Possible Association between Behçet’s Disease and Periodontal diseases

Abstract

Aim

This study explores the connection between Behçet’s disease (BD), characterized by persistent oral and genital ulcers alongside iritis, and periodontal disease. It examines the levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and nitric oxide (NO) in gingival crevicular fluid (GCF) and saliva.

Methods

Forty Behçet’s patients with gingivitis or periodontitis and 47 patients with either gingivitis or periodontitis but without BD were studied. Periodontal status was recorded with standard clinical indexes. GCF and saliva samples were obtained. NO, IL-1β and TNF-α levels were analysed. Current Behçet’s symptoms and medications usage were recorded.

Results

Mean salivary IL-1β was elevated (p = .045), and mean NO level was decreased in BD patients with gingivitis compared to patients without BD (p = .000). In contrast, mean NO level in crevicular fluid was higher in Behçet’s patients with periodontitis than in patients without BD (p = .009). Furthermore, among Behçet’s patients, those with vascular involvement had lower salivary NO level compared to patients without vascular involvement (p = .000).

Conclusions

Based on our findings, the elevated levels of IL-1β in the saliva of Behçet’s patients with gingivitis, along with the decreased NO level, indicate an altered inflammatory response in the oral cavity.

Supplementary Information

The online version contains supplementary material available at 10.1186/s12903-024-04749-x.

Introduction

Behçet’s Disease (BD) is a rare disorder, and most patients present with a characteristic triad of recurrent aphtous ulcers in the oral and genital regions, along with ocular lesions. Cardiovascular involvement, which may be arterial or venous, can also be observed. Occasionally, gastrointestinal tract and central nervous system can be affected [1–3]. BD was first described in 1937 by a Turkish dermatologist Dr. Hulusi Behçet. The exact cause of BD is not fully understood, but genetic, geographic, and environmental factors are believed to play significant roles in its etiology [3].

Indeed, numerous experimental and clinical studies have indicated that chronic inflammation localized in periodontitis can have implications for systemic health. Additionally, it has been observed that oral infections can potentially lead to systemic inflammation [4]. BD and periodontal diseases share certain genetic and environmental risk factors and immunologic responses play a role in the development of both conditions, it is hypothesized that there might be an association between BD and periodontal diseases [5]. This association could arise from shared underlying mechanisms involving the immune system and inflammatory responses. Moreover, systemic inflammation may be proposed to explain the potential relationship between BD and periodontitis. Supporting this hypothesis, certain systemic therapies used to treat autoimmune diseases like BD, such as tumor necrosis factor alpha (TNF-α) inhibitors and corticosteroids, have been observed to result in clinical improvement in periodontal diseases [6]. This indicates that controlling the systemic inflammation associated with BD may have a positive effect on periodontal health [7–10].

Absolutely, an insufficient innate immune response in both BD and periodontal disease can create an environment conducive to the accumulation of proinflammatory cytokines, such as interleukin-1β (IL-1β) and TNF-α, as well as soluble antigens in the oral cavity [11, 12]. Nitric oxide (NO) in periodontium and saliva, has antimicrobial activity and plays an important role in the immune response of the oral cavity against pathogenic bacteria [13]. Previous studies have, so far, evaluated the relationship between BD and periodontitis [14, 15]; however, to date, there is no available data on the impact of BD on gingivitis or periodontitis with respect to gingival crevicular fluid (GCF) and salivary immunologic mediators based on current knowledge.

The objective of this research was to compare various parameters related to periodontitis and gingivitis between patients with and without BD. Specifically, we focused on evaluating the levels of IL-1β, TNF-α, and NO in GCF and saliva in the presence of periodontal diseases. Our aim was to determine if these biomarkers differ significantly in individuals with BD.

Our hypothesis is that BD patients may display distinct immunological responses to bacterial inflammation in their periodontal tissues, depending on their systemic conditions and symptoms. By investigating these immunological differences, we seek to gain a better understanding of the potential associations between BD and periodontal diseases.

