Label-free quantitative proteomics reveals differentially regulated proteins in experimental gingivitis.

We investigated the sequential protein expression in gingival crevicular fluid samples during the induction (I) and resolution (R) of experimental gingivitis. Periodontally and systemically healthy volunteers (n = 20) participated in a three-week experimental gingivitis protocol, followed by debridement and two weeks of regular plaque control. Gingival crevicular fluid (GCF) samples were collected at baseline, Day 7, 14, and 21 (induction; I-phase), and at Day 21, 25, 30, and 35 (resolution; R-phase). Liquid chromatography-tandem mass spectrometry (LC-MS/MS) for label-free quantitative proteomics was applied. A total of 287 proteins were identified including 254 human, 14 bacterial, 12 fungal, and 7 yeast proteins. Ontology analysis revealed proteins primarily involved in cytoskeletal rearrangements, immune response, antimicrobial function, protein degradation, and DNA binding. There was considerable variation in the number of proteins identified, both among subjects and within subjects across time points. After pooling of samples between subjects at each time point, the levels of 59 proteins in the I-phase and 73 proteins in the R-phase were quantified longitudinally. Our data demonstrate that LC-MS/MS label-free quantitative proteomics is valuable in the assessment of the protein content of the GCF and can facilitate a better understanding of the molecular mechanisms involved in the induction and resolution of plaque-induced gingival inflammation in humans.

Evaluation of the antiplaque efficacy of two cetylpyridinium chloride-containing mouthwashes.

OBJECTIVE: The purpose of this clinical study was to evaluate the efficacy in reducing dental plaque regrowth of two mouthwashes containing 0.075% cetylpyridinium chloride (CPC), one with 6% alcohol and one alcohol-free, as compared to a negative control mouthwash without CPC, using the Modified Gingival Margin Plaque Index (MGMPI). METHODS: The study was a double-blind, randomized, three-way crossover, controlled design. Following a washout period, subjects reported to the dental clinic where they were instructed to brush their teeth, used their assigned mouthwash, and were scored by the examining dentist for plaque using the MGMPI method. Subjects were instructed to refrain from all oral hygiene for the next 24 hours, except for rinsing with their assigned mouthwash 12 hours post-brushing. After this 24-hour period, subjects returned to the dental clinic and were once again scored for plaque. This sequence of washout followed by mouthwash use and plaque scoring was repeated until each subject had used all three mouthwashes. An ANOVA was conducted to assess between-group differences. RESULTS: The two test mouthwashes significantly reduced plaque regrowth over a 24-hour period (p < 0.05) as compared to the negative control mouthwash. The difference between the CPC-containing mouthwashes was not significant (p = 0.4868). CONCLUSION: Two mouthwashes containing 0.075% CPC, one with 6% alcohol and the other alcohol-free, were found to be safe and effective in reducing plaque accumulation when compared a negative control mouthwash without CPC. In short-term studies, the MGMPI appears useful for evaluating the antiplaque efficacy of mouthwash products.

Comparison of the short-term antiplaque/antibacterial efficacy of two commercial dentifrices.

