Letter Cite This: ACS Biomater. Sci. Eng. XXXX, XXX, XXX−XXX
pubs.acs.org/journal/abseba
Rapid Coating Process Generates Omniphobic Dentures in Minutes to Reduce C. albicans Biofouling Anna Waterhouse,†,‡ Daniel C. Leslie,† Kayla Lightbown,† Daniel Antonoff,† Shanda Lightbown,† Nikolaos Dimitrakakis,† Julia B. Hicks-Berthet,† Cheyene N. Leslie,† Michael Super,† Donald E. Ingber,*,†,§,⊥,△ and Marc B. Ackerman*,∥,#,△
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†
Wyss Institute for Biologically Inspired Engineering at Harvard University, 3 Blackfan Circle, Boston, Massachusetts 02115, United States § Vascular Biology Program, Boston Children’s Hospital and Harvard Medical School, Boston, Massachusetts 02115, United States ⊥ John A. Paulson School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts 02138, United States ∥ Department of Dentistry, Boston Children’s Hospital, Boston, Massachusetts 02115, United States # Department of Developmental Biology, School of Dental Medicine, Harvard University, Boston, Massachusetts, United States S Supporting Information *
ABSTRACT: Localized infections caused by biofilm formation on dentures pose a serious health risk for patients, especially the elderly, as they can lead to complications such as pneumonia. Daily enzymatic denture cleaners do not fully prevent biofilm formation on dentures. Here we developed a rapid coating process to apply a liquid repellent surface to dentures in ∼5 min and demonstrated a significant 225-fold reduction of Candida albicans adhesion over 6 days, compared to uncoated dentures. This rapid coating process could be applied to dentures and other dental devices chair-side and allow the research community to quickly and easily generate ominphobic surfaces. KEYWORDS: denture, fungi, biofilm, immobilized liquid surface, medical devices, biomaterials
Candida albicans (C. albicans) is the most ubiquitous fungal species in the human microbiome.1 Although C. albicans and other Candida species are a component of the healthy microbiota colonizing the gastrointestinal tract, oral cavity and skin, alterations in other species of the host microbiome caused by administration of antibiotics, a diminution of the host immune response or pH changes may initiate C. albicans overgrowth and cause infection.2−4 C. albicans biofilms can form on medical devices such as urinary and venous catheters, cardiac pacemakers, prosthetic heart valves, joint prostheses, contact lenses, and dentures.5 Localized Candidal infections in the oral cavity are seen up to ∼70% of denture wearers and are termed denture stomatitis (DS). C. albicans is implicated as the primary causative species in this disease.6 A growing body of work suggests DS is a polymicrobial disease in which bacterial and fungal interplay may be responsible for disease pathogenesis.7−9 Edentulousness (lack of teeth) is estimated to affect between 7 and 69% of the world’s adult population based on geographic region.10 The elderly are the largest subset of the population using complete dentures. C. albicans can readily adhere to denture base acrylic and imbed in microscopic fissures on the acrylic surface. These C. albicans biofilms pose a significant health risk to patients;11 for example, C. albicans has been reported to be one of the fungi that may induce aspiration © XXXX American Chemical Society
pneumonia in the lungs of the elderly via secretions from the oral cavity.12,13 Clinically recommended, daily denture hygiene is intended to remove Candida biofilm from the denture base resin. Commercially available denture cleaners utilize neutral enzymatic peroxides that interfere with biofilm development without impacting the surface properties of the resin.14,15 However, a study examining daily use of a commercially available denture cleaner reported that daily use of enzymatic denture cleaners is not successful in halting the development of C. albicans biofilms on denture base resins.16 Furthermore, there is currently no biofilm-specific drug for C. albicans which makes treatment of a biofilm-based infection exceedingly difficult.17 An alternative approach to reducing or eliminating Candida biofilms on dentures is the modification of denture base resin with compounds known to have antifungal and antimicrobial properties. One in vitro study investigated the addition of zinc oxide nanoparticles (ZnO-NP’s) into poly(methyl methacrylate) (PMMA) resin. The authors found that with increasing Received: October 4, 2018 Accepted: December 21, 2018
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DOI: 10.1021/acsbiomaterials.8b01214 ACS Biomater. Sci. Eng. XXXX, XXX, XXX−XXX
Letter
