42. Diagnosis / Indices-Gingival Fluid & Saliva

42. Diagnosis / Indices-Gingival Fluid & Saliva                                  

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  1. Gingival Crevicular Fluid - Overview
  2. Association of GCF with health or disease
  3. The composition of gingival fluid
  4. GCF and the susceptibility or resistance to periodontal disease
  5. antibiotics in the gingival crevicular fluid
  6. periodontitis disease markers in GCF

Gingival Crevicular Fluid - Overview

Is the measurement of gingival fluid easy, repeatable, and dependable?

1. Griffiths, G. Formation, collection and significance of gingival crevice fluid. Periodontl 2000. 2003; 31: 32-42

2. Goodson, J. Gingival crevice fluid flow. Periodontol 2000. 2003; 31:43-54

3. Delima, A, Van Dyke, T. Origin and function of the cellualr componenents in gingival crevice fluid. Peridontol 2000. 2003; 31: 55-76

4. Uitto V., Overall, C, McCulloch, C. Proteolytic host cell enzymes in gingvial crevice fluid. Periodontol 2000. 2003; 31:77-104

5. Eley, G., Cox, S. Proteolytic an hydrolytic enzymes from putative periodotal pathogens: characterization, molecular genetics, effects on host defecses and tissues and detection in gingival crevice fluid. Periodotol 2000. 2003; 31:105-24

Gingival Crevicular Fluid

Discuss the composition of gingival fluid.   What is the diagnostic significance of increased amounts of gingival fluid?

6. Orban, J, Stallard, R: Gingival crevicular fluid: A reliable predictor of gingvial health: J Periododontol 40:231-235, 1969

7. Hancock, E et al: The relationship between gingival crevicular fluid and gingival inflammation. A Clinical and histologic study. J Periodontol 50: 13-19, 1979

8. Offenbacher, S et al: The use of crevicular fluid prostaglandidn E2 levels as a predictor of periodontal attachment loss. J Periodont Res 21:101, 1986

9. Lamster, I et al: Development of a risk profile for periodontal disease: Microbial and host response factors. J Periodontol 65:511-520, 1994

10. Giannopoulou, C., Kamma J., Mombelli, A. Effect of inflammation, smoking and stress on gingival crevicular fluid cytokine level. J Clinical Periodontol 2003; 30: 145-153

11. Safkan-Sppparla, B., Sorsa, T et al: Collagenases in gingival crevicular fluid in type I diabetes mellitus. J Clin Periodontol 2006; 77:189-194

12. Bostanci N. et al. Ginvial crevicular fluidlevels of RANKL and OPG in periodontal diseases: Implications of the relative ratio. J Clin Periodontol 2007; 34:370-376

Passage of Antibiotics

Which antibiotics are concentrated in the gingival crevicular fluid?

13. Gordon JM, Walker CB, Murphy JC, Goodson JM, Socransky SS. Concentration of tetracycline in human gingival fluid after single doses. J Clin Periodontol 8:117-121,1981.

14. Gordon JM, et al. Tetracycline: Levels achievable and in vitro effect on subgingival organisms. Part I. Concentrations in crevicular fluid after repeated doses. J. Periodontol. 52:609, 1981.

15. Pascale D, et al: Concentration of doxycycline in human gingival fluid. J. Clin. Periodontol. 13: 841- , 1986

16. Ciancio S, Mather M, McCullen J : An evaluation of minocycline in patients with periodontal disease. J. Periodontol. 51:530, 1980.

17. Britt MR, Pohlod DJ. Serum and crevicular fluid concentrations after a single dose of metronidazole. J. Periodontol. 57:104-107, 1986.

18. Conway TB, Beck FM, Walters JD. Gingival fluid ciprofloxacin levels at healthy and inflammed human periodontal sites. J Periodontol 71:1448-1452, 2000.

Indicators of Disease Activity

Can gingival fluid contents be used to determine severity of periodontal conditions or disease activity?

19. Leibur E, Tuhkanen A, Pintson U, Soder P-O. Prostaglandin E2 levels in blood plasma and in crevicular fluid of advanced periodontitis patients before and after surgical therapy. Oral Diseases 5:223-228, 1999.

20. Lamster IB, Oshrain RL, Harper DS, et al : Enzyme activity in crevicular fluid for detection and prediction of clinical attachment loss in patients with chronic adult periodontitis. Six month results. J. Periodontol. 59: 516-523, 1988.

21. Bader HI, Boyd RL. Long-term monitoring of adult periodontitis patients in supportive therapy: Correlation of gingival crevicular fluid proteases with probing attachment loss. J Clin Perio 26:99-105,1999.

22. Lamster IB, Ahlo JK. Analysis of gingival crevicular fluid as applied to the diagnosis of oral and systemic diseases. Ann N Y Acad Sci. 1098:216-29, 2007. Review

Saliva

How does saliva play a role in a patient's susceptibility or resistance to periodontal disease ?

23. Giannobile WV. Salivary diagnostics for periodontal diseases.J Am Dent Assoc. 2012 Oct;143(10 Suppl):6S-11S. Review.

25. Guentsch A, Preshaw PM, Bremer-Streck S, Klinger G, Glockmann E, Sigusch BW. Lipid peroxidation and antioxidant activity in saliva of periodontitis patients: effect of smoking and periodontal treatment. Clin Oral Investig. 12(4):345-52, 2008. Epub 2008 May 29.

26. Reher VG, Zenóbio EG, Costa FO, Reher P, Soares RV. Nitric oxide levels in saliva increase with severity of chronic periodontitis. J Oral Sci. 49(4):271-6. 2007

27. Frodge BD, Ebersole JL, Kryscio RJ, Thomas MV, Miller CS. Bone remodeling biomarkers of periodontal disease in saliva. J Periodontol. 79(10):1913-9, 2008.

28. Tobón-Arroyave SI, Jaramillo-González PE, Isaza-Guzmán DM. Correlation between salivary IL-1beta levels and periodontal clinical status. Arch Oral Biol. 53(4):346-52, 2008.

29. Rai B, Kharb S, Jain R, Anand SC. Biomarkers of periodontitis in oral fluids. J Oral Sci. 50(1):53-6, 2008

30. Miller CS, King CP Jr, Langub MC, Kryscio RJ, Thomas MV. Salivary biomarkers of existing periodontal disease: a cross-sectional study. J Am Dent Assoc. 137(3):322-9, 2006.

Gingival Crevicular Fluid - Overview

Is the measurement of gingival fluid easy, repeatable, and dependable?

Topic: Gingival crevicular fluid (GCF)

Authors: Griffiths, G.                                                            ARTICLE

Title: collection and significance of gingival crevice fluid.

Source: Periodontl 2000. 2003; 31: 32-42
Type: Discussion

Rating: Good

Keywords: gingival crevice fluid

Background:  The exact nature of GCF, its origins and composition has been subject of controversy.  The principle questions that remain unanswered are whether GCF is a transudate or an exudate, and whether GCF can form in a clinically healthy site.  The answers to these questions could yield therapeutic approaches.  An increase in GCF flow may have physical protective effects through flushing the pocket, as well as facilitating the passage of immunoglobulins.

Purpose:  To discuss the formation, collection, and significance of GCF.


Discussion: How is GCF formed?

Is GCF a transudate of interstitial fluid?

Controversy exists whereas GCF is a transudate or inflammatory exudate.

Collection Methods:

Several techniques have been employed and the technique chosen will depend upon the objectives of the study as each technique has advantages and disadvantages. Three basic categories:

  1. Gingival washing: the gingival crevice is perfused by isotonic solution. The fluid collected represents a dilution of crevicular fluid and contain both cells and soluble constituents such as plasma proteins. This technique is valuable for harvesting cells from the gingival crevice region. Major disadvantage is that all fluid may not be recovered and accurate quantification of GCF volume or composition is not possible.

  2. Capillary tubing and micropipettes: following isolation and drying of the site capillary tubes are inserted into the entrance of the gingival crevice. GCF from the crevice migrates into the tube and because the internal diameter is known the volume of the fluid collected can be accurately determined by measuring the distance which the GCF has migrated. However, it is difficult to collect an adequate volume of GCF in a short period unless the sites are inflamed and contain large quantities of GCF. It is difficult to conceive that holding a capillary tube at the entrance to the gingival crevice for long time ensures an atraumatic collection. Also, it is difficult to remove the complete sample from the tubing.

  3. Absorbent filter paper strips: quick and easy to use. Can be applied to individual sites and is possibly the least traumatic.

Methods of collection can be broadly divided into intracrevicular and extracrevicular techniques. The former depends on the strip being inserted into the gingival crevice, whereas in the latter the strips are overlaid on the gingival crevice region in an attempt to minimize trauma. The intracrevicular method is the method used most frequently and can be further subdivided depending upon whether the strip is inserted just at the entrance of the crevice or periodontal pocket or whether the strip is inserted to the base of the pocket or until minimum resistance is felt.

