Research Article - Biomedical Research (2017) Volume 28, Issue 20
Impact of glucocorticoid receptor gene Bcl-1 variant on temporomandibular disordersMehmet Kemal Tumer1,2*, Kaan Yerliyurt3, Ayse Nursal4, Nevin Karakus2, Akin Tekcan5 and Serbulent Yigit2
- *Corresponding Author:
- Mehmet Kemal Tumer
Department of Oral and Maxillofacial Surgery
Accepted date: September 27, 2017
Objectives: Temporomandibular Disorders (TMD) constitute a heterogeneous group of disorders characterized by alterations in mandibular movement. The aim of this study was to investigate the association between the Bcl1 variant of NR3C1 gene and TMD susceptibility in Turkish population.
Method: NR3C1 gene BcI1 variant of 100 TMD patients and 105 healthy controls was genotyped by polymerase chain reaction-restriction fragment length polymorphism method (PCR-RFLP).
Results: There was also no significant difference in regard to genotype and allele frequencies between the patients and the controls (OR 0.216 (95% Cl: 0.85-2.04); p=0.216). However, present study found that numeric pain rating scale was higher in patients with CC and CG genotypes.
Discussion: Although the NR3C1 Bcl1 variant did not show any difference between the TMD and the control groups, we thought that this variant could be correlated with pain intensity in patients. Further studies with different ethnic subjects are needed to confirm the results.
Temporomandibular disorders, Glucocorticoid receptor gene, Bcl1 variant.
Temporomandibular disorders (TMD) are a kind of disease group that manifest itself as musculoskeletal and neuromuscular conditions . It is characterized by recurrent or chronic pain in the temporomandibular region, disc displacement with clicking and crepitus noises produced during mandibular movement. The clear etiology of TMD has not been established yet. It is believed that genetic risk factors can play a role in the etiology of TMD, as suggested by a twin study that ascribed 44% of TMD to genetic inheritance of variations . In epidemiologic studies, it has been shown that women have a higher risk for TMD .
Glucocorticoids (GCs) play a crucial role in the regulation of several processes in the human body. They display their influences by binding to the glucocorticoid receptor . The human glucocorticoid receptor gene (NR3C1) is localized on chromosome 5q31-32, includes 9 exons and it regulates the coding of the NR3C1 protein, which binds glucocorticoid hormones in the liver, muscle, and vasculature, affecting the metabolism and the cardiovascular function . GCs may lower inflammatory gene expression by interfering with the inflammatory transcription factors and by facilitating the transcription of genes with anti-inflammatory activity. Inflammatory proteins cause an increase in bone turnover by inducing osteoclastic activity. The amount of Glucocorticoid receptors, which could be affected by the polymorphisms on NR3C1 gene, will determine the amount of free GCs and consequently the amount of inflammatory proteins . The increase in inflammatory proteins may lead to TMD as well as to other joint disorders. On the other hand, TMD is also thought to be one of the physical results of stress. Stress is one of the causes of elevated free cortisol levels. Cortisol, the basic glucocorticoid in circulating, shows its effect by binding especially the glucocoticoid receptors in the hippocampus .
The NR3C1 gene has several polymorphic sites. The Bcl1 variant (rs41423247) of the NR3C1 gene, a C to G substitution located in intron 2, is correlated with the enhanced glucocorticoid sensitivity and higher cortisol levels .
The aim of this present study was to investigate the association between Bcl1 variant of NR3C1 gene and TMD susceptibility in a Turkish population.
One hundred patients with TMD (77 females and 23 males; aged 18-71) were included in the study. Patients were recruited from the Department of Oral and Maxillofacial Surgery during the eight months period between July 2015 to March 2016. The diagnosis of TMD was based on the criteria described by Schiffman et al. (Table 1) . A detailed medical history was taken, followed by a complete oral examination. Additionally, a control group consisting of 105 unrelated healthy subjects (69 females and 36 males; aged between 19 and 69) with similar ethnic background and residing in the same geographic area with the patients was formed. Subjects with no evidence of chronic disease were included in the control group. Informed written consent was obtained from each subject according to the Declaration of Helsinki, and design of the work (15-KAEK-124) was approved by the Local Ethical Committee.
|Temporomandibular joint (TMJ) disorders||42 (42.0)|
|Masticatory muscle disorders||2 (2.0)|
|TMJ disorders+Masticatory muscle disorders||5 (5.0)|
|TMJ disorders+Headache||38 (38.0)|
|Masticatory muscle disorders+Headache||3 (3.0)|
|TMJ disorders+Masticatory muscle disorders+Headache||10 (10.0)|
Table 1. The diagnosis of TMD patients according to criteria described by Schiffman et al. .
