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Representative pictures of caspase-activated apoptosis at 1 (top row), 5 (middle row), and 10 (bottom row) days of free-standing (left panel) and fibrin-embedded (right panel) culture for βTC3-only (left), 2:1 (middle), and 1:1 (right) ratio of EPIs.
Source publication
Despite the recent achievements in cell-based therapies for curing type-1 diabetes (T1D), capillarization in beta (β)-cell clusters is still a major roadblock as it is essential for long-term viability and function of β-cells in vivo. In this research, we report sprouting angiogenesis in engineered pseudo islets (EPIs) made of mouse insulinoma βTC3...
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Citations
... Leveraging Additive manufacturing designed structures guided vessel formation in vitro and in vivo [80,89,93]. Shifting to bioprinting complex branching conduits in supportive hydrogels facilitated clinical translation for diverse cell therapies [94][95][96][97][98]. Researchers focused on developing a 3D scaffold platform to improve the transplantation outcomes of islet cells in T1D. ...
... This supported the potential for inducing angiogenesis within bioengineered islet constructs. Future work may combine patient-specific stem cell-derived human beta cells with endothelial cells using this approach to promote long-term graft survival for treating type 1 diabetes [98]. While, large-scale 3D printed vascularized structures are currently limited for the islet transplantation, advancements in leveraging additive manufacturing for the optimization vascularization conditions through the pore sizes and material choices, may facilitate translation to β-cell therapy in type 1 diabetes. ...
Diabetes mellitus is a significant global public health challenge, with a rising prevalence and associated morbidity and mortality. Cell therapy has evolved over time and holds great potential in diabetes treatment. In the present review, we discussed the recent progresses in cell-based therapies for diabetes that provides an overview of islet and stem cell transplantation technologies used in clinical settings, highlighting their strengths and limitations. We also discussed immunomodulatory strategies employed in cell therapies. Therefore, this review highlights key progresses that pave the way to design transformative treatments to improve the life quality among diabetic patients.
... Various strategies and advancements in recent years towards tissue engineering approaches may offer a viable alternative [190]. Towards this goal, the presence of ECs has been shown to influence the specific differentiation of pancreatic β-cells and sprouting angiogenesis was reported in these 3D β-cell spheroids [191][192][193]. These spheroids confirmed neovascularization with improved β-cell viability and functionality over time. ...
Biofabricated tissues have found numerous applications in tissue engineering and regenerative medicine in addition to the promotion of disease modeling and drug development and screening. Although three-dimensional (3D) printing strategies for designing and developing customized tissue constructs have made significant progress, the complexity of innate multicellular tissues hinders the accurate evaluation of physiological responses in vitro. Cellular aggregates, such as spheroids, are 3D structures where multiple types of cells are co-cultured and organized with endogenously secreted extracellular matrix and are designed to recapitulate the key features of native tissues more realistically. 3D Bioprinting has emerged as a crucial tool for positioning of these spheroids to assemble and organize them into physiologically- and histologically-relevant tissues, mimicking their native counterparts. This has triggered the convergence of spheroid fabrication and bioprinting, leading to the investigation of novel engineering methods for successful assembly of spheroids while simultaneously enhancing tissue repair. This review provides an overview of the current state-of-the-art in spheroid bioprinting methods and elucidates the involved technologies, intensively discusses the recent tissue fabrication applications, outlines the crucial properties that influence the bioprinting of these spheroids and bioprinted tissue characteristics, and finally details the current challenges and future perspectives of spheroid bioprinting efforts in the growing field of biofabrication.
... To recapture the changes occurring in the native pancreas, except the 3D surrounding, the pancreas model also requires spatial organization. Scientists are focusing on developing pancreatic β-cells able to secrete insulin [32][33][34] as well as developing co-cultured models as an attempt to mimic the interactions between cells in the islet organogenesis [35][36][37] and with endothelial cells to investigate the influence of vascularization [33,[38][39][40][41]. ...
