Archives of Oral Biology 120 (2020) 104957 Available online 22 October 2020 0003-9969/© 2020 Elsevier Ltd. All rights reserved. Cytokine profles of healthy and diseased sites in individuals with periodontitis Tamires Szeremeske Miranda a , Nathalia de Freitas Figueiredo a , Luciene Cristina Figueiredo a , H´ elio Doyle Pereira da Silva a , Fernanda Regina Godoy Rocha c , Poliana Mendes Duarte a, b, * a Department of Periodontology, Dental Research Division, Guarulhos University, Praça Tereza Cristina, 229, Guarulhos, S˜ ao Paulo, Brazil b Department of Periodontology, College of Dentistry, University of Florida, 1600 SW Archer Rd., Room D10-6, Gainesville, FL, USA c Department of Oral Biology, College of Dentistry, University of Florida, 1600 SW Archer Rd., Room D10-6, Gainesville, FL, USA A R T I C L E INFO Keywords: Periodontitis Cytokines immunology infammation ABSTRACT Objective: The aims of this study were: 1) to compare the levels of cytokines between healthy and diseased sites, in patients with untreated periodontitis; 2) to correlate cytokine levels with each other and with key periodontal pathogens, in healthy and diseased sites. Methods: Paired gingival crevicular fuid (GCF) samples were obtained from two healthy (probing depth (PD) and clinical attachment level (CAL) ≤3 mm without bleeding) and two diseased sites (PD and CAL ≥5 mm with bleeding on probing [BoP]) of patients with generalized stage III/IV grade B/C periodontitis. GCF levels of eighteen cytokines and subgingival levels of seven periodontal pathogens were assessed by multiplex immuno- assay and qPCR, respectively. Results: A total of 112 subjects and 448 GCF samples were analyzed. The GCF levels of GM-CSF, IL-17, IL-1β, IL-2, IL-21, IL-23 and TGF-β were signifcantly higher in the diseased than in the healthy sites (p < 0.05). Levels of IL-8 and MIP-1α were signifcantly higher in the healthy than in the diseased sites (p < 0.05). In the healthy sites, IL-8 and MIP-1α formed an independent cluster of cytokines and, MIP-1α positively correlated with Porphyromonas gingivalis (p < 0.05). In deep sites, smoking negatively associated with GM-CSF, IL-10, IL-17, IL-23, IL-5, IL-6, IL- 7, IL-8 and MIP-1α levels (p < 0.05). Conclusions: Diseased sites exhibited increased levels of T helper 17-related cytokines and TGF-β while healthy sites presented increased levels of the chemokines, IL-8 and MIP-1α. Patients with periodontitis may not only have infammation in diseased deep sites, but also present signifcant hidden subclinical infammation in their shallow clinically healthy sites. 1. Introduction Periodontitis is an infectious-infammatory disease that results in periodontal tissue collapse, as a consequence of the interaction between dysbiotic subgingival microbial communities and host immune- infammatory responses (Cekici, Kantarci, Hasturk, & Van Dyke, 2014; Van Dyke, Bartold, & Reynolds, 2020). Patients with periodontitis generally present shallow, moderate and deep categories of probing depth (PD), including disease-affected sites and not-yet-affected healthy sites (Teles et al., 2018). Longitudinal studies investigating the devel- opment of periodontal diseases have demonstrated that prevalence of progression is higher overall at deep sites, compared to shallow sites. However, due to the larger number of shallow sites in most periodontitis subjects, the number of progressing shallow sites is much higher than that of deeper sites (Lindhe, Okamoto, Yoneyama, Haffajee, & Soc- ransky, 1989; Teles et al., 2018; Thomson, Broadbent, Poulton, & Beck, 2006). Dysregulated production of cytokines is related to the development of various infectious and infammatory diseases. A complex cascade of cytokines, which can act antagonistically or synergistically, is released as a result of cellular crosstalk during response to pathogens (Dinarello, 2007). Over recent years, research has advanced in understanding the role of cytokines in the pathogenesis of periodontal diseases, demon- strating that cytokines play crucial functions in the onset and * Corresponding author at: University of Florida, Department of Periodontology, 1600 SW Archer Rd., Room D10-6, Gainesville, FL 32610, USA. E-mail addresses: szeremeske@yahoo.com.br (T.S. Miranda), nathaliaf.fgueiredo@gmail.com (N.F. Figueiredo), lucienedefgueiredo@gmail.com (L.C. Figueiredo), helio.silva@prof.ung.br (H.D.P. Silva), FGodayRocha@dental.uf.edu (F.R.G. Rocha), PMendesDuarte@dental.uf.edu (P.M. Duarte). Contents lists available at ScienceDirect Archives of Oral Biology journal homepage: www.elsevier.com/locate/archoralbio https://doi.org/10.1016/j.archoralbio.2020.104957 Received 25 August 2020; Received in revised form 16 October 2020; Accepted 18 October 2020