Materials and methods

The study described was a clinical study and consisted of 40 patients with BD (24 female and 16 male) who received treatment at Gazi University Faculty of Medicine, Department of Rheumatology, Ankara. Forty-seven systemically healthy gingivitis or periodontitis patients without BD (26 female, 21 male) from Gazi University Faculty of Dentistry, Department of Periodontology were included as controls (Fig. 1). Periodontitis was diagnosed as generalized with a severity of Stage II, pocket depth 3–4 mm according to the criteria by the criteria 2017 World Workshop on the Classification of Periodontal and Peri-Implant Diseases and Conditions [16]. Gingivitis was diagnosed as generalized according to a BOP score ≥ 10%, without attachment loss and radiographic bone loss [17]. The participants were divided into 4 groups; systemically healthy patients with gingivitis (HG), systemically healthy patients with periodontitis (HP), BD patients with gingivitis (BG), and BD patients with periodontitis (BP). This study was approved by the human subject’s ethics board of Gazi University Clinical Research Ethics Committee (2590160043) and was conducted in accordance with the Helsinki Declaration of 1975, as revised in 2013. All patients signed an informed consent form. The study was registered in Clinical Trials with the TCTR identification number is TCTR20240722003 and the web site is https://www.thaiclinicaltrials.org/show/TCTR20240722003. Exclusion criteria were smoking, pregnancy, lactation, other systemically diseases, and previous periodontal and/or antibiotic treatment within the last 6 months.

Fig. 1.

Flow chart of participants

All examinations were conducted by a single experienced dental examiner (FO). GCF samples and clinical measurements were obtained. The clinical examination included plaque index (PI) [18], gingival index (GI) [19], clinical attachment level (CAL), probing pocket depth (PPD), and bleeding on probing (BOP) [20]. The measurements were performed at 6 sites per tooth (mesio-buccal, mid-buccal, disto-buccal, mesio-lingual, mid-lingual and disto-lingual) using a periodontal probe (Williams periodontal probe, Nordent Manufacturing, Elk Grove Village, IL). GCF was obtained before recording clinical measurements. The deepest 4 pockets of non-adjacent single rooted teeth were selected for GCF sampling (Fig. 2).

Fig. 2.

The sampling area was isolated, and a gingival crevicular fluid sample was taken

Diagnosis of BD

Patients were diagnosed according to the International Criteria for Behçet’s Disease (ICBD). ICBD is based on ocular lesions, genital ulcers, oral aphthous ulcers, skin manifestations, vascular involvement and positive pathergy test. Systemic involvement associated with BD, current symptoms and the use of any systemic medications were recorded.

Saliva sampling and processing

Saliva sampling was performed before the periodontal examination and GCF collection. For each patient, unstimulated whole mixed saliva was collected using a modified method. Participants were asked to rinse her/his mouth with water for a minute, 10 min before the sampling. The saliva samples were collected into a sterile plastic container for 5 min. Samples were centrifuged for 10 min at 15,000 x g at 4 °C and the supernatant was frozen at -20 °C until analyses.

GCF sampling and processing

The GCF sampling site was gently air dried and supragingival plaque was removed. The area was carefully isolated with cotton rolls in order to prevent from contamination. Standardized paper strips (Periopaper, OraFlow, Amityville, NY, USA) were inserted into the sulcus until slight resistance was felt and left in place for 30 s. Strips contaminated by bleeding were discarded. GCF volumes were determined by using a calibrated Periotron 8000^®. The readings from the Periotron 8000^® were converted into microliter (µL) by referencing to the standard curve with MILCONVERT.exe program ( MILCONVERT.exe software version 2.52, Oraflow, Amityville, NY, USA). Strips were placed into coded micro centrifuge tubes and stored at -80 °C until processing. Before biochemical analyses, paper strips were placed in 300 µL of phosphate buffered saline solution containing 0.5% bovine serum albumin in Eppendorf tubes, and GCF was eluted from the strips by centrifugation for 6 min at 5000 x g at 4 °C.

Biochemical analyses

NO level of samples were determined using colorimetric nitric oxide assay kit (Oxford Biomedical Research, Oxford, MI, USA) (Product number: NB98). In aqueous solution, NO rapidly degrades to nitrate and nitrite. This kit employs the NADH-dependent enzyme reductase (NaR) for conversion of nitrate to nitrite prior to quantification of nitrate using Griess reagent-thus providing for accurate determination of total NO production. This colorimetric nitric oxide assay kit can be used to accurately measure as little as 1 pmol/µL (∼1µM) NO produced in aqueous solutions. The completed reaction is read at 540 nm. NO concentration was expressed as µM.

