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vol. 15
Letter to the Editor

Association of polymorphisms of the TNFRSF11B and TNFSF11 genes with bone mineral density in postmenopausal women from western Mexico

Anahi González-Mercado
Josefina Y. Sánchez-López
Francisco J. Perea-Díaz
Maria T. Magaña-Torres
Mario Salazar-Páramo
Laura González-López
Mirna Gisel González-Mercado
Bertha Ibarra-Cortés

Arch Med Sci 2019; 15 (5): 1352–1356
Online publish date: 2019/08/22
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Osteoporosis (OP) is a disease with reduced bone mass and deterioration of the spatial distribution of the trabecular bone structure, leading in consequence to increased bone fragility and risk of fractures [1]. In Mexico its frequency in postmenopausal women is 8.3% to 31% [2]. OP is a multifactorial disease because its development involves physiological, environmental and genetic factors [3]. Genes of the tumor necrosis factor (TNF) family like TNFRSF11B (TNF receptor superfamily member 11b) and TNFSF11 (TNF superfamily member 11) encode for osteoprotegerin (OPG) and receptor activator for nuclear factor B ligand (RANKL) proteins, respectively, that participate in the signaling pathway OPG/RANK/RANKL, balancing the activity between osteoblasts and osteoclasts to prevent bone loss and to ensure normal bone turnover [4, 5]. Different single nucleotide polymorphisms (SNPs) in genes of this pathway are related to bone mineral density (BMD) or OP [6, 7]. In Mexican postmenopausal women, SNPs of the TNFRSF11B gene were studied but no association with BMD [8, 9] or OP [10] was observed; SNPs of the TNFSF11 gene have not been studied for OP.
The aim of this study was to investigate the association of the polymorphisms 1181 G>C (rs2073618) and 1217 C>T (rs3102734) of TNFRSF11B; as well as -708 T>A, -693 C>G (rs9533155), -657 C>A, -643 C>T (rs9533156) and -290 C>T (rs9525641) of TNFSF11 with BMD.
Five hundred and thirteen postmenopausal Mexican-Mestizo women from western Mexico agreed to participate in the study. They were recruited from Hospital General Regional No. 110 of the Instituto Mexicano del Seguro Social in Guadalajara city. BMD of the lumbar spinal L1-L4 region and of the femoral neck (left and right) was determined by means of DEXA (dual-energy X-ray absorptiometry, Prodigy Advance, GE). A hundred and seventy-four women were matched by age (range was between 42 to 69 years) and body mass index (BMI) (range from 19.7 to 42.1 kg/m2) in two groups by the WHO criteria: women with values of the T-score less than –2.5 SD (OP group, n = 87) and those with values of the T-score above -1 SD (non-OP group, n = 87). A questionnaire that included different biological (age, BMI, age at onset of menarche and menopause, family history of OP, personal and family history of fractures) and lifestyle (smoking, alcoholism, coffee consumption and sedentarism) variables was applied to all participants. This study was approved by the local ethical committee; informed consent was obtained from all women.
Identification of polymorphisms was performed by PCR followed by Sanger DNA sequencing with the BigDye terminator kit v3.1 (Applied Biosystems Foster City). To identify the rs2073618 and rs3102734 variants, a fragment of 274 bp was amplified using the primers forward 5´-GATCAAAGGCAGGCGATACTTCC-3´ and reverse 5´-CTGGGAGGGAGCGAGTGGAGC-3´. The PCR mixture (25 l) contained: genomic DNA (200 ng), 10 pmol each primer, Taq polymerase 1 U, MgCl2 2.0 mM, dNTPs 250 mol and buffer 1X. The thermocycling conditions were: initial denaturation 95°C/5 min, denaturation 95°C/1 min, annealing 57°C/1 min, extension 72°C/1:30 min, 35 cycles and a final extension 72°C/5 min. The -708 T>A, rs9533155, -657 C>A, rs9533156 and rs9525641 polymorphisms were identified according to Mencej et al. [11].
For categorical variables, the frequencies or percentages were obtained; for continuous variables, the average and standard deviation were calculated. Chi square and Fisher exact tests were used for comparison of genotypic and allelic frequencies of each polymorphism between both groups. The risk for OP was estimated through odds ratio, considering classic models of inheritance. Haplotypes were established inferring phase of family segregation. Linkage disequilibrium (LD) was significant when r2  0.33, using Arlequin v3.01 software. ANOVA and Student t-tests were used to relate quantitative variables with genotypes and alleles, respectively. Other tests used were Spearman and Pearson correlations and linear regression models. Statistical analyses were carried out using SPSS version 20.0 and p < 0.05 was significant.
As expected, BMD values in the group of postmenopausal women with OP (0.834 ±0.052 g/cm2 in lumbar spine L1–L4; 0.788 ±0.086 g/cm2 in left femoral neck; 0.787 ±0.085 g/cm2 in right femoral neck) were lower than in the non-OP group (1.173 ±0.109 g/cm2 in lumbar spine L1-L4; 1.005 ±0.087 g/cm2 in left femoral neck; 1.003 ±0.083 g/cm2 in right femoral neck) (p < 0.001). The other biological and lifestyle variables did not show significant differences between the groups, even when they have been reported as high risk for the development of OP in some populations [12, 13].
