Gcf Level of Il‑35 (interleukin) in Chronic Periodontitis

Research progress

In the present study, GCF level of IL-35 and clinical parameters were evaluated in chronic periodontitis patients with and without type 2 diabetes before and after SRP. To the best of our knowledge, this is the first study to compare the GCF IL-35 level before and after SRP in chronic periodontitis patients with and without type 2 diabetes. A large number of studies have linked inflammation to the development of insulin resistance as seen in type 2 diabetes. There is production of inflammatory cytokines, such as tumor necrosis factor (TNF), interleukins (ILs), and cytokine-like proteins known as adipokines in type 2 diabetes.

Homeostasis is maintained by balancing both pro and anti-inflammatory cytokines. IL-12 is one of the well studied cytokine family comprising of both pro and anti-inflammatory cytokines, which include IL-12, IL-23, IL-27, and IL-35. Out of these ILs, IL-35 has suppressive effect and it acts as inhibitory cytokine generated by Treg cell populations. IL-35 has a potential to activate Treg cells especially at high inflammation sites. Treg cells are necessary in the maintenance of immune homeostasis and the prevention of autoimmune disease.

There is evidence to suggest that anti inflammatory Treg cells also play an important role in the development of periodontal disease and are involved in the subsequent inflammation and bone resorption. The infiltration of Treg cells into periodontal tissues reflects their ability to inhibit tissue damage. Nakijima have stated that patients with chronic periodontitis exhibited increased frequency of T lymphocytes and CD4 CD25 T cells in the inflammatory infiltrate of gingival tissues and also exhibited the phenotypic markers of Tregs, such as Foxp3. These are the potential markers associated with the severity, as well as the susceptibility of periodontal disease. Various studies have concluded that periodontal therapy helps in improving HbA1C level.

In the present study, GCF samples were used for estimation of IL-35 as it is non invasive, easy to collect, and it has got local and systemic biomarkers due to host bacterial interactions. The volume of fluid coming out of the pocket increases together with raising vascular wall permeability caused by the action of inflammatory mediators.

Its composition changes during the development of inflammation. As a result of inflammation, when GCF leaks from dilated blood vessels within the gingival connective tissue, this fluid flows through the inflamed connective tissue, along with the enzymes and other mediators involved in immune response. This leaked fluid in GCF is an “inflammatory soup” containing subgingival bacteria and host cells. So, it offers a great potential as a source of factors that may be associated with disease activity.

According to several researchers, GCF protein level obtained from the sulcus with clinical symptoms of inflammation is much higher and has a concentration similar to the concentration of proteins in serum. In the present study, clinical parameters like plaque index (PI), gingival index (GI), probing depth (PD) and clinical attachment level (CAL) were assessed in all the three groups. The values of PI, GI, PD and CAL were higher in Group II and Group III compared to Group I.

There was no significant difference in PI scores and CAL between Group II and Group III. However after 6 weeks of SRP, there was significant reduction in all the clinical parameters in both the Groups. In the present study, the baseline IL-35 values were higher in experimental groups such as group II and group III compared to the group I (control group). Similar observations were made by Kalburgi, Mitani, Koseoglu in their respective studies.

Kalburgi (2013) analyzed the expression profile of IL-35 mRNA in gingiva of chronic periodontitis and aggressive periodontitis patients by semiquantitative real time PCR. They observed that IL-35 levels were higher in chronic periodontitis group compared to the aggressive periodontitis group and healthy group. The investigators stated that the increased expression of IL-35 in chronic and aggressive periodontitis suggests its possible role in pathogenesis of periodontitis. In the present study also, higher level of IL-35 levels was recorded in CP group compared to healthy group, thus supporting the findings in the previous study.

Mitani (2015) conducted a study on expression of IL-35 and IL-17, in gingival tissues with chronic periodontitis. They found that IL-35 levels were higher in GCF from chronic periodontitis patients compared to the healthy subjects. They concluded that IL-35 plays an important role in the pathogenesis of periodontitis. The findings of our study were consistent with the observations made by Mitani.

