Native Extracellular Matrix Research Articles

Engineering biomimetic scaffolds by combining silk protein nanofibrils and hyaluronic acid

Nanofibrous scaffolds mimicking important features of the native extracellular matrix (ECM) provide a promising strategy for tissue regeneration. However, 3D scaffolds mimicking natural protein nanofibers and bioactive glycosaminoglycans remain poorly developed. In this study, a biomimetic nanofibrous scaffold composed of natural silk protein nanofibers and glycosaminoglycan hyaluronic acid (HA) was developed. HA functionalization significantly improved the hydrophilicity and bioactivity of silk nanofibers (SNFs). SNFs can be assembled into nanofibrous aerogel scaffolds with low density and desirable shapes on a large scale. More importantly, with the assistance of HA, the silk nanofibrous aerogel scaffolds with ultra-high porosity, natural bioactivity, and structural stability in aqueous environment can be fabricated. In the protease/hyaluronidase solution, the SNF scaffolds with 10.0 % HA can maintain their monolithic shape for >3 weeks. The silk nanofibrous scaffolds not only imitate the composition of ECM but also mimic the hierarchical structure of ECM, providing a favorable microenvironment for cell adhesion and proliferation. These results indicate that this structurally and functionally biomimetic system is a promising tissue engineering scaffold.

Does tissue fixation change the mechanical properties of dry ovine bone extracellular matrix?

Tissue fixation is a prevalent method for bone conservation. Bone biopsies are typically fixed in formalin, dehydrated in ethanol, and infiltrated with polymethyl methacrylate (PMMA) Since some experiments can only be performed on fixed bone samples, it is essential to understand how fixation affects the measured material properties. The aim of this study was to quantify the influence of tissue fixation on the mechanical properties of cortical ovine bone at the extracellular matrix (ECM) level with state-of-the-art micromechanical techniques.A small section from the middle of the diaphysis of two ovine tibias (3.5 and 5.5 years old) was cut in the middle and polished on each side, resulting in a pair of mirrored surfaces. For each pair, one specimen underwent a fixation protocol involving immersion in formalin, dehydration with ethanol, and infiltration with PMMA. The other specimen (mirrored) was air-dried. Six osteons were selected in both pairs, which could be identified in both specimens. The influence of fixation on the mechanical properties was first analyzed using micropillar compression tests and nanoindentation in dry condition. Additionally, changes in the degree of mineralization were evaluated with Raman spectroscopy in both fixed and native bone ECM. Finally, micro tensile experiments were conducted in the 3.5-year fixed ovine bone ECM and compared to reported properties of unfixed dry ovine bone ECM.Interestingly, we found that tissue fixation does not alter the mechanical properties of ovine cortical bone ECM compared to experiments in dry state. However, animal age increases the degree of mineralization (p = 0.0159) and compressive yield stress (p = 0.041). Tissue fixation appears therefore as a valid conservation technique for investigating the mechanical properties of dehydrated bone ECM.

Development and Characterization of Decellularized Lung Extracellular Matrix Hydrogels.

The use of extracellular matrix (ECM)-derived hydrogels in tissue engineering has become increasingly popular, as they can mimic cells' natural environment in vitro. However, maintaining the native biochemical content of the ECM, achieving mechanical stability, and comprehending the impact of the decellularization process on the mechanical properties of the ECM hydrogels are challenging. Here, a pipeline for decellularization of bovine lung tissue using two different protocols, downstream characterization of the effectiveness of decellularization, fabrication of reconstituted decellularized lung ECM hydrogels and assessment of their mechanical and cytocompatibility properties were described. Decellularization of the bovine lung was pursued using a physical (freeze-thaw cycles) or chemical (detergent-based) method. Hematoxylin and Eosin staining was performed to validate the decellularization and retention of major ECM components. For the evaluation of residual collagen and sulfated glycosaminoglycan (sGAG) content within the decellularized samples, Sirius red and Alcian blue staining techniques were employed, respectively. Mechanical properties of the decellularized lung ECM hydrogels were characterized by oscillatory rheology. The results suggest that decellularized bovine lung hydrogels can provide a reliable organotypicalternative to commercial ECM products by retaining most native ECM components. Furthermore, these findings reveal that the decellularization method of choice significantly affects gelation kinetics as well as the stiffness and viscoelastic properties of resulting hydrogels.