OBJECTIVE: The objective of these three clinical trials was to compare the impact of two commercial products, Colgate Total and Crest Pro-Health, on the formation of dental plaque over a 24-hour period of time. The studies utilized the Modified Gingival Margin Plaque Index (MGMPI), a validated and reliable clinical method for assessing the efficacy of products in reducing plaque build-up. METHODS: Colgate Total and Crest Pro-Health were the test products for all three clinical trials. Colgate Great Regular Flavor (CR) was used as the universal washout product. Colgate Total, as the only toothpaste approved by the FDA under an NDA for antiplaque, antigingivitis, and anticaries benefits, contains 0.3% triclosan/2.0% PVM/MA copolymer for antigingivitis and antiplaque, as well as 0.243% sodium fluoride (NaF) for anticaries. Crest Pro-Health contains 0.454% stannous fluoride (SnF2) as both a monographed anticaries agent and a monographed antigingivitis agent, along with sodium hexametaphosphate and zinc lactate. Twenty-five healthy subjects meeting all study criteria were included into each of the double-blind studies. Product assignment was randomized and a crossover design was implemented. Informed consent was obtained from all subjects prior to commencement of each of the studies. The studies followed published MGMPI procedures, which require subjects to receive a dental scaling/prophylaxis followed by a one-week washout period prior to use of test products. A baseline MGMPI score was calculated following use of the test products in the dental clinic. Subjects refrained from all oral hygiene for 24 hours following use of each test product, and returned to the clinic for a 24-hour MGMPI score. Following a washout period, subjects repeated the procedure with the other test product as per the crossover design. The differences (delta) between baseline plaque scores and 24-hour plaque scores were independently calculated for each study, and the delta values were compared for the two test products in each of the studies. RESULTS: In all three clinical trials, Colgate Total significantly reduced plaque regrowth over a 24-hour time period (p < or = 0.05) compared to Crest Pro-Health. Existing differences were determined via a paired t-test, which confirmed that Colgate Total was statistically significantly different from Crest Pro-Health. CONCLUSION: These in vivo data support the antiplaque benefit of the 0.3% triclosan/2.0% PVM/MA copolymer/0.243% sodium fluoride dentifrice. Additionally, the results support that Colgate Total provides superior efficacy in inhibiting the formation of dental plaque compared to Crest Pro-Health.

The use of the Modified Gingival Margin Plaque Index (MGMPI) method to investigate the inhibitory effect of various toothpastes on dental plaque formation.

OBJECTIVE: Colgate Total (CTT) is the only FDA-approved toothpaste for antiplaque and antigingivitis benefits. The objective of this study was to compare the impact of Colgate Total Pharma (CTP), a new variant of Colgate Total, with Colgate Regular Toothpaste (CRT) on plaque formation over a 24-hour period following a single use of the dentifrice. METHODS: CTP and CRT were the two test products. CRT was used for a washout product as well. Fifteen male/female subjects who met the inclusion/exclusion criteria were included into this single-blind (preliminary phase) and double-blind (randomized phase) crossover study. Ethical approval and written informed consent were obtained. Preliminary phase: After a one-week washout with CRT, subjects brushed in the dental clinic with CRT before a one-minute use of a test dentifrice. A baseline Modified Gingival Margin Plaque Index (MGMPI) score was calculated. Subjects refrained from oral hygiene for 24 hours, and returned to the clinic for their 24-hour MGMPI score. Subjects entered the second washout phase to repeat as per the crossover design. The above procedures were conducted three times by three independent examiners. Randomized phase: Subjects were randomized to the groups according to a computer-generated randomization schedule. The procedure was carried out as in the preliminary phase, except the washout period between the two products was at least one week and the products (CTP or CRT) were used in a randomized double-blind manner. Plaque scores were recorded as above. RESULTS: CTP provided a significant (p = 0.01) antiplaque effect versus CRT. The results are consistent with previously reported data for CTT. All three examiners demonstrated a strong correlation for this clinical study utilizing the MGMPI methodology. CONCLUSION: This clinical investigation examined the efficacy of a new variant of a commercial dentifrice, historically shown to provide antiplaque and antigingivitis efficacy. It is important to confirm the continued efficacy of new products to consumers and to the profession. Additionally, this clinical trial demonstrated the usefulness of the clinical methodology with respect to consistency in results by three independent clinical examiners. Because this methodology is often employed to document antiplaque benefits of new and existing technologies, it is important to periodically evaluate and confirm its reliability and reproducibility.

Comparison of two clinical methods for evaluating the anti-plaque efficacy of a dentifrice.

