RESUMEN
Micro- and nanoencapsulation of cells has been studied as a strategy to protect cells from environmental stress and promote survival during delivery. Hydrogels used in encapsulation can be modified to influence cell behaviors and direct assembly in their surroundings. Here, we report a system that conformally encapsulated stem cells using hyaluronic acid (HA). We successfully modified HA with lipid, thiol, and maleimide pendant groups to facilitate a hydrogel system in which HA was deposited onto cell plasma membranes and subsequently crosslinked through thiol-maleimide click chemistry. We demonstrated conformal encapsulation of both neural stem cells (NSCs) and mesenchymal stromal cells (MSCs), with viability of both cell types greater than 90% after encapsulation. Additional material could be added to the conformal hydrogel through alternating addition of thiol-modified and maleimide-modified HA in a layering process. After encapsulation, we tracked egress and viability of the cells over days and observed differential responses of cell types to conformal hydrogels both according to cell type and the amount of material deposited on the cell surfaces. Through the design of the conformal hydrogels, we showed that multicellular assembly could be created in suspension and that encapsulated cells could be immobilized on surfaces. In conjunction with photolithography, conformal hydrogels enabled rapid assembly of encapsulated cells on hydrogel substrates with resolution at the scale of 100 µm.
Asunto(s)
Supervivencia Celular , Ácido Hialurónico , Hidrogeles , Células Madre Mesenquimatosas , Células-Madre Neurales , Ácido Hialurónico/química , Ácido Hialurónico/farmacología , Animales , Células-Madre Neurales/efectos de los fármacos , Células-Madre Neurales/citología , Hidrogeles/química , Hidrogeles/farmacología , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Encapsulación Celular/métodos , Ratones , Propiedades de Superficie , Células CultivadasRESUMEN
One of the goals of tissue engineering and regenerative medicine is restoring primary living tissue function by manufacturing a 3D microenvironment. One of the main challenges is protecting implanted non-autologous cells or tissues from the host immune system. Cell encapsulation has emerged as a promising technique for this purpose. It involves entrapping cells in biocompatible and semi-permeable microcarriers made from natural or synthetic polymers that regulate the release of cellular secretions. In recent years, droplet-based microfluidic systems have emerged as powerful tools for cell encapsulation in tissue engineering and regenerative medicine. These systems offer precise control over droplet size, composition, and functionality, allowing for creating of microenvironments that closely mimic native tissue. Droplet-based microfluidic systems have extensive applications in biotechnology, medical diagnosis, and drug discovery. This review summarises the recent developments in droplet-based microfluidic systems and cell encapsulation techniques, as well as their applications, advantages, and challenges in biology and medicine. The integration of these technologies has the potential to revolutionise tissue engineering and regenerative medicine by providing a precise and controlled microenvironment for cell growth and differentiation. By overcoming the immune system's challenges and enabling the release of cellular secretions, these technologies hold great promise for the future of regenerative medicine.
Asunto(s)
Encapsulación Celular , Medicina Regenerativa , Ingeniería de Tejidos , Humanos , Encapsulación Celular/métodos , Medicina Regenerativa/métodos , Animales , Microfluídica/instrumentación , Técnicas Analíticas Microfluídicas/instrumentaciónRESUMEN
Delivery of therapeutic stem cells to treat bone tissue damage is a promising strategy that faces many hurdles to clinical translation. Among them is the design of a delivery vehicle which promotes desired cell behavior for new bone formation. In this work, we describe the use of an injectable microporous hydrogel, made of crosslinked gelatin microgels, for the encapsulation and delivery of human mesenchymal stem cells (MSCs) and compared it to a traditional nonporous injectable hydrogel. MSCs encapsulated in the microporous hydrogel showed rapid cell spreading with direct cell-cell connections whereas the MSCs in the nonporous hydrogel were entrapped by the surrounding polymer mesh and isolated from each other. On a per-cell basis, encapsulation in microporous hydrogel induced a 4 × increase in alkaline phosphatase (ALP) activity and calcium mineral deposition in comparison to nonporous hydrogel, as measured by ALP and calcium assays, which indicates more robust osteogenic differentiation. RNA-seq confirmed the upregulation of the genes and pathways that are associated with cell spreading and cell-cell connections, as well as the osteogenesis in the microporous hydrogel. These results demonstrate that microgel-based injectable hydrogels can be useful tools for therapeutic cell delivery for bone tissue repair.