(a) extracrevicular method; (b) intracrevicular method ‘superficial’ (c) intracrevicular method ‘deep’

Association of GCF with health or disease: The association of an increased volume of GCF with an increase in the severity of inflammation is well supported by evidence from literature.

Topic: Gingival crevicular fluid

Authors: Goodson, J                                                           ARTICLE

Title: Gingival crevice fluid flow

Source: J. Periodontol 2000. 2003; 31:43-54

Type: Discussion article

Rating: Good

Keywords: Gingival fluid


Purpose: To focus attention on the importance of GCF flow and the effect that small streams of fluid flowing out of the periodontal pocket have on the periodontal environment.

Discussion:

GCF flow (or flow rate): is the process of fluid moving into and out of the gingival crevice or pocket.

Fluid flow: is a rate measure of the volume that crosses a defined boundary over a given time.

Resting volume is the amount of fluid within a given space.

3 Methods of Measurement are available to differentiate fluid flow from the resting GCF volume.

Method 1: Samples are collected (w/ filter paper strips) rapidly under strict timing protocol so that the resting volume is removed by the first sample and the volume of the newly formed GCF can be measured in subsequent samples. The key to this measurement is to select a sampling time that is small enough so that the resting volume of the pocket is not allowed to re-establish. Ideally, one would remove the entire contents of the pocket with the first sample so that all subsequent samples would be repetitive estimates of the GCF volume flow over the sampling time.

Method 2 : Collect samples (w/ filter paper strips) after equilibrium has been re-established in the pocket over several different time intervals. In this case, each GCF sample includes both the resting volume (Vr) and the volume of GCF that entered the pocket over the sampling period lends itself directly to linear regression analysis in which the volume and time of each sample contribute to the analysis to determine the slope which is the GCF influx (fi) and the intercept which is the resting volume of the pocket (Vr).

Method 3: Measure the equilibrium concentration of a marker substance pumped into a pocket at a constant rate. By pumping a marker substance (ie Tetracycline) into a periodontal pocket at a constant rate, an equilibrium concentration will be established which is the result of the fluid flow rate and the pump delivery rate. To date, no published study has been conducted to evaluate GCF flow in this manner.

Lamster et al :

Described changes in GCF sample volumes taken during experimental gingivitis that are directly amenable to analysis by method 1. In the GCF sampling protocol of this study, a filter paper strip was placed in the sulcus for 30s and removed for volume determination (V1). Thirty seconds was allowed to elapse and the GCF volume was determined by introducing a second filter paper strip into the site for 3s (V2). The results of this study clearly indicated that the two sample volumes collected increased linearly over the period of development of experimental gingivitis. The GCF flow increases as inflammation becomes more severe and vascular permeability increases.

One site from each of 56 systemically healthy subjects with periodontal disease was assigned to each clinical category (Health, gingivitis, mod Periodontitis and advanced periodontitis). The data indicate that broadly defined categories of health and disease can be distinguished by differences in GCF flow.

Conclusion: Given the evidence that change in GCF flow may be a sensitive measure of local inflammation, this type of evaluation could increase the diagnostic potential of this measure being used chair-side. GCF flow measurement could provide added benefit in establishing a diagnosis and monitoring the response to therapy.

Topic: Gingival crevicular fluid

Authors: Delima, A, Van Dyke, T.                                                           ARTICLE

Title: Origin and function of the cellualr componenents in gingival crevice fluid.

Source: Peridontol 2000. 2003; 31: 55-76

Type: Discussion/Review

Rating: Good

Keywords: gingival crevicular fluid


 

Purpose: To review the origin and function of the cellular components in gingival crevicular fluid


Discussion: Periodontitis is initiated by bacterial colonization, followed by proliferation, and extension of the plaque into the subgingival environment. While diseases of the periodontium are bacterial in origin, the extent and severity of the disease depend upon the interaction between pathologic microbes and host response. The outflow of GCF helps to cleanse the dentogingival space of non-adherent microbes and to reduce the concentrations of toxins and byproducts. GCF contains microbes and antimicrobial compounds and enzymes directed at them. We often think about the flushing mechanism of GCF, but it also serves as an entry point to bacteria. One of the initial responses of the host to bacterial plaque is an increase in the vascular permeability of the subepithelial blood vessels. This leads to edema in the gingival crevice (via the JE) and results in increased crevicular fluid flow. Since this exudate is essentially a growth medium that supports host cells and tissues, the GCF can also act as an excellent source of nutrients for subgingival microbes and may actually contain factors that are necessary for the proliferation of some pathologic bacterial species.

The GCF is rich in cellular elements and also delivers antibodies and the complement system to fight the plaque front. The epithelia of the sulcus are constantly renewing and this rapid turnover appears to aid in the clearance of bacteria that adhere to these cells. One major set of cells found here are the PMN’s – these are the major cellular defense system in the gingival crevice. These cells leave the connective tissue and migrate through the JE into the crevicular space where they accumulate at the interface of the subgingival plaque. PMN’s compromise about 90% of the cells in the GCF, and their defense mechanism includes phagocytosis and release of bactericidal enzymes. It also has been shown that the release of granules from PMN’s that are capable of disengaging bacterial plaque adherence to the tooth. Other cells present in the GCF include monocytes and macrophages. These are usually seen against the oral epithelium, and they function in the cellular defense and acquired immunity of the periodontal tissues.

Conclusion: The nature and extent of bacterial challenge is modulated and attenuated by the host immune response. Inflammatory and immune responses can also contribute to the destruction of host tissues. The narrow balance between periodontal homeostasis and disease depends upon a qualitatively and quantitatively appropriate response of the host defense mechanism to infection of the periodontal tissue. The outflow of GCF helps to cleanse the dentogingival space of non-adherent microbes and to reduce the concentrations of toxins and metabolic byproducts. Constant cellular turnover and the presence of PMNs help to aid in the clearance and protection of the sulcus. Understanding the components of GCF will help to clarify the initial events in the pathogenesis of periodontal disease and aid in the monitoring of the disease process.

Cell Type

Source

Function

Bacteria

Adjacent plaque mass

Etiologic factors of periodontal disease. Initiates the host immune response.

Epithelial cells

Oral sulcular and junctional epithelium

Represents the high turnover rate of the epithelium that comprise the gingival sulcus.

Leukocytes

Gingival plexus of blood vessels

Effector cells of host response. PMNs play a role in innate immunity. Monocytes/macrophages and lymphocytes play roles in cell-mediated immunity.

Erythrocytes

Blood vessels

Incidental finding. Results from damage to the small blood vessels and capillaries of gingival connective tissue.

Topic: GCF

Authors: Uitto V., Overall, C, McCulloch, C                                                           ARTICLE

Title: Proteolytic host cell enzymes in gingvial crevice fluid.

Source: Periodontol 2000. 2003; 31:77-104

Type: Discussion

Rating: Good

Keywords: GCF


Purpose: This is a review of literature on findings on host cell-derived enzymes that relate to periodontal disease processes. Special emphasis is placed on MMPs. The potential use of tissue-derived GCF in enzymes in clinical periodontology is also discussed.

Discussion: Neutrophils form the first line of defense and are attracted to infected tissues by chemoattractants released from bacteria, host cells or degraded tissue. Over 90% of leukocytes in GCF are neutrophils.

They contain vesicles where molecules used for host defense are stored. These granules are generally classified into azurophilic (primary), specific (secondary) and gelatinase (tertiary) types. Gelatinase granules are released first, then specific and lastly the azurophilic. The major MMPs in neutrophils are MMP-8 and -9.

The serine proteinases (neutrophil elastase, cathepsin G and proteinase 3) are secreted from the azurophilic granules during phagocytosis, stimulation and cell lysis. Stimulation by LPS, TNF-α and IL-8 results in increased (20-fold) binding of these proteinases with cell membranes. Their actions is inhibited by the serpins. When there is increased population of neutrophils there is increased concentration of active forms of the enzymes in the tissues, causing degradations of extracellular matrix. Elastase and cathepsin G are capable of activating epithelial cells to produce IL-8, IL-6 and prostaglandin E2, which further increase chemotaxis, immune cell proliferation, and tissue degradation in inflamed tissues.

Epithelial cell proteinases: Although epithelial cells were considered as relatively passive cells whose function was to protect body surfaces, it now clear that they strongly respond to exogenous factors and therefore exhibit different morphotypes. When activated they behave aggressively by migrating, proliferating and producing various cytokines and proteolytic enzymes. They were found to produce collagenases (MMP-13, produced by the basal cells of pocket epithelium) when stimulated in vitro by TNF-α, TGF-β or keratinocyte growth factor. MMPs -2, -9, -7 have also been found to being produced by epithelial cells.

