Cancer cell migration depends on adjacent ASC and adipose spheroids in a 3D bioprinted breast cancer model.

Breast cancer develops in close proximity to mammary adipose tissue and interactions with the local adipose environment have been shown to drive tumor progression. The specific role, however, of this complex tumor microenvironment in cancer cell migration still needs to be elucidated. Therefore, in this study, a 3D bioprinted breast cancer model was developed that allows for a comprehensive analysis of individual tumor cell migration parameters in dependence of adjacent adipose stroma. In this co-culture model, a breast cancer compartment with MDA-MB-231 breast cancer cells embedded in collagen is surrounded by an adipose tissue compartment consisting of adipose-derived stromal cell (ASC) or adipose spheroids in a printable bioink based on thiolated hyaluronic acid. Printing parameters were optimized for adipose spheroids to ensure viability and integrity of the fragile lipid-laden cells. Preservation of the adipogenic phenotype after printing was demonstrated by quantification of lipid content, expression of adipogenic marker genes, the presence of a coherent adipo-specific extracellular matrix, and cytokine secretion. The migration of tumor cells as a function of paracrine signaling of the surrounding adipose compartment was then analyzed using live-cell imaging. The presence of ASC or adipose spheroids substantially increased key migration parameters of MDA-MB-231 cells, namely motile fraction, persistence, invasion distance, and speed. These findings shed new light on the role of adipose tissue in cancer cell migration. They highlight the potential of our 3D printed breast cancer-stroma model to elucidate mechanisms of stroma-induced cancer cell migration and to serve as a screening platform for novel anti-cancer drugs targeting cancer cell dissemination.

Assessment and process optimization of high throughput biofabrication of immunocompetent breast cancer model for drug screening applications.

Recent advancements in 3D cancer modeling have significantly enhanced our ability to delve into the intricacies of carcinogenesis. Despite the pharmaceutical industry's substantial investment of both capital and time in the drug screening and development pipeline, a concerning trend persists: drug candidates screened on conventional cancer models exhibit a dismal success rate in clinical trials. One pivotal factor contributing to this discrepancy is the absence of drug testing on pathophysiologically biomimetic 3D cancer models during pre-clinical stages. Unfortunately, current manual methods of 3D cancer modeling, such as spheroids and organoids, suffer from limitations in reproducibility and scalability. In our study, we have meticulously developed 3D bioprinted breast cancer model utilizing decellularized adipose tissue-based hydrogel obtained via a detergent-free decellularization method. Our innovative printing techniques allows for rapid, high-throughput fabrication of 3D cancer models in a 96-well plate format, demonstrating unmatched scalability and reproducibility. Moreover, we have conducted extensive validation, showcasing the efficacy of our platform through drug screening assays involving two potent anti-cancer drugs, 5-Fluorouracil and PRIMA-1Met. Notably, our platform facilitates effortless imaging and gene expression analysis, streamlining the evaluation process. In a bid to enhance the relevance of our cancer model, we have introduced a heterogeneous cell population into the DAT-based bioink. Through meticulous optimization and characterization, we have successfully developed a biomimetic immunocompetent breast cancer model, complete with microenvironmental cues and diverse cell populations. This breakthrough paves the way for rapid multiplex drug screening and the development of personalized cancer models, marking a paradigm shift in cancer research and pharmaceutical development.

Enhancing scaffold-free spheroid models: 3D cell bioprinting method for metastatic HSC3-Oral squamous carcinoma cell line.

3D in vitro systems offer advantages over the shortcomings of two-dimensional models by simulating the morphological and functional features of in vivo-like environments, such as cell-cell and cell-extracellular matrix interactions, as well as the co-culture of different cell types. Nevertheless, these systems present technical challenges that limit their potential in cancer research requiring cell line- and culture-dependent standardization. This protocol details the use of a magnetic 3D bioprinting method and other associated techniques (cytotoxicity assay and histological analysis) using oral squamous cell carcinoma cell line, HSC3, which offer advantages compared to existing widely used approaches. This protocol is particularly timely, as it validates magnetic bioprinting as a method for the rapid deployment of 3D cultures as a tool for compound screening and development of heterotypic cultures such as co-culture of oral squamous cell carcinoma cells with cancer-associated fibroblasts (HSC3/CAFs).

3D bioprinted mesenchymal stem cell laden scaffold enhances subcutaneous vascularization for delivery of cell therapy.

