|Year : 2015 | Volume
| Issue : 6 | Page : 628-632
Association of serum immunoglobulin-G to Porphyromonas gingivalis with acute cerebral infarction in the Chinese population
Zhang Zheng1, Ning Ma2, Youli Zheng3, Li Zhang4
1 Department of Periodontology, Stomatological Hospital, Jilin University, Changchun; Department of Periodontology, Tianjin Stomatological Hospital, Nankai University, Tianjin, China
2 Department of Periodontology, Stomatological Hospital, Jilin University, Changchun, China
3 Department of General Dentistry, Stomatological Hospital, Tianjin Medical University, Tianjin, China
4 Department of Emergency, Stomatological Hospital, Jilin University, Changchun, China
|Date of Web Publication||28-Dec-2015|
Department of Periodontology, Stomatological Hospital, Jilin University, Changchun 130021, Jilin Province
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background/Purpose: There is evidence supporting an association between ischemic stroke and periodontitis in western countries. Differing genetic backgrounds and lifestyles among populations may affect this association. The aim of our study was to determine whether antibody titers to Porphyromonas gingivalis are associated with acute cerebral infarction in the Chinese population. Materials and Methods: This case-control study was conducted on 88 acute cerebral infarction patients and 40 healthy control subjects. Serum immunoglobulin-G (IgG) antibody to P. gingivalis was analyzed by enzyme-linked immune sorbent assay. Serum lipids were determined with the automatic biochemical analyzer. Fibrinogen was measured using automated coagulation analyzer. High-sensitivity C-reactive protein (hs-CRP) and interleukin-6 (IL-6) were quantified using commercial ELISA kits. The intima-media thickness of the common carotid arteries (IMT-CCA) was measured by ultrasonography. Results: The results showed that P. gingivalis IgG antibody levels were significantly higher in acute cerebral infarction cases than in healthy controls (mean ± standard deviation, 11.06 ± 1.49 vs. 9.15 ± 1.70, P < 0.001). There were significant correlations of P. gingivalis IgG titer with total cholesterol (r = 0.34, P = 0.001), low-density lipoprotein (r = 0.39, P < 0.001), apolipoprotein-B (r = 0.30, P = 0.004), hs-CRP (r = 0.35, P = 0.001), IL-6 (r = 0.27, P = 0.011), and IMT-CCA (left: r = 0.306, P = 0.004; right: r = 0.241, P = 0.024). Conclusion: Antibody titers to P. gingivalis are associated with acute cerebral infarction in the Chinese population.
Keywords: Antibody, cerebral infarction, periodontitis, Porphyromonas gingivalis
|How to cite this article:|
Zheng Z, Ma N, Zheng Y, Zhang L. Association of serum immunoglobulin-G to Porphyromonas gingivalis with acute cerebral infarction in the Chinese population. J Indian Soc Periodontol 2015;19:628-32
|How to cite this URL:|
Zheng Z, Ma N, Zheng Y, Zhang L. Association of serum immunoglobulin-G to Porphyromonas gingivalis with acute cerebral infarction in the Chinese population. J Indian Soc Periodontol [serial online] 2015 [cited 2019 Nov 16];19:628-32. Available from: http://www.jisponline.com/text.asp?2015/19/6/628/164750
| Introduction|| |
Cerebral infarction is a type of ischemic stroke resulting from a blockage in the blood vessels supplying blood to the brain. Periodontitis, one of the most common chronic infections, has been reported as an independent risk factor for ischemic stroke in the previous studies.,, The diagnosis of periodontitis in those studies was based on self-reported questionnaire or clinical examination. However, self-reported or clinical periodontitis do not display systemic effects evoked by the periodontal disease directly., Systemic immune response to periodontitis, which can be measured as elevated serum antibody levels to periodontal pathogens, may give a better reflection of the systemic effects of the disease than the clinical examination does. Nevertheless, the immune response to periodontal pathogens in patients with cerebral infarction is poorly studied, especially in China.
Porphyromonas gingivalis, a Gram-negative bacterium, has been considered as one of the important periodontal pathogens. Elevated levels of immunoglobulin-G (IgG) to P. gingivalis have been found to be associated with carotid atherosclerosis which is the major mechanism of cerebral infarction. Dyslipidemia and raised levels of acute phase proteins like C-reactive protein (CRP), fibrinogen, and pro-inflammatory cytokines such as interleukin-6 (IL-6) are considered to play an active role in the development and progression of atherosclerotic plaques. In addition, ultrasound measurements of the intima-media thickness of the common carotid arteries (IMT-CCA) are often used as an indicator of carotid atherosclerosis. However, the literature is lacking in reports on the relationship of IgG to P. gingivalis with inflammatory markers, lipids, and IMT-CCA in cerebral infarction patients.