Blood samples were collected in Ethylene Diamine Tetra- Acetic acid (EDTA)-coated tubes from the patients with TMD and control subjects, and stored at -20°C until use. Genomic DNA was extracted using commercial kit (Sigma–Aldrich, Taufkirchen, Germany) according to manufacturer protocol. The BclI variant in the intron 2 of NR3C1 gene was evaluated using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. PCR reaction was performed as described previously . PCR products were cut with Ksp 221 restriction enzyme (Sigma-Aldrich, Taufkirchen, Germany). Then, the digestion products were separated on 3% agarose gels stained with ethidium bromide and photographed with a standard ultraviolet transilluminator. Wild-type (CC) individuals had 2 fragments of 90 and 116 bp. Individuals with the GG genotype had an uncut fragment of 206 bp. The heterozygotes (GC) had fragments of 90, 116, and 206 bp.
All statistical analyses were performed using IBM SPSS 20.0 and OpenEpi 2.3.1 software. Continuous data were presented as mean ± SD (standard deviation). Chi square test was used to detect the significance of differences in the allele frequencies and genotype distributions between the two study groups. The association between Bcl1 variant and the clinical and demographic characteristics of patients and clinical characteristics of pain of TMD patients were analyzed by using χ2 test or analysis of variance (ANOVA) statistics. Hardy- Weinberg equilibrium test was performed for both study groups. Odds ratio (OR) and 95% confidence intervals (CIs) were calculated. A p value of <0.05 was considered statistically significant.
Two hundred and five Turkish individuals (105 controls and 100 patients) participated in the study. There were 69 female and 36 male participants in the control group, and the patient group consisted of 77 females and 23 males. The females constituted the majority of cases in both patient and control groups. The mean age was 34.92 ± 13.343 and 36.76 ± 11.255 in the patient and in the control group, respectively. The demographic features of the study subjects are shown in Table 2.
|Demographical characteristics||TMD patients||Healthy controls||p|
|Age, mean ± SD (y)||34.92 ± 13.343||36.76 ± 11.255||0.286|
|Gender, n (%)||0.09|
|Male||23 (23.0)||36 (34.3)|
|Female||77 (77.0)||69 (65.7)|
Table 2. The demographical characteristics of TMD patients and healthy controls.
Genotypic and allelic distributions of the NR3C1 Bcl1 variant in patient and control groups are presented in Table 3. The CC, CG, GG genotypes of the NR3C1 Bcl1 variant were observed in 49.5%, 40.0%, and 10.5% of control subjects and in 57%, 36% and 7.0% of patients, respectively. Genotype distribution did not show any significant difference between patients and controls according to NR3C1 Bcl1 variant (p˃0.05). The frequency of Bcl1 C allele was 75% (n=150) and that of G allele was 25% (n=50) in the patient group. There were not any significant differences of allele frequencies between patients and controls (OR 0.216 (95% Cl: 0.85-2.04); p=0.216).
|Polymorphism||Patients n=100 (%)||Controls n=105 (%)||p||OR (CI 95%)|
|CC||57 (57.0)||52 (49.5)||0.482|
|CG||36 (36.0)||42 (40.0)|
|GG||7 (7.0)||11 (10.5)|
|CC:CG+GG||57 (57.0):43 (43.0)||52 (49.5):53 (50.5)||0.285||1.35 (0.78-2.35)|
|CC+CG:GG||93 (93.0):7 (7.0)||94 (89.5):11 (10.5)||0.379||1.55 (0.57-4.42)|
|C||150 (75.0)||146 (69.5)||0.216||1.31 (0.85-2.04)|
|G||50 (25.0)||64 (30.5)|
Table 3. Genotype and allele frequencies of NR3C1 Bcl1 variant in patient and control groups.