TNF-ɑ Picokine ELISA (human, Catalog Number: EK0525) and IL-1β Picokine ELISA (human, Catalog Number: EK0392) kits are sandwich ELISA kits for quantitative detection of human TNF-ɑ and human IL-1β. Detection range for TNF-α Picokine ELISA kit was 7.8–500 pg/mL and for IL-1β 3.9–250 pg/mL. Sensitivities for TNF-ɑ and IL-1β kits were < 1 pg/mL and < 0,15 pg/mL, respectively. Analyses and microplate readings were performed using an ELISA autoanalyzer (ChemWell 2900, Awareness Technology, Palm City, FL, USA).

TNF-ɑ (human) and IL-1β (human) Picokine ELISA kits (Boster Biological Technology Co., Pleasanton, Ltd., CA, USA) were based on similar standard sandwich ELISA technology. A monoclonal antibody (MoAb) from mouse specific TNF-ɑ or IL-1β has been precoated onto 96 well plates. Following the addition of standards and samples to the wells, a biotinylated detection polyclonal antibody from goat specific TNF-ɑ or IL-1β was added and then washed. Avidin-Biotin-Peroxidase Complex was added and unbound conjugates were washed away. Horseradish peroxidase (HRP) substrate 3,3’,5,5’-tetramethylbenzidine (TMB) was used to visualize HRP enzymatic reaction. The blue color product changed into yellow after adding stop solution. The density of the yellow color which absorbs at 450 nm is proportional to the human TNF-ɑ or IL-1β amount of sample captured in plate. Results were expressed as pg/mL.

Statistical analysis

Power analysis was based on a previous studies that reported GCF chemokines in individuals with periodontal diseases. A sample size of 15 participants per group was deemed sufficient to achieve 80% power with a 5% alpha level [21, 22].

Data were analyzed using the SPSS statistical package (version 23.0, IBM SPSS, Inc., Chicago, IL, USA). The results were expressed as mean ± standard deviation and median (minimum-maximum). PI, GI, CAL, BOP, PD, GCF volume and involvement types of Behçet’s disease were analyzed using Independent Sample t test. The assumption of homogeneity of variance was tested using Levene’s Test of Equality of Variances. The differences in the use of medications among the groups were compared by using ANOVA with Bonferroni correction. The correlation between clinical parameters was evaluated with Pearson Correlation test. p < .05 was considered statistically significant.

Results

Demographic data of the study groups are outlined in Table 1. Eighteen patients used colchicine, four patients used TNF-ɑ inhibitor, infliximab, the others used azathioprine, cyclophosphamide, interferon, and prednisolone only or a combination of those for BD therapy (Fig. 3).

Table 1.

The demographic data of the participants and comparison between Behcet gingivitis (BG), systemically healthy gingivitis (HG), Behcet periodontitis (BP) and systemically healthy periodontitis patients’ (HP) full mouth clinical parameters

	HG (n = 24)		BG (n = 26)		HP (n = 23)		BP (n = 14)			HG vs. BG	HP vs. BP	BG vs. BP			HG vs. HP
	Mean ± SD	Median (Min-Max)	Mean ± SD	Median (Min-Max)	Mean ± SD	Median (Min-Max)	Mean ± SD	Median (Min-Max)	P value		P value		P value	P value
Gender (F/M)	9/15		15/11		17/6		9/5
PI	1.27 ± 0.06	1.27 (0.32–1.89)	1.35 ± 0.09	1.44 (0.18–2.11)	1.38 ± 0.11	1.36 (0.16–2.18)	1.56 ± 0.15	1.47 (0.77–2.73)	p = .508		p = .363		p = .238	p = .372
GI	1.28 ± 0.07	1.27 (0.33–1.79)	1.25 ± 0.08	1.35 (0.26–1.88)	1.47 ± 0.06	1.56 (0.70–1.96)	1.43 ± 0.08	1.5 (0.94–1.87)	p = .796		p = .648		p = .178	p = .052
PPD	2.30 ± 0.06	2.20 (1.20–2.89)	2.05 ± 0.06	2.04 (1.14–2.63)	3.03 ± 0.10	3.02 (2.22–4.05)	2.92 ± 0.17	2.67 (2.30–4.36)	p = .009**		p = .570		p = .000***	p = .000***
CAL	2.49 ± 0.08	2.38 (2.05–3.74)	2.14 ± 0.06	2.13 (1.22–2.72)	3.28 ± 0.11	3.21 (2.36–4.4)	3.25 ± 0.20	3.01 (2.30–4.48)	p = .002**		p = .907		p = .000***	p = .000***
BOP	61.91 ± 20.23	65 (3–90)	56.81 ± 22.8	56 (3-100)	81.26 ± 17.12	87 (48–100)	75.64 ± 17.67	77 (45–100)	p = .408		p = .346		p = .011*	p = .001**