ACS Biomaterials Science & Engineering concentration of ZnO-NP’s, the antifungal activity increased. However, they cautioned that zinc oxide release from the denture at the investigated concentrations may induce mucosal irritation.18 Another in vitro study added silver nanoparticles to denture base resin and examined C. albicans adhesion and biofilm formation. The authors found that nanosilver at higher concentrations appears to demonstrate an antifungal property and an inhibitory effect on C. albicans adhesion.19 These higher concentrations may also increase toxicity in the oral environment. New approaches to combat biofilm formation focus solely on preventing adhesion of microbes that lead to colonization and biofilm formation, without the use of potentially harmful products. Liquid-immobilized surface coatings are an example of this method.20 This technology was originally inspired by the Nepenthes pitcher plant, which employs a layer of liquid water held in place on a nanostructured surface to generate a low friction surface which inhibits attachment of insects.21 Liquid-immobilized surfaces have been used to repel surface biological fouling and represent a promising surface coating for medical applications.20 Specifically, they have shown reduced bacterial adhesion and biofilm formation under varying conditions in vitro and in vivo.20,22−24 The TLP layer consists of an immobilized tethered perfluorocarbon (TP) that is overlaid by a thin liquid perfluorocarbon (LP) layer, which forms a liquid film that generates an omniphobic surface capable of repelling both hydrophobic and hydrophilic liquids, including living microbial cells such as bacteria, fungi and yeast (Figure 1a). Relevant for denture materials, the tethered-liquid perfluorocarbon (TLP) surface coating was applied to PMMA and successfully prevented both thrombus and biofilm formation in vitro.25 In this current study, we developed a rapid coating process for TLP, reducing surface treatment time from >16 h to ∼5 min, to allow application to denture acrylic and fully formed dentures to prevent C. albicans adhesion and subsequent biofilm development. Previously developed methods for coating materials with TLP took approximately >70 min to 17 h and involved 4 basic steps: (1) creating reactive groups on the surface by exposing the materials to low-pressure oxygen plasma25,26 or phosphoric acid.24 (2) Creating a perfluorinated surface by immersing the surface in a fluorinated silane and ethanol solution24,25 or vapor26 to create the TP layer, (3) sometimes curing it overnight and (4) then applying the LP before use (Figure 1b). These methods required substantial, time-consuming incubation and/or washing steps, complicating the process of translating this coating for the dental market. Via these methods, slippery coatings would need to be applied to dentures during the denture manufacturing process and would require specific laboratory equipment. Given the denture fitting process can require multiple visits and sizing adjustments, it would be beneficial for a slippery coating to be applied to the final denture in the dentists’ facility, chair-side, once the fitting process is complete. We developed a rapid TLP coating process using clinically available dental reagents currently used in the oral cavity (Figure 1b). This method took advantage of clinically used porcelain repair kits which are include phosphoric acid and silane products and are carried out rapidly in the dentists’ office. We activated the surface with a 20 s treatment of phosphoric acid (20 s per surface/side). After rinsing with water and ethanol, a silane mixture was applied for 1 min, which consisted of the fluorinated silane used to make TLP,
Figure 1. Rapid TLP coating process. (a) Schematic showing TLP applied to dental materials repelling pathogen adhesion (modified from Leslie et al.25). (b) Methods showing steps and time frames for coating materials with fluorinated silanes that have been previously published, including Leslie et al., ∼17 h;25 Yin et al., ∼72 min;24 and Badv et al., ∼17.5 h26 compared to the current method presented in this manuscript, which takes approximately 5 min.
and the clinically used silane bonding agent from a porcelain repair kit. Finally, the surfaces were rinsed, air-dried and coated with LP (by simple dip coating) before use. This allowed the full TLP process to be completed within approximately 5 min (Figure 1b), compared to the previous shortest method using phosphoric acid, that took >70 min to apply a slippery coating directly to tooth enamel.24 Our rapid TLP coating method could be rapidly translated as it uses existing dental materials and reagents from Porcelain repair kits, which have been developed to be used in the oral cavity and work quickly. The dental silane contains 5−15% methacryloxy propyl trimethoxysilane, 92% isopropyl alcohol and