Furthermore, this present study also analyzed if any differences existed in clinical and demographic characteristics of patients according to genotype distribution. The clinical and demographic characteristics according to genotype distribution are presented in Table 4. There was not any significant difference between genotype and age, gender, disease duration, or clinical parameters such as bruxism, jaw joint clicking or popping, joint locking and sounds related to these findings (p>0.05).
|NR3C1 Bcl1 variant|
|Characteristics||Total n=100||CC n=57||CG n=36||GG n=7||P value|
|Age, mean ± SD (y)||34.92 ± 13.343||33.88 ± 12.490||36.67 ± 13.879||34.43 ± 18.137||0.619|
|Gender, n (%)|
|Male||23 (23.0)||15 (26.3)||7 (19.4)||1 (14.3)||0.634|
|Female||77 (77.0)||42 (73.7)||29 (80.6)||6 (85.7)|
|Duration of disease, n (%)|
|<1 y||27 (27.0)||18 (31.6)||6 (16.7)||3 (42.9)||0.374|
|1-5 y||36 (36.0)||21 (36.8)||13 (36.1)||2 (28.6)|
|>5 y||37 (37.0)||18 (31.6)||17 (47.2)||2 (28.6)|
|Family history of TMD, n (%)|
|Yes||49 (49.0)||24 (42.1)||22 (61.1)||3 (42.9)||0.192|
|No||51 (51.0)||33 (57.9)||14 (38.9)||4 (57.1)|
|History of systemic disease, n (%)|
|Yes||63 (63.0)||40 (70.2)||18 (50.0)||5 (71.4)||0.13|
|No||37 (37.0)||17 (29.8)||18 (50.0)||2 (28.6)|
|Bruxism, n (%)|
|Yes||55 (55.0)||31 (54.4)||21 (58.3)||3 (42.9)||0.746|
|No||45 (45.0)||26 (45.6)||15 (41.7)||4 (57.1)|
|Eating disorders, n (%)|
|Yes||33 (33.0)||15 (26.3)||16 (44.4)||2 (28.6)||0.188|
|No||67 (67.0)||42 (73.7)||20 (55.6)||5 (71.4)|
|Sound in TMJ (jaw joint clicking or popping), n (%)|
|Yes||73 (73.0)||42 (73.7)||25 (69.4)||6 (85.7)||0.664|
|No||27 (27.0)||15 (26.3)||11 (30.6)||1 (14.3)|
|TMJ locking (open or closed), n (%)|
|Yes||8 (8.0)||5 (8.8)||2 (5.6)||1 (14.3)||0.7|
|No||92 (92.0)||52 (91.2)||34 (94.4)||6 (85.7)|
Table 4. Clinical and demographical characteristics of TMD patients stratified according to NR3C1 Bcl1 variant.
Clinical characteristics of pain of TMD patients were stratified according to NR3C1 Bcl1 variant are shown in Table 5. Pain intensity in patients was measured using the numeric pain rating scale (NPRS) (1-10). A "numeric pain" score ranging from 0 (no pain) to 10 (maximum pain) was constructed. Eleven patients who had rare pain were not evaluated. In terms of correlation between genotype distribution of the NR3C1 Bcl1 variant and clinical parameters such as pain during sleep, chewing or speaking, pain localization, pain period, factors that trigger pain, pain type and pain duration, present study didn't find any statistical significance among the groups as well. However, we noted a significant association of NR3C1 Bcl1 variant with numeric pain rating scale (p=0.048). The numeric pain rating score was higher in patients with CC and CG genotype.