... After 30 min of crosslinking in the incubator, cell media was deposited on top of scaffolds. The constructs culture medium was formulated in 1:1 ratio of βTC3 media and EGM-2V media (Lonza) based on previous study [38]. ...
... In this study, the aim was to understand the role of increasing simulated matrix complexity on the mechanical properties and cell behavior of the sECM. Therefore, the first group was a simple fibrin hydrogel, sECMf (see scheme in figure 1(A)) with a concentration that was increased by a factor of two over that shown previously [38]. The increase in concentration aided in stiffening the final matrix, creating a more stable perfusable device. ...
The extracellular matrix (ECM) influences cellular behavior, function, and fate. The ECM surrounding Langerhans islets has not been investigated in detail to explain its role in the development and maturation of pancreatic β-cells. Herein, a complex combination of the simulated ECM (sECM) has been examined with a comprehensive analysis of cell response and a variety of controls. The most promising results were obtained from group containing fibrin, collagen type I, Matrigel®, hyaluronic acid, methylcellulose, and two compounds of functionalized, ionically crosslinking bacterial cellulose (sECMbc). Even though the cell viability was not significantly impacted, the performance of group of sECMbc showed 2 to 4x higher sprouting number and length, 2 to 4x higher insulin secretion in static conditions, and 2 to 10x higher gene expression of VEGF-A, Endothelin-1, and NOS3 than the control group of fibrin matrix (sECMf). Each material was tested in a hydrogel-based, perfusable, pancreas-on-a-chip device and the best group - sECMbc has been tested with the drug Sunitinib to show the extended possibilities of the device for both diabetes-like screening as well as PDAC chemotherapeutics screening for potential personal medicine approach. It proved its functionality in 7 days dynamic culture and is suitable as a physiological tissue model. Moreover, the device with the pancreatic-like spheroids was 3D bioprintable and perfusable.
... Ensemencés dans un hydrogel avec des cellules supports, comme des CSM, les HUVEC ont, de plus, la capacité de former un réseau endothélial autour des îlots de rat [47]. L'encapsulation des Langerhanoïdes vascularisés dans un hydrogel favorise l'échappement des cellules endothéliales endogènes en formant des bourgeonnements (en anglais, sproutings) visibles en microscopie [37,48]. Ainsi, une anastomose entre les cellules endothéliales constituant le Langerhanoïde et celles présentes dans l'hydrogel semble facilitée. ...
Résumé
Les organoïdes sont des assemblages multicellulaires auto-organisés résultant de la prolifération et de la différenciation principalement des cellules souches. Des organoïdes d’îlots pancréatiques – récemment nommés Langerhanoïdes – ont été développés, dans l’espoir d’apporter une solution supplémentaire aux patients diabétiques. Pour construire des organoïdes ressemblant à des îlots humains, les cellules, les biomatériaux et la structure tridimensionnelle sont trois éléments clés à considérer. Le potentiel des Langerhanoïdes en tant que plateforme pour des études de développement, la modélisation des maladies, le dépistage de la toxicité et les traitements personnalisés et préventifs pour le diabète semble évident. Nous augurons qu’ils auront aussi un impact clinique substantiel dans un proche avenir et offriront de nouvelles perspectives thérapeutiques pour les patients.
... Islet-specific work in the AM-vascularization space has primarily focused on fundamental biological discoveries, with groups like Hospodiuk et al. using FDM technology to explore vessel sprouting from pseudo islets. [93] While publications creating macroscale, AM-manufactured vascularized constructs for islet transplantation are currently limited, advancements made in leveraging AM to support favorable vascularization conditions, such as the selection of optimal pore sizes and optimal material choices, facilitate their translation to -cell therapy in T1DM. ...
