Tissue Engineering Part A最新文献

Adipose tissue is a highly plastic organ whose remodeling dynamics are central to whole-body metabolic health. Expansion of white adipose tissue occurs through either hyperplasia, which preserves tissue function, or hypertrophy, which causes local hypoxia, inflammation, and pathological extracellular matrix (ECM) accumulation. Under hypertrophic conditions, the ECM stiffens and transitions from a supportive scaffold to a fibrotic barrier that limits expansion and perpetuates metabolic dysfunction. Understanding how mechanical cues regulate adipose tissue remodeling is, therefore, essential for identifying new therapeutic strategies. Two mechanosensitive cell populations, adipose stem cells (ASC) and mature adipocytes, are central to this process. ASC interpret ECM stiffness and compositional changes, which determine lineage outcomes. Soft and flexible matrices favor adipogenesis, whereas stiff matrices drive fibroblast-like activation and matrix deposition. Adipocytes, though differentiated, retain mechanosensitive signaling capabilities that shape their function. Under chronic mechanical stresses, cytoskeletal remodeling pathways lead to changes in gene expression and partial dedifferentiation toward a fibroblast-like phenotype. Reciprocal signaling between ASC and adipocytes amplifies these processes, establishing feedback loops that reinforce either healthy or pathological remodeling. Cell and tissue engineering approaches are essential for dissecting these processes, with hydrogel substrates, 3D scaffolds, compression assays, and atomic force microscopy offering physiologically relevant platforms to model progenitors and adipose tissue cellular mechanics. Emerging tools, including nanotopography and mechanical stimulation devices, have the capacity to further clarify how mechanical signals influence adipose remodeling. By positioning ASC and adipocytes as active regulators of ECM mechanics, we underscore the importance of mechanotransduction pathways in adipose tissue health and point to bioengineering strategies that may help discover ways to restore tissue flexibility and improve metabolic outcomes.

Fish collagen is gaining attention as a sustainable biomaterial for three-dimensional (3D) scaffold fabrication in tissue engineering. In this study, a biomimetic, one-pot crosslinking strategy for native fish skin collagen was developed and compared with a conventional periodate oxidation-Schiff base approach using oxidized maltose. Both approaches increased viscosity and thermal stability while preserving native structural features. The oxidized maltose-crosslinked collagen demonstrated Schiff-base crosslinking, and the one-pot crosslinking method produced a covalent bond that was not a Schiff base. Precrosslinked collagens were processed into microgels and incorporated into calcium alginate to yield a shear-recovering, extrudable ink suitable for 3D extrusion printing. The printed scaffolds maintained structural resilience under physiological conditions, exhibited shear-recovery behavior confirmed by rheological analysis, and supported high cell viability. To enhance biofunctionality, vascular endothelial growth factor was conjugated to the 3D scaffolds, which were subsequently seeded with human bone marrow-derived mesenchymal stem cells. Immunofluorescence staining indicated endothelial lineage differentiation, suggesting that this platform may support the development of vascularized 3D tissue constructs. Impact Statement This study presents a one-pot crosslinking approach to enable three-dimensional extrusion printing of a shear-recovery, precrosslinked fish collagen ink design, eliminating the need for postprinting treatments. Functionalization with vascular endothelial growth factor further enhanced the bioactivity of the printed scaffolds by promoting angiogenic response. Collectively, these findings demonstrate a sustainable and biocompatible strategy that broadens the applicability of fish collagen-based inks for vascularized tissue engineering applications.

