Effect of initial periodontal therapy on gingival crevicular fluid cytokine profile in subjects with chronic periodontitis.

Cytokines are thought to play an important role in the pathogenesis of periodontal disease. Because periodontal disease is known for its inhomogeneous distribution within the dentition, it is unclear to what extent the detection of various cytokines at different sites correlates with presence of disease. We evaluated whether levels of 12 cytokines in gingival crevicular fluid (GCF) discriminated periodontally diseased sites from healthy ones, or periodontally diseased persons from healthy ones, and assessed the impact of nonsurgical periodontal therapy on these readings. This study included 20 periodontally healthy persons (H) and 24 patients with chronic periodontitis (P). In every participant, we measured the plaque index, gingival index, probing pocket depth (PD), bleeding on probing, and recession at six sites of every tooth. GCF was collected with Durapore® filter strips from two healthy sites (PD<4 mm; HH) in group H, and from two periodontally diseased sites (PD≥5 mm; PP) and two periodontally healthy sites (PD≤3 mm; PH) in group P. The periodontally diseased participants underwent comprehensive nonsurgical periodontal therapy including deep scaling and root planing under local anesthesia. In these participants, GCF samples were again collected at the same sites 1 and 3 months after therapy. Twelve cytokines (il-1ß, il-1ra, il-6, il-8, il-17, b-fgf, g-csf, gm-csf, ifn-γ, mip-1ß, vegf, and tnf-α) were assessed using the Bio-Plex suspension array system. Mean plaque index, gingival index, bleeding on probing, PD, and recession were significantly different between groups H and P. Differences between PP and PH sites were not significant for any of the cytokines. Il-1ra, il-6, il-17, b-fgf, gm-csf, mip-1ß, and tnf-α differed significantly between HH sites and both PH and PP sites, whereas il-8 was significantly higher only at PP sites. Periodontal treatment increased gm-csf and decreased il-1ra levels in PP sites. Il-1ra, il-6, il-8, il-17, b-fgf, gm-csf, mip-1ß, and tnf-α identified patients with chronic periodontitis, rather than diseased sites, suggesting a generalized inflammatory state that is not limited to clinically diseased sites only.

Systemic Biomarkers in 2-Phase Antibiotic Periodontal Treatment: A Randomized Clinical Trial.

Accumulating evidence suggests that periodontal infections may have an impact on systemic health. In patients with untreated periodontitis, very high values for several inflammatory markers in serum are expressed simultaneously. We investigated to what extent these peak values change after nonsurgical and surgical periodontal treatment, with adjunctive antibiotics administered during the first or the second treatment phase. In a single-center, randomized, placebo-controlled, and double-masked clinical trial, 80 patients with chronic or aggressive periodontitis were randomized into 2 treatment groups: group A, receiving systemic amoxicillin and metronidazole during the first, nonsurgical phase of periodontal therapy (phase 1), and group B, receiving the antibiotics during the second, surgical phase (phase 2). Serum samples were obtained at baseline (BL), 3 mo after phase 1 (M3), and 6 and 12 mo after phase 2 (M6, M12). Samples were evaluated for 15 cytokines and 9 acute-phase proteins using the Bio-Plex bead array multianalyte detection system. For each analyte, peak values were defined as greater than mean +2 SD of measurements found in 40 periodontally healthy persons. Sixty-six patients showed a peak value of at least 1 analyte at BL. At M12, the number of these patients was only 36 (P = 0.0002). This decrease was stronger in group A (BL: 35, M12: 19, P = 0.0009) than in group B (BL: 31, M12: 17, P = 0.14). Twenty patients displayed peak values of at least 4 biomarkers at BL. The nonsurgical therapy delivered in the first phase reduced most of these peaks (group A, BL: 9, M3: 4, P = 0.17; group B, BL: 11, M3: 2, P = 0.01), irrespective of adjunctive antibiotics. The reductions obtained at M3 were maintained until M12 in both groups. Initial, nonsurgical periodontal therapy reduced the incidence of peak levels of inflammatory markers. Antibiotics and further surgical therapy did not enhance the effect (Clinicaltrials.gov NCT02197260).

