Alterations in crevicular fluid flow during healing following gingivectomy and flap procedures

Gingival fluid was collected from the mesial aspects of the anterior teeth of twelve patients before initial preparation and two weeks after completion of this. All pockets from which fluid was collected were measured prior to initial preparation. On twelve occasions pockets were eliminated by conventional gingivectomy and on eight flaps were reflected followed by curettage and osteoplasties. Gingival fluid was collected immediately prior to operation and at two, three and four weeks post-operatively. A strong correlation (r =+0.93: P < 0.001) was found between pocket depth and the amount of the gingival fluid exudate. Initial preparation reduced the amount of fluid flow, the extent of the reduction being significant (P < 0.01) where pockets were deep. No significant change in the amount of fluid flow had occurred two weeks after either type of operation but there were significant reductions (P < 0.01) between the third and fourth post-operative weeks when flap procedures had been performed and in the fourth post-operative week in the case of gingivectomies. The level of gingival fluid flow was the same in both groups four weeks after the operations even though it was twice as great in the flap procedures group as in the gingivectomy group before operation.     

Changes in soluble adhesion molecules in gingival crevicular fluid following periodontal surgery

BACKGROUND: Inflammation of periodontal tissues during postoperative wound healing is mediated by cell surface adhesion molecules. Soluble forms of these antigens have also been identified and shown to be important in immunoregulatory processes, but have previously not been investigated during periodontal repair and regeneration. The present study has examined the presence and possible changes in soluble intercellular adhesion molecule-1 (sICAM-1; CD54) and lymphocyte function-associated antigen-3 (sLFA-3; CD58) in gingival crevical fluid (GCF) following periodontal surgery. METHODS: GCF samples were collected from four groups: 1) a guided tissue regeneration (GTR) test; 2) a GTR control, at least one complete tooth unit away from the periodontal defect; 3) a conventional flap (CF) surgery; and 4) a crown lengthening (CL). Sandwich enzyme-linked immunosorbent assay (ELISA) was used to measure the levels of sICAM-1 and sLFA-3 in the GCF samples. RESULTS: A marked increase in GCF volumes was found in all sites after surgery, although a persistent increase was associated only with the period of membrane retention at the GTR test sites. In addition, sICAM-1 and sLFA-3 were found in the GCF of healthy as well as diseased sites prior to treatment and the total amounts of both increased transiently following surgical intervention, especially sLFA-3. However, the concentrations of these GCF components, particularly sICAM-1, tended to decrease. CONCLUSIONS: The temporal decrease in the concentration of sICAM-1 and sLFA-3 in GCF may serve to enhance inflammatory reactions at surgically-treated periodontal sites, thereby limiting repair and regeneration in the periodontium. These soluble adhesion molecules may thereby be of potential therapeutic value and might also be useful markers for monitoring periodontal wound healing.     

Changes in transforming growth factor-beta1 in gingival crevicular fluid following periodontal surgery

OBJECTIVES: Growth factors play a major part in wound healing, including in the periodontium. However, the presence of growth factors in gingival crevicular fluid (GCF) in humans during periodontal wound healing has not yet been determined. Our hypothesis is that such factors are present in GCF and that changes in their levels might be of value as a prognostic marker of wound-healing activity and therapeutic progress following periodontal surgery. The aim of this study was therefore to measure transforming growth factor-beta1 (TGF-beta1) in GCF collected from sites that have undergone guided tissue regeneration (GTR) and conventional flap (CF) surgery and to compare these with GCF collected from unaffected healthy sites. MATERIALS AND METHODS: GCF samples were collected, using filter paper strips, at baseline (pre-surgical) and then at intervals up to 26 weeks from 16 patients undergoing GTR and from 11 patients undergoing CF surgery. After elution and acid treatment, TGF-beta1 levels were measured by ELISA. RESULTS: Treatment of periodontal defect sites significantly reduced the mean probing pocket depth (PPD) and improved the mean lifetime cumulative attachment loss (LCAL). Average GCF volumes also significantly increased at all sites at 2 weeks post-surgery and thereafter declined to baseline levels, except at the GTR test sites that were still elevated at 7 weeks. TGF-beta1 could be detected in almost all GCF samples, and 2 weeks after surgery, the average levels increased two-fold at the surgically treated but not at the control sites, which remained unchanged. CONCLUSION: TGF-beta1 is readily detectable in GCF and increases transiently following periodontal surgery. This suggests that changes in the levels of this growth factor in GCF might be useful for monitoring the progress of periodontal repair and regeneration.     