Given the number of rotator cuff (RC) repairs performed annually and the high rate of structural failure, there remains a significant clinical need for new approaches to augment the repair by enhancing the rate and quality of the tendon healing processes. Tissue-engineering approaches that combine the use of scaffolds and bioactive molecules represent promising new solutions for RC repair. In this study, we investigated the effect of the incorporation of two innate immune pattern recognition receptor agonists (PRRAs) into surgically implanted hydrogels on healing in vitro using ovine RC tendon tissues and in vivo in a translational rat model of RC injury. To address the impact of these innate immune agonists on shoulder healing, we assessed gait function, surgical site histopathology, and quantification of local immune cell infiltrates. We also treated tendon tissues in vitro to assess the impact on tendon transcriptomic responses. We hypothesized that early stimulation of innate immune responses at the site of tendon injury would improve functional and structural tendon healing. We found that of the three PRRAs evaluated, only polyinosine-polycytidylic acid [Poly(I:C)] improved functional gait quality in the postinjury period. However, PRRA injection exerted minimal effects on tendon histology or the density of immune infiltrates. In vitro transcriptomic analysis of tendon blocks treated with PRAAs provided evidence of activation of interferon pathways by Poly(I:C)-treated tissues, suggesting a role of these innate immune cytokines in the pain reduction response. Thus, we conclude that incorporation of certain PRRAs in hydrogels may improve functional recovery after shoulder tendon repair surgery, but also recognize that the timing and release kinetics of agonists delivered in gels at the surgery site can be further optimized. Impact Statement The immunological cascade of healing rotator cuff tissue is a large determinant of whether the tissue will heal or scar. Immunomodulation through biologics has shown mixed success in clinical applications for rotator cuff repair, perpetuating high retear rates. As such, there is a need to investigate novel, immunologically instructive therapies. Herein, we demonstrate that incorporating Toll-like receptor 3 agonist, polyinosine-polycytidylic acid, into a methylcellulose/hyaluronic acid blend hydrogel can induce functional, but interestingly, not tissue, level changes in a rat model of rotator cuff damage. Indicating initial efficacy for a novel potential immunotherapy for rotator cuff injury.

Cell morphology is not only integral to its function within the body but also plays a critical role in cellular behavior and fate. In tissue engineering, cell-scaffold interactions play a critical role because scaffold physical and biochemical characteristics, such as pore size, fiber alignment, and surface architecture, directly influence cellular morphology and behavior. These interactions impact key biological processes, including adhesion, proliferation, migration, and differentiation of the cells, ultimately influencing tissue formation and regeneration. This study investigated how scaffold topography and culture time influence fibroblast morphology and behavior in a bilayer scaffold consisting of randomly oriented fiber layer and aligned fiber layer. Fibroblasts were seeded onto the scaffolds and cultured for 1, 3, 6, or 9 days, and nuclear and cytoskeletal morphologies were quantified using shape descriptors, including nuclear and cellular roundness, eccentricity, aspect ratio, and area ratio. The results demonstrate that scaffold fiber alignment significantly modulates cellular morphology, with aligned fibers promoting elongated, aligned morphologies and randomly oriented fibers favoring branched, multidirectional spreading. Culture time emerged as a key factor, as cells on both surfaces exhibited more rounded, stabilized morphologies by day 6, suggesting time-dependent remodeling and interaction with the scaffold microarchitecture. Specifically, aligned fiber-like scaffold surfaces may benefit regeneration of uniaxially aligned tissues, such as tendon, ligament, or nerve, whereas random fiber-like scaffold surfaces may support stromal or bone environments requiring isotropic spreading. Furthermore, the bilayer scaffold architecture holds promise for complex tissue interfaces, such as the periodontium or osteochondral units, where region-specific topographical cues are essential for functional tissue integration.

