In vitro antimicrobial effects of grape seed extract on peri-implantitis microflora in craniofacial implants

To determine the antimicrobial effects of grape seed on peri-implantitis microflora.Methods:The grape seed extract was tested against peri-implantitis microflora most commonly found in craniofacial implants including reference strains of Staphylococcus aureus (S. aureus), Escherichia coli (E. coli), Candida albicans (C. albicans) and clinical strains of S. aureus, Klebsiella pneumonia (K. pneumonia) and Candida parapsilosis (C. parapsilosis) by disk diffusion test. Minimum inhibitory concentrations (MIC) and minimum cidal concentrations (MCC) were determined using modified agar dilution millpore method. The extract was further combined with polyethylene glycol and propylene glycol, and was tested for antimicrobial effects. Results: Grape seed extract showed positive inhibitory effects with S. aureus at MIC of 0.625 mg/mL and MCC of 1.25 mg/mL respectively. However the extracts showed minimal or no reactivity against strains of E. coli, K. pneumonia, C. parapsilosis and C. albicans. The use of grape seed extract in combination with polyethylene glycol and propylene glycol also showed dose dependent inhibitory effect on S.aureus. Conclusions: The results of the study showed that grape seed has potential antimicrobial effects which can be further studied and developed to be used in the treatment of infected skin-abutment interface of craniofacial implants.

Fermentation of various sugars and sugar substitutes by oral microorganisms

Objective: To examine acid production of caries-associated strains of oral microorganisms and salivary microorganisms from sugar and sugar substitutes. Methods:Standard and clinical strains of Streptococcus mutans (S. mutans), Lactobacillus casei (L. casei) and Candida albicans were incubated in peptone-yeast-extract media containing 1% test sugar (sucrose, glucose, fructose) or sugar substitutes (xylitol, sorbitol, trehalulose and palatinose) at 37 ℃in 5% CO2 for 24-48 h. The pH of each culture was measured and microbial growth was determined as optical density at 660 nm. Paraffin-stimulated saliva collected from high caries-risk persons were added to media containing 10%test sugar or sugar substitutes. The pH of medium was measured at each time interval from 0-90 minutes. Results:All types of sugar and trehalulose could be fermented by all test microorganisms in pH lower than 5.5 except sucrose by standard strain of L. casei. All sugar and sugar substitutes supported growth of all organisms except xylitol for S. mutans. In the fermentation assay by salivary microorganisms, all sugar could be utilized and produced pH< 5.5 within 10 minutes of incubation and the pH drop was prolonged to until 90 minutes. Conversely, xylitol and palatinose were not fermented by microorganisms in saliva. Conclusions:All test microorganisms could ferment sucrose, glucose, fructose and trehalulose to pH lower than 5.5. Sugar alcohols and palatinose were not utilized well by organisms and may be used as sugar substitutes to reduce dental caries incidence. However, further studies particularly clinical investigations are required to evaluate the cariogenicity of these sugar substitutes.

Remineralization of enamel subsurface lesions by xylitol chewing gum containing funoran and calcium hydrogenphosphate.

The aim of the present study was to determine the remineralization effects of xylitol chewing gum containing funoran and calcium hydrogenphosphate on enamel subsurface lesions in humans. The study was a double-blind, randomized, cross-over design, with 4 types of gum: (1) xylitol gum, (2) xylitol gum containing funoran and calcium hydrogenphosphate, (3) sugar gum, and (4) gum base as a control. Seven subjects were instructed to wear removable lingual appliances, with half-slab insets of human enamel containing demineralized subsurface lesions. They were told to chew gum for 20 minutes 4 times per day for 7 days. Upon completion of each treatment the enamel half-slabs were paired with their respective demineralized control half-slabs, embedded, sectioned, and subjected to microradiography and densitometric image analysis, for measurement of the level of remineralization. The mean area of remineralization (deltaZd-deltaZr) and mean percent remineralization (%R) in those chewing xylitol gum containing funoran and calcium hydrogenphosphate were significantly higher than the corresponding values for xylitol gum, sugar gum and gum base. Chewing xylitol gum containing funoran and calcium hydrogenphosphate has a significant effect on the remineralization of initial caries-like lesions of the teeth.

Salivary secretory IgA, pH, flow rates, mutans streptococci and Candida in children with rampant caries.

The aim of this study was to determine the levels of secretory IgA (SIgA), pH, flow rates, mutans streptococci (MS) and Candida in saliva of children with rampant caries compared to those caries-free. Thirty children (age 62-123 months) were enrolled and divided into two groups: Group I, children with rampant caries, Group II, caries-free children. The average salivary flow rate was measured from the volume yielded within 5 minutes and the pH was determined using a pH-electrode. Measurement of SIgA was performed using an immunoassay kit. The levels of MS and Candida were determined by culture on Mitis-Salivarius Bacitracin agar and Sabouraud dextrose agar. It was found that children with rampant caries presented with significantly higher levels of salivary SIgA, MS and Candida. However, the mean values for salivary flow rates and pH were similar between the groups. The results reveal that children with rampant caries had significantly higher levels of SIgA, MS and Candida in their oral cavities. This finding tends to support the hypothesis that higher levels of salivary SIgA may reflect a past exposure of the host to cariogenic microorganisms.