Hyaluronic Acid Hydrogels with Phototunable Supramolecular Cross-Linking for Spatially Controlled Lymphatic Tube Formation.

The dynamics of the extracellular matrix (ECM) influences stem cell differentiation and morphogenesis into complex lymphatic networks. While dynamic hydrogels with stress relaxation properties have been developed, many require detailed chemical processing to tune viscoelasticity, offering a limited opportunity for in situ and spatiotemporal control. Here, a hyaluronic acid (HA) hydrogel is reported with viscoelasticity that is controlled and spatially tunable using UV light to direct the extent of supramolecular and covalent cross-linking interactions. This is achieved using UV-mediated photodimerization of a supramolecular ternary complex of pendant trans-Brooker's Merocyanine (BM) guests and a cucurbit[8]uril (CB[8]) macrocycle. The UV-mediated conversion of this supramolecular complex to its covalent photodimerized form is catalyzed by CB[8], offering a user-directed route to spatially control hydrogel dynamics in combination with orthogonal photopatterning by UV irradiation through photomasks. This material thus achieves spatial heterogeneity of substrate dynamics, recreating features of native ECM without the need for additional chemical reagents. Moreover, these dynamic hydrogels afford spatial control of substrate mechanics to direct human lymphatic endothelial cells (LECs) to form lymphatic cord-like structures (CLS). Specifically, cells cultured on viscoelastic supramolecular hydrogels have enhanced formation of CLS, arising from increased expression of key lymphatic markers, such as LYVE-1, Podoplanin, and Prox1, compared to static elastic hydrogels prepared from fully covalent cross-linking. Viscoelastic hydrogels promote lymphatic CLS formation through the expression of Nrp2, VEGFR2, and VEGFR3 to enhance the VEGF-C stimulation. Overall, viscoelastic supramolecular hydrogels offer a facile route to spatially control lymphatic CLS formation, providing a tool for future studies of basic lymphatic biology and tissue engineering applications.

Recent advances in the development of bioartificial pancreas using 3D bioprinting for the treatment of type 1 diabetes: a review

Type 1 diabetes is a chronic condition that results from the destruction of insulin-producing β-cells in the pancreas. Current treatments for type 1 diabetes, such as insulin therapy and pancreatic islet transplantation, have several limitations and, hence not quite effective in the long run. As current therapy methods fail to slow disease development, novel strategies such as the development of a bioartificial pancreas are being seriously considered. Over the last decade, research has focused on tissue engineering, which aids in the design of biological alternatives for the repair and replacement of non-functional or damaged organs. Three dimensional (3D) bioprinting technology which employs 3D printing technology to generate 3D tissue-like structures from biomaterials and cells, offers a promising solution for the treatment of type 1 diabetes by providing the ability to generate functional endocrine pancreatic tissue. Bioprinted structures are therefore an important aspect of tissue engineering because they have been found to replicate the native extracellular matrix, promoting cell survival and proliferation. In this review, recent developments in 3D bioprinting of endocrine pancreas for the treatment of type 1 diabetes particularly focussing on the choice of cells, biomaterials, growth factors, and essential considerations have been discussed in detail. Additionally, the key challenges and perspectives towards recapitulation of the pancreatic function of the pancreatic organ engineering technologies have also been discussed.

Decellularized extracellular matrix: A promising strategy for skin repair and regeneration

The skin is an important organ of the human body that resists external threats but lacks sufficient self-regeneration ability when severe damage occurs. However, most of the available skin substitutes cannot achieve ideal restoration of complex structures and multiple functions of native skin tissues. Fortunately, the advent of decellularized extracellular matrix (dECM) offers a promising approach to overcome these obstacles. The dECM, derived from the natural extracellular matrix (ECM), possesses a similar structure and composition, which constructs an environment favorable for cell performance in regeneration. Moreover, dECM retains good bioactivity, low immunogenicity, and high availability, making it a suitable biomaterial for skin repair and regeneration. In this review, various decellularization methods and subsequent evaluations of dECM are introduced first, and the main sources of dECM are then presented. Furthermore, the recent progress of dECM-based biomaterials applied in skin regeneration and future perspectives are summarized.