Traumatic injuries lead to volumetric muscle loss (VML) and nerve damage that cause chronic functional deficits. Due to the inability of mammalian skeletal muscle to regenerate after VML damage, engineered scaffolds have been explored to address this challenge, but with limited success in functional restoration. We introduce novel bioactive amorphous silicon oxynitride (SiONx) biomaterials with surface properties and Si ion release to accelerate muscle and nerve cell differentiation for functional tissue regeneration. Micropatterned scaffolds were designed and developed on Si-wafer to test the effect of SiONx on myogenesis and neurogenesis. The scaffolds were created using UV photolithography to first pattern their surface, followed by the deposition of SiONx through plasma enhanced chemical vapor deposition (PECVD). X-ray diffraction (XRD) and energy dispersive spectroscopy (EDS) confirmed the uniform chemical structure of an amorphous SiONx film on the patterned surfaces. Atomic force microscopy and scanning electron microscopy (SEM) elucidated the surface morphology with a uniform 2 μm grating microstructure. The 2 µm pattern size is within the range of cellular dimensions, allowing for effective cell-surface interactions. Further, 2 µm features provide sufficient contact points for cell adhesion without overwhelming the cell's ability to interact with the surface. Two separate studies were conducted with SiONx biomaterials and Si ions alone. This was done to understand how Si ions impact cell response separate from the surfaces. C2C12 mouse myoblasts and NG108 neuronal cells were cultured on SiONx biomaterials. In separate studies, we tested the effect of Si ion treatments with these cells (cultured on tissue culture plastic). Cell culture studies demonstrated enhanced C2C12 myoblast attachment and proliferation on SiONx surfaces. High-resolution SEM and fluorescence images revealed highly aligned myotubes (from C2C12 cells) and axons (from NG108 cells) in a parallel direction to the micropatterned SiONx scaffolds. GAP43 expression, neurite outgrowth, and alignment were significantly increased with the Si-ions and SiONx biomaterials. These findings suggest that SiONx scaffolds enhance muscle and nerve cell adhesion and growth and promote the formation of aligned myotubes and axons on the pattern surfaces.

Primary open-angle glaucoma is a prevalent type of degenerative eye disease that results in lifelong blindness, and its critical pathogenic cause is trabecular meshwork (TM) dysfunction or decreased TM cellularity. Considering that TM develops from neural crest cells (NCCs), we investigate the potential of human embryonic stem cell (hESC)-derived NCCs transplantation for TM regeneration. We used a chemically defined method to induce the differentiation of NCCs and injected 1.0 × 10⁶ hESC-derived NCCs combined with 100 μmol/L Y-27632 into the anterior chamber of rabbit. Intraocular pressure (IOP), TM, and corneal changes of rabbits with cell transplantation were examined with TonoPEN AVIA, slit lamp microscope, dual-immunofluorescence staining, and optical coherence tomography. The hESC-derived NCCs underwent homogenous differentiation over the course of 5 days' induction, which expressed the typical neural crest markers HNK-1, P75, SOX10, and AP-2α. NOD/SCID mice received injections of hESC-derived NCCs in the groin or axilla. There was no teratoma formation. When intracamerally injected, hESC-derived NCCs integrated into the TM tissue and expressed mature TM cell markers Aqp1, Chi3l1, and Timp3 after 7 days transplantation in rabbit eyes. The IOP and central corneal thickness basically maintained at normal levels within 2 weeks. No significant adverse effects in rabbits with hESC-derived NCC injection were observed after 5 weeks of cell transplantation. Our findings indicate that hESC-derived NCCs could integrate into the TM tissue and differentiate into mature TM cells after being injected intracamerally, showing a potential therapeutic approach to addressing TM dysfunction in the treatment of glaucoma.

Insufficient vascularization is the main barrier to creating engineered bone grafts for treating large and ischemic defects. Modular tissue engineering approaches have promise in this application because of the ability to combine tissue types and localize microenvironmental cues to drive desired cell function. In direct bone formation approaches, it is challenging to maintain sustained osteogenic activity, since vasculogenic cues can inhibit tissue mineralization. This study harnessed the physiological process of endochondral ossification to create multiphase tissues that allowed concomitant mineralization and vessel formation. Mesenchymal stromal cells in pellet culture were differentiated toward a cartilage phenotype, followed by induction to chondrocyte hypertrophy. Hypertrophic pellets (HPs) exhibited increased alkaline phosphatase activity, calcium deposition, and osteogenic gene expression relative to chondrogenic pellets. In addition, HPs secreted and sequestered angiogenic factors, and supported new blood vessel formation by cocultured endothelial cells and undifferentiated stromal cells. Multiphase constructs created by combining HPs and vascularizing microtissues and maintained in an unsupplemented basal culture medium were shown to support robust vascularization and sustained tissue mineralization. These results demonstrate a promising in vitro strategy to produce multiphase-engineered constructs that concomitantly support the generation of mineralized and vascularized tissue in the absence of exogenous osteogenic or vasculogenic medium supplements.

