10.59987/ads/2013.2.191-195
Aim. Periodontal diseases entail a variety of conditions affecting the periodontium. The pathogenesis results from a complex interaction of genetic and environmental factors. Although there are evidences to confirm a role of genetic determinants, the outcome of the available studies is controversial and the largest part of the research has been carried out in Asian populations.
Methods. We investigated two polymorphisms in the genes encoding Interelukin-1β (IL-1β +3953 C&gt;T; rs1143634) and vitamin D receptor (VDR Taq1; rs731236) in 42 Caucasian patients with chronic periodontal disease and 39 Caucasian subjects, matched for age and gender.
Results. The IL-1β C allele was present in 100% of cases and 92% of controls (p=0.07), the T allele was present in 19% of cases and in 44% controls (p=0.017). The prevalence of the VDR Taq1 tt genotype was lower in patients as compared with controls (i.e., 10 versus 59%; p&lt;0.01), whereas the tT and TT genotypes were disproportionally higher in patients than in cases (i.e., 62 versus 33% for tT and 29% versus 8% for TT; p&lt;0.01). The t allele was present in 71% of cases and 92% of controls (p=0.016), whereas the T allele was present in 90% of patients with periodontal disease and in 41% controls (p&lt;0.01).
Conclusion. The results of this case control study attest that the T allele of VDR Taq1 is strongly associated with periodontal disease, whereas the t allele of the IL-1β +3953 confers a slightly protection against the risk of periodontitis.
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Annali di stomatologia
gene expression
Association between periodontal disease and Interleukin-1β +3953 and vitamin D receptor Taq1 genetic polymorphisms in an Italian caucasian population
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Association between periodontal disease and Interleukin-1β +3953 and vitamin D receptor Taq1 genetic polymorphisms in an Italian caucasian population
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                                                                                                                  Original article



                   Association between periodontal disease and
                   Interleukin-1β +3953 and vitamin D receptor Taq1
                   genetic polymorphisms in an Italian caucasian




                                                                                                                                    li
                   population




                                                                                na
                   Alberto Baldini, PhD, DDS1                             41% controls (p<0.01).
                   Alessandro Nota, DDS1                                  Conclusion. The results of this case control study at-




                                                                              io
                   Elena Fanti, BSc2                                      test that the T allele of VDR Taq1 is strongly associ-
                   Francesco Saverio Martelli, DMD, IRF3                  ated with periodontal disease, whereas the t allele of
                   Cosimo Ottomano, MD4                                   the IL-1β +3953 confers a slightly protection against
                   Giuseppe Lippi, MD4                                    the risk of periodontitis.




                                                                           az
                                                                          Key words: periodontal diseases, periodontitis, ge-
                   1 Department of Orthodontics,                          netics, interleukins, gene expression.
                     University of Rome “Tor Vergata”, Italy




                                                                        rn
                   2 Biomolecular Diagnostic, Firenze, Italy
                   3 IRF Microdentistry, Firenze, Italy                   Introduction
                   4 U.O. Ematochimica Diagnostics,




                                                                     te
                     University Hospital of Parma, Italy                  Periodontal diseases encompass a variety of conditions
                                                                          affecting the periodontium. A simple classification of this
                                                                          condition has been agreed upon in 1999, during the Inter-
                   Corresponding author:                                  national Workshop for a Classification of Periodontal Dis-
                   Alberto Baldini, PhD, DDS
                   Centro Medico Polispecialistico Baldini Srl
                                                                  In      eases and Conditions, and is now widely used by clini-
                                                                          cians and research scientists worldwide (1).
                   Via S. Orsola, 5                                       Basically, this classification ranks periodontal disease and
                   24048 Bergamo, Italy                                   conditions into eight leading categories, that are gingival
                   Phone: +39 035 271935                                  diseases (including dental plaque-induced gingival dis-
                                                             ni
                   E-mail: studiomedicobaldini@gmail.com                  eases, non-plaque-induced gingival lesions), chronic peri-
                                                                          odontitis (either localized or generalized), aggressive peri-
                                                                          odontitis (either localized or generalized), periodontitis as
                   Summary                                                a manifestation of systemic diseases (i.e., associated with
                                             o