Plaque pH response to snack foods in children with different levels of mutans streptococci.

This study aimed to investigate the effects of some snack foods on plaque pH in children with different levels of mutans streptococci (MS). Six children, aged 9-12 years, with low (<10(4)) and 6 children, aged 10-12 years, with high (>106) numbers of MS/ml saliva participated in the study. Dental plaque pH changes, after the consumption of milk chocolate, sweet biscuit, instant noodle, sticky rice with banana and a 10% sucrose positive control were measured using pH-electrode. The measurements of plaque pH were made on forty-eight-hour accumulated plaque, at baseline to determine the resting pH of the fasted plaque and at time intervals of 2, 5, 10, 20 and 30 minutes after food consumption. The plaque pH curves, delta pH values and area under curve for pH 6.0 for each test food were determined. Plaque acidogenicity was more pronounced for the high-MS group at almost all test periods compared to the low-MS group with all test foods. The test foods were ranked according to maximum pH drop in about the same order in both groups as follows: 10% sucrose > milk chocolate > sweet biscuit > sticky rice with banana > instant noodle. The plaque pH also stayed below pH 6.00 for a longer period in the high-MS group with sweet biscuit, milk chocolate, and sticky rice with banana. Findings suggest that pH responses were more acidic in high-MS group than low-MS group.

Antimicrobial activity of Cratoxylum formosum on Streptococcus mutans.

The gum of Cratoxylum formosum, commonly known as mempat, is a natural agent that has been used extensively for caries prevention by hill tribe people residing in Thailand. The objective of this study was to investigate the antimicrobial activity of Cratoxylum formosum gum on Streptococcus mutans (S. mutans) in vitro. The gum extracted from stem bark of Cratoxylum formosum was investigated for antimicrobial activity against different strains of S. mutans, including S. mutans KPSK2 and 2 clinical isolates. Inhibition of growth was primarily tested by agar diffusion method. A two-fold broth dilution method was then used to determine the minimum inhibitory concentration (MIC) of the extract. The extract of Cratoxylum formosum was effective against S. mutans with the inhibition zones ranging from 9.5 to 11.5 mm and MIC values between 48 microg/ml and 97 microg/ml. The gum of Cratoxylum formosum has high antimicrobial activity against S. mutans and may become a promising herbal varnish against caries.

Effects of sonicated Prevotella intermedia, Fusobacterium nucleatum and Lactobacillus casei extracts on interleukin-8 production by human dental pulp cells.

The objective of this study was to determine the effects of Prevotella intermedia, Fusobacterium nucleatum and Lactobacillus casei on the production of IL-8 by human dental pulp cells. Human dental pulp cells from teeth of young patients (aged 18-25 years) were cultured and tested with sonicated P. intermedia ATCC 25611, F. nucleatum ATCC 25586 and L. casei ATCC 4646 extracts. IL-8 secreted into the culture supernatants were measured at 6, 12 and 24 hours using a quantitative sandwich enzyme immunoassay technique. Cell viability was evaluated using trypan blue exclusion technique. IL-8 production by human dental pulp cells increased significantly at 12 and 24 hours after exposure to P. intermedia and F. nucleatum, whereas L. casei extract exhibited low IL-8 production. The sonicated bacterial extracts did not significantly affect viability or total number of dental pulp cells.

A modified dip-slide test for microbiological risk in caries assessment.

This study was undertaken to evaluate the validity of a new modified dip-slide test kit for the estimation of salivary mutans streptococci, lactobacilli and Candida levels by comparison with the results obtained from conventional agar plate counts. Five milliliters of paraffin-stimulated saliva sample was collected from 118 6-12-year old children attending 4 primary schools in Thailand. Saliva was poured over a 3-compartment slide containing Mitis-Salivarius Bacitracin agar, Rogosa agar and Sabouraud dextrose agar. A slide was incubated in a 5% CO2 incubator for 48 hours. The results of the modified test kit were compared with those from the conventional saliva sampling with dilution method. The salivary mutans streptococci, lactobacilli and Candida counts obtained from the conventional methods were significantly correlated (p < 0.0001) with the modified dip-slide estimates of these organisms. (Kendall Tau = 0.71, 0.58 and 0.76, respectively). The correlation between a modified dip-slide test kit and the conventional method indicates that this new test kit is suitable for the screening of salivary mutans streptococci, lactobacilli and Candida levels in patients. The test is simple and would be useful for the early selection of patients for dental examination. In addition, it would be a valuable educational aid for the motivation and dietary counseling among children.

Lead in saliva and its relationship to blood in the residents of Klity Village in Thailand.

Lead concentrations in whole blood and saliva were examined in 16 females and 13 males living in Klity village, a highly contaminated area from lead mining, Thailand. The geometric mean for the lead content in the blood was 24.03 microg/dl (range 11.80-46.60 microg/dl) while the lead content in the saliva was 5.69 microg/dl (range 1.82-25.28 microg/dl). No significant differences were found between the concentrations of lead in blood and saliva in relation to the age of the subject. Males were found to have higher blood lead levels than females. The coefficient of correlation gamma between salivary and blood lead levels was -0.025. Our data suggests that saliva is not suitable material for biological monitoring with respect to lead exposure.