Skin aging involves changes in extracellular matrix components, such as wrinkles and pigmentation. Caviar extract (CE) is a promising compound for skin rejuvenation, but effective topical delivery requires optimized carriers. This study evaluated polyvinyl alcohol/carboxymethyl chitosan (PVA/CMC) hydrogels loaded with CE at concentrations of 2%, 3.5%, and 5% as scaffolds to influence the epithelial differentiation of adipose-derived mesenchymal stem cells (ADMSCs). Hydrogels were synthesized using a freeze-thaw method and characterized by Fourier-transform infrared spectroscopy (FTIR), scanning electron microscopy, swelling and degradation tests, and mechanical analysis. Biocompatibility and cell migration were assessed using MTT and scratch assays; at the same time, expression of cytokeratin-18 (CK-18) and pan-cytokeratin (pan-CK) was measured via reverse transcription-quantitative polymerase chain reaction and immunocytochemistry (ICC), respectively. FTIR confirmed successful CE incorporation, and SEM revealed a porous structure. Hydrogels with 3.5% and 5% CE demonstrated a good balance between swelling and degradation over 336 h. The biocompatibility tests showed that 5% CE supported enhanced long-term cell growth. The scratch assay indicated improved cell migration, and transcriptional analysis revealed significantly higher CK-18 levels in ADMSCs treated with PVA/CMC/CE 5% (p < 0.001). ICC results showed significantly higher pan-CK expression at 3.5% CE (41.82%) and 5% CE (48.16%), suggesting that CE promotes repair processes. These findings suggest that 5% CE-loaded PVA/CMC hydrogel could be an effective option for skin regeneration and antiaging. Impact Statement Caviar extract (CE) was considered a bioactive ingredient, along with polyvinyl alcohol (PVA) and carboxymethyl chitosan (CMC) polymers, to prepare a functional and practical hydrogel without hazardous components for anti-aging and cosmetic applications. In the present study, the PVA/CMC hydrogel contains various concentrations of CE (3.5% and 5%), is biocompatible, and enhances cellular viability and migration of adipose-derived mesenchymal stem cell. Our results demonstrated that the synergistic effect of CE and CMC could promote the expression of cytokeratin-18 gene and pan-cytokeratin protein and play a critical role in stimulating skin regeneration.

Biological materials composed of extracellular matrix (ECM) or its components have been successfully used for tissue repair and reconstruction. Preclinical studies, along with a cohort study following stage T1A esophageal adenocarcinoma (EAC) resection, have shown that ECM biomaterials can restore esophageal mucosa and submucosa without cancer recurrence. However, the molecular mechanisms underlying these effects remain largely unexplored. The present study investigates the in vitro effects of ECM degradation products from nonmalignant esophageal (eECM) and urinary bladder (ubECM) sources on EAC cell proliferation, migration, and associated signaling pathways. Both eECM and ubECM significantly inhibited OE33 cell proliferation, with eECM exhibiting a stronger effect-reducing proliferation to 25% at 24 h and 7% at 72 h compared with pepsin control (p < 0.001). A high-throughput cell surface marker screen followed by gene and protein expression analysis revealed that both ECM sources downregulated CD164 and CXCR4, reducing CXCR4 protein levels by approximately 50% (p = 0.006 for eECM, p = 0.007 for ubECM). Notably, only eECM significantly suppressed OE33 cell migration (p ≤ 0.0001) and downregulated bone morphogenetic protein 4 BMP4 expression, along with its downstream targets pSMAD1/5/8, ID2, and SNAI2, thereby reducing epithelial-mesenchymal transition. These findings support the concept that biochemical cues from nonmalignant ECM modulate neoplastic cell behavior. Given the involvement of PI3K-Akt and BMP4 signaling in EAC progression, ECM-based strategies may warrant further investigation as potential therapeutic approaches following esophageal cancer resection.