Independent T test. Statistically significant difference ***p < .001, **p < .01 and *p < .05

PI: Plaque index. GI: Gingival index. CAL: Clinical attachment level (mm). BOP: Bleeding on probing (%). PPD: Probing pocket depth (mm)

Fig. 3.

The numbers and percentages of therapies of Behçet’s Disease

Full mouth clinical periodontal index comparisons are shown at Table 1. PPD, CAL values, and BOP were found significantly higher in BP group when compared with BG group (p = .000, p = .000, p = .011, respectively) (Table 1). The differences in PI (p = .363) and GI (p = .648) between both periodontitis groups (BP and HP) and gingivitis groups (BG and HG) were not statistically significant (PI; p = .508, GI; p = .052). PPD and CAL were significantly higher in HG group than in BG group (p = .009, p = .002 respectively). None of the clinical parameters was found to be significant between BP and HP groups.

The periodontal parameters at sampled sites, saliva and GCF of IL-1β, TNF-α and NO levels for HG, BG, HP and BP groups were shown at Table 2. PPD, BOP and GCF volume (p = .000, p = .009, p = .001, respectively). Similarly, PPD and BOP were measured to be higher in HP group compared to HG group (p = .000, p = .02, respectively). GCF volume, GCF IL-1β and salivary IL-1β levels were significantly higher in HP group than in HG group (p = .000, p = .025, p = .005, respectively). GCF NO values were significantly elevated in HG group than in HP group (p = .007). In BG subjects, salivary IL-1β level was increased (p = .045), while salivary NO levels were determined to be lower in HG subjects (p = .000). NO levels in GCF were detected to be higher in BP group than in HP group (p = .009), while salivary TNF-α levels were significantly higher in BP subjects than in HP subjects (p = .022).

Table 2.

Comparison of sampling sites’ clinical parameters and biochemical parameters in GCF and in saliva between systemically healthy gingivitis (HG), Behcet gingivitis (BG), systemically healthy periodontitis (HP) and Behcet periodontitis (BP)

