Journal of biomedical materials research. Part A最新文献

Cardiovascular diseases are a major global health challenge. Blood vessel disease and dysfunction are major contributors to this healthcare burden, and the development of tissue-engineered vascular grafts (TEVGs) is required, particularly for the replacement of small-diameter vessels. Silk fibroin (SF) is a widely used biomaterial for TEVG fabrication due to its high strength and biocompatibility. However, the stiffness of SF is much higher than that of native blood vessels (NBVs), which limits its application for vascular tissue engineering. In this study, SF was plasticized with glycerol to produce TEVGs exhibiting similar stiffness and ultimate tensile strength to those of NBVs. The electrospun SF/glycerol TEVGs exhibited mechanical properties comparable to NBVs and supported the in vitro proliferation of essential vascular cells—endothelial and smooth muscle cells. After 5 days of culture, the TEVGs exhibited an endothelial monolayer in the lumen, demonstrating their potential for functional vascular tissue regeneration. Our study demonstrates the feasibility of producing TEVGs from SF with tailored mechanical properties, paving the way for more functional and durable TEVGs for future clinical applications.

Cell therapies harnessing the pro-vascular regenerative capacities of mesenchymal stromal cell (MSC) populations, including human adipose-derived stromal cells (hASCs), have generated considerable interest as an emerging treatment strategy for peripheral arterial disease (PAD) and its progression to critical limb ischemia (CLI). There is evidence to support that polysaccharide hydrogels can enhance therapeutic efficacy when applied as minimally-invasive delivery systems to support MSC survival and retention within ischemic tissues. However, there has been limited research to date on the effects of hydrogel composition on the phenotype and function of encapsulated cell populations. Recognizing this knowledge gap, this study compared the pro-angiogenic function of hASCs encapsulated in distinct but similarly-modified natural polysaccharide hydrogels composed of methacrylated glycol chitosan (MGC) and methacrylated hyaluronic acid (MHA). Initial in vitro studies confirmed high viability (>85%) of the hASCs following encapsulation and culture in the MGC and MHA hydrogels over 14 days, with a decrease in the cell density observed over time. Moreover, higher levels of a variety of secreted pro-angiogenic and immunomodulatory factors were detected in conditioned media samples collected from the hASCs encapsulated in the MGC-based hydrogels compared to the MHA hydrogels. Subsequent testing focused on comparing hASC delivery within the MGC and MHA hydrogels to saline controls in a femoral artery ligation-induced CLI (FAL-CLI) model in athymic nu/nu mice over 28 days. For the in vivo studies, the hASCs were engineered to express tdTomato and firefly luciferase to quantitatively compare the efficacy of the two platforms in supporting the localized retention of viable hASCs through longitudinal cell tracking with bioluminescence imaging (BLI). Interestingly, hASC retention was significantly enhanced when the cells were delivered in the MHA hydrogels as compared to the MGC hydrogels or saline. However, laser Doppler perfusion imaging (LDPI) indicated that the restoration of hindlimb perfusion was similar between the treatment groups and controls. These findings were corroborated by endpoint immunofluorescence (IF) staining showing similar levels of CD31⁺ cells in the ligated limbs at 28 days in all groups. Overall, this study demonstrates that enhanced MSC retention may be insufficient to augment vascular regeneration, emphasizing the complexity of designing biomaterials platforms for MSC delivery for therapeutic angiogenesis. In addition, the data points to a potential challenge in approaches that seek to harness the paracrine functionality of MSCs, as strategies that increase the secretion of immunomodulatory factors that can aid in regeneration may also lead to more rapid MSC clearance in vivo.

Regenerated fibrous cellulose possesses a unique set of properties, including biocompatibility, biodegradability, and high surface area potential, but its applications in the biomedical sector have not been sufficiently explored. In this study, nanofibrous cellulose matrices were fabricated via a wet-electrospinning process using a binary system of the solvent ionic liquid (IL) 1-butyl-3-methylimidazolium acetate (BMIMAc) and co-solvent dimethyl sulfoxide (DMSO). The morphology of the matrices was controlled by varying the ratio of BMIMAc versus DMSO in the solvent system. The most effective ratio of 1:1 produced smooth fibers with diameters ranging from 200 to 400 nm. The nanofibrous cellulose matrix showed no cytotoxicity when tested on mouse fibroblast L929 cells whose viability remained above 95%. Human triple-negative breast cancer MDA-MB-231 cells also exhibited high viability even after 7 days of seeding and were able to penetrate deeper layers of the matrix, indicating high biocompatibility. These properties of nanofibrous cellulose demonstrate its potential for tissue engineering and cell culture applications.

