Vitamin D Deficiency and Interleukin Levels in Allergic Rhinitis: A Case–Control Study

Abstract

Various studies have shown a positive co-relation between Vitamin D deficiency and severity of Allergic Rhinitis (AR) based on subjective symptoms. AR is also associated with serum eosinophilia and raised levels of various interleukins (IL)—particularly IL-4, IL-5 and IL-13. To compare serum Vitamin D levels, IL-4, IL-5, and IL-13 levels, and eosinophilia in AR patients with healthy controls and co-relate disease severity using Sino-nasal Outcome Test-22 (SNOT-22) score in patients with Vitamin D deficiency. 30 patients and 30 healthy controls were recruited. 10 ml blood sample was drawn from each patient and healthy control. It was then processed to evaluate absolute eosinophil count, serum levels of Vitamin D, and IL-4, IL-5, and IL-13. 93.33% of patients with AR and 70% of healthy controls had decreased Vitamin D levels (below 25 ng/ml). The mean Vitamin D levels was 10.50 ± 2.34 ng/ml and 17.54 ± 2.84 ng/ml in the patient and control group, respectively (p = 0.001). But there was no significant co-relation between SNOT-22 score and Vitamin D level, and between interleukin levels in patient and control group. Vitamin D deficiency is associated with Allergic Rhinitis and therefore, checking Vitamin D levels in patients with AR can be considered as routine practice in outpatient clinics. However, co-relation between severity of Allergic Rhinitis and Vitamin D levels and the proven therapeutic role of Vitamin D in Allergic Rhinitis is still debatable and thus, requires large sample size randomised controlled trials.

Introduction

Allergic Rhinitis (AR) is a global health problem. Moderate to severe forms of AR can affect work performance and daily activities. Its prevalence is up to 50% in high income countries and is relatively low in middle- and low-income countries but there is a gradual rising trend [1]. In India, the prevalence is up to 30% but is gradually on the rise since the past 2 decades [2, 3]. It is a multifactorial disease. Various studies have shown a positive association between Vitamin D and AR symptoms. However, there is ambiguity in literature when correlating deficiency of Vitamin D with severity of AR [4], 5–10. Vitamin D deficiency is seen in up to 80–90% of the population of all age groups in India [11]. In addition to Vitamin D deficiency, AR is also associated with serum eosinophilia and raised levels of various interleukins (IL)—particularly IL-4, IL-5 and IL-13 and eosinophilic proteins [12–16]. On exposure to allergen, activated Th2 cells lead to production of specific cytokines like IL-3, IL-4, IL-5, IL-13, etc. which causes increased synthesis of eosinophils and transformation of B-lymphocytes into IgE secreting plasma cells. Out of these IL-4 plays a major role in allergic immune response and production of IgE antibody and IL-5 is seen in seasonal AR and viral rhinitis and is responsible for survival of eosinophils in tissues [16–18]. However, there is lack of substantial evidence which compares vitamin D with interleukin levels in AR and healthy controls.

The prevalence of vitamin D deficiency in India is remarkably high and hence, we conducted a study to compare serum Vitamin D levels, eosinophilia, and IL-4, IL-5, and IL-13 levels in patients of AR with those of healthy controls. Sino-nasal Outcome Test-22 (SNOT-22) score was calculated in all patients to assess the disease severity. The possibility of a severe form of AR with Vitamin D deficiency was expected prior to the start of the study.

Methodology

This was a prospective case–control study conducted in the Department of Otorhinolaryngology and Head-Neck Surgery at an apex tertiary care hospital. The patients were recruited from the outpatient clinics and were diagnosed on the basis of ARIA (Allergic Rhinitis and its Impact on Asthma) guidelines [19]. The age-matched healthy controls were defined as patients who had never experienced AR symptoms and any other diagnosed disease which can affect the level of interleukins and vitamin D. The subjects who were not willing to participate, below 18 years of age, suffering from other diseases (rheumatoid arthritis, thyroid diseases, etc.) and received vitamin D supplementation which can modify the level of vitamin D and interleukins, were excluded from the study.

30 patients (Group 1) and 30 healthy controls (Group 2) were recruited. SNOT-22 score was calculated in all patients to assess the disease severity. 10 ml blood sample was drawn from each patient and healthy control. It was then processed to evaluate absolute eosinophil count, serum levels of 25 hydroxy cholecalciferol (Vitamin D) and Interleukins (IL)—IL-4, IL-5, and IL-13.

The blood for vitamin D was separated on a centrifuge at 3000 rpm for 5 min and incubated using a pre-treatment reagent. The 25 hydroxy cholecalciferol (Vitamin D) level was then measured using chemiluminescence. Vitamin D level below 25 ng/ml was considered abnormal. The serum sample for Interleukins was separated and stored at − 80 °C. Serum levels were quantified using Human Interleukin antibody coated Enzyme linked immunosorbent Assay (ELISA) kits. The separated serum was applied in the ELISA wells and fixed with reagent and buffers provided. The values were taken at 450 nm spectrometer and then compared to standard human Interleukin given in the kit.

Results

The mean age in the Allergic Rhinitis (AR) group (Group1) was 32.8 ± 4.45 years and there were 60% males and 40% females. Among healthy adults (Group 2), the mean age was 33.6 ± 4.67 years and there were 43.3% males and 56.6% females.