Anti-apolipoprotein A-1 autoantibodies as biomarker for atherosclerosis burden in patients with periodontitis.

BACKGROUND AND OBJECTIVE: Anti-apolipoprotein A-1 (anti-apoA-1) IgG is a potential marker of atherosclerotic plaque vulnerability and cardiovascular complications. In patients with periodontitis the presence of anti-apoA-1 IgGs in serum and their association with atherosclerosis is unknown. MATERIAL AND METHODS: One-hundred and thirty subjects with periodontal disease and 46 healthy subjects, matched for age and gender, participated in this study. Anti-apoA-1 IgG, high-sensitivity C-reactive protein (hsCRP) and matrix metalloproteinase (MMP) -2, -3, -8 and -9 were measured in serum samples. An ankle-brachial index (ABI) value below 1.11 served as a surrogate marker of atherosclerosis. Predictive accuracies of biomarkers for abnormal ABI were determined using receiver-operating characteristics curves and logistic regression analyses. RESULTS: Compared with healthy controls, periodontitis patients showed lower median ABI values (1.10 vs. 1.15; p < 0.0001), a higher prevalence of anti-apoA-1 IgG positivity (23.8% vs. 6.5%; p = 0.009) and higher concentrations of hsCRP (1.62 mg/L vs. 0.85 mg/L; p = 0.02) and MMP-9 (435 µg/mL vs. 283 µg/mL; p < 0.0001). In patients younger than 50 years of age (n = 66), anti-apoA-1 IgG was found to be the best predictor for an abnormal ABI (area under the curve = 0.63; p = 0.03). Anti-apoA-1 IgG positivity increased the risk of having an abnormal ABI (odds ratio = 4.20; p = 0.04), independently of diabetes, smoking and body mass index. CONCLUSIONS: Anti-apoA-1 IgG positivity and atherosclerosis, as reflected by abnormal ABI, were more prevalent in periodontitis patients than in age- and gender-matched controls. In younger periodontitis patients, anti-apoA-1 IgG was found to be the best predictor of atherosclerosis burden.

Association of chronic plaque psoriasis and severe periodontitis: a hospital based case-control study.

BACKGROUND: Both chronic plaque psoriasis and periodontitis have an increasing prevalence worldwide and have been associated with the metabolic syndrome; however limited information is available on their association. OBJECTIVE: To evaluate the possible association of severe periodontitis and chronic plaque psoriasis. METHODS: This was a hospital based case-control study. Chronic plaque psoriasis patients and age- and gender-matched controls have been recruited. Baseline demographic data have been recorded. To explore correlations between different dichotomous variables the Sperman Rho correlation coefficient was used. Correlations were further explored non-parametrically and univariate and multivariate logistic regression was utilized after adjustment for the effect of confounders. RESULTS: During the study enrollment period 100 patients with CPP and 100 age- and gender-matched controls were included in this study. Mean age for both groups was 57.2 ± 5.3 years. 43% of patients and controls were males. Significant correlations where noted between psoriasis and 1) periodontitis (rho = 0.219, P = 0.02) and 2) metabolic syndrome (rho = 0.191, P = 0.07) using Spearman's Rho correlation co-efficient. Univariate logistic regression reported significant relations between psoriasis and periodontitis (OR = 3.329, 95%CI: 1.513-7.324, P = 0.003) and psoriasis and metabolic syndrome (OR = 2.293, 95%CI: 1.250-4.207, P = 0.007). On the contrary, a non-significant relation between psoriasis and active smoking status was detected (OR = 1.041, 95%CI: 0.597-1.817, P = 0.887). In a multivariate analysis model we found a significant correlation of psoriasis and periodontitis when controlled for the presence of metabolic syndrome (OR: 2.486, 95%CI: 1.002-5.842, P = 0.049). CONCLUSION: Periodontitis may be associated with psoriasis but further studies are required to elucidate their relationship in the context of the biologic plausibility.

