目标:在过去的几年中,白细胞介素-22(IL-22)和T辅助细胞(Th)22淋巴细胞在牙周炎发病机制中的重要性已变得明显;还有一些方面尚未解决。表达IL-22和芳烃受体(AhR)的细胞,转录因子主开关基因与Th22淋巴细胞的分化和功能有关,已在牙周炎患者的牙周组织中检测到。此外,IL-22在体外与破骨细胞分化及其骨吸收活性相关。然而,在牙周炎期间,表达IL-22的AhRTh22淋巴细胞在牙周组织上的破坏潜力尚未在体内得到证实。因此,本研究旨在分析在实验性牙周炎过程中,牙周病灶中检测到的表达IL-22的CD4+AhR+T淋巴细胞是否与牙槽骨吸收相关。
方法:使用小鼠牙周炎模型,IL-22和AhR的表达水平,以及Th1-,Th2-,Th17和T调节相关细胞因子,使用qPCR分析牙周病。使用流式细胞术分析牙周病变和引流这些牙周病变的颈部淋巴结中CD4+IL-22+AhR+T淋巴细胞的检测。此外,通过qPCR分析称为核因子κB配体受体活化因子(RANKL)的破骨细胞介质的表达,westernblot,和免疫组织化学。最后,使用显微计算机断层扫描和扫描电子显微镜分析牙槽骨吸收,骨吸收水平与IL-22和RANKL表达相关。
结果:更高水平的IL-22,AhR,还有RANKL,以及IL-1β,IL-6、IL-12、IL-17、IL-23和TNF-α,与假感染和未感染对照的牙周组织相比,在感染小鼠的牙周病中表达。同样,在感染小鼠的牙周组织中观察到高RANKL免疫反应;然而,在对照组中观察到很少或没有RANKL免疫反应。蛋白质印迹证实了RANKL与牙周感染之间的这种关联。此外,与未感染的对照组相比,在感染小鼠的牙周病损和引流这些牙周病损的颈部淋巴结中发现了更高的CD4+IL-22+AhR+T淋巴细胞检测.最后,IL-22和RANKL表达的增加与实验性牙周病中观察到的牙槽骨吸收增加呈正相关。
结论:本研究显示牙周炎患者组织中表达IL-22的CD4+AhR+T淋巴细胞增加,IL-22、RANKL表达呈正相关,和牙槽骨吸收。
OBJECTIVE: Over the past few years, the importance of interleukin-22 (IL-22) and T-helper (Th)22 lymphocytes in the pathogenesis of periodontitis has become apparent; however, there are still aspects that are not addressed yet. Cells expressing IL-22 and aryl hydrocarbon receptor (AhR), transcription factor master switch gene implicated in the differentiation and function of Th22 lymphocytes, have been detected in periodontal tissues of periodontitis-affected patients. In addition, IL-22 has been associated with osteoclast differentiation and their bone resorptive activity in vitro. However, the destructive potential of IL-22-expressing AhR+ Th22 lymphocytes over periodontal tissues during periodontitis has not been demonstrated in vivo yet. Therefore, this study aimed to analyze whether IL-22-expressing CD4+ AhR+ T lymphocytes detected in periodontal lesions are associated with alveolar bone resorption during experimental periodontitis.
METHODS: Using a murine model of periodontitis, the expression levels of IL-22 and AhR, as well as the Th1-, Th2-, Th17- and T regulatory-associated cytokines, were analyzed in periodontal lesions using qPCR. The detection of CD4+ IL-22+ AhR+ T lymphocytes was analyzed in periodontal lesions and cervical lymph nodes that drain these periodontal lesions using flow cytometry. In addition, the expression of the osteoclastogenic mediator called receptor activator of nuclear factor-κB ligand (RANKL) was analyzed by qPCR, western blot, and immunohistochemistry. Finally, alveolar bone resorption was analyzed using micro-computed tomography and scanning electron microscopy, and the bone resorption levels were correlated with IL-22 and RANKL expression.
RESULTS: Higher levels of IL-22, AhR, and RANKL, as well as IL-1β, IL-6, IL-12, IL-17, IL-23, and TNF-α, were expressed in periodontal lesions of infected mice compared with periodontal tissues of sham-infected and non-infected controls. Similarly, high RANKL immunoreaction was observed in periodontal tissues of infected mice; however, few or absent RANKL immunoreaction was observed in controls. This association between RANKL and periodontal infection was ratified by western blot. Furthermore, a higher detection of CD4+ IL-22+ AhR+ T lymphocytes was found in periodontal lesions and cervical lymph nodes that drain these periodontal lesions in infected mice compared with non-infected controls. Finally, the increased IL-22 and RANKL expression showed positive correlation between them and with the augmented alveolar bone resorption observed in experimental periodontal lesions.
CONCLUSIONS: This study demonstrates the increase of IL-22-expressing CD4+ AhR+ T lymphocytes in periodontitis-affected tissues and shows a positive correlation between IL-22, RANKL expression, and alveolar bone resorption.