Smoking cessation increases gingival blood flow and gingival crevicular fluid

OBJECTIVES: The purpose of the present study was to determine the effect of smoking cessation on gingival blood flow (GBF) and gingival crevicular fluid (GCF). MATERIAL AND METHODS: Sixteen male smokers (aged 22-39 (25.3+/-4.0) years), with no clinical signs of periodontal and systemic diseases, were recruited. The experiment was performed before (baseline) and at 1, 3 and 5 days, and at 1, 2, 4 and 8 weeks after smoking cessation. The status of smoking and smoking cessation was verified by exhaled carbon monoxide (CO) concentration, and by serum nicotine and cotinine concentrations. A laser Doppler flowmeter was used to record relative blood flow continuously, on three gingival sites of the left maxillary central incisor (mid-labial aspect of the gingival margin and bilateral interdental papillae). The GCF was collected at the mesio- and disto-labial aspects of the left maxillary central incisor and the volume was calculated by the Periotron 6000(R) system. The same measurements except for the GBF were performed on 11 non-smoking controls (four females and seven males), aged 23-27 (24.4+/-1.2) years. RESULTS: Eleven of 16 smokers successfully completed smoking cessation for 8 weeks. At 1 day after smoking cessation, there was a significantly lower CO concentration than at baseline (p<0.01). Also, nicotine and cotinine concentrations markedly decreased at the second measurement. The GBF rate of smokers was significantly higher at 3 days after smoking cessation compared to the baseline (p<0.01). While the GCF volume was significantly increased at 5 days after smoking cessation compared to the baseline (p<0.01), it was significantly lower than that of non-smokers until 2 weeks after smoking cessation (p<0.01). CONCLUSION: The results show that the gingival microcirculation recovers to normal in the early stages of smoking cessation, which could activate the gingival tissues metabolism/remodeling, and contribute to periodontal health.     

Osteopontin in gingival crevicular fluid

Osteopontin (OPN) is a major glycosylated phosphoprotein in bone matrix and is produced by several cells including osteoblasts, osteoclasts and macrophages. OPN levels increase in active sites of bone metabolism. Recently, several bone-related proteins were identified in gingival crevicular fluid (GCF) to seek markers of alveolar bone resorption in periodontal disease. In this study, we investigated the existence of OPN in GCF and the correlation between OPN level in GCF and probing depth (PD) of sampling sites in 98 periodontitis patients and 35 healthy subjects. An immunoblotting analysis using 10% polyacrylamide gel showed that two forms of OPN with molecular masses of 54 and 66 kDa and several degraded fragments were detected in most GCF samples from diseased sites (PD > 4 mm). In GCF samples from healthy sites (PD < or = 3 mm), only one form (54 kDa) was observed, but any degraded fragments were not detected. When OPN amounts in GCF samples were determined by ELISA, a weak. but significant correlation was observed between OPN amount in GCF and PD (r=0.32, p=0.0013). These results demonstrate that OPN exists in GCF and that OPN level in GCF increases with the progression of periodontal disease.     

Dentine phosphoproteins in gingival crevicular fluid during root resorption

External apical root resorption is a common, yet unexplained, phenomenon associated with orthodontic treatment. Available methods of clinical evaluation are radiographic. Biochemical assays offer the advantage of being non-invasive, as well as being diagnostic and potentially prognostic. The hypotheses are firstly that during the process of root resorption, organic matrix proteins are released into the gingival crevicular fluid (GCF) and, secondly, that there is a difference in the levels of these proteins between a group of patients with mild root resorption and a control group. GCF was collected from the permanent central incisors of untreated subjects (controls, n = 20), primary second molars with half of the root resorbed (primary group and positive controls, n = 20) and permanent central incisors with mild root resorption in patients undergoing active orthodontic treatment (orthodontic group, n = 20). Dentine phosphoproteins (DPP) were measured in the GCF using an enzyme-linked immunosorbent assay developed with DPP isolated from human first premolars and an antibody against rat incisor DPP. The primary group showed the highest levels of DPP in the GCF compared with the orthodontic (P = 0.296) and control (P = 0.001) groups. The orthodontic group showed elevated levels relative to the control group (P = 0.046). It is concluded that root resorption can be studied using a biochemical immunoassay and that this method can provide quantitative measurement of DPP in GCF.     