The distribution of genotypic and allelic frequencies of the seven SNPs was similar between the groups (p > 0.05). The SNPs –708 T>A and –657 C>A were observed with low frequencies in both groups, and the –708A and –657A alleles were not detected in homozygote state in the study (Table I). The most widely studied polymorphism of TNFRSF11B is 1181 G>C; the distribution of allelic frequencies is variable in women with OP from several populations [14–16]. Under classical models of inheritance (dominant, recessive, codominant or additive) neither genotypes nor alleles were associated with risk for OP (p > 0.05). Regarding TNFSF11, in the Egyptian population, the –290T allele was associated with increased risk of developing OP (OR = 1.7, IC: 1.1–2.7, p = 0.019) [17].
Haplotypes for TNFRSF11B were constructed with two polymorphisms (1181 G>C and 1217 C>T) and three haplotypes were observed; the most frequent was CC (OP 54.6% and non-OP 54.0%). In TNFSF11, the haplotypes were constructed with five SNPs: –708 T>A, –693 C>G, –657 C>A, –643 C>T and –290 C>T, and 13 haplotypes were observed, the most frequent in both groups being TCCTT (OP = 49.4% and non-OP = 54.1%). The distribution of haplotype frequencies was compared between the two groups and no statistically significant differences were found (p > 0.05), with the exception of ACATT haplotype (OP group = 9.8%, non-OP group = 4.0%; p = 0.03). The same combinations of SNPs for the two genes are not reported in the literature.
The SNPs of TNFRSF11B were in linkage equilibrium in the two groups (OP: r2 = 0.089; non-OP: r2 = 0.127). For the five sites of TNFSF11, we found that four pairs of loci are in LD in both groups, since r2 values ranged from 0.6587 to 0.9058 with p < 0.001: 1) –708 T>A/-657 C>A, 2) –693 C>G/–643 C>T, 3) –693 C>G/-290 C>T and 4) –643 C>T/–290 C>T. In women with OP and non-OP from Slovenia, LD was also observed between pairs 2, 3 and 4 [18, 19].
The relationship between genotypes and alleles of the studied polymorphisms with the biological variables in each group was analyzed; only the BMD and BMI presented statistically significant results with some SNPs (Table II). For BMD, no association was found with 1181 G>C and 1217 C>T SNPs of TNFRSF11B; similar results were observed in postmenopausal women from Australia [20], Ireland [14, 21], and Malta [22]; however, the 1181GG genotype was associated with lower BMD in Chinese [7, 16], Slovenian [15, 23], and Koreans [24]; in China, the 1217T allele was related to increased risk of osteoporotic fracture (OR = 1.35; 95% CI: 1.17–1.55; Bonferroni p = 2.6 × 10–4) [25]. Regarding BMI, in the present study, the 1181GG genotype was significantly associated with lower BMI in women with OP (Table II). Although this relationship has not been reported, we can suggest that this polymorphism could have an indirect effect on the variation of the BMD, as was demonstrated by Mendez et al. [26], in which a high BMI is positively correlated with higher BMD. It has been proposed that obesity protects against bone loss through mechanical load, and it has been suggested that the adipose tissue is involved in the homeostasis of the skeleton, possibly through the role of some adipokines in bone remodeling. Also, adipocytes secrete estrogens that help to regulate homeostasis and contribute to the increase of bone mass [26].
On the other hand, the –693CC, –643TT and –290CT genotypes of TNFSF11 reveal lower BMD in the left and/or right femoral neck than the other genotypes in postmenopausal women with OP, which demonstrates the important role of these SNPs in the variation of BMD (Table II). The linear regression analysis with all studied polymorphisms and BMD of the lumbar spine, and the right and left femoral necks, showed only a significant model between the –693G allele and BMD of the right femoral neck (constant = 0.738,  = 0.028, p = 0.032), indicating that there is a positive relationship since the BMD is increased slightly. These three polymorphisms in Slovenian women with OP were associated with BMD of the lumbar spine and –693 C>G also with BMD of the femoral neck [18]. In the Chinese population –693 C>G or –643 C>T polymorphism was not related to BMD in peri- and postmenopausal women [7]. The –643TT and –290TT genotypes of TNFSF11 in the non-OP group were related to lower BMI, which could suggest a possible influence on the BMD variation as explained above for 1181 G>C [26].
In conclusion, the 1181 G>C and 1217 C>T polymorphisms of TNFRSF11B and –708 T>A, –693 C>G, –657 C>A, –643 C>T and –290 C>T of TNFSF11 were not associated with OP. However, –693CC, –643TT and –290CT genotypes have an effect of lowering BMD in the left and/or right femoral neck. The 1181GG genotype was significantly associated with lower BMI in women with OP as well as –643TT and –290TT genotypes of TNFSF11 in the non-OP group. The –708 T>A and –657 C>A polymorphisms have not been described in the literature.


