Association between dental caries experience and salivary profile among autoimmune thyroid disease subjects - a cross-sectional comparative study

Background: Autoimmune thyroid disease (AITD) is an inflammatory condition that primarily affects women between the ages of 30 and 50. It has been hypothesised that AITD causes salivary glands to produce less saliva due to its endocrine effects. Studies showing the effect of AITD on salivary glands are very scarce. The aim of this study was to compare AITD patients with controls who were of a similar age and gender in terms of salivary metrics and the prevalence of dental caries. Methods: 200 subjects in Bangalore city (100 AITD subjects and 100 healthy subjects as controls) participated in a cross-sectional study. Subjective oral dryness was assessed using Farsi’s criteria. Salivary parameters and caries were assessed using GC Saliva-Check BUFFER kit and WHO caries criteria respectively. Descriptive and analytical statistics were done. Significant data was defined as a p value of <0.05. Results: When compared to controls, the AITD group had substantially more subjective mouth dryness and dental caries. Unstimulated (USFR) and stimulated salivary flow rate (SFR), pH and buffering capacity were reduced in AITD group whereas viscosity was high. There was a significant negative correlation between USFR, SFR, subjective oral dryness and dental caries. In a linear regression, there was no association between age, gender, socioeconomic status (SES), thyroid stimulating hormone (TSH), salivary viscosity, pH, buffering capacity and dental caries. Dental caries, USFR and SFR demonstrated a substantial relationship. Conclusions: The present research may indicate an underlying association between thyroid and salivary gland dysfunction resulting in oral dryness and high dental caries experience.


Introduction
The immune system is dysregulated in autoimmune thyroid disease (AITD), which leads to an attack on the thyroid gland.Its two primary clinical symptoms are Graves disease (GD) and Hashimoto's thyroiditis (HT).An estimated 5% of the general population is thought to be affected by HT and GD.AITD develops when genetically susceptible people lose their tolerance to thyroid antigens in conjunction with environmental factors such altered microbiota.Both symptoms are marked by aberrant thyroid functioning and T and B cell infiltration into the thyroid that is responsive to thyroid antigens. 1TD is unique to thyroid gland and women are affected 4-10 times more frequently than males. 2 The most prevalent forms, Hashimoto's thyroiditis (HT) and Graves' disease (GD), have many immune characteristics in common.The primary characteristic of AITD is the emergence of antibodies against thyroid peroxidase (TPO), thyroglobulin (TG), and thyroid stimulating hormone receptor (TSH-R). 3 A combination of main and minor salivary gland secretions, gingival crevicular fluid, cellular debris, and bacteria are all present in saliva.Saliva serves a wide range of purposes, including the defence of the mouth cavity, digesting owing to salivary amylase, immunity to microbes, and, most critically, for use in diagnostic activities.The use of saliva in diagnostic procedures is expanding since it is simple, non-invasive, and affordable.Enzymes, hormones, antibodies, antibacterial components, growth factors, and other substances that enter from the blood through the gaps between cells can all be found in saliva.Some of the conditions for which saliva has significant diagnostic potential are oncological, endocrine, cardiovascular, rheumatic, autoimmune, neurological, or viral illnesses. 1ntal caries is seen as a complex microbiological illness.It is a multifactorial disease of the teeth that results in a localised loss of tooth structure.It is caused by the interaction of dietary carbohydrates, tooth substrate, and cariogenic bacteria in the dental biofilm, which produces acid after carbohydrate fermentation and causes changes in the pH of the biofilm, which leads to mineral loss (demineralization) as a result of disturbances in the physiologic equilibrium between the biofilm and tooth.Previous studies have discovered an increase in the prevalence of dental caries in people with thyroid dysfunction, either as a result of the disease process itself, as a result of the surgical treatment (thyroidectomy), or as a result of medication taken that worsens oral and dental conditions. 4e amount and quality of saliva generated are both impacted by AITD, which either directly or tangentially affects salivary gland secretory function.The primary determinants of salivary defence are its composition, buffering ability, pH and flow rate.Saliva production, when reduced, has an impact on mouth health and can cause complications like oral candidiasis, gum disease, tooth cavities, and other pharyngeal and oral disorders. 3evious studies have examined the major involvement of the salivary glands in instances with AITD, which suggests that the development of thyroid and salivary gland immunological diseases may have a same mechanism. 4According to animal research, thyroid malfunction alters basal metabolic rate, which may therefore have an impact on the salivary gland's secretary unit. 5ere are very few studies demonstrating how AITD affects the salivary ducts.Because of this, the following research topic guided the execution of this study: Do AITD participants' experiences with tooth caries and their salivary characteristics correlate?The link between dental caries history and salivary profile among AITD patients was postulated to exist.The objectives were to evaluate the relationship between tooth caries and salivary profile in AITD individuals and to contrast it with controls who were of a similar age and gender.