Granulocyte elastase activity in static and flow gingival crevicular fluid

OBJECTIVES: This study aimed to evaluate the volume of gingival crevicular fluid (GCF) and granulocyte elastase activity in static GCF (sGCF) and flow GCF (fGCF) from subjects with various periodontal conditions. METHODS: Eleven periodontally healthy, 10 gingivitis and 12 periodontitis subjects were recruited and the sites investigated consisted of healthy sites from healthy subjects (HH); healthy (HG) and gingivitis sites (GG) from gingivitis subjects; and healthy (HP), gingivitis (GP) and periodontitis sites (PP) from periodontitis subjects. fGCF samples were collected either 1 min or 5 min following sGCF collection by paper strip technique. GCF volume was determined by Periotron 6000 and granulocyte elastase activity was assayed with a specific substrate [l-pyroglutamyl-l-prolyl-l-valine-p-nitroanilide(pGluProVal-pNA)]. RESULTS: At baseline, no significant differences existed in clinical and GCF parameters between the two matched sites for subsequent collection of fGCF samples either 1 min or 5 min after sGCF sampling in all subjects. The flow exudate in HG and HP sites quickly replenished to sGCF levels, while a delayed replenishment was found in HH sites, despite the similar sGCF volumes of these sites. The GCF volume and elastase levels in the fGCF at 1 min were higher in GP sites than in GG sites (P < 0.05). Overall, depletion of elastase levels in the fGCF at 1 min was observed in all subjects, whereas elastase levels in the fGCF at 5 min had replenished to sGCF levels in HP, GP, PP sites and GG sites, but had remained at a lower level in HH and HG sites. An overall positive correlation was found between sGCF and fGCF for GCF volume and elastase activity (P < 0.001); however, this correlation varied with GCF parameters and with site conditions of the subjects concerned. CONCLUSIONS: This study shows that patterns of dynamic changes in GCF flow and elastase activity varied under different periodontal conditions. Assessment of both sGCF and fGCF may allow better insight into the dynamic change of the target components in GCF.     

Glycosaminoglycans in human gingival crevicular fluid during orthodontic movement

Glycosaminoglycans (GAG) in gingival crevicular fluid (GCF) were investigated by cellulose acetate electrophoresis of simultaneously collected samples from the mesial and distal surfaces of teeth in 3 groups of young persons. In a control group, which had not undergone orthodontic treatment, a major band of hyaluronic acid (HA) and a minor band of chondroitin sulphate (CS) were present. No differences in the mean content of either GAG between the mesial and distal surfaces were detected. From teeth undergoing movement by fixed appliances (active group), a raised mean level of CS was present in GCF from the surface towards which movement was directed. Teeth held passively by an appliance following cessation of active movement (retention group) showed raised levels of CS at mesial and distal surfaces. A heparan sulphate-like GAG was commonly present in this group only. No significant increase in the levels of HA were detected at the mesial and distal surfaces of either the active or the retention groups, despite increased GCF flow rates unassociated with more severe gingival inflammation. The GAG composition of GCF, particularly CS, appears to reflect changes occurring in the deeper periodontal tissues of alveolar bone and periodontal ligament during orthodontic treatment.     

Granulocyte elastase in gingival crevicular fluid

The granulocyte elastase activity and the immuno-reactive (antigenic) granulocyte elastase of gingival crevicular fluid (GCF) were studied in 16 periodontitis patients and in 10 gingivitis patients. The elastase activity was measured with a low molecular weight substrate specific for granulocyte elastase. The antigenic elastase was determined with specific antibodies against granulocyte elastase. Intracrevicular sampling of GCF with paper strips for 30 s seemed to provide representative values of elastase. The elastase activity correlated with probing depth and attachment loss and appeared to be a measure of the degree of tissue destruction. Antigenic elastase represents the number of granulocytes in GCF and should thus be related to the degree of inflammation. The periodontitis patients and the gingivitis patients both had a similar degree of inflammation as measured by antigenic elastase per microliter GCF and gingival index. The elastase activity per microliter GCF, however, was higher in the periodontitis group. Elevated granulocyte elastase activity in GCF seems to be independent of inflammation and could thus be an indicator of patients at risk for periodontitis.     