The work was conducted in División de Genética, CIBO, IMSS, Guadalajara, Jalisco.
The research was supported by a research grant from Consejo Nacional de Ciencia y Tecnología #2003-107.

Conflict of interest

The authors declare no conflict of interest.


1. Bączyk G, Samborski W, Jaracz K. Evaluation of the quality of life of postmenopausal osteoporotic and osteopenic women with or without fractures. Arch Med Sci 2016; 12: 819-27.
2. González-Mercado A, Sánchez-López JY, Ibarra B. Factores de riesgo para osteoporosis en mujeres posmenopáusicas de Guadalajara, Jalisco. Salud Publica Mex 2013; 55: 627-30.
3. McCormick RK. Osteoporosis: integrating biomarkers and other diagnostic correlates into the management of bone fragility. Altern Med Rev 2007; 12: 113-45.
4. Kearns AE, Khosla S, Kostenuik PJ. Receptor activator of nuclear factor kappa b ligand and osteoprotegerin regulation of bone remodeling in health and disease. Endocr Rev 2008; 29: 155-92.
5. Trouvin AP, Goëb V. Receptor activator of nuclear factor-kappaB ligand and osteoprotegerin: maintaining the balance to prevent bone loss. Clin Interv Aging 2010; 5: 345-54.
6. Tu P, Duan P, Zhang RS, et al. Polymorphisms in genes in the RANKL/RANK/OPG pathway are associated with bone mineral density at different skeletal sites in post-menopausal women. Osteoporos Int 2015; 26: 179-85.
7. Shang M, Lin L, Cui H. Association of genetic polymorphisms of RANK, RANKL and OPG with bone mineral density in Chinese peri- and postmenopausal women. Clin Biochem 2013; 46: 1493-501.
8. Canto-Cetina T, Polanco Reyes L, González Herrera L, et al. Polymorphism of LRP5, but not of TNFRSF11B, is associated with a decrease in bone mineral density in postmenopausal Maya-Mestizo women. Am J Hum Biol 2013; 25: 713-8.
9. Rojano-Mejía D, Coral-Vázquez RM, Espinosa LC, et al. TNFRSF11B gene haplotype and its association with bone mineral density variations in postmenopausal Mexican-Mestizo women. Maturitas 2012; 71: 49-54.
10. Zavala-Cerna MG, Moran-Moguel MC, Cornejo-Toledo JA, et al. Osteoprotegerin polymorphisms in a mexican population with rheumatoid arthritis and generalized osteoporosis: a preliminary report. J Immunol Res 2015; 2015: 376197.
11. Mencej S, Prezelj J, Kocijancic A, Ostanek B, Marc J. Association of TNFSF11 gene promoter polymorphisms with bone mineral density in postmenopausal women. Maturitas 2006; 55: 219-26.
12. Rossini M, Adami S, Bertoldo F, et al. Guidelines for the diagnosis, prevention and management of osteoporosis. Reumatismo 2016; 68: 1-39.
13. Gronholz MJ. Prevention, diagnosis, and management of osteoporosis-related fracture: a multifactoral osteopathic approach. J Am Osteopath Assoc 2008; 108: 575-85.