Fibroblast proteinases: Fibroblasts are responsible for the turnover of connective tissue in normally functioning tissues and they have therefore the capacity to degrade all the components of CT. The major MMPs produced are 1, 2, 13, 3, 14. It appears that MMP- 1 is secreted in the crevicular fluid of patients with localized juvenile periodontitis, while neutrophil type collagenase MMP-8 is prevalent in adult periodontitis. Expression of MMPs can be regulated by the protein composition of the extracellular matrix and following ligation of integrin receptors.

Matrix metalloproteinases: MMPs form the most important family of proteinases that participate in the normal turnover of periodontal tissues as well as their degradative aspects during periodontal diseases. As MMPs can potentially destroy tissues, their activity is strictly controlled at different levels. First, specific inhibition of MMPs can be mediated by the four members of the tissue inhibitor of metalloproteinase (TIMP) family, proteins that regulate the extracellular activity of MMPs. Second, MMPs are synthesized as latent zymogens. Activation of MMP zymogens is a critical step for regulating MMP activity and hence the composition, structure, and function of periodontal connective tissue matrices. Third, most MMPs are secreted from cells as a soluble proform. For some soluble MMPs, activation occurs at the cell surface following proteolytic cleavage by MT-MMPs, often in a TIMP dependent pathway. For other MMPs, activation occurs in the extracellular environment in an activation cascade initiated by tissue proteinases, such as plasmin, kallikrein, and tryptase, a process that is often amplified by the activated MMPs functioning as pro-MMP activators.

TIMPs: Inhibition of MMPs by TIMPs is largely interchangeable, except for the MT-MMPs, which are not inhibited by TIMP-1. TIMP-2 also plays a paradoxical role in mediating MMP-2 activation.

Detection methods for GCF MMPs: collagenases (MMP-1, -8, 13), gelatinases (MMP-2, -9), and stromelysin (MMP-3) have all been measured in GCF. Immunological assays, including ELISA and immunoblots, offer highly sensitive and specific testing methods. Immunoblots are time consuming, ELISA using antibodies is accurate and significant numbers of anti-MMPs exist on the market. Although these methods of detection are highly specific, they might not be able to distinguish between active and inactive MMPs, since MMPs can still bind the substrate without cleavage. Immunohistochemical examinations can help identify MMPs but without actually quantifying them. Another possible but untried method would be neo-epitope antibodies that can detect degradation fragments from matrix proteins or MMPs, thus help identify, quantify, and determine activity.

Substrate degradation: measure primarily the ability of MMPs to degrade the substrate, estimating enzyme activity depending on the relative abundance of the substrate degradation products by dot assays. sodium dodecyl sulfate–polyacrylamide gel electrophoresis, radioactivity, zymography, or fluorescence. The rationale for examining enzyme activity compared to the presence or abundance of MMPs (measured typically by immunochemical methods) is that in periodontitis the temporal progression of lesions is more strongly associated with the presence of active MMPs than with the total amount of enzyme. Collagen degradation products: measuring collagen metabolites in GCF can help and more directly quantify MMPs activity

GCF enzymes as indicators of periodontal health: it has been shown that PMNs enzymes detected in the GCF reflect the number of leukocytes rather than tissue destructions, so directing our attention to MMP-13 for example rather than MMP-8 (collagenase-2) appears more rational, since MMP-13 (collagenase-3) can also be produced by fibroblasts and pocket epithelial cells during tissue destruction. Neutrophil elastase, and B-Glucuronidase have been extensively studied as well, both are involved in tissue destruction. A simple mouth rinse assay has determined that total crevicular fluid elastase levels were indicated that there was a good correlation between the oral elastase activity and the number of deep pockets and the average CPITN scores. Longitudinal studies have shown the risk ratio of clinical attachment loss in patients with high B-glucuronidase activity in GCF is about 10-fold, this relation is also high when considering progressive sites vs. non progressive. Another test that has been developed is detection of aspartate aminotransferase activity GCF, this test yielded an OR of 12 for attachment loss.

Topic: Host defense

Author: Eley, G., Cox, S.                                                            ARTICLE

Title: Proteolytic and hydrolytic enzymes from putative periodontal pathogens: characterization, molecular genetics, effects on host defenses and tissues and detection in gingival crevice fluid.

Source: Periodontology 2000. 2003; 31:105-24

Type: Discussion article

Rating: Good

Keywords: proteolytic, hydrolytic, enzymatic reactions, host defense, pathogenicity


 

Purpose: Discussion article that describes the characteristics of proteolytic and hydrolytic enzymes produced by various bacteria, (Pg, Pi, Aa, Fn, Treponema, Eikenella corrodens, and capnocytophaga).

Discussion:

BL: GCF arg-gingipain appears to be an excellent predictor and GCF dipeptidylpeptidase is a moderately good predictor of future progressive CAL loss. A chair-side system has been developed to detect these bacterial proteases.


 

Gingival Crevicular Fluid

Discuss the composition of gingival fluid.

What is the diagnostic significance of increased amounts of gingival fluid?

Topic: Gingival crevicular fluid

Authors: Orban, J, Stallard, R                                                            ARTICLE

Title: Gingival crevicular fluid: A reliable predictor of gingvial health

Source: J Periododontol 40:231-235, 1969

Type: Clinical study

Rating: Good

Keywords: gingival crevicular fluid, hyaluronidase, biopsy


 

Purpose:  To evaluate the accuracy of GCF as an indicator of gingival inflammation.

Method: Dental school patients were chosen at random and evaluated by using Ramfjord's criteria for Perio Dx Index (PDI) & Greene & Vermillion's Oral Hygiene Index-Simplified (OHI-S).  GCF measurements were made using filter strips 1mm wide and several mm long, placed intracrevicularly for 3 minutes, removed and allowed to dry. The area that absorbed crevicular fluid was stained by 2% ninhydrin solution. The area stained was then measured and mean scores were calculated.  Calculus was scored for OHI-S and plaque for Ramfjord's Index using disclosing tablets.  Biopsies taken from distal half of each area evaluated. Each biopsy was evaluated according to amount and extent of inflammatory infiltrate and scored on a 0-10 scale.

Results:  From the data obtained it was apparent that GCF scores were not directly related to biopsy scores. Thus, crevicular fluid did not prove to be an accurate predictor of gingival inflammation. Plaque scores revealed a high degree of correlation on both an individual and overall evaluation when compared to biopsy scores of inflamed gingiva.

Discussion: Enzymes commonly found in plaque, primarily hyaluronidase, cause an increase in crevicular fluid flow without altering the inflammatory condition of the gingiva. Apparently, the intercellular cementing substance is modified, resulting in an increased permeability of the crevicular epithelium. Other factors including chewing, brushing, gingival massage, circadian rhythm and hormones all affect crevicular fluid flow without necessarily altering the inflammatory condition of the gingival tissues. However, it is a measure of the intactness and permeability of the gingival tissues due to previously mentioned factors. Further, as a single measurement on a patient, crevicular fluid recordings may have little significant value. Consecutive measurements, however, are of value in relating the response of the gingival tissues to environmental, physiologic and pathologic conditions and changes.

Conclusion: Dental plaque is a better indicator of the inflammatory status of the gingival tissues than GCF levels.


 

Topic: gingival fluid

Authors: Hancock EB, Cray RJ, O'Leary TJ.                                                           ARTICLE

Title: The relationship between gingival crevicular fluid and gingival inflammation. A clinical and histologic study.

Source: J Periodontol. 1979 Jan;50(1):13-9. DOI: 10.1902/jop.1979.50.1.13

Type: clinical

Rating: good

Keywords: GCF, gingival crevicular fluid, gingival inflammation,

Background: Past research reported that the flow of GCF begins a few days before other clinical signs of inflammation are apparent. Therefore, it is suggested that its flow might be measured to evaluate gingival inflammation.

Purpose: To evaluate the relationship between GI, GCF flow, and histology in order to see if GCF could be an indicator of disease severity in form of gingival inflammation.


Methods: 60 patients (26M, 34F, 18-72 years old); considered themselves in good health without known systemic diseases. No periodontal therapy within the last 90 days. Teeth #5-#12 were examined in each patient and evaluated for selection by the following criteria:

1) Presence of an adequate zone of gingiva, 2) No cervical or interproximal carious lesions. 3) No defective interproximal restorations. 4) No facial cervical restorations. 5) No acute gingivitis or periodontitis condition.

