|Year : 2015 | Volume
| Issue : 6 | Page : 618-623
Relationship between interleukin 1α levels in the gingival crevicular fluid in health and in inflammatory periodontal disease and periodontal inflamed surface area: A correlative study
Kalaichelvi Govindarajan, Santhanakrishnan Muthukumar, Suresh Rangarao
Department of Periodontics, Sri Ramachandra University, Chennai, Tamil Nadu, India
|Date of Web Publication||28-Dec-2015|
Department of Periodontics, Sri Ramachandra University, Porur, Chennai - 600 116, Tamil Nadu
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background: Periodontitis has been suggested as a source of inflammation for pathological changes in distant sites. Interleukin-1 alpha (IL-1α) has shown to have specific roles in inflammation, immunity, tissue breakdown, and tissue homeostasis. This study assessed the correlation of periodontal inflamed surface area (PISA) index with the gingival crevicular fluid (GCF) levels of IL-1α, which would be helpful in evaluating the validity of PISA index in terms of reflection of the disease. Materials and Methods: A total of 40 subjects were recruited for this study and 20 subjects with healthy gingiva (Group I) served as controls and 20 subjects served as cases with periodontitis (Group II). Samples of GCF were obtained from one site in each patient by placing color-coded, calibrated, volumetric microcapillary pipettes extracrevicularly, and subjected to ELISA test. Results: There was a statistical significance for mean probing depth (PD) and periodontal epithelial surface area (PESA) (P < 0.01), mean bleeding on probing and PISA, IL-1α (P < 0.01) and PESA (P < 0.05) in Group I. Statistical significance was found between PISA and IL-1α in Group I (P < 0.01). A positive correlation was found in Group II between mean PD and mean attachment loss (P < 0.01), PISA, IL-1α and PESA (P < 0.01), PISA and IL-1α levels (P < 0.01) which was statistically significant. Conclusion: The inflammatory burden index was statistically significant in the periodontitis group correlating with higher IL-1α levels, which clearly indicates the validity of PISA index.
Keywords: Gingival crevicular fluid, interleukin alpha, periodontal inflamed surface area
|How to cite this article:|
Govindarajan K, Muthukumar S, Rangarao S. Relationship between interleukin 1α levels in the gingival crevicular fluid in health and in inflammatory periodontal disease and periodontal inflamed surface area: A correlative study. J Indian Soc Periodontol 2015;19:618-23
|How to cite this URL:|
Govindarajan K, Muthukumar S, Rangarao S. Relationship between interleukin 1α levels in the gingival crevicular fluid in health and in inflammatory periodontal disease and periodontal inflamed surface area: A correlative study. J Indian Soc Periodontol [serial online] 2015 [cited 2020 Jun 1];19:618-23. Available from: http://www.jisponline.com/text.asp?2015/19/6/618/162197
| Introduction|| |
The plaque bacteria pose a microbial challenge to the periodontium and in turn stimulate the host immune response.
Among the cells of the immune system, the monocytes and macrophages are agranulocytes that are powerful phagocytes. In addition to mediating and performing phagocytic function, the monocytes produce two important cytokines namely Interleukin-1(IL-1) (Ani Biotech Oy, Orgenium Laboratories, Vantaa, Finland) (α and β) and tumor necrosis factor alpha (TNF-α) which causes tissue damage and bone resorption as seen in periodontal disease.
Interleukin-1 is a polypeptide with a wide variety of activities and roles in tissue homeostasis, inflammation, immunity, and tissue breakdown. Following activation, it is synthesized by various cell types, including monocyte, macrophage, T-lymphocyte, fibroblast, vascular cell, brain cell, and skin cell. There are two subtypes of IL-1, designated IL-1α, and IL-1β, produced mainly by monocytes and macrophages but also by other types of cells.
Interleukin-1α is described as “pro-inflammatory” because it stimulates the activity of genes involved in inflammation and immunity. This protein plays a critical role in protecting the body from foreign invaders such as bacteria and viruses. It is also involved in bone resorption, the breakdown and removal of bone tissue that is no longer needed.