14. Langdahl BL, Carstens M, Stenkjaer L, Eriksen EF. Polymorphisms in the osteoprotegerin gene are associated with osteoporotic fractures. J Bone Miner Res 2002; 17: 1245-55.
15. Mencej-Bedrač S, Preželj J, Marc J. TNFRSF11B gene polymorphisms 1181G > C and 245T > G as well as haplotype CT influence bone mineral density in postmenopausal women. Maturitas 2011; 69: 263-7.
16. Zhao HY, Liu JM, Ning G, et al. The influence of Lys3Asn polymorphism in the osteoprotegerin gene on bone mineral density in Chinese postmenopausal women. Osteoporos Int 2005; 16: 1519-24.
17. Mohamed RH, Mohamed RH, El-Shahawy EE. Relationship between RANK and RANKL gene polymorphisms with osteoporosis in rheumatoid arthritis patients. Genet Test Mol Biomarkers 2016; 20: 249-54.
18. Mencej S, Albagha OME, Preželj J, Kocjan T, Marc J. Tumour necrosis factor superfamily member 11 gene promoter polymorphisms modulate promoter activity and influence bone mineral density in postmenopausal women with osteoporosis. J Mol Endocrinol 2008; 40: 273-9.
19. Mencej-Bedrač S, Preželj J, Kocjan T, et al. The combinations of polymorphisms in vitamin D receptor, osteoprotegerin and tumour necrosis factor superfamily member 11 genes are associated with bone mineral density. J Mol Endocrinol 2009; 42: 239-47.
20. Ueland T, Bollerslev J, Wilson SG, et al. No associations between OPG gene polymorphisms or serum levels and measures of osteoporosis in elderly Australian women. Bone 2007; 40: 175-81.
21. Wynne F, Drummond F, O’Sullivan K, et al. Investigation of the genetic influence of the OPG, VDR (Fok1), and COLIA1 Sp1 polymorphisms on BMD in the Irish population. Calcif Tissue Int 2002; 71: 26-35.
22. Vidal C, Brincat M, Xuereb Anastasi A. TNFRSF11B gene variants and bone mineral density in postmenopausal women in Malta. Maturitas 2006; 53: 386-95.
23. Arko B, Preželj J, Kocijancic A, Komel R, Marc J. Association of the osteoprotegerin gene polymorphisms with bone mineral density in postmenopausal women. Maturitas 2005; 51: 270-9.
24. Choi JY, Shin A, Park SK, et al. Genetic polymorphisms of OPG, RANK, and ESR1 and bone mineral density in Korean postmenopausal women. Calcif Tissue Int 2005; 77: 152-9.
25. Wang C, Zhang Z, Zhang H, et al. Susceptibility genes for osteoporotic fracture in postmenopausal Chinese women. J Bone Miner Res 2012; 27: 2582-91.
26. Méndez JP, Rojano-Mejía D, Pedraza J, et al. Bone mineral density in postmenopausal Mexican-Mestizo women with normal body mass index, overweight, or obesity. Menopause 2013; 20: 568-72.
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