	Gingivitis				Periodontitis				HG vs. BG	HP vs. BP	HG vs. HP	BG vs. BP
	HG(n = 24)		BG(n = 26)		HP(n = 23)		BP(n = 14)
	Mean ± SD	Median (Min-Max)	Mean ± SD	Median (Min-Max)	Mean ± SD	Median (Min-Max)	Mean ± SD	Median (Min-Max)	P value	P value	P value	P value
PI	1.30 ± 0.16	1.00 (0.00–3.00)	1.21 ± 0.19	1,50 (0.00–3.00)	1.53 ± 0.1	1.50 (1.00–2.00)	1.78 ± 0.2	2.00 (0.00–3.00)	p = .720	p = .117	p = .778	p = .073
GI	1.36 ± 0.08	1.50 (0.50-2.00)	1.38 ± 0.11	1,50 (0.00–2.00)	1.56 ± 0.1	1.50 (0.50-3.00)	1.53 ± 0.1	1.5 (1.00–2.00)	p = .909	p = .877	p = .208	p = .400
PPD	2.62 ± 0.1	2.50 (2.00–4.00)	2.09 ± 0.12	2.00 (1.00–3.50)	3.89 ± 0.2	4 (2.5–5.5)	3.82 ± 0.3	3,75 (2.5-6.00)	p = .002**	p = .831	p = .000***	p = .000***
BOP	63.16 ± 27.28	66 (13–100)	63.26 ± 31	58 (0-100)	81.04 ± 23	83 (16–100)	87.6 ± 15	91.5 (50–100)	p = .990	p = .351	p = .020*	p = .009**
GCF Volume	0.13 ± 0.02	0.09 (0.03–0.33)	0.16 ± 0.02	0.14 (0.03–0.4)	0.39 ± 0.1	0.32 (0.09–1.12)	0.44 ± 0.1	0.34 (0.09–1.23)	p = .395	p = .613	p = .000***	p = .001**
GCF IL-1β	37.88 ± 7.62	31.07 (2.66–139.1)	35.24 ± 7.6	18.96 (2.42–147.7)	70.56 ± 12	31.07 (6.43-198.24)	67.3 ± 18	45.05 (4.79-268.96)	p = .808	p = .878	p = .025*	p = .065
GCF TNF-α	27.81 ± 2	27.33 (14.95–50.8)	41.55 ± 9.3	24.28 (15.29–252)	24.0 ± 1.9	21.99 (14.95–47.41)	35.6 ± 9.9	25.35 (17.39–161)	p = .172	p = .191	p = .292	p = .687
GCF NO	19.33 ± 2.93	15,94 (0.05–68.52)	20.89 ± 2	19.08 (4.23–48.82)	9.45 ± 1.0	6.63 (0.01–38.81)	17.2 ± 1.9	15.72 (8.68–32.02)	p = .661	p = .009**	p = .007**	p = .250
Saliva IL-1β	142.01 ± 31.86	97.64 (1.53–596.8)	229.5 ± 28	194.6 (51.78–567)	294.2 ± 42	286.98 (32.9-624.58)	272 ± 53	178.83 (38.76–617)	p = .045*	p = .748	p = .005**	p = .438
Saliva TNF-α	23.53 ± 0.57	22.63 (18.77–31.8)	36.36 ± 5.1	25.25 (21.2-136.1)	26 ± 1.69	23.59 (17.16–53.61)	36.6 ± 5	28.58 (21.17–81.15)	p = .155	p = .022*	p = .145	p = .486
Saliva NO	213.15 ± 25.47	207.16 (16.63–430)	77.78 ± 15.9	58.89 (1.2-277.67)	154 ± 27.7	116.71 (2.47- 423.41)	82.8 ± 23	39.44 (1.75-223.55)	p = .000***	p = .084	p = .124	p = .856

Independent T test. Statistically significant difference ***p < .001, **p < .01 and *p < .05

PI: Plaque index. GI: Gingival index. CAL: Clinical attachment level (mm). BOP: Bleeding on probing (%). PPD: Probing pocket depth (mm). GCF volume (µg)

The Pearson correlation analysis results for the full mouth and sampled sites in HG and BG groups which were shown in Table 3. A positive correlation was shown in HG group only between full mouth CAL value and salivary IL-1 β levels (p < .05; r = .446). A strong positive correlation was found between full mouth PI and salivary IL-1β value in BG group (p < .01). Moreover, in this group, a positive correlation was found among PI, GI and GCF IL-1β in the sampling site, whilst a negative correlation was observed between PPD and GCF NO (p < .05).

Table 3.

The correlations between clinical and biochemical parameters of systemically healthy gingivitis (HG) and Behcet gingivitis (BG) groups

		HG(n = 24)			BG(n = 26)
Full mouth		Saliva IL-1β	Saliva TNF-α	Saliva NO	Saliva IL-1β	Saliva TNF-α	Saliva NO
PI	PCC Sig.	-0.236 0.267	0.014 0.948	-0.235 0.268	0.542 0.004**	0.106 0.608	-0.016 0.937
GI	PCC Sig.	0.000 0.999	0.036 0.869	-0.021 0.924	0.240 0.238	0.129 0.530	-0.004 0.985
PPD	PCC Sig.	0.237 0.264	-0.083 0.700	-0.125 0.559	0.119 0.563	0.101 0.624	-0.034 0.869
CAL	PCC Sig.	0.446 0.029*	0.070 0.747	0.243 0.253	0.095 0.643	0.207 0.311	-0.085 0.679
BOP	PCC Sig.	-0.085 0.693	0.141 0.511	0.074 0.731	0.263 0.194	0.225 0.269	-0.071 0.732
Sampled sites		GCF IL-1β	GCF TNF-α	GCF NO	GCF IL-1β	GCF TNF-α	GCF NO
GCF Volume	PCC Sig.	-0.105 0.626	0.125 0.561	-0.297 0.158	0.244 0.230	0.077 0.708	0.176 0.390
PI	PCC Sig.	0.169 0.430	0.257 0.225	0.01 0.998	0.392 0.048*	0.091 0.660	0.315 0.117
GI	PCC Sig.	0.160 0.454	0.013 0.953	0.090 0.676	0.493 0.011*	0.255 0.208	0.155 0.450
PPD	PCC Sig.	0.258 0.223	0.247 0.245	-0.002 0.994	-0.118 0.567	-0.128 0.533	-0.460 0.018*
BOP	PCC Sig.	0.075 0.727	0.226 0.288	0.065 0.765	0.335 0.094	0.101 0.625	0.145 0.479

