Assessment of serum levels of triglycerides, total cholesterol, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol in periodontitis patients.

BACKGROUND AND OBJECTIVES: Periodontal disease is a destructive inflammatory disease inducing profound changes in the plasma concentrations of cytokines leading to a catabolic state characterized by altered lipid metabolism and hypertriglyceridemia. The main objective of the present study was to evaluate the effect of periodontal infection on serum levels of triglycerides (TGL), total cholesterol (TC), high-density lipoprotein (HDL) cholesterol, and low-density lipoprotein (LDL) cholesterol.
MATERIALS AND METHODS: A sample of 90 subjects; 30 periodontally healthy individuals, 30 chronic gingivitis cases (n=30), and 30 chronic periodontitis cases (n=30) with an age range of 25 to 65 years were included in the study. Periodontal parameters including Plaque Index, Gingival Index, Probing Depth, and Clinical Attachment Level were recorded. Venous blood samples were obtained after 12 hours fasting period from antecubital vein and serum levels of TGL, TC, HDL, and LDL cholesterol were measured.
RESULTS: The levels of TGL, TC, and LDL cholesterol were significantly higher for periodontitis group (P<0.05) as compared to gingivitis and periodontally healthy groups. HDL cholesterol levels were significantly lower in periodontitis group (P<0.05) as compared to periodontally healthy and gingivitis groups.
CONCLUSION: The results of the present study indicate that periodontal infection has a definite role in altering lipid metabolism leading to hyperlipidemia. However, further studies are required to clarify the relationship between periodontitis and serum lipid levels and to determine whether oral healthcare has the potential to reduce serum lipid levels in otherwise systemically healthy individuals.

INTRODUCTION

Chronic inflammatory periodontal disease represents a primarily anaerobic Gram-negative oral infection that leads to gingival inflammation, destruction of periodontal tissues, loss of alveolar bone, and eventual exfoliation of teeth in severe cases.[12]

Several studies have indicated that subjects with periodontal disease may have a higher risk for cardiovascular disease.[3-6] Thus, one can speculate that periodontal disease as a chronic infection may be related to cardiovascular disease through infection-related mediators, hyperactivity of white blood cells, and altered lipid metabolism leading to hyperlipidemia. Chronic local and acute systemic infections have been demonstrated to induce profound changes in the plasma concentrations of cytokines leading to a catabolic state characterized by altered lipid metabolism. The main features of this altered metabolism are hypertriglyceridemia and lipid oxidation.[7]

Hyperlipidemia is a condition where there is an elevation of the serum levels of total cholesterol (TC) and triglycerides (TGL) due to the lipid metabolism alteration, with an increase in the liver lipogenesis and lipolysis in the adipocytes.[8] TGL are the glycerol esterified at each of its three hydroxyl groups by a fatty acid and are most abundant lipids comprising 85-90% of body lipids. Cholesterol is the most abundant steroid in animal tissues, especially food rich in animal fats; circulates in the plasma complexed to proteins of various densities and plays an important role in the pathogenesis of atheroma formation in the arteries; a precursor of steroid hormones. Low-density lipoprotein (LDL) is the compound containing both lipid and protein, which transport cholesterol to tissues other than the liver. High-density lipoprotein (HDL) is the compound containing both lipid and protein, which transport cholesterol to the liver for excretion in the bile.[9]

There have been several proinflammatory cytokines implicated in the immunopathology of periodontitis; however, some of the most convincing evidence for destruction of periodontium involves IL-1β and TNF-α. These cytokines are significantly elevated in diseased periodontal ligament sites demonstrating inflammation and during periods of active tissue destruction.[10]

Conversely, systemic exposure to infectious challenges such as bacterial lipopolysaccharide can alter fat metabolism and promote hyperlipidemia.[710]

Various studies have shown that P. gingivalis can invade deep connective tissues/vascular endothelium associated with the periodontium, can be found within vascular pathological plaques, and can elicit circulating antibody response.[11] This suggests that even in a localized oral infection, such as periodontitis, the potential exists for chronic low-level systemic exposure to microorganisms/LPS leading to generalized alterations in lipid metabolism.