|NR3C1 Bcl1 variant|
|Characteristics||Total n=89||CC n=48||CG n=34||GG n=7||P value|
|The severity of pain (The numeric pain rating scale (1-10)), mean ± SD||3.89 ± 1.957||3.77 ± 1.753||4.35 ± 2.116||2.43 ± 1.902||0.048|
|Pain during sleep, n (%)|
|Yes||49 (55.1)||27 (56.3)||20 (58.8)||2 (28.6)||0.332|
|No||40 (44.9)||21 (43.7)||14 (41.2)||5 (71.4)|
|Pain during chewing and speaking, n (%)|
|Yes||55 (61.8)||31 (64.6)||22 (64.7)||2 (28.6)||0.169|
|No||34 (38.2)||17 (35.4)||12 (35.3)||5 (71.4)|
|The localization of pain, n (%)|
|Muscle||9 (10.1)||5 (10.4)||4 (11.8)||0||0.866|
|Joint||69 (77.5)||38 (79.2)||25 (73.5)||6 (85.7)|
|Muscle and joint||11 (12.4)||5 (10.4)||5 (14.7)||1 (14.3)|
|Period of pain, n (%)|
|Chronic||27 (30.3)||10 (20.8)||15 (44.1)||2 (28.6)||0.077|
|At regular intervals||62 (69.7)||38 (79.2)||19 (55.9)||5 (71.4)|
|Factors that trigger pain|
|Movement||53 (59.6)||26 (54.2)||21 (61.8)||6 (85.7)||0.48|
|Cold||17 (19.1)||11 (22.9)||5 (14.7)||1 (14.3)|
|Movement and cold||19 (31.3)||11 (22.9)||8 (23.5)||0|
|Types of pain|
|Blunt||43 (48.3)||23 (47.9)||16 (47.1)||4 (57.1)||0.859|
|Sharp||43 (48.3)||24 (50.0)||16 (47.1)||3 (42.9)|
|Pulse type||3 (3.4)||1 (2.1)||2 (5.9)||0|
|The duration of pain|
|<1 h||39 (43.8)||21 (43.8)||14 (41.2)||4 (57.1)||0.935|
|≥ 1 h||39 (43.8)||21 (43.8)||16 (47.1)||2 (28.6)|
|Constant||11 (12.4)||6 (12.5)||4 (11.8)||1 (14.3)|
Table 5. Clinical characteristics of pain of TMD patients stratified according to NR3C1 Bcl1 variant.
GCs are an essential class of endogenous steroid hormones that modulate vital biological processes such as growth, development, metabolism, behavior and apoptosis . They have been considered to hinder activation of the hypothalamicpituitary- adrenocortical (HPA) axis through a delayed feedback system that is regulated by GC levels and that involves genomic changes. The glucocorticoid receptor, a 94kD cytosolic protein, mediates the physiological effects of GCs. NR3C1, also known as glucocorticoid receptor gene, has been widely studied in various inflammatory and autoimmune diseases such as Graves opthalmopathy, asthma, cystic fibrosis, and inflammatory bowel disease [12-15]. There are several variants in the NR3C1 gene. NR3C1 Bcl1 variant has been associated with hypersensitivity to GCs and regulation of the HPA axis activity . The Bcl1 variant acts as part of a conserved haplotype and has frequently been associated with mood and mental diseases . Numerous studies suggested associations of this haplotype with alterations in metabolism including hyperinsulinemia, increased abdominal fat, higher mass index (BMI), raised leptin levels, and larger increases in body weight after experimentally induced overfeeding, in C genotype carriers . Watt et al. noticed that the higher allele of Bcl1 variant is more frequent in individuals genetically predisposed to develop hypertension .
The temporomandibular joint, which is one of the most complicated joints in human body, plays an important role in functions such as jaw motion, speaking, chewing, and swallowing. TMD is a common term covering a number of clinical conditions that involve the masticatory masculature, the temporomandibular joint, and the related structures. Even though clear etiology of TMD has not been established, there is a common belief for a bio-psychosocial and multifactorial background, indicating the complex interaction between biological mechanism, mental conditions and psychological traits, environmental factors, and macro-microtrauma .
TMD can lead to pain and functional impairment, impede quality of life and frequently result in depressive symptoms . Recent studies suggest that shifted basal and stressinduced HPA activity can be associated with painful idiopathic conditions like fibromyalgia and irritable bowel syndrome [21,22]. These clinical conditions may be also associated with inter-individual variations in HPA axis activity and modified glucocorticoid impacts. TMD is often seen together with other psychosomatic symptoms such as sleep disorders, headache, fatigue, and depression, all of which belong to somatic syndromes . It was also reported that the NR3C1 major allele exists in higher frequency in patients with chronic fatigue syndrome . Rossum et al. reported that the NR3C1 Bcl1 variant is associated with major depression .