                                                                          hematological disorders, genetic disorders, or not other-
                   Aim. Periodontal diseases entail a variety of condi-   wise specified), necrotizing periodontal diseases (which
                                         izi


                   tions affecting the periodontium. The pathogenesis     can be either necrotizing ulcerative gingivitis or necrotiz-
                   results from a complex interaction of genetic and      ing ulcerative periodontitis), abscesses of the periodon-
                   environmental factors. Although there are evi-         tium, periodontitis associated with endodontic lesions, as
                   dences to confirm a role of genetic determinants,      well as developmental or acquired deformities and condi-
                          Ed




                   the outcome of the available studies is controver-     tions (1, 2).
                   sial and the largest part of the research has been     According to the Research, Science and Therapy Com-
                   carried out in Asian populations.                      mittee of the American Academy of Periodontology, the
                   Methods. We investigated two polymorphisms in          prevalence of severe generalized periodontitis is com-
                   the genes encoding Interelukin-1β (IL-1β +3953         prised between 5% and 15% of the general population,
                   C>T; rs1143634) and vitamin D receptor (VDR Taq1;      although moderate disease may affect a majority of
               C




                   rs731236) in 42 Caucasian patients with chronic pe-    adults (2). Accordingly, the prevalence of gingivitis
                   riodontal disease and 39 Caucasian subjects,           among school children in the US ranges from 40% to
    CI




                   matched for age and gender.                            60%, whereas 47% of males and 39% of females aged
                   Results. The IL-1β C allele was present in 100% of     18 to 64 exhibit at least one site which bled on probing
                   cases and 92% of controls (p=0.07), the T allele was   (BOP). Although reliable data attests that relatively few
                   present in 19% of cases and in 44% controls            sites with gingivitis progress to develop manifest peri-
                   (p=0.017). The prevalence of the VDR Taq1 tt geno-     odontitis, a genetically determined response has been
                   type was lower in patients as compared with con-       hypothesized, with non-smoking or former smoking, in-
©




                   trols (i.e., 10 versus 59%; p<0.01), whereas the tT    terleukin (IL)-1 genotype-positive individuals having
                   and TT genotypes were disproportionally higher in      greater risk of BOP than non-smoking and former
                   patients than in cases (i.e., 62 versus 33% for tT     smoking IL-1 negatives (3).
                   and 29% versus 8% for TT; p<0.01).                     As specifically regards the pathogenesis, the old view
                   The t allele was present in 71% of cases and 92% of    of periodontitis as the outcome of infection is now over-
                   controls (p=0.016), whereas the T allele was present   come by the new concept that the disease would
                   in 90% of patients with periodontal disease and in     emerge from a complex interaction of several genetic

                   Annali di Stomatologia 2013; IV (2): 191-195                                                                    191
4-Baldini_- 19/06/13 10:47 Pagina 192




                   A. Baldini et al.


                   and environmental factors (4-6), including bacterial in-       Table 1. Leading characteristics of the study population.
                   fection, host response and inflammation, with tobacco          Values are shown as mean ± standard deviation.
                   smoking, ageing, male gender and low socioeconomic             PPD, probing pocket depth; BOP, bleeding on probing.
                   status being considered other important risk factors (7,
                   8). The host response mediated by several cytokines is                                 Chronic                Healthy




                                                                                                                                             li
                   now regarded as a leading aspect in the clinical expres-                               periodontal            controls
                   sion of periodontitis, and several inflammatory biomark-                               disease




                                                                                      na
                   ers have been consistently associated with periodontal         n                       42                     39
                   disease, especially prostaglandin E2 (PGE2), tumor             Age (yrs)               60±9                   55±15
                   necrosis factor-alpha (TNF-α), Interleukin-1 alpha (IL-
                   1α), and Interleukin-1 beta (IL-1β) (9, 10). Interestingly,    Gender, females (%)     27 (64%)               30 (77%)




                                                                                    io
                   only 20% of periodontal diseases is now attributed to          PPD (mm)                4,17                   1,34
                   bacterial variance, 20% to tobacco use, whereas 50%
                                                                                  Presence of PUS (%)     58                     0
                   of disease expression has been attributed to genetic
                                                                                  BOP (%)                 99                     26




                                                                                 az
                   variance at several loci, including specific polymor-
                   phisms at the loci encoding for some cytokines (i.e., IL-
                   1α, IL-1β, IL-6, TNF-α) (9), as well as for the vitamin D
                   receptor (VDR) gene (10).                                      Hilden, Germany). Allelic analysis was performed using
                   Although some studies (11) and meta-analyses (12, 13)




                                                                              rn
                                                                                  Taq Man1 SNP Genotyping Assays rs731236 and
                   are now available to attribute a certain the role to these     rs1143634 (Applied Biosystems, Forster City, CA, USA),
                   genetic determinants in the pathogenesis of periodontal        which are based on a predesigned mix of unlabeled
                   disease, the outcome is often controversial. As such,
                                                                                  polymerase chain reaction (PCR) primers and the Taq