The objectives of this study were to investigate whether: (i) Serum IgG levels against P. gingivalis differ between acute cerebral infarction patients and health controls; and (ii) antibody levels are associated with atherogenic dyslipidemia, inflammation, and IMT-CCA in patients with cerebral infarction.
| Materials and Methods|| |
We consecutively recruited a total of 88 patients with first-ever cerebral infarction who were admitted to the Department of Neurology, the second Hospital of Jilin University (Changchun, China). The diagnosis of cerebral infarction was confirmed by a computed tomography scan or magnetic resonance imaging. A total of 40 healthy controls were selected from the health check-up center. The control group was matched to patients for age and gender distribution and time of examination. Exclusion criteria for patients and controls were recurrent cerebral infarction, hemorrhagic stroke, coronary heart disease, tumor, hypertension, diabetes, long-term use of antibiotics, serious infections elsewhere in the body. All study subjects were >18 years of age and provided informed written consent for study participation. This study was approved by the ethics committees, School of Stomatology, Jilin University.
Data on age, sex, weight, height, and system history were collected by a questionnaire. Trained nurses measured blood pressure on admission of subjects. Fasting blood was taken from the peripheral venous circulation in the next morning after admission. Blood samples of patients and control subjects were prepared, centrifuged, and stored in 0.5-ml aliquots within the first 2 h after being drawn. Sera were kept frozen at − 80°C until analysis.
Antibody to Porphyromonas gingivalis
Specific serum IgG levels against sonicated P. gingivalis ATCC33277 whole cells were measured using enzyme-linked immune sorbent assay (ELISA). Briefly, each well of a flat bottom microplate was coated with 100 μl of P. gingivalis preparation (10 μg/ml, Beijing Institute of Dental Research, China) at 4°C overnight. Before being tested, these wells were washed with phosphate-buffered saline (PBS), pH 7.2, containing 0.05% Tween 20 (PBST). Serial 2-fold dilutions of serum sample were prepared in PBS from starting dilutions of 1:25. Then, 200 μl of each dilution was added to the well and incubated at 37°C for 2 h. After washing 3 times with PBST, a 200 μl aliquot of a 5000-fold-diluted horseradish peroxidase-labeled rabbit anti-human IgG (Boster, Wuhan, China) was added to each well. The plates were incubated at 37°C for 1 h, and washed 3 times with PBST. Then, a 200 μl aliquot of o-Phenylenediamine, dissolved in phosphate-citrate buffer (pH 5.0) containing 0.4 μl/ml H2O2 in a concentration of 0.4 mg/ml, was added to each well. After 30 min incubation at room temperature, the reaction was stopped by the addition of 50 μl of 2 mol/L H2 SO4. The plates were read at 490 nm using a Microplate Reader (Labsystems, Helsinki, Finland). The final dilution is reported as the highest dilution that had an optical density two-times over background (no human serum added). The P. gingivalis antibody titer could be defined as log2 of the inverse value of the final dilution.
Laboratory measures in cerebral infarction patients
The fasting plasma glucose, triglycerides (TG), total cholesterol (TC), low-density lipoprotein (LDL), high-density lipoprotein (HDL), apolipoprotein-A (ApoA), and apolipoprotein-B (ApoB) in serum were determined with a Hitachi 7600-020 automatic biochemical analyzer (Hitachi, Tokyo, Japan). Fibrinogen was measured using a Sysmex CA-1500 automated coagulation analyzer (Sysmex, Kobe, Japan). High-sensitivity CRP (hs-CRP, BlueGene, Shanghai, China) and IL-6 (Boster) were quantified using ELISA kits obtained commercially.
Evaluation of intima-media thickness of the common carotid arteries in cerebral infarction patients
Ultrasonic evaluation of the IMT-CCA was performed with a Philips IE33 color Doppler ultrasonic diagnostic apparatus (Philips Medical System, Andover, USA) and a 7.5-MHz Phased array probe. Both right and left carotid arteries were assessed by trained sonologists.