PCC: Pearson correlation coeeficient. Statistically significant difference **p < .01 and *p < .05. PI: Plaque index. GI: Gingival index. CAL: Clinical attachment level. BOP: Bleeding on probing. PPD: Probing pocket depth

Table 4 presented statistically significant positive correlations among full mouth PI, GI and saliva IL-1 β values (p < .05). A positive correlation was seen between PPD and GCF IL-1β (p < .05) in the sampled site, adversely, there was a strong negative correlation in GCF NO (p < .01) in HP group. When CAL was getting higher in BP group, salivary IL-1β values increased (p < .05). Similarly, a positive correlation was found between PPD and GCF IL-1β (p < .05).

Table 4.

The correlations between clinical and biochemical parameters of systemically healthy periodontitis (HP), Behcet periodontitis (BP)

		HP(n = 23)			BP(n = 14)
Full mouth		Saliva IL-1β	Saliva TNF-α	Saliva NO	Saliva IL-1β	Saliva TNF-α	Saliva NO
PI	PCC Sig.	0.446 0.026*	0.363 0.075	-0.132 0.529	0.463 0.096	0.124 0.673	-0.388 0.170
GI	PCC Sig.	0.421 0.036*	0.140 0.504	-0.184 0.380	0.292 0.311	0.088 0.764	-0.191 0.512
PPD	PCC Sig.	0.210 0.313	-0.167 0.424	− 0.049 0.814	0.467 0.092	0.366 0.198	-0.022 0.941
CAL	PCC Sig.	0.360 0.077	0.065 0.758	0.071 0.737	0.532* 0.050	0.486 0.078	0.051 0.863
BOP	PCC Sig.	0.277 0.180	0.016 0.938	-0.256 0.216	0.461 0.097	0.347 0.224	-0.125 0.670
Sampled site		GCF IL-1β	GCF TNF-α	GCF NO	GCF IL-1β	GCF TNF-α	GCF NO
GCF Volume	PCC Sig.	0.239 0.250	-0.029 0.892	-0.244 0.239	0.060 0.838	-0.146 0.618	-0.260 0.369
PI	PCC Sig.	0.228 0.273	-0.149 0.479	-0.203 0.329	0.437 0.118	-0.171 0.559	0.122 0.678
GI	PCC Sig.	0.247 0.233	-0.046 0.827	-0.034 0.870	0.420 0.135	-0.303 0.292	-0.004 0.990
PPD	PCC Sig.	0.385 0.050*	-0.051 0.808	-0.526 0.007**	0.611 0.020*	-0.045 0.880	-0.084 0.776
BOP	PCC Sig.	0.244 0.239	-0.042 0.842	-0.315 0.125	0.383 0.177	-0.394 0.164	0.223 0.443

PCC: Pearson correlation coeeficient Statistically significant difference **p < .01 and *p < .05. PI: Plaque index. GI: Gingival index. CAL: Clinical attachment level. BOP: Bleeding on probing. PPD: Probing pocket depth

In HG group, a positive correlation was found between salivary TNF-α and GCF TNF-α levels (p < .05; r = .502), and between salivary TNF-α and salivary IL-1 β levels (p < .01; r = .595). In BP group, a very strong positive correlation was found between salivary IL-1β and salivary TNF-α levels (p < .001) (see supplementary Table 1 S).

It was recorded that among 40 patients with Behcet, 38 had oral ulcers, 15 had genital ulcers, 14 had ocular lesions, 10 had articular involvement, 4 had nervous system involvement, and 5 had vascular involvement. When analyzing variables related to the presence of genital, vascular, and other lesions, as well as medication use, Behcet patients with gingivitis and periodontitis were grouped together.

When the patients with (n = 15) and without (n = 25) genital aphthae were compared, sampling site of GI, full mouth GI, and PI values were significantly higher in patients with genital aphthae (Independent T-test, p < .05). When the patients with (n = 5) and without (n = 35) vascular involvement were compared, the salivary NO level was significantly lower in patients with vascular involvement (p < .001). There was no difference in clinical and immunological status between patients with and without eye, nerve and joint involvement.