                                                                           te
                   the aim of this article was to investigate whether two
                                                                                  Man® minor groove binding group (MGB) probe (FAM™
                   polymorphisms in the genes encoding IL-1β (i.e., IL-1β
                                                                                  and VIC® dye-labeled), on the instrument Step One (Ap-
                   +3953 C>T; rs1143634) and VDR (i.e., VDR Taq1; rs
                                                                                  plied Biosystems). Thermocycler conditions were an ini-
                   731236) are associated with periodontal disease in a
                                                                                  tial 35 s denaturation at 95° C, followed by 40 cycles of
                   Caucasian population.
                                                                        In        95° C for 10 s and 60° C for 45 s. The significance of
                                                                                  differences between cases and controls was evaluated
                                                                                  with Chi-Square test using Analyse-it for Microsoft Ex-
                   Materials and methods
                                                                                  cel (Analyse-it Software Ltd, Leeds, UK).
                                                          ni
                   The study population consisted in 42 Caucasian pa-
                   tients (27 females and 12 males, mean age of 60±9
                   years) with chronic periodontal disease diagnosed ac-
                                                                                  Results
                   cording to the well-established criteria of the Interna-
                                            o

                   tional Workshop for a Classification of Periodontal Dis-       The results of IL-1β +3953 are shown in Figure 1. A sig-
                   eases and Conditions (1).                                      nificant difference was observed in the overall distribution
                                        izi


                   Accordingly, each of the 42 patients with chronic periodon-    of C/T polymorphisms (Pearson Chi-Square = 7.106;
                   tal disease had severe and generalized form of periodon-       DF=2; p=0.029), in that patients with periodontal disease
                   tal disease, showing at least 5 sites with probing pocket      exhibited a higher prevalence of the CC genotype (81
                   depth (PPD) >6 mm located in different teeth and distrib-      versus 56%), whereas the TT genotype was present in
                          Ed




                   uted among the four quadrants, BOP and pus. The control        8% of controls but in none of the cases (Fig. 1). The het-
                   group consisted in 39 Caucasian subjects matched for           erozygous allele CT was present in 19% of cases versus
                   age and gender (30 females and 9 males, mean age               36% of controls. Globally, the C allele was present in
                   56±15 years) not affected by periodontal disease, i.e., with   100% of cases and 92% of controls (Pearson Chi-
                   no history of periodontal disease, without periodontal         Square = 3.36; DF=1; p=0.07), whereas the T allele was
                   pockets >3 mm and radiographic evidence of bone loss           present in 19% of patients with periodontal disease and
               C




                   (Tab. 1).                                                      in 44% without (Pearson Chi-Square = 5.71; DF=1;
                   All the subjects were enrolled in the study after signing      p=0.017).
    CI




                   an inform consent form, and the experimental protocol          An even more statistically significant associations was
                   was accepted by the ethical committee.                         however found for the VDR Taq1 polymorphism (Fig. 2)
                   No case or control subject was affected by systemic            (Pearson Chi-Square = 23.024; DF = 2; p<0.01), in that
                   diseases that are known to influence development or pro-       the prevalence of the tt genotype very low in patients
                   gression of periodontal disease (e.g., diabetes), or was       with periodontal disease as compared with controls (i.e.,
                   pregnant, currently smoking or using anti-inflammatory         10 versus 59%), whereas the tT and TT genotypes were
©




                   drugs. Each patient signed the informed consent to be          disproportionally higher in patient than in cases (i.e., 62
                   included in the study, which was carried out in accor-         versus 33% for tT and 29% versus 8% for TT). Overall,
                   dance with the Declaration of Helsinki, and under the          the prevalence of the t allele was present in 71% of cas-
                   terms of all relevant local legislation. Genomic testing       es and 92% of controls (Pearson Chi-Square = 5.84;
                   was carried out after collecting material with a sterile       DF=1; p=0.016), whereas the T allele was present in
                   foam tipped applicator. The DNA was extracted with a           90% of patients with periodontal disease and in 41%
                   commercial kit purchased from Qiagen (Qiagen GmbH,             controls (Pearson Chi-Square = 22.25; DF=1; p<0.01).

                   192                                                                              Annali di Stomatologia 2013; IV (2): 191-195


             192
4-Baldini_- 19/06/13 10:47 Pagina 193




                   Association between periodontal disease and Interleukin-1β +3953 and vitamin D receptor Taq1 genetic polymorphisms
                   in an Italian caucasian population

                                                                                                                    Figure 1. Results for
                                                                                                                    IL-1β +3953.