Remaining microtia tissue as a source for 3D bioprinted elastic cartilage tissue constructs, potential use for surgical microtia reconstruction.

The absence of ears in children is a global problem. An implant made of costal cartilage is the standard procedure for ear reconstruction; however, side effects such as pneumothorax, loss of thoracic cage shape, and respiratory complications have been documented. Three-dimensional (3D) printing allows the generation of biocompatible scaffolds that mimic the shape, mechanical strength, and architecture of the native extracellular matrix necessary to promote new elastic cartilage formation. We report the potential use of a 3D-bioprinted poly-ε-caprolactone (3D-PCL) auricle-shaped framework seeded with remaining human microtia chondrocytes for the development of elastic cartilage for autologous microtia ear reconstruction. An in vivo assay of the neo-tissue formed revealed the generation of a 3D pinna-shaped neo-tissue, and confirmed the formation of elastic cartilage by the presence of type II collagen and elastin with histological features and a protein composition consistent with normal elastic cartilage. According to our results, a combination of 3D-PCL auricle frameworks and autologous microtia remnant tissue generates a suitable pinna structure for autologous ear reconstruction.

Self-organizing behaviors of cardiovascular cells on synthetic nanofiber scaffolds.

In tissues and organs, the extracellular matrix (ECM) helps maintain inter- and intracellular architectures that sustain the structure-function relationships defining physiological homeostasis. Combining fiber scaffolds and cells to form engineered tissues is a means of replicating these relationships. Engineered tissues' fiber scaffolds are designed to mimic the topology and chemical composition of the ECM network. Here, we asked how cells found in the heart compare in their propensity to align their cytoskeleton and self-organize in response to topological cues in fibrous scaffolds. We studied cardiomyocytes, valvular interstitial cells, and vascular endothelial cells as they adapted their inter- and intracellular architectures to the extracellular space. We used focused rotary jet spinning to manufacture aligned fibrous scaffolds to mimic the length scale and three-dimensional (3D) nature of the native ECM in the muscular, valvular, and vascular tissues of the heart. The representative cardiovascular cell types were seeded onto fiber scaffolds and infiltrated the fibrous network. We measured different cell types' propensity for cytoskeletal alignment in response to fiber scaffolds with differing levels of anisotropy. The results indicated that valvular interstitial cells on moderately anisotropic substrates have a higher propensity for cytoskeletal alignment than cardiomyocytes and vascular endothelial cells. However, all cell types displayed similar levels of alignment on more extreme (isotropic and highly anisotropic) fiber scaffold organizations. These data suggest that in the hierarchy of signals that dictate the spatiotemporal organization of a tissue, geometric cues within the ECM and cellular networks may homogenize behaviors across cell populations and demographics.

Perfusability and immunogenicity of implantable pre-vascularized tissues recapitulating features of native capillary network.

Vascularization is a key pre-requisite to engineered anatomical scale three dimensional (3-D) constructs to ensure their nutrient and oxygen supply upon implantation. Presently, engineered pre-vascularized 3-D tissues are limited to only micro-scale hydrogels, which meet neither the anatomical scale needs nor the complexity of natural extracellular matrix (ECM) environments. Anatomical scale perfusable constructs are critically needed for translational applications. To overcome this challenge, we previously developed pre-vascularized ECM sheets with long and oriented dense microvascular networks. The present study further evaluated the patency, perfusability and innate immune response toward these pre-vascularized constructs. Macrophage-co-cultured pre-vascularized constructs were evaluated in vitro to confirm micro-vessel patency and perturbations in macrophage metabolism. Subcutaneously implanted pre-vascularized constructs remained viable and formed a functional anastomosis with host vasculature within 3 days of implantation. This completely biological pre-vascularized construct holds great potential as a building block to engineer perfusable anatomical scale tissues.

Human Adipose Tissue Lysate-Based Hydrogel for Lasting Immunomodulation to Effectively Improve Spinal Cord Injury Repair.