Thus, it has been stated that a relationship exists between chronic periodontitis and hyperlipidemia that involves the inflammatory response to LPS from periodontal pathogens such as Porphyromonas gingivalis.[7]

Hence, an attempt was made to evaluate the linkage between periodontal disease and changes in serum lipid parameters.

MATERIALS AND METHODS

Study design

A Sample of 90 subjects (30 periodontally healthy subjects [n=30], 30 chronic gingivitis cases [n=30], and 30 chronic periodontitis cases [n=30]) were selected from patients attending Department of Periodontics, Vishnu Dental College, Bhimavaram. Prior ethical committee approval was obtained before commencement of the study.

Inclusion criteria

Patients aging between 25 to 65 of years and having ≥10 teeth were included in the study.

Exclusion criteria

Patients with systemic diseases, history of drug treatment for hyperlipidemia, pregnant women, alcoholics, and smokers were excluded from the study.

Methodology

Patient's detailed case history was recorded which included general information of the patient, medical history, drug history, history of smoking and alcoholism. Anthropometric measurements such as height in meters and weight in kilograms were recorded. Body mass index was calculated using the formula BMI = Weight/(Height)2. Plaque Index (Silness and Loe 1964)[12] and Gingival Index (Loe and Silness 1963)[13] were recorded. Probing Depth (PD) and Clinical Attachment Level (CAL) were measured by using by using UNC–15 periodontal probe.

Even though patients aging between 25 to 65 of years and having ≥10 teeth were included in the study, These patients were divided into 3 groups, i.e., healthy group, gingivitis group, and periodontitis group. Individuals with a gingival index score of 0 were considered as periodontally healthy and individuals with gingival index score of more than 1 without periodontal pockets were considered as chronic gingivitis patients. All periodontitis patients had pocket depth of ≥ 6 mm involving more than 30% of sites or clinical attachment loss of 3 to 4 mm and radiographic evidence of bone loss.

PD and CAL measurements were recorded at six sites (mesiobuccal, mid buccal, distobuccal, mesiolingual/palatal, midlingual/palatal, distolingual/palatal) by using UNC-15 periodontal probe.

Metabolic parameters

Enzymatic analysis of cholesterol levels and TGL was done using enzymes manufactured by Ensure Biotechprivate limited™ and ELICO UV-Vis™ spectrophotometer SL 159 version 6.1.

Venous blood samples were obtained after 12 hours fasting period from antecubital vein. TC and HDL cholesterol levels were determined by Trinder CHOD/POD End Point method under the principle of enzymatic determination of TC using the following reactions: Cholesterol ester is converted into cholesterol and fatty acids in the presence of cholesterol esterase. Cholesterol is oxidized in the presence of cholesterol oxidase to produce hydrogen peroxide. Hydrogen peroxide reacts with Phenol and 4-Aminoantipyrin in the presence of peroxidases to give Quinone and water molecule to produce red color. The optical density is read at 500 nm against blank.

TGL were determined by GPO/POD method. TGL are hydrolyzed by lipase to glycerol and free fatty acids. Glycerol is phosphorylated by ATP in the presence of glycerol kinase to Glycerol-3-phosphate which is oxidized by the enzyme Glycerol-3-phosphate oxidase producing hydrogen peroxide. Hydrogen peroxide so formed reacts with 4-Aminoantipyrine and 4-Chlorophenol in the presence of enzyme peroxidases to produce red Quinoneimine. The intensity of color developed is proportional to the TGL concentration.

LDL cholesterol was assessed according to the formula by Friedewald et al. in 1972, in which LDL = TC – (HDL + TG/5).

Cutoff points were taken as follows: For TGL, ≥150 mg/dl; TC, ≥200 mg/dl; HDL-cholesterol, ≤60 mg/dl; and LDL–cholesterol, ≥130 mg/dl.[1415]

RESULTS

This study included 90 subjects (47 male and 43 female subjects) with age group ranging from 25 to 65 years who satisfied the selection criteria. The selected subjects were explained about the study and an informed consent was obtained. The demographic data, periodontal parameters, and serum lipid profile were obtained. The obtained data were statistically analyzed using Microsoft Excel and Statistical software (SPSS version 16). For all these tests, a P value of 0.05 or less was set for statistical significance (S). Kruskal-Wallis ANOVA test was done for multiple group comparisons. The Mann Whitney U test was done to compare between two groups. Unpaired T test was done to compare lipid parameters between male and female subjects. Pearson correlation coefficient (r) was used to describe correlations between age, BMI, lipid parameters, and periodontal parameters.

