now i will provide you with data from five studies. Can you interpret together the data and the results of the three studies and especially concerning HBD-2 ? Study 1: 1. Study design
“The study protocol was approved by the Ethics Committee on Human Research of the School of Dentistry at Araraquara - UNESP (protocol number 01394712.5.0000.5416). The purpose and content of the study were explained to eligible subjects, and written informed consent was obtained from all participants for inclusion in the study. A total of 70 participants were recruited from June 2016 to March 2018 at the Periodontology Clinic, School of Dentistry at Araraquara - UNESP. The diagnosis of periodontitis was performed according to the classification of the 2017 World Workshop on the Classification of Periodontal and Peri-implant Diseases and Conditions 12 . All patients should present a minimum of 15 natural teeth. Twenty patients were included in the periodontally healthy control group. All subjects were free of systemic and periodontal diseases. They should present probing depth (PD) ≤ 3 mm, no bleeding on probing (BOP), no clinical attachment loss, plaque index (PI) and gingival index (GI) ≤ 15%. Fifty patients with periodontitis stage III or IV were included in periodontitis groups. In addition to the diagnostic criteria 12 , they should present at least five sites in non-adjacent teeth with PD ≥ 4 mm, BOP, and clinical attachment level (CAL) ≥ 4 mm, except for third molars. From the 50 patients, 25 had never smoked and 25 were smokers who should smoke at least 10 cigarettes/day for a minimum of 5 years.
None of the patients had a history of systemic diseases or immunological disorders. Subjects who had received antibiotics and anti-inflammatory drugs within the last 3 months or received periodontal treatment within the last 6 months were not included in the study. Patients using hormone replacement therapy, pregnant and lactating women and orthodontic patients were also excluded.
The sample size was calculated using statistical software (GPower 3.1 Statistical Power Analysis for Windows, Düsseldorf, Germany). Considering mean values of hBD expression and standard deviation from previous studies 13 , 14 , 15 , an effect size of 0.3 was obtained. Based on this effect size, a significance level of 0.05 and a study power of 0.80, a sample size of 20 individuals per group were determined to be necessary.”,
2.Clinical examination
“One calibrated examiner performed all clinical examinations. The intra-examiner reproducibility was assessed by measuring the PD in six teeth (16, 21, 24, 36, 41 and 44), according to the Ramfjord index, in a total of six patients (36 sites/patient) on two different occasions in a time interval of 7 days. Data were submitted to Kappa concordance analysis (k=0.91). The parameters PD, BOP and CAL were assessed at six sites per tooth (mesio-buccal, mid-buccal, disto-buccal, mesio-lingual, mid-lingual, disto-lingual), while PI and GI were registered as absence (negative site) or presence (positive site) of visible plaque and gingival bleeding at four sites per tooth (mesial, distal, buccal and lingual), excluding the third molars. Measurements were performed with a manual periodontal probe (UNC15; Hu-Friedy, Chicago, IL, USA).”, 3. Gingival crevicular fluid (GCF) sampling :