Data was collected on 57 sites; amount of GCF flow, gingival health status, GI, histologic evaluation, and histologic evaluation. After isolation with cotton rolls, GCF collected from mid-facial with filter paper in the opening of the gingival crevice for 3 min. A second sample was taken to verify fluid flow rate. Gingiva at these areas was evaluated for inflammation. GI assessed using modified GI by Löe. One tooth from each patient had gingival biopsy, trying to take equal numbers of healthy and inflamed tissues. Histology looked for inflammatory cell density. Histologic analysis looked at extent of inflammatory infiltrate using a gravimetric method

Results: results show that GCF flow tended to increase as the degree of inflammation became more severe. The quantity of GCF, by itself, was a poor indicator of the severity of gingival inflammation. The highest correlation was seen between GI and the gingival status (mainly BOP based). A high correlation was seen between clinical and histologic scores, but a very weak correlation was seen between GCF and either of these factors.

Conclusion: GCF was weakly correlated with GI and histologic evidence of inflammation; it did, however, tend to increase when inflammation increased.

Topic: Indices

Author: Offenbacher S.                                                           ARTICLE

Title: The use of Crevicular fluid prostaglandin E2 levels as a predictor of periodontal attachment loss

Source: J Periodont Res 21:101, 1986

Type: Longitudinal study

Rating: Good

Keywords: Indices; prostaglandin E2; periodontal disease

Purpose: To report results of a three-year longitudinal study, which demonstrate that crevicular fluid PGE levels can be used to reliably indicate ongoing tissue destruction, and may also be used to predict future acute periodontal attachment loss.


Methods: 7 healthy patients, 41 adult periodontitis patients and 12 juvenile periodontitis patients were assessed. All patients had a negative history of major systemic illness. All patients had at least 20 remaining teeth. Data was obtained over a period of 18 months up to 3 years for analysis. CF fluid was collected and clinical measurements (redness, edema, suppuration, BOP, pain on probing, Attach loss) were obtained at baseline and repeated one week later. SRP was performed on all diseased patients. CF collection and clinical measurements were repeated one month following initial therapy. Patients were placed on a three-month recall and CF collection was repeated at each maintenance. When a site demonstrated a statistically significant loss of periodontal attachment, after the data collection (CF and clinical parameters), local curettage and root planning was performed. One month after therapy data was again collected, and then patients were exited from the study and referred to periodontist for additional therapy. During the visit at which AL was identified, 6 consecutive CF samples were obtained at attachment loss site and 6 CF samples at the contralateral control site. Mean CF-PGE level was determined.


Results: The mean CF-PGE2 of the healthy individuals (26.9 ng/mL) was significantly lower than the one of the adult periodontitis patients (56.6± 5.4 ng/mL) and the juvenile periodontitis patients (139.4±15.3 ng/mL). The CF- PGE2 was also elevated at the time of detectable AL comparing to the cross-sectional adult periodontitis MCF-PGE.

It was observed that CF-PGE levels were increased prior to the development of the acute lesion (immediately preceding the AL episode). There was a high degree of overall and specific agreement between high CF-PGE2 levels and attachment loss. The elevated levels of CF-PGE are highly specific (0.94) and sensitive (0.76) of ongoing AL.

Conclusion: PGE levels in GCF reliably indicate ongoing disease activity and could predict upcoming attachment loss.


 

Topic: risk factors

Authors: Lamster I, et al                                                           ARTICLE

Title: Development of a risk profile for periodontal disease: Microbial and host response factors

Source: J Periodontol 65:511-520, 1994

Type: report

Rating: Good

Keywords: periodontitis/microbiology; periodontal diseases/microbiology; risk factors, host response, gingival crevicular fluid, glucuronidase

Purpose: To list risk factors associated with the host-microbial interaction in periodontal disease

Conclusion: 4 factors and their association with active chronic periodontal disease were explored in this review: Elevated GCF levels of β-glucuronidase (a lysosomal hydrolase that can serve as a marker for primary granule release from PMNs), elevated levels of periodontal pathogens, reduced levels of IgA in GCF, and reduced levels of serum IgG antibody to putative periodontal pathogens. These pathogen associated risk factors, as well as patient- associated risk factors (DM, smoking), can provide diagnostic information to aid in the treatment of patients with periodontal disease.


Saliva

How does saliva play a role in a patient's susceptibility or resistance to periodontal disease ?

Topic: Smoking on gingival crevicular fluid

Authors: Giannopoulou, C., Kamma J., Mombelli                                                           ARTICLE

Title: A. Effect of inflammation, smoking and stress on gingival crevicular fluid cytokine level.

Source: J Clinical Periodontol 2003; 30: 145-153
Type: Clinical study

Rating: Good

Keywords: smoking, interleukins, gingival crevicular fluid

Purpose: To determine the levels of interleukin (IL)-1β, IL-4, IL-6 and IL-8 in gingival crevicular fluid of periodontally healthy and diseased individuals and to study their association to environmental factors such as smoking and stress.

Material and method:

Results: In subjects with periodontitis, the total amounts of IL-1β, IL-6 and IL-8 were significantly elevated compared to healthy subjects.  IL-4 showed an inverse relationship to periodontal status and higher amounts were found in the healthy group. The amounts of all four cytokines were positively correlated with probing depths. IL-4, IL-6 and IL-8 were significantly correlated to smoking while stress was associated with IL-1β, IL-6 and IL-8 levels.

Conclusion: IL-1β, IL-6 and IL-8 reflect the activity of periodontal destruction, whereas IL-4 shows an inverse correlation to it.

Topic: Gingival Crevicular Fluid

Authors: Safkan-Sppparla, B., Sorsa, T                                                            ARTICLE

Title: Collagenases in gingival crevicular fluid in type I diabetes mellitus.

Source: Implant Dent. 2008 Mar;17(1):16-23

Type: Clinical study

Reviewer: Laura Porras

Rating: Good

Keywords:

Purpose: To analyze GCF collagenases of poorly and well-controlled type 1 DM and chronic periodontitis subjects compared to non-diabetic with chronic periodontitis and periodontally healthy control subjects

Method: Collagenase activity was studied in human GCF. 22 pts with chronic periodontitis, poorly controlled diabetes (with history of problems related to the control of their diabetes, such as hypo-, or hyperglycemias, recurrent infections, ketoacidosis, diabetic coma, glucosuria, nephropathies, neuropathies - average HbA1c 9.6%) and 5 pts with chronic periodontitis, well controlled diabetic pts (average HbA1c 8.4%) were compared to 6 chronic periodontitis, non-diabetic pts and 5 periodontally healthy, non-diabetic controls. PI, bleeding index, loss of attachment, PD, bone loss (approximal loss of marginal alveolar bone was measured from panoramic radiographs. The distance from the cemento-enamel junction to the alveolar bone crest was measured mesially and distally on all teeth to the nearest millimeter using a transparent scale. A mean score based on these measurements was calculated for each individual.were evaluated).

GCF was collected. Sites (two to five sites per subject) were cleaned and dried gently and supragingivally with sterile curettes and kept dry with cotton wool rolls. Visible plaque was carefully removed. One to two filter paper strips were placed at the opening of the gingival margin 0 to 1 mm into the sulcus for 3 minutes for gingival crevicular fluid collection. Strips were placed into Eppendorf tubes. Collagenase activity against type 1 collagen was measured using gel electrophoresis.

Results: Poorly controlled diabetic patients had more alveolar bone loss (3.7 mm) than the well-controlled diabetic patients (2.4 mm) and controls.

Conclusion: Poorly controlled diabetes is strongly related to periodontal tissue destruction, and collagenases in GCF may mediate and reflect this effect.

Critique: HbA1c of 8.4% considered as controlled diabetics. Used PANOs for bone loss measurement.

Topic: RANKL and OPG

Authors: Bostanci N et al                                                           ARTICLE

Title: Ginvial crevicular fluidlevels of RANKL and OPG in periodontal diseases: Implications of the relative ratio.

Source: J Clin Periodontol 2007; 34:370-376

Type: Retrospective study

Rating: Good

Keywords: bone resorption; ELISA; gingival crevicular fluid; OPG; RANKL

Background: RANKL is expressed predominantly as a membrane-bound ligand on osteoblasts, fibroblasts and activated T and B cells, and its osteoclastogenic action can be blocked by its soluble decoy receptor OPG.

Purpose: To investigate RANKL and OPG levels, as well as their relative ratio in GCF of patients with gingivitis, chronic periodontitis, generalized aggressive periodontitis, and chronic periodontitis under immunosuppressive treatment, as well as healthy subjects.

Method: 107 subjects: 21 healthy group, 22 gingivitis group, 26 G-AgP (GAP), 28 chronic periodontitis group, and 11 patients who were taking immunosuppressive drugs for renal transplants (cyclosporin-A) that had significant gingival enlargement. GCF samples were collected from mesio-buccal single-rooted teeth with PD 6-8 mm, in the gingivitis group the GCF was taken from teeth with 3 mm or less PD with BOP, in healthy GCF was taken from areas of no BOP. Commercially available ELISA kits used.

