Serum high-sensitivity C-reactive protein levels in co- morbid patients with type-2 diabetes mellitus and periodontal disease

Purpose: To investigate the relationship between serum levels of high-sensitivity C-reactive protein (hsCRP) and the severity of periodontal disease in diabetics with periodontitis. Methods: Ninety patients were recruited for this study. They were divided into three groups, namely, group 1 (30 patients with type 2 diabetes mellitus (T2DM) and periodontal disease), group II (30 patients with T2DM only) and control (30 healthy individuals). Serum levels of hs-CRP and glycosylated hemoglobin (HbAc) were determined. Moreover, blood glucose (BG) and insulin (FNS) levels were determined in the fasted state, and their values used to compute insulin resistance index (Homa-IR). Results: Serum levels of FNS, FPG, HbAc and Homa-IR in group I patients were significantly higher (p < 0.05) than those of control group. While the levels of BG and Homa-IR in the serum of patients in groups I and II were significantly higher (p < 0.05) than those of control, marked reductions were seen in their values in group II, relative to group I. The serum levels of hs-CRP in group I and II were significantly increased (p < 0.05) relative to control, but were lower in group II than in group I (p < 0.05). Homa-IR was positively correlated with serum hs-CRP, FNS, BG, HbAc, and Homa-IR in groups I and II. Results from multiple regression analysis revealed significant effects of hs-CRP and HbAc on HomaIR. Conclusion: Serum levels of hs-CRP in patients with T2DM and periodontitis are closely related to disease severity, insulin resistance and blood glucose level.


INTRODUCTION
Studies have shown that insulin resistance and abnormal expression of inflammatory factors are the major features of T2DM [1].High-sensitivity C-reactive protein (hs-CRP) is one of the common non-specific inflammatory markers that can effectively predict the incidence of cardiovascular disease and T2DM [2,3].Periodontitis is one of the risk factors for progression of T2DM [4].However, not much is known about the relationship between serum levels of hs-CRP and the severity of periodontal disease in patients with T2DM and periodontitis.The aim of this study was to investigate the relationship between serum levels of hs-CRP and the gravity of periodontitis in patients with T2DM and periodontitis.

EXPERIMENTAL Materials
The immuno-chromatography kits used in this study was purchased from Wuhan Mingde Biotechnology Co., Ltd.

General characteristics of subjects
Ninety patients participated in this study.They were divided into three groups: group I (thirty patients with T2DM and periodontal disease), group II (thirty patients with T2DM only), and the control group (30 healthy individuals).Approval for this study was given by the Ethical Committee of Stomatology Department, Zhejiang Hospital, Hangzhou City, Zhejiang Province, 310013, China.The study was carried out in conformity with the amended Helsinki Declaration of 1964 [5].

Inclusion criteria
The inclusion criteria used were: (1) patients with periodontitis who presented with severe gingivitis, attachment loss > 5 mm, and depth of adventitia bag > 6 mm; (2) patients with T2DM consistent with the World Health Organization (WHO) classification and diagnostic criteria [6]; (3) patients with alveolar bone resorption more than 1/2; (4) patients who did not receive antibiotics 6 months before the commencement of the study; and (5) the patients who agreed to participate in the study by signing written informed consent.

Exclusion criteria
The excluded patients were: (1) patients who had endocrine system diseases that affect lipid and glucose metabolism; (2) patients who had heart, liver or kidney disease; (3) patients with malignant tumors; (4) lactating or pregnant patients; and (5) patients who voluntarily withdrew from the study.

Biochemical assays
Fasting venous blood (3 ml) was collected from each patient and subjected to centrifugation for 10 min at 3000 g to obtain serum used for immuno-chromatography.Levels of BG, HbAc and FNS were determined in the serum.The calculation for Homa-IR was done using the formula:

Statistical analysis
The results obtained are presented as mean ± standard error of the mean (SEM).Comparison between groups was done with Student's t-test, using SPSS 20.0.Statistical significance was fixed at p < 0.05.

RESULTS General characteristics of subjects
There were no significant differences in sex and age of patients in the three groups (Table 1).The characteristics of the patients are shown in Table 1, while the clinico-pathological features of periodontitis are shown in Figure 1.

Serological profile
The serum levels of FNS, BG, HbAc and Homa-IR in group I patients were significantly higher (p < 0.05) than those in control group.While the serum levels of BG and Homa-IR in patients in groups I and II were significantly higher (p < 0.05) than those of control, they were markedly lower in group II than in group I (Table 2).

Serum levels of hs-CRP
The serum levels of hs-CRP in groups I and II were significantly higher (p < 0.05) than those of  3).

Severity of periodontitis correlated with serum hs-CRP
The serum levels of hs-CRP in groups A and B were positively correlated with their FNS, FPG, HbAc and Homa-IR levels.Results from multiple regression revealed significant influence of hs-CRP and HbAc on Homa-IR (Table 4).

DISCUSSION
Under physiological or pathological conditions, the liver synthesizes and secretes large amounts of hs-CRP as an acute phase protein which serves as a non-specific biomarker of inflammation [7].The resultant increase in hs-CRP level can effectively trigger a number of signaling pathways, stimulate the inflammatory response, and reduce the sensitivity of peripheral tissues to insulin [8].As a biologically sensitive marker of systemic disease, hs-CRP is affected by factors such as systemic infection, obesity and immune system diseases [9].There were higher levels of serum FNS, FPG, HbAc and Homa-IR in group I patients than in the control group.While the levels of FPG and Homa-IR in the serum of patients in groups I and II were significantly higher than those of control, they were significantly reduced in group II, when compared to I. The serum level of hs-CRP in group II was significantly increased relative to control, but was significantly lower than that of group I.
These results suggest that T2DM may be closely related to inflammation.Some studies have suggested that elevated tumor necrosis factor affects the phosphorylation of insulin receptor substrate (IRS), and enhances the formation and release of hs-CRP [10].In addition, it has been reported that serum hs-CRP levels in patients are increased by simultaneous presence of T2DM and periodontal disease [10].The results of this study are in agreement with reports of previous studies [11].
Studies have shown that the periodontal pocket ulcer area of patients with severe periodontitis could be as high as 72 cm 2 or more, while in chronic periodontal tissue infection, the endotoxin produced by inflammatory cytokines and gram-negative bacteria can enter the systemic circulation, leading to acute inflammation [12][13][14][15][16][17].In the present study, there was a positive correlation between serum hs-CRP, FNS, BG, HbAc and Homa-IR in groups I and II patients.Results from multiple regression indicated that hs-CRP and HbAc significantly affected Homa-IR.Hence, high levels of hs-CRP in the serum of patients with T2DM and periodontitis may increase insulin resistance.Therefore, Hs-CRP can be used as a sensitive marker of inflammation.

Limitations of the study
The sample size used in this study was small.Thus, there will be need to expand the sample size in further studies and analyze the link amongst inflammation, Homa-IR and hyperglycemia.

CONCLUSION
The levels of hs-CRP in the serum of patients with T2DM and periodontitis are closely linked with the severity of periodontal disease, resistance to insulin, and blood glucose level.Thus, the drugs used can be guided possibly by monitoring the levels of hs-CRP in the serum of patients with T2DM.

Table 1 :
General clinical data of subjects Figure 1: The manifestations of periodontitis

Table 2 :
Serological profiles of the groups studied

Table 3 :
Serum levels of hs-CRP

Table 4 :
Analysis of correlation between serum hs-CRP and serological parameters