“GCF samples were collected from healthy (n=5) sites of each periodontally healthy patient and from healthy (n=5) and diseased (n=5) sites of each periodontitis patient. Sites at different non-adjacent teeth, preferably one site per quadrant, were selected for GCF collection in each subject, one day after the periodontal examination. Healthy sites had no gingival inflammation, PD ≤ 3 mm, no BOP or clinical attachment loss; and diseased sites had gingival inflammation, PD and CAL ≥ 4 mm with BOP. Diseased sites were selected according to the deepest probing depth in each quadrant. A single examiner performed GCF sampling. The sites to be sampled were isolated with cotton rolls, the supragingival plaque was removed with a sterile curette and the surfaces were gently air-dried. GCF was sampled by inserting absorbent paper strips (Periopaper®, Oraflow Inc., New York, USA) into the gingival sulcus or periodontal pocket for 30 seconds. Paper strips contaminated with blood and saliva were rejected. An electronic device Periotron 8000 determined the GCF volume absorbed into each strip (Periopaper, ProFlow, Inc., Amityville, NY, USA), which was calibrated based on a protocol described before 16 (calibration of the electronic device by polynomial regression). The GCF was obtained as a pooled sample of the same category (healthy or diseased site) in each subject. The papers strips were immediately placed into a dry sterile polypropylene tube (5 paper strips per tube) and kept at -80 °C until analysis. The readings from the Periotron 8000 were converted to an actual volume (μL) by reference to a standard curve.”,
4. Immunosorbent assay for hBD 1 and hBD 2 quantification:
“First, the absorbed GCF was eluted from the paper strips by adding 750 μL (150 μL per strip) of 1x Phosphate-Buffered Saline (PBS) to each tube. The tubes were left on ice in a horizontal shaker for 30 minutes and then centrifuged at 4 °C, 13.000 g for 10 minutes. The GCF levels of hBD 1 and 2 were measured using the sandwich ELISA technique according to the manufacturer’s guidelines (Peprotech, Rocky Hill, NJ, USA). The absorbance was measured by a spectrophotometer at 450 nm wavelength. The levels of AMPs in each sample were determined using the concentration values of standards included in the kit contents. The results were expressed in pg/mL.” and
5. Statistical analysis
“The statistical analysis was performed using the software GraphPad Prism 7 (San Diego, CA, USA). For clinical data, normal distribution was analyzed using the D’Agostino and Pearson test. For normally distributed data, it was used One-way ANOVA followed by Tukey for healthy sites and unpaired t test for diseased sites on inter-group analyses and paired t test for intra-group analyses. For non-normally distributed data, Kruskal-Wallis and Dunn were used for healthy sites and Mann-Whitney tests was used for diseased sites on inter-group analyses, while the Wilcoxon test was performed for intra-group comparison. A p-value of less than 0.05 was considered significant. The effect size was estimated for the primary outcome of the study (AMPs level in GCF). Since non-parametric tests were used for intra and inter-group comparisons, the eta squared and the correlation coefficient r were calculated.” 1. Demographics and periodontal parameters
“A significant age difference was found between periodontally healthy subjects (25.79 ± 3.95) and periodontitis patients (NS [40.32 ± 12]; S [43.88 ± 10.64]) (p<0.05). The distribution of the male and female participants had no significant difference between the study groups, periodontally healthy subjects (5 males/15 females) and periodontitis patients (NS - 9 males/16 females; S - 12 males/13 females) (p = 0.28). The periodontal clinical parameters are presented in table 1. Regarding the full mouth periodontal clinical parameters, PI, BOP, PD and CAL were significantly lower in periodontally healthy subjects than in periodontitis patients (p<0.0001). Only for GI, a significant difference was observed between periodontally healthy patients and smoker patients (p=0.0001). The sampled sites of periodontally healthy subjects presented significantly lower values of PD and CFV than healthy sampled sites of smokers and non-smokers with periodontitis (p<0.05), as well as lower CAL than healthy sampled sites of smokers (p<0.05) (Table 2). In patients with periodontitis, the intra-group comparison revealed lower levels of PD, CAL and CFV in the healthy sampled sites than in the diseased sites (p<0.0001) (Table 2).” ,