Study design and population
In Bangalore metropolis, a cross-sectional research study involving AITD participants was carried out from June to August (pilot) and August to September (full study) in 2018.

Ethics
The Institutional Ethical Committee (GDCRI/IEC-ACM(2)/9/2018-19, dated 21/08/2018) granted their approval for this study.The appropriate approval was received from the hospital's administrators.The pilot study was conducted prior to ethical approval as this is part of the approval application process.After explicitly outlining the study's goal and methodology, participants gave their written informed consent.Prior to the commencement of the research, the investigator was trained and calibrated to guarantee reliability (k = 0.80).

Questionnaire
The Farsi oral dryness test, which consists of four queries and answer options, was used. 6During the pilot research, readability and understanding were evaluated.Necessary corrections and modifications were made.Internal consistency (α) was found to be good (0.87).

Sample size
To determine the sample number and assess the study's viability, a pilot study with 10 participants was carried out.The sample size was determined based on the frequency of dental caries using the following formula: "Where P = Prevalence of reduced unstimulated salivary flow rate i.e., 80.0%, statistical power = 80%, Z α = 1.96 at 95% confidence interval, E = margin of error-10%, Design effect, D = 1" The sample size obtained was 96.04 which was rounded up to 100.Hence, 100 AITD subjects and 100 healthy, age-and gender-matched subjects (subjects' companions) were chosen based on the eligibility criteria.

Inclusion criteria
The inclusion criteria were subjects aged 18 years and above diagnosed with autoimmune thyroid disease (both hypo and hyper AITD), and subjects with no other systemic illness.

Exclusion criteria
The exclusion criteria were subjects who smoked or drank alcohol, subjects with a history of/undergoing radio-active iodine, and subjects taking any xerogenic medication.

Inclusion criteria
The inclusion criteria were those 18 years or older without systemic illness, whose age and gender matched subjects in the AITD group.

Exclusion criteria
Excluded from the study were subjects with any habits like smoking and alcohol consumption.