OBJECTIVE: The objective of this research was to assess two different published methods of plaque level evaluation on human subjects, and to determine whether both methods can produce similar findings to the point that the methods can be used interchangeably. METHODOLOGY: Healthy human volunteers entered into a number of double-blind, cross-over clinical trials. Two plaque scoring methods were used: The Modified Gingival Margin Plaque Index (MGMPI) and the Turesky Modified Quigley-Hein (TMQH). With the MGMPI, plaque was evaluated 24 hours after a single product use. With the TMQH, plaque was scored after four days of product use. Three dentifrices were studied: Colgate Total, Colgate Total plus Whitening, and Colgate Regular Dental Cream. Additionally, two mouthrinses were evaluated: PerioGard and Fluorigard. In all studies conducted, there was a one-week wash-out period between each product use. RESULTS: There were no side effects observed or reported in any study. In all studies, both the plaque indices used found that the active products Colgate Total, Colgate Total plus Whitening and PerioGard produced significantly lower mean plaque scores compared to Colgate Regular Dental Cream and Fluorigard (p < 0.05). CONCLUSION: The results of this investigation indicate that the two plaque evaluation methods continue to demonstrate that an active product is significantly more efficacious than a control product. Therefore, either method can be used to investigate the anti-plaque efficacy of a product. Additionally, the MGMPI method can be used 24 hours after product use, which is a clear advantage over a four-day product use and will yield the same results.

Analysis of the antibacterial activity and plaque control benefit of colgate total dentifrice via clinical evaluation and real-time polymerase chain reaction.

OBJECTIVE: This study analyzed, from a combined clinical and molecular biologic perspective, the antibacterial and antiplaque efficacy of Colgate Total dentifrice (CTD). METHODOLOGY: A single-blind crossover study design utilized 11 healthy human subjects. After a one-week washout period, subjects donated dental plaque, received a dental prophylaxis, and subsequently brushed with a test product. Twenty-four hours postbrushing, dental plaque was collected and a clinical plaque score determined. Dental plaque was submitted for Real-time Polymerase Chain Reaction (Real-time PCR) analysis. The same procedure was repeated in accordance with a crossover design for the use of the second test product. Following a one-week washout, a plaque donation, prophylaxis, and brushing with the test product ensued for each subject. Twenty-four hours post-brushing, the subjects returned for a plaque score and plaque donation. RESULTS: Twenty-four hours after brushing, dental plaque coverage increased 17.88% +/- 8.27% with CTD, compared to 30.42% +/- 9.97% with Colgate Cavity Protection (CCP; p = 0.005). Real-time PCR found plaque collected 24 hours after brushing with CTD exhibited, on average, fewer representative periodontal pathogens (Fusobacterium nucleatum, Actinobacillus actinomycetemcomitans, Tannerella forsythensis, and Porphyromonas gingivalis) and fewer early colonizers (Actinomyces naeslundii) than plaque collected before brushing, whereas CCP showed a moderate effect on oral bacteria. CONCLUSION: The study provides clinical and molecular biological evidence to substantiate the antibacterial and plaque control benefits of Colgate Total, and suggests the value of combining a molecular biological method with clinical research to corroborate clinical benefits.

Effectiveness of a triclosan/copolymer dentifrice on microbiological and inflammatory parameters.