ANOVA analysis showed a difference in PPD and GCF volume only in patients using colchicine (n = 18), not in those who do not use any systemic medications (n = 7) (p < .05).

Discussion

The chronic inflammatory nature of both BD and periodontal disease, along with their close association with proinflammatory cytokines, suggests the possibility of a bidirectional interaction between these conditions [23, 24]. Despite this, there is limited existing research on the periodontal health of individuals with BD [14, 15]. Moreover, no previous studies have investigated the clinical parameters, systemic therapy effects, and IL-1β, TNF-α and NO levels in GCF and saliva samples in BD patients.

In this study, we assessed both saliva and GCF as crucial oral secretions that can provide valuable insights into the impact of systemic and local diseases. Among the cytokines evaluated in periodontal diseases, IL-1β was of particular interest due to its well-established association with bone resorption [25]. Interestingly, the levels of salivary IL-1β were found to be significantly higher in the BG group compared to HG group. The existence of IL-1β gene polymorphism in BD could potentially modify the inflammatory response, leading to increase in periodontal inflammation in the initial stages of lesions [14]. In genetic investigations, IL-1β gene polymorphism has been identified as a significant factor influencing the development of BD, thereby playing a crucial role in its etiology [26]. According to Akman et al., there was a possibility of an association between BD and periodontal disease with IL-1 gene polymorphism [14]. Additionally, it has been suggested that the etiopathogenesis of BD may be related to TNF-α gene polymorphism [27]. Consistent with these findings, our present study revealed significantly higher salivary TNF-α levels in the Behçet’s periodontitis compared to the healthy periodontitis. The results of our study are in agreement with Turkcu et al. who also reported that patients with BD exhibited significantly elevated serum TNF-α levels compared to the systemic healthy control group [27]. The increase in TNF-α, released from Th1 cells and known to play a role in delayed-type hypersensitivity in BD, could potentially contribute to the higher salivary TNF-α levels observed in individuals with BD [28]. NO is among the most frequently encountered molecules exhibiting antimicrobial activity in various infectious diseases, including periodontitis [29]. In the present study, significantly lower salivary NO levels were found in BG compared to HG. According to a study conducted by Turkmen et al. comparing serum NO levels between individuals with BD and healthy controls, it was found that BD patients had lower serum NO levels while experiencing increased oxidative stress [30]. It was further proposed that measuring serum NO levels could be a potentially useful approach for diagnosing and monitoring patients with BD. Similarly, Orem et al. also found that the level of NO was reduced in active Behçet’s patients [31]. This decrease in NO led to endothelial dysfunction, which in turn contributed to the development of pathophysiological conditions in BD. Saliva is a fluid that reflects the plasma levels of many metabolites, therefore low salivary NO may be an indicator of serum NO levels. It might be hypothesized that low salivary NO levels in Behçet’s patients may lead to impaired defensive functions of saliva and increased susceptibility to periodontal diseases. NO level in BD could also be a potential biomarker of vascular inflammation [32]. It has been reported that there was eNOS gene polymorphism in BD patients [33]. NO synthesized by eNOS has crucial functions such as maintaining vascular integrity, vasodilation, and adhesion of leukocytes to endothelial cells. Since the serum NO level has already been reported to be lower in BD patients, our findings of the low salivary NO level in gingivitis could be attributed to the reflection of the low serum NO level.

GCF is a fluid that contains cytokines secreted through the periodontal tissues and proposed to be a site-specific reflection of periodontal inflammation compared to saliva [34]. Oh et al. reported that GCF volume and IL-1β levels were higher in deeper pockets which could be a marker showing the severity of periodontal disease [35]. In the current study, similar results were observed concerning the levels of IL-1β in GCF. The levels of IL-1β were found to be significantly higher in healthy patients with periodontitis (HP) compared to those with healthy with gingivitis (HG). Considering the increased GCF IL-1β at sampled sites, irrespective of BD, suggests a primary role in GCF IL-1β levels in periodontal disease. Significant differences between Behçet’s patients and systemic healthy patients were observed in salivary IL-1β and TNF-α levels. These discrepancies may be attributed to the fact that saliva tends to reflect the overall systemic inflammation rather than solely the presence of periodontal disease [36]. Conversely, GCF is more closely associated with periodontal inflammation rather than reflecting the systemic health status of the individual [37].