The long-term inflammatory microenvironment is one of the main obstacles to inhibit acute spinal cord injury (SCI) repair. The natural adipose tissue-derived extracellular matrix hydrogel shows effective anti-inflammatory regulation because of its unique protein components. However, the rapid degradation rate and removal of functional proteins during the decellularization process impair the lasting anti-inflammation function of the adipose tissue-derived hydrogel. To address this problem, adipose tissue lysate provides an effective way for SCI repair due to its abundance of anti-inflammatory and nerve regeneration-related proteins. Thereby, human adipose tissue lysate-based hydrogel (HATLH) with an appropriate degradation rate is developed, which aims to in situ long-term recruit and induce anti-inflammatory M2 macrophages through sustainedly released proteins. HATLH can recruit and polarize M2 macrophages while inhibiting pro-inflammatory M1 macrophages regardless of human or mouse-originated. The axonal growth of neuronal cells also can be effectively improved by HATLH and HATLH-induced M2 macrophages. In vivo experiments reveal that HATLH promotes endogenous M2 macrophages infiltration in large numbers (3.5 × 105/100µL hydrogel) and maintains a long duration for over a month. In a mouse SCI model, HATLH significantly inhibits local inflammatory response, improves neuron and oligodendrocyte differentiation, enhances axonal growth and remyelination, as well as accelerates neurological function restoration.

Phosphoproteomic Changes Induced by Cell-Derived Matrix and Their Effect on Tumor Cell Migration and Cytoskeleton Remodeling.

Increased fibrotic extracellular matrix (ECM) deposition promotes tumor invasion, which is the first step of the metastatic cascade. Yet, the underlying mechanisms are poorly understood as conventional studies of tumor cell migration are often performed in 2D cultures lacking the compositional and structural complexity of native ECM. Moreover, these studies frequently focus on select candidate pathways potentially overlooking other relevant changes in cell signaling. Here, we combine a cell-derived matrix (CDM) model with phosphotyrosine phosphoproteomic analysis to investigate tumor cell migration on fibrotic ECM relative to standard tissue culture plastic (TCP). Our results suggest that tumor cells cultured on CDMs migrate faster and in a more directional manner than their counterparts on TCP. These changes in migration correlate with decreased cell spreading and increased cell elongation. While the formation of phosphorylated focal adhesion kinase (pFAK)+ adhesion complexes did not vary between TCP and CDMs, time-dependent phosphoproteomic analysis identified that the SRC family kinase LYN may be differentially regulated. Pharmacological inhibition of LYN decreased tumor cell migration and cytoskeletal rearrangement on CDMs and also on TCP, suggesting that LYN regulates tumor cell migration on CDMs in combination with other mechanisms. These data highlight how the combination of physicochemically complex in vitro systems with phosphoproteomics can help identify signaling mechanisms by which the fibrotic ECM regulates tumor cell migration.

Biomimetic Conductive Hydrogel Scaffolds with Anisotropy and Electrical Stimulation for In Vivo Skeletal Muscle Reconstruction.

The nature of the hydrogel scaffold mimicking extracellular matrix plays a crucial role in tissue engineering like skeletal muscle repair. Herein, an anisotropic and conductive hydrogel scaffold is fabricated using gelatin methacryloyl (GelMA) as the matrix hydrogel and silver nanowire (AgNW) as the conductive dopant, through a directional freezing technique for muscle defect repair. The scaffold has an anisotropic structure composed of a directional longitudinal section and a honeycomb cross-section, with high mechanical strength of 10.5kPa and excellent conductivity of 0.26 S m-1 . These properties are similar to native muscle extracellular matrix (ECM) and allow for cell orientation under the guidance of contact cues and electrical stimulation synergistically. In vitro experiments show that the scaffold's oriented structure combined with electrical stimulation results in enhanced myotube formation, with a length of up to 863µm and an orientation rate of 81%. Furthermore, the electrically stimulated scaffold displays a promoted muscle reconstruction ability when transplanted into rats with muscle defects, achieving a muscle mass and strength restoration ratio of 95% and 99%, respectively, compared to normal levels. These findings suggest that the scaffold has great potential in muscle repair applications.

