Tissue Engineering Research Articles

Three-Dimensional Bioprinting of GelMA Hydrogels with Culture Medium: Balancing Printability, Rheology and Cell Viability for Tissue Regeneration

Three-dimensional extrusion bioprinting technology aims to become a fundamental tool for tissue regeneration using cell-loaded hydrogels. These biomaterials must have highly specific mechanical and biological properties that allow them to generate biosimilar structures by successive layering of material while maintaining cell viability. The rheological properties of hydrogels used as bioinks are critical to their printability. Correct printability of hydrogels allows the replication of biomimetic structures, which are of great use in medicine, tissue engineering and other fields of study that require the three-dimensional replication of different tissues. When bioprinting cell-loaded hydrogels, a small amount of culture medium can be added to ensure adequate survival, which can modify the rheological properties of the hydrogels. GelMA is a hydrogel used in bioprinting, with very interesting properties and rheological parameters that have been studied and defined for its basic formulation. However, the changes that occur in its rheological parameters and therefore in its printability, when it is mixed with the culture medium necessary to house the cells inside, are unknown. Therefore, in this work, a comparative study of GelMA 100% and GelMA in the proportions 3:1 (GelMA 75%) and 1:1 (GelMA 50%) with culture medium was carried out to determine the printability of the gel (using a device of our own invention), its main rheological parameters and its toxicity after the addition of the medium and to observe whether significant differences in cell viability occur. This raises the possibility of its use in regenerative medicine using a 3D extrusion bioprinter.

How framing bias impacts preferences for innovation in bone tissue engineering.

It is currently unknown if surgeons and biomaterial scientists &or tissue engineers (BS&orTE) process and evaluate information in similar or different (un)biased ways. For the gold standard of surgery to move "from bench to bedside", there must naturally be synergies between these key stakeholders' perspectives. Because only a small number of biomaterials & tissue engineering innovations have been translated into the clinic today, we hypothesised this lack of translation is rooted in the psychology of surgeons and BS&orTE. Presently, both clinicians and researchers doubt the compatibility of surgery and research in their daily routines. This has led to the use of a metaphorical expression "squaring of the circle," which implies an unsolvable challenge. As bone tissue engineering belongs to the top 5 research areas in tissue engineering we choose the field of bone defect treatment options for our bias study. Our study uses an online survey instrument for data capture: incorporating a behavioural economics cognitive framing experiment methodology. Our study sample consisted of surgeons (n=208) and BS&orTE (n=59). And we employed a convenience sampling method, with participants (conference attendants) being approached both in person and via email - 22 October 2022-13 March 2023. We find no distinct positive-negative cognitive framing differences by occupation. That is, any framing bias present in this surgical decision-making setting does not appear to differ significantly between surgeon and BS&orTE specialisation. When we explored within group differences by frames, we see statistically significant (p<0.05) results for surgeons in the positive frame ranking autologous bone graft transplantation lower compared to surgeons in the negative frame. Further, surgeons in the positive frame rank Ilizarov bone transport method higher compared to surgeons in the negative frame (p<0.05).

Bibliometric Analyses of the Research Trends of Female Pelvic Organ Prolapse.

The study was aimed at systematically analyzing the research status and trends of pelvic organ prolapse (POP) using bibliometrics. We retrieved documents published between 1975 and 2022 from the Web of Science Core Collection (WoSCC) database, and manually selected them for bibliometric analyses of country, institution, journal, highly locally cited documents and research trends based on co-citation clustering and keywords using the R Bibliometricx package and CiteSpace software. A total of 5,703 publications were included. Although the number of annual publications on POP increased, the trend of annual publication reached an obvious plateau in the first half of the 2010s. The USA, China, the UK, the University of Michigan, the University of Pittsburgh, and the University of Sydney were the top three countries and institutions with the most publications respectively. International Urogynecology Journal, American Journal of Obstetrics and Gynecology, and Obstetrics and Gynecology were the journals with the most extensive academic influence on the field of POP research. The international cooperation was lacking and the highly cited documents focused on high-level, evidence-based studies. Epidemiological studies and surgical treatment have achieved a plateau or decline. Recent studies have focused on conservative treatment, physical therapy, and minimally invasive surgery. In addition to evidence-based medicine studies, tissue engineering is the future direction of POP. This study used bibliometric analyses to provide insights into the status and potential research directions of POP. More high-quality, evidence-based medicine studies and in-depth tissue engineering research should be propelled forward.