Any classification of periodontitis as a risk factor for other diseases should quantify the amount of inflamed periodontal tissue in order to quantify the inflammatory burden. Therefore, the periodontal inflamed surface area (PISA) was proposed as a classification of periodontitis that quantifies the amount of inflamed periodontal tissue and as such, quantifies the systemic inflammatory burden.
Gingival crevicular fluid (GCF) provides a unique window for analysis of periodontal condition as the levels of inflammatory mediators, which results due to the increased local destruction of connective tissue structural elements represent the ideal markers of disease activity can be estimated in the GCF.
Thus, the aim of this study was to correlate the PISA index values with the GCF cytokine levels of IL-1α, which would be helpful in evaluating the validity of PISA index.
| Materials and Methods|| |
This is a correlative study were two groups healthy (Group I) and chronic periodontitis (Group II) participated. The sample of convenience was used and 40 subjects were recruited for this study and 20 subjects were selected as cases with chronic periodontitis (Group II) and 20 subjects with healthy gingiva (Group I) served as controls after obtaining informed consent from the patients and ethical approval from the Institutional Ethics Committee and Review Board of Sri Ramachandra University.
Patients who had healthy gingiva as determined by absence of bleeding on probing (BOP), no attachment loss (AL), no mobility or furcation involvement, and were otherwise systemically healthy with no previous history of periodontal disease were selected as controls representing Group I.
The chronic periodontitis group was represented by patients presenting with the presence of at least 10 natural teeth, chronic generalized periodontitis as evidenced by the presence of AL >1 mm in more than 30% of sites examined, mobile teeth, furcation involvement, and should be otherwise systemically healthy.
In both the groups, CAL, recessions, and BOP were recorded to calculate the PISA values using a freely downloadable Microsoft excel spreadsheet was constructed to facilitate PISA calculation from the site www.parsprototo.info., described by Nesse et al.
The criteria for exclusion included; subjects who use tobacco in any form, alcoholics, subjects who have taken antibiotics in the past 6 months, subjects who have taken analgesics in the past 1-week, pregnant and lactating women, subjects who had undergone any periodontal therapy in the last 6 months, subjects with a history of any systemic diseases.
| Sample Collection|| |
Gingival crevicular fluid sample collection
Patients were asked to gargle with a glass of cold sterile water to cleanse the teeth of loosely adherent debris. The sites selected randomly were dried and isolated with cotton rolls. Samples of GCF were obtained from one site in each patient by placing color-coded, calibrated, volumetric microcapillary pipettes extracrevicularly. Pipettes that were contaminated with blood/saliva were discarded. From each site, a standardized volume of 2 μl was collected and immediately transferred to a eppendrof tube and frozen at −70°C and subjected to ELISA (Ani Biotech Oy, Orgenium Laboratories Business Unit, Tiilite 3, FIN-07120 Vantaa Finland) test.
The assay was performed using IL-1α kit. The manufacturer's instructions were carefully followed and each plate was checked before use to ensure the calibration curve measured IL-1α standards (3–250 pg/ml) within the stated limits of the assay. The kit made use of biotinylated anti-human IL-1α. The substrate color reaction was measured at 450 nm by an ELISA reader. The optical density values obtained with the known samples were used to calculate the quantity of IL-1α.
The demographic characteristics of the study population were calculated using t-test. The correlation between the parametric values and IL-1α levels was obtained using Pearson's Correlation and Fisher exact-2 tailed test.
| Results|| |
The study comprised of 40 subjects (19 males and 21 females, mean age 27.75–36.20 years). Group I (n = 20) and Group II (n = 20). The patients characteristics such as age, mean probing depth (PD), mean BOP, oral hygiene index (OHI) score, PISA, and IL-1α are reported in [Table 1] [Figure 1] and [Figure 2] and characteristic of gender in [Table 2] [Figure 3]. T-test was used to calculate the mean and standard deviation of the demographic data.