Listen to Your Gut: Key Concepts for Bioengineering Advanced Models of the Intestine.

The intestine performs functions central to human health by breaking down food and absorbing nutrients while maintaining a selective barrier against the intestinal microbiome. Key to this barrier function are the combined efforts of lumen-lining specialized intestinal epithelial cells, and the supportive underlying immune cell-rich stromal tissue. The discovery that the intestinal epithelium can be reproduced in vitro as intestinal organoids introduced a new way to understand intestinal development, homeostasis, and disease. However, organoids reflect the intestinal epithelium in isolation whereas the underlying tissue also contains myriad cell types and impressive chemical and structural complexity. This review dissects the cellular and matrix components of the intestine and discusses strategies to replicate them in vitro using principles drawing from bottom-up biological self-organization and top-down bioengineering. It also covers the cellular, biochemical and biophysical features of the intestinal microenvironment and how these can be replicated in vitro by combining strategies from organoid biology with materials science. Particularly accessible chemistries that mimic the native extracellular matrix are discussed, and bioengineering approaches that aim to overcome limitations in modelling the intestine are critically evaluated. Finally, the review considers how further advances may extend the applications of intestinal models and their suitability for clinical therapies.

Metabolic reprogramming driven by EZH2 inhibition depends on cell–matrix interactions

EZH2 (Enhancer of Zeste Homolog 2), a subunit of Polycomb Repressive Complex 2 (PRC2), catalyzes the trimethylation of histone H3 at lysine 27 (H3K27me3), which represses expression of genes. It also has PRC2-independent functions, including transcriptional coactivation of oncogenes, and is frequently overexpressed in lung cancers. Clinically, EZH2 inhibition can be achieved with the FDA-approved drug EPZ-6438 (tazemetostat). To realize the full potential of EZH2 blockade, it is critical to understand how cell-cell/cell-matrix interactions present in three-dimensional (3D) tissue and cell culture systems influences this blockade in terms of growth-related metabolic functions. Here, we show that EZH2 suppression reduced growth of human lung adenocarcinoma A549 cells in two-dimensional (2D) cultures but stimulated growth in 3D culture. To understand the metabolic underpinnings, we employed [13C6]-glucose Stable Isotope-Resolved Metabolomics (SIRM) to determine the effect of EZH2 suppression on metabolic networks in 2D versus 3D A549 cultures. The Krebs cycle, neoribogenesis, γ-aminobutyrate (GAB) metabolism, and salvage synthesis of purine nucleotides were activated by EZH2 suppression in 3D spheroids but not in 2D cells, consistent with the growth effect. Using simultaneous 2H7-glucose + 13C5,15N2-Gln tracers and EPZ-6438 inhibition of H3 trimethylation, we delineated the effects on the Krebs cycle, γ-aminobutyrate metabolism, gluconeogenesis, and purine salvage to be PRC2 dependent. Furthermore, the growth/metabolic effects differed for mouse Matrigel versus self-produced A549 extracellular matrix. Thus, our findings highlight the importance of the presence and nature of extracellular matrix in studying the function of EZH2 and its inhibitors in cancer cells for modeling the in vivo outcomes.

Engineered 3D tumor microenvironment recapitulating stiffness of lung tissue to explore drug resistance of lung carcinoma

Despite clinical advances, the development of anticancer drugs faces high failure rates owing to cancer drug resistance and insufficient understanding of the native tumor microenvironment. Polymeric hydrogels are promising engineered 3D cancer models owing to their tunable properties and structural similarities to the native extracellular matrix. However, precisely mimicking the native cancer microenvironment remains challenging. In this study, we present interpenetrating polymer network (IPN) hydrogels with independent stiffness control as an engineered 3D lung carcinoma model for drug screening and basic cancer research. These IPN hydrogels are formed via horseradish peroxidase/hydrogen peroxide (H2O2)-mediated dual-crosslinking reactions, showing independently tunable stiffness at various H2O2 concentrations. Using this ability, we create engineered 3D lung cancer models that simulate normal and cancerous lung tissue stiffness with encapsulation of non-small cell lung cancer cells. We perform drug resistance tests using this system and compare the drug responses in various preclinical cancer models in vitro and in vivo. Finally, we investigate the biological mechanism underlying the acquired resistance of lung carcinoma cells to epidermal growth factor receptor inhibitors. In summary, our engineered 3D cancer model can serve as an advanced preclinical platform for drug screening and cancer biology, including the biological mechanisms of acquired drug resistance.