Asunto(s)
Diferenciación Celular , Hidrogeles , Células Madre Mesenquimatosas , Osteogénesis , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Humanos , Hidrogeles/química , Porosidad , Fosfatasa Alcalina/metabolismo , Células Cultivadas , Encapsulación Celular/métodos , Trasplante de Células Madre Mesenquimatosas/métodos , InyeccionesRESUMEN
Cell encapsulation into spherical microparticles is a promising bioengineering tool in many fields, including 3D cancer modelling and pre-clinical drug discovery. Cancer microencapsulation models can more accurately reflect the complex solid tumour microenvironment than 2D cell culture and therefore would improve drug discovery efforts. However, these microcapsules, typically in the range of 1 - 5000 µm in diameter, must be carefully designed and amenable to high-throughput production. This review therefore aims to outline important considerations in the design of cancer cell microencapsulation models for drug discovery applications and examine current techniques to produce these. Extrusion (dripping) droplet generation and emulsion-based techniques are highlighted and their suitability to high-throughput drug screening in terms of tumour physiology and ease of scale up is evaluated.
3D microencapsulation models of cancer offer a customisable platform to mimic key aspects of solid tumour physiology in vitro. However, many 3D models do not recapitulate the hypoxic conditions and altered tissue stiffness established in many tumour types and stages. Furthermore, microparticles for cancer cell encapsulation are commonly produced using methods that are not necessarily suitable for scale up to high-throughput manufacturing. This review aims to evaluate current technologies for cancer cell-laden microparticle production with a focus on physiological relevance and scalability. Emerging techniques will then be touched on, for production of uniform microparticles suitable for high-throughput drug discovery applications.
Asunto(s)
Descubrimiento de Drogas , Neoplasias , Humanos , Neoplasias/patología , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Descubrimiento de Drogas/métodos , Encapsulación Celular/métodos , Modelos Biológicos , Cápsulas , Animales , Composición de Medicamentos/métodos , Microambiente Tumoral/efectos de los fármacosRESUMEN
Microfluidic cell encapsulation has provided a platform for studying the behavior of individual cells and has become a turning point in single-cell analysis during the last decade. The engineered microenvironment, along with protecting the immune response, has led to increasingly presenting the results of practical and pre-clinical studies with the goals of disease treatment, tissue engineering, intelligent control of stem cell differentiation, and regenerative medicine. However, the significance of cell-substrate interaction versus cell-cell communications in the microgel is still unclear. In this study, monodisperse alginate microgels were generated using a flow-focusing microfluidic device to determine how the cell microenvironment can control human bone marrow-derived mesenchymal stem cells (hBMSCs) viability, proliferation, and biomechanical features in single-cell droplets versus multi-cell droplets. Collected results show insufficient cell proliferation (234 % and 329 %) in both single- and multi-cell alginate microgels. Alginate hydrogels supplemented with poly-l-lysine (PLL) showed a better proliferation rate (514 % and 780 %) in a comparison of free alginate hydrogels. Cell stiffness data illustrate that hBMSCs cultured in alginate hydrogels have higher membrane flexibility and migration potency (Young's modulus equal to 1.06 kPa), whereas PLL introduces more binding sites for cell attachment and causes lower flexibility and migration potency (Young's modulus equal to 1.83 kPa). Considering that cell adhesion is the most important parameter in tissue engineering, in which cells do not run away from a 3D substrate, PLL enhances cell stiffness and guarantees cell attachments. In conclusion, cell attachment to PLL-mediated alginate hydrogels is crucial for cell viability and proliferation. It suggests that cell-cell signaling is good enough for stem cell viability, but cell-PLL attachment alongside cell-cell signaling is crucial for stem cell proliferation and self-renewal.