Cellularised Biomaterials Used as Gingival Connective Tissue Substitutes In Vivo: Systematic Review.

Developing an in vitro model of gingival connective tissue, mimicking the original structure and composition of gingiva for clinical grafting is relevant for personalised treatment of missing gingiva. Using tissue engineering techniques allows to bypass limitations encountered with existing solutions to increase oral soft tissue volume. This review aims to systematically analyse the different currently existing cellularised materials and technologies used to engineer gingival substitutes for in vivo applications. The Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines were followed. An electronic search on PubMed, Scopus, Web of Science and Cochrane Library databases was conducted to identify suitable studies. In vivo studies about gingival substitute and graft containing oral cells compared with a control to investigate the graft remodelling were included. Risk of bias in the included studies was assessed using the Systematic Review Centre for Laboratory animal Experimentation (SYRCLE) 10-item checklist. Out of 631 screened studies, 19 were included. Animal models were mostly rodents, and the most used implantation was subcutaneous. According to the SYRCLE tool, low-to-unclear risk of bias were prevalent. Studies checked vascularisation and extracellular remodelling up to 60 days after implantation of the cellularized biomaterial. Cells used were mostly fibroblasts and stem cells from oral origin. Grafts presenting vascularisation potential after implantation were produced by tissue engineering technologies including cell seeding or embedding for 14, cell sheets for 2, microsphere for 1 and extrusion 3D bioprinting for 2. Components used to build the scaffold containing the cells are all naturally derived and are mainly fibrin, gelatine, collagen, agarose, alginate, fibroin, guar gum, hyaluronic acid, and decellularised extracellular matrix. The most recurring crosslinking method was using chemicals. All studies except 1 reported vascularisation of the graft after implantation and some detailed extracellular matrix remodelling. Current solutions are not efficient enough. By assessing the relevant studies on the subject, this systematic review showed that a diversity of cellularised biomaterials substituting gingival connective tissue enable vascularisation and extracellular remodelling. Taking results of this review in count could help improving current bio-inks used in 3D bioprinting for in vivo applications compensating for gingival loss.

Integration of BMP-2/PLGA microspheres with the 3D printed PLGA/CaSO4 scaffold enhances bone regeneration

Treatment of large and complex irregular bone defects is a major clinical challenge in orthopedic surgery. The current treatment includes bone transportation using the Ilizarov technique and bone cement repair using the Masquelet technique, but they require long-term manual intervention or secondary operation. To improve this situation, we compared the different implanting materials in the literature published in the past 10 years, finding that glycolic acid copolymer (PLGA) and Calcium sulfate (CaSO4) are appropriated to be used as synthetic bone materials due to their advantages of easy-availability, nontoxicity, osteogenic properties and rapid degradation. Meanwhile, the development of 3D printing technique and devices makes it relatively easier to synthetize customized bio-mimetic porous scaffolds, thus facilitating the release of modified protein. In this study, we compounded BMP-2/PLGA microspheres with polylactic glycolic acid copolymer/CaSO4 (PC) 3D printed scaffold to improve the osteogenic properties of the scaffold. The result of our in vitro experiment demonstrated that the prepared PCB scaffold not only had satisfactory bio-compatibility, but also promoted osteogenic differentiation. This 3D printed scaffold is capable to accelerate the repair of complex bone defects by promoting new bone formation, suggesting that it may prove to be a potential bone tissue engineering substitute.