Clinical and laboratory study of ocular rosacea in northern Greece.

BACKGROUND: The prevalence of ophthalmic involvement in rosacea is probably higher than previously presumed and varies considerably among several studies. OBJECTIVE: This study aimed to determine the incidence of ocular disease among a population of rosacea patients in Northern Greece, to objectively determine the presence of eye dryness in rosacea patients with and without clinical ophthalmic involvement and correlate the severity of ocular disease with the severity of cutaneous rosacea. METHODS: One hundred patients with rosacea were assessed for the stage of their disease and examined for ocular symptoms and signs. In 24 of them the tear break up time (TBUT) and Schirmer test were performed in each eye, along with 24 controls. RESULTS: A total of 33 patients (33%) were positive for ophthalmic findings. The most frequent symptoms and signs were burning sensation and tearing, and conjunctivitis and blepharitis, respectively. Eleven patients with ophthalmic manifestations had mild to moderate erythematotelangiectatic rosacea, 17 had moderate papulopustular rosacea and four exhibited findings of phymatous rosacea. The total mean value of patients' Schirmer tests was significantly lower compared with the healthy controls (P < 0.0001). Mean TBUT was shorter in the rosacea group than that in the age-matched controls (P < 0.0001). CONCLUSION: Ocular involvement in rosacea is a common phenomenon with eye dryness being an early sign. Tear function tests, like Schirmer test and TBUT, although not specific, could contribute to the screening and early diagnosis of the disease, to prevent the potential development of sight-threatening conditions.

Cytokines in gingival crevicular fluid of adolescents and young adults.

BACKGROUND/AIM: The purpose of this study was to compare the levels of the cytokines interleukin-1beta (IL-1beta), IL-4, and IL-8 in the gingival crevicular fluid (GCF) of adolescents and young adults. METHODS: Twenty-five adolescents aged between 14 and 16 years (Group A) and 20 periodontally healthy young adults aged between 25 and 35 years (Group B) were selected from two private dental clinics limited to pedodontics and periodontics respectively in Piraeus Greece. All subjects were systemically healthy. Clinical examination included probing pocket depth (PPD), presence or absence of plaque, and bleeding on probing (BOP). GCF was collected from four sites per subject. IL-1beta, IL-4, and IL-8, measured as total amounts (pg/30 s), were evaluated in 180 samples using a commercially available sandwich enzyme-linked immunosorbent assay. RESULTS: IL-1beta mean levels of Groups A and B were adjusted for BOP and PPD. Differences of IL-1beta mean levels between the two age groups were statistically significant (F = 50.245, P < 0.001) in favour of Group A. Adolescents showed statistically significantly lower mean levels of IL-4 than young adults in the presence of BOP (F = 10.690, P = 0.001). There was no statistically significant difference between adolescents and adults for the means of IL-8 adjusted for BOP and plaque presence (F = 2.032, P = 0.161). CONCLUSIONS: Within the limits of this study the differences reported in mean levels of IL-1beta and IL-4 may be attributed to the different age status.

Enamel matrix derivative and systemic antibiotics as adjuncts to non-surgical periodontal treatment: biologic response.

BACKGROUND: Short-term clinical observations suggest an anti-inflammatory effect of enamel matrix derivative (EMD). The purpose of this study was to evaluate the anti-inflammatory capacity of EMD, used as an adjunct to non-surgical periodontal treatment of deep lesions in chronic periodontitis patients, by monitoring inflammatory markers in gingival crevicular fluid (GCF). METHODS: Sixteen subjects were randomly assigned to treatment with EMD or placebo in contralateral dentition areas. Half of the subjects received 250 mg metronidazole and 375 mg amoxicillin three times a day for 7 days; the other half received a placebo. GCF samples were collected from one interproximal lesion in each of the contralateral quadrants before treatment and after 10 days and 2, 6, and 12 months. Total protein content was determined according to the Bradford method. Myeloid-related protein (MRP) 8/14 and interleukin (IL)-1beta were analyzed quantitatively by enzyme-linked immunosorbent assay (ELISA), and elastase activity was determined using a low molecular weight fluorogenic substrate. RESULTS: No significant differences were observed between sites treated with or without EMD for any biochemical parameter. Two months after treatment, subjects treated with antibiotics exhibited less clinical signs of inflammation. Furthermore, these subjects had lower MRP 8/14 levels only at day 10 compared to those receiving the placebo. For total protein, IL-1beta, and elastase, no statistically significant differences were noted for subjects with or without antibiotic therapy at any time point. CONCLUSIONS: Improved healing of the soft tissues has been noted clinically in non-surgically treated sites in subjects treated with antibiotics. The expression of inflammatory mediators in GCF corroborated this finding only in part. EMD did not seem to further affect the expression of inflammatory mediators.