Data collection
Based on inclusion and exclusion factors, research participants were chosen at random from a hospital that was randomly selected from the list of hospitals in Bangalore. 7The study period was from June to September 2018.Patients who met the eligibility criteria were approached in the hospital based on the medical records and their bystanders were included as the control group.The participants were given an information sheet outlining the purpose of the study, the specifics of the research study, and the processes, after which they consented to participate in the study.Their queries regarding the study were addressed.Written informed consent was acquired.The study's participants were assured their right to withdraw and were free to leave at any moment.From the data that were accessible from the medical records, thyroid disorders were screened for in all research participants.Among 100 AITD subjects, equal distribution of hyper and hypo AITD were maintained.Most of them were diagnosed within the past year.Data regarding thyroid hormone levels were obtained from the hospital records.Increased blood levels of thyroid stimulating hormone (TSH) > 5mIU/l, low serum free tetraiodothyroxine (FT4) < 0.61 ng/dl, and elevated thyroid peroxidase (TPO) > 34 IU/ml were used to identify autoimmune hypothyroiditis.Based on reduced (TSH) < 0.3 mIU/l, TSH receptor antibody, and elevated FT4 > 2 ng/dl, autoimmune hyperthyroidism was identified. 8Data were gathered by a single calibrated observer using an organised questionnaire, clinical evaluation, and recording by a trained assistant.In addition to demographic information, dentist appointments and oral care routines were logged.Socioeconomic status (SES) was evaluated using an adapted Kuppuswamy categorization system. 9roughout the research, infection control practises were followed.All the data were collected at one instance from each patient which comprised of demographic details, subjective assessment of saliva using Farsi's questionnaire, dental caries examination and objective assessment of saliva using GC Saliva-Check BUFFER kit.
Demographic details were first collected by the principal examiner followed by the administration of Farsi's questionnaire.Oral dryness was assessed using Farsi's oral dryness questionnaire which comprised of 4 items where a response of 'Yes' was scored as 1 and 'No' was scored as 0. Positive response (Yes) was indicative of subjective dryness.Utilizing WHO 2013 caries criteria, 10 clinical evaluation of oral caries was conducted.
The criteria for diagnosing a tooth status and the coding are as follows: Clinical examination of dental caries was followed by saliva collection of each patient.
To avoid confounding brought on by diurnal fluctuation, saliva was taken from each subject at the same time of day (between 9 and 11 AM).Viscosity was assessed visually in the oral region by observing the consistency of unstimulated saliva.Normal viscosity was indicated by watery, transparent spit.Saliva that was sticky, foamy, and effervescent was a sign of increased viscosity.
Salivary parameters were assessed using GC Saliva-Check BUFFER kit (GC India Dental Pvt.Ltd.), comprised of graduated collecting cylindrical tube with pipettes, pH strips, buffer strips, colour indicator chart and paraffin wax. 11aliva was stimulated with paraffin wax.The participants in the research were instructed to chew on a portion of paraffin wax for 5 minutes while spitting into a gathering cup every 30 seconds.Patients were handed graded collecting cylindrical tubes and instructed to sit up straight for five minutes.They were told to spit into the conduit for five minutes without ingesting (drooling method).The salivary composition, which included USFR, SFR, pH, and buffering capacity, was estimated from the collected saliva.Normal range of USFR is 0.2-0.3ml/ min and below 0.2ml/ min was considered as reduced salivary flow.Normal range of SFR is 1-3 ml/min and below 1ml/min was considered as reduced salivary flow.
The pH was measured using pH strips with unstimulated saliva.The participants in the research were told to expectorate any accumulated saliva into the gathering cup.For 10 seconds, the pH strip was inserted into the saliva.The strip's hue was contrasted with the colour indicator chart, where green represented healthy saliva.Yellow and red colours indicated moderately and highly acidic saliva respectively.Buffering capacity was measured with stimulated saliva using buffer strips.
Stimulated saliva was drawn from the collection cup using a pipette and one drop was dispensed to each of the three test pads.The colour of the strip was compared with the colour indicator chart where green colour denoted normal/high buffering capacity.Yellow and red colours indicated low and very low buffering capacity respectively.

Statistical analysis
The gathered information was put into a Microsoft Excel spreadsheet.The Statistical Package for Social Sciences was used to analyse the data (IBM SPSS Statistics V22.0).Statistics were analysed using both descriptive and inferential methods.Subgroup analysis was performed and analysis was done between autoimmune hyperthyroidism and autoimmune hypothyroidism.Pearson's association, the unpaired t-test, the chi square test and Fisher exact test, as well as linear regression, were used.Dental caries was the dependent variable in a linear regression analysis with unstimulated saliva, stimulated saliva, salivary viscosity, salivary pH, and salivary buffering capacity as the independent factors.By using an online calculator, 9 Kuppuswamy's salary categories were revised with the Consumer Price Index for Industrial Workers (CPI-IW) at 291 for June 2018.A 5% level of significance (p < 0.05) was deemed acceptable.

