Comparative study of hyperpure chlorine dioxide with two other irrigants regarding the viability of periodontal ligament stem cells

Clinical Oral Investigations https://doi.org/10.1007/s00784-020-03618-5 ORIGINAL ARTICLE Comparative study of hyperpure chlorine dioxide with two other irrigants regarding the viability of periodontal ligament stem cells Orsolya Láng 1 & Krisztina S. Nagy 2,3 & Julia Láng 1 & Katalin Perczel-Kovách 2,4 & Anna Herczegh 5 & Zsolt Lohinai 5 & Gábor Varga 2 & László Kőhidai 1 Received: 16 June 2020 / Accepted: 1 October 2020 # The Author(s) 2020 Abstract Objectives Periodontal ligament stem cells (PDLSCs) have an underlined significance as their high proliferative capacity and multipotent differentiation provide an important therapeutic potential. The integrity of these cells is frequently disturbed by the routinely used irrigative compounds applied as periodontal or endodontic disinfectants (e.g., hydrogen peroxide (H2O2) and chlorhexidine (CHX)). Our objectives were (i) to monitor the cytotoxic effect of a novel dental irrigative compound, chlorine dioxide (ClO2), compared to two traditional agents (H2O2, CHX) on PDLSCs and (ii) to test whether the aging factor of PDLSC cultures determines cellular responsiveness to the chemicals tested. Methods Impedimetry (concentration-response study), WST-1 assays (WST = water soluble tetrazolium salt), and morphology analysis were performed to measure changes in cell viability induced by the 3 disinfectants; immunocytochemistry of stem cell markers (STRO-1, CD90, and CD105) measured the induced mesenchymal characteristics. Results Cell viability experiments demonstrated that the application of ClO2 does not lead to a significant decrease in viability of PLDSCs in concentrations used to kill microbes. On the contrary, traditional irrigants, H2O2, and CHX are highly toxic on PDLSCs. Aging of PLDSC cultures (passages 3 vs. 7) has characteristic effects on their responsiveness to these agents as the increased expression of mesenchymal stem cell markers turns to decreased. Conclusions and clinical relevance While the active ingredients of mouthwash (H2O2, CHX) applied in endodontic or periodontitis management have a serious toxic effect on PDLSCs, the novel hyperpure ClO2 is less toxic providing an environment favoring dental structure regenerations during disinfectant interventions. Keywords Chlorine dioxide . Dental stem cells . PDLSC . Viability . Toxicity . Hydrogen peroxide . Chlorhexidine Introduction Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00784-020-03618-5) contains supplementary material, which is available to authorized users. * László Kőhidai 1 Department of Genetics, Cell- and Immunobiology, Semmelweis University, Budapest, Hungary 2 Department of Oral Biology, Semmelweis University, Budapest, Hungary 3 Laboratory of Nanochemistry, Department of Biophysics and Radiation Biology, Semmelweis University, Budapest, Hungary 4 Department in Community Dentistry, Semmelweis University, Budapest, Hungary 5 Department of Conservative Dentistry, Semmelweis University, Budapest, Hungary For the last two decades, the discovery of the dental stem cells (DSCs) has opened new perspectives in regenerative dentistry and medicine. The first source of these oral cells with mesenchymal stem cell (MSC) properties was the human dental pulp [1], and subsequently, four more types of DSCs were gained from different tooth-related tissues, such as pulp of exfoliated deciduous teeth [2], periodontal ligament (PDL) [3], dental follicle [4], and apical papilla [5]. The common feature of these DSCs is that they exhibit fibroblast-like morphology with good proliferative potential and fulfill the minimal criteria of MSC characteristics, such as adherence to plastic surface, expression of certain surface antigens (e.g., more than 95% of the cell population express CD73, CD90, and CD105, and less than 2% express hematopoietic markers), and capacity for multipotent differentiation in vitro [6]. Recent studies Clin Oral Invest provided evidence for a wide range of plasticity of these SCs and their ability to repair tooth-related tissues or bone in vivo. Additionally, as DSCs are easily accessible and lack strict ethical concern conversely from their embryonic counterparts, they represent favorable tools also for the therapy of neurodegenerative disorders (e.g., Alzheimer or Parkinson diseases) or cardiac ischemia [7, 8]. Of the abovementioned DSCs, the PDLSCs are of great significance both in theoretical and practical aspects. Due to the lack of consensus criteria defining dental stem cells based on the surface antigen expression pattern, in most of the studies, PDLSCs are characterized by positivity for MSC markers. PDLSCs were found to express the STRO-1 antigen [3, 9]— identified first in bone marrow stromal cells [10]—and other MSC markers, such as CD13, CD29, CD44, CD59, CD90, and CD105 [7]. However, some investigation also revealed embryonic stem cell marker positivity (e.g., Oct-4) of these cells [9]. PDLSCs exhibit multipotent differentiation capacity. In vitro, these cells are able to differentiate into osteogenic, adipogenic [3], chondrogenic [11], neurogenic [12], and myogenic [13] lineages. In vivo, they have fundamental importance in the physiology of PDL, which does not only anchor the cementum covering the root to the alveolar bone but also contributes to its nutrition, homeostasis, and repair. The significant regenerative potential of PLDSCs allows these cells to contribute the spontaneous or medically induced restorative mechanisms of the periodontal region [14, 15]. The differentiation potential of these stem cells is similar to pericytes, while their immunomodulatory character is also well described [16, 17]. The integrity of these cells in the PDL is vital for the whole life of the tooth. One of the longstanding goals of dental care is to keep the periodontium in good health and to reconstruct it when destroyed by the periodontal disease. Therefore, it is of paramount importance to know the effect of the disinfectant substances used routinely in the oral cavity on the physiological processes of these stem cells. In conventional dental care, irrigative agents are frequently used to eliminate the bacteria from different regions, e.g., outer root surface in case of periodontal or root canal surface in endodontic treatment. Due to the multispecies composition of the biofilm, the effective antibacterial irrigative have a relatively broad spectrum and multiple intracellular targets, which reduce the frequency of resistant cases. Topical antiseptics, such as chlorhexidine (CHX) and hydrogen peroxide (H2O2), which are routinely applied as disinfectants in dentistry, correspond to these criteria. However, antiseptics, unlike antibiotics, are potentially toxic not only to the infectious microbes but to the host cells as well. Recently, a well-known biocide, chlorine dioxide (ClO2), has been invented in the dental care, as its application was suggested for disinfection of the air o (...truncated)