RESUMO
Alginate hydrogel beads are a common platform for generating 3D cell cultures in biomedical research. Simple methods for bead generation using a manual pipettor or syringe are low-throughput and produce beads showing high variability in size and shape. To address these challenges, we designed a 3D printed bead generator that uses an airflow to cleave beads from a stream of hydrogel solution. The performance of the proposed alginate bead generator was evaluated by changing the volume flow rates of alginate (QAlg) and air (QA), the diameter of device nozzle (d) and the concentration of alginate gel solution (C). We identified that the diameter of beads (D = 0.9 -2.8 mm) can be precisely controlled by changing QA and d. Also the bead generation frequency (f) can be tuned by changing QAlg. Finally, we demonstrated that viability and biological function (pericellular matrix deposition) of chondrocytes were not adversely affected by high f using this bead generator. Because 3D printing is becoming a more accessible technique, our unique design will allow greater access to average biomedical research laboratories, STEM education and industries in cost- and time-effective manner.
Assuntos
Alginatos , Técnicas de Cultura de Células , Hidrogéis , Impressão TridimensionalRESUMO
Substantial, involved, and expensive efforts to promote the dissemination of scientific knowledge and career interest in Science, Technology, Engineering, and Mathematics (STEM) are enthusiastically supported by many scientific, federal, and local organizations. The articulated underlying goals for these efforts include an enhanced public understanding of science and science-related policy, an increased diversity in STEM careers, and an increase in the future STEM workforce. This effort is primarily driven by an underperformance of the United States that includes poor test performance and limited number of students pursuing STEM degrees. Despite this investment, attitudes toward STEM have not notably changed. The goal of this project was to determine students' attitudes toward STEM in response to a previously established scientific outreach event. This event was used to address three common goals in STEM outreach: STEM literacy, diversity and inclusion, and career preparedness. We found there was a notable difference in the attitudes toward scientific activities and interest in pursuing a "Science Career" after participation in this event. Strikingly, interest in hypothesis development, the keystone of all STEM disciplines, was the least liked of all the activities offered during the event. Our data suggest that events designed to enhance interest in pursuing a STEM career may benefit from different elements compared with events designed to increase understanding of STEM literacy concepts, such as hypothesis development.
Assuntos
Estudantes , Tecnologia , Atitude , Escolha da Profissão , Humanos , Matemática , Estudos RetrospectivosRESUMO
Spatial and temporal regulation of chondrocyte maturation in the growth plate drives growth of many bones. One essential event to generate the ordered cell array characterizing growth plate cartilage is the formation of chondrocyte columns in the proliferative zone via 90-degree rotation of daughter cells to align with the long axis of the bone. Previous studies have suggested crucial roles for cadherins and integrin ß1 in column formation. The purpose of this study was to determine the relative contributions of cadherin- and integrin-mediated cell adhesion in column formation. Here we present new mechanistic insights generated by application of live time-lapse confocal microscopy of cranial base explant cultures, robust genetic mouse models, and new quantitative methods to analyze cell behavior. We show that conditional deletion of either the cell-cell adhesion molecule Cdh2 or the cell-matrix adhesion molecule Itgb1 disrupts column formation. Compound mutants were used to determine a potential reciprocal regulatory interaction between the two adhesion surfaces and identified that defective chondrocyte rotation in a N-cadherin mutant was restored by a heterozygous loss of integrin ß1. Our results support a model for which integrin ß1, and not N-cadherin, drives chondrocyte rotation and for which N-cadherin is a potential negative regulator of integrin ß1 function.
Assuntos
Caderinas , Cartilagem , Lâmina de Crescimento , Integrina beta1 , Animais , Camundongos , Caderinas/metabolismo , Cartilagem/metabolismo , Adesão Celular/fisiologia , Lâmina de Crescimento/metabolismo , Integrina beta1/metabolismoRESUMO
In the past decade, cartilage tissue engineering has arisen as a promising therapeutic option for degenerative joint diseases, such as osteoarthritis, in the hope of restoring the structure and physiological functions. Hydrogels are promising biomaterials for developing engineered scaffolds for cartilage regeneration. However, hydrogel-delivered mesenchymal stem cells or chondrocytes could be exposed to elevated levels of reactive oxygen species (ROS) in the inflammatory microenvironment after being implanted into injured joints, which may affect their phenotype and normal functions and thereby hinder the regeneration efficacy. To attenuate ROS induced side effects, a multifunctional hydrogel with an innate anti-oxidative ability was produced in this study. The hydrogel was rapidly formed through a dynamic covalent bond between phenylboronic acid grafted hyaluronic acid (HA-PBA) and poly(vinyl alcohol) and was further stabilized through a secondary crosslinking between the acrylate moiety on HA-PBA and the free thiol group from thiolated gelatin. The hydrogel is cyto-compatible and injectable and can be used as a bioink for 3D bioprinting. The viscoelastic properties of the hydrogels could be modulated through the hydrogel precursor concentration. The presence of dynamic covalent linkages contributed to its shear-thinning property and thus good printability of the hydrogel, resulting in the fabrication of a porous grid construct and a meniscus like scaffold at high structural fidelity. The bioprinted hydrogel promoted cell adhesion and chondrogenic differentiation of encapsulated rabbit adipose derived mesenchymal stem cells. Meanwhile, the hydrogel supported robust deposition of extracellular matrix components, including glycosaminoglycans and type II collagen, by embedded mouse chondrocytesin vitro. Most importantly, the hydrogel could protect encapsulated chondrocytes from ROS induced downregulation of cartilage-specific anabolic genes (ACAN and COL2) and upregulation of a catabolic gene (MMP13) after incubation with H2O2. Furthermore, intra-articular injection of the hydrogel in mice revealed adequate stability and good biocompatibilityin vivo. These results demonstrate that this hydrogel can be used as a novel bioink for the generation of 3D bioprinted constructs with anti-ROS ability to potentially enhance cartilage tissue regeneration in a chronic inflammatory and elevated ROS microenvironment.