2. hBD 1 and hBD 2 quantification
“The GCF levels of hBD 1 and hBD 2 are presented in Figure 1. The hBD 1 levels were significantly higher in healthy sites of the patients without periodontal disease than in healthy sites of smokers and non-smokers with periodontitis (p<0.0001), with a mean difference of 31.62 (95% CI: 17.15 - 46.08) between SH and H and a mean difference of 31.20 between NSH and H (95% CI: 13.81 - 48.57). The lowest GCF hBD 2 level was observed in healthy sites of non-smokers (p=0.0003). A mean difference of 33.61 was observed between SH and H (95% CI: -3.41 - 70.63), and a mean difference of 6.56 was found between NSH and H (95% CI: -1.68 - 14.79). The magnitude of the effect observed for GCF levels of hBDs in healthy sites was large (eta squared= 0.497 and r= 0.70 for hBD 1; eta squared= 0.176 and r= 0.42 for hBD 2). The comparison between diseased sites of patients with periodontitis revealed significantly higher levels of hBD 2 and lower levels of hBD 1 in the GCF of smokers compared to non-smokers (p<0.05). The mean difference between SD and NSD was 74.67 (95% CI: -50.82 - 200.16) for hBD 2 and 6.89 (95% CI: 0.64 - 13.14) for hBD 1. The magnitude of the effect observed for GCF levels of hBDs in diseased sites of smokers and non-smokers was intermediate (hBD 2 eta squared= 0.073 and r= 0.27; hBD 1 eta squared= 0.081 and r= 0.28) 17 . In non-smokers, higher levels of hBD 1 and 2 were observed in diseased sites than in healthy sites (p<0.05). A mean difference of 4.34 for hBD 1 (95% CI: -6.74 - 15.41) and 118.95 for hBD 2 (95% CI: -3.14 - 241.04) was observed between NSD and NSH. The magnitude of the effect observed was intermediate for hBD 2 (eta squared= 0.097 and r= 0.31) and small for hBD 1 (eta squared= 0.021 and r= 0.14). However, in smokers, no significant difference in hBDs level was found between healthy and diseased sites (p>0.05). A mean difference of 2.13 for hBD 1 (95% CI: -2.71 - 6.96) and 10.67 for hBD 2 (95% CI: -36.24 - 57.58) was observed between SD and SH. The magnitude of the effect observed was small for hBD 2 (eta squared= 0.019 and r= 0.14) and intermediate for hBD 1 (eta squared= 0.108 and r= 0.33) .” . Study 2: 1. Materials and methods
"The study population consists of patients from the Department of Periodontology, Dr.D.Y.Patil Dental College and Hospital, Pimpri, Pune. The study protocol was designed in accordance with Helsinki declara- tion of 1975, as revised in 2013 and was approved by the university ethics committee (DYPV/EC/122/17).All participants willing to participate in our study signed an informed consent, were in good general heath, had no history of periodontal therapy, nor had they taken antibiotics/anti-inflammatory drugs within 6 months. Smokers, pregnant women, patients with underlying systemic disease were excluded from the study. Sample size estimation was based on data provided by Chen et al., 2014 for expected means of GCF hBD-2 levels as measured by ELISA in chronic periodontitis, gingivitis and healthy control subjects (chronic periodontitis = 266.91 pg/mL ± 311.81 pg/mL, patients with chronic gingivitis:487.80 ± 420.66 pg/mL and healthy controls was 99.20 ± 99.98 pg/mL).14 Using a sample size of 25 subjects per group the overall estimated effect size was determined as f = 0.52 (F-test ANOVA) at p = 0.05 and 80% power, which was in the large range (> 0.40) and thus deemed acceptable.15 Since there are no preliminary data for correlation of IL- 22 and hBD-2 in each these groups, we estimated the power of this sample to detect a significant large correlation effect (>0.5) and at n = 25, the sample had 80% power at p = 0.05 to determine a Pearson’s correlation coefficient of 0.53.” , can you produce the risk of bias according to the Agency for