Result: Gingivitis and healthy groups showed low levels of RANKL, some patients had no detectable RANKL. In all periodontitis patients RANKL was always detected, no significant difference between G-AgP and chronic, but the immunosuppressed group had lower RANKL levels than either periodontitis forms (2x lower). OPG was detected in all patients at all sites, highest in healthy patients, lowest in chronic and G-AgP patients, whereas it was mildly lowered in gingivitis patients. RANKL/OPG was lowest in health and gingivitis, and elevated in all three diseased groups, with no SSD between them.

Conclusion: GCF concentrations of RANKL and OPG may be indicative of disease occurrence.

Passage of Antibiotics

Which antibiotics are concentrated in the gingival crevicular fluid?

Topic: Passage of Antibiotics

Authors: Gordon JM, Walker CB, Murphy JC, Goodson JM, Socransky SS                                                           ARTICLE

Title: Concentration of tetracycline in human gingival fluid after single doses.

Source: J Clin Periodontol 8:117-121,1981

Type: Clinical

Rating: Good

Keywords: gingival fluid, antibiotics, tetracycline

Purpose: To measure the concentration of tetracycline (TTC) in the GCF following oral administration of single doses.

Materials and methods: 6 volunteers were administered single doses of 250-500mg of TTC. GCF was sampled every 15 minutes for the first 2 hours, every 30 mins for the next 2 hours and at 5, 6, and 7 hours. Blood was sampled at 0, 3.5 and 7 hours. A second group of 4 volunteers were administered a single-dose of 250 or 500mg TTC and GCF was sampled every hour for 24 hours. Sample sites were the mesial sites of the first molars. Blood samples were obtained every hour for the first 6 hours and at 9,12,16,20 and 24 hours. Samples were taken with filter paper strips and GCF was measured with GCF meter. The concentration of TTC was determined by comparing the inhibition zones on the paper strips with standard strips containing known concentrations of TTC.

Results: The concentration of TTC in GCF after a single oral dose of 250 or 500mg peaked at 3.5-7 hours achieving average peak levels of 5-12µm/ml. Blood levels peaked at 3-4 hours and had concentrations of 1.0-2.6 µm/ml. After a brief lag period TTC persisted at higher levels in the GCF than in blood for at least 19 hours after administration however by 24 hours was rarely detectable. GCF levels varied from between the individuals and between each site in the same individual.

Conclusion: TTC in GCF was typically 2-10 times the amount in blood levels after a single dose. The presented investigation opens alternative means for the clinician and research worker to optimize therapeutic benefit derived from a given antimicrobial agent. Rational selection of the antibiotic clearly depends on knowledge of levels typically attained in the periodontal pocket and the intrinsic antibiotic susceptibility of pathogens present in the individual patient.

Topic: Host defense

Author: Gordon JM, et al                                                           ARTICLE

Title: Tetracycline: Levels achievable and in vitro effect on subgingival organisms. Part I. Concentrations in crevicular fluid after repeated doses.

Source: J. Periodontol. 52:609, 1981

Type: Clinical trial

Rating: Good

Keywords: gingival crevicular fluid, tetracycline, antibiotic, biomodulation

Purpose: To measure the concentration of tetracycline in gingival fluid following the oral administration of multiple doses of 250 mg.

Materials and methods:

Results:

Discussion:

BL: After repeated doses of TCN, the GCF is typically 2-4 times blood levels.


 

Topic: Gingival crevicular fluid

Authors: Pascale D et al                                                           ARTICLE

Title: Concentration of doxycycline in human gingival fluid.

Source: J. Clin. Periodontol. 13: 841, 1986

Type: Clinical study

Rating: Good

Keywords: doxycycline, tetracycline, antibiotics, gingival fluid


Background: The recognition that periodontal disease is caused by specific microrganisms has led to increased interest in the use of antibiotics as adjunctive treatment. To be effective in the treatment of periodontal disease, an antibiotic must penetrate to the site of infection. Tetracycline hydrochloride and minocycline have been shown to have gingival fluid levels in excess of blood levels. Doxycycline is a synthetic tetracycline compound with advantages over tetracycline hydrochloride, increased oral absorption, prolonged serum half-life and decreased GI side effects.

Purpose:  To measure the concentration of doxycycline in gingival fluid and blood after oral administration.

Methods: 4 volunteers were given doses of 100 mg doxycycline q 12 h on 1st day of antibiotic regimen, then a maintenance dose of 100 mg per day for an additional 4 days. 3 of these volunteers were also given 250 mg tetracycline hydrochloride q 6 h for 5 days either 1 month before or after doxycycline to compare gingival fluid levels of these 2 tetracycline compounds. Gingival fluid was sampled from 4 gingival sites (MB sites of 1st molars) in each volunteer at hours 0 - 6 then 9, 24, 27, 48 - 54, 57, 72, 75, 96 - 102 and 105. Blood was sampled (finger puncture) at hours 0, 3, 6, 24, 48, 54, 72, 96 and 102. Gingival fluid levels from the 4 sites (mesiobuccal surface of first molars) were averaged for each time interval. Antibiotic levels in GCF and blood were measured using an agar diffusion assay method.

Results: Gingival fluid levels of doxycycline were higher than blood levels at all times sampled and were significantly higher at 48, 72, 96 and 102 hrs. Overall, tetracycline achieved slightly higher gingival fluid levels than doxycycline. The difference was not stastistically significant. There was also no SSD between blood levels of the two antibiotics.  Doxycycline levels in gingival fluid ranged between 1.2 µg/ml and 8.1 µg /ml in the first 24 h and generally achieved 3-10 µg /ml after 48 h. Blood levels after 48 h ranged between 2.1 µg /ml and 2.9 µg /ml. Tetracycline levels in gingival fluid after 48 h were generally in the range of 4 µg /ml-10 micrograms/ml with blood levels between 2.2 µg /ml and 3.4 µg /ml.

Conclusion: Doxycycline will achieve higher levels in the GCF than in the blood. The levels detected in gingival fluid for doxycycline were comparable to tetracycline hydrochloride gingival fluid levels. Due to diminished side effects and efficacious levels in GCF, doxycycline may be favorably considered as an alternative to tetracycline in periodontal therapy.

Topic: gingival fluid and saliva

Authors: Ciancio SG, Mather ML, McMullen JA.                                                           ARTICLE

Title: An evaluation of minocycline in patients with periodontal disease.

Source: J Periodontol. 1980 Sep;51(9):530-4. DOI: 10.1902/jop.1980.51.9.530

Type: clinical

Rating: good

Keywords: gingival crevicular fluid, minocycline, saliva, drug concentration in saliva and GCF,

Purpose: To determine the passage into and concentration of minocycline in gingival crevicular fluid (GCF) and the relationship between its concentration in saliva, GCF, serum and changes in periodontal health.

Methods: 20 adults with gingivitis and/or periodontitis were included. Over an 8-day period, 10 subjects received 100 mg minocycline twice a day (200mg) orally (Group 1), and 10 subjects received 50 mg minocycline in the A.M and 100 mg minocycline in P.M (150mg). Patients were told to take medication 1 hr prior to morning appointments. The parameters evaluated: DMF, GI, PI, crevice/pocket depth, soft tissue, SMA-12 (a blood chemistry screen test), CBC, prothrombin times, and concentrations of minocycline in serum, saliva and GCF (by modified Bennet method) from 1 to 7 hrs after taking the medication. DMF score, PD, CBC and prothrombin time were determined on days 1 and 8. All other parameters were evaluated on days 1, 2, 3, 5 and 8.

Results: Minocycline administration resulted in no significant changes in blood chemistry or blood counts. Prothrombin time was not altered (but diet was not considered). Minocycline was present in GCF>serum>saliva. The antibiotic levels in serum and GCF reached bacteriostatic concentrations on day 1 and remained bacteriostatic throughout the study. GI scores were markedly reduced in this study. PI was also reduced. Significant reduction in PD in group 1 but not in group 2. The remaining data showed no significant difference between groups 1 and 2. 4 people in group 1 reported vertigo by the 3rd day. A dose of 150 mg/day should be adequate. GCF concentration of minocycline was 500% of that of serum and its concentration in saliva was approximately 6%.

Conclusion: Minocycline is effective against oral microorganism. It is present in serum at therapeutic levels with either 150 or 200 mgs/day, is concentrated in GCF 5x higher than serum and produces an improvement in gingival health.