Statistical analysis of data was performed with the SPSS package version 13.0 (SPSS, Chicago, IL, USA). First of all, the normality test was used to determine whether a data set resembles the normal distribution. Variable with skewed distribution, such as IL-6 concentration, was log transformed before testing. Continuous variables are presented as mean ± standard deviation (SD) and were compared by t-tests or One-way ANOVA, whereas categorical variables are presented as relative frequencies and were compared by the Chi-square test. The associations of P. gingivalis antibody titer with lipids, hs-CRP, fibrinogen, IL-6, and IMT-CCA were examined using Pearson correlation. Statistical significance was set at 0.05.
| Results|| |
We analyzed 88 cerebral infarction cases and 40 age and sex matched healthy controls. Besides age and sex, no significant differences were observed between groups in the body mass index, blood pressure, and glucose levels. After ELISA determination, a significantly higher P. gingivalis IgG antibody titer was found in cerebral infarction cases than in controls (mean ± SD, 11.06 ± 1.49 vs. 9.15 ± 1.70, P < 0.001) [Table 1].
|Table 1: Basic characteristics and P. gingivalis IgG titers in acute cerebral infarction cases and healthy controls|
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In analyses limited to cerebral infarction cases (n = 88), the data were divided into three tertiles according to the titers of P. gingivalis antibodies. The result revealed that increased P. gingivalis antibody (IgG) titer was associated with increased TC (P = 0.047), LDL (P = 0.015), ApoB (P = 0.04), hs-CRP (P = 0.002), and IL-6 (P = 0.047). Although not statistically significant, a trend was noted for greater TG and fibrinogen in the highest compared to the lowest P. gingivalis antibody (IgG) titer tertiles [Table 2].
|Table 2: Serum concentrations of lipids, hs-CRP, fibrinogen, and IL-6 stratified by P. gingivalis antibody (IgG) titer|
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In cerebral infarction cases, there were significant correlations of P. gingivalis IgG titer with TC (r = 0.34, P = 0.001), LDL (r = 0.39, P < 0.001), ApoB (r = 0.30, P = 0.004), hs-CRP (r = 0.35, P = 0.001), and IL-6 (r = 0.27, P = 0.011). No correlation was observed between P. gingivalis IgG titers and concentrations of TG, HDL, ApoA, and fibrinogen [Table 3].
|Table 3: Correlations of P. gingivalis antibody (IgG) titer with lipids, hs-CRP, fibrinogen, and IL-6 in acute cerebral infarction cases (n=88)|
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After the evaluation of IMT-CCA in cerebral infarction patients, an increased IMT-CCA was found in high antibody titer to P. gingivalis [Figure 1]. In addition, Pearson's correlation analysis showed a significant correlation between P. gingivalis antibody (IgG) titer and IMT-CCA (left: r = 0.306, P = 0.004; right: r = 0.241, P = 0.024) [Figure 2].
|Figure 1: Changes of the intima-media thickness of the common carotid arteries stratified by Porphyromonas gingivalis antibody (immunoglobulin-G) titer. Data are presented as mean ± standard deviation. *P < 0.05, compared to 1st tertile and 2nd tertile|
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|Figure 2: Correlation between Porphyromonas gingivalis antibody titer and the intima-media thickness of the common carotid arteries, A: Left side (r = 0.306, P = 0.004); B: Right side (r = 0.241, P = 0.024)|
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| Discussion|| |
The present study was the first time to evaluate the association between antibody titers to P. gingivalis and cerebral infarction in the Chinese population. We demonstrated that serum IgG antibody levels against P. gingivalis in cerebral infarction patients were significantly higher than that of healthy subjects. Levels of antibody to periodontopathic bacteria, especially P. gingivalis, reflect current, and past infection status as being a useful systemic biomarker for periodontitis in the patients who have not received periodontal therapy. The result observed in our study suggests that, in comparison to healthy controls, cerebral infarction cases may have a stronger host immune response induced by P. gingivalis. In previous studies, the association between antibody titers to P. gingivalis and the incident of stroke has been demonstrated in a western population. High IgG antibodies to P. gingivalis were a significant stroke risk factor in females and IgG antibody levels against P. gingivalis were linearly associated with an increased risk of stroke in seropositive males. In Japanese male subjects, IgG antibodies to P. gingivalis SU63 were significantly elevated in cerebral infarction patients. The lifestyle and ethnicity of the study population were different, but the results were consistent. These findings suggest that P. gingivalis has specific biological effects or may induce specific host responses related to cerebral infarction.