                                                                                                                                        li
                                                                                     na
                                                                                   io
                                                                                az
                                                                             rn
                                                                          te
                                                                                                                    Figure 2. Results for
                                                                                                                    VDR Taq1.
                                                                       In
                                             o               ni
                                         izi
                          Ed




                   Discussion and conclusion                                     Asian populations did not allow to draw definitive con-
                                                                                 clusions on this topic.
               C




                   Periodontal disease is a public health problem afflicting     The results of our investigation, which were limited to
                   the vast majority of adults in mild to moderate forms         the two genetic polymorphisms for which more solid ev-
    CI




                   and being associated with important clinical implica-         idence are available (i.e., IL-1β +3953 and VDR Taq1),
                   tions not only for dental medicine, but also for its causal   attest that patients who are more prone to develop
                   relationship with a variety of other important disorders,     chronic periodontitis may have an important genetic
                   namely cardiovascular disease (13, 14). Recent and re-        predisposition.
                   markable advances in our understanding of the patho-          This finding may have important clinical implications in
                   genesis of periodontal disease have led to a radical ap-      terms of genotype-based risk assessment, early pre-
©




                   proach for prevention and treatment, which now deeply         vention of predisposed individuals (e.g., smoking and
                   involve the assessment of personal susceptibility. The        genetic interactions are important contributory factor in
                   outcome of several epidemiological investigations have        severity of periodontitis), and more aggressive treat-
                   allowed to identify a variety of genetic polymorphisms        ment of patients at greater risk.
                   that may be associated with periodontal disease (7,           The two genotypes that have been assessed in this
                   15), but the heterogeneity of the studies, the different      study exert different but synergic role in the pathogene-
                   diagnostic criteria as well as the prevalent enrolment of     sis of periodontal disease. The pivotal role of the IL-1

                   Annali di Stomatologia 2013; IV (2): 191-195                                                                       193
4-Baldini_- 19/06/13 10:47 Pagina 194




                   A. Baldini et al.


                   family of interleukins as mediators of inflammatory re-        odontal disease, so that mediators of bone metabolism
                   actions has been well established. Both IL-1α and IL-          like the VDR and its genetic polymorphisms may play a
                   1β exert similar proinflammatory activities, whereas IL-       role in the pathogenesis (19). Unfortunately, the poten-
                   1 receptor antagonist (IL-1Ra) is an anti-inflammatory         tial mechanism/s linking VDR and its gene polymor-
                   cytokine that counteract the former interleukins by bind-      phisms with periodontal disease are still largely ob-




                                                                                                                                                  li
                   ing to the IL-1 receptor (16).                                 scure. Some hypothesis were postulated, including
                   Although some similarities exist between IL-1α and IL-         negative influences on bone mineral density, turnover,




                                                                                      na
                   1β, the latter is a different molecular form, which is also    and bone loss (10, 20). In 2011 Deng et al. carried out
                   known as catabolin. IL-1β exerts a kaleidoscope of bio-        a database search retrieving 15 studies involving 1338
                   logical functions; it is an important mediator of the in-      cases and 1302 controls, and found that that chronic
                   flammatory response and actively involved in several           periodontitis cases had a weak significantly higher fre-




                                                                                    io
                   cellular activities such as cell proliferation, differentia-   quency of TT genotype of VDR Taq1 (OR = 1.86, 95%
                   tion, and apoptosis. In a seminal article by Liu et al., the   CI, 1.00, 3.46; p = 0.049) in Asians (10). These results
                   severity of periodontal disease (in terms of gingival in-      are globally in agreement with those obtained in our




                                                                                 az
                   dex, probing depth and gingival crevicular fluid flow)         case control study in a Caucasian population, since we
                   was first associated with IL-1β activity (17). Hou et al.      observed that the prevalence of the T allele was more
                   also reported that the tissue concentration of IL-1β de-       than twice in with patients with periodontal disease as
                   termined from diseased gingivae were remarkably high-          compared with healthy controls (i.e., 90 versus 41% P
                   er than those of controls from both healthy sites of peri-     < 0.01).




                                                                              rn
                   odontitis and non-periodontitis subjects (18), thereby         Taken together, the results of this case control study at-
                   confirming that IL-1β plays a significant role in the path-    test that the T allele of VDR Taq1 are strongly associat-
                   ogenic mechanisms of periodontal tissue destruction            ed with periodontal disease, whereas the t allele of the




                                                                           te
                   and that its assessment in periodontal tissue might be a       IL-1β +3953 confers a slightly protection against the
                   valuable aid and useful diagnostic marker of disease.          risk of periodontitis.
                   The important role played by IL-1β on periodontal dis-
                   ease has been attributed to the potency of inducing
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                   Annali di Stomatologia 2013; IV (2): 191-195                                                                                          195