Subcutaneous delivery of cell therapy is an appealing minimally-invasive strategy for the treatment of various diseases. However, the subdermal site is poorly vascularized making it inadequate for supporting engraftment, viability, and function of exogenous cells. In this study, we developed a 3D bioprinted scaffold composed of alginate/gelatin (Alg/Gel) embedded with mesenchymal stem cells (MSCs) to enhance vascularization and tissue ingrowth in a subcutaneous microenvironment. We identified bio-ink crosslinking conditions that optimally recapitulated the mechanical properties of subcutaneous tissue. We achieved controlled degradation of the Alg/Gel scaffold synchronous with host tissue ingrowth and remodeling. Further, in a rat model, the Alg/Gel scaffold was superior to MSC-embedded Pluronic hydrogel in supporting tissue development and vascularization of a subcutaneous site. While the scaffold alone promoted vascular tissue formation, the inclusion of MSCs in the bio-ink further enhanced angiogenesis. Our findings highlight the use of simple cell-laden degradable bioprinted structures to generate a supportive microenvironment for cell delivery.

[Printing Process Quality Control of Bioprinting Medical Devices].

Objective: This study analyzes the risk points in the quality control of bioink and the main processes of bioprinting, clarifies and explores the quality control and supervision model for bioprinting medical devices, and provides theoretical and practical guidance to ensure the safety and effectiveness of bioprinting medical devices. Methods: The quality control risk points throughout the bioprinting process were comprehensively analyzed, with a particular focus on bioprinting materials and key processes. The regulatory model and methods for bioprinting medical devices were examined. This research concentrated on critical technologies such as extrusion, laser-assisted, and in situ bioprinting, assessing their potential for clinical applications and regulatory challenges. Results: Bioink from different sources should meet regulatory requirements. It is essential to ensure aseptic handling of raw materials and to validate sterilization under "worst-case" conditions. Conclusion: As bioprinting technology advances rapidly, corresponding research into materials, processes, and quality risk control should be conducted to ensure the concurrent development of the regulatory system. This will continuously contribute to the orderly progression of the entire industry and human health.

Revitalizing liver function in mice with liver failure through transplantation of 3D-bioprinted liver with expanded primary hepatocytes.

The utilization of three-dimensional (3D) bioprinting technology to create a transplantable bioartificial liver emerges as a promising remedy for the scarcity of liver donors. This study outlines our strategy for constructing a 3D-bioprinted liver, using in vitro-expanded primary hepatocytes recognized for their safety and enhanced functional robustness as hepatic cell sources for bioartificial liver construction. In addition, we have developed bioink biomaterials with mechanical and rheological properties, as well as printing capabilities, tailored for 3D bioprinting. Upon heterotopic transplantation into the mesentery of tyrosinemia or 90% hepatectomy mice, our 3D-bioprinted liver effectively restored lost liver functions, consequently extending the life span of mice afflicted with liver injuries. Notably, the inclusion of an artificial blood vessel in our 3D-bioprinted liver allowed for biomolecule exchange with host blood vessels, demonstrating, in principle, the rapid integration of the bioartificial liver into the host vascular system. This model underscores the therapeutic potential of transplantation for the treatment of liver failure diseases.

Characterization of two different alginate-based bioinks and the influence of melanoma growth within.

Extrusion-based bioprinting is an established method in biofabrication. Suitable bioinks have fundamentally different compositions and characteristics, which should be examined, in order to find a perfect model system. Here, we investigate the effect of two alginate-based, yet unalike 3D-printed bioinks, pre-crosslinked alginate-dialdehyde gelatin (ADA-GEL) and a mixture of alginate, hyaluronic acid, and gelatin (Alg/HA/Gel), on the melanoma cell line Mel Im and vice versa in terms of stiffness, shrinkage, cellular behavior and colony formation over 15 days. Rheological stiffness measurements revealed two soft gels with similar storage moduli. The cells did not have a significant impact on the overall stiffness, whereas ADA-GEL (2.5/2.5%) was significantly stiffer than Alg/HA/Gel (0.5/0.1/3%). Regarding the shrinkage of printed constructs, cells had a significant influence, especially in ADA-GEL, which has covalent bonds between the oxidized alginate and gelatin. Multi-photon microscopy exhibited proliferation, cell spreading and migration in ADA-GEL with cell-cell and cell-matrix interaction, dissimilarly to Alg/HA/Gel, in which cells formed spherical, encapsulated colonies. Scanning electron microscopy and histology showed degradation and multi-layered growth on ADA-GEL and fewer examples of escaped cells on Alg/HA/Gel. Both gels serve as proliferation bioink for melanoma with more necrosis in deeper Alg/HA/Gel colonies and differences in spreading and matrix interaction. These findings show the importance of proper characterization of the bioinks for different applications.