Multifunctional ADM hydrogel containing endothelial cell-exosomes for diabetic wound healing

Non-healing wound, with limited treatment options, remains a prevalent complication of diabetes mellitus. The underlying causes wherein include oxidative stress injury, bacterial infection, cellular dysfunction, and persistent inflammation. Acellular Dermal Matrix (ADM), a wound dressing composed of natural extracellular matrix and abundant bioactive factors, has been successfully developed to treat various wounds, including burns and diabetic ulcers. Protocatechualdehyde (PA) & trivalent iron ion (Fe3+) complex (Fe3+@PA) exhibits potential antioxidant and antibacterial properties. In this study, we developed a dual hydrogel network by combining Fe3+@PA complex-modified ADM with light-cured gelatin (GelMA), supplemented with exosomes derived from human umbilical vein endothelial cells (HUVEC-Exos), to create an ADM composite hydrogel system (ADM-Fe3+@PA-Exos/GelMA) with antioxidant, antibacterial, and cell-promoting functions for diabetic wound treatment. Through in vitro experiments, we investigated the biosafety, antioxidant and antibacterial properties of ADM composite hydrogel. Furthermore, we examined the protective effects of ADM composite hydrogel on diabetic wound. The above experiments collectively demonstrate that our ADM-Fe3+@PA-Exos/GelMA hydrogel promotes diabetic wound healing by eliminating bacterial infection, reduced the reactive oxygen species (ROS) levels, protecting cells against oxidative stress damage, promotingcollagen deposition and angiogenesis, which provides a promising strategy to optimize ADM for diabetic wound treatment.

Introducing a novel ex-vivo calcification model for human aortic valves

Abstract Background Aortic stenosis (AS) is a narrowing of the aortic valve opening due to calcification and thickening of the leaflets. There are currently no effective pharmacological treatments for AS, and surgery or percutaneous interventions are the only therapeutic options. The cellular and molecular mechanisms underlying AS are complex and not fully understood due to the complexity and multi-factorial nature of AS and the absence of suitable test systems. In the current in-vitro models to study valvular calcification, isolated valvular interstitial cells (VICs) are cultured in 2D and in the absence of its native extracellular matrix. In addition, other cell types like valvular endothelial cells (VECs) and macrophages, which have been shown to impact the behavior of VICs, are absent. Purpose Our goal is to establish an ex-vivo calcification model for human aortic valves which maintains the original valve structure and cellular components. Methods Diseased aortic valves were harvested from patients undergoing aortic valve replacement. A 5mm x 7mm piece of this aortic valve tissue was dissected and embedded in agarose, then cut into 300 mm slices using a vibratome (Figure 1). In this way VICs remained in their valvular environment allowing the most native response of VICs to stimuli. The use of multiple slices from a single patient ensured proper internal controls. The slices were placed on a cell culture insert and cultured for 14 days. To induce calcification, the slices were cultured in the presence of 3 mM phosphate (n=23). As a control, valvular slices were cultured in the absence of 3mM phosphate (n=12). To validate if the calcification induced in the ex vivo culture system can be molecularly modulated the glucocorticoid dexamethasone was added to cultures containing 3mM phosphate (n=8). The slices were assessed for the presence of calcification using Alizarin Red staining and for the presence of osteogenic and inflammatory markers using (Immuno) histochemistry. Results In the presence of 3mM Phosphate, 74% of the slices demonstrated calcification, whereas no calcification was observed in all slices cultured without 3mM Phosphate (p<0.0001; Figure 2a). Furthermore, osteogenic genes such as Runx and alkaline phosphatase-1, as well as the inflammation marker NFkb were expressed in the calcified slices (Figure 2a). Dexamethason inhibited the calcification in the slices treated with 3 mM phosphate (Figure 2b). Conclusions We developed a novel ex-vivo calcification model for human aortic valves in which the initiation and progression of aortic valve calcification can be studied. In addition, the calcification can be modulated at a molecular level providing a unique opportunity to uncover the molecular signaling pathways involved in valvular calcification and to test compounds facilitating pre-clinical translational studies.Figure 1Figure 2

Using Decellularized Magnetic Microrobots to Deliver Functional Cells for Cartilage Regeneration.