Healthcare Research and Quality for the study “Gingival crevicular fluid levels of Interleukin-22 (IL-22) and human β Defensin-2 (hBD-2) in periodontal health and disease : A correlative study” while i provide you with the following data from the study : 1. Materials and methods “The study population consists of patients from the Department of Periodontology, Dr.D.Y.Patil Dental College and Hospital, Pimpri, Pune. The study protocol was designed in accordance with Helsinki declara- tion of 1975, as revised in 2013 and was approved by the university ethics committee (DYPV/EC/122/17).All participants willing to parti- cipate in our study signed an informed consent, were in good general heath, had no history of periodontal therapy, nor had they taken anti- biotics/anti-inflammatory drugs within 6 months. Smokers, pregnant women, patients with underlying systemic disease were excluded from the study. Sample size estimation was based on data provided by Chen et al., 2014 for expected means of GCF hBD-2 levels as measured by ELISA in chronic periodontitis, gingivitis and healthy control subjects (chronic periodontitis = 266.91 pg/mL ± 311.81 pg/mL, patients with chronic gingivitis:487.80 ± 420.66 pg/mL and healthy controls was 99.20 ± 99.98 pg/mL).14 Using a sample size of 25 subjects per group the overall estimated effect size was determined as f = 0.52 (F-test ANOVA) at p = 0.05 and 80% power, which was in the large range (> 0.40) and thus deemed acceptable.15 Since there are no preliminary data for correlation of IL- 22 and hBD-2 in each these groups, we estimated the power of this sample to detect a significant large correlation effect (>0.5) and at n = 25, the sample had 80% power at p = 0.05 to determine a Pearson’s correlation coefficient of 0.53.”, 2. Clinical measures : “Clinical measurements made were plaque index (PI), gingival index (GI), bleeding on probing (BOP), probing depth (PD) and clinical attachment level (CAL). The measurements were taken at 6 sites per tooth and entered manually in the data sheet. Probing pocket depth and attachment level measurements were made to the nearest millimeter using an UNC 15 Probe. A total of 79 subjects were recruited in the study and were grouped according to their periodontal status. Group 1 (Healthy controls) included 27 subjects with a plaque index (PI) of < 1, gingival index (GI) of < 1, probing depth (PD) of ≤ 3 mm, and no loss of attachment. Group 2 (Gingivitis) included 25 subjects with a PI of ≥1, GI of ≥1PD ≤ 3 mm, clinical evidence of inflammation based on presence of BOP ≥ 50% of sites and no clinical attachment loss. Group 3 (Chronic periodontitis) had 27 subjects with a PI of ≥1, GI of ≥1, Probing depth (PD) ≥ 5 mm and clinical attachment level(CAL) ≥ 3 mm and presence of BOP ≥ 50% of sites with radiographic evidence of bone loss.” , 3. GCF collection :
“GCF collection was done a day after the periodontal examination. In healthy individuals, sites with PD ≤ 3 mm and no signs of inflamma- tion, in gingivitis patients, sites with bleeding on probing and in chronic periodontitis subjects, the sites with PD ≥ 5 mm were chosen for ac- quiring GCF. The area was isolated with gauze to avoid saliva con- tamination, after drying the area with air, supragingival plaque was removed with curette (Hu Friedy, USA), a calibrated volumetric mi- crocapillary pipette (Sigma Aldrich) was placed at the entrance of the gingival sulcus, gently touching the gingival margin. From each test site, 2 μl of GCF was collected. The collected GCF was then transferred to a sterile Eppendorf tube and stored at −70oc until the assay. Micropipettes with contaminants like blood or saliva were discarded.18 The GCF level of IL-22, hBD-2 was assayed using commercially available ELISA kit (E Lab Biosciences) [Catalog no: E-EL-H0106 (IL- 22), Catalog no: E-EL-H0996 (hBD-2]. The procedure was performed according to the instructions given in the kit.” ? 1. Quantification of hBD-2