The increasing importance of regenerative medicine has resulted in a growing need for advanced tissue replacement materials in head and neck surgery. Allo- and xenogenic graft processing is often time-consuming and can deteriorate the extracellular matrix (ECM). High hydrostatic pressure (HHP)-treatment could allow specific devitalization while retaining the essential properties of the ECM. Porcine connective tissue and cartilage were HHP-treated at 100–400 MPa for 10 min. Structural modifications following HHP-exposure were examined using electron microscopy, while devitalization was assessed through metabolism and cell death analyses. Furthermore, ECM alterations and decellularization were evaluated by histology, biomechanical testing, and DNA content analysis. Additionally, the inflammatory potential of HHP-treated tissue was evaluated in vivo using a dorsal skinfold chamber in a mouse model. The devitalization effects of HHP were dose-dependent, with a threshold identified at 200 MPa for fibroblasts and chondrocytes. At this pressure level, HHP induced structural alterations in cells, with a shift toward late-stage apoptosis. HHP-treatment preserved ECM structure and biomechanical properties, but did not remove cell debris from the tissue. This study observed a pressure-dependent increase of markers suggesting the occurrence of immunogenic cell death. In vivo investigations revealed an absence of inflammatory responses to HHP-treated tissue, indicating a favorable biological response to HHP. In conclusion, application of HHP devitalizes fibroblasts and chondrocytes at 200 MPa while retaining the essential properties of the ECM. Prospectively, HHP may simplify the preparation of allo- and xenogenic tissue replacement materials and increase the availability of grafts in head and neck surgery.

The electrospinning technique is a commonly employed approach to fabricate fibers intended for various tissue engineering applications. The aim of this study is to develop a novel strategy for tendon repair through the use of aligned poly(ε-caprolactone) (PCL) and poly(glycerol sebacate) (PGS) fibers fabricated in benign solvents, and further explore the potential application of PGS in tendon tissue engineering (TTE). The fibers were characterized for their morphological and physicochemical properties; amniotic epithelial stem cells (AECs) were used to assess the fibers teno-inductive and immunomodulatory potential due to their ability to teno-differentiate undergoing first a stepwise epithelial to mesenchymal transition, and due to their documented therapeutic role in tendon regeneration. The addition of PGS to PCL improved the spinnability of the polymer solution, as well as the uniformity and directionality of the so-obtained fibers. The mechanical properties were in the range of most TTE applications, specifically in the case of PCL/PGS 4:1 and 2:1 ratios. Compared to PCL alone, the same ratios also allowed a better AECs infiltration and growth over 7 days of culture, and triggered the activation of tendon-related genes (SCX, COL1, TNMD) and the expression of tenomodulin (TNMD) at the protein level. Concerning the immunomodulatory properties, both PCL and PCL/PGS fibers negatively affected the immunomodulatory profile of AECs, up-regulating both anti-inflammatory (IL-10) and pro-inflammatory (IL-12) cytokines over 7 days of culture. Overall, PCL/PGS 2:1 fibers fabricated with benign solvents proved to be the most suitable composition for TTE application based on their topographical cues, mechanical properties, biocompatibility, and teno-inductive properties.

Macrophages are involved in several critical activities associated with tissue repair and regeneration. Current approaches in regenerative medicine are focusing on leveraging the innate immune response to accelerate tissue regeneration and improve long-term healing outcomes. Of particular interest in this regard are the currently known, four main M2 macrophage subtypes: M2^{interleukin (IL)-4,IL-13}, M2^IC, M2^IL-10, M2^{non-selective adenosine receptor agonists (NECA)} (M2^IL-4,IL-13 → M2^NECA). In this study, rat bone marrow-derived macrophages (M₀) were polarized to each of the four subtypes M2^IL-4,IL-13 → M2^NECA and cultured for 72 h in vitro. Luminex assay results highlighted increased production of tissue inhibitor of metalloproteinases-1 (TIMP-1) for M2^IL-4,IL-13, higher amounts of transforming growth factor-beta 1 (TGF-β1) for M2^IL-10, and elevated vascular endothelial growth factor A (VEGF-A) from M2^NECA. Co-culture experiments performed with M2^IL-10 macrophages and L929 fibroblasts highlighted the increased production of soluble collagen within the media as well as higher amounts of collagen in the extracellular matrix. Human umbilical vein endothelial cells (HUVECs) were co-cultured with M2^NECA macrophages, which demonstrated an increase in intercellular adhesion molecule (ICAM) and platelet endothelial cell adhesion molecule (PECAM), as well as increased formation of endothelial tubes. The findings of this study emphasize a critical demand for further characterization and analyses of distinct M₂ subtypes and careful selection of specific macrophage populations for regeneration of specific tissue types. The current, broad classification of “M₂” may be sufficient in many general tissue engineering applications, but, as conditions are constantly in flux within the microenvironment in vivo, a higher degree of specificity and control over the initial M₂ subtype could result in more consistent long-term outcomes where macrophages are utilized as part of an overall regenerative strategy.