Asunto(s)
Alginatos , Adhesión Celular , Proliferación Celular , Células Madre Mesenquimatosas , Microgeles , Polilisina , Alginatos/química , Alginatos/farmacología , Polilisina/química , Polilisina/farmacología , Humanos , Adhesión Celular/efectos de los fármacos , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Proliferación Celular/efectos de los fármacos , Microgeles/química , Microfluídica/métodos , Comunicación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Hidrogeles/química , Hidrogeles/farmacología , Encapsulación Celular/métodos , Análisis de la Célula Individual , Autorrenovación de las Células/efectos de los fármacos , Diferenciación Celular/efectos de los fármacosRESUMEN
AIMS: This study aimed to encapsulate natural killer (NK) cells in a hydrogel to sustain their function within the hypoxic tumour microenvironments. METHODS: An alginate-gelatine hydrogel was generated via electrospray technology. Hydrogel biocompatibility was assessed through cell counting kit-8 and Live/Dead assays to ascertain cell. Moreover, we analysed lactate dehydrogenase assays to evaluate the cytotoxicity against tumours and utilised RT-qPCR to analyse cytokine gene level. RESULTS: Alginate and gelatine formed hydrogels with diameters ranging from 489.2 ± 23.0 µm, and the encapsulation efficiency was 34.07 ± 1.76%. Encapsulated NK cells exhibited robust proliferation and tumour-killing capabilities under normoxia and hypoxia. Furthermore, encapsulation provided a protective shield against cell viability under hypoxia. Importantly, tumour-killing cytotoxicity through cytokines upregulation such as granzyme B and interferon-gamma was preserved under hypoxia. CONCLUSION: The encapsulation of NK cells not only safeguards their viability but also reinforces anticancer capacity, countering the inhibition of activation induced by hypoxia.
Asunto(s)
Alginatos , Proliferación Celular , Gelatina , Hidrogeles , Células Asesinas Naturales , Microesferas , Alginatos/química , Alginatos/farmacología , Hidrogeles/química , Hidrogeles/farmacología , Humanos , Proliferación Celular/efectos de los fármacos , Gelatina/química , Supervivencia Celular/efectos de los fármacos , Hipoxia de la Célula/efectos de los fármacos , Encapsulación Celular/métodos , Animales , Microambiente Tumoral/efectos de los fármacos , Línea Celular Tumoral , RatonesRESUMEN
Liver failure represents a critical medical condition with a traditionally grim prognosis, where treatment options have been notably limited. Historically, liver transplantation has stood as the sole definitive cure, yet the stark disparity between the limited availability of liver donations and the high demand for such organs has significantly hampered its feasibility. This discrepancy has necessitated the exploration of hepatocyte transplantation as a temporary, supportive intervention. In light of this, our review delves into the burgeoning field of hepatocyte transplantation, with a focus on the latest advancements in maintaining hepatocyte function, co-microencapsulation techniques, xenogeneic hepatocyte transplantation, and the selection of materials for microencapsulation. Our examination of hepatocyte microencapsulation research highlights that, to date, most studies have been conducted in vitro or using liver failure mouse models, with a notable paucity of experiments on larger mammals. The functionality of microencapsulated hepatocytes is primarily inferred through indirect measures such as urea and albumin production and the rate of ammonia clearance. Furthermore, research on the mechanisms underlying hepatocyte co-microencapsulation remains limited, and the practicality of xenogeneic hepatocyte transplantation requires further validation. The potential of hepatocyte microencapsulation extends beyond the current scope of application, suggesting a promising horizon for liver failure treatment modalities. Innovations in encapsulation materials and techniques aim to enhance cell viability and function, indicating a need for comprehensive studies that bridge the gap between small-scale laboratory success and clinical applicability. Moreover, the integration of bioengineering and regenerative medicine offers novel pathways to refine hepatocyte transplantation, potentially overcoming the challenges of immune rejection and ensuring the long-term functionality of transplanted cells. In conclusion, while hepatocyte microencapsulation and transplantation herald a new era in liver failure therapy, significant strides must be made to translate these experimental approaches into viable clinical solutions. Future research should aim to expand the experimental models to include larger mammals, thereby providing a clearer understanding of the clinical potential of these therapies. Additionally, a deeper exploration into the mechanisms of cell survival and function within microcapsules, alongside the development of innovative encapsulation materials, will be critical in advancing the field and offering new hope to patients with liver failure.