Acute-phase proteins in gingival crevicular fluid during experimentally induced gingivitis

The dynamics of four acute-phase proteins were investigated in gingival crevicular fluid (GCF) during the course of a 21 day experimental gingivitis study. These acute-phase proteins were the protease inhibitors α2-macroglobulin (α2-M) and α1-antitrypsin (α1-AT) and the iron-binding proteins transferrin (TF) and lactoferrin (LF). 6 healthy volunteers ceased all oral hygiene procedures for 3 weeks. GCF was sampled at seven day intervals from two sites per subject by paper strips for 30 s during the experimental gingivitis period and for two additional weeks after the reinstitution of oral hygiene. The mainly serum derived α2-M, αl-AT and TF exhibited very similar dynamics which reflects their common origin in GCF. Their levels increased significantly from baseline and remained high for at least one week after the reinstitution of oral hygiene measures (repeated measures MANOVA; α2-M: p=0.015; α1-AT: p=0.012; TF: p=0.02). This probably reflects increased vascular permeability in the gingivae and, to a lesser degree, local production by gingival inflammatory cells. In contrast to the serum derived acute-phase proteins, the neutrophil derived LF rose significantly from baseline (repeated measures MANOVA; p=0.001) but dropped rapidly after the reinstitution of oral hygiene measures. This could be because dental plaque was removed and thus neutrophil chemotactic agents in the crevice were decreased.     

Alkaline phosphatase activity in gingival crevicular fluid during canine retraction

OBJECTIVES: The aim of the study was to investigate alkaline phosphatase activity in the gingival crevicular fluid (GCF) during orthodontic tooth movement in humans. SETTING AND SAMPLE POPULATION: Postgraduate orthodontic clinic. Ten female patients requiring all first premolar extractions were selected and treated with standard edgewise mechanotherapy. EXPERIMENTAL VARIABLE: Canine retraction was done using 100 g sentalloy springs. Maxillary canine on one side acted as experimental site while the contralateral canine acted as control. OUTCOME MEASURE: Gingival crevicular fluid was collected from mesial and distal of canines before initiation of canine retraction (baseline), immediately after initiation of retraction, and on 1st, 7th, 14th and 21st day and the alkaline phosphatase activity was estimated. RESULTS: The results show significant (p < 0.05) changes in alkaline phosphatase activity on the 7th, 14th and 21st day on both mesial and distal aspects of the compared experimental and control sides. The peak in enzyme activity occurred on the 14th day of initiation of retraction followed by a significant fall in activity especially on the mesial aspect. CONCLUSIONS: The study showed that alkaline phosphatase activity could be successfully estimated in the GCF using calorimetric estimation assay kits. The enzyme activity showed variation according to the amount of tooth movement.     

Protease activity in gingival crevicular fluid

Abstract. Our aim was to study protease activity in GCF from inflamed sites with or without tissue destruction. 19 patients with both periodontitis and gingivitis sites and 12 patients having gingivitis alone participated in the study. GCF samples were collected by an intracrevicular washing method. The protease activity was measured as degradation of FITC-conjugated casein. To obtain a semiquantitative estimate of the harvested GCF volume, we measured the transferrin concentration in the wash-fluid. The protease activity was significantly higher in the deep pockets in periodontitis patients than in shallow pockets in the same patients. This difference was still higher when the ratio of protease activity to the amount of transferrin in the sample was plotted. Although protease activity was lower in samples from gingivitis patients than in the deep pockets in periodontitis patients, the difference was not significant. About 90% of the activity could be inhibited by the addition of an excess amount of α-1-antitrypsin (A1AT). This study shows that protease activity is higher in inflamed sites with tissue destruction than in inflamed sites without. Most of this activity could be inhibited by A1AT, which suggests that the activity is due to an imbalance between protease and antiprotease rather than to proteases insensitive to A1AT.     

Arylsulphatase activity in human gingival crevicular fluid

Arylsulphatase activity in fluid collected from non-inflamed (n = 5), gingivitis (n = 5) and periodontitis (n = 5) subjects was assayed. The mean volume activity for each group was: non-in-flamed = 768 ± 165 nm/ml per h; gingivitis = 2431 ± 1118 nm/ml per h; periodontitis = 2860 ± 1839 nm/ml per h. The mean total unit activity for each group was: non-inflamed = 0.326 ± 0.076 nm; gingivitis = 1.394 ± 0.411 nm; and periodontitis = 3.571 ± 1.700 nm. Analysis of fluid from isolated sites in periodontitis suggests that reporting total unit activity (absolute amount of enzyme activity) is more meaningful than reporting volume activity (enzyme concentration) for arylsulphatase activity.     

