Indian Journal of Dental Research

ORIGINAL RESEARCH
Year
: 2020  |  Volume : 31  |  Issue : 2  |  Page : 252--256

Estimation of salivary calcium level as a screening tool for the osteoporosis in the post-menopausal women: A prospective study


Amit Wasti1, Jyoti Wasti2, Ritunja Singh3,  
1 Department of Oral Pathology, Government Dental College and Hospital, Raipur, Chhattishagrh, India
2 Department of Periodontology, Government Dental College and Hospital, Raipur, Chhattishagrh, India
3 Department of Periodontology, New Horizon Dental College, Bilaspur, Chhattishagrh, India

Correspondence Address:
Dr. Ritunja Singh
New Horizon Dental College and Research Institute, Bilaspur, Chhattisgarh - 495001
India

Abstract

Aims and Objectives: The aim and objective of the present study was to explore the use of salivary calcium levels as a diagnostic biochemical marker for osteoporosis in menopausal women and also to find the correlation among oestrogen level, bone density and salivary calcium level. Materials and Methods: The study included 180 individuals and they were divided into three groups with 60 individuals in each group, comprised of healthy women, pregnant women and post-menopausal women. All the women were asked to collect at least 2 ml of unstimulated whole saliva in the sterile plastic sample containers. The samples were immediately subjected to biochemical estimation of calcium. Similarly, estimation was done for oestrogen level and bone density among all the groups. The results were obtained by one-way analysis of variance (ANOVA) using Statistical Software SPSS version 17. For the correlation among the bone density, salivary calcium level and serum oestrogen levels, Pearson's correlation was used. Results: The mean salivary calcium level in the healthy women group was found to be 3.0 ± 0.50 μg/ml. Similarly, pregnant women and post-menopausal group, it was found to be 3.20 ± 0.72 and 7.5 ± 0.90 μg/ml, respectively. When the intergroup comparison was done in the three groups, it was found to be highly significant (P = 0.001). Similarly, the difference in mean value for oestrogen level and bone density was highly significant among all the groups (P = 0.001). Conclusion: Among all the three groups, the salivary calcium levels do exhibit the correlation with bone mineral density. In the post-menopausal group, there was significant increase in salivary calcium level compared to other groups. Similarly, the study showed a negative correlation between salivary calcium and serum oestrogen. This substantiates the point that salivary calcium levels can definitely indicate the possibility of the presence or absence of osteoporosis in post-menopausal women.



How to cite this article:
Wasti A, Wasti J, Singh R. Estimation of salivary calcium level as a screening tool for the osteoporosis in the post-menopausal women: A prospective study.Indian J Dent Res 2020;31:252-256


How to cite this URL:
Wasti A, Wasti J, Singh R. Estimation of salivary calcium level as a screening tool for the osteoporosis in the post-menopausal women: A prospective study. Indian J Dent Res [serial online] 2020 [cited 2020 Jul 5 ];31:252-256
Available from: http://www.ijdr.in/text.asp?2020/31/2/252/284589


Full Text



 Introduction



Bone is a living tissue that is constantly being broken down and rebuilt. When the balance between breakdown and rebuilding is disturbed, for example, by hormonal changes or dietary changes, the bone may lose some of the minerals that contribute to its density and strength.[1] A condition of diminished bone density is called osteopenia. When a significant loss in bone density occurs, such that the bone is markedly weakened and susceptible to fracture, the condition is termed osteoporosis (porous bone). Most of the people at risk for osteoporosis are not aware of it. It is called a “silent disease” because there are usually no symptoms until a person has a bone fracture. This breakage, frequently in the hip, the vertebrae of the spine, or in the wrist, can occur with very little pressure and can cause the person significant pain and protracted or permanent disability.[2],[3]

In most women, bone mass reaches its peak in the third decade of life and declines thereafter. This decline in bone mass is accelerated with the onset of menopause and might lead to excessive bone resorption and ultimately, osteoporosis.[4] Osteoporosis affects over 20 million women and leads to approximately 1.5 million fractures each year, making it one of the leading public health problems. The most important risk factor for bone loss in midlife women is the menopause. Women lose about 50% of their trabecular bone and 30% of their cortical bone during the course of their lifetime, about half of which is lost during the first 10 years after the menopause.[5] Menopausal and post-menopausal women may experience an increased rate of bone mass loss with a decrease in the hormone oestrogen.[5] Due to the depletion of primordial follicles, ovarian oestradiol secretion ceases at menopause and is followed by very low levels of circulating oestradiol generating the typical menopausal symptoms.