|Figure 1: Comparison of periodontal inflamed surface area between groups|
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Pearson correlation and Fisher exact-2-tailed tests were used to assess the strength of association between parametric and nonparametric variables in Group I [Table 3] and Group II [Table 4] [Figure 4], respectively. There was a statistical significance for mean BOP, PISA, and IL-1α (P < 0.01) in Group I. Statistical significance was found between PISA and IL-1α in Group I (P < 0.01).
|Table 3: Correlation between PISA and IL-1 alpha in Group I (healthy group)|
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|Table 4: Correlation between PISA and IL-1 alpha in Group II (chronic periodontitis group)|
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|Figure 4: Scatter plot correlating periodontal inflamed surface area and interleukin-1 alpha|
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In [Table 4], a positive correlation was found in Group II between mean PD and mean AL (P < 0.01), PISA, PISA, and IL-1α levels (P < 0.01), which was statistically significant.
| Discussion|| |
Periodontitis results in the loss of connective tissue attachment of a tooth to the surrounding bone. Previous studies have noted that periodontal disease exhibits periods of quiescence and exacerbation during its course of progression. A critical problem in periodontal disease is a lack of any reliable criteria to determine the extent of disease activity or rate of disease progression at a given time., The traditional clinical parameters such as PD, gingivitis score, GCF flow, and radiographic appearance of alveolar bone are not considered to provide accurate diagnostic aids. It is thought that there is a potent local mediator of tissue destruction associated with inflammatory pathologies such as rheumatoid arthritis and periodontitis. IL-1 has been suggested to have a key regulatory influence on these disease processes, since a significant elevation of this factor has been found in these diseased tissues.,
Lipopolysaccharide is a key microbial stimulus that initiates the host response at periodontal disease sites and triggers monocytes to release inflammatory mediators (prostaglandin E2, thromboxane B, IL-1, -6 and -8, TNF, and collagenase) that increase the local destruction of connective tissue structural element. Therefore, the levels of monocytic inflammatory mediators in GCF may be ideal markers of disease activity at a particular site. As there are several studies regarding the other pro-inflammatory cytokines such as IL-1β, TNF α, and IL-6, we decided to estimate the IL-1α levels, in healthy and diseased conditions as it is considered to be a cytokine that is also a powerful mediator of inflammation.
It is fair to expect that the bacterial infection induces a destructive immunologic reaction dominated by mononuclear inflammatory cells and resident tissue cells, all producing pro-inflammatory cytokines interfering with the regeneration process. IL-1α is one of the most potent pro-inflammatory cytokines, which is mainly synthesized by monocytes and play an important role in periodontal pathogenesis and destruction, however, the course and extent of the host response is influenced by genetic factors.,
A major drawback of the studies published on the periodontal inflammation systemic disease interaction is the lack of a tool that adequately assesses the inflammatory burden of periodontitis. Therefore, a new measure of periodontitis as a risk factor for other diseases was developed, the PISA. PISA reflects the surface area of bleeding pocket epithelium in square millimeters. PISA is calculated using conventional CAL, recession, and BOP measurements. PISA quantifies the amount of inflamed periodontal tissue, thereby quantifying the inflammatory burden posed by periodontitis. Broad application of PISA may provide decisive conclusions on periodontitis as a risk factor for other diseases. An additional advantage of the PISA is that it can be retrospectively calculated using existing research data containing CAL, recession, and BOP measurements.
It has been previously reported that both IL-1 activity in GCF and IL-1 activity index which uniquely were closely associated with the severity of periodontal disease as classified by alveolar bone resorption. Many reports have indicated that IL-1 activity was higher in GCF obtained from inflamed gingival sites than in that from inflammation-free sites, and several reports have shown a relationship among the effects of periodontal therapy, IL-1 activity in GCF and the localization of IL-1 in inflamed gingival tissue. However, conflicting results have been reported regarding the effect of periodontal treatment on IL-1 levels. One possible explanation for these conflicting results may be the dilution effects of increased GCF volumes in inflammation that may confound measures. Therefore, the levels of monocytic inflammatory mediators in GCF may be ideal markers of disease activity at a particular site.