“Briefly, to a human hBD-2 antibody precoated ELISA plate 100 μL of the standard or sample is added and incubated for 90 minutes at 37 °C and the free components are washed. Add 100 μL biotinylated detection antibody is added and incubated for 1 hour at 37 °C and washed. Then 100 μL of horse radish peroxide (HRP) conjugate is added and incubated for 30 minutes at 37 °C and washed. 90 μL of substrate reagent is added and incubated for 15 minutes at 37 °C followed by the addition of 50 μL stop solution. The optical density is measured with a spectrometry at a wave length of 450 nm immediately. The minimum detection limit for hBD-2 is 37.50 pg/mL. The levels of hBD-2 in each sample were determined using the concentration values of standards included in the kit contents. The results were expressed in pg/mL.”, 2. Statistical analysis
“The statistical analysis was done using the Statistical package for the Social science (SPSS) for windows, version 19; SPSS Inc. Chicago, IL, USA. All the periodontal parameters were described and reported as mean values ( ± SD). The distribution and comparison of clinical parameters and gingival crevicular fluid IL-22, hBD-2 levels in healthy controls, gingivitis and chronic periodontitis groups were assessed using one way ANOVA test. Post Hoc Bonferroni test was used for intergroup comparison of clinical parameters and gingival crevicular fluid (GCF) IL-22, hBD-2 levels in healthy controls, gingivitis and chronic periodontitis groups. Association among the IL-22 and hBD-2 and clinical parameters in all three groups were examined using Pearson’s correlation test. Probability value of less than 0.05 was considered statistically significant for all analyses.”, 3. Clinical analysis
“The healthy control group had 27 subjects with a mean age of 25.14 years and 25 gingivitis subjects with a mean age of 31.92 years and 27 patients with periodontitis with a mean age of 47.00 years. Periodontal parameters of the study groups are shown in Table 1. All clinical parameters were significantly higher in the chronic periodontitis group than the healthy and gingivitis groups (P < 0.05).”, 4. GCF hBD-2 levels
” The mean GCF levels of hBD-2(Table 1, Graph 1) and the mean difference between groups are represented (Table 2). The hBD−2 concentration was significantly higher in the chronic periodontitis group compared to gingivitis group (P = 0.003) and healthy control group (P < 0.05).However there were no statistical differences be- tween healthy and gingivitis groups in terms of total amount of hBD-2 in GCF. " , 5. Correlations "There was significant positive correlation between GCF total amount of IL-22 and GI (P < 0.01), PI (P < 0.01), PD (P < 0.00.01), CAL (P < 0.05) [Table 3, Graph 2(A, B, C, D)] in chronic periodontitis group (Group 3). Positive correlation between GCF total amount of hBD-2 and PI (P < 0.05) and CAL (P < 0.01) [Table 3, Graph 3(B, D)] was observed in chronic periodontitis group. Positive correlation was observed between IL-22 and hBD-2 levels in GCF of CP Group (P < 0.005) [Table 3, Graph 3(E)]. The correlation between clinical parameters and gingival crevicular fluid levels of hBD-2 and IL-22 in healthy controls and gingivitis group was statistically insignificant (data not shown). 3rd Study : 1. Participant selection and clinical evaluations
“In this cross-sectional study, 42 CP patients and 59 controls were recruited from either patients attending to the Department of Periodontology, Faculty of Dentistry, or from the personal patient populations of the academic faculty and postgraduate students at the Ondokuz Mayis University between June 2014 and May 2015. Ethics committee approval was received from Ondokuz Mayıs University clinical research ethics committee with decision number 2014/556-630(2013/416-447).