Going through menopause early can exacerbate the loss. According to the NOF, some women can lose up to 20% of their bone mass in the first 5 to 7 years following menopause.[6]

There are various theories proposed for the inductions of the high rate of bone turn over after the period of menopause. The most accepted is the direct action of oestradiol on the activation of bone-resorbing cells that is osteoclast.[7] Among the various methods used for the determination of osteoporosis, DEXA that is dual-energy X-ray absorptiometry is considered as the gold standard.[8] However, all these methods involve a high cost to benefit ratio coupled with exposure to radiation. Hence, a diagnostic tool that eliminates both these factors or at least considerably reduces them is desirable.

Calcium is an important skeletal mineral. Several studies indicate that salivary calcium and phosphate concentrations show a clear increase with increasing age. Calcium is the only electrolyte which does not show correlation with salivary flow rate.[9],[10],[11]

In recent years, saliva-based diagnostic tests have increased in popularity because of their non-invasive nature. Salivary biomarkers have been used to assess the risk of: developing oral, ovarian and breast cancers; HIV infection; dental caries; and periodontal diseases. Using saliva rather than serum has benefits: it is non-invasive, easy to obtain, painless and there is no need to employ specially trained personal for sample collection.[9],[10],[11]

Calcium and phosphorous are present as the main inorganic components of saliva, which quantitatively account for the main mineral components of the human skeletal system.[10] Hence, the present study was planned to explore the use of salivary calcium levels as a diagnostic biochemical marker for osteoporosis.

 Materials and Methods



The present study was conducted among all the patients visiting in the outpatient department of the medical college from April 2019 to June 2019. Individuals were randomly selected from outpatient department comprising a total of 180 individuals (convenience sample). The included individuals were divided into three groups with 60 individuals in each group, comprised of healthy women, pregnant women and post-menopausal women. All the women were informed about the procedure of the study and the written informed consent was obtained from each of the participants. The institutional ethical committee were informed about the study and the ethical clearance certificate was obtained from them. We have included healthy woman, pregnant woman, and post-menopausal woman as our three different groups so that we can also compare the salivary calcium, oestrogen level and bone density in all the three phases of woman's life.

Group's criteria

Group A comprised of healthy women. All the women with the age ranging from 20 years to 35 years with the regular menstrual cycle and having the body mass index <24 were included in the study. Women with the presence of any systemic diseases, salivary gland diseases, and patient taking topical steroids, drug intake (e.g. hormonal replacement therapy [HRT] and calcium supplements) and usage of tobacco were excluded from the study.

Group B comprised of pregnant women. All the women with the age ranging from 20 years to 35 years with their first pregnancy were included in the study. Women with the history of oral contraceptives (as they may affect salivary calcium level),[12] drug intake (e.g. hormonal replacement therapy [HRT] and calcium supplements), salivary gland diseases and patient taking topical steroids and tobacco were excluded from the study, as they may affect the salivary content of individuals in the present study.

Group C comprised of post-menopausal women. All the women with the age ranging from 45 years to 56 years with the onset of menopause at least 1 year prior and BMI less than 30 were included in the study. Women with the presence of any systemic illness, drug intake and intake of calcium supplement were excluded from the study. Patients on drugs that alter salivary flow such as diuretics and anti-cholinergics were also excluded from the study.

Body mass index

BMI was calculated using the equation as below[13]

BMI = (kg)/m2

Where kg is weight in kilograms and m is height in meters.