No association was found between adult/juvenile rheumatoid arthritis (RA) and the NR3C1 Bcl1 variant in studies conducted on the basis of thought that inappropriately low endogenous cortisol production and impaired HPA axis response involved in pathogenesis of RA [10,26,27]. Holliday et al. investigated whether genetic variations that play a role in HPA axis affect the predisposition to musculoskeletal pain, however, they did not find any correlation between the NR3C1 variants and musculoskeletal pain . The NR3C1 gene was one of the genes that was evaluated in OPPERA (Orofacial Pain: Prospective Evaluation and Risk Assessment) trial, in which 358 genes were investigated to illuminate the basis of TMD in recent years .
In present study, Bcl1 variant of NR3C1 was genotyped in a group of patients with TMD to analyze the correlation between the genotypes and the clinico-pathologic features of the patients. The current data suggested that this variant do not affect the development and clinical course of TMD in Turkish population (Table 3). However, this present study found that numeric pain scale was significantly higher in patients with NR3C1 Bcl1 CC and CG genotype (Table 5).
Although current data did not support that the Bcl1 variant of the NR3C1 gene is associated with TMD susceptibility, these findings serve to advise that the Bcl1 variant of the NR3C1 gene may affect pain intensity in TMD patients. However, further studies with a larger sample size are needed to verify these findings.
Conflict of Interest
No potential conflict of interest was reported by the authors.
This work (15-KAEK-124) was approved by the Local Ethical Committee.
- Slade GD, Diatchenko L, Bhalang K, Sigurdsson A, Fillingim RB. Influence of psychological factors on risk of temporomandibular disorders. J Dent Res 2007; 86: 1120-1125.
- Smith SB, Mir E, Bair E, Slade GD, Dubner R, Fillingim RB. Genetic variants associated with development of TMD and its intermediate phenotypes: the genetic architecture of TMD in the OPPERA prospective cohort study. J Pain 2013; 14: T91-101.
- Lee DG, Kim TW, Kang SC, Kim ST. Estrogen receptor gene polymorphism and craniofacial morphology in female TMJ osteoarthritis patients. Int J Oral Maxillofac Surg 2006; 35: 165-169.
- Van Rossum EF, Roks PH, De Jong FH, Brinkmann AO, Pols HA, Koper JW. Characterization of a promoter polymorphism in the glucocorticoid receptor gene and its relationship to three other polymorphisms. Clin Endocrinol 2004; 61: 573-581.
- Ash GI, Kostek MA, Lee H, Angelopoulos TJ, Clarkson PM, Gordon PM. Glucocorticoid Receptor (NR3C1) Variants Associate with the Muscle Strength and Size Response to Resistance Training. PLoS One 2016; 11: e0148112.
- Schindler R, Mancilla J, Endres S, Ghorbani R, Clark SC. Correlations and interactions in the production of interleukin-6 (IL-6), IL-1, and tumor necrosis factor (TNF) in human blood mononuclear cells: IL-6 suppresses IL-1 and TNF. Blood 1990; 75: 40-47.
- Yang D, Ye L. Temporomandibular disorders and declarative memory. Med Hypotheses 2011; 76: 723-725.
- Srivastava N, Prakash J, Lakhan R, Agarwal CG, Pant DC, Mittal B. Influence of Bcl-1 Gene Polymorphism of Glucocorticoid Receptor Gene (NR3C1, rs41423247) on Blood Pressure, Glucose in Northern Indians. Ind J Clin Biochem 2011; 26: 125-30.
- Schiffman E, Ohrbach R, Truelove E, Look J, Anderson G, Goulet JP. Diagnostic Criteria for Temporomandibular Disorders (DC/TMD) for Clinical and Research Applications: recommendations of the International RDC/TMD Consortium Network and Orofacial Pain Special Interest Group. J Oral Facial Pain Headache 2014; 28: 6-27.
- Kostik MM, Klyushina AA, Moskalenko MV, Scheplyagina LA, Larionova VI. Glucocorticoid receptor gene polymorphism and juvenile idiopathic arthritis. Pediatr Rheumatol Online J 2011; 9: 2.
- Charmandari E, Tsigos C, Chrousos G. Endocrinology of the stress response. Annu Rev Physiol 2005; 67: 259-284.
- Boyle B, Korányi K, Patocs A, Liko I, Szappanos A. Polymorphisms of the glucocorticoid receptor gene in graves ophthalmopathy. Br J Ophthalmol 2008; 92: 131-134.