Individuals included in the study were informed about the purpose and content of the study and signed an informed consent form indicating they voluntarily participated in the study.”,
2. Inclusion criteria
“Individuals included were over 18 years old and willing to provide in- formed consent. In addition, they did not have any systemic diseases including cardiovascular diseases, diabetes, chronic renal failure, and hepatitis B and C, and were not using any chronic anti-inflammatory drugs. Moreover, participants who are pregnant or lactating at the time of the study or receive any periodontal treatment within the preceding six months were excluded from the study. All subjects were evaluated clinically and radiographically at the first clinical visit; periapical and panoramic radiographs were obtained from all patients to verify the clinical periodontal diagnosis and the extent of alveolar bone loss. All clinical parameters were evaluated by a single investigator (S.K). The examiner was calibrated against an expert periodontist. Calibration was validated if 90% of the recordings could be reproduced within a 1.0-mm difference (Badersten, Nilvéaus, & Egelberg, 1984). The following clinical parameters were assessed at six sites per tooth: probing pocket depth (PD); clinical attachment loss (CAL); bleeding on probing (BOP) (Ainamo & Bay, 1975); plaque index (PI) (Silness & Löe, 1964); gingi- val index (GI) (Löe & Silness, 1963); and the number of missing teeth, mobility, and furcation involvement. PD was defined as the distance between the coronal-most margin of the free gingiva to the most apical penetration of the Michigan-0 probe. CAL was defined as the distance from the cemento-enamel junction to the most apical penetration of the Michigan-O probe. The presence of supragingival plaque was recorded dichotomously during PD measurements. Bleeding on probing (BOP) within 20s was recorded dichotomously.Clinical and radiographic criteria proposed by the 1999 International World Workshop were used for diagnosis.”,
3. Control group
“Patients had no history of periodontitis, and no furcation involvement and mobility. Their PD was smaller than 3 mm.”,
4. CP group inclusion criteria
“Radiographic evidence of alveolar bone loss and at least ≥4 mm PDD in two interdental areas, not on the same tooth (Eke, Page, Wei, Thornton-Evans, & Genco, 2012).”,
5. Collection and storage of samples
“GCF samples were obtained for analyses (mainly from the maxillary premolar area for standardization). The supragingival plaque was removed without touching the gingival margin. The area was care- fully washed with water to prevent salivary contamination, the tooth was isolated with cotton rolls, and the area was dried with air for 10 s. For sampling, paper strips (Periopaper, ProFlow, Amityville, NY,USA) were carefully placed in the gingival sulcus and held there for 30 s. If the paper strips were contaminated with blood or saliva, they were discarded and replaced. The volume of each paper strip was determined with Periotron 8,000 (ProFlow, Amityville, NY, USA). The samples were placed with 50 microliters of phosphate-buffered saline (pH 7.0) in an Eppendorf tube. Two milliliters of peripheral venous blood samples were taken from each subject, using a standard venipuncture method. The sam- ples were then transferred to tubes containing 8% liquid EDTA. GCF samples were centrifuged at 1,000 × g for 10 min; blood samples were centrifuged at 3,000 × g for 10 min by a centrifuge device (Shimadzu UV160A, SNo: 28006648, Japan). The supernatants obtained from GCF and serum samples were separated for the study. The samples were mixed thoroughly with a vortex and stored at −80oC until the working day. On the working day, the samples were brought to room temperature.” .1.Determination of hBD-2 concentrations
“Serum and GCF hBD-2 concentrations were studied by the commercially available Human BioD2 ELISA Kit (Sun-Red Bio Company, Cat No. 201-12-1937, Shanghai, China), in the Ondokuz Mayıs University, Faculty of Medicine, by the Medical Biochemistry Research Laboratory. The “double-antibody sandwich” enzyme-linked immunosorbent assay (ELISA) procedure was used for this analysis. All working solutions were freshly prepared and stored at room temperature (25°C) until use.