Serum Oestrogen assessment

For the estimation of serum oestrogen, 5 ml of venous blood was drawn from the cubital vein and was transferred to the laboratory. Each of the samples was then centrifuged for about 10 min at 3000 rpm. The serum obtained was then subjected to quantitative estimation of oestrogen.

Salivary calcium level estimation

Salivary sample collection: Prior to the sample collection, all the participants were asked not to eat, drink or perform any other oral hygiene procedures so that morning salivary samples can be collected. Participants were asked to rinse their mouth with distal drinking water. All the women were asked to collect at least 2 ml of unstimulated whole saliva in the sterile plastic sample containers. The samples were immediately subjected to biochemical estimation of calcium.

The kit used for biochemical estimation of the calcium was the Erba Mannheim, Trans Asia Biochemical's Ltd. The basic principle behind this test is of O-cresolphthalein complexone method. The purple colour complex is formed by the binding of calcium in alkaline medium with OCPC. The amount of the intensity of the purple colour complex depends on the amount of calcium present in saliva.

The bone mineral density (BMD) was measured with the help of ultrasound (Paltech System CM200). All the women were asked to sit in a chair and place their foot on the machine. Later on, readings were recorded. T scores are used for the categorisation of bone mineral density. For T score of 1, it was considered as normal, for T score of -1 to -2.5, it was considered as Osteopenic and for T score less than -2.5, it was considered as Osteoporotic.

Statistical analysis

The results obtained were tabulated and subjected to statistical analysis by one-way analysis of variance (ANOVA) using Statistical Software SPSS version 17. The data were expressed as mean and standard deviation. P values were considered to be statistically significant (P < 0.0005). For the correlation among the alveolar bone's mass, the salivary calcium level and serum oestrogen levels, Pearson's correlation was used.

 Results



The mean serum oestrogen level was found to be 116.67 ± 79 pg/ml in the healthy women, in pregnant women, it was found to be 6500.25 ± 80 pg/ml and it was found to be 49.15 ± 70 pg/ml in the post-menopausal groups. When the intergroup comparison was done for the means of serum oestrogen among all the three groups, there was a highly significant difference with the P value = 0.001 [Table 1].{Table 1}

The mean age of women in the healthy group was 22 years, whereas it was 24 years in the pregnant group. The mean age was 58 years in the last post-menopausal group [Table 2].{Table 2}

The mean salivary calcium level in the healthy women group was found to be 3.0 ± 0.50 μg/ml. In the pregnant women and post-menopausal group, it was found to be 3.20 ± 0.72 and 7.5 ± 0.90 μg/ml, respectively. When the intergroup comparison was done among the three groups, it was found to be highly significant with the P value = 0.001 [Table 3].{Table 3}

The mean bone mineral density as expressed as T score was found to be 0.15 ± 1.00 for healthy groups, whereas it was found to be -0.59 ± 0.60 and -2.45 ± 0.63, respectively, in the pregnant and post-menopausal women. When the intergroup comparison was done among all the groups, it was found to be highly significant with the P value = 0.001 [Table 4]. A highly significant negative correlation was found between the salivary calcium levels and bone mineral density and also between the salivary calcium level and serum oestrogen levels among all the three groups [Table 5].{Table 4}{Table 5}

 Discussion



Menopause is defined as the point in time when menstrual cycles permanently cease due to the natural depletion of ovarian oocytes from aging. The diagnosis is typically made retrospectively after the woman has missed menses for 12 consecutive months. The average age was found to be 55 years.[14] There is great fluctuation in the women hormones throughout their lives. This hormones' fluctuation affects their entire body including their oral cavity.[15] Thus, menopauses directly affect the metabolism of the human body that leads to change in the parameter of saliva and affecting the quality of life.