Device Design and Advanced Computed Tomography of 3D Printed Radiopaque Composite Scaffolds and Meniscus

Abstract3D‐printed biomaterial implants are revolutionizing personalized medicine for tissue repair, especially in orthopedics. In this study, a radiopaque bismuth oxide (Bi2O3) doped polycaprolactone (PCL) composite is developed and implemented to enable the use of diagnostic X‐ray technologies, especially spectral photon counting X‐ray computed tomography (SPCCT), for comprehensive tissue engineering scaffold (TES) monitoring. PCL filament with homogeneous Bi2O3 nanoparticle (NP) dispersion (0.8 to 11.7 wt%) is first fabricated. TES are then 3D printed with the composite filament, optimizing printing parameters for small features and severely overhung geometries. These composite TES are characterized via micro‐computed tomography (µCT), tensile testing, and a cytocompatibility study, with 2 wt% Bi2O3 NPs providing improved tensile properties, equivalent cytocompatibility to neat PCL, and excellent radiographic distinguishability. Radiographic performance is validated in situ by imaging 4 and 7 wt% Bi2O3 doped PCL TES in a mouse model with µCT, showing excellent agreement with in vitro measurements. Subsequently, CT image‐derived swine menisci are 3D printed with composite filament and re‐implanted in corresponding swine legs ex vivo. Re‐imaging the swine legs via clinical CT allows facile identification of device location and alignment. Finally, the emergent technology of SPCCT unambiguously distinguishes the implanted meniscus in situ via color K‐edge imaging.

Effect of Physicochemical Surface Properties of Silicon-Substitute hydroxyapatite on Angiogenesis.

Synthetic hydroxyapatite (HA) is a widely studied bioceramic for bone tissue engineering (BTE) due to its similarity to the mineral component of bone. As bone mineral contains various ionic substitutions that play a crucial role in bone metabolism, the bioactivity of HA can be improved by adding small amounts of physiologically relevant ions into its crystal structure, with silicate-substituted HA (Si-HA) showing particularly promising results. Nevertheless, it remains unclear how distinct material characteristics influence the bioactivity due to the intertwined nature of surface properties. A co-culture methodology was optimised and applied for in vitro quantification of the biological response. Initially, HA and Si-HA samples were produced and characterised. To compare the bioactivity of the samples, a method was developed to measure interactions in an increasingly complex environment, first including fibronectin (FN) adsorption and subsequently cell adhesion in mono- and co-culture using primary human osteoblasts (hOBs) and human dermal microvascular endothelial cells (HDMECs), with- and without FN pre-coating. An experimental set-up was designed to assess to what extent different surface features of the samples contribute to the induced biological response. An 8 nm gold-sputter coating was applied to eradicate the electrochemical differences and polishing and abrading was used to reduce the differences in surface topographies. Overall, 1.25wt%Si-HA exhibited most nano-scale variations in surface potential. In terms of bioactivity, 1.25wt%Si-HA samples induced the highest osteoblast attachment and vessel formation. Additionally, in vitro vessel formation was established on Si-HA surfaces using a hOB;HDMEC cell ratio of 70:30 and a methodology was established that enabled the assessment of the relative effect of topographical and electrochemical features induced by silicon substitution in the HA lattice on their bioactivity. It was found that the difference in the amount of protein attached to HA and 1.25wt%Si-HA after 2 hours was affected by topographical differences. Conversely, electrochemical differences induced different vessel-like structure formation in co-culture with a FN pre-coating. Without a FN pre-coating, both topographical and electrochemical differences dictated the differences in angiogenic response. Overall, 1.25wt%Si-HA surface features appear to induce the most favourable protein adsorption and cell adhesion in mono- and co-culture with- and without FN pre-coating.

Directed biomechanical compressive forces enhance fusion efficiency in model placental trophoblast cultures.

The syncytiotrophoblast is a multinucleated structure that arises from fusion of mononucleated cytotrophoblasts, to sheath the placental villi and regulate transport across the maternal-fetal interface. Here, we ask whether the dynamic mechanical forces that must arise during villous development might influence fusion, and explore this question using in vitro choriocarcinoma trophoblast models. We demonstrate that mechanical stress patterns arise around sites of localized fusion in cell monolayers, in patterns that match computational predictions of villous morphogenesis. We then externally apply these mechanical stress patterns to cell monolayers and demonstrate that equibiaxial compressive stresses (but not uniaxial or equibiaxial tensile stresses) enhance expression of the syndecan-1 and loss of E-cadherin as markers of fusion. These findings suggest that the mechanical stresses that contribute towards sculpting the placental villi may also impact fusion in the developing tissue. We then extend this concept towards 3D cultures and demonstrate that fusion can be enhanced by applying low isometric compressive stresses to spheroid models, even in the absence of an inducing agent. These results indicate that mechanical stimulation is a potent activator of cellular fusion, suggesting novel avenues to improve experimental reproductive modelling, placental tissue engineering, and understanding disorders of pregnancy development.