Results
This study included 100 AITD subjects and 100 healthy subjects (controls).The mean age for AITD group and control group were 32.20 AE 4.21years and 33.93 AE 5.55 years respectively.There was a higher proportion of female participants than males.The majority of the study subjects belonged to lower middle class (85.8%).Between the research groups, there was no statistically significant variation in terms of age (p = 0.06), gender (p = 0.30), or socioeconomic position (p = 0.29) (Table 1).
The majority of research subjects (AITD: 75%, control: 68%) had never been to the dentist.The majority of those who had attended a dentist had done so within the previous year (AITD: 72%, control: 53.1%).Tooth extraction (AITD: 40%, control: 46.9%) was the most frequently reported dental procedure, and pain was the primary factor driving dental visits (AITD: 60%, control: 62.5%).Between the research groups, there was no statistically significant variation in the duration since last dental visit (p = 0.05), the causes for the dental appointment (p = 0.82), or the type of treatment received (p = 0.76).
The majority of the AITD subjects (81%) were under anti-thyroid drugs for the past 1 year.Mean TSH values among AITD hyper and hypo groups were 0.20 AE 0.07 and 6.43 AE 1.30 respectively (Table 2).Comparing the AITD group to the control group, a substantially greater percentage of respondents reported subjective dryness (Table 3).Regarding subjective dryness, there was no statistically significant difference between the AITD hyper and hypo groups (Table 4).
When compared to the control group, the amount of stimulated and unstimulated saliva was considerably lower in the AITD group.In comparison to the control group, a considerably greater percentage of research participants in the AITD group had increased salivary viscosity.When compared to the control group, salivary pH and buffering capacity were considerably lower in the AITD group (Table 5).Regarding salivary measures, there was no statistically significant variation between the AITD hyper and hypo groups (Table 6).