Asunto(s)
Encapsulación Celular , Supervivencia Celular , Hepatocitos , Animales , Humanos , Encapsulación Celular/métodos , Hepatocitos/trasplante , Hepatocitos/citología , Fallo Hepático/terapia , Trasplante HeterólogoRESUMEN
This study presents the development and characterization of a novel double-network self-healing hydrogel based on N-carboxyethyl chitosan (CEC) and oxidized dextran (OD) with the incorporation of crosslinked collagen (CEC-OD/COL-GP) to enhance its biological and physicochemical properties. The hydrogel formed via dynamic imine bond formation exhibited efficient self-healing within 30 min, and a compressive modulus recovery of 92 % within 2 h. In addition to its self-healing ability, CEC-OD/COL-GP possesses unique physicochemical characteristics including transparency, injectability, and adhesiveness to various substrates and tissues. Cell encapsulation studies confirmed the biocompatibility and suitability of the hydrogel as a cell-culture scaffold, with the presence of a collagen network that enhances cell adhesion, spreading, long-term cell viability, and proliferation. Leveraging their unique properties, we engineered assemblies of self-healing hydrogel modules for controlled spatiotemporal drug delivery and constructed co-culture models that simulate angiogenesis in tumor microenvironments. Overall, the CEC-OD/COL-GP hydrogel is a versatile and promising material for biomedical applications, offering a bottom-up approach for constructing complex structures with self-healing capabilities, controlled drug release, and support for diverse cell types in 3D environments. This hydrogel platform has considerable potential for advancements in tissue engineering and therapeutic interventions.
Asunto(s)
Adhesión Celular , Quitosano , Dextranos , Hidrogeles , Hidrogeles/química , Hidrogeles/farmacología , Quitosano/química , Dextranos/química , Humanos , Adhesión Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Colágeno/química , Animales , Liberación de Fármacos , Proliferación Celular/efectos de los fármacos , Encapsulación Celular/métodos , Materiales Biomiméticos/química , Materiales Biomiméticos/farmacología , Ratones , Biomimética/métodos , Materiales Biocompatibles/química , Materiales Biocompatibles/farmacología , Andamios del Tejido/químicaRESUMEN
Droplet-based microfluidic technologies for encapsulating single cells have rapidly evolved into powerful tools for single-cell analysis. In conventional passive single-cell encapsulation techniques, because cells arrive randomly at the droplet generation section, to encapsulate only a single cell with high precision, the average number of cells per droplet has to be decreased by reducing the average frequency at which cells arrive relative to the droplet generation rate. Therefore, the encapsulation efficiency for a given droplet generation rate is very low. Additionally, cell sorting operations are required prior to the encapsulation of target cells for specific cell type analysis. To address these challenges, we developed a cell encapsulation technology with a cell sorting function using a microfluidic chip. The microfluidic chip is equipped with an optical detection section to detect the optical information of cells and a sorting section to encapsulate cells into droplets by controlling a piezo element, enabling active encapsulation of only the single target cells. For a particle population including both targeted and non-targeted particles arriving at an average frequency of up to 6000 particles per s, with an average number of particles per droplet of 0.45, our device maintained a high purity above 97.9% for the single-target-particle droplets and achieved an outstanding throughput, encapsulating up to 2900 single target particles per s. The proposed encapsulation technology surpasses the encapsulation efficiency of conventional techniques, provides high efficiency and flexibility for single-cell research, and shows excellent potential for various applications in single-cell analysis.
