Frontiers in Bioengineering and Biotechnology最新文献

Hemophilia is an inