Proinflammatory and Anti-InflammatoryCytokines in Gingival Crevicular Fluidand Serum of Patients With RheumatoidArthritis and Patients With ChronicPeriodontitisBurcu Cetinkaya,* Esra Guzeldemir,† Ersin Ogus,‡ and Sule Bulut*

Background: The aim of this study is to evaluate proinflammatory andanti-inflammatory cytokine levels in gingival crevicular fluid (GCF) andserum of rheumatoid arthritis (RA) and chronic periodontitis (CP) pa-tients to assess whether cytokine profiles distinguish patients with RAand patients with CP while using healthy patients as background con-trols.

Methods: A total of 49 patients, 17 patients with RA (three males and14 females; mean age: 47.82 – 10.74 years), 16 patients with CP (10males and six females; mean age: 44.00 – 7.00 years), and 16 controls(eight males and eight females; mean age: 28.06 – 6.18 years) were en-rolled. Patients with RA were under the supervision of rheumatologists;15 of the patients with RA were being treated with methotrexate–sulfasa-lazine combined therapy, and two of the patients were being treated withleflunomid therapy. Periodontal parameters (plaque index, gingival in-dex, probing depth, and clinical attachment level) were recorded. Inter-leukin (IL)-1b, IL-4, IL-10, and tumor necrosis factor-a (TNF-a) weredetermined in GCF and IL-1b and IL-10 in serum by enzyme-linked im-munosorbent assay.

Results: There were significant differences found among RA, CP, andcontrol groups for all periodontal parameters (P <0.05). The total amountand concentration of GCF IL-1 b, IL-4, IL-10, and TNF-a were similar inRA and CP patients (P >0.05). Although the total amount and concentra-tion of serum IL-10 was not significantly different among the groups(P >0.05), serum IL-1b was significantly lower in the RA group comparedto CP patients and controls and was higher in GCF of the RA groupcompared to the CP group.

Conclusions: Although clinical periodontal disease parameters indi-cated more severe periodontal disease in CP compared to RA patients,immunologic evaluation did not reveal consistent results regardingproinflammatory and anti-inflammatory cytokine levels. This might bea result of the use of non-steroidal anti-inflammatory drugs and rheuma-toid agents by patients with RA. J Periodontol 2013;84:84-93.

KEY WORDS

Arthritis, rheumatoid; chronic periodontitis; interleukin-1beta;interleukin-4; interleukin-10; tumor necrosis factor-alpha.

Chronic periodontitis (CP)is a bacterially inducedchronic inflammatory

disease resulting in connec-tive tissue and bone de-struction.1 The pathogenesisof periodontal disease in-volves innate and adaptiveimmune responses.2

Recent studies indicate thatperiodontal disease might becorrelated to various systemicdiseases, including adversepregnancy outcome,3 diabe-tes,4 coronary heart disease,5

myocardial infarction, stroke,and respiratory disease, andhas been proposed as havingan etiologic ormodulating rolein these diseases.6 Most ofthese associations are attrib-uted to excessive productionof proinflammatory cytokinesand other inflammatory medi-ators, suchas interleukin (IL)-1,IL-6, IL-11, IL-17, prostaglan-din E2 (PGE2), and tumor ne-crosis factor-a (TNF-a) and animbalance between proinflam-matory and anti-inflammatorycytokines.7-9 It was shownthat inflammatory response

* Department of Periodontology, Faculty of Dentistry, Baskent University, Ankara, Turkey.† Department of Periodontology, Faculty of Dentistry, Kocaeli University, Kocaeli, Turkey.‡ Department of Biostatistics, Faculty of Medicine, Baskent University. doi: 10.1902/jop.2012.110467

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of proinflammatory cytokines spreads to regionsclose to bone.10 Inflammatory mediators that leadto bone resorption depend on the expression ofproinflammatory cytokines, and to the contrary, anti-inflammatory cytokines, such as IL-4, IL-10, IL-12,IL-13, and IL-18, serve to inhibit bone resorption.7

Rheumatoid arthritis (RA) is a chronic, immune-mediated inflammatory disease characterized by sy-novial inflammation and early cartilage and focalbone loss implemented by bone resorbing osteo-clasts that often results in structural damage, dis-ability, and loss of function.11 RA affects �1% ofthe world population in a female/male ratio of 3:1and has a peak incidence of onset in females intheir fourth and fifth decades of life.12 It was con-cluded that the majority of clinical and epidemi-ologic studies indicate that patients with RA havean increased prevalence of periodontitis and toothloss.11,13

There are remarkable similarities regarding thepathogenesis of periodontal disease and arthritis withevidence of coexistence.9 In both periodontal diseaseand RA, a persistent inflammatory reaction occurs inareas composed of connective tissue and bone to-gether with the activation of complement, productionof cytokines, and release of other inflammatory cellproducts.14 Although the etiology of periodontal dis-ease has been well established,15 the etiology ofRA remains uncertain. However, the association be-tween RA and periodontitis implies that certainfeatures of the inflammatory response might becommon to both diseases;11 moreover, pathogenesisof these two diseases indicates a common mech-anism for dysregulation of the inflammatory re-sponse.12 Periodontitis contains remarkably similarcytokine profiles to those of RA.12,16

IL-4 and IL-10 are potent anti-inflammatoryagents.17 The lack of IL-4 may cause the increasedproduction of inflammatory mediators, such as TNF-aand IL-1b, in periodontal tissue, and it has beensuggested that IL-10 has regulatory effects in peri-odontal inflammation. Also, the lack of IL-4 andIL-10 may contribute to the initiation and pro-gression of inflammation during chronic synovitisin RA joints.18,19 Proinflammatory cytokines, suchas IL-1 and TNF-a, are abundant in RA, but IL-4 ispresent in low concentrations within the rheumatoidsynovial membrane. Furthermore, a ‘‘balance’’ be-tween the effects of proinflammatory and anti-inflammatory cytokines is thought to determine theoutcome of disease, in either the short or long term.

The data seem to indicate that a relationship ex-ists between the extent and severity of periodontaldisease and RA.9,12 In literature, a possible rela-tionship was shown between RA and periodontaldiseases.11,12,14,20-25 However, the findings regarding

the relationship between RA and periodontal dis-eases are not always concordant.8,23,26

The aim of this study is to evaluate whether thepatients with CP and the patients with RA share sim-ilar proinflammatory and anti-inflammatory cytokineprofiles in their serum and gingival crevicular fluid(GCF) and to distinguish these patients from in-dividuals free of disease.

MATERIALS AND METHODS

This study was conducted at Baskent University,Faculty of Dentistry, Periodontology Clinics, Ankara,Turkey, from 2007 to 2009. The study protocol wasapproved by the Ethics Committee of the MedicalFaculty of Baskent University according to the Hel-sinki declarations. After completion of personal,medical, and dental questionnaires, written informedconsent was obtained.

Study PopulationForty-nine patients were enrolled in the cross-sectional study. Three study groups were conductedas follows; 17 patients with RA (three males and 14females; mean age: 47.82 – 10.74 years; range: 29to 70 years), 16 otherwise healthy patients with CP(10 males and six females; mean age: 44.00 – 7.00years; range: 34 to 54 years), and 16 healthy controls(eight males and eight females; mean age: 28.06 –6.18 years; range: 20 to 43 years).

All patients with RA were diagnosed according tothe American Rheumatology Association 1987 re-vised criteria for the classification of RA. The cri-teria are as follows: 1) morning stiffness in andaround joints lasting ‡1 hour before maximal im-provement; 2) soft-tissue swelling (arthritis) of ‡3joint areas observed by a physician; 3) swelling(arthritis) of the proximal interphalangeal, meta-carpophalangeal, or wrist joints; 4) symmetric swell-ing (arthritis); 5) rheumatoid nodules; 6) the presenceof rheumatoid factor; and 7) radiographic erosionsand/or periarticular osteopenia in hand and/or wristjoints.27 Criteria 1 through 4 must have been pre-sent for ‡6 weeks. RA is defined by the presenceof ‡4 criteria.

Patients with RA were excluded if they had anyknown systemic diseases other than RA and a historyof systemic or local disease with an influence onthe immune system (cancer, cardiovascular or re-spiratory diseases, etc.).

All patients with RA were under the supervisionof rheumatologists, and 15 of the patients with RAwere being treated with methotrexate–sulfasalazinecombined therapy ([mean dosage: 22.27 – 0.52mg/week; range: 15 to 25 mg/week] - [mean dos-age: 2.54 – 0.52 g/day; range: 2 to 3 g/d]). Becausetwo of the patients did not respond to this treatment

J Periodontol • January 2013 Cetinkaya, Guzeldemir, Ogus, Bulut

85

protocol, leflunomid therapy (20 mg/day) was con-ducted during the last 6 months. All of the patientswere using non-steroidal anti-inflammatory drugs(NSAIDs) (flurbiprofen, meloxicam, diclofenac, pa-racetamol, or sodium) regularly. During the clinicalevaluation period, it was seen that 15 of the patientsused corticosteroids (1 mg/kg/day). Erythrocytesedimentation rate of the patients with RA was16.00 – 9.08 mm/hour (range, 2 to 35 mm/hour),mean C-reactive protein level was 8.97 – 9.52mg/L (range, 0.20 to 29.00 mg/L), and RA dura-tion was 6.08 – 6.14 years (range, 2 to 24 years).

The patients in the CP group were recruited frompatients seeking dental treatment. The periodontaldiagnosis of patients was established on the basisof clinical and radiographic criteria defined by the1999 International World Workshop for a Classifi-cation of Periodontal Diseases and Conditions.28,29

Briefly, patients with CP had ‡20 teeth exhibiting>5 mm clinical attachment loss (AL) on ‡3 anteriorteeth (incisors and/or canines). The bone lossestimation was radiographically performed ineach patient for the assessment of the extent andseverity of alveolar bone loss. Patients with CPwere excluded if they had any known systemicdiseases that could influence the periodontalstatus, oral diseases other than CP, and ongoingorthodontic therapy.

Individuals in the control group were systemicallyand periodontally healthy and had neither a historynor sign of periodontal disease, i.e., probing depths(PDs) <3 mm with no AL, no obvious clinical in-flammation, and no bleeding on probing.

Individuals who were evaluated for the presentstudy were excluded if they had conservative orprosthetic restorations and caries at the anterior re-gion, history of systemic or local disease with aninfluence on the immune system (cancer, cardio-vascular and respiratory diseases), history of hep-atitis or human immunodeficiency virus infection,immunosuppressive chemotherapy or current preg-nancy or lactation, requirement for antibiotic pro-phylaxis, a history of antibiotic therapy within thepreceding 3 months, or periodontal treatment within6 months. To be included in the study, individualshad to be >18 years of age. Smokers were excludedfrom the study.

Periodontal ExaminationThe full-mouth clinical periodontal measurementswere recorded at six sites per tooth (mesio-buccal,mid-buccal, disto-buccal, disto-lingual, mid-lingual,and mesio-lingual), including plaque index (PI),30

gingival index (GI),31 PD, and clinical attachmentlevel (CAL). A periodontal probe§ was used forperiodontal measurements.

GCF Sampling and AnalysisAt least 24 hours later, after periodontal measure-ments,GCFsampleswereobtained fromthemaxillaryanterior region using standardized paper stripsi asdescribed previously.4,32 Briefly, samples were col-lected after isolation of the tooth with cotton rolls andremoval of any supragingival deposits on the toothsurfaces. The samples were collected from themesio-buccal and disto-buccal of each of six teeth. Thecollection region was air dried gently for 2 seconds toreduce any contaminationwith plaque and saliva. Thestrip was left at the entrance of the gingival sulcus for30seconds.After thecollectionofGCF, thestripsweremoved to a micro-moisture meter device,¶ which wasprecalibrated with a dry, sterile strip. Samples withevidence of bleeding were not included. After volumedetermination, GCF samples were placed in sterileEppendorf tubes that were wrapped securely andstored at -70�C until the day of laboratory analysis.

Blood Sampling and AnalysisVenous blood samples were collected in vacutainertubes on the day of the GCF sampling. All patientsfasted overnight. Samples were centrifuged at 4,000rpm for 5minutes, and serumwas isolated and placedin sterile Eppendorf tubes that were wrapped se-curely and stored at -70�C until the day of laboratoryanalysis.

Quantification of IL-1b, TNF-a, IL-4,and IL-10 ContentAlthough GCF samples were analyzed for IL-1b,#

TNF-a,** IL-4,†† and IL-10,‡‡ serum samples wereanalyzed for IL-1b and IL-10 using enzyme-linkedimmunosorbent assay§§ following the instructions ofthe manufacturer. The assays were run separately foreach cytokine using the standard curves.

Statistical AnalysesStatistical analyses were performed with a softwareprogram.ii Power analysis was used, and sample sizewas calculated as 17 for the power 76.4. For the sta-tistical analyses, normality was tested with theShapiro-Wilk test. For normally distributed groups,parametric tests were used to compare group dif-ferences among groups. One-way analysis of vari-ance (ANOVA) was used to compare differencesamong groups. The Tukey test was used in con-junction with ANOVA to find which means are sig-nificantly different from one another. For the groups

§ Williams periodontal probe, Hu-Friedy, Chicago, IL.i Periopaper, Oraflow, Smithtown, NY.¶ Periotron 8000, Pro Flow, Amityville, NY.# Human IL-1b ELISA kit, BioSource Europe, Nivelles, Belgium.** Human TNF-a ELISA kit, BioSource Europe.†† Human IL-4 ELISA kit, BioSource Europe.‡‡ Human IL-10 ELISA kit, BioSource Europe.§§ Enzyme-linked immunosorbent assay, Oraflow.ii SPSS v.15 for Windows, IBM, Chicago, IL.

Cytokine Profiles in Rheumatoid Arthritis and Chronic Periodontitis Volume 84 • Number 1

86

that were not distributed normally, the Kruskal-Wallis test was used to compare differences amonggroups. The Dunn test was used to test each of theindividual tests for their significance. Values of P <0.05 were considered statistically significant.

RESULTS

Clinical FindingsDemographic variables and clinical periodontal pa-rameters are shown in Table 1. Periodontal pa-rameters were found to be significantly differentbetween the groups (P <0.05). Although PI wassignificantly higher in the RA group comparedwith the CP and control groups (P <0.05), GI, PD,and CAL were significantly higher in the CP group(P <0.05) compared with the RA and control groups.The number of missing teeth was highest in theRA group, then the CP group, and then the controlgroup (P <0.05), respectively.

Levels of IL-1b, TNF-a, IL-4, and IL-10 in GCFTable 2 summarizes the total amount and concen-trations of GCF IL-1b, TNF-a, IL-4, and IL-10 in thegroups. The total amount and concentration ofIL-1b, IL-4, and IL-10 were found to be statisticallydifferent for RA versus controls and CP versuscontrols (P <0.05). However, there was no differencebetween the RA and CP groups for all cytokines(P >0.05). The concentrations of IL-1b, TNF-a,IL-4, and IL-10 in the RA and CP groups were sta-tistically lower than those of the controls (P <0.05).

Levels of IL-1b and IL-10 in SerumThe total amount and concentration of IL-1b in theRA group were significantly lower than those ofother groups (P >0.05) (Table 3). The total amountand concentration of IL-10 were not significantlydifferent among the groups (P >0.05).

Cytokine Ratios in GCF and SerumAlthough the IL-1b/IL-10 ratio was higher in GCFof the RA and CP groups than that of controls(P >0.05) (Table 4), it was significantly lower inserum of the RA group compared with the CP andcontrol groups (P >0.05) (Table 5). The TNF-a/IL-4 ratio in GCF was significantly lower in theCP group (P >0.05) compared with the RA and controlgroups (Table 4). The IL-1b + TNF-a/IL-4 + IL-10GCF cytokine ratio (for concentrations) was not sig-nificantly different among the groups (Table 4).

DISCUSSION

In the present study, we evaluate the levels ofproinflammatory and anti-inflammatory cytokinesin the GCF and serum of patients with RA and pa-tients with CP to identify potential similarities in theimmunopathology of these diseases while usingT

able

1.

Clin

icalCharacteristicsoftheStudyGroups

RA

(n=17)

CP(n

=16)

Controls(n

=16)

Test

PValue

RA–Control

CP–Control

RA–CP

Age

(years)†

47.82–10.74(29to

70)

44.00–7.00(34to

54)

28.6

–6.18(20to

43)

ANOVA

0.02

**

‡

Females/m

ales

14/3

6/10

8/8

Num

ber

ofteeth†

22–4.47(14to

28)

25–2.60(20to

28)

27.75–0.57(26to

28)

ANOVA

0.03

**

*

RA

duration(years)†

6.08–6.14(2

to24)

NA

NA

Clinicalperiodontalparam

eters

PI†

1.44–0.49(1

to2.66)

1.34–0.43(0.5

to2.0)

0.36–0.41(0

to1)

ANOVA

0.02

**

*GI†

1.28–0.39(0.66to

2)

1.85–0.31(1.16to

2.33)

0.32–0.22(0

to0.66)

ANOVA

0.02

**

*PD

(mm)†

2.21–0.75(1

to4.33)

6.18–0.62(5.16to

7.33)

1.57–0.27(1.33to

2.33)

Kruskal-W

allis

0.03

**

*AL(m

m)†

2.80–1.16(1.33to

6.32)

7.28–0.95(5.16to

8.82)

1.57–0.68(1.33to

2.33)

Kruskal-W

allis

0.02

**

*

NA

=notapplic

able.

*P<0

.05.

†Mea

n–SD

(range).

‡Nosignifica

ntdifference

.

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87

otherwise healthy individuals asbackground controls. The peri-odontal parameters were signifi-cantly different among groups.However, there were no differ-ences between the RA andCP groups for all evaluated cyto-kines in GCF. Conversely, serumIL-1b had a significantly differentlevel between RA and CP.

The sex distribution in theRA group reflected the epidemi-ology of the disease; females(82.35%) are three times morelikely to develop RA than males(17.65%).8,27,33,34 Consistentwith previous reports, the agerange was 29 to 70 years (mean,47.82 years).8,34 Hence, ourevaluated RA population was rep-resentative.

RA and CP have remarkablysimilar pathobiology.9,12,35

Studies evaluating theassociationbetween CP and RA were con-ducted, but the findings wereinconsistent.8,23,34,36-40 Thesedifferences may be arising fromdifferent study designs and dif-ferent disease classifications. Re-cent evidence is accumulatingthat individuals with RA havehigher odds of periodontal dis-ease compared with controls,8,13,18,20,34,41 which is consistentwith our results.

The results of the present studyshow a relatively lower numberof missing teeth in the patientswith RA, contrary to other stud-ies.8,13,34,42,43 However, Mirandaet al.44 reported no differencebetween RA and controls for thenumber of missing teeth. Thepresent study shows a low level ofperiodontal disease prevalence inthe RA group compared with theCP group. This could be explainedby the regular use of NSAIDs bypatients with RA.14,41,45,46 In thepresent study, RA does not seemto have an additional detrimentaleffect on periodontal health alonewhen considering mean RA du-ration, which was 6 years in ourgroup.T

able

2.

TheTotalAmount(pg)andConcentrations(pg/m

L)ofIL-1

b,TNF-a,IL-4

,andIL-1

0in

GCFSamples

Cytokines

RA

(n=17)

CP

(n=16)

Controls

(n=16)

Test

PValue

RA–Control

CP–Control

RA–CP

IL-1b(pg)

131.42–111.04

(31.15to

385.09)

207.26–98.59

(37.33to

391.76)

119.69–118.29

(32.67to

357.40)

Kruskal-W

allis

0.01

**

†

IL-1b(pg/mL)

173.25–175.57

(19.79to

726.59)

163.98–107.33

(23.93to

421.24)

568.43–472.82

(60.51to

1,624.56)

Kruskal-W

allis

0.00

**

†

TNF-a(pg)

11.28–0.32

(11.05to

12.07)

11.47–0.33

(11.11to

12.33)

11.55–0.85

(11.12to

14.28)

ANOVA

0.363

††

†

TNF-a(pg/mL)

15.15–6.73

(5.66to

28.56)

9.41–4.26

(3.85to

21.59)

71.92–45.53

(21.20to

159.08)

Kruskal-W

allis

0.02

**

†

IL-4 (p

g)2,211.17–1,383.97

(342.92to

4,669.56)

2,996.49–1,061.12

(646.39to

4,849.04)

2,018.35–860.11

(569.69to

3,223.13)

Kruskal-W

allis

0.01

**

†

IL-4

(pg/mL)

3,041.73–2,355.73

(225.60to

8,810.50)

2,481.24–1,543.03

(429.84to

6,541.96)

10,616.14–7,338.97

(1,345.84to

28,150.19)

Kruskal-W

allis

0.00

**

†

IL-10(pg)

54.35–21.34

(44.76to

114.25)

71.65–83.79

(45.45to

385.14)

66.38–76.38

(31.95to

351.25)

ANOVA

0.03

**

†

IL-10(pg/mL)

74.63–46.83

(23.23to

202.67)

60.03–78.44

(15.95to

346.98)

338.03–219.91

(59.16to

798.29)

Kruskal-W

allis

0.03

**

†

Data

are

prese

ntedasmea

n–SD

(range).

*P<0

.05.

†Nosignifica

ntdifference

.

Cytokine Profiles in Rheumatoid Arthritis and Chronic Periodontitis Volume 84 • Number 1

88

In the present study, there are significant differ-ences among groups with regard to PI, GI, PD,and CAL. These parameters were higher in the CPgroup than the RA and control groups, exceptPI. The reason for the impaired oral hygiene andplaque accumulation may be the articular de-struction in patients with RA. Bozkurt et al.47

also reported higher PI scores in patients with RA,although other clinical periodontal parametersdid not reach significant difference between thegroups. However, the same group18 later reportedthat they found more severe gingival inflammation inCP patients than the patients with RA. Havemose-Poulsen et al.46 reported no difference betweenthe RA and control groups for plaque scores; how-ever, AL, PD, alveolar bone loss, and mean numberof missing teeth were significantly higher in theRA group compared with controls. Miranda et al.44

reported no difference between the RA and controlgroups, and Biyiko�glu et al.41 reported no differencebetween the RA and CP groups in clinical periodontalparameters.

In this study, the total amounts and concentrationsof evaluated cytokines in GCF are not significantlydifferent between the RA and CP groups. However,the RA and CP groups are significantly different com-pared with controls for all evaluated cytokines, ex-cept for the amount of TNF-a.

Bozkurt et al.18 reported higher levels of IL-4 andIL-10 in GCF of healthy controls compared with RAand CP patients and higher levels of IL-4 and IL-10 inRA patients than CP patients. Our findings confirmthe results of this study. Biyiko�glu et al.41 found nodifferences between RA and CP patients for the GCFlevels of IL-1bwith long-termusage ofmethotrexate.Miranda et al.44 found lower amounts of IL-1b in theGCF of patients with RA compared with controls.This result is compatible with our findings.

Contrary to a previous report by Havemose-Poulsen et al.,20 who reported no difference be-tween RA and controls for plasma IL-1b but notIL-10, the authors of the present study found thatthe serum IL-1b was lowest in patients with RAcompared with the CP and control groups, and,similarly, there was no difference for IL-10.

Similar to a recent paper by Esen et al.,40 patientswho have applied to rheumatology clinics consist ofindividuals who have already used NSAIDs or dis-ease-modifying anti-rheumatic drugs, such asmethotrexate, or who were unaware of which drugsthey used. Although RA is an inflammatory diseasethat has painful episodes, the patients with RA aretaking regular medication; accordingly, all patientsin the present study were using NSAIDs regularly.Hence, when assessing the results of the studies,one should consider the positive effects of theT

able

3.

TheTotalAmount(pg)andConcentration(pg/m

L)ofIL-1

bandIL-1

0in

Serum

Samples

Cytokines

RA(n

=17)

CP(n

=16)

Controls(n

=16)

Test

PValue

RA–Control

CP–Control

RA–CP

IL-1b(pg)

1,459.93–51.05

(1,398.20to

1,600.40)

1,602.41–143.76

(1,431.81to

1,941.20)

1,742.79–250.72

(1,428.40to

2,396.40)

Kruskal-W

allis

0.00

*†

*

IL-1b(pg/mL)

7.30–0.26

(6.99to

8.00)

8.01–0.72

(7.16to

9.71)

8.71–1.25

(7.14to

11.98)

Kruskal-W

allis

0.01

*†

*

IL-10(pg)

1,186.35–193.39

(1,088.40to

1,814.60)

1,358.17–710.40

(1,048.10to

3,961.50)

1,393.21–909.92

(1,082.50to

4,793.40)

Kruskal-W

allis

0.586

††

†

IL-10(pg/mL)

11.86–1.93

(10.88to

18.15)

13.58–7.10

(10.48to

39.62)

11.68–0.76

(10.83to

13.11)

Kruskal-W

allis

0.068

††

†

Data

are

prese

ntedasmea

n–SD

(range).

*P<0

.05.

†Nosignifica

ntdifference

.

J Periodontol • January 2013 Cetinkaya, Guzeldemir, Ogus, Bulut

89

rheumatologic agents on periodontal condition andgingival inflammation.8,41,44,48,49 NSAIDs were re-ported to suppress the synthesis and secretion ofIL-1b, IL-6, and PGE2,47,50 whereas methotrexateand corticosteroids have inhibitory effects on IL-1.41

Conversely, the use of NSAIDs for the treatment ofperiodontal disease has been studied. There areconflicting results; whereas Jeffcoat et al.51 reportedpositive effects of NSAIDs on periodontal diseaseprogression, Heasman and Seymour45 did not ob-serve significant differences for PI, GI, PD, and ALbetween the RA and control groups. NSAIDs aredocumented to limit periodontal bone loss in long-term use.8,52 Thomas and Carol53 showed thatmethotrexate reduces the levels of IL-1b in sy-novial fluid of patients with RA. Previously, it wasshown that the heavy medication used by juvenileidiopathic arthritis patients during 2 years resultedin decreased GCF IL-1b levels and improved peri-odontal condition.48 In two other studies, becausethe gingival inflammation did not decrease bythat time in the RA group, the authors concludedthat the decreased level of IL-1b was associated

with the rheumatologic agents.42,50 The findings ofthe current study are consistent with these reports.Mean RA duration of this group was 6 years, whichmeans that these patients with RA were undergoinglong-term antirheumatic therapy, and this may ex-plain the lower cytokine concentrations in patientswith RA.

In the present study, 15 of 17 patients with RAwere being treated with methotrexate–sulfasalazinecombined therapy. In recent years, methotrexate isthe most preferred drug in the treatment of RA.54 Theanti-inflammatory effect of methotrexate was ap-proved; methotrexate inhibits granulocyte functions,chemotaxis, protease activity, and leukotriene B4production.55 It was also shown that the use ofmethotrexate resulted in increases in IL-1b con-centration, leukocyte count, and ratio in synovialfluid of patients with RA.53

Sulfasalazine is an antimicrobial agent that hasanti-inflammatory effects. Anti-inflammatory effectsarise from inhibition of nuclear factor kappa a activa-tion that is an early stimulation for releasing proin-flammatory cytokines.56 Sulfasalazine stimulates

Table 4.