Osteoporosis affects a large proportion of world over the age of 50. The third National Health and Nutrition Examination Survey estimated that 12 million white women over age 50 met the WHO criterion for osteopenia and 5 million for osteoporosis.[16] There are low bone mass and change in the micro-architectural deterioration in the presence of osteoporosis. This can eventually lead to pathological fractures. There is lot of influence of osteoporosis in the quality of life as also the dental health in particular.[17],[18],[19]

In the present study, 180 females were subdivided into three groups. The study participants in the pregnant group comprised only those in their first pregnancy and those in the post-menopausal group comprised participants who had attained menopause within a minimum of 1 year prior to the conduct of the study. This was done so as to minimise the effect of calcium deficiency which was time-dependent and strongly correlate to the functional levels of oestrogen in both these groups when compared to controls.

The mean oestrogen level was found to be highest in the pregnant women group and lowest in the post-menopausal women group. These differences in the levels in the oestrogen level were found to be due to various physiological changes in the various stages of the life of women. During pregnancy, the placenta secretes oestrogen with the levels increased by about 30 times the normal towards parturition. Menopause is a burn out of the ovaries resulting in a cessation of oestrogen synthesis resulting in a decrease below the critical levels and nearing almost to zero. Oestrogen is produced in subcritical levels but only for a short time.

Oestrogen is believed to directly affect the physiologic absorption of calcium from the gut. A drastic fall or deficiency in estrogen levels as seen in post-menopausal phase triggers a release of parathormone which in turn actively induces calcium resorption from the skeletal system increasing the serum levels of calcium which in turn reflects in the raised levels of calcium in the saliva. The result of the present study clearly revealed a highly significant increase in the mean salivary calcium of post-menopausal women group compared to healthy and pregnant women. Nagler and Hershkovich have also observed that salivary calcium levels are significantly higher in the elderly age group compared to the young.[20]

There was a highly significant negative correlation (P = 0.0000) between the BMD score and salivary calcium levels indicative of the fact that a reduction in the mean BMD scores is associated with a concurrent increase in the salivary calcium levels. It was also found that there was also a highly significant negative correlation (P = 0.0000) between serum oestrogen levels and salivary calcium. This establishes the scientific link among the three entities, that is, serum oestrogen, BMD and salivary calcium.

The basal metabolic index was not taken into consideration with pregnant woman due to added weight of embryo that may affect the BMI of individual leading to false data. Similarly, menopausal woman were placed at higher BMI in the recent study so that the lowered cutoffs fails to capture most post menopausal women whose actual body fat percentage would classify them as obese, let alone those whose body fat percentage would classify them as overweight. To improve the sensitivity of BMI in identifying post-menopausal women at the risk of obesity-related diseases, the authors of the recent study concluded that the obesity cutoff might need to be set as low as 24.9, which is currently the top of the normal BMI range for the general adult population.[21]

 Conclusion



Among all the three groups, the salivary calcium levels do exhibit the correlation with bone mineral density. In the post-menopausal group, there was a significant increase in salivary calcium level compared to other groups. Similarly, the study showed a negative correlation between salivary calcium and serum oestrogen. This study also showed us a significant negative correlation exists between bone density and calcium level, calcium level in saliva and oestrogen level. This substantiates the point that salivary calcium levels can definitely indicate the possibility of the presence or absence of osteoporosis in post-menopausal women. Limitations: first, our study was done on a small sample size. The same needs to be re-examined and substantiated in a larger sample of population. Second, convenience sampling method was used in our study; a more and detailed study with different sampling method or controlled trials must be done to obtain exact and more valid results. However, they can be extrapolated to arrive at a scientific understanding of the interrelationship that exists between the triad of serum oestrogen, salivary calcium and bone density among healthy, pregnant and post-menopausal women population so that they can be applied as routine chair-side diagnostic procedure for osteoporosis.