The increasing global prevalence of endocrine diseases like type 1 diabetes mellitus (T1DM) elevates the need for cellular replacement approaches, which can potentially enhance therapeutic durability and outcomes. Central to any cell therapy is the design of delivery systems that support cell survival and integration. In T1DM, well‐established fabrication methods have created a wide range of implants, ranging from 3D macro‐scale scaffolds to nano‐scale coatings. These traditional methods, however, are often challenged by their inherent limitations in reproducible and discrete fabrication, particularly when scaling to the clinic. Additive manufacturing (AM) techniques provide a means to address these challenges by delivering improved control over construct geometry and microscale component placement. While still early in development in the context of T1DM cellular transplantation, the integration of AM approaches serves to improve nutrient material transport, vascularization efficiency, and the accuracy of cell, matrix, and local therapeutic placement. This review highlights current methods in T1DM cellular transplantation and the potential of AM approaches to overcome these limitations. In addition, emerging AM technologies and their broader application to cell‐based therapy are discussed.
... Ensemencés dans un hydrogel avec des cellules supports, comme des CSM, les HUVEC ont, de plus, la capacité de former un réseau endothélial autour des îlots de rat [47]. L'encapsulation des Langerhanoïdes vascularisés dans un hydrogel favorise l'échappement des cellules endothéliales endogènes en formant des bourgeonnements (en anglais, sproutings) visibles en microscopie [37,48]. Ainsi, une anastomose entre les cellules endothéliales constituant le Langerhanoïde et celles présentes dans l'hydrogel semble facilitée. ...
Les îlots de Langerhans isolés de donneurs en état de mort encéphalique constituent actuellement la seule source de cellules pour la transplantation de patients atteints de diabète de type 1. Cette approche thérapeutique reste cependant compromise par la rareté des donneurs et par certains aspects techniques. L’utilisation de sources alternatives de cellules productrices d’insuline est donc un enjeu tant thérapeutique que pour la recherche pharmacologique. Plusieurs équipes dans le monde, dont la nôtre, développent des modèles de culture cellulaire en 3D, les Langerhanoïdes , qui sont physiologiquement proches des îlots pancréatiques humains. Dans cette revue, nous décrivons les récentes avancées mimant la niche pancréatique (matrice extracellulaire, vascularisation, microfluidique), permettant ainsi d’accroître la fonctionnalité de ces Langerhanoïdes .
... Microtissues in the form of tissue spheroids and other small cell aggregates in 3D are ideal candidates to simulate tissue microenvironments in vivo, which can be reconstituted to generate reproducible complex tissues such as bone [158] and pancreas [159] and cancer tissue models for therapeutic purposes [160]. The assembly of this kind of spheroids can occur in a controlled manner by 3D bioprinting and microfluidic devices [160][161][162]. ...
Efficient strategies to promote microvascularization in vascular tissue engineering, a central priority in regenerative medicine, are still scarce; nano- and micro-sized aggregates and spheres or beads harboring primitive microvascular beds are promising methods in vascular tissue engineering. Capillaries are the smallest type and in numerous blood vessels, which are distributed densely in cardiovascular system. To mimic this microvascular network, specific cell components and proangiogenic factors are required. Herein, advanced biofabrication methods in microvascular engineering, including extrusion-based and droplet-based bioprinting, Kenzan, and biogripper approaches, are deliberated with emphasis on the newest works in prevascular nano- and micro-sized aggregates and microspheres/microbeads.
... Agarose Mold with 3D Printing Technology Agarose multiwell molds provide a suitable environment for spheroid formation, and are an easier and cheaper way to create a non-adhesive surface on which to form spheroids compared to other 3D culture methods. Agarose is biocompatible and non-toxic, which makes it very suitable for cell culture [65]. In other 3D cell culture methods, the size uniformity properties of the spheroids can be a problem. ...
Introduction
Three-dimensional (3D) cell culture studies are becoming extremely common because of their capability to mimic tumor architecture, such as cell-cell and cell-ECM interactions, more efficiently than 2D monolayer systems. These interactions have important roles in defining the tumor cell behaviors, such as proliferation, differentiation, and most importantly, tumor drug response.