Skin Grafts in Plastic Surgery: A Comprehensive Review of Procedures, Techniques, and Clinical Considerations

Objective: This study aims to analyze the progress and challenges related to the use of skin grafts in plastic surgery, with the purpose of providing a comprehensive understanding of the associated techniques and clinical considerations. Theoretical Framework: A review of the fundamentals of plastic surgery is presented, emphasizing the importance of skin grafts in wound and deformity reconstruction, as well as exploring the impact of tissue engineering on the evolution of these techniques. Method: This study is based on a systematic review of the scientific literature, following recognized guidelines. Relevant articles addressing different types of skin grafts, their clinical applications, and associated procurement techniques were selected. Results and Discussion: The results of the review highlight the variety of skin grafts available and their respective indications. The discussion addresses the challenges in selecting the appropriate type of graft and the ethical issues related to its procurement and use. Implications of the Research: This study offers important insights for plastic surgeons and researchers on best practices in skin graft utilization, emphasizing the need for ethical considerations and specific care in graft selection and procurement. Originality/Value: It contributes to the literature by providing a comprehensive analysis of skin graft techniques in plastic surgery, including recent advancements and emerging issues. Its relevance lies in the practical guidance it offers to professionals in the field.

Comparing fabrication techniques for engineered cardiac tissue.

Tissue engineering can provide in vitro models for drug testing, disease modeling, and perhaps someday, tissue/organ replacements. For building 3D heart tissue, the alignment of cardiac cells or cardiomyocytes (CMs) is important in generating a synchronously contracting tissue. To that end, researchers have generated several fabrication methods for building heart tissue, but direct comparisons of pros and cons using the same cell source is lacking. Here, we derived cardiomyocytes (CMs) from human induced pluripotent stem cells (hiPSCs) and compare the assembly of these cells using three fabrication methods: cardiospheres, muscle rings, and muscle strips. All three protocols successfully generated compacted tissue comprised of hiPSC-derived CMs stable for at least 2 weeks. The percentage of aligned cells was greatest in the muscle strip (55%) and the muscle ring (50%) compared with the relatively unaligned cardiospheres (35%). The iPSC-derived CMs within the muscle strip also exhibited the greatest elongation, with elongation factor at 2.0 compared with 1.5 for the muscle ring and 1.2 for the cardiospheres. This is the first direct comparison of various fabrication techniques using the same cell source.

The Effect of Assam Silk Fibroin on the Bacterial Cellulose Cytocompatibility as the Extracellular Matrix

AbstractThe growing interest in natural polymers for biomedical and tissue engineering has fueled the search for materials with superior mechanical and biological properties. Bacterial cellulose (BC) and silk fibroin (SF) emerge as promising candidates meeting these criteria. This study focuses on enhancing BC through high‐pressure homogenization (HPH) and subsequent functionalization with Assam silk fibroin (ASF) using an ex‐situ approach. Analysis via attenuated total reflection–Fourier transform infrared (ATR–FTIR) spectroscopy confirms the successful integration of ASF into the HPH‐treated BC scaffold. The diffractogram of HPH‐BC/ASF indicates the prevalence of type I cellulose crystalline structures, with variations in lamellar and porous architecture based on component ratios. Mechanical testing, particularly the compressive test, reveals that the HPH‐BC/ASF formulation exhibits the highest compressive stress and modulus compared to other samples. Supplementary analyses, including swelling ratio and porosity measurements, support the superior compressive properties of HPH‐BC/ASF. Moreover, the cell viability of chondrocytes demonstrates compatibility with the BC‐based scaffold material. These findings underscore the potential applications of HPH‐BC and ASF in areas such as scaffolding for the development of extracellular matrix.