Declaration of patient consent

The authors certify that they have obtained all appropriate patient consent forms. In the form, the patients have given their consent for their images and other clinical information to be reported in the journal. The patients understand that their names and initials will not be published and due efforts will be made to conceal their identity, but anonymity cannot be guaranteed.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

References

1Qin QH. Mechanics of Cellular Bone Remodeling: Coupled Thermal, Electrical, and Mechanical Field Effects. Unites states: CRC Press; 2013.
2Hall JC, Riley RE. Nutritional strategies to reduce the risk of osteoporosis. Medsurg Nurs 1999;8:281-93.
3Kawalkar AC. A comprehensive review on osteoporosis. J Trauma 2015;10:3-12.
4Jeffcoat MK, Lewis CE, Reddy MS, Wang CY, Redford M. Post-menopausal bone loss and its relationship to oral bone loss. Periodontology 2000 2000;23:94-102.
5Civitelli R, Pilgram TK, Dotson M, Muckerman J, Lewandowski N, Armamento-Villareal R, et al. Alveolar and postcranial bone density in postmenopausal women receiving hormone/estrogen replacement therapy: A randomized, double-blind, placebo-controlled trial. Arch Intern Med 2002;162:1409-15.
6Frackiewicz EJ, Cutler NR. Women's health care during the perimenopause. J Am Pharm Assoc 2000;40:800-11
7Parfitt AM. Skeletal Heterogeneity and the Purposes of Bone Remodeling: Implications for the Understanding of Osteoporosis. Osteoporosis: Elsevier; 2001. p. 433-47.
8Hans D, Barthe N, Boutroy S, Pothuaud L, Winzenrieth R, Krieg MA. Correlations between trabecular bone score, measured using anteroposterior dual-energy X-ray absorptiometry acquisition, and 3-dimensional parameters of bone microarchitecture: An experimental study on human cadaver vertebrae. J Clin Densitom 2011;14:302-12.
9Vissink AJ, Wolf A, Veerman ECI. “Slaiva Collectors”. In: Wong DT, editor. Salivary Diagnostics. 1st ed. Ames, IO, USA: Wiley-Blackwell; 2008. p. 37-59.
10Navazesh M, Kumar SKS. Measuring salivary flow: Challenges and opportunities. J Am Dent Assoc 2008;139:35S-40S.
11Sevón L, Laine MA, Karjalainen S, Doroguinskaia A, Helenius H, Kiss E, et al. Effect of age on flow-rate, protein and electrolyte composition of stimulated whole saliva in healthy, non-smoking women. Open Dent J 2008;2:89-92.
12Sewón L, Laine M, Karjalainen S, Leimola-Virtanen R, Hiidenkari T, Helenius H. The effect of hormone replacement therapy on salivary calcium concentrations in menopausal women. Arch Oral Biol 2000;45:201-6.
13Lee YH, Wong DT. Saliva: An emerging biofluid for early detection of diseases. Am J Dent 2009;22:241-8.
14Agha-Hosseini F, Mirzai-Dizgah I, Moosavi MS. Relationship of serum and saliva calcium, phosphorus and alkaline phosphatase with dry mouth feeling in menopause. Gerodontology 2012;29:e1092-e7.
15Singh SP, Sikri G, Garg MK. Body mass index and obesity: Tailoring “cut-off” for an Asian Indian male population. Med J Armed Forces India 2008;64:350-3.
16Butler L, Santoro N. The reproductive endocrinology of the menopausal transition. Steroids 2011;76:627-35.
17Prasanna JS, Karunakar P, Sravya MN, Madhavi B, Manasa A. Detrimental consequences of women life cycle on the oral cavity. J Oral Res Rev 2018;10:39-44.
18Looker AC, Melton III LJ, Harris TB, Borrud LG, Shepherd JA. Prevalence and trends in low femur bone density among older US adults: NHANES 2005–2006 compared with NHANES III. J Bone Miner Res 2010;25:64-71.
19Alghamdi HS, Jansen JA. Bone regeneration associated with nontherapeutic and therapeutic surface coatings for dental implants in osteoporosis. Tissue Eng Part B Rev 2012;19:233-53.
20Nagler RM, Hershkovich O. Relationships between age, drugs, oral sensorial complaints and salivary profile. Arch Oral Biol 2005;50:7-16.
21Rubin R. Postmenopausal women with a “Normal” BMI might be overweight or even obese. JAMA 2018;319:1185-7.