Many studies have reported that GCF IL-1 levels are significantly elevated in all forms of periodontitis, compared to health or gingivitis. Studies have also confirmed an association between elevated GCF IL-1 levels and gingival inflammation, as well as a relationship between the severity of periodontitis and elevated GCF IL-1 levels. Matsuki et al. reported IL-1 activity in GCF from inflamed gingiva was higher than that extracted from the healthy gingiva. Ishihara et al. reported that the degree of periodontitis classified according to alveolar bone resorption was correlated with the volume of IL-1α and IL-1β in GCF and the level of IL-1 activity index.
To our knowledge, this is the first study, correlating the PISA index with the GCF cytokine levels which would be helpful in evaluating the validity of PISA index.
The present study was performed to examine the relationship between clinical parameters, PISA, and IL-1α in GCF of healthy and chronic periodontitis patients. It has been shown by many investigators that IL-1α, IL-1β, IL-8, and TNF-α can be detected in GCF in periodontal disease and that the cytokine levels in GCF are closely associated with the severity of gingival inflammation and/or periodontal tissue destruction. It has been shown that IL-1α, IL-1β, IL-6, IL-8, and TNF-α can be detected in the GCF. Higher levels of these mediators in the GCF were detected in the periodontal disease. Each subjects were evaluated in terms of OHI, PD, AL, BOP. GCF was collected from each patient in the deepest probing pocket depth of about 2 μl.
The mean PD, Mean BOP, and OHI scores were increased in Group II compared to Group I. The periodontal epithelial surface area and PISA scores were also higher in Group II compared to Group I. However, conflicting results were obtained by Yoshinari et al. when they compared clinical parameters and the volume of IL-1α, -1β and -1ra in GCF, and IL-1 activity index. They found that both PPD and CAL increased, that is, the severity of inflammation became clinically serious, and the amount of IL-1 β in GCF also increased. However, the amounts of IL-1α, -1ra, and IL-1 activity index did not change.
In our study, commercially available ELISA test kit was used to estimate IL-1α in GCF collected with microcapillary pipettes. Micropipettes were used to rule out the possibility of polypeptides getting adsorbed to cellulose. Our study showed that the concentration of IL-1α in the GCF of periodontitis was higher (P < 0.0001) than those with healthy.
The values obtained in patients with healthy gingiva range between 13 pg/ml and 163 pg/ml. This wide range may be attributed to the variation in plaque accumulation or variation in subsequent inflammation due to plaque. However, another possibility put forth by Kinane et al. that needs to be considered is the inherent variation in the ability to produce IL-1 and that could result in heterogenous responses accounting to intraspecies differences. Looking into the results obtained with samples in Group II, it may be noted that the values fall between a wide range (121 pg/ml and 250 pg/ml). Similarly, higher levels of these mediators in the GCF were detected in the periodontal disease.
There was a positive correlation between IL-1α and PISA in periodontitis groups compared to healthy in the present study. Similarly, cytokine levels in GCF were closely associated with the severity of gingival inflammation and/or periodontal tissue destruction. It has been previously reported that both IL-1 activity in GCF and IL-1 activity index were closely associated with the severity of periodontal disease as classified by alveolar bone resorption. Many reports have indicated that IL-1 activity was higher in GCF obtained from inflamed gingival sites than in that from inflammation-free sites.
Therefore, determination of the presence of inflammatory products in GCF may be useful for evaluating periodontal disease status. Studies have reported a positive correlation between level of GCF inflammatory mediators and clinical periodontal conditions, whereas other studies have reported a poor correlation between such levels and the site clinical status. Similarly, the inflammatory burden index was statistically significant in the periodontitis group, correlating with higher IL-1α levels, which clearly indicates the validity of PISA index. However, the limitations of the study include the nonestimation of serum IL-1α levels, which would have given the impact of periodontal disease on systemic inflammation and the effect of periodontal treatment on the serum and GCF levels could have proved an importance of periodontal therapy in reducing the systemic inflammatory burden.
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Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4]
[Table 1], [Table 2], [Table 3], [Table 4]