Effect of surface stiffness in initial adhesion of oral microorganisms under various environmental conditions.

Biofilms are three-dimensional structures formed as a result of microorganism's adhesion on a biotic or abiotic surface. Once a biofilm is established, it is onerous to eradicate it or kill the pathogens therein. Thus, targeting the microbial adhesion process, the initial stage of biofilm formation, is a reasonable approach to avoid challenges associated with subsequently formed biofilms. While many properties of interacting material that play significant roles in initial bacterial adhesion have been widely studied, the effect of surface stiffness on bacterial adhesion was relatively underexplored. In this study, we aimed to investigate the effect of surface stiffness on the adhesion of microbial species found in the oral cavity by employing representative oral bacteria, Streptococcus mutans and Streptococcus oralis, and the fungus, Candida albicans. We compared the adhesion behavior of these species alone or in combination toward various surface stiffness (0.06 - 3.01 MPa) by assessing the adhered and planktonic cell numbers at an early (4 h) adhesion stage under various carbon sources and the presence of conditioning film. Our data revealed that in general, a higher amount of microbial cells adhered to softer PDMS surfaces than stiffer ones, which indicates that surface stiffness plays a role in the adhesion of tested species (either single or co-cultured). This pattern was more obvious under sucrose conditions than glucose + fructose conditions. Interestingly, in monospecies, saliva coating did not alter the effect of surface stiffness on S. mutans adhesion while it diminished S. oralis and C. albicans adhesion. However, in the multispecies model, saliva coating rendered the percentage of all adhered microbes to varied PDMS not distinct. The data provide new insights into the role of surface stiffness on microbial mechanosensing and their initial adhesion behavior which may set a scientific foundation for future anti-adhesion strategies.

A dental implant-on-a-chip for 3D modeling of host-material-pathogen interactions and therapeutic testing platforms.

The precise spatiotemporal control and manipulation of fluid dynamics on a small scale granted by lab-on-a-chip devices provide a new biomedical research realm as a substitute for in vivo studies of host-pathogen interactions. While there has been a rise in the use of various medical devices/implants for human use, the applicability of microfluidic models that integrate such functional biomaterials is currently limited. Here, we introduced a novel dental implant-on-a-chip model to better understand host-material-pathogen interactions in the context of peri-implant diseases. The implant-on-a-chip integrates gingival cells with relevant biomaterials - keratinocytes with dental resin and fibroblasts with titanium while maintaining a spatially separated co-culture. To enable this co-culture, the implant-on-a-chip's core structure necessitates closely spaced, tall microtrenches. Thus, an SU-8 master mold with a high aspect-ratio pillar array was created by employing a unique backside UV exposure with a selective optical filter. With this model, we successfully replicated the morphology of keratinocytes and fibroblasts in the vicinity of dental implant biomaterials. Furthermore, we demonstrated how photobiomodulation therapy might be used to protect the epithelial layer from recurrent bacterial challenges (∼3.5-fold reduction in cellular damage vs. control). Overall, our dental implant-on-a-chip approach proposes a new microfluidic model for multiplexed host-material-pathogen investigations and the evaluation of novel treatment strategies for infectious diseases.

A 3D-Printed Customizable Platform for Multiplex Dynamic Biofilm Studies.

Biofilms are communities of microbes that colonize surfaces. While several biofilm experimental models exist, they often have limited replications of spatiotemporal dynamics surrounding biofilms. For a better understanding dynamic and complex biofilm development, this manuscript presents a customizable platform compatible with off-the-shelf well plates that can monitor microbial adhesion, growth, and associated parameters under various relevant scenarios by taking advantage of 3D printing. The system i) holds any substrate in a stable, vertical position, ii) subjects samples to flow at different angles, iii) switches between static and dynamic modes during an experiment, and iv) allows multiplexing and real-time monitoring of biofilm parameters. Simulated fluid dynamics is employed to estimate flow patterns around discs and shear stresses at disc surfaces. A 3D printed peristaltic pump and a customized pH measurement system for real-time tracking of spent biofilm culture media are equipped with a graphical user interface that grants control over all experimental parameters. The system is tested under static and dynamic conditions with Streptococcus mutans using different carbon sources. By monitoring the effluent pH and characterizing biochemical, microbiological, and morphological properties of cultured biofilms, distinct properties are demonstrated. This novel platform liberates designing experimental strategies for investigations of biofilms under various conditions.

Bimodal Nanocomposite Platform with Antibiofilm and Self-Powering Functionalities for Biomedical Applications.

Advances in microelectronics and nanofabrication have led to the development of various implantable biomaterials. However, biofilm-associated infection on medical devices still remains a major hurdle that substantially undermines the clinical applicability and advancement of biomaterial systems. Given their attractive piezoelectric behavior, barium titanate (BTO)-based materials have also been used in biological applications. Despite its versatility, the feasibility of BTO-embedded biomaterials as anti-infectious implantable medical devices in the human body has not been explored yet. Here, the first demonstration of clinically viable BTO-nanocomposites is presented. It demonstrates potent antibiofilm properties against Streptococcus mutans without bactericidal effect while retaining their piezoelectric and mechanical behaviors. This antiadhesive effect led to ∼10-fold reduction in colony-forming units in vitro. To elucidate the underlying mechanism for this effect, data depicting unfavorable interaction energy profiles between BTO-nanocomposites and S. mutans using the classical and extended Derjaguin, Landau, Verwey, and Overbeek theories is presented. Direct cell-to-surface binding force data using atomic force microscopy also corroborate reduced adhesion between BTO-nanocomposites and S. mutans. Interestingly, the poling process on BTO-nanocomposites resulted in asymmetrical surface charge density on each side, which may help tackle two major issues in prosthetics-bacterial contamination and tissue integration. Finally, BTO-nanocomposites exhibit superior biocompatibility toward human gingival fibroblasts and keratinocytes. Overall, BTO-embedded composites exhibit broad-scale potential to be used in biological settings as energy-harvestable antibiofilm surfaces.