Elevated levels of IgG to P. gingivalis have been found to be associated with elevated TC and LDL, but not TG and HDL, in type 2 diabetic subjects. A similar relationship has been demonstrated in our study, serum TC, LDL, and ApoB levels were significantly associated with antibody titers to P. gingivalis. The cerebral infarction subjects with “serologically defined periodontitis,” that is, high antibody response to P. gingivalis, had higher serum TC, LDL, and ApoB levels than those without it. However, no significant association was observed between serum TG, HDL, ApoA, and antibody titer to P. gingivalis. In previous studies, increased TC and LDL have also been observed in clinically diagnosed periodontitis,, but the effect of periodontitis on TG is still controversial at present. Lim et al. reported that the percentage of sites with bleeding on probing was correlated with increased TC, LDL, and TG in diabetic subjects. A study by Katz et al. found that probing depth was correlated with hypercholesterolemia, but not hypertriglyceridemia, in subjects with coronary heart disease. Similarly, Craig et al. showed higher TC, LDL, and lower HDL in the periodontally diseased group, but no significant difference in TG. As we known, high levels of TG, TC, and LDL are correlated with the occurrence of atheromatous cerebral infarction. The association of P. gingivalis infection with dyslipidemia suggests that infection with this bacterium or response of the host against the infection may increase the risk for dyslipidemia and play a role in the pathogenesis of cerebral infarction.
In addition to lipids, circulating markers of systemic inflammation are also thought to be involved in the pathogenesis of atherosclerotic plaques and increase the risk for cerebral infarct. Furthermore, elevated concentrations of CRP and IL-6 have been observed in ischemic stroke, and associated with cerebral infarct volume., In the present study, we demonstrated that levels of hs-CRP and IL-6 were higher in subjects with elevated titers to P. gingivalis than in those with lower titers to P. gingivalis. Furthermore, higher levels of such markers were associated with higher titers to P. gingivalis. The effect of periodontitis, assessed by clinical measures, on CRP and IL-6 level has been reported in a number of studies. It is a fairly consistent finding that periodontitis is associated with increased circulating CRP and IL-6 concentrations which could be reduced after periodontal treatment.,, In addition, a significant positive association between IgG to P. gingivalis and CRP has also been found in the previous studies.,, The results of the present study are in agreement with the observations of the previous researchers. P. gingivalis infection could result in higher systemic levels of CRP and IL-6 in cerebral infarction case as well, which may increase inflammatory activity in atherosclerotic lesions and potentially increase the risk of cerebral infarction.
Dyslipidemia and inflammation are established risk factors for atherosclerosis which can be measured by IMT-CCA. Periodontal disease has been identified as a principal independent predictor of carotid arterial wall thickness. In the Atherosclerosis Risk in Communities study, Beck et al. demonstrated that clinically measured severe chronic periodontitis is associated with IMT in 6017 adults. In our study, P. gingivalis infection has also been found associated with IMT-CCA. Consistent with our findings, Beck et al. reported a positive dose-response type relationship between IgG antibody to oral organisms and carotid IMT. These findings shed light on a potential causal pathway for the association between host immune response induced by P. gingivalis and cerebral infarction.
There are some limitations in this study that need to be discussed. The primary limitation of the study is the small sample size. Clearly, this exploratory study needs to be replicated with a larger sample size to confirm the reported association in the Chinese population. Another study limitation is that the possibility of the influence of residual risk factors cannot be excluded. Many unknown risk factors could potentially impact on the relationship between IgG antibody to P. gingivalis and cerebral infarction. Third, many other kinds of Igs (e.g., IgA, IgM) was not assessed, they may be similarly associated with cerebral infarction.
| Conclusion|| |
Our study found that, compared to healthy controls, acute cerebral infarction cases had a significantly higher P. gingivalis IgG antibody titer, which was associated with high risk of cerebral infarction. Therefore, we speculated that it would have important clinical significance to prevent cerebral infarction by arresting periodontal infection and maintaining a healthy periodontium in the high-risk group of cerebral infarction.
Financial support and sponsorship
This work was supported by Natural Science Foundation of Jilin Province, China, No.201115107.
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2]
[Table 1], [Table 2], [Table 3]