Among the various clinical manifestations in BD, aphthous ulcers are the most common findings and these ulcers can present challenges in maintaining proper oral hygiene for affected individuals [38]. Coit et al. have reported that PI values were significantly higher in BD patients compared to systemically healthy control groups [39]. In our study, we found that full mouth PI values were non-significantly higher in BD with periodontal diseases group (BG and BP) compared to systemically healthy HG and HP groups. In Coit et al.’s study, the BD patients were not categorized according to their periodontal status and the sample size was too small to make a comparison with our study [39]. Similar to our results, Seoudi et al. did not find any difference between PI values both in BD patients, patients with reccurent oral apthous ulcers and systemically healthy subjects [40]. In other studies, it was reported that PD measurements were higher in BD patients than in systemically healthy ones [15, 38, 41]. In our study, there was no difference in full mouth PPD and CAL values between HP and BP patients, however HG had significantly higher PPD and CAL than BG patients (p < .05). The prominent influence of local factors in periodontitis may mask any noticeable clinical differences caused by the systemic effects of BD. As a result, the impact of BD on the clinical presentation of periodontitis might not be easily apparent.

In instances where gingival inflammation does not resolve, the root cause is often systemic. A thorough examination of the oral cavity for additional mucosal changes may uncover signs indicative of an underlying systemic condition [42]. Gingival inflammation, in terms of GI and BOP values, did not differ in BG and HG patients, as well as BP and HP patients. However, an increase in PPD due to edema in the gingiva might indicate a reflection of the systemic status, which did not appear to have the same impact on gingival bleeding. The immune-suppressing medications commonly used in the treatment of BD, such as colchicine, azathioprine, and prednisolone, have the potential to influence immuno-inflammatory mechanisms [43]. Despite this, our study findings indicate that these medications did not appear to have a significant impact on the clinical indicators of gingival inflammation. In this study, we also examined the variations in clinical and immunological findings related to periodontal diseases based on the types of systemic involvement observed in BD patients. When the correlation between vascular involvement and clinical and immunological findings in BD patients were examined, salivary NO level was found significantly lower in patients with vascular involvement (p < .001). It has been reported that low serum NO levels play an important role in the etiology of vasculitis in Behçet’s patients [44]. Salivary NO level can be considered as closely related with the serum NO level and the measurement of salivary NO level may become important in diagnosis and prognosis of vascular involvement in BD. Salivary TNF-α was the only parameter which correlated with age in BD. In our study, BD patients were categorized into 3 groups: those who were using colchicine, those taking other medications, and those not on any medication. Out of the participants, 18 were using colchicine, 15 were on other medications, and 7 were not taking any medication. Interestingly, we found no significant differences in full mouth PPD and CAL values, as well as immunological parameters, among these 3 groups. This suggests that the use of colchicine or other medications did not lead to distinct variations in the periodontal clinical and immunological findings compared to those not using any medication.

Regarding the limitations, a significant challenge in such studies is the difficulty in controlling confounding factors. However, these results are based on a limited number of participants. Additional research with larger sample sizes and longer follow-up periods is needed to confirm these findings.

Conclusion

In conclusion, the findings of our study indicate that immunopathogenetic mechanisms in BD may influence the nature of periodontal diseases in affected individuals. However, to better understand the potential impact and associations, further prospective studies are required. These studies can explore whether treating the periodontal condition in BD patients could potentially lead to improvements in the systemic manifestations of BD.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Author contributions

F.O., N.O., S.E. and S.G. made substantial contributions to conception and design of the study. S.E. performed the biochemical analysis. F.O., S.G., H.K., B.B. and A.T. involved in data collection. F.O., N.O., S.G. and B.G. carried out data analysis and drafted the manuscript. All authors reviewed the manuscript and approved the final version of the manuscript.

Funding

This study was funded by the Gazi University Research Grant at Gazi University Scientific Research Council (03/2016–01).

Data availability

The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.

Declarations

Ethical approval

This study was approved by the human subjects ethics board of Gazi University Clinical Research Ethics Committee (22122014/579). All procedures applied in studies involving human participants were under the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.

Consent for publication

Not Applicable.

Consent to participate

Informed consent was obtained from all individual participants included in the study.

Competing interests

The authors declare no competing interests.