Induction of type I collagen and osteocalcin in human dental pulp cells by retinoic acid.

Retinoic acid has been known to play a key role in the regulation of bone cell differentiation and function. The effects of retinoic acid on human dental pulp cells, which contain several characteristics similar to those of bone cells, has yet to be elucidated extensively. The effects of retinoic acid on human dental pulp cells in terms of type I collagen and osteocalcin induction were investigated in vitro. Dental pulp cells obtained from the teeth of young patients (age between 18-22 years) were cultured and subsequently treated with various concentrations of retinoic acid (0, 10(-7), 10(-6), 10(-5) M) in serum-free DMEM. At different time intervals (8, 12 and 24 hours), the levels of type I collagen and osteocalcin secreted were determined using Type I Procollagen C-Peptide and Gla-type Osteocalcin EIA kits, respectively. Induction effects were evaluated using analysis of variance and the Duncan's multiple rank test. Retinoic acid at concentrations of 10(-5), 10(-6), 10(-7) M was able to induce type I collagen and osteocalcin production in human dental pulp cells within 12 hours of exposure. Dose-dependent induction was observed only after 24 hours. A two-fold increase in osteocalcin level was detected after exposed to 10(-5) M retinoic acid within 24 hours. Our data suggest that retinoic acid at concentrations of 10(-5), 10(-6), 10(-7) M has the ability to induce type I collagen and osteocalcin secretions in human dental pulp cells in vitro.

Salivary mutans streptococci and lactobacilli after self arresting caries treatment.

Previous studies have shown that Self Arresting Caries Treatment (SACT) may be a useful technique to arrest active caries in Thai preschool children in remote areas. The objective of this study was to examine the effect of SACT on the levels of salivary mutans streptococci and lactobacilli in Thai children. Eighty-two subjects aged 4-6 years from primary school in a remote area of Thailand were recruited for this study. Each subject has at least two pairs of interproximal carious cavities between the first and second deciduous molars, and no history of severe pain. The subjects were divided into the control and SACT groups. The SACT technique was performed by removing food remnants and the surrounding overhang enamel of the carious cavity walls to enlarge the space between the carious cavities. The levels of mutans streptococci and lactobacilli in paraffin-stimulated whole saliva were determined at baseline, 2 and 4 months after treatment compared to those of the control group by the spatula method. The bacterial growth score was divided into three categories: 1 = 0-20 CFU, 2 = 21-100 CFU, and 3 > or = 100 CFU. At baseline, no differences in the mutans streptococci and lactobacilli scores were found between the SACT and control groups (p>0.05). Lower salivary lactobacilli levels were found in the SACT group on follow-up at 2 and 4 months (p<0.05), whereas the mutans streptococci level remained unchanged during these time periods. Our data confirms that the SACT technique can promote a self cleansing condition for interproximal lesions and can reduce the retentive sites in the oral cavity due to a reduction in salivary lactobacilli.

The effect of xylitol chewing gum on mutans streptococci in saliva and dental plaque.

Dental caries are one of the most common infectious diseases occurred in Thai children. The chewing of xylitol, sorbitol, and even sugar gum has been suggested to reduce caries rates. The aim of the present investigation was to evaluate the effect of xylitol chewing gum on mutans streptococci (MS) in saliva and dental plaque. Ninety-one children, aged 10-12 years, with more than 1 x 10(5) MS per milliliter of saliva were included in this study. They were divided into three groups balanced according to their MS counts at baseline: one control group (no supervised gum use), and two xylitol groups (supervised 55% and 100% xylitol gum use). Whole saliva and pooled plaque samples were obtained after 90 school days. When comparing the MS counts between the groups, those chewing 100% and 55% xylitol gum showed significant reductions (p < 0.025), but a dose response effect was not demonstrated. Chewing 100% xylitol gum caused significant reductions on salivary MS scores (p < 0.025) which was little different from the 55% xylitol group. The results suggest that the use of xylitol chewing gum can reduce the levels of MS in plaque and saliva.

Effect of fluoride on human dental pulp cells in vitro.

Human dental pulp cells were cultured in fluoride containing medium of various concentrations (0, 1, 2, 5, 10, 20, 25, 30, 40, 50, 60 and 80 ppm) in order to study the biological effect on the cells' proliferation and alkaline phosphatase (ALP) activities. It was found that fluoride at 5 ppm concentration significantly stimulated cell proliferation and ALP activity between 24 and 48 hours after exposure whereas at higher concentrations (40 - 80 ppm), fluoride significantly inhibited cell growth and ALP activity after 48 hours (Student's t test). The maximum effect was around 80 ppm. These observations suggest that fluoride, if used at a low concentration, may be a useful therapeutic agent for the treatment of pulpal disease by means of stimulating the proliferation and differentiation of dental pulp cells. At higher concentrations, it will have negative effects on this kind of cell.