Several investigators have also showed that osteoblasts incubated with Ti and CoCr particles/ions exhibit upregulated IL-8 chemokine expression [32; 41; 42]. These data indicate that the peri-implant-stimulated osteoblasts, although not the major source of chemotactic factors in the periprosthetic space, may be early and direct contributors to the initiation of the chronic peri-prosthetic inflammatory responses [11; 12; 41]. Interestingly, the release of COX-2 was upregulated, and the effect lasted for extended periods of exposure in most test groups, being more pronounced in the platform-matched couplings. It has been documented that COX-2 is dramatically upregulated during inflammation and usually increases in parallel with prostaglandin (PG) production and clinical inflammation [43]. [PERSON] et al. [42] also observed an increase in COX-2 secretion from osteoblasts exposed to Co and Cr ions, which corroborates the present results. This suggests that one of the pathogenic mechanisms of peri-implant tissue inflammation in vivo may be the synthesis of COX-2 by osteoblastic cells in response to ion release, in conjunction with PGE2 production [12; 42]. Increased levels of PGE2 have been demonstrated in the peri-prosthetic area and have been shown to play a significant role in the pathogenesis of peri-prosthetic osteolysis [11; 12]. To our knowledge, this is the first in vitro test that examined the effect of different IAC on the different osteoblast production levels of inflammatory cytokines and chemokines related to bone resorption; therefore, the results could not be precisely compared to previous investigations. However, it appears that the element release from alloys is necessary but not a sufficient condition for all cytotoxicity, as stated by [PERSON] et al. [44; 45]. In other words, elements must be released for cytotoxicity to occur, but not all element releases cause cytotoxicity. Therefore, the observed adverse reactions in osteoblasts were the result of ion release from the surfaces of immersed samples and the pronounced responses in the platform-matched couplings were due to the higher amounts of ion release from those groups. It is obvious then that the proposed theories explaining peri-implant bone loss, the biologic width theory [46], the bacterial theory [47] and the mechanical theory [48], cannot be proposed here to explain the adverse biological responses of the bone cells. Therefore, if we consider the corrosion product theory as explaining the present in vitro situation, it would then be logical to consider applying theory to the in vivo peri-implant bone changes around dental implants.