Enamel matrix proteins and systemic antibiotics as adjuncts to non-surgical periodontal treatment: clinical effects.

OBJECTIVES: The purpose of the present study was to evaluate the clinical effects of non-surgical periodontal treatment, supplemented with enamel matrix derivative (EMD) and/or systemic antibiotics, in deep periodontal pockets of patients with chronic periodontitis. METHODS: This was a randomized, placebo-controlled longitudinal clinical trial of 12 months duration. Using a split-mouth design, 16 subjects were randomly assigned to scaling and root planing (SRP) with EMD or placebo in contra-lateral dentition areas. One half of the subjects received 250 mg metronidazole and 375 mg amoxicillin three times a day for 7 days and the other half received a placebo. One inter-proximal periodontal lesion was chosen as study site in each of the contra-lateral quadrants. RESULTS: Subjects treated with systemic antibiotics yielded significantly better clinical results than those treated with placebo. In these cases, probing pocket depth was reduced significantly more after 6 months (3.0+/-2.1 mm versus 1.6+/-1.4 mm, p=0.05), and the mean clinical attachment gain was significantly greater after 6 months (2.3+/-1.9 mm versus 0.7+/-1.6 mm, p=0.02) and 12 months (2.3+/-3.5 mm versus 0.4+/-3.8 mm, p=0.02). Sites treated with the antibiotics plus EMD gained the largest amount of clinical attachment. There was no significant benefit of EMD adjunctive to SRP in subjects not treated with antibiotics. CONCLUSIONS: The present study supports the notion that optimal repair and regeneration of the periodontium requires suppression of the microbiota causing periodontal disease.

Effect of nicotine-treated epithelial cells on the proliferation and collagen production of gingival fibroblasts.

BACKGROUND, AIMS: Several in vitro and in vivo studies have indicated that tobacco smoking may be an important risk factor for the development and severity of inflammatory periodontal disease. METHOD: In the present study, we developed an in vitro model to study the interactions between nicotine-treated epithelial cells (EC) and gingival fibroblasts (GF) derived from the same patient. EC were treated with nicotine concentrations varying from 1 microg/ml to 500 microg/ml and their effect on different functions of GF was studied. The proliferation of GF was evaluated by the incorporation of 3H-thymidine. A dose-dependent inhibition was observed with nicotine concentrations > or =100 microg/ml. Similar results were observed when studying the total protein synthesis of GF by incorporation of 3H-proline into non-dialyzable material. RESULTS: When collagen production was evaluated by 3H-proline incorporation into collagenase-sensitive protein, a dose-dependent reduction was observed: the degree of inhibition varied from 25% with 50 microg/ml nicotine, to almost 60% with 500 microg/ml. Interestingly, the production of non-collagenous proteins decreased by almost 50% only when EC were treated with the highest concentration of nicotine. CONCLUSIONS: The results suggest that epithelial cells, acting as mechanical barrier, can reduce but not completely eliminate the deleterious effect of nicotine on gingival fibroblasts.

Effects of nicotine on periodontal ligament fibroblasts in vitro.