Discussion
Autoimmune thyroiditis is a chronic disease in which the body interprets the thyroid glands and its hormones T3, T4 and TSH as threats. 12Sex hormones play a big role in triggering or protecting from autoimmunity.Estrogen has the ability to enhance the inflammatory process of the immune system and could contribute to AITD. 12,13The mean age of the AITD group was 32.20 AE 4.21 years which is similar to a previous study (30.06 years). 5The majority of participants were female which is in line with two previous studies. 5,14e to the chronic nature of autoimmune illness, there will be significant financial expenses as well as a negative effect on the patient's health and quality of life.Dental caries are more likely to occur when socioeconomic factors, lower levels of schooling, and lower revenue are combined. 15Most of the research participants were from the lower middle class.Age, gender, and socioeconomic position among the research groups did not vary numerically significantly, indicating homogeneity.
Dental visits are of utmost importance to high-risk groups like autoimmune thyroid disease subjects as they are more prone to a variety of dental problems.The majority of the research participants had never been to a dentist.Most of the trips were motivated by symptoms (pain), which shows that the respondents did not fully understand the value of a preventive dental appointment.
Autoimmune thyroid disease has a detrimental effect on salivary glands and cause reduction in salivary secretion. 12,16asal metabolic rate is impacted by thyroid malfunction, and the secretary unit of the salivary duct is subsequently impacted. 5Salivary secretion is reduced in AITD hypothyroidism.This is believed to be connected to the hypothyroidism-related decreased metabolism.Although the cause is unclear, AITD hyperthyroidism can reduce saliva production when it is untreated or only partly managed. 17livary mucins have an important role in maintaining rheological properties of saliva.AITD impairs secretary unit of salivary gland and cause decreased quality of salivary mucins which in turn affects viscosity, pH and buffering capacity. 18rsi's criteria have been used to evaluate subjective dryness among patients with thyroid disorders.A higher proportion of AITD subjects reported subjective dryness than in the control group, suggesting the impact of AITD on salivary flow.Unstimulated and stimulated saliva were significantly reduced in the AITD group which is similar to two previous studies, 5,14 suggesting consistent association between salivary flow and AITD.A higher proportion of study subjects in the AITD group experienced increased salivary viscosity and reduced salivary pH and buffering capacity, which was also similar to a previous study. 5Hence, salivary parameters are significantly affected with patients with AITD.
A person with systemic hypothyroidism may have slower-healing cells and tissues, making them more vulnerable to illness.As a result, they are more likely to develop mouth and tooth conditions like dental caries, frequent mandible spasms, and bleeding lips.People who have autoimmune hyperthyroidism are more susceptible to dental caries, teeth sensitivity and pain in the jaw. 13Autoimmune thyroid disease increases the risk of dental caries which has been attributed to the reduced salivary flow caused by the condition. 19Dental caries experience was significantly higher in AITD group.Indicating the efficacy of using Farsi's criteria to assess oral dryness, a moderately significant negative association was discovered between "unstimulated salivary flow," "stimulated salivary flow rate," and "subjective oral dryness."This indicates a connection between salivary flow and caries, and a marginally significant negative association was discovered between "unstimulated salivary flow," "stimulated salivary flow rate," and "dental caries."Therefore, perceived dryness may be a sign of cavities development and salivary flow rate.
Dental caries was used as the result variable in a study using linear regression.In contrast to demographic factors, TSH, salivary viscosity, pH, and buffering capacity, which were not linked with dental caries, the current research discovered a highly significant correlation between salivary flow and dental caries.
The current research has some strengths and limitations.We believe that this research is the first of its kind to link salivary parameters and tooth caries in AITD.Standard kits and criteria were used to evaluate salivary parameters reflecting high internal validity.
Although it can be seen that there are risk factors, a cross-sectional research design does not allow for the evaluation of causation between study variables.Undiagnosed systemic disease could be present and could have affected the salivary metrics.
Prospective studies are recommended to understand changes in salivary parameters and the impact of drugs over a period of time among hypo and hyper AITD patients.There must be proper maintenance of oral hygiene and diet modifications.
Periodic dental management should be conducted by well trained dentists.Even if there are no comorbid conditions, dental therapy adjustments may be required for individuals who are receiving medical supervision and follow-up for an autoimmune thyroid disease.
Thyroid dysfunction can affect every system in the body, including the mouth.An overabundance or underabundance of these hormones has a negative impact on the oral cavity.The endocrinologist must be aware with the oral symptoms of thyroid dysfunctions before treating a patient with a thyroid disease.Prior to receiving dental care from a dentist, patients who have thyroid dysfunction and those who are on medication for it need to properly manage their risks.Thus, in order to preserve the patient's thyroid and oral health, contact between the dentist and endocrinologist must be two-way.

Conclusion
The current study may indicate an underlying association between thyroid and salivary gland dysfunction.Dental caries is a multi-factorial condition where diet and oral hygiene habits play key roles.Hence, future studies should take these factors into consideration to determine whether thyroid disorder alone can cause salivary gland dysfunction resulting in oral dryness and high dental caries experience.
figshare: Data key.xlsx,https://doi.org/10.6084/m9.figshare.23585319.v1. 22is project contains the data key for the underlying data file.Results: It explains the differences between the AITD group and the control group and further explains the hyperthyroid and hypothyroid conditions.Linear regression analysis with dental caries as a dependent variable shows a significant relationship between unstimulated and stimulated saliva.The author might have also considered the duration of diagnosis and duration of anti-thyroid drug usage, as they will influence the salivary secretion and process of dental caries.Discussion: The discussion is very well articulated and mentions study limitations and recommendations.

Conclusion:
The conclusion is more general in nature.The authors can be specific with the study results obtained, even though some results might not be statistically significant but indicate a difference.