The Cytokine Ratios for GCF Samples

Cytokines RA (n = 17) CP (n = 16) Controls (n = 16) Test P Values RA–Control CP–Control RA–CP

IL-1b/IL-10 2.43 – 1.99(0.70 to 6.89)

3.97 – 2.20(0.32 to 8.59)

2.11 – 2.04(0.64 to 7.11)

ANOVA 0.02 * * †

IL-1b/IL-4 0.06 – 0.03(0.02 to 0.13)

0.08 – 0.06(001 to 0.26)

0.05 – 0.04(0.02 to 0.14)

ANOVA 0.369 † † †

TNF-a/IL-10 0.22 – 0.05(0.10 to 0.25)

0.22 – 0.06(0.03 to 0.27)

0.24 – 0.07(0.03 to 0.36)

ANOVA 0.078 † † †

TNF-a/IL-4 0.009 – 0.009(0.0 to 0.03)

0.005 – 0.004(0.0 to 0.02)

0.007 – 0.004(0.0 to 0.02)

ANOVA 0.00 † * *

IL-1b + TNF-a/IL-4 + IL-10 0.07 – 0.03(0.02 to 0.11)

0.08 – 0.06(0.01 to 0.27)

0.06 – 0.03(0.02 to 0.14)

ANOVA 0.524 † † †

Data are presented as mean – SD (range).* P <0.05.† No significant difference.

Table 5.

Cytokine Ratio for Serum Samples

Cytokines RA (n = 17) CP (n = 16) Controls (n = 16) Test P Value RA–Control CP–Control RA–CP

IL-1b/IL-10 0.63 – 0.08(0.40 to 0.70)

0.66 – 0.16(0.19 to 0.89)

0.72 – 0.18(0.16 to 0.92)

ANOVA 0.00 * † *

Data are presented as mean – SD (range).* P <0.05.† No significant difference.

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apoptosis of human T lymphocytes, which dependson concentration and duration of medication.

The strengths of the present study and the exami-nation of all evaluated patients were assessed bya periodontist (BC) who works in Baskent UniversityHospital Periodontology Clinics, and the patients withRA were diagnosed and controlled by staff rheumatolo-gists in the Baskent University Hospital and AdanaHospital Rheumatology Clinics. All included in-dividuals were non-smokers. Contrary to another re-port,34 all control individuals were recruited fromuniversity clinics; hence, our control population wasrepresentative of the general population.

Among the limitations of our study is the cross-sectional design47 and the relatively small size of oursample, which may limit the precision of the riskestimates.34 However, the present study does notaim to determine the epidemiologic aspect of thedisease.

Second, possible effects of NSAIDs used bypatients with RA might have an effect on the peri-odontal condition. Our conclusion about rheumaticdrugs, which were used by patients with RA in thepresent study, is that they have an effect on evalu-ated inflammatory mediators. We think that the ef-fects of rheumatic medication are not only determinedin systemic circulation and synovial fluid but also inGCF. When considering the higher levels of PI in RApatients, patients with RA are susceptible to peri-odontal diseases and maybe this was masked bythe NSAIDs.

In literature review, it was very difficult to compareour results to different studies because of a lack ofstandardization in classifications used for periodontaldisease and RA, disease severity, microbial exposure,informationaboutdrugregimen,RAduration, inclusioncriteria, and evaluated parameters in different bodycomponents, such as saliva, GCF, blood serum, andwhole blood cells.21,57

Toeliminatecontradictionsamongdifferent studiesand verify the associations between RA and peri-odontal disease, well-defined inclusion criteria andclassification systems should be used and controlled,and longitudinal clinical and laboratory study set-upsshould be designed.

CONCLUSIONS

The following conclusions could be drawn withinthe limitations of the present study. 1) No significantdifferences in proinflammatory and anti-inflammatorycytokine levels between patients with RA and pa-tientswithCPwereobserved.2)However, theclinicalperiodontal parameters were found to be signifi-cantly different between the groups. 3) The reasonfor these inconsistent results may be explained by

the possibility that patients with RA may benefitfrom NSAIDs and rheumatoid agents for hostmodulations.

ACKNOWLEDGMENTS

The authors thank Ms. Mine Yildirim and Mrs. BurakAkcakanat, Istanbul, Turkey, for editing for Englishgrammar. The authors report no conflicts of interestrelated to this study.

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Correspondence: Dr. Sule Bulut, Baskent University,Faculty of Dentistry, Department of Periodontology, 11.Sokak, No. 26 Bahcelievler, 06490 Ankara, Turkey. Fax:90-312-215-2962; e-mail: suleb@baskent.edu.tr.

Submitted August 9, 2011; accepted for publicationFebruary 2, 2012.

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