Objective
This review aims to provide an overview of the methods for 3D tumor spheroid formation to model human tumors, specifically concentrated on studies using hepatocellular carcinoma (HCC) cells.
Method
We obtained information from previously published articles. In this review, there is discussion of the scaffold and non-scaffold-based approaches, including hanging drop, bioreactors and 3D bioprinting.
Results and Conclusion
The mimicking of the tumor microenvironment (TME) as tumor spheroids could provide a valuable platform for studying tumor biology. Multicellular tumor spheroids are self-assembled cultures of mixed cells (tumor and stromal cells) organized in a 3D arrangement. These spheroids closely mimic the main features of human solid tumors, such as structural organization, central hypoxia, and overall oxygen and nutrient gradients. Hepatocellular carcinoma (HCC) is the most common liver malignancy, and most difficult to overcome because of its drug resistance and tumor heterogeneity. In order to mimic this highly heterogeneous environment, 3D cell culture systems are needed.
... 12,32 Investigations into the combination of different cell types at different ratios have led to the understanding that mimicking the composition of the native islet commonly results in superior β cell function as measured by their glucose responsiveness. 21,33 In rodent islets there is an inner core consisting predominantly of β cells, while α and δ cells reside in the periphery, like a mantle surrounding the islet. Conversely, the human islet has a more random arrangement of the β and α cells with some intermittent clustering of similar cell types. ...
The pancreatic islet of Langerhans is a multicellular system that relies on cell–cell interaction and communication for its function. The most abundant cells in the islets are alpha, beta and endothelial cells, all of which have been shown to positively support each other by generating direct cell–cell interactions, extracellular matrix proteins, or through the secretion of soluble factors into the extracellular space. Knowing how these cell types assemble and support each other to improve viability, migration, and function (glucose responsiveness) is important for the aim of re-establishing beta cell mass with a de novo cell source derived from human cells.
... The surface tension of hMSC-only and hMSC/HUVEC spheroids was measured using a micropipette aspiration technique, as previously described [27]. The customized micropipettes were prepared from borosilicate Pasteur pipettes (vWR, 14 673-043, Radnor, PA) on a P1000 Flaming/Brown micropipette puller (Sutter Instrument, Novato, CA). ...
Conventional top-down approaches in tissue engineering involving cell seeding on scaffolds have been widely used in bone engineering applications. However, scaffold-based bone tissue constructs have had limited clinical translation due to constrains in supporting scaffolds, minimal flexibility in tuning scaffold degradation, and low achievable cell seeding density as compared with native bone tissue. Here, we demonstrate a pragmatic and scalable bottom-up method, inspired from embryonic developmental biology, to build three-dimensional (3D) scaffold-free constructs using spheroids as building blocks. Human umbilical vein endothelial cells (HUVECs) were introduced to human mesenchymal stem cells (hMSCs) (hMSC/HUVEC) and spheroids were fabricated by an aggregate culture system. Bone tissue was generated by induction of osteogenic differentiation in hMSC/HUVEC spheroids for 10 d, with enhanced osteogenic differentiation and cell viability in the core of the spheroids compared to hMSC-only spheroids. Aspiration-assisted bioprinting (AAB) is a new bioprinting technique which allows precise positioning of spheroids (11% with respect to the spheroid diameter) by employing aspiration to lift individual spheroids and bioprint them onto a hydrogel. AAB facilitated bioprinting of scaffold-free bone tissue constructs using the pre-differentiated hMSC/HUVEC spheroids. These constructs demonstrated negligible changes in their shape for two days after bioprinting owing to the reduced proliferative potential of differentiated stem cells. Bioprinted bone tissues showed interconnectivity with actin-filament formation and high expression of osteogenic and endothelial-specific gene factors. This study thus presents a viable approach for 3D bioprinting of complex-shaped geometries using spheroids as building blocks, which can be used for various applications including but not limited to, tissue engineering, organ-on-a-chip and microfluidic devices, drug screening and, disease modeling.