Cigarette smoking is associated with increased incidence of periodontal disease and poor response to therapy. In the present study, we examined the effects of nicotine on several functions of periodontal ligament fibroblasts (PDLF): proliferation, attachment, alkaline phosphatase production and chemotaxis. Nicotine concentrations varying from 5 ng/ml to 250 microg/ml were tested. Proliferation of cells was studied by the incorporation of 3H-thymidine, and a dose-dependent inhibition was observed with concentrations > or =100 ng/ml. Similar results were observed when studying the attachment of the cells on plastic surfaces, using a colorimetric method. The inhibition of attachment was even more evident after 6 h incubation of the cells with nicotine. The activity of alkaline phosphatase, as determined with the substrate p-nitrophenyl phosphate, in both conditioned medium (CM) and cellular extract (CE), was also significantly decreased in a concentration-related fashion. Finally, the chemotaxis of PDLE as examined by a modification of the Boyden's blind-well chamber technique, was inhibited in a dose-dependent manner. The degree of inhibition varied from 15% with the lowest concentration of nicotine (50 ng/ml), to almost 90% with the highest (5 microg/ml). The results show that nicotine can have direct adverse effects on various functions of the periodontal cells.

A longitudinal study of various crevicular fluid components as markers of periodontal disease activity.

In order to examine the relationship of possible crevicular biochemical parameters to attachment loss (ALOSS), 330 sites from 8 untreated adult patients were monitored longitudinally at 3-month intervals, for up to 1 year. Attachment levels were measured with a force-sensing probe and an acrylic stent in duplicates at each study point. Crevicular samples were collected and used for the determination of the following 11 markers: number of polymorphonuclear leukocytes (PMNs), prostaglandin E2 (PGE2), osteocalcin (OC), alkaline phosphatase (ALP), collagenase (COL), beta-glucuronidase (BG), antigenic and functional elastase (AEL and FEL), alpha-1 antitrypsin (a1AT), alpha-2 macroglobulin (a2M) and aspartate aminotransferase (AST). 10 sites with ALOSS of > or = 1.5 mm per 3 months (active sites) and 43 sites with negligible changes (inactive sites) were identified. Total amounts of ALP, BG and COL were found to be significantly higher in active as compared to inactive sites, prior to significant ALOSS, without any significant differences in crevicular fluid volume and clinical indices. When biochemical parameters were expressed as ratios to the number of PMNs, PGE2/ PMNs was significantly elevated in active sites. The capacity of such individual parameters to distinguish between active and inactive sites was limited. However, linear discriminant analysis using total amounts of PGE2, COL, ALP, a2M, OC and AEL showed more significant diagnostic values (sensitivity: 80%, specificity: 91%). These findings suggest that the combination of several biochemical parameters in crevicular fluid could give more information to predict future clinical ALOSS.

Functional characteristics of gingival and periodontal ligament fibroblasts.

In periodontal surgery, healing after guided tissue regeneration (GTR) may be explained by differences in functional activities of gingival and periodontal ligament fibroblasts (GF and PDLF). Several studies in vitro have supported this hypothesis, but much remains to be defined. In the present work, gingival and periodontal ligament fibroblasts derived from five healthy subjects were isolated and compared in vitro. The morphology of the cells was observed under scanning electron microscopy (SEM). Several extracellular matrix components (ECM) were studied to compare the effects on fibroblast attachment, proliferation, and protein synthesis. Several biochemical markers were examined in both cellular extract (CE) and conditioned medium (CM). We also examined the muscle differentiation markers alpha-smooth muscle actin, desmin, and smooth-muscle myosin. Finally, we studied the effects of epithelial cells on the proliferation and protein synthesis of the two types of fibroblasts. GF and PDLF appeared identical under the SEM. All ECM components enhanced attachment; however, while collagen types I and IV promoted the attachment of GF, gelatin, laminin, and vitronectin promoted that of PDLF. Most ECM components increased the proliferation rate of GF and the biosynthetic activity of PDLF. The biochemical markers were similarly distributed between the two cell types, except for alkaline phosphatase, which was detected only in the CE of PDLF. Both GF and PDLF strongly expressed alpha-smooth-muscle actin and were negative for desmin; only PDLF were positive for smooth-muscle myosin. Epithelial cells increased the proliferation of both GF and PDLF but had no effect on their biosynthetic activity. These in vitro results may better explain the in vivo functional differences between GF and PDLF.