3D-bioprinted GelMA/gelatin/amniotic membrane extract (AME) scaffold loaded with keratinocytes, fibroblasts, and endothelial cells for skin tissue engineering.

Gelatin-methacryloyl (GelMA) is a highly adaptable biomaterial extensively utilized in skin regeneration applications. However, it is frequently imperative to enhance its physical and biological qualities by including supplementary substances in its composition. The purpose of this study was to fabricate and characterize a bi-layered GelMA-gelatin scaffold using 3D bioprinting. The upper section of the scaffold was encompassed with keratinocytes to simulate the epidermis, while the lower section included fibroblasts and HUVEC cells to mimic the dermis. A further step involved the addition of amniotic membrane extract (AME) to the scaffold in order to promote angiogenesis. The incorporation of gelatin into GelMA was found to enhance its stability and mechanical qualities. While the Alamar blue test demonstrated that a high concentration of GelMA (20%) resulted in a decrease in cell viability, the live/dead cell staining revealed that incorporation of AME increased the quantity of viable HUVECs. Further, gelatin upregulated the expression of KRT10 in keratinocytes and VIM in fibroblasts. Additionally, the histological staining results demonstrated the formation of well-defined skin layers and the creation of extracellular matrix (ECM) in GelMA/gelatin hydrogels during a 14-day culture period. Our study showed that a 3D-bioprinted composite scaffold comprising GelMA, gelatin, and AME can be used to regenerate skin tissues.

Advanced optical assessment and modeling of extrusion bioprinting.

In the context of tissue engineering, biofabrication techniques are employed to process cells in hydrogel-based matrices, known as bioinks, into complex 3D structures. The aim is the production of functional tissue models or even entire organs. The regenerative production of biological tissues adheres to a multitude of criteria that ultimately determine the maturation of a functional tissue. These criteria are of biological nature, such as the biomimetic spatial positioning of different cell types within a physiologically and mechanically suitable matrix, which enables tissue maturation. Furthermore, the processing, a combination of technical procedures and biological materials, has proven highly challenging since cells are sensitive to stress, for example from shear and tensile forces, which may affect their vitality. On the other hand, high resolutions are pursued to create optimal conditions for subsequent tissue maturation. From an analytical perspective, it is prudent to first investigate the printing behavior of bioinks before undertaking complex biological tests. According to our findings, conventional shear rheological tests are insufficient to fully characterize the printing behavior of a bioink. For this reason, we have developed optical methods that, complementarily to the already developed tests, allow for quantification of printing quality and further viscoelastic modeling of bioinks.

Nylons with Applications in Energy Generators, 3D Printing and Biomedicine.

Linear polyamides, known as nylons, are a class of synthetic polymers with a wide range of applications due to their outstanding properties, such as chemical and thermal resistance or mechanical strength. These polymers have been used in various fields: from common and domestic applications, such as socks and fishing nets, to industrial gears or water purification membranes. By their durability, flexibility and wear resistance, nylons are now being used in addictive manufacturing technology as a good material choice to produce sophisticated devices with precise and complex geometric shapes. Furthermore, the emergence of triboelectric nanogenerators and the development of biomaterials have highlighted the versatility and utility of these materials. Due to their ability to enhance triboelectric performance and the range of applications, nylons show a potential use as tribo-positive materials. Because of the easy control of their shape, they can be subsequently integrated into nanogenerators. The use of nylons has also extended into the field of biomaterials, where their biocompatibility, mechanical strength and versatility have paved the way for groundbreaking advances in medical devices as dental implants, catheters and non-absorbable surgical sutures. By means of 3D bioprinting, nylons have been used to develop scaffolds, joint implants and drug carriers with tailored properties for various biomedical applications. The present paper aims to collect evidence of these recently specific applications of nylons by reviewing the literature produced in recent decades, with a special focus on the newer technologies in the field of energy harvesting and biomedicine.

Scaffolding fundamentals and recent advances in sustainable scaffolding techniques for cultured meat development.