According to the US Surgeon General's report, "Oral Health in America," published in 2000, most adults in the United States show some degree of periodontal pathology, with severe periodontal diseases affecting about 14% of middle-aged adults. Periodontal diseases are polymicrobial-induced inflammatory diseases, and they vary from mild gingival inflammation to severe deterioration of the periodontium, ie, loss of periodontal supportive tissues and, ultimately, tooth loss. New evidence shows that periodontal diseases may impact systemic health. For this reason, the maintenance of a healthy mouth is becoming increasingly important for the overall health of the body. This article summarizes laboratory research conducted during the development of a novel, multibenefit, oral-care technology based on triclosan--a broad-spectrum antibacterial agent--and a polyvinylmethylether/maleic acid copolymer. This unique combination of agents is found in Colgate Total, a clinically proven efficacious dentifrice for control of dental plaque and gingivitis. Data are presented that demonstrate the unique antibacterial properties of this dentifrice: (1) a broad-spectrum antimicrobial profile; (2) the long-lasting retention of triclosan on hydroxyapatite and epithelial cells; and (3) molecular evidence of antibacterial activity against specific pathogens in clinical dental plaque. In addition, data are presented that demonstrate the anti-inflammatory effects of triclosan on specific cytokines, the interruption of inflammatory pathways, and the inhibition of bone resorption. Overall, these data support the multibenefit clinical effects of Colgate Total and suggest a plurality of mechanisms of action.

The development of a new dental probe and a new plaque index.

OBJECTIVE: A curved periodontal probe has been developed to measure the length of newly formed plaque along the gingival margin of a tooth and the length of the entire gingival margin of the tooth. Plaque is expressed as a percent of the tooth's gingival margin, and is defined as the Modified Gingival Margin Plaque Index (MGMPI). The purpose of this research was to evaluate the efficacy of three marketed antiplaque products using the MGMPI. METHODOLOGY: A series of three clinical studies, involving five separate trials, was conducted. Three marketed antiplaque products were tested; PerioGuard mouthrinse, Colgate Total dentifrice and Colgate Total Plus Whitening dentifrice. Using a cross-over design, the efficacy of these test products on plaque formation was compared to Colgate Regular dentifrice. Plaque was evaluated before, and 24 hours after a single use of the above products using the MGMPI. RESULTS: In all studies conducted, plaque formation, as measured by the MGMPI, significantly increased over a 24-hour period. There were no significant differences in 24-hour scores between the test and control products evaluated. When the change from baseline to 24 hours was analyzed, the test products resulted in a consistently and significantly (p < 0.05) lower MGMPI mean score than the Colgate Regular dentifrice control. CONCLUSION: A new method has been developed that can measure plaque formation along the gingival margin.

Triclosan blocks MMP-13 expression in hormone-stimulated osteoblasts.

BACKGROUND: Matrix metalloproteinase-13 (MMP-13) is an important enzyme for the modulation of bone turnover and gingival recession. Elevated levels of MMP-13 are associated with alveolar bone resorption, periodontal ligament breakdown, and gingival attachment loss, which are the clinical symptoms of periodontal disease. Evidence continues to suggest that periodontal disease contributes to oral tissue breakdown and is linked to numerous systemic conditions. Triclosan (TCN) is a long-standing, proven antibacterial and anti-inflammatory agent found in the only Food and Drug Administration-approved dentifrice for the treatment of plaque and gingivitis. METHODS: This study examines the inhibitory effects of TCN on lipopolysaccharide-, parathyroid hormone (PTH)-, and prostaglandin E2 (PGE2)-induced expression of MMP-13 in UMR 106-01 cells, an osteoblastic osteosarcoma cell line. The cells were stimulated with PTH or PGE2 to induce MMP-13 mRNA expression, and real-time reverse transcription-polymerase chain reaction was performed to determine gene expression levels. Western blot analysis assessed the presence or absence of protein degradation or inhibition of protein synthesis. MMP-13 promoter reporter assay was used to explore possible direct effects of TCN on the MMP-13 promoter. RESULTS: TCN significantly reduced PTH or PGE2 elevated expression of MMP-13 in osteoblastic cells without affecting basal levels of the mRNA. Surprisingly, TCN enhanced the expression of c-fos and amphiregulin mRNA. A promoter assay indicated that TCN directly inhibits the activation of the PTH-responsive minimal promoter of MMP-13. CONCLUSION: The present study appears to have identified a nuclear mechanism of action of TCN that accounts for the ability of TCN to inhibit PTH- or PGE2-induced MMP-13 expression in osteoblastic cells.