- Lovén J, Svitacheva N, Jerre A, Miller-Larsson A, Korn SH. Anti-inflammatory activity of beta2-agonists in primary lung epithelial cells is independent of glucocorticoid receptor. Eur Respir J 2007; 30: 848-856.
- Corvol H, Nathan N, Charlier C, Chadelat K, Le Rouzic P, Tabary O. Glucocorticoid receptor gene polymorphisms associated with progression of lung disease in young patients with cystic fibrosis. Respir Res 2007; 8: 88.
- De Iudicibus S, Stocco G, Martelossi S, Drigo I, Norbedo S, Lionetti P. Association of BclI polymorphism of the glucocorticoid receptor gene locus with response to glucocorticoids in inflammatory bowel disease. Gut 2007; 56: 1319-1320.
- Koetz KR, Van Rossum EF, Ventz M, Diederich S, Quinkler M. BclI polymorphism of the glucocorticoid receptor gene is associated with increased bone resorption in patients on glucocorticoid replacement therapy. Clin Endocrinol (Oxf) 2013; 78: 831-837.
- DeRijk R, de Kloet ER. Corticosteroid receptor genetic polymorphisms and stress responsivity. Endocrine 2005; 28: 263-270.
- Watt GC, Harrap SB, Foy CJ, Holton DW, Edwards HV. Abnormalities of glucocorticoid metabolism and the renin-angiotensin system: a four-corner approach to the identification of genetic determinants of blood pressure. J Hypertens 1992; 10: 473-482.
- Jason LA, Richman JA, Rademaker AW, Jordan KM, Plioplys AV. A community-based study of chronic fatigue syndrome. Arch Intern Med 1999; 159: 2129-2137.
- Quartana PJ, Buenaver LF, Edwards RR, Klick B, Haythornthwaite JA, Smith MT. Pain catastrophizing and salivary cortisol responses to laboratory pain testing in temporomandibular disorder and healthy participants. J Pain 2010; 11: 186-194.
- Bonifazi M, Suman AL, Cambiaggi C, Felici A, Grasso G, Lodi L. Changes in salivary cortisol and corticosteroid receptor-alpha mRNA expression following a 3-week multidisciplinary treatment program in patients with fibromyalgia. Psychoneuroendocrinology 2006; 31: 1076-1086.
- Dinan TG, Quigley EM, Ahmed SM, Scully P, O'Brien S, O'Mahony L. Hypothalamic-pituitary-gut axis dysregulation in irritable bowel syndrome: plasma cytokines as a potential biomarker? Gastroenterology 2006; 130: 304-311.
- Ojima K, Watanabe N, Narita N, Narita M. Temporomandibular disorder is associated with a serotonin transporter gene polymorphism in the Japanese population. Biopsychosoc Med 2007; 1: 3.
- Rajeevan MS, Smith AK, Dimulescu I, Unger ER, Vernon SD, Heim C. Glucocorticoid receptor polymorphisms and haplotypes associated with chronic fatigue syndrome. Genes Brain Behavior 2007; 6: 167-176.
- Van Rossum EF, Binder EB, Majer M, Koper JW, Ising M, Modell S. Polymorphisms of the glucocorticoid receptor gene and major depression. Biol Psychiatry 2006; 59: 681-688.
- Donn R, Payne D, Ray D. Glucocorticoid receptor gene polymorphisms and susceptibility to rheumatoid arthritis. Clin Endocrinol (Oxf) 2007; 67: 342-345.
- Aydeniz A, Sever T, Pehlivan S, Pehlivan M, Altindag O, Budeyri S. Investigation of Glucocorticoid Receptor Gene Bcl-1 Polymorphism in Rheumatoid Arthritis. Turk J Rheumatol 2011; 26: 199-203.
- Holliday KL, Nicholl BI, Macfarlane GJ, Thomson W, Davies KA. Genetic variation in the hypothalamic-pituitary-adrenal stress axis influences susceptibility to musculoskeletal pain: results from the EPIFUND study. Ann Rheum Dis 2010; 69: 556-560.
- Smith SB1, Maixner DW, Greenspan JD, Dubner R, Fillingim RB. Potential genetic risk factors for chronic TMD: genetic associations from the OPPERA case control study. J Pain 2011; 12: T92-101.