In cultured meat (CM) production, Scaffolding plays an important role by aiding cell adhesion, growth, differentiation, and alignment. The existence of fibrous microstructure in connective and muscle tissues has attracted considerable interest in the realm of tissue engineering and triggered the interest of researchers to implement scaffolding techniques. A wide array of research efforts is ongoing in scaffolding technologies for achieving the real meat structure on the principality of biomedical research and to replace serum free CM production. Scaffolds made of animal-derived biomaterials are found efficient in replicating the extracellular matrix (ECM), thus focus should be paid to utilize animal byproducts for this purpose. Proper identification and utilization of plant-derived scaffolding biomaterial could be helpful to add diversified options in addition to animal derived sources and reduce in cost of CM production through scaffolds. Furthermore, techniques like electrospinning, modified electrospinning and 3D bioprinting should be focused on to create 3D porous scaffolds to mimic the ECM of the muscle tissue and form real meat-like structures. This review discusses recent advances in cutting edge scaffolding techniques and edible biomaterials related to structured CM production.

Alternative Therapies in Transplantology as a Promising Perspective in Medicine.

Despite continuous and rapid progress in the transplantation of cells, tissues, and organs, many patients die before receiving them. This is because of an insufficient number of donors, which leads to a significant disproportion between the need for donors and their availability. This review aims to present the possibilities offered by alternative therapies. We use the term "functional transplantology" to describe such alternative methods of transplantation that could help change the current state of transplantation medicine. Its purpose is not to replace a defective or removed organ with another but to replace its functions using complementary biological, mechanical, or biomechanical structures or devices. Implementation of many innovative solutions shown in the work for clinical applications is already a fact. In the case of others, it should be considered a future vision. We hope that the role of a defective or damaged tissue or a group of tissues will be taken over by different structures that are functionally complementary with the organ being substituted. Undoubtedly, developing the described methods based on functional transplantology will change the face of transplantation medicine. Thus, we show current trends and new directions of thinking and actions in transplantation medicine that combine technology and transplantology. The review considers the latest technologies, including 3D bioprinting, nanotechnology, cell encapsulation, and organoids. We discuss not only the advantages of new approaches but also the limitations and challenges that must be overcome to achieve significant progress in transplantation. That is the only option to provide a safe and efficient way of improving the quality of life of many patients.

[Research progress of three-dimensional bioprinting technology in auricle repair and reconstruction].

Objective: To review the research progress on the application of three-dimensional (3D) bioprinting technology in auricle repair and reconstruction. Methods: The recent domestic and international research literature on 3D printing and auricle repair and reconstruction was extensively reviewed, and the concept of 3D bioprinting technology and research progress in auricle repair and reconstruction were summarized. Results: The auricle possesses intricate anatomical structure and functionality, necessitating precise tissue reconstruction and morphological replication. Hence, 3D printing technology holds immense potential in auricle reconstruction. In contrast to conventional 3D printing technology, 3D bioprinting technology not only enables the simulation of auricular outer shape but also facilitates the precise distribution of cells within the scaffold during fabrication by incorporating cells into bioink. This approach mimics the composition and structure of natural tissues, thereby favoring the construction of biologically active auricular tissues and enhancing tissue repair outcomes. Conclusion: 3D bioprinting technology enables the reconstruction of auricular tissues, avoiding potential complications associated with traditional autologous cartilage grafting. The primary challenge in current research lies in identifying bioinks that meet both the mechanical requirements of complex tissues and biological criteria.

Innovative technologies for the fabrication of 3D/4D smart hydrogels and its biomedical applications - A comprehensive review.

Repairing and regenerating damaged tissues or organs, and restoring their functioning has been the ultimate aim of medical innovations. 'Reviving healthcare' blends tissue engineering with alternative techniques such as hydrogels, which have emerged as vital tools in modern medicine. Additive manufacturing (AM) is a practical manufacturing revolution that uses building strategies like molding as a viable solution for precise hydrogel manufacturing. Recent advances in this technology have led to the successful manufacturing of hydrogels with enhanced reproducibility, accuracy, precision, and ease of fabrication. Hydrogels continue to metamorphose as the vital compatible bio-ink matrix for AM. AM hydrogels have paved the way for complex 3D/4D hydrogels that can be loaded with drugs or cells. Bio-mimicking 3D cell cultures designed via hydrogel-based AM is a groundbreaking in-vivo assessment tool in biomedical trials. This brief review focuses on preparations and applications of additively manufactured hydrogels in the biomedical spectrum, such as targeted drug delivery, 3D-cell culture, numerous regenerative strategies, biosensing, bioprinting, and cancer therapies. Prevalent AM techniques like extrusion, inkjet, digital light processing, and stereo-lithography have been explored with their setup and methodology to yield functional hydrogels. The perspectives, limitations, and the possible prospects of AM hydrogels have been critically examined in this study.