Asunto(s)
Dispositivos Laboratorio en un Chip , Análisis de la Célula Individual , Análisis de la Célula Individual/instrumentación , Humanos , Técnicas Analíticas Microfluídicas/instrumentación , Diseño de Equipo , Ensayos Analíticos de Alto Rendimiento/instrumentación , Animales , Encapsulación Celular/métodos , Encapsulación Celular/instrumentaciónRESUMEN
Droplet microfluidics-based single-cell encapsulation is a critical technology that enables large-scale parallel single-cell analysis by capturing and processing thousands of individual cells. As the efficiency of passive single-cell encapsulation is limited by Poisson distribution, active single-cell encapsulation has been developed to theoretically ensure that each droplet contains one cell. However, existing active single-cell encapsulation technologies still face issues related to fluorescence labeling and low throughput. Here, we present an active single-cell encapsulation technique by using microvalve-based drop-on-demand technology and real-time image processing to encapsulate single cells with high throughput in a label-free manner. Our experiments demonstrated that the single-cell encapsulation system can encapsulate individual polystyrene beads with 96.3 % efficiency and HeLa cells with 94.9 % efficiency. The flow speed of cells in this system can reach 150 mm/s, resulting in a corresponding theoretical encapsulation throughput of 150 Hz. This technology has significant potential in various biomedical applications, including single-cell omics, secretion detection, and drug screening.
Asunto(s)
Análisis de la Célula Individual , Humanos , Análisis de la Célula Individual/métodos , Células HeLa , Procesamiento de Imagen Asistido por Computador , Poliestirenos/química , Técnicas Analíticas Microfluídicas/instrumentación , Dispositivos Laboratorio en un Chip , Encapsulación Celular/métodosRESUMEN
Encapsulation of single cells is a powerful technique used in various fields, such as regenerative medicine, drug delivery, tissue regeneration, cell-based therapies, and biotechnology. It offers a method to protect cells by providing cytocompatible coatings to strengthen cells against mechanical and environmental perturbations. Silk fibroin, derived from the silkworm Bombyx mori, is a promising protein biomaterial for cell encapsulation due to the cytocompatibility and capacity to maintain cell functionality. Here, THP-1 cells, a human leukemia monocytic cell line, were encapsulated with chemically modified silk polyelectrolytes through electrostatic layer-by-layer deposition. The effectiveness of the silk nanocoating was assessed using scanning electron microscopy (SEM) and confocal microscopy and on cell viability and proliferation by Alamar Blue assay and live/dead staining. An analysis of the mechanical properties of the encapsulated cells was conducted using atomic force microscopy nanoindentation to measure elasticity maps and cellular stiffness. After the cells were encapsulated in silk, an increase in their stiffness was observed. Based on this observation, we developed a mechanical predictive model to estimate the variations in stiffness in relation to the thickness of the coating. By tuning the cellular assembly and biomechanics, these encapsulations promote systems that protect cells during biomaterial deposition or processing in general.
Asunto(s)
Bombyx , Supervivencia Celular , Humanos , Supervivencia Celular/efectos de los fármacos , Animales , Seda/química , Células THP-1 , Fibroínas/química , Materiales Biocompatibles/química , Materiales Biocompatibles/farmacología , Proliferación Celular/efectos de los fármacos , Encapsulación Celular/métodosRESUMEN
Encapsulated cell technology (ECT) is a targeted delivery method that uses the genetically engineered cells in semipermeable polymer capsules to deliver cytokines. Thus far, ECT has been extensively utilized in pharmacologic research, and shows enormous potentials in the treatment of posterior segment diseases. Due to the biological barriers within the eyeball, it is difficult to attain effective therapeutic concentration in the posterior segment through topical administration of drug molecules. Encouragingly, therapeutic cytokines provided by ECT can cross these biological barriers and achieve sustained release at the desired location. The encapsulation system uses permeable materials that allow growth factors and cytokines to diffuse efficiently into retinal tissue. Moreover, the ECT based treatment can be terminated timely when we need to retrieve the implant, which makes the therapy reversible and provides a safer alternative for intraocular gene therapy. Meanwhile, we also place special emphasis on optimizing encapsulation materials and enhancing preservation techniques to achieve the stable release of growth factors and cytokines in the eyeball. This technology holds great promise for the treatment of patients with dry AMD, RP, glaucoma and MacTel. These findings would enrich our understandings of ECT and promote its future applications in treatment of degenerative retinopathy. This review comprises articles evaluating the exactness of artificial intelligence-based formulas published from 2000 to March 2024. The papers were identified by a literature search of various databases (PubMed/MEDLINE, Google Scholar, Cochrane Library and Web of Science).
Asunto(s)
Citocinas , Oftalmopatías , Humanos , Citocinas/metabolismo , Animales , Oftalmopatías/terapia , Oftalmopatías/tratamiento farmacológico , Sistemas de Liberación de Medicamentos/métodos , Encapsulación Celular/métodosRESUMEN
Diabetes is a prevalent chronic disease affecting millions of people globally. To address this health challenge, advanced beta cell therapy using biomaterials-based macroscale, microscale, and nanoscale encapsulation devices must tackle various obstacles. First, overcoming foreign body responses is a major focus of research. Strategies such as immunomodulatory materials and physical immunoshielding are investigated to reduce the immune response and improve the longevity of the encapsulated cells. Furthermore, oxygenating strategies, such as the use of oxygen-releasing biomaterials, are developed to improve oxygen diffusion and promote cell survival. Finally, yet importantly, promoting vascularization through the use of angiogenic growth factors and the incorporation of pre-vascularized materials are also explored to enhance nutrient and oxygen supply to the encapsulated cells. This review seeks to specifically highlight the emerging research strategies developed to overcome these challenges using micro and nanoscale biomaterial encapsulation devices. Continuously improving and refining these strategies make an advance toward realizing the improved therapeutic potential of the encapsulated beta cells.
Asunto(s)
Materiales Biocompatibles , Diabetes Mellitus Tipo 1 , Células Secretoras de Insulina , Humanos , Células Secretoras de Insulina/citología , Células Secretoras de Insulina/metabolismo , Diabetes Mellitus Tipo 1/terapia , Animales , Materiales Biocompatibles/química , Encapsulación Celular/métodos , Oxígeno/químicaRESUMEN
Hydrogel cell encapsulation devices are a common approach to reduce the need for chronic systemic immunosuppression in allogeneic cell product transplantation. Macroencapsulation approaches are an appealing strategy, as they maximize graft retrievability and cell dosage within a single device; however, macroencapsulation devices face oxygen transport challenges as geometries increase from preclinical to clinical scales. Device design guided by computational approaches can facilitate graft oxygen availability to encapsulated cells in vivo but is limited without accurate measurement of oxygen levels within the transplant site and graft. In this study, we engineer pO2 reporter composite hydrogels (PORCH) to enable spatiotemporal measurement of oxygen tension within macroencapsulation devices using the proton Imaging of siloxanes to map tissue oxygenation levels (PISTOL) magnetic resonance imaging approach. We engineer two methods of incorporating siloxane oximetry reporters within hydrogel devices, an emulsion and microbead-based approach, and evaluate PORCH cytotoxicity on co-encapsulated cells and accuracy in quantifying oxygen tension in vitro. We find that both emulsion and microbead PORCH approaches enable accurate in situ oxygen quantification using PISTOL magnetic resonance oximetry, and that the emulsion-based PORCH approach results in higher spatial resolution.
Asunto(s)
Hidrogeles , Imagen por Resonancia Magnética , Oxígeno , Imagen por Resonancia Magnética/métodos , Oxígeno/análisis , Oxígeno/metabolismo , Hidrogeles/química , Humanos , Encapsulación Celular/métodos , Oximetría/métodosRESUMEN
An ideal hydrogel for stem cell therapy would be injectable and efficiently promote stem cell proliferation and differentiation in body. Herein, an injectable, single-component hydrogel with hyaluronic acid (HA) modified with phenylboronic acid (PBA) and spermidine (SM) is introduced. The resulting HAps (HA-PBA-SM) hydrogel is based on the reversible crosslinking between the diol and the ionized PBA, which is stabilized by the SM. It has a shear-thinning property, enabling its injection through a syringe to form a stable hydrogel inside the body. In addition, HAps hydrogel undergoes a post-injection "self-curing," which stiffens the hydrogel over time. This property allows the HAps hydrogel to meet the physical requirements for stem cell therapy in rigid tissues, such as bone, while maintaining injectability. The hydrogel enabled favorable proliferation of human mesenchymal stem cells (hMSCs) and promoted their differentiation and mineralization. After the injection of hMSCs-containing HAps into a rat femoral defect model, efficient osteogenic differentiation of hMSCs and bone regeneration is observed. The study demonstrates that simple cationic modification of PBA-based hydrogel enabled efficient gelation with shear-thinning and self-curing properties, and it would be highly useful for stem cell therapy and in vivo bone regeneration.
Asunto(s)
Regeneración Ósea , Ácidos Borónicos , Diferenciación Celular , Hidrogeles , Células Madre Mesenquimatosas , Animales , Regeneración Ósea/fisiología , Ratas , Hidrogeles/química , Células Madre Mesenquimatosas/citología , Humanos , Ácido Hialurónico/química , Ratas Sprague-Dawley , Encapsulación Celular/métodos , Proliferación Celular , Osteogénesis/fisiología , Modelos Animales de Enfermedad , Espermidina/farmacología , Espermidina/químicaRESUMEN
Tissue engineering at single-cell resolution has enhanced therapeutic efficacy. Droplet microfluidics offers a powerful platform that allows deterministic single-cell encapsulation into aqueous droplets, yet the direct encapsulation of cells into microgels remains challenging. Here, the design of a microfluidic device that is capable of single-cell encapsulation within polyethylene glycol norbornene (PEGNB) hydrogels on-chip is reported. Cells are first ordered in media within a straight microchannel via inertial focusing, followed by the introduction of PEGNB solution from two separate, converging channels. Droplets are thoroughly mixed by passage through a serpentine channel, and microgels are formed by photo-photopolymerization. This platform uniquely enables both single-cell encapsulation and excellent cell viability post-photo-polymerization. More than 90% of singly encapsulated mesenchymal stromal cells (MSCs) remain alive for 7 days. Notably, singly encapsulated MSCs have elevated expression levels in genes that code anti-inflammatory cytokines, for example, IL-10 and TGF-ß, thus enhancing the secretion of proteins of interest. Following injection into a mouse model with induced inflammation, singly encapsulated MSCs show a strong retention rate in vivo, reduce overall inflammation, and mitigate liver damage. These translational results indicate that deterministic single-cell encapsulation could find use in a broad spectrum of tissue engineering applications.
Asunto(s)
Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas , Norbornanos , Polietilenglicoles , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Animales , Polietilenglicoles/química , Ratones , Trasplante de Células Madre Mesenquimatosas/métodos , Norbornanos/química , Microgeles/química , Encapsulación Celular/métodos , Hidrogeles/química , Hidrogeles/farmacología , Supervivencia Celular/efectos de los fármacos , HumanosRESUMEN
Mesenchymal stem/stromal cells (MSC) promote recovery in a wide range of animal models of injury and disease. They can act in vivo by differentiating and integrating into tissues, secreting factors that promote cell growth and control inflammation, and interacting directly with host effector cells. We focus here on MSC secreted factors by encapsulating the cells in alginate microspheres, which restrict cells from migrating out while allowing diffusion of factors including cytokines across the capsules. One week after intrathecal lumbar injection of human bone marrow MSC encapsulated in alginate (eMSC), rat IL-10 expression was upregulated in distant rat spinal cord injury sites. Detection of human IL-10 protein in rostrally derived cerebrospinal fluid (CSF) indicated distribution of this human MSC-secreted cytokine throughout rat spinal cord CSF. Intraperitoneal (IP) injection of eMSC in a rat model for endotoxemia reduced serum levels of inflammatory cytokines within 5 h. Detection of human IL-6 in sera after injection of human eMSC indicates rapid systemic distribution of this human MSC-secreted cytokine. Despite proof of concept for eMSC in various disorders using animal models, translation of encapsulation technology has not been feasible primarily because methods for scale-up are not available. To scale-up production of eMSC, we developed a rapid, semi-continuous, capsule collection system coupled to an electrosprayer. This system can produce doses of encapsulated cells sufficient for use in clinical translation.
Asunto(s)
Antiinflamatorios , Encapsulación Celular , Citocinas , Células Madre Mesenquimatosas , Animales , Humanos , Ratas , Alginatos , Encapsulación Celular/métodos , Citocinas/metabolismo , Interleucina-10/metabolismoRESUMEN
Monolayer cultures of cell lines and derived-patient cells have long been the in vitro model of choice in oncology. In particular, these models have made it possible to decipher the mechanisms that determine tumor proliferation and invasion. However these 2D models are insufficient because they do not take into account the spatial organization of cells and their interactions with each other or with the extracellular matrix. In the context of cancer, there is a need to develop new 3D (tumoroid) models in order to gain a better understanding of the development of these pathologies but also to assess the penetration of drugs through a tissue and the associated cellular response. We present here the cell capsule technology (CCT), which allows the production of different tumoroid models: simple or more complex 3D culture models including co-culture of tumor cells with components of the microenvironment (fibroblasts, matrix, etc.). The development of these new 3D culture systems now makes it possible to propose refined physiopathological models that will allow the implementation of improved targeted therapeutic strategies.
Asunto(s)
Técnicas de Cultivo Tridimensional de Células/métodos , Encapsulación Celular/métodos , Organoides , Esferoides Celulares , Alginatos , Fibroblastos Asociados al Cáncer , Comunicación Celular , Proliferación Celular , Técnicas de Cocultivo/métodos , Transición Epitelial-Mesenquimal , Matriz Extracelular/química , Humanos , Invasividad Neoplásica , Células Tumorales Cultivadas , Microambiente TumoralRESUMEN
Smart hydrogels with versatile properties, including a tunable gelation time, nonswelling attributes, and biocompatibility, are in great need in the biomedical field. To meet this urgent demand, we explored novel biomaterials with the desired properties from sessile marine organisms. To this end, a novel protein, Sbp9, derived from scallop byssus was extensively investigated, which features typical epidermal growth factor-like (EGFL) multiple repetitive motifs. Our current work demonstrated that the key fragment of Sbp9 (calcium-binding domain (CBD) and 4 EGFL repeats (CE4)) was able to form a smart hydrogel driven by noncovalent interactions and facilitated by disulfide bonds. More importantly, this smart hydrogel demonstrates several desirable and beneficial features, which could offset the drawbacks of typical protein-based hydrogels, including (1) a redox-responsive gelation time (from <1 to 60 min); (2) tunable mechanical properties, nonswelling abilities, and an appropriate microstructure; and (3) good biocompatibility and degradability. Furthermore, proof-of-concept demonstrations showed that the newly discovered hydrogel could be used for anticancer drug delivery and cell encapsulation. Taken together, a smart hydrogel inspired by marine sessile organisms with desirable properties was generated and characterized and demonstrated to have extensive applicability potential in biomedical applications, including tissue engineering and drug release.
Asunto(s)
Proteínas de Unión al Calcio/química , Encapsulación Celular/métodos , Portadores de Fármacos/química , Hidrogeles/química , Pectinidae/química , Materiales Inteligentes/química , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Animales , Antineoplásicos/química , Proteínas de Unión al Calcio/toxicidad , Línea Celular Tumoral , Doxorrubicina/química , Portadores de Fármacos/toxicidad , Liberación de Fármacos , Humanos , Hidrogeles/toxicidad , Peróxido de Hidrógeno/química , Masculino , Células Madre Mesenquimatosas/efectos de los fármacos , Oxidación-Reducción , Porosidad , Dominios Proteicos , Ratas Sprague-Dawley , Materiales Inteligentes/toxicidadRESUMEN
Drought is a major abiotic stress imposed by climate change that affects crop production and soil microbial functions. Plants respond to water deficits at the morphological, biochemical, and physiological levels, and invoke different adaptation mechanisms to tolerate drought stress. Plant growth-promoting bacteria (PGPB) can help to alleviate drought stress in plants through various strategies, including phytohormone production, the solubilization of mineral nutrients, and the production of 1-aminocyclopropane-1-carboxylate deaminase and osmolytes. However, PGPB populations and functions are influenced by adverse soil factors, such as drought. Therefore, maintaining the viability and stability of PGPB applied to arid soils requires that the PGPB have to be protected by suitable coatings. The encapsulation of PGPB is one of the newest and most efficient techniques for protecting beneficial bacteria against unfavorable soil conditions. Coatings made from polysaccharides, such as sodium alginate, chitosan, starch, cellulose, and their derivatives, can absorb and retain substantial amounts of water in the interstitial sites of their structures, thereby promoting bacterial survival and better plant growth.