 

Topic: Pharmacological efficacy

Author: Britt MR, Pohlod DJ                                                           ARTICLE

Title: Serum and Crevicular fluid concentrations after a single dose of metronidazole

Source: J. Periodontol. 57:104-107, 1985

Type: Clinical trial

Rating: Good

Keywords: Metronidazole; GCF concentrations; Serum concentrations

Background: No study at that time analyzed the concentrations of metronidazole in serum or GCF for an extended period of time, greater than 4 hours, with dose levels of 100-150mg p.o. as used in clinical trials

Purpose: To demonstrate the effectiveness of metronidazole as a site-specific therapeutic agent in treating periodontal disease over 18 hours.

Methods: 6 female patients (age 22 to 47) with no clinical or radiographic evidence of advancing periodontitis, each with14 maxillary teeth. To increase the flow rate of gingival fluid they were told to stop brushing the upper teeth for 14 days. One 250mg tablet of metronidazole was taken at 5am on test day without eating after midnight. Samples of GCF and serum were taken at 1, 2, 3, 4, 5, 6, 7, 8, 12, 18 hours after the single oral dose. Metronidazole and its metabolites were assayed and statistical analysis was performed.

Results: Drug levels were detectable by the first hour in both serum and GCF. In serum the mean concentration peaked by the 2nd hours. In GCF, the mean concentration peaked at the 2nd hour and again at 7th hour. Mean drug concentrations in GCF never exceeded those of serum (NSSD). In both fluids drug levels were detectable for up to 18 hours after the single dose.

Discussion: The second peak observed at 7 hours, has been shown in other drugs also. This second peak is possibly due to enterohepatic recycling of the drug.

Conclusion: Metronidazole can be found in sufficient concentrations in both serum and GCF to inhibit a wide range of suspected periodontopathogens. Metronidazole demonstrates site specificity as a potential chemotherapeutic agent in treating periodontal infections caused by obligate anaerobes.


 

Topic: drug therapy

Authors: Conway TB, Beck FM, Walters JD                                                           ARTICLE

Title: Gingival fluid ciprofloxacin levels at healthy and inflammed human periodontal sites

Source: J Periodontol 71:1448-1452, 2000

Type: cross-sectional study

Rating: Good

Keywords: gingival crevicular fluid/analysis; ciprofloxacin/therapeutic use; serum/analysis; neutrophils; periodontal diseases/drug therapy; follow-up studies; cross-sectional studies


Purpose: PMNs take up and accumulate ciprofloxacin. This may allow them to enhance drug delivery in the inflamed periodontium. The purpose of this study was to determine if systemic ciprofloxacin attains higher concentrations in GCF from inflamed periodontal tissues than it does in the blood stream or GCF from healthy sites. The effect of SRP on GCF ciprofloxacin levels was also investigated.

Methods: 2 groups were recruited: 7 subjects without periodontal disease and good OH and 8 subjects with untreated adult periodontitis (PD ≥5 mm, mod-adv bone loss in at least 2 quads). Three doses of 500 mg of ciprofloxacin were given to both groups to establish steady state tissue levels of the drug and samples were taken 28 hours after the first dose of Cipro. The diseased group continued Cipro (500 mg bid) for another 7 days. GCF was collected from 12 interproximal sites with paper strips in healthy patients. In diseased subjects, 2 quads with the most severe disease were selected and 12 paper strip samples were obtained. Venous blood was drawn on all subjects. After initial collection, 1 quadrant in the disease group was selected for SRP (split-mouth design). These patients returned 196 hours after the first Cipro dose for GCF and serum samples. GCF and serum Cipro content was measured with liquid chromatography

Results: In both healthy and diseased patients, GCF Cipro levels were significantly higher than the serum levels. There was no difference in GCF Cipro levels in the healthy and diseased groups. At 196 hours, the mean pooled GCF volume at root planed sites was 16% lower than at untreated control sites.

Conclusion: The results of this study suggest that inflammation has no significant impact on GCF levels of Cipro. Mean levels of GCF Cipro were 4-5 fold higher than in serum, regardless of periodontal status. The increased availability of this drug in GCF should enhance its antimicrobial effects against susceptible subgingival microorganisms. The total amount of ciprofloxacin delivered to inflamed tissues was greater than at healthy sites because GF flow was higher.


 

Indicators of Disease Activity

Can gingival fluid contents be used to determine severity of periodontal conditions or disease activity?

Topic: Prostaglandin E2 in crevicular fluid

Authors: Leibur E, Tuhkanen A, Pintson U, Soder P-O                                                           ARTICLE

Title: Prostaglandin E2 levels in blood plasma and in crevicular fluid of advanced periodontitis patients before and after surgical therapy.

Source: Oral Diseases 5:223-228, 1999.
Type: clinical study

Rating: Good

Keywords: prostaglandin E2, blood plasma, gingival crevicular fluid, periodontitis surgical treatment.

Purpose: To determine PGE2 levels in venous blood plasma (VBP), gingival blood plasma (GBP) and GCF in advanced periodontal patients before and after surgical treatment.

Material and methods: 12 periodontal patients, 28-45 years old and 7 periodontally healthy patients, same age, were used as controls. During the pre-study period of 7 days patients in both groups received professional cleaning. Clinical parameters (PI, GI, BI, PD, AL) and radiographic (panoramic) bone height were measured before and 6 months after surgical therapy (surgery involved autogenous bone graft in periodontal defects).  PGE2 were determined by radioimmunoassay, and measured prior to surgery in venous blood plasma, gingival blood plasma, and GCF, and after surgery as well.


R: 

Before treatment

After Treatment

  • Diseased sites of periodontal patients had higher plaque levels and more severe gingival inflammation than healthy sites.

  • Diseased sites had significantly deeper mean PD and lower BH (bone height) % than that in control subjects. At baseline means of PI, GI, BI had higher values than that of control.

  • The mean VBP, GBP and GCF PGE2 levels in periodontal patients were higher than healthy controls.

  • The values of clinical parameters as PI, GI, BI, PD and AL diminished and there was no statistically significant differences between the data of clinical parameters of the control subjects and the patients after treatment.

  • After 6 months treatment there was no significant differences between the means of BH% as compared with the same parameter of the control.

  • 6 months after Treatment, the mean levels of PGE2 in VBP, GBP and GCF were significantly reduced in patients with improvement of clinical and radiographic parameters

The mean levels of PGE2 in GBP and GCF before treatment were significantly higher compared to the mean level of PGE2 after treatment.

C:  The present study showed that PGE2 is involved in the pathogenesis of periodontal disease. The inflamed periodontal tissues may produce significant amounts of PGE2 and the degree of inflammation might be determined by the ratios of PGE2.

BL: The lower levels of PGE2 in blood plasma and in gingival crevicular fluid after treatment are signs of improvement of periodontal disease.

Topic: Antibiotics

Authors Lamster IB, Oshrain RL, Harper DS                                                           ARTICLE

Title: Enzyme activity in crevicular fluid for detection and prediction of clinical attachment loss in patients with chronic adult periodontitis. Six month results.

Source: J. Periodontol. 59: 516-523, 1988.

Type: Clinical study

Rating: Good

Keywords: Antibiotics, GCF


Purpose: To study the relationship of changes in GCF levels of the vertebrate lysosomal enzymes B-glucuronidase, arylsulfatase and the cytoplasmic enzyme lactate dehydrogenase in reference to attachment loss in patients with existing periodontitis.


Method: Longitudinal study 36 patients were followed up for 6 months. Patients had existing chronic adult periodontitis (minimal 20 natural teeth, and at least two teeth in each quadrant ≥5mm attachment loss). CAL (baseline-3m-6m), GCF (baseline-3m) were recorded. GCF was collected using filter paper strips, from mesial surface of the 4-5 most distal teeth. 681 samples analyzed at baseline and 3 months. Three groups of patients were identified based on disease progression:

Group I (5 pts) – generalized form of disease activity (CAL > 2 mm at least 3 sites)

Group II (4 pts) – localized form (CAL > 2.5 mm at 1-2 sites)

Group III (27 pts) – did not display CAL

Enzyme analysis was evaluated as a whole mouth score (the percent of samples from a patient in which enzyme activity was at least twice the population mean) and at individual samples.

Results:

Conclusion: An exuberant PMN response is related to clinical attachment loss in patients with existing chronic periodontitis.

Topic: Gingival crevicular fluid proteases

Authors: Bader HI, Boyd RL.                                                            ARTICLE

Title: Long-term monitoring of adult periodontitis patients in supportive therapy: Correlation of gingival crevicular fluid proteases with probing attachment loss.

Source: J Clin Perio 26:99-105,1999.

Type: Longitudinal retrospective study

Rating: Fair

Keywords: periodontits; proteases; probing attachment loss; disease activity; longitudinal studies

Purpose: To determine if a chair-side assay for neutral protease activity in GCF could provide an early indication of site-specific disease activity as defined by probing attachment.