3D-Bioprinted Hepar-on-a-Chip Implanted in Graphene-Based Plasmonic Sensors.

Precise three-dimensional (3D) bioprinting designs enable the fabrication of unique structures for 3D-cell culture models. There is still an absence of real-time detection tools to effectively track in situ 3D-cell performance, hindering a comprehensive understanding of disease progression and drug efficacy assessment. While numerous bioinks have been developed, few are equipped with internal sensors capable of accurate detection. This study addresses these challenges by constructing a 3D-bioprinted hepar-on-a-chip embedded with graphene quantum dot-capped gold nanoparticle-based plasmonic sensors, featuring strong surface-enhanced Raman scattering (SERS) enhancement, biostability, and signal consistency. Such an integrated hepar-on-a-chip demonstrates excellent capability in the secretion of liver function-related proteins and the expression of drug metabolism and transport-related genes. Furthermore, the on-site detection of cell-secreted biomarker glutathione transferase α (GST-α) was successfully achieved using the plasmonic probe, with a dynamic linear detection range of 20-500 ng/mL, showcasing high anti-interference and specificity for GST-α. Ultimately, this integrated hepar-on-a-chip system offers a high-quality platform for monitoring liver injury.

Embedded Bioprinting of Tissue-like Structures Using κ-Carrageenan Sub-Microgel Medium.

Embedded three-dimensional (3D) bioprinting utilizing a granular hydrogel supporting bath has emerged as a critical technique for creating biomimetic scaffolds. However, engineering a suitable gel suspension medium that balances precise bioink deposition with cell viability and function presents multiple challenges, particularly in achieving the desired viscoelastic properties. Here, a novel κ-carrageenan gel supporting bath is fabricated through an easy-to-operate mechanical grinding process, producing homogeneous sub-microscale particles. These sub-microgels exhibit typical Bingham flow behavior with small yield stress and rapid shear-thinning properties, which facilitate the smooth deposition of bioinks. Moreover, the reversible gel-sol transition and self-healing capabilities of the κ-carrageenan microgel network ensure the structural integrity of printed constructs, enabling the creation of complex, multi-layered tissue structures with defined architectural features. Post-printing, the κ-carrageenan sub-microgels can be easily removed by a simple phosphate-buffered saline wash. Further bioprinting with cell-laden bioinks demonstrates that cells within the biomimetic constructs have a high viability of 92% and quickly extend pseudopodia, as well as maintain robust proliferation, indicating the potential of this bioprinting strategy for tissue and organ fabrication. In summary, this novel κ-carrageenan sub-microgel medium emerges as a promising avenue for embedded bioprinting of exceptional quality, bearing profound implications for the in vitro development of engineered tissues and organs.

Droplet bioprinting of acellular and cell-laden structures at high-resolutions.

Advances in digital light projection(DLP) based (bio) printers have made printing of intricate structures at high resolution possible using a wide range of photosensitive bioinks. A typical setup of a DLP bioprinter includes a vat or reservoir filled with liquid bioink, which presents challenges in terms of cost associated with bioink synthesis, high waste, and gravity-induced cell settling, contaminations, or variation in bioink viscosity during the printing process. Here, we report a vat-free, low-volume, waste-free droplet bioprinting method capable of rapidly printing 3D soft structures at high resolution using model bioinks and model cells. A multiphase many-body dissipative particle dynamics model was developed to simulate the dynamic process of droplet-based DLP printing and elucidate the roles of surface wettability and bioink viscosity. Process variables such as light intensity, photo-initiator concentration, and bioink formulations were optimized to print 3D soft structures (∼0.4-3 kPa) with a typical layer thickness of 50µm, an XY resolution of 38 ± 1.5µm and Z resolution of 237 ± 5.4µm. To demonstrate its versatility, droplet bioprinting was used to print a range of acellular 3D structures such as a lattice cube, a Mayan pyramid, a heart-shaped structure, and a microfluidic chip with endothelialized channels. Droplet bioprinting, performed using model C3H/10T1/2 cells, exhibited high viability (90%) and cell spreading. Additionally, microfluidic devices with internal channel networks lined with endothelial cells showed robust monolayer formation while osteoblast-laden constructs showed mineral deposition upon osteogenic induction. Overall, droplet bioprinting could be a low-cost, no-waste, easy-to-use, method to make customized bioprinted constructs for a range of biomedical applications.

