Serum ferritin level and red blood cell parameters in healthy controls and chronic periodontitis patients.

Periodontitis, which is a chronic inflammatory disease causes reduction in the number of erythrocytes and hemoglobin. It is found to be caused by specific pathogenic subgingival plaque bacteria. Periodontitis is host mediated through release of pro inflammatory cytokines by local tissues and immune cells in response to bacterial flora and its products, especially lipopolysacharides. Periodontitis is found to have systemic effect and the cytokines produced inhibit proliferation and differentiation of erythrocytes leading to anaemia. This study evaluate level of hemoglobin erythrocytes, hematocrit and serum ferritin levels in healthy subjects and periodontitis patient.

Periodontitis is a chronic inflammatory disease which causes a reduction in a number of erythrocytes and consequently lowers hemoglobin level in periodontitis patients. It has been recognized as a chronic inflammatory oral disease caused by specific pathogenic subgingival plaque bacteria. Periodontal destruction is host-mediated through release of pro-inflammatory cytokines by local tissues and immune cells in response to bacterial flora and its products, especially lipopolysaccharides. In periodontitis, the levels of interleukin-β (IL-1β) and tumor necrosis factor-α (TNF-α)[1] are significantly elevated in gingival crevicular fluid and enters systemically, falling within the detectable range of biological serum assays.[2]

Periodontitis-induced pro-inflammatory cytokines play a major role in development of variety of systemic disease[3] epidemiological studies[4] suggest that periodontitis is associated with an increased risk for systemic diseases such as cardiovascular diseases, cerebrovascular ischemia, and atherosclerosis. These associations indicate that periodontitis has systemic effects, and most likely systemic inflammation must also be present. It has been observed that infection is associated with profound disturbances in iron metabolism and decreased incorporation of iron into hemoglobin give rise to anemia. A pro-inflammatory cytokines from the chronic disease processes are involved in the pathogenesis of anemia. These cytokines inhibit proliferation and differentiation of erythrocyte progenitors,[5] modulate iron metabolism,[6] and suppress erythropoietin production[7] with resultant anemia. Hence, the present study is undertaken to substantiate that periodontitis may cause anemic state.

Materials and Methods

The subjects for this study were selected from the outpatients attending the Department of Periodontics, Tamil Nadu government dental college and hospital Chennai, after informed consent.

Inclusion criteria (for both groups):

Age groups 40–50 years

Either sex

Systemically healthy subjects.

Exclusion criteria (for both groups):

History of peptic ulcer, tuberculosis, bronchiectasis, pneumonia, lung abscess, bacterial endocarditis, chronic renal failure, hepatic disease, rheumatoid arthritis, systemic lupus erythematosus, malaria, viral infection, diabetes, menorrhagia, hodgkins disease, carcinoma, traumatic injuries, gastric surgeries, allergy, pregnancy, smokers and alcoholic.

Control group

Control group included any periodontal sight which had probing depth (PD) of ≤ 3 mm and with no clinical attachment loss (CAL).

Study group

Study group included any periodontal site which had PD of ≥4 mm and with CAL.

Study design

Examination was preceded by a thorough medical history of both he groups. A complete periodontal examination was undertaken using a mouth mirror, and Williams graduated periodontal probe. Periodontal status was assessed using the clinical parameters such as PD and level of the CAL. Full mouth intraoral periapical radiographs were taken for both the groups and 5 mm of venous blood samples were collected from antecubetal fossa.

Hematological investigations are done with blood samples, consisting erythrocyte count, white blood cell count, estimation of hemoglobin, erythrocyte sedimentation rate (ESR), and peripheral smear examination and for biochemical investigation such as blood glucose, blood urea, serum creatinine, and serum ferritin.

Probing depth

Probing depth is measured from the gingival margin to the base of the pocket using a calibrated periodontal probe. The probe is passed under the gingiva along the circumference of the tooth. Three measurements are made in the buccal aspect and three on the lingual aspect of each tooth total of six sites per tooth (mesio buccal, midbuccal, distobuccal, mesiolingual, midlingual, distolingual).

Clinical attachment level

Clinical attachment level is measured from the cemento-enamel junction to the base of the pocket using a calibrated periodontal probe.

When the gingival margin is located on the anatomical crown, the level of attachment is determined by subtracting from the pocket the distance from the gingival margin to cemento-enamel junction. If both are the same, the loss of attachment is zero.

Collection of Blood Sample

Five ml of venous blood samples was obtained by venipuncture of cubital vein in the anticubibetal fossa using 5 ml disposable syringe 23 gauge needles. The blood was then transferred to sterile vacuum tube containing ethylene diamine tetra acetic acid and transported to the clinical laboratory for processing within 4 h of venepuncture.

Results

Twenty-eight subjects were selected from the outpatients attending the Department of Periodontics, Tamil Nadu government dental college and hospital, Chennai. Fourteen subjects with PD ≤ 3 mm and no CAL were taken as a control group and the remaining 14 patients with PD ≥ 4 mm and with CAL ≥ 1 mm were taken as study group.

Clinical periodontal parameters used in this study consisted of measuring the PD and clinical attachment level in mm. Hematological and biochemical investigations performed for both the groups included erythrocyte count, estimation of hemoglobin, hematocrit, ESR, and estimation of serum ferritin. Statistical analyses were done using Student's t-test and Pearson correlation analysis.

Tables 1 and 2 show the values of clinical parameters, hematological and biochemical investigations in control and study group.