Elastase release from gingival crevicular and peripheral neutrophils in periodontitis and health.

The activity of elastase increases significantly in the gingival sulcus during inflammation. The release of this enzyme from crevicular and peripheral polymorphonuclear leucocytes (PMNs) was studied in seven patients with gingivitis, six with adult periodontitis, seven with rapidly progressive periodontitis and in nine healthy subjects. Peripheral PMNs were isolated from blood and crevicular PMNs from gingival washings. After preincubation with cytochalasin B, the same numbers of crevicular and peripheral cells were incubated either in phosphate-buffered saline (spontaneous release) or in the same buffer containing formyl methionyl-leucyl-phenyl-alanine (stimulated release). Elastase activity was measured in the supernatant by a fluorimetric technique. The results confirm that compared to peripheral PMNs, crevicular cells show a higher spontaneous release of elastase and a lower stimulated release. The activity of elastase released either spontaneously or after stimulation of crevicular cells was significantly lower in the controls as compared with cells obtained from patients with gingivitis or periodontitis. Crevicular PMNs collected from patients with deeper pockets appear to release larger amounts of elastase than those from controls.

Neutrophil elastase and its inhibitors in human gingival crevicular fluid during experimental gingivitis.

The relative concentrations and absolute amounts of neutrophil elastase and its two inhibitors, alpha 2-macroglobulin (alpha 2-M) and alpha 1-antitrypsin (alpha 1-AT), were determined in gingival crevicular fluid (GCF) collected from six dental students who refrained from brushing the upper left or right quadrant during three weeks. Plaque and gingival indices and flow of GCF were measured before, during, and after the three weeks of no brushing. Functional elastase, representing the enzyme complexed with alpha 2-M, was measured by use of a low-molecular-weight fluorogenic substrate. Elastolytic activity in GCF was also assayed by use of elastin as substrate. Antigenic elastase, representing the enzyme complexed with alpha 1-AT, as well as the inhibitors alpha 2-M and alpha 1-AT were measured by ELISA. After three weeks of plaque accumulation, the concentrations of both functional and antigenic elastase increased by a factor of about 3, whereas the concentrations of the inhibitors increased in a much higher proportion. No free elastase could be detected in GCF, as evidenced by the Sephadex G-75 elution profile of GCF, by the negative results obtained when elastin was used as substrate, and by the demonstration that pure enzyme kept its activity against the low-molecular-weight substrate after being saturated by alpha 2-M.

Reversible CT findings of the liver after radiotherapy.

We report the computed tomographic (CT) findings in a case of radiation therapy of the abdomen which affected the left lobe of the liver. The post-radiotherapy CT demonstrated a low density area involving the left lobe of the liver. One month later in a new CT this finding disappeared. We conclude that low density areas in the liver parenchyma following radiotherapy should be analysed with caution, especially when secondary deposits are considered in the differential diagnosis.

Synthesis of alpha 2-macroglobulin in human gingiva: a study of the concentration of macroglobulin and albumin in gingival fluid and serum.

Albumin and alpha 2-macroglobulin were analysed in gingival fluid collected by filter paper strips from 6 volunteers before, during and after 10 days of dental plaque accumulation. The two proteins were also determined in samples of venous blood at the beginning of abstention from tooth brushing and 10 days later. A nephelometric technique was used for the analysis of albumin, but alpha 2-macroglobulin was determined by ELISA. The plaque and gingival indices, as well as the flow of gingival fluid, increased significantly during the period of plaque accumulation and then returned to baseline values. The concentrations of the two proteins in blood remained within normal limits. In gingival fluid, however, the concentration of albumin increased about two-fold at the peak of inflammation, while that of alpha 2-macroglobulin increased as much as 6 times. The enhanced passage of alpha 2-macroglobulin into the gingival sulcus in comparison to the entirely plasma-derived albumin was consistent with the concept that the greater excretion of globulin during inflammation must be due to an increased local synthesis.