Implication of Surface Properties, Bacterial Motility, and Hydrodynamic Conditions on Bacterial Surface Sensing and Their Initial Adhesion.

Biofilms are structured microbial communities attached to surfaces, which play a significant role in the persistence of biofoulings in both medical and industrial settings. Bacteria in biofilms are mostly embedded in a complex matrix comprised of extracellular polymeric substances that provide mechanical stability and protection against environmental adversities. Once the biofilm is matured, it becomes extremely difficult to kill bacteria or mechanically remove biofilms from solid surfaces. Therefore, interrupting the bacterial surface sensing mechanism and subsequent initial binding process of bacteria to surfaces is essential to effectively prevent biofilm-associated problems. Noting that the process of bacterial adhesion is influenced by many factors, including material surface properties, this review summarizes recent works dedicated to understanding the influences of surface charge, surface wettability, roughness, topography, stiffness, and combination of properties on bacterial adhesion. This review also highlights other factors that are often neglected in bacterial adhesion studies such as bacterial motility and the effect of hydrodynamic flow. Lastly, the present review features recent innovations in nanotechnology-based antifouling systems to engineer new concepts of antibiofilm surfaces.

Synergism of Streptococcus mutans and Candida albicans Reinforces Biofilm Maturation and Acidogenicity in Saliva: An In Vitro Study.

Early childhood caries, a virulent-form of dental caries, is painful, difficult, and costly to treat that has been associated with high levels of Streptococcus mutans (Sm) and Candida albicans (Ca) in plaque-biofilms on teeth. These microorganisms appear to develop a symbiotic cross-kingdom interaction that amplifies the virulence of plaque-biofilms. Although biofilm studies reveal synergistic bacterial-fungal association, how these organisms modulate cross-kingdom biofilm formation and enhance its virulence in the presence of saliva remain largely unknown. Here, we compared the properties of Sm and Sm-Ca biofilms cultured in saliva by examining the biofilm structural organization and capability to sustain an acidic pH environment conducive to enamel demineralization. Intriguingly, Sm-Ca biofilm is rapidly matured and maintained acidic pH-values (~4.3), while Sm biofilm development was retarded and failed to create an acidic environment when cultured in saliva. In turn, the human enamel slab surface was severely demineralized by Sm-Ca biofilms, while there was minimal damage to the enamel surface by Sm biofilm. Interestingly, Sm-Ca biofilms exhibited an acidic environment regardless of their hyphal formation ability. Our data reveal the critical role of symbiotic interaction between S. mutans and C. albicans in human saliva in the context of pathogenesis of dental caries, which may explain how the cross-kingdom interaction contributes to enhanced virulence of plaque-biofilm in the oral cavity.

A Dynamic Culture Method to Produce Ovarian Cancer Spheroids under Physiologically-Relevant Shear Stress.

The transcoelomic metastasis pathway is an alternative to traditional lymphatic/hematogenic metastasis. It is most frequently observed in ovarian cancer, though it has been documented in colon and gastric cancers as well. In transcoelomic metastasis, primary tumor cells are released into the abdominal cavity and form cell aggregates known as spheroids. These spheroids travel through the peritoneal fluid and implant at secondary sites, leading to the formation of new tumor lesions in the peritoneal lining and the organs in the cavity. Models of this process that incorporate the fluid shear stress (FSS) experienced by these spheroids are few, and most have not been fully characterized. Proposed herein is the adaption of a known dynamic cell culture system, the orbital shaker, to create an environment with physiologically-relevant FSS for spheroid formation. Experimental conditions (rotation speed, well size and cell density) were optimized to achieve physiologically-relevant FSS while facilitating the formation of spheroids that are also of a physiologically-relevant size. The FSS improves the roundness and size consistency of spheroids versus equivalent static methods and are even comparable to established high-throughput arrays, while maintaining nearly equivalent viability. This effect was seen in both highly metastatic and modestly metastatic cell lines. The spheroids generated using this technique were fully amenable to functional assays and will allow for better characterization of FSS's effects on metastatic behavior and serve as a drug screening platform. This model can also be built upon in the future by adding more aspects of the peritoneal microenvironment, further enhancing its in vivo relevance.

Cerium Oxide Nanoparticles: A Brief Review of Their Synthesis Methods and Biomedical Applications.

Cerium oxide nanoparticles (CeNPs) exhibit antioxidant properties both in vitro and in vivo. This is due to the self-regeneration of their surface, which is based on redox-cycling between 3+ and 4+ states for cerium, in response to their immediate environment. Additionally, oxygen vacancies in the lattice structure allow for alternating between CeO2 and CeO2-x during redox reactions. Research to identify and characterize the biomedical applications of CeNPs has been heavily focused on investigating their use in treating diseases that are characterized by higher levels of reactive oxygen species (ROS). Although the bio-mimetic activities of CeNPs have been extensively studied in vitro, in vivo interactions and associated protein corona formation are not well understood. This review describes: (1) the methods of synthesis for CeNPs, including the recent green synthesis methods that offer enhanced biocompatibility and a need for establishing a reference CeNP material for consistency across studies; (2) their enzyme-mimetic activities, with a focus on their antioxidant activities; and, (3) recent experimental evidence that demonstrates their ROS scavenging abilities and their potential use in personalized medicine.