Method: The cohorts in this longitudinal retrospective study were selected from one private practice limited to periodontics. 38 subjects that had undergone periodontal therapy and were selected from a maintenance program with 3 or 6 months recall intervals. All sites selected had ≥4mm CAL loss. Samples of GCF were collected from 71 selected sites using paper strips. The assay is designed to detect only the active and not the total amount of enzyme present in the GCF sample. A score of 0 and 1 was considered negative and a score of 2 was positive. Positive BOP and positive neutral protease activity (NPA) scores were classified as true positives if sites subsequently lost at least 1mm of CAL over the next 12 months. Sites were monitored at 6-month intervals for 24-36 months.

Result: As a predictor of breakdown the NPA assay had an accuracy of 94% and a risk ratio of 37.6 as compared to values of 58% and 1.5 for BOP. When only the subset of sites ≥7mm were considered the NPA assay had a calculated accuracy of 92% vs. a value of 50% for BOP.

Conclusion: The results indicate that the NPA assay appears to differentiate between bleeding at sites exhibiting only chronic inflammation with no CAL loss and bleeding at sites undergoing active CAL loss.


Cr: Why not have active loss be defined as CAL ≥2mm?

Topic: Indicators of Disease Activity

Authors: Lamster IB, Ahlo JK                                                           ARTICLE

Title: Analysis of gingival crevicular fluid as applied to the diagnosis of oral and systemic diseases.

Source: Ann N Y Acad Sci. 1098:216-29, 2007

Type: Discussion

Rating: Good

Keywords: gingival crevicular fluid; diagnosis; periodontal disease

Purpose: A review of GCF and its applications in diagnosing oral and systemic diseases.

Discussion: GCF can be either a serum transudate or more commonly as an inflammatory exudate. It reflects the constituents of blood, cells and tissues of the peridontium. As such, it has been studied to identify active periodontitis.

Collection of GCF includes using micropipettes, appliances to isolate and collect fluid, and filter paper strips (methylcellulose). Attention must be given during paper strip placement in order not to be contaminated with plaque, blood or saliva. The strip must remain in the sulcus long enough to obtain adequate amount of fluid. The volume of GCF may be related to inflammation and permeability/ulceration of crevicular epithelium. 65 GCF components have been examined as potential diagnostic markers of disease progression which are classified as enzymes, inflammatory mediators and tissue breakdown by-products. Loos and Tjoa identified 8 possibly valuable markers: alkaline phosphatase, β-glucuronidase, cathepsin B, MMP-8, MMP-9, DPP II and III, and elastase. The predominance of enzymes associated with tissue breakdown clearly emphasizes the importance of the inflammatory response to the pathogenesis of periodontitis. Limitations of application of using this for diagnostic testing include needing to collect samples from multiple sites on a patient, isolation of GCF is difficult, and laboratory tests are not routinely used.

-Systemic Diseases

Diabetes: PGE2 and IL-1B have been found in higher concentrations in type 1 DM patients, regardless of severity of the disease. Higher IL-6 levels have been seen in type 2 DM compared to adult periodontitis and healthy controls.

Smoking: Smokers demonstrate elevated levels of IL-8 and lower levels of IL-4. IL-6 was elevated in with aggressive periodontitis that smoked. IL-1α was lower in smokers.

CVD: Elevated leukotrienes were found in GCF of patients with atherosclerosis (with or without periodontitis).

HIV: High levels of IL-1B, IL-6 and TNF-A and IFN-gamma have been found in HIV patients with periodontitis suggesting the elevated inflammatory mediators may play a role in the bony destruction in these patients.

Conclusion: Analysis of GCF has contributed to understanding the role of inflammation in the periodontal disease process but sampling is impractical in the clinical setting. Saliva collection is a less technique sensitive and more amenable to chairside utilization.

Topic: Host defense

Author: Giannobile WV.                                                          NO ARTICLE

Title: Salivary diagnostics for periodontal diseases.

Source: J Am Dent Assoc. 2012 Oct;143(10 Suppl):6S-11S.

Type: Review

Rating: Good

Keywords: gingival crevicular fluid, tetracycline, antibiotic, biomodulation


Purpose: To review the literature with respect to currently available salivary diagnostics used to identify bacteria prevalent in periodontal disease, and to focus on the future development and use of a variety of rapid detection platforms, such as lab-on-a-chip, as a point-of-care (POC) device for identification of a pt’s risk.

Discussion:

BL: Saliva-based diagnostics offers a promising future for diagnosing perio dz and monitoring tx outcomes.


 

Topic: Gingival crevicular fluid

Authors: Guentsch A, Preshaw PM, Bremer-Streck S, Klinger G, Glockmann E, Sigusch BW.                                                            ARTICLE

Title: Lipid peroxidation and antioxidant activity in saliva of periodontitis patients: effect of smoking and periodontal treatment.

Source: Clin Oral Investig. 12(4):345-52, 2008. Epub 2008 May 29.

Type: Clinical study

Rating: Good

Keywords: Periodontitis, Oxidative stress, Lipid peroxidation, Antioxidant capacity, Smoking, Saliva

Background: PMNs constitute the first line of cellular host defense against bacteria in the gingival sulcus. PMNs produce reactive oxygen species (ROS), molecules which are capable of initiating periodontal tissue destruction. ROS can cause tissue damage via DNA damage, lipid peroxidation, protein damage, and enzyme oxidation. Several ROS and lipid peroxidation products are produced in physiological quantities in the human body, but an overproduction of ROS occurs at sites of chronic inflammation.  During gingival inflammation, GCF flow increases, and components of the inflammatory response are detectable in saliva, including lipid peroxidation products.

Purpose: To examine lipid peroxidation (as an end product of oxidative stress) and corresponding antioxidant activity in patients with periodontitis and assess the influence of smoking and periodontal treatment on these parameters.

Methods: 30 subjects with generalized chronic periodontitis were included.  Patients did not have perio tx, abx, or immunosuppressive agents in the preceding 6 months. Periodontally healthy control subjects (n=30) with no evidence of periodontal disease were also recruited. Test and control groups each contained the same proportion of smokers and non-smokers. Subjects with significant systemic dz (diabetes, cancer, coronary heart dz), pregnant or lactating females were excluded. Clinical data were recorded after sample collection (saliva and blood). PDs, BOP were assessed. Non-surgical tx (full mouth sc/rp, maintenance, monitor OH) given in the periodontitis patients. Sample collection and clinical data was recorded following hygiene phase and at 6 months. Malondialdehyde (MDA) (final end product of lipid breakdown caused by oxidative stress), glutathione peroxidase (GSHPx) (antioxidant parameter), and total antioxidant capacity (TAOC) were measured. 

Results: Patients with periodontal dz demonstrated significantly higher mean PDs and BOP. There were no SSD in mean PDs between smokers and non-smokers. Mean PDs significantly reduced after periodontal treatment in the periodontitis patients.

MDA: The level of MDA increased progressively from the non-smokers to the smokers of the periodontally healthy controls and then to the non-smokers and smokers of the periodontitis group. Lowest in saliva of the non-smoking periodontally healthy subjects (0.065±0.05 μmol/l). Significantly higher in periodontitis patients who smoked (0.123±0.08 μmol/l).  Periodontal tx led to a significant reduction in lipid peroxidation products.

GSHPx: Patients with periodontitis (smokers and non-smokers) demonstrated significantly elevated glutathione peroxidase activity compared to periodontally health controlled groups. GSHPx activity was significantly reduced after periodontal tx.

TAOC flow rate: Significantly lower in patients with periodontitis (0.34±0.26 μmol/ml) in comparison to the controls (0.62±0.24 μmol/ml; p<0.05). TAOC flow rated did not increase in perio patients after treatment.

- Patients with periodontitis demonstrated more lipid peroxidation than healthy subjects and smoking enhanced this effect.

Conclusion: Patients who smoke and have periodontal dz demonstrate more lipid peroxidation in saliva than healthy subjects. There was an increase in glutathione peroxidase activity in periodontal dz and an additive effect of smoking was identified. A reduced antioxidant capacity in periodontitis patients was noticed. Successful periodontal therapy had an effect on MDA and GSHPx activity in saliva, but not the antioxidant flow rate.

Topic: gingival fluid and saliva

Authors: Reher VG1, Zenóbio EG, Costa FO, Reher P, Soares RV.                                                           ARTICLE

Title: Nitric oxide levels in saliva increase with severity of chronic periodontitis.

Source: J Oral Sci. 2007 Dec;49(4):271-6.

Type: clinical

Rating: good

Keywords: chronic periodontitis; saliva; nitric oxide; biological marker; diagnostic.

Background: Nitric oxide (NO) has been linked to the etiopathogenesis of periodontal disease. Cytokines and other bacterial products stimulate the expression of inducible NO synthase (iNOS) and interfere with periodontal disease progression.