Koseoglu (2015) assessed the levels of IL-35 in GCF, saliva, and plasma in periodontal health and disease. IL-35 levels were higher in chronic periodontitis group compared to gingivitis and healthy group. They concluded that IL-35 could have an important role in suppressing periodontal inflammation and maintaining periodontal health. Similar observations were made in our study. This increase in IL-35 levels in chronic periodontitis can be attributed to its property of anti-inflammatory cytokine. Another study conducted by Jin concluded that increased level of IL 35 plays a protective role in periodontal disease by maintaining immune system homeostasis and dampening the inflammatory response.

Kaustubh Thakre (2017) compared the GCF level of IL-35 in patients with chronic gingivitis and chronic periodontitis. They found that IL-35 levels were higher in chronic gingivitis as compared with chronic periodontitis group, indicating that the levels of IL-35 decrease with increase in the inflammatory status, so it might play an important role in suppressing gingival inflammation and maintaining periodontal health. However, the findings in our study didn’t support the observations made by Kaustubh Thakre.

Various studies validated the potential approach of targeting of inflammatory mediators as a treatment for type 2 diabetes and support a causative role for inflammation in the pathogenesis of this disease. This offers the opportunity to simultaneously targeting several features of the disease with anti-inflammatory cytokines (either alone or in combination) to alter the course of the disease. Based on preclinical studies, three anti-inflammatory approaches have been clinically tested: TNF antagonism, IL-1β antagonism and salsalate treatment.

In the present clinico-biochemical study, a new anti-inflammatory cytokine, IL-35 was investigated for its role in periodontal inflammation and type 2 diabetes. As per our literature serach many studies were conducted on IL-35 and its effects on inflammatory conditions like tuberculosis, laryngeal carcinoma, Hashimotos thyroiditis and infectious conditions like hepatits. Though there are few studies describing the effects of IL-35 on type 1 diabetes, no references of the studies are available on the role of IL-35 in the pathogenesis of periodontal disease in type 2 diabetes. Our main emphasis in this study was the comparative evaluation of the effects of SRP on the GCF IL-35 level as correlated to the clinical parameters.

In the present study, IL-35 levels were found to be increased with the severity of chronic periodontitis and the values were decreased with the resolution of inflammation after the periodontal therapy (SRP). In group III subjects, the IL-35 levels (110.27± 25.97) were higher compared to the group II (87.65 ± 33.52) which depicts its inflammatory component along with altered host immune response. Another important observation made in this study was that the IL-35 levels were not reduced to a great extent after periodontal therapy in group III (83.26 ± 18.77) subjects compared to group II (47.04 ± 18.38), which suggests that the anti-inflammatory cytokines levels have a role in the pathogenesis of periodontitis and type 2 diabetes.

The results of the present study indicate that IL 35 expression increases with progression of chronic periodonititis, which may help to attenuate its progression. More specifically, gram negative anaerobic bacteria at the bottom of periodontal pockets may invade epithelial cells and hide in host cells, aggravating the destruction of periodontal tissue; they can potentially also invade endothelial cells and access the blood circulation to stimulate a host immune response and cause systemic inflammation. IL 35, as a negative regulator of immune factors, may slow or inhibit the development of periodontal disease and thus, indirectly slows down the systemic disease process.

Conclusion

On the basis of results in the present study, it was concluded that the GCF level of IL-35 can be used as a reliable indicator for the severity of periodontal disease. As IL 35 is a relatively recently identified cytokine which has not been studied in many disease models, further studies are required to prove this. Since the parameters investigated in the present study are few, it is unclear whether IL 35 enhances or antagonizes the effects of other cytokines or immune cells in CP with diabetes. In future studies, it will be necessary to increase the sample size and a greater number of parameters to understand the role of IL 35 in chronic periodontitis with diabetes and its use in the treatment of chronic periodontitis.