Significant differences were found when comparison of Age (P=0.0), BMI (P=0.01), TGL (P=0.001), TC (P=0.005), LDL (P=0.001), and HDL (P=0.0) were done between 3 groups as shown in Table 1 and Figures 1–6.

Table 1

Comparison of demographic data and lipid parameters between periodontally healthy, gingivitis, and periodontitis groups

Figure 1

Comparison of mean age between periodontally healthy, gingivitis and periodontitis groups. Age in years

Figure 6

Comparison of mean high-density lipoprotein values between periodontally healthy, gingivitis, and periodontitis groups. HDL(mg/dL)

Comparison of mean body mass index between periodontally healthy, gingivitis, and periodontitis groups. BMI (kg/m2)

Comparison of mean TGL values between periodontally healthy, gingivitis, and periodontitis groups TGL (mg/dL)

Comparison of mean total cholesterol. TC(mg/dL) values between periodontally healthy, gingivitis, and periodontitis groups. TC(mg/dL)

Comparison of mean low-density lipoprotein values between periodontally healthy, gingivitis, and periodontitis groups. LDL(mg/dL)

Significant difference were found when comparison of Age (P=0.001), BMI (P=0.016), TGL (P=0.02), and HDL (P=0.004) were done between group I and II. When compared between Group I and III, significant differences were found with Age (P=0.0), BMI (P=0.005), TGL (P=0.001), TC (P=0.003), LDL (P=0.001), and HDL (P=0.00). When compared between Group II and III, significant differences were found with Age (P=0.0), TC (P=0.019), LDL (P=0.003), and HDL (P=0.00) as shown in Table 2.

Table 2

Comparison of age, BMI and lipid parameters between groups

When comparison of lipid parameters between males and females within group I, group II, and group III was done, none of the parameters are significant, as shown in Table 3.

Table 3

Comparison of lipid parameters between male and female subjects

When comparison of lipid parameters between age, gender, and BMI within group I, group II, and group III was done, none of the parameters are significant, as shown in Table 4.

Table 4

Correlations between age, gender, BMI, and lipid parameters

When comparison of lipid parameters between GI and PI within group I, group II, and group III was done, none of the parameters are significant. Probing Pocket Depth (P=0.01) and CAL (P=0.004) were significant within group 3 only, as shown in Table 5.

Table 5

Correlations between periodontal and lipid parameters

DISCUSSION

Over many years, the prevailing view was that periodontal infections were localized only to the periodontium and rarely had systemic implications in healthy individuals. More recent evidence, however, has shown that periodontitis induces increased systemic inflammation, as indicated by raised serum levels of various inflammatory markers leading to development of variable systemic complications such as cardiovascular disease, cerebrovascular disease, respiratory diseases, pregnancy complications, altered glycemic control, and renal disease.[16]

A significant association between periodontitis and cholesterol has been reported.[710] Hence, this study was designed for better understanding the association between periodontal disease and serum lipid levels.

Ninety subjects with age range between 25 to 65 years having ≥10 natural teeth were included. Age matching could not be done as most of the periodontitis patients were of higher age group.

The mean BMI for all the three study groups was <25. This suggests that the subjects were in normal weight range.[17] Thus, the potential effect of obesity on lipid profile levels was eliminated.

Highest mean values of TGL, TC, and LDL were observed for periodontitis group and least mean values were observed for periodontally healthy group. On the other hand, HDL value was highest in the periodontally healthy group and least for periodontitis group after eliminating potential confounders. This elevated lipid parameters in periodontitis group is mainly due to the elevated proinflammatory cytokines produced by chronic periodontitis.

