Dynamic Communication Systems Based on Soft Hydrogel Microbial Modulators

New hierarchical hydrogel architectures have been pioneered to create multi-layered soft hydrogels in a concentric fashion as providing fundamental insights at the molecular level. Yet the strategies might not be standardized because different design considerations are required due to a wide range of chemical processes involved. At small scales, the fabrication of micro-size architectures requires higher reaction rates than that of macroscopic objects, which causes agglomeration between gels in a reaction bath. Due to the agglomeration issue, there has been an absence of reliably scale-down models for mass production. Only a few methods have been introduced to produce a single multi-layered capsule with diameters between 4 and 20 mm. Even with predictive mechanisms in various hydrogels, challenges exist in both mass production and scaling-down of the size of hydrogel architectures, and multi-layered techniques for cell encapsulation studies are even sparse.<br/>Soft hydrogel-based modalities have been designed to study cell-cell communication systems including bacterial quorum-sensing by confining genetically engineered microorganisms, which allows the diffusion of solutes<i> </i>across the hydrogel matrix<i>. </i>The communication modes typically use chemical signals mediated by intra/extracellular messenger molecules which not only governs the basic activity of cells but also regulates their physiological functions in a milieu. Despite numerous efforts, technical obstacles to create confined architectures for cell communications has remained elusive. Above all, an effective platform for bacterial confinement should be to retain functionally engineered cells inside the hydrogel architectures without disruption of the structures. Herein, microbial colonization occurs ensuring cell longevity and production of designed signaling molecules for quorum sensing through diffusive biological gels. However, to the best of our knowledge, most of the previous strategies failed to demonstrate an effective platform to confine bacterial communities effectively. In some cases, to constrain cell leakage, cytotoxic processes and/or chemicals such as acrylamide polymerization were utilized to form a size-exclusive barrier, which is not suitable for bacterial biocontainment. Even if bacterial cells could be contained and are able to survive in adverse conditions, chemical toxicity and cell damage are inevitable. Unfavorable procedures lead to rapid cell death within two days in such confined environments, impeding the potential to study the basic dynamics of cellular responses and interactions between microbial communities.<br/>To address the challenges, we have designed and developed a biocompatible confinement model utilizing alginate-based hierarchical architectures. Importantly, our methodology can be used to develop hydrogel architectures that can be reliably scale-down and controlled for mass production. The confinement technology conceived has excellent permeable properties to allow diffusion of signaling molecules and essential nutrients, including oxygen, and byproducts, to maintain their physiological function for a prolonged period. Numerous small ecological systems in a hydrogel environment can be generated to serve as independent microbial modulator with direct or indirect activation of a response regulator by stimuli, including signaling molecules. Specifically, we demonstrate three representative models (<i>i.e.</i>, bead-to-bead, stratified communication and bacteria-mammalian cell interaction system) based on hydrogel modulators by entrapping specific microorganisms. This platform will have significant impact in intra/multicellular communications by evaluating interspecies’ signaling in a highly specific manner.