Five standards were prepared with the serial dilution method using human hBD-2 standard (S1–6.4 ng/ml, S2–3.2 ng/ml, S3–1.6 ng/ml, S4–0.8 ng/ml, and S5–0.4 ng/ml). The wells were identified on the ELISA plate for blank, standards, and samples. Nothing was added to the blank well except Chromogen A, Chromogen B, and a stop solution. The same procedure used with the sample was applied to the standards. Fifty μL of standard (S1–S5) was pipetted into each well, and 40 μl + 10 μl of hBD-2 antibody was pipetted from each sample. Then, 50 μl Streptavidin–Horseradish peroxidase was added to the standards and samples and incubated at 370C for 60 min. After incubation, the plate was washed 5 times with 350 μl washing solution with the help of an automatic washer. 50 μl Chromogen A and 50 μl Chromogen B were added to all wells and incubated at 37°C for 10 min. Then, the reaction was stopped by pipetting 50 μl of stop solution. At the end of the study, absorbance at 450 nm wavelength was read by a microplate reader. Sample hBD-2 concentrations were calculated according to the standard curve generated using standard values, and the concentrations obtained were expressed in ng/ml. Kit sensitivity 0.05 ng/ml and assay range 0.1 ng/ml-10 ng/ml were given. High concentration samples were confirmed by working twice.”,
2. Statistical analysis
"Data analysis was performed using R program version 3.5.1. (R Core Team, 2019). Data are expressed as means and standard deviations (SD) or medians and interquartile ranges (IR) as appropriate. Data were analyzed for normal distribution by the Shapiro–Wilk test. A chi-square test was used for comparison of categorical variables. An independent- sample t test was used to detect the statistical significance between the two groups, based on the clinical variables obtained, such as age, probing depth, clinical attachment loss, and others. Univariable and multivariable logistic regressions were used to assess the association between the outcome (periodontal disease) with serum and GCF hBD-2 levels and other covariates. Correlation between each clinical parameter and serum and GCF hBD-2 levels was analyzed using the Spearman correlation coefficient. A p-value of <.05 was considered statistically significant. The estimated power of the test was 0.80.
3. Clinical findings
“A total of 101 participants, 59 controls and 42 patients with CP, were assessed. Of the volunteers included in the study, only 8 in the control group and only 5 in the CP group were smokers. Thirty- seven (37) females and 22 males (mean age 27.69 ± 7.78) were included in the control group, and 28 females and 14 males (mean age 42.67 ± 10.62) were included in the CP group. There was no statisti- cal difference between the groups in terms of smoking and gender distribution (p > .05). The distribution of demographic data is shown in Table 1.
Age was found to be significantly higher in the CP group (p < .001). The mean age of all participants was 33.93 (±11.68) years; the mean age of the control group was 27.69 (±7.78), and the mean age of the CP group was 42.67 (±10.62) (Table 1).”,
4. Clinical variables
“All recorded clinical parameters (BOP, CAL, PI, GI, and PD) were significantly higher in the CP group than in the control group (p < .001) (Table 2). CP was reclassified according to the modification of the new international classification of periodontitis (Tonetti, Greenwell, & Kornman, 2018). Accordingly, cases were classified as Stage II (n = 3; 7%); Stage III (n = 34; 81%); Stage IV (n = 5; 12%); and Grade B (n = 25; 60%) to C (n = 17; 40%) periodontitis. Of these, 67% of CP patients were generalized CP, and 33% were localized CP.The mean hBD-2 levels of GCF samples in the CP group were 2.77 (±0.74) ng/30 s and 2.51 (±0.54) ng/30 s in the control group. The GCF levels of the mean hBD-2 were significantly higher in the CP group compared with those in healthy controls (p = .047; Figure 1). Adjusting for potential confounders such as smoking, age, and gender did not alter this association (p = .036). Conversely, the mean hBD-2 levels of serum samples in the CP group were lower (2.92 ± 3.07 ng/ ml) compared with those of the control group (7.75 ± 6.37 ng/ml; p < .001; Figure 2). Similarly, adjusting for smoking status, age, and gender did not alter this association (p < .001). Additionally, we tested the difference of hBD-2 level between generalized and localized CP. The difference between the groups both in serum (p = .25) and in GCF hBD-2 levels (p = .51) was not statistically significant.”,
5. Correlations
“The correlation between serum and GCF hBD-2 expression and clinical periodontal measures was analyzed. No correlation was observed between serum and GCF hBD-2 levels (r = .004, n = 99, p = .97). In both GCF and serum samples, no correlations were ob- served between hBD-2 and the clinical parameters of periodontal disease such as PD, GI, PI, CAL, and number of teeth present. On the other hand, both localized GCF and serum hBD-2 were correlated with BOP within the periodontitis group. However, after adjustment by the Bonferroni method for multiple comparisons, a statistically significant correlation remained only between GCF hBD-2 and BOP (r = .54, p = .0002; Table 3).” .