Commercially pure titanium (c.p. Ti) and Ti6Al4V alloys are the most widely used metallic biomaterials in the biomedical sector. However, their high rigidity and the controversial toxicity of their alloying elements often compromise their clinical success. The use of porous β-Titanium alloys is proposed as a solution to these issues. In this regard, it is necessary to implement economic, repetitive, and non-destructive measurement techniques that allow for the semi-quantitative evaluation of the chemical nature of the implant, its microstructural characteristics, and/or surface changes. This study proposes the use of simple measurement protocols based on electrical impedance measurements, correlating them with the porosity inherent to processing conditions (pressure and temperature), as well as the chemical composition of the implant. Results revealed a clear direct relationship between porosity and electrical impedance. The percentage and/or size of the porosity decrease with an increase in compaction pressure and temperature. Moreover, there is a notable influence of the frequency used in the measurements obtained. Additionally, the sensitivity of this measurement technique has enabled the evaluation of differences in chemical composition and the detection of intermetallics in the implants. For the first time in the literature, this research establishes relationships between stiffness and electrical impedance, using approximations and models for the observed trends. All the results obtained corroborate the appropriateness of the technique to achieve the real-time characterization of Titanium implants, in an efficient and non-invasive way.

To enhance the treatment of patients' urethral defects, such as strictures and hypospadias, we investigated the potential of using artificial urethral tissue. Our study aimed to generate this tissue and assess its effectiveness in a rabbit model. Two types of bioprinted grafts, based on methacrylated gelatin-silk fibroin (GelMA-SF) hydrogels, were produced: acellular, as well as loaded with autologous rabbit stem cells. Rabbit adipose stem cells (RASC) were differentiated toward smooth muscle in the GelMA-SF hydrogel, while rabbit buccal mucosa stem cells (RBMC), differentiated toward the epithelium, were seeded on its surface, forming two layers of the cell-laden tissue. The constructs were then reinforced with polycaprolactone-polylactic acid meshes to create implantable multilayered artificial urethral grafts. In vivo experiments showed that the cell-laden tissue integrated into the urethra with less fibrosis and inflammation compared to its acellular counterpart. Staining to trace the implanted cells confirmed integration into the host organism 3 months postsurgery.

The periodontal tissue comprises alveolar bone, cementum, and periodontal ligament (PDL), forming a highly hierarchical architecture. Although current therapies could regenerate the hard tissue well, the simultaneous reconstruction of hard and soft tissue remains a great clinical challenge with the major difficulty in highly orientated PDL regeneration. Using the unidirectional freeze-casting method and biomimetic mineralization technique, we construct a hierarchical bilayer scaffold with the aligned chitosan scaffold with ZIF-8 resembling PDL, and intrafibrillarly mineralized collagen resembling alveolar bone. The hierarchical bilayer scaffold exhibits different geomorphic clues and chemical microenvironments to realize a perfect simulation of the natural periodontal hierarchical architecture. The aligned scaffold with ZIF-8 could induce the fibrogenic differentiation of bone mesenchymal stromal cells (BMSCs), and the mineralized scaffold could induce osteogenic differentiation of BMSCs. The hierarchical bilayer scaffold could simulate periodontal complex tissue, exhibiting great promise for synchronized multi-tissue regeneration of periodontal tissue.

The avascular structure and low cell migration to the damaged area due to the low number of cells do not allow spontaneous repair of the articular cartilage tissue. Therefore, functional scaffolds obtained from biomaterials are used for the regeneration of cartilage tissue. Here, we functionalized one of the self-assembling peptide (SAP) scaffolds KLD (KLDLKLDLKLDL) with short bioactive motifs, which are the α1 chain of type II collagen binding peptide WYRGRL (C1) and the triple helical collagen mimetic peptide GFOGER (C2) by direct coupling. Our goal was to develop injectable functional SAP hydrogels with proper mechanical characteristics that would improve chondrogenesis. Scanning electron microscopy (SEM) was used to observe the integration of peptide scaffold structure at the molecular level. To assure the stability of SAPs, the rheological characteristics and degradation profile of SAP hydrogels were assessed. The biochemical study of the DNA, glycosaminoglycan (GAG), and collagen content revealed that the developed bioactive SAP hydrogels greatly increased hMSCs proliferation compared with KLD scaffolds. Moreover, the addition of bioactive peptides to KLD dramatically increased the expression levels of important chondrogenic markers such as aggrecan, SOX-9, and collagen Type II as evaluated by real-time polymerase chain reaction (PCR). We showed that hMSC proliferation and chondrogenic differentiation were encouraged by the developed SAP scaffolds. Although the chondrogenic potentials of WYRGRL and GFOGER were previously investigated, no study compares the effect of the two peptides integrated into 3-D SAP hydrogels in chondrogenic differentiation. Our findings imply that these specifically created bioactive peptide scaffolds might help enhance cartilage tissue regeneration.