Smitha B Kulkarni
Dayananda Sagar College of Dental Science, Bengaluru, Karnataka, India Title: Title is apt.Age group of the study subjects could have been mentioned.
The introduction gives a detailed outline of the need for the study and the research questions along with objectives are clearly stated.
Methodology: Inclusion and exclusion criteria are clearly stated.Sample size determination is clear.
Methodology is explained in detail and gives a very good understanding of the study.Calibration and validation of the tools are explicitly mentioned Most of the confounding factors are eliminated.
Statistical analysis is explained clearly.
Results are well structured with good detailed explanation with tables related to all the parameters.
Discussion is elaborate and conclusions are supported by the results.

Is the study design appropriate and is the work technically sound? Yes
Are sufficient details of methods and analysis provided to allow replication by others?Yes

If applicable, is the statistical analysis and its interpretation appropriate? Yes
Are all the source data underlying the results available to ensure full reproducibility?Yes

Are the conclusions drawn adequately supported by the results? Yes
Competing Interests: No competing interests were disclosed.
Reviewer Expertise: Xerostomia related oral health problems, different mouthwashes for the treatment of gingivitis and prevention of dental caries I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.
The results section is logically structured and provides detailed data analysis of variables related to demographics, dental caries, salivary parameters, thyroid function, correlations, and linear regressions.
The discussion section adequately summarizes the study findings, provides relevant pointers from previous studies, and graciously acknowledges limitations.Its strength lies in suggesting prospective studies in future and a note on how dentists and endocrinologists can collaborate to address oral health problems in thyroid dysfunction patients.
However, in the text description regarding anti-thyroid drugs (table 2), it would be better if the results are presented as 'less than one year' and 'one year and greater' instead of "The majority of the AITD subjects (81%) were under anti-thyroid drugs for the past 1 year."for better clarity and understanding.
In table 7, the p value for DT (AITD group vs Control group should also be made bold) as it is statistically significant.

Is the work clearly and accurately presented and does it cite the current literature? Yes
Is the study design appropriate and is the work technically sound?Yes

Are sufficient details of methods and analysis provided to allow replication by others? Yes
If applicable, is the statistical analysis and its interpretation appropriate?Yes Are all the source data underlying the results available to ensure full reproducibility?Yes

Are the conclusions drawn adequately supported by the results? Yes
Competing Interests: No competing interests were disclosed.
Reviewer Expertise: Epidemiology, Oral health promotion and disease prevention, Tobacco control, Research methodology and Statistics.
I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.
The benefits of publishing with F1000Research: Your article is published within days, with no editorial bias • You can publish traditional articles, null/negative results, case reports, data notes and more • The peer review process is transparent and collaborative • Your article is indexed in PubMed after passing peer review • Dedicated customer support at every stage • For pre-submission enquiries, contact research@f1000.com Data are available under the terms of the Creative Commons Zero "No rights reserved" data waiver (CC0 1.0 Public domain dedication).Introduction: It explains AITD and its relation to salivary secretions.It establishes the role of salivary secretions changes and their effects on oral health, thus establish the need for study Methodology: It explicitly mentions sample size and sampling, inclusion and exclusion criteria, cases and controls, data collection, and a statistical plan.

Table 1 .
Demographic characteristics of the study participants.

Table 2 .
History of autoimmune thyroid disease among study participants.

Table 3 .
Subjective oral dryness among study participants according to Farsi's criteria.

Table 4 .
Subjective oral dryness among AITD groups according to Farsi's criteria.

Table 5 .
Salivary parameters among study participants.

Table 10 .
Linear regression analysis with dental caries as dependent variable.

the current literature? Yes Is the study design appropriate and is the work technically sound? Yes Are sufficient details of methods and analysis provided to allow replication by others? Yes If applicable, is the statistical analysis and its interpretation appropriate? Yes Are all the source data underlying the results available to ensure full reproducibility? Yes Are the conclusions drawn adequately supported by the results? Partly Competing Interests:
No competing interests were disclosed.

have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.
Reviewer Report 14 December 2023 https://doi.org/10.5256/f1000research.148815.r223549© 2023 B Kulkarni S. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.