Steering Piezocatalytic Therapy for Optimized Tumoricidal Effect

AbstractPiezocatalysts, because of their mechano‐electrical conversion properties, are exploited for various medical applications, such as sterilization, tissue engineering, biosensing, and disease theranostics. In particular, based on the unique advantage of the piezoelectric effect, piezocatalytic therapy (PCT) has been developed as a novel and promising candidate for tumor therapy. To optimize the utilization of piezocatalysts in tumor therapy, a comprehensive understanding of the antitumor mechanism associated with these materials is imperative. Here, the piezocatalytic action principle is elucidated by investigating piezocatalysts, reactants, energy inputs, and products. Subsequently, the antitumor mechanisms of PCT have been extensively discussed and are recapitulative as follows: restraining cell proliferation, inducing cell programmed death, hindering tumor metastasis, inhibiting tumor angiogenesis, and enhancing antitumor immunity. Additionally, the optimized therapeutic outcomes of PCT‐centric synergistic cancer therapy are systematically described. Finally, the main challenges and future research directions of piezocatalysis applied in cancer therapy are envisioned. It is believed that PCT will serve as a new‐generation ingenious tool for cancer treatment.

Biomimetic grafts from ultrafine fibers for collagenous tissues.

The ligament is the soft tissue that connects bone to bone and, in case of severe injury or rupture, it cannot heal itself mainly because of its poor vascularity and dynamic nature. Tissue engineering carries the potential to restore the injured tissue functions by utilization of scaffolds mimicking the structure of native ligament. Collagen fibrils in the anterior cruciate ligament (ACL) have a diameter ranging from 20 to 300nm, which defines the physical and mechanical properties of the tissue. Also, the ACL tissue exhibited a bimodal distribution of collagen fibrils. Currently, the ability to fabricate scaffolds replicating this structure is a significant challenge. This work aims at i) measuring the diameter of collagens of bovine ACL tissue, ii) investigating the fabrication of sub-100nm fibers, and iii) fabricating aligned scaffolds with bimodal diameter distribution (with two peaks) resembling the healthy ACL structure. It is hypothesized that such scaffolds can be produced by electrospinning polycaprolactone (PCL) solutions. To test the hypothesis, various PCL solutions were formulated in acetone and formic acid in combination with pyridine, and electrospun to generate sub-100nm fibers. Next, this formulation was adjusted to produce nanofibers with a diameter between 100nm and 200nm. Finally, these solutions were combined in the co-electrospinning process, i.e., two-spinneret electrospinning, to fabricate biomimetic scaffolds with a bimodal distribution. Electrospinning of 8% and 15% PCL solutions, respectively, resulted in the production of fibers with diameters below and above 100nm. The combined scaffold exhibited a bimodal distribution of aligned fibers with peaks around 80 and 180nm, thus mimicking the collagen fibrils of healthy ACL tissue. This research is expected to have a society-wide impact because it aims to enhance the health condition and life quality of a wide range of patients.

Enhancing Surface Morphology, Mechanical, and Wettability Properties of PVA/Chitosan/HAp Nanofiber Scaffold through Dielectric Barrier Discharge Plasma Treatment

Abstract Dielectric barrier discharge (DBD) plasma treatment has been widely used for surface functionalization, allowing for precise modification of surface chemistry and morphology. This study investigates the efficacy of DBD plasma treatment in enhancing the surface morphology and wettability of electrospun nanofiber scaffolds composed of polyvinyl alcohol (PVA), chitosan, and hydroxyapatite (HAp), with potential applications in bone tissue engineering. Scanning electron microscopy (SEM) revealed significant alterations in surface morphology after treatment, including a reduction in average fiber diameter and the presence of uneven, damaged, and even broken fibers. Interestingly, the ultimate strength of the nanofibers increased from 1.13 ± 0.05 MPa to 6.99 ± 0.07 MPa despite the decrease in diameter. Contact angle measurements confirmed a remarkable improvement in wettability, with the contact angle decreasing from 39.46° to 7.45° following increasing treatment time. This enhanced wettability suggests improved cell adhesion, potentially leading to more effective bone tissue regeneration.

Biomaterials and emerging technologies for tissue engineering and in vitro models

Biomaterials and emerging technologies for tissue engineering and in vitro models

Recreating 3D cell environments in the lab

Advancements in 3D cell culturing and bioprinting platforms will allow a better understanding of cellular behavior for tissue engineering and biomedical research.