The use of natural cartilage extracellular matrix (ECM) has gained widespread attention in the field of cartilage tissue engineering. However, current approaches for delivering functional scaffolds for osteoarthritis (OA) therapy rely on knee surgery, which is limited by the narrow and complex structure of the articular cavity and carries the risk of injuring surrounding tissues. This work introduces a novel cell microcarrier, magnetized cartilage ECM-derived scaffolds (M-CEDSs), which are derived from decellularized natural porcine cartilage ECM. Human bone marrow mesenchymal stem cells are selected for their therapeutic potential in OA treatments. Owing to their natural composition, M-CEDSs have a biomechanical environment similar to that of human cartilage and can efficiently load functional cells while maintaining high mobility. The cells are released spontaneously at a target location for at least 20 days. Furthermore, cell-seeded M-CEDSs show better knee joint function recovery than control groups 3 weeks after surgery in preclinical experiments, and ex vivo experiments reveal that M-CEDSs can rapidly aggregate inside tissue samples. This work demonstrates the use of decellularized microrobots for cell delivery and their in vivo therapeutic effects in preclinical tests.

Advanced Hydrogel-Based Strategies for Enhanced Bone and Cartilage Regeneration: A Comprehensive Review.

Bone and cartilage tissue play multiple roles in the organism, including kinematic support, protection of organs, and hematopoiesis. Bone and, above all, cartilaginous tissues present an inherently limited capacity for self-regeneration. The increasing prevalence of disorders affecting these crucial tissues, such as bone fractures, bone metastases, osteoporosis, or osteoarthritis, underscores the urgent imperative to investigate therapeutic strategies capable of effectively addressing the challenges associated with their degeneration and damage. In this context, the emerging field of tissue engineering and regenerative medicine (TERM) has made important contributions through the development of advanced hydrogels. These crosslinked three-dimensional networks can retain substantial amounts of water, thus mimicking the natural extracellular matrix (ECM). Hydrogels exhibit exceptional biocompatibility, customizable mechanical properties, and the ability to encapsulate bioactive molecules and cells. In addition, they can be meticulously tailored to the specific needs of each patient, providing a promising alternative to conventional surgical procedures and reducing the risk of subsequent adverse reactions. However, some issues need to be addressed, such as lack of mechanical strength, inconsistent properties, and low-cell viability. This review describes the structure and regeneration of bone and cartilage tissue. Then, we present an overview of hydrogels, including their classification, synthesis, and biomedical applications. Following this, we review the most relevant and recent advanced hydrogels in TERM for bone and cartilage tissue regeneration.

Effect of collagen denaturation degree on mechanical properties and biological activity of nanofibrous scaffolds.

Further progress in regenerative medicine and bioengineering highly depends on the development of 3D polymeric scaffolds with active biological properties. The most attention is paid to natural extracellular matrix components, primarily collagen. Herein, nonwoven nanofiber materials with various degrees of collagen denaturation and fiber diameters 250-500 nm were produced by electrospinning, stabilized by genipin, and characterized in detail. Collagen denaturation has been confirmed using DSC and FTIR analysis. The comparative study of collagen and gelatin nonwoven materials (NWM) revealed only minor differences in their biocompatibility with skin fibroblasts and keratinocytes in vitro. In long-term subcutaneous implantation study, the inflammation was less evident on collagen than on gelatin NWM. Remarkably, the pronounced calcification was revealed in the collagen NWM only. The results obtained can be useful in terms of improving the electrospinning technology of collagen from aqueous solutions, as well as emphasize the importance of long-term study to ensure proper implementation of the material, taking into account the ability of collagen to provoke calcification.