A comparative analysis of 3D bioprinted gelatin-hyaluronic acid-alginate scaffold and microfracture for the management of osteochondral defects in the rabbit knee joint.

OBJECTIVES: This study aims to compare the radiological, biomechanical, and histopathological results of microfracture treatment and osteochondral damage repair treatment with a new scaffold product produced by the three-dimensional (3D) bioprinting method containing gelatin-hyaluronic acid-alginate in rabbits with osteochondral damage. MATERIALS AND METHODS: A new 3D bioprinted scaffold consisting of gelatin, hyaluronic acid, and alginate designed by us was implanted into the osteochondral defect created in the femoral trochlea of 10 rabbits. By randomization, it was determined which side of 10 rabbits would be repaired with a 3D bioprinted scaffold, and microfracture treatment was applied to the other knees of the rabbits. After six months of follow-up, the rabbits were sacrificed. The results of both treatment groups were compared radiologically, biomechanically, and histopathologically. RESULTS: None of the rabbits experienced any complications. The magnetic resonance imaging evaluation showed that all osteochondral defect areas were integrated with healthy cartilage in both groups. There was no significant difference between the groups in the biomechanical load test (p=0.579). No statistically significant difference was detected in the histological examination using the modified Wakitani scores (p=0.731). CONCLUSION: Our study results showed that 3D bioprinted scaffolds exhibited comparable radiological, biomechanical, and histological properties to the conventional microfracture technique for osteochondral defect treatment.

Assessing Bioprinted Functionalized Grafts for Biological Tendon Augmentation In Vitro.

Tendinopathy, characterized by inflammatory and degenerative changes, presents challenges in sports and medicine. In addressing the limitations of conservative management, this study focuses on developing tendon grafts using extrusion bioprinting with platelet-rich plasma (PRP)-infused hydrogels loaded with tendon cells. The objective is to understand paracrine interactions initiated by bioprinted tendon grafts in either inflamed or non-inflamed host tissues. PRP was utilized to functionalize methacrylate gelatin (GelMA), incorporating tendon cells for graft bioprinting. Bioinformatic analyses of overexpressed proteins, predictive of functional enrichment, revealed insights into PRP graft behavior in both non-inflamed and inflamed environments. PRP grafts activated inflammatory pathways, including Interleukin 17 (IL-17), neuroinflammation, Interleukin 33 (IL-33), and chemokine signaling. Interleukin 1 beta (IL-1b) in the graft environment triggered p38 mitogen-activated protein kinase (MAPK) signaling, nuclear factor kappa light chain enhancer of activated B cells (NF-kB) canonical pathway, and Vascular Endothelial Growth Factor (VEGF) signaling. Biological enrichment attributed to PRP grafts included cell chemotaxis, collagen turnover, cell migration, and angiogenesis. Acellular PRP grafts differed from nude grafts in promoting vessel length, vessel area, and junction density. Angiogenesis in cellular grafts was enhanced with newly synthesized Interleukin 8 (IL-8) in cooperation with IL-1b. In conclusion, paracrine signaling from PRP grafts, mediated by chemokine activities, influences cell migration, inflammation, and angiogenic status in host tissues. Under inflammatory conditions, newly synthesized IL-8 regulates vascularization in collaboration with PRP.

Silk fibroin increases the elasticity of alginate-gelatin hydrogels and regulates cardiac cell contractile function in cardiac bioinks.

Silk fibroin (SF) is a natural protein extracted fromBombyx morisilkworm thread. From its common use in the textile industry, it emerged as a biomaterial with promising biochemical and mechanical properties for applications in the field of tissue engineering and regenerative medicine. In this study, we evaluate for the first time the effects of SF on cardiac bioink formulations containing cardiac spheroids (CSs). First, we evaluate if the SF addition plays a role in the structural and elastic properties of hydrogels containing alginate (Alg) and gelatin (Gel). Then, we test the printability and durability of bioprinted SF-containing hydrogels. Finally, we evaluate whether the addition of SF controls cell viability and function of CSs in Alg-Gel hydrogels. Our findings show that the addition of 1% (w/v) SF to Alg-Gel hydrogels makes them more elastic without affecting cell viability. However, fractional shortening (FS%) of CSs in SF-Alg-Gel hydrogels increases without affecting their contraction frequency, suggesting an improvement in contractile function in the 3D cultures. Altogether, our findings support a promising pathway to bioengineer bioinks containing SF for cardiac applications, with the ability to control mechanical and cellular features in cardiac bioinks.