Quantification of pro-inflammatory cytokines and osteoclastogenesis markers in successful and failed orthodontic mini-implants

Abstract Objectives: Miniscrew has been frequently used, considering that anchorage control is a critical point in orthodontic treatment, and its failure, the main adverse problem. Using two groups of stable (successful) and unstable (failed) mini-implants, this in vivo study aimed to quantify proinflammatory cytokines IL-1 α, IL-6, IL-17, and TNF-α and osteoclastogenesis marker RANK, RANKL, and OPG in gingival tissue, using the real-time polymerase chain reaction technique. Methodology: Thirteen patients of both sexes (11-49 years old) under orthodontic treatment were selected, obtaining 11 successful and 7 failed mini-implants. The mini-implants were placed and removed by the same surgeon, in both jaws. The mean time of permanence in the mouth was 29.4 months for successful and 7.6 months for failed mini-implants. At removal time, peri-mini-implant gingival tissue samples were collected and processed for quantification of the proinflammatory cytokines and osteoclastogenesis markers. Nonparametric Wilcoxon rank-sum test considering the clusters and Kruskal-Wallis test were used for statistical analysis (α=0.05). Results: No significant difference (p>0.05) was observed between the groups for either quantification of cytokines or osteoclastogenesis markers, except for IL-6 (p<0.05). Conclusions: It may be concluded that the expression of IL-1α, IL-17, TNF-α, RANK, RANKL, and OPG in peri-implant gingival tissue were not determinant for mini-implant stability loss, but the higher IL-6 expression could be associated with mini-implant failure.


Introduction
Mini-implants have been widely used in orthodontics 1,2 with a high clinical success rate. 3 Even so, loss of devices may still occur in some cases. The reasons for failures are not entirely clear. According to a recent revision, 4 failure was associated with initial loading, inadequate hygiene, brushing, or thrusting in the area, and persistent inflammation. Moreover, inflammation is frequently mentioned as one of the factors involved in implant loss. 3,5,6 The installation of dental implants promotes the activation of molecular mechanisms involved in bone remodeling for osteointegration and can also trigger a cascade of inflammatory reactions by the stimulus of cytokine and chemokine production, contributing to the establishment of a unique biochemical environment. [7][8][9] Moreover, inflammatory and immune response increases in peri-implant periodontal tissues with increased microbial colonization of implants after installation in the oral cavity, resulting in greater production of proinflammatory cytokines. 7,10 The expression of these proinflammatory cytokines and osteoclastogenesis-related factors plays an important role in the development and severity of peri-implantitis, a major cause of dental implant loss. [11][12][13] A balance between pro-inflammatory and antiinflammatory cytokines regulates immune response.
These mediators are also key regulating factors of osteoclast and osteoblast differentiation and activation, which modulate osteoclastogenesis and maintain bone homeostasis, especially in response to aggressive agents. 7 19,20 Due to the scarcity of studies, it is not known whether cytokines can cause development of periimplantitis in orthodontic mini-implants as well. The evaluation of these mediators involved in inflammation is critical to increase the stability of temporary anchorage devices in orthodontics. 8 Only few studies evaluated the expression of some cytokines -IL-1 β, IL-2, IL-6, and IL-8 -in peri-implant crevicular fluid in response to orthodontic tooth movement, 21,22

Discussion
Pro-inflammatory and anti-inflammatory cytokines regulate osteoclast and osteoblast differentiation and activation, which modulate osteoclastogenesis and maintain bone homeostasis, especially in response to aggressive agents. 6,7 Cytokine overexpression involved in inflammation is critical to increase the stability of temporary anchorage devices in orthodontics. 8 Inflammatory events are commonly identified as key elements of healing processes, inflammation mechanism remains unclear. 25 Then, this study evaluated the gene expression of cytokines and osteoclastogenese markers to verify their role in failed mini-implants.
In this study, no significant differences were found between successful and failed mini-implants regarding IL-1α expression in peri-implant gingival tissue samples of both groups, but the group with failed mini-implants showed higher numerical values of this cytokine than In orthodontics, RANK/RANKL/OPG system has been shown to play an important role in the bone remodeling process. In this study, no differences were observed between the groups regarding RANK, RANKL, and OPG levels. For RANKL, our results agree with those of Güncü, et al. 10  Although some authors suggest that inflammation of the mucosa adjacent to mini-implants, periimplantitis and even bone loss mediated by the host response can be decisive for mini-implant failure, 6,8,30 no other studies published quantified pro-inflammatory cytokines and osteoclastogenesis mediators in gingival

Conclusion
The expression of pro-inflammatory cytokines IL-1α, IL-17, and TNF-α and osteoclastogenesis markers RANK, RANKL, and OPG in peri-implant gingival tissue were not determinant for mini-implant stability loss. The higher expression of the IL-6 pro-inflammatory cytokine could be associated with mini-implant stability loss.