Purpose: To measure and compare salivary NO levels in patients with and without generalized chronic periodontitis (GCP); to evaluate correlations between these levels and probing depth as a clinical diagnostic parameter; and to determine usefulness of NO measurements as biological markers of perio disease.

Methods: 30 individuals; exclusion criteria: current smokers, pregnant, recent use of Abs or recent trauma. Inclusion: periodontitis with no perio tx in past year. Subjects were divided into three groups: control group (GC) of subjects without periodontitis; group (GM) with moderate generalized chronic periodontitis, and group (GA) with advanced generalized chronic perio. Salivary samples were collected and NO levels measured.

Results:

  1. GCP group NO mean concentrations (GM: 7.78 μM; GA: 15.79 μM) were higher than in the control group (GC: 5.86 μM). 

  2. NO levels in the GA group were significantly higher than in the GC group

  3. Positive correlations between NO level and the number of teeth with a probing depth of ≥ 4 mm and ≥ 7 mm were observed.

Conclusion: NO levels are elevated in individuals with GCP, and are associated with periodontal severity. NO may serve as a potential biological marker for detection and monitoring of GCP.


 

Topic: Indices

Author: Frodge BD, Ebersole JL, et al.                                                           ARTICLE

Title: Bone remodeling biomarkers of periodontal disease in saliva

Source: J Periodontol. 79(10):1913-9, 2008

Type: Clinical Trial

Rating: Good

Keywords: C-telopeptide pyridinoline; periodontal diseases; receptor activator of nuclear factor-kappa B ligand; tumor necrosis factor-alpha


Background: Tumor necrosis factor-alpha (TNF-a), C-telo-peptide pyridinoline cross-links of type I collagen (ICTP), and receptor activator of nuclear factor-kappa B ligand (RANKL) have been associated with bone remodeling and periodontal tissue destruction.

Purpose: To evaluate the hypothesis that biomolecules involved in bone remodeling (TNF-α, RANKL, ICTP) are increased in the saliva of patients with periodontal disease compared to control subjects.

Methods:

Experimental group: 35 patients with moderate to severe periodontitis

Control group: 39 healthy adults of similar age, race and gender.

Un-stimulated saliva was collected from each subject. One examiner recorded clinical indices for each patient: 1-PD (measured at six locations per tooth); 2-BOP; 3-AL (interproximal sites only).

Concentrations of TNF-a were determined in all 74 adults. Salivary levels of RANKL and ICTP of a subset of 21 subjects and 21 matched controls were examined (levels were below the limit of detection in >80% of subjects in both groups).

Results: RANKL and ICPT were detected only in a minority of patients. The small numbers of subjects with these two markers detected did not allow meaningful comparisons of these analyses between groups.

TNF-α was detected in all samples. Mean levels of TNF-α were significantly higher in subjects with periodontal disease than in controls. Subjects with salivary TNF-a levels above a threshold of 5.75 pg/ml (i.e., two standard deviations above the mean of the controls) had significantly more sites with BOP, PD ≥ 4 mm, and AL ≥ 2 mm.

Bottom Line: Salivary levels of TNF-α above a threshold might help to identify patients who have periodontal disease.

Topic: immunology/IL-1β

Authors: Tobón-Arroyave SI, Jaramillo-González PE, Isaza-Guzmán DM                                                            ARTICLE

Title: Correlation between salivary IL-1beta levels and periodontal clinical status

Source: Arch Oral Biol. 53(4):346-52, 2008

Type: clinical study

Rating: Good

Keywords: aggressive periodontitis, chronic periodontitis, IL-1β, immunology, saliva

Purpose: To assess the concentration of proinflammatory cytokine IL-1β in saliva of periodontally diseased and healthy patients and their relationship with periodontal status.

Methods: 66 patients were recruited for this study. Clinical parameters recorded included PD and CAL. These values were used to classify the extent of periodontal disease. 30 patients had chronic periodontitis, 18 had aggressive, and 18 were used as healthy controls. About 10-ml of unstimulated whole saliva was collected from each subject. ELISA was used to analyze the amount of IL-1β in the samples.

Results: No differences were noted between men and women. The chronic group was statistically higher in age than the aggressive and healthy groups. Detectable levels of IL-1β were found in all groups. Aggressive and chronic periodontitis groups had significantly higher amounts of IL-1β in their saliva, however no statistical difference was noted between the chronic and aggressive groups.

Conclusion: The study suggests that whole saliva sampling could play an important role in terms of immunological purposes in periodontal disease and that elevated IL-1β concentration may be one of the host-response components associated with the clinical manifestation of periodontal disease.

Topic: Horizontal augmentation

Authors: Rai B, Kharb S, Jain R, Anand SC.                                                           ARTICLE

Title: Biomarkers of periodontitis in oral fluids.

Source: J Oral Sci. 50(1):53-6, 2008
Type: clinical study

Rating: Good

Keywords: Matrix metalloproteinases, gingivitis, periodontitis, healthy.


Background: MMP’s (Matrix metalloproteinases) are the major group of enzymes responsible for degradation of extracellular matrix (ECM). The onset of collagen destruction is caused by the action of collagenases, a subgroup of MMPs. MMP-1,2,3,8 and 9 have been found in biopsy specimens of human inflammatory periodontal tissues whereas healthy gingiva contains only pro-MMP-2. MMP-2 is secreted by gingival fibroblasts and MMP-9 is mainly secreted by PMNs and they degrade type IV collagen present in gingival tissues (basement membrane). MMP-8 degrades type I and type III collagen which is critical for periodontal destruction.

Purpose: The aim of the present study was to compare the levels of MMP-2 and MMP-9 in GCF and MMP-8 in saliva samples from healthy subjects, and patients with gingivitis and periodontitis.

Materials and methods:

Results: Elevated salivary levels of MMP-8 and crevicular levels of MMP-9 were observed in the periodontitis and gingivitis patients compared to the controls. Crevicular MMP-2 levels were lower in gingivitis and periodontitis as compared to healthy controls. PPD, CAL and BOP were correlated with elevated levels of MMP-8. CAL loss and BOP were significantly correlated with elevated levels of MMP-2 and MMP-8.

Discussion: MMP-9 levels in GCF were higher in patients with chronic periodontitis than in patients with gingivitis and healthy subjects, while MMP-2 levels in GCF were lower in patients with chronic periodontitis than in patients with gingivitis and healthy subjects. Salivary MMP-8 levels were higher in patients with chronic periodontitis than in patients with gingivitis and healthy subjects.


BL: MMP-2, MMP-8 and MMP-9 levels were highly correlated with the CAL loss and PDs.

Topic: Salivary biomarkers

Authors: Miller CS, King CP Jr, Langub MC, Kryscio RJ, Thomas MV                                                            ARTICLE

Title: Salivary biomarkers of existing periodontal disease: a cross-sectional study.

Source: J Am Dent Assoc. 137(3):322-9, 2006.

Type: Case Report

Rating: Good

Keywords: GCF flow, Salivary biomarkers, saliva


Background: Interleukin 1 beta (IL-1β)- a proinflammatory cytokine that stimulates the induction of adhesion molecules and other mediators that facilitate and amplify the inflammatory response that occurs in periodontal disease.

Matrix metalloproteinase (MMP)-8- a key enzyme in extracellular collagen matrix degradation derived predominantly from PMNs during acute stages of periodontal disease.

OPG- a glycoprotein that acts as an osteoblast- secreted decoy receptor and competitively inhibits osteoclast differentiation and activity by preventing osteoclast differentiation factor or the receptor activator of NF-κB ligand (RANKL) from binding to osteoclast precursors and promoting the formation of bone-resorbing osteoclasts.


Purpose: To test the hypothesis that levels of salivary biomarkers specific for three aspects of periodontitis: inflammation, collagen degradation and bone turnover correlate with the clinical features of the disease.

Method: The experimental group constisted 28 pts (12 M, 16 F). Inclusion criteria: generalized BOP, at least 20% of sites with ≥4mm PD, at least 5% of sites with interproximal CAL loss >2mm, and evident radiographic bone loss. 29 healthy subjects of similar age were used as controls.

Unstimulated whole expectorated saliva was collected and was immediately frozen and analyzed within 6 months. Subjects rinsed their mouths with tap water, and then expectorated whole saliva into sterile tubes.

PD, BOP, and CAL were recorded immediately after saliva sampling. Saliva samples were analyzed for the concentration of IL-1b, MMP-8 and OPG using ELISA or EIA. Statistical analysis was done.

Results:

Conclusion: Levels of salivary biomarkers, for inflammation, collagen degradation and bone turnover correlate with the clinical features of periodontal disease and suggest that elevated salivary levels of MMP- 8 and IL-1b are candidate biomarkers of periodontal disease.

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