Spectroscopic Characterization of Biomaterials for Tissue Engineering Applications

Spectroscopic Characterization of Biomaterials for Tissue Engineering Applications

Multiphoton Lithography of Interpenetrating Polymer Networks for Tailored Microstructure Thermal and Micromechanical Properties.

Multiphoton lithography (MPL), an emerging truly 3D microfabrication technique, exhibits substantial potential in biomedical applications, including drug delivery and tissue engineering. Fabricated micro-objects are often expected to undergo shape morphing or bending of the entire structure or its parts. Furthermore, ensuring precise property tuning is detrimental to the realization of the functionality of MPL microstructures. Herein, novel MPL materials based on interpenetrating polymer networks (IPNs) are presented that effectively combine the advantages of acrylate and epoxy systems. IPNs with varying component ratios are investigated for their microfabrication performance and structural integrity with respect to thermal and micromechanical properties. A variety of high-resolution techniques is applied to comprehensively evaluate IPN properties at the bulk, micron, and segmental levels. This study shows that the MPL laser scanning velocity and power, photoinitiator content, and multi-step exposure can be used to tune the morphology and properties of the IPN. As a result, a library of 3D MPL IPN microstructures with high 3D structural stability and tailored thermal and micromechanical properties is achieved. New IPN microstructures with Young's moduli of 3-4MPa demonstrate high-to-fully elastic responses to deformations, making them promising for applications in morphable microsystems, soft micro-robotics, and cell engineering.

ATPSpin: A Single Microfluidic Platform that Produces Diversified ATPS-Alginate Microfibers.

Microfluidic spinning is emerging as a useful technique in the fabrication of alginate fibers, enabling applications in drug screening, disease modeling, and disease diagnostics. In this paper, by capitalizing on the benefits of aqueous two-phase systems (ATPS) to produce diverse alginate fiber forms, we introduce an ATPS-Spinning platform (ATPSpin). This ATPS-enabled method efficiently circumvents the rapid clogging challenges inherent to traditional fiber production techniques by regulating the interaction between alginate and cross-linking agents like Ba2+ ions. By varying system parameters under the guidance of a regime map, our system produces several fiber forms─solid, hollow, and droplet-filled─consistently and reproducibly from a single device. We demonstrate that the resulting alginate fibers possess distinct features, including biocompatibility. We also encapsulate HEK293 cells in the microfibers as a proof-of-concept that this versatile microfluidic fiber generation platform may have utility in tissue engineering and regenerative medicine applications.

Microfluidic Assembly of Degradable, Stereocomplexed Hydrogel Microparticles.

Hydrogel microparticles (HMPs) have been investigated widely for their use in tissue engineering and drug delivery applications. However, translation of these highly tunable systems has been hindered by covalent cross-linking methods within microparticles. Stereocomplexation, a stereospecific form of physical cross-linking, provides a robust yet degradable alternative for creating translationally relevant HMPs. Herein, 4-arm polyethylene glycol (PEG) stars were used as macromolecular initiators from which oligomeric poly(l-lactic acid) (PLLA) was polymerized with a degree of polymerization (DPn) of 20 on each arm. Similarly, complementary propargyl-containing ABA cross-linkers with enantiomeric poly(d-lactic acid) (PDLA) segments (DPn = 20) on each arm. Droplets of these gel precursors were formed via a microfluidic organic-in-oil-in-water system where microparticles self-assembled via stereocomplexation and were stabilized after precipitation in deionized water. By varying the flow rate of the dispersed phase, well-defined microparticles with diameters of 33.7 ± 0.5, 62.4 ± 0.6, and 105.7 ± 0.8 μm were fabricated. Gelation due to stereocomplexation was confirmed via wide-angle X-ray scattering in which HMPs exhibited the signature diffraction pattern of stereocomplexed PLA at 2θ = 12.2, 21.2, 24.2°. Differential scanning calorimetry also confirmed stereocomplexation by the appearance of a crystallization exotherm (Tc = 37 °C) and a high-temperature endotherm (Tm = 159 °C) that does not appear in the homocrystallization of PLLA or the hydrogel precursors. Additionally, the propargyl handle present on the cross-linker allows for pre- or post-assembly thiol-yne "click" functionalization as demonstrated by the addition of thiol-containing fluorophores to the HMPs.