Table 1

Values of clinical periodontal parameters, hematological investigations, and biochemical investigations in control group

Table 2

Values of clinical periodontal parameters, hematological investigations, and biochemical investigations in study group

Tables 3 and 4 show the peripheral blood smear examination in control and study group. Four subjects in the control group and 2 patients in study group had microcytes, the remaining patients had macrocytes.

Table 3

Peripheral blood smear examination in the control group

Table 4

Peripheral blood smear examination in study group

Table 5 shows number of the control group and study group with values red blood cell parameters and serum ferritin levels above or below the reference range. In hemoglobin levels, 100% of the control and study group were anemic. 42.9% controls and 50% of the study group were anemic concerning hematocrit levels. Further, 14.2% of the controls and 28.4% of the study group were anemic concerning number of erythrocytes. Finally, slightly a higher proportion of study group (42.9%) had an elevated ESR compared to the control (21.4%).

Table 5

Number an percentage control and study groups with values with RBC parameters and serum ferritin below or above reference range

Table 6 shows the mean, standard deviation and test of significance of mean values between control and study group. Statistical analysis by Student's independent t-test showed than the mean PD in the study group (4.7 ± 0.5) is significantly higher than the control group (1.6 ± 0.2) (P < 0.001). Similarly, the mean CAL in the study group (5.5 ± 0.8) is significantly higher than control group (0.0 ± 0.0) (P < 0.0001). However, no other variables are statistically significant between the control and study group (P > 0.005).

Table 6

Mean, SD, and test off significance of mean values between control and study group

Table 7 shows the mean, standard deviation, and test of significance of mean values between the control and study group males. Statistical analysis showed that the mean PD and CAL are significantly higher in the study group males (PD = 4.8 ± 0.6, CAL = 6.0 ± 0.7) than the control group.

Table 7

Mean, SD, and test off significance of mean values in control and study males

Males (PD = 1.6 ± 0.2; CAL = 0.0 ± 0.0) (P < 0.0001).

Table 8 shows the mean, standard deviation, and test of significance of mean values between the control and study group females. Statistical analysis showed that the mean PD and CAL are significantly higher in the study group females (PD = 4.7 ± 0.5, CAL = 4.9 ± 0.6) than control group females (PD = 1.6 ± 0.2, CAL = 0.0 ± 0.0) (P < 0.0001).

Table 8

Mean, SD and test off significance of mean values in control and study females

Table 9 shows the correlation analysis of clinical periodontal parameters with hematological and biochemical investigations in control and study group. Pearson correlation analysis showed that there is a significant positive linear relationship between hemoglobin and CAL in study group (r = 0.45, P = 0.05).

Table 9

Correlation analysis of clinical periodontal parameters with RBC parameters in control and study group

Table 10 shows the correlation analysis of clinical periodontal parameters with hematological and biochemical investigations in control and study group males.

Table 10

Correlation analysis of clinical periodontal parameters with RBC parameters in control and study group males

Table 11 shows the correlation analysis of clinical periodontal parameters with hematological and biochemical investigations in control and study group females. There is a significant positive linear relationship between packed cell volume and PD for females in the study group (r = 0.70, P = 0.04)

Table 11

Correlation analysis of clinical periodontal parameters with RBC parameters in control and study group females

Discussion

Periodontitis is initiated by a specific bacteria, predominantly Gram-negative anaerobes[8] that activate tissue mechanism that produces a series of inflammatory and immunologic changes leading to destruction[910] of periodontium. Periodontal tissue destruction is host-mediated through release of proinflammtory cytokines by local tissue and immune cells in response to bacterial flora and its product. There have been several proinflammatory cytokines implicated in the immunopathology of periodontitis, however recent studies[11] have shown that IL-1β and TNF-α are most commonly involved in the destruction of the periodontium.

Earlier paradigms have maintained that periodontitis is an oral disease and that the tissue destruction respond remains localized between the periodontium, limiting it effects to oral tissues supporting the teeth. Recent studies have indicated that periodontitis also produces alterations in the systemic health. It has significant association with acute cerebral infarction, failure of organ replacement and kidney dialysis,[12] coronary heart disease,[13] preterm low birth weight,[14] aspiration pneumonia,[15] and diabetes.[16] It has been shown that patients with moderate to severe periodontitis had a lower number of erythrocytes, lower levels of hemoglobin and hematocrits indicating possible anemia.

In our study, the hematological investigations performed include erythrocyte count, estimation of hemoglobin, hematocrit, and ESR. And the present study also includes the estimation value of the white blood cells and peripheral smear examination. Total leukocytes count was taken into consideration to rule out parasitic infections and allergic reactions. Peripheral smear examination was performed to differentiate iron deficiency, macrocytic and normocytic anemia. Biochemical investigations such as blood glucose, blood urea, and serum creatinine were done to exclude the diabetic and renal failure patients. Estimation of serum ferritin was performed to show that in spite of low serum iron level in anemia due to chronic diseases also accompanied by a normal serum ferritin level.[17]

Pearson correlation analysis is used in this study to see the correlation between the clinical periodontal parameters and red blood parameters among the control and study groups. It showed a significant positive linear relationship between hemoglobin and CAL in study group (r = 0.45, P = 0.05), the significant relationship may indicate the beginning of subclinical or clinically evident anemic status.

Conclusion

Both the groups showed iron deficiency and macrocytic anemia, no statistically significant correlation was found to exist between periodontitis and erythrocyte count, levels of hemoglobin, hematocrit, and serum ferritin since there is no normocytic anemia.