Periodontal infection induces the APR leading to multiple alterations in lipid and lipoprotein metabolism. Plasma TGL levels increase from increased VLDL secretion as a result of adipose tissue lipolysis, increased de novo hepatic fatty acid synthesis, and suppression of fatty acid oxidation. Hypercholesterolemia occurs attributable to increased hepatic cholesterol synthesis and decreased LDL clearance, conversion of cholesterol to bile acids, and secretion of cholesterol into the bile.[18]

IL-1β and TNF-α exert an effect on lipid metabolism by influencing production of other cytokines, altering hemodynamics/amino acid utilization of various tissues involved in lipid metabolism, or modifying the hypothalamic-pituitary-adrenal axis increasing plasma concentrations of adrenocorticotropic hormone, cortisol, adrenaline, noradrenaline, and glucagon.[10]

Further studies have explained this concept by highlighting the fact that periodontal disease results in elevated serum levels of aspartate aminotransferase, alanine aminotransferase, cholinesterase, and an aspartate aminotransferase-to-alanine aminotransferase ratio less than one, suggesting that they have a tendency toward steatohepatitis leading to hepatic dyslipidemia.[18]

In contrast to the results of the present study, no raise in levels of TGL and cholesterol were observed in periodontitis group when compared to control group.[19] This could be due to the collection of non-fasting venous blood in that study that might have altered the plasma lipid values. In our present study, blood was drawn after 12 hours of fasting as food intake may alter the serum TGL levels.

The present study showed a significant increase in the TGL value and a significant decrease in the HDL value in the gingivitis group compared to the periodontally healthy group. This increase in TGL value may be because of the difference in the physical activity, food habits, and socioeconomic conditions of the subjects in the two groups. But, the difference in mean TC and LDL values between the two groups was not significant. This might be due to the less severity of proinflammatory cytokine production in gingivitis compared to periodontitis condition leading to minimal systemic manifestations. To the best of our knowledge, no study has been reported in the literature comparing the lipid profile in periodontally healthy group and gingivitis group.

A significant increase in TC, LDL values and a significant decrease in HDL values were seen in periodontitis group when compared to gingivitis group. This signifies that inflammation of the superficial structures of the periodontium may not alter the lipid metabolism. However, absence of significant difference in TGL values may be due to a variation in dietary and exercise pattern which could not be excluded as dietary records were not kept, that might have affected along with socioeconomic conditions.

When comparing the lipid profile between male and female subjects in periodontally healthy group and gingivitis group, TGL and HDL values were more for females when compared to male subjects. However, TC and LDL values were more for male subjects than females. Moreover, in periodontitis group, TGL, TC, and HDL values were more for male subjects and LDL value was more for female subjects. But, the values were not statistically significant. This shows the absence of consistent pattern of variation among male and female subjects. This can be due to the unequal distribution of male and female subjects in each group.

The results of the present study demonstrated a positive correlation between probing depth and clinical attachment level with TGL, TC, LDL and a negative correlation with HDL values. This shows that with increasing probing depth and CAL, the values of TGL, TC, and LDL increased but the value of HDL decreased. This is in accordance with the study conducted by Katz et al. who hypothesized that there is a strong positive statistical association between the existence of periodontal pockets and plasma lipid levels, thereby confirming a positive relationship between periodontitis and hyperlipidemia.[20]

Machado et al. demonstrated a positive correlation between tooth loss and the levels of TGL, TC, and LDL and a negative correlation between tooth loss and the levels of HDL.[21] Similar results were obtained by Morita T et al. who showed that positive conversions of blood pressure and the blood lipid index were significantly associated with the presence of periodontal pockets.[22]

In contrast to the results of present study, the results of the study conducted by Chen L et al. showed no positive correlation of TC, HDL cholesterol, LDL cholesterol, and TGL with the mean probing depth. This might be due to the inclusion of diabetic patients in the study population, as diabetes acts as a potential risk factor for the development of hyperlipidemia. The advantage of present study is the elimination of all confounding factors.[23]

CONCLUSION

The present study indicates that periodontal infection has a definite role in altering lipid metabolism which supports the hypothesis that a relationship exists between chronic periodontitis and hyperlipidemia.

Based on our results, following conclusions can be drawn:

Periodontitis patients have a significant increase in TGL, TC, and LDL levels and decrease in HDL levels compared to periodontally healthy group and gingivitis group

A positive correlation exists between probing depth and CAL with TGL, TC, and LDL

A negative correlation exists between probing depth and CAL with HDL value

Gingival inflammation alone in the absence of probing depth and attachment loss has no effect on the serum lipid profile.

However, further studies are required to clarify the relationship between periodontitis and serum lipid levels and to determine whether oral healthcare has the potential to reduce serum lipid levels in otherwise systemically healthy individuals.