Eosinophilia was defined as absolute eosinophil count > 500 cells/µL [20]. 60% of the patients in the allergic rhinitis group had eosinophilia but it was not seen in any of the controls (p = 0.001). 93.33% of patients with AR and 70% of healthy controls were having decreased Vitamin D levels (below 25 ng/ml). The mean Vitamin D levels was 10.50 ± 2.34 ng/ml and 17.54 ± 2.84 ng/ml in the patient and control group, respectively (p = 0.001). There was no statistically significant difference between patient and control group with respect to interleukin levels. Table 1 shows a summary of various parameters tested in the patient and control group.

Table 1.

Comparison of various parameters in patient and control group

Serial no	Parameters	Group 1 (allergic rhinitis)	Group 2 (healthy controls)	p value
1	Eosinophilia (> 500 cells/µL)	60%	0	0.001
2	Vitamin D deficiency (mean level ng/ml)	10.50 ± 2.34	17.54 ± 2.84	0.001
3	Interleukins levels (mean level pg/ml)
a	IL-4	6.48 ± 0.25	6.35 ± 0.48	0.482
b	IL-5	174.11 ± 34.63	146.63 ± 11.61	0.150
c	IL-13	20.13 ± 2.29	23.62 ± 3.36	0.130

In AR group, there was a weak negative correlation between Vitamin D levels and serum eosinophil count, however it was not statistically significant (Pearson correlation coefficient ‘r’ = − 0.2513, p = 0.18). There was no significant correlation of SNOT-22 score with any of the parameters as shown in Table 2.

Table 2.

Corelation coefficient between SNOT-22 score and various parameters in Allergic Rhinitis group

Parameter	Pearson correlation coefficient (r)	p value
Serum eosinophilia	0.167	0.377
Vitamin D level	0.006	0.972
IL-4 level	− 0.198	0.292
IL-5 level	− 0.012	0.946
IL-13 level	0.067	0.722

SNOT-22 score sino-nasal outcome test-22 score, IL interleukin

Discussion

Allergic rhinitis is an immune (IgE) mediated inflammatory reaction of the upper airway after allergen exposure. Vitamin D deficiency is considered as a potential etiologic or disease-modifying factor in various allergic conditions including allergic rhinitis. Observational studies confirm the association of Vitamin D deficiency with AR with prevalence ranging from 30 to 90% as compared to healthy controls but interventional studies to study the effect of Vitamin D on control of allergic symptoms yield ambiguous results [8, 21–24]. Similarly, the association between severity of Vitamin D deficiency with severity of AR is controversial as studies show different results [8–10]. Our observational study in the North Indian population showed significantly lower levels of Vitamin D in AR patients as compared to healthy controls. Some similar studies from the Italian and Korean population show contrasting results and do not show any association between Vitamin D levels and allergic rhinitis [25, 26]. We estimated the severity of AR using SNOT-22 scoring but did not find any correlation between SNOT 22 and any of the parameters including Vitamin D deficiency.

A systematic review and meta-analysis to review observational studies investigating the relationship between vitamin D levels and AR concluded that prior vitamin D levels were not related to developing AR, but lower vitamin D levels were associated with a higher AR prevalence only in children. Some studies have shown potential benefit of Vitamin D supplementation on symptom control, but the meta-analysis failed to show any significant effect on AR symptoms [21, 27, 28].

Serum eosinophilia is commonly seen in allergic disorders including AR [14, 29, 30]. Some studies have shown an inverse relationship between Vitamin D levels and serum eosinophil count; however, this association requires further large sample studies [31, 32]. Our study also showed a weak negative correlation between Vitamin D level and serum eosinophil count, but it was not statistically significant. Recent studies have shown the role of nasal or local eosinophilia in AR more than serum eosinophilia [33–35].

Meta-analysis studies have shown increased risk of AR in patients with IL-13 single nucleotide polymorphism [36, 37]. It has been shown that there is positive correlation between serum IL-4 levels and AR and drugs like Dupilumab, which is anti-IL-4 receptor α monoclonal antibody, result in significantly improved AR-associated nasal symptoms [38–40]. In our study, we did not find any significantly higher levels of IL-4, IL-5 or IL-13 in AR patients as compared to healthy controls. This could be because of the inverse association of Vitamin D with pro-inflammatory markers and its positive association with anti-inflammatory markers [41]. 70% of the healthy controls in our study had low Vitamin D levels leading to raised interleukin (pro-inflammatory) levels thus, negating any significant difference of interleukin levels when compared with the patient group.

Conclusion

Allergic Rhinitis is a major health problem with increasing prevalence in developing nations. Vitamin D deficiency is associated with Allergic Rhinitis and therefore, checking Vitamin D levels in patients with Allergic Rhinitis can be considered as a routine practice in outpatient clinics. However, correlation between severity of Allergic Rhinitis and Vitamin D levels and the proven therapeutic role of Vitamin D in Allergic Rhinitis is still debatable and thus, requires large sample size randomised controlled trials.

Authors' contribution

SK and PS: Writing Original draft, data collection, and review of literature. YG and RD: Data analysis and manuscript review. HV: Study design, conception, and final revision of the manuscript. All authors read and approved the final manuscript.

Funding

The authors did not receive support from any organization for the submitted work.

Availability of data and materials

The data that support the findings of this study are available from the corresponding author, Hitesh Verma, upon reasonable request.

Code availability

Not applicable.

Declarations

Ethics approval

The authors assert that all procedures contributing to this work comply with the ethical standards of Institute Ethics Committee, All India Institute of Medical Sciences, New Delhi, India (Reference number IECPG-618/31.01.2018) on human experimentation and with the Helsinki Declaration of 1975, as revised in 2008.

Consent to participate

Informed consent was obtained from all individual participants included in the study.

Consent for publication

Patients signed informed consent regarding publishing their data.

Conflict of interest

The authors declare that they have no competing interests.