Real-Time Analysis of Antiproliferative Effects of Mouthwashes Containing Alcohol, Sodium Fluoride, Cetylpyridinium Chloride, and Chlorhexidine In Vitro

Abstract

Objectives. In this study, the cytotoxic responses of six different over-the-counter mouthwashes on L929 cells were analyzed by two different techniques: the traditional colorimetric tetrazolium-based reduction assay (MTT) and the modern impedance-based real-time cell analysis (RTCA) system to investigate their biocompatibility in vitro. Thus, the investigation of the antiproliferative effects of the specified materials via different techniques is vital to reach this goal. Materials and Methods. First, L929 mouse fibroblasts were exposed to the dilutions of mouthwashes for 2 minutes. After incubation, the tetrazolium reduction method was used to assess the metabolic viability of cells measured by colorimetric MTT assay and morphological inspection of cells was performed via phase-contrast microscopy. Furthermore, the effect of each mouthwash on the proliferation, morphology, and adhesion of L929 cells was monitored continuously by a noninvasive and label-free RTCA system for 140 h. Results. Our data showed that all of the mouthwashes had varying cytotoxic effects on fibroblasts compared to the control group in MTT assay. In addition to that, RTCA technology has provided the growth kinetic profiles that can be used to analyze if the treatment is causing antimitotic or DNA-damaging effect on cells. Thus, analysis via this system can tell us the mechanism of toxicity behind the cell growth inhibition in vitro. Here, we found that only mouthwash 1 moderately maintained the viability of the L929 cells, yet displaying antimitotic effects and the other mouthwashes (mouthwash 2-mouthwash 6) showed toxicity via DNA-damaging effects. Conclusions. Of the six types of mouthwash tested, the most biocompatible result was obtained from a mouthwash containing alcohol (i.e., mouthwash 1). On the other hand, sodium fluoride- (NaF-) and cetylpyridinium chloride- (CPC-) containing mouthwash (i.e., mouthwash 2) showed the most cytotoxic effect.

Effect of Commercial Children’s Mouthrinses and Toothpastes on the Viability of Neonatal Human Melanocytes: An In Vitro Study

“…Our results of cytotoxicity by CPC containing mouthrinse and that of pure CPC as assessed by MTS assay that evaluates mitochondrial function indicates that compromised viability might be due to the inhibitory effects of CPC on mitochondrial complex I and mitochondrial O 2 consumption as shown previously [105,106]. An earlier report also showed that a 2 min exposure to CPC containing mouthrinse induced cytotoxicity to an L929 fibroblast cell line [107]. The immediate examination of mouthrinse cytotoxicity after a 2 min exposure was not conducted in our study.…”

“…The detergent SLS in toothpastes induced comparable cytotoxic effects in primary human keratinocytes from the skin and oral mucosa, suggesting that skin cells could be used as a substitute for oral cells [120]. Furthermore, the use of immortalized human skin keratinocytes (HaCaT cells) or mouse skin fibroblasts (L929 cells) has been common in previous studies that examined toothpastes [82], mouthwashes [107,121], or dental materials [122][123][124][125][126]. Based on these justifications, it can be inferred that utilizing skin melanocytes obtained from a neonatal lightly pigmented donor can serve as a model for oral melanocytes of children.…”

Efficacy of a mouthwash containing ε-poly-L-lysine, funme peptides and domiphen in reducing halitosis and supragingival plaque: a randomized clinical trial

“…Mouthwash has been recommended to be the essential for cleaning teeth and gum and freshening the breath. At present, commercial mouthwashes commonly contain some active ingredients including chlorhexidine, cetylpyridonium chloride, fluoride, and alcohol that act as antibacterial substances, however some users may encounter with cytotoxicity and various side effects caused by such active ingredients [24]. In this study, the newly developed antibacterial combination of domiphen and other two non-toxic AMPs namely ɛ-PL and FP was investigated for its inhibitory and bactericidal effects against some oral pathogens, and the mouthwash supplemented with this antibacterial combination in reducing The antimicrobial effects and safety of the efficacy ingredients used in this study have been evaluated in several studies.…”

Effect of Three Chlorhexidine-Based Mouthwashes on Human Gingival Fibroblasts: An In Vitro Study

“…Owing to its well-documented antibacterial activity, CHX is frequently employed to reduce oral bacterial load [20]. However, mouthwashes containing a high percentage of CHX have been associated with cytotoxic effects in vitro [12,16,44,45]. Therefore, usage of lower concentrations of CHX in combination with CPC have been proposed [24][25][26][27][28].…”