Cytotoxicity of three commercial mouthrinses on extracellular matrix metabolism and human gingival cell behaviour.

This study evaluated the effects of commercially available antiseptic mouthrinses on human gingival fibroblast and keratinocyte behaviour and metabolism. Three mouthrinses containing essential oil (EO), chlorhexidine (CHX) and amine fluoride/stannous fluoride (AFSF), were tested in an in vitro study. Human gingival fibroblasts and keratinocytes were washed with 10% or 30% concentration of the commercial mouthrinses and their effects on cell adhesion and proliferation were investigated as well as the specific gene expression of markers involved in oral mucosa metabolism. As markers of cell metabolism, type I and IV collagens, laminin, fibronectin, fibromodulin and integrins were studied with real-time PCR. Moreover, interleukin-1 secretion, one of the major pro-inflammatory cytokines, was evaluated. The results showed that CHX significantly reduced fibroblast and keratinocyte substrate adhesion capacities and CHX and EO inhibited cell proliferation better than AFSF rinse. The gene expression of several matrix components and cell adhesion receptors was downregulated in cells washed with CHX and EO compared with those washed with AFSF rinse. In conclusion, the AFSF mouthrinse does not induce or induces to a lesser extent the onset of irritation and/or cytotoxicity than CHX or EO. These findings and those of future studies will enable us to gain further insight into the clinical significance and effects of commercial mouthrinses. Pending further investigations, clinicians should be aware of the potentially adverse effects of mouthrinses and warn their patients against making improper use of these products.

The peritoneal explant test for evaluating tissue tolerance to mouthrinses.

The tissue cultures of explants of neonatal rat peritoneum have been demonstrated to be a sensitive test for tissue compatibility with wound antiseptics. The present study investigated the suitability of this method to assess the relative toxicity of mouthrinses to tissue. Mouthrinses containing 0.1% chlorhexidine (Chlorhexamed Fluid 0.1%) (A), 0.3% triclosan (Colgate) (B), essential oil in ethanolic solution (Listerine) (C), and amine/stannous fluoride (Meridol) (D) were tested at use concentration and in dilutions of 10, 1, and 0.1% with exposure times of 1, 10, and 30 min, respectively. The mouthrinses (test) and Ringer's solutions (control) were applied to opened rat peritoneum. After thorough irrigation with Ringer's solution, a piece of peritoneum was removed and 1 x 1 mm explants were cut. The explants were cultivated with a bovine serum culture medium in 24-well plates at 37 degrees C in a CO2 incubator (95% air, 5% CO2). After 10 days, the tissue proliferation for the explants was assessed by a stereo microscope at 10x magnification after ethanol fixing and hemalaun staining. With 24 grafts per test, the proliferation rate was calculated relative to a control, which was run for each mouthrinse and concentration/time combination. Data were analyzed using ANOVA (SPSS 11.0) and post-hoc paired t test. Statistical significance of all correlations was tested by setting the significance level at p < 0.05. At most concentrations, D caused significantly less tissue damage than A or B. There was no difference between C and A or C and B at 100%. However, the toxicity of C was significantly less than A or B at 10, 1, and 0.1%. C and D behaved similarly except for the 10% (30 min) and the 1% (10 min) solutions in which C was significantly less toxic. We concluded that the rat peritoneum explant test was demonstrated to be a sensitive test to assess the relative toxicity of mouthrinses to tissue.

Amine fluoride gel affects the viability and the generation of superoxide anions in human polymorphonuclear leukocytes: an in vitro study.

Amine hydrofluorides are widely used to prevent caries. As an acidulated gel, they were also studied for their applicability to reduce pathogenic bacteria in periodontal pockets. We assessed the toxicity of this pharmaceutical amine hydrofluoride preparation on human polymorphonuclear leukocytes in vitro by measuring Trypan blue exclusion and the generation of superoxide anions (O2) by the chemotactic peptide N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP) after a 3-min contact with gel. Depending on the experimental conditions, gel dilutions up to 1.3 x 10(4) resulted in an increase in Trypan blue-colored cells and liberation of beta-glucuronidase. Dilutions between 3 x 10(4) and 1 x 10(5) augmented the fMLP-mediated O2- generation, which could be prevented by Ca2+ chelation with BAPTA-AM (1,2'-bis (o-aminophenoxyethane-N.N.N'.N'-tetraacetic acid tetra (acetoxymethyl) ester) and ethyleneglycoltetraacetic acid (EGTA) or inhibition of protein kinase C (PKC) with staurosporine and bisindolylmaleimide I. respectively. Compared with data published on the minimal inhibitory concentration for periodontal pathogenic bacteria, the cytotoxicity of amine hydrofluorides on eukaryotic cells is much greater and thus of consequence for their clinical use.

Assessment of the stannous fluoride and phytic acid effect in the yeast Saccharomyces cerevislae.

Stannous fluoride (SnF2) is a powerful reducing agent in 99mTc-labelled radiopharmaceuticals for nuclear medicine procedures. SnF2 may enhance reactive oxidative species (ROS) in prokaryotic cells. Phytic acid (PA) is a wide-ranging regulator of many important cellular functions such as intracellular regulations of surface receptions channels and it is known to have antioxidant and chelating properties. In order to analyze whether membrane transporters of the facilitator or the ABC type (SNQ1 and SNQ2) have an influence on Sn2+ toxicity in yeast we used the respective mutants and compared their responses to the wild type (WT). Since ABC transporters are YAP1p transcription activator inducible, we included a yap1 mutant in our Sn2+ toxicity assay. Finally, we tested the PA influence on Sn2+ toxicity in these strains. Yeast cells in stationary growth phase were exposed to different concentrations of SnF2 (ranging from 2 to 6 mg/ml) and PA (0.1 M) for one hour. The snq1 mutant exhibited the highest sensitivity to SnF2 while the snq2 and snq3/yap1 mutants had an equally intermediate sensitivity. The presence of PA was not able to produce a significant protection against the cytotoxicity of SnF2. This is probably due to its reduced chelating power in complex liquid media Our results with yeast support the genotoxic effects described for SnF2 in bacteria andindicate that the biological effect of this reducing agent could be related to the generation of reactive oxygen species.

Comparison of sodium and stannous fluoride nephrotoxicity.

Single i.p. injections of 0.5 mmol F-/kg and repeated i.p. injections of 0.25 mmol F-/kg as NaF or SnF2 to male rats showed that both compounds produced early lesions of kidney tubule demonstrated by an intense increase of urine gamma-glutamyl transferase (GGT). Disturbance of kidney function with an increase of diuresis and phosphaturia and a decrease of natriuria and kaliuria were more marked in SnF2-treated animals as was the decrease of urine GGT after a few days. These results indicate that tin nephrotoxicity adds to that of fluoride and antagonizes tubule regeneration.

[Comparison of embryotoxic effects of inorganic fluorides]. / Vergleich der embryotoxischen Wirkungen anorganischer Fluoride.

A comparison of the toxicity of SnF2, BaF2, KF, CaF2, and MgF2 shows that the hard-to-dissolve fluorides CaF2 and MgF2 are absolutely safe. The other fluorides tested showed no toxic effects in dosages relevant for caries prophylaxis and with oral application.