Lactate dehydrogenase activity in gingival crevicular fluid during orthodontic treatment.

During orthodontic treatment, the early response of periodontal tissues to mechanical stress involves several metabolic changes that allow tooth movement. Many studies have evaluated such modifications by analysis of various host metabolites released into the gingival crevicular fluid (GCF). This study used a cross-sectional design to examine the lactate dehydrogenase (LDH) activity in GCF to assess whether GCF LDH can be proposed as a sensitive marker for periodontal tissue modifications during orthodontic tooth movement. Thirty-seven subjects, 16 males and 21 females (mean age 18.7 years, range 14.0 to 26.7 years), participated in this study. Each subject underwent a session of professional oral hygiene and received oral hygiene instructions; 2 weeks later, a fixed orthodontic appliance was placed on the maxillary arch. A randomly selected maxillary canine was considered as the test tooth, and its antagonist, which had no appliance, was used as the control tooth. From 2 to 12 weeks after orthodontic appliance placement, GCF was harvested from both experimental teeth at the mesiobuccal angle, for GCF volume and LDH activity determinations. Clinical monitoring consisted of recording supragingival plaque presence, bleeding on probing, and probing depth at the same collection sites. The results showed that no differences in clinical conditions and GCF volume occurred between the experimental teeth. On the contrary, GCF LDH activity in the test teeth was significantly greater than that of the control teeth (P <.01). Moreover, no differences were found in the enzymatic activity between the sexes by experimental tooth, and no significant correlation was present between GCF LDH activity and patients' ages within experimental teeth. Our enzymatic results initially indicated a possible role of GCF LDH during the early phases of orthodontic treatment and therefore warrant further study as a possible diagnostic tool for tissue response during orthodontic treatment.     

The periotron gingival crevicular fluid meter

The present study was undertaken to compare the calibration curve of the periotron* (HAR-6000) with those of two earlier models (HAR-600) instrument, which was found to be appreciably more sensitive in detecting small volumes of fluid than the two older models, is curvilinear in shae while the latter two models display linear calibration characteristics Calibration curves were also obtained for one of the HAR-600 models with the periopaper strip inserted up to and beyond the manufactures recommendations. The results obtained in this case differed from those published by others. Using fluids possessing different dielectric dissipation constants, calibration curves were determined for the HAR-6000 model over a wide range of volumes, The calibration data obtained best fitted quadratic functions and the relative positions of the curves on the axes are in the correct relationship to their dielectric constants Inter-instrument and other variations have once more stressed the importance of individual instrument calibration by uers.     

Comparison of gingival health and gingival crevicular fluid flow in children with and without diabetes.

GCF flow measurements and a clinical scoring of gingival health were both recorded for a group of 56 children with diabetes and 41 children without diabets. The children with diabetes had significantly more gingival disease than the children without diabetes when compared with either measure. A small but significant correlation was found between the GCF flow and the clinical scores with the children with diabetes but not with the children without diabetes.     

Hydroxyproline determination in serum and gingival crevicular fluid

A high-performance liquid chromatographic assay for total hydroxyproline in serum and gingival crevicular fluid has been developed. The Clq component of the first complement factor was precipitated out of the samples with a 0.02 M sodium acetate solution. Aliquots of the supernatants were dried, vapor hydrolyzed with 6 M hydrochloric acid at 105°C for 24 hours and derivatized with phenylisothiocyanate. The derivatives were chromatographed using a multilinear solvent gradient with detection at 254 nm. The regression of peak areas on the concentrations of hydroxyproline was linear with a correlation coefficient of 0.99 within the range of assayed quantities (2.5 to 25 ng). Analytical recovery was 90.5 ± 2.54% (± SD) and the determination level was 2.5 ng (19.1 pmol). The precision of the assay as determined by the coefficient of variation for five consecutive runs of six concentrations of hydroxyproline in serum hydrolysate ("within-run" precision) and five series run on different days ("between-run" precision) was 1.8 ± 1.28% and 4.2 ± 2.59% (± SD), respectively. Hydroxyproline concentrations in GCF from single sites during developing experimental gingivitis in the beagle dog showed an irregular pattern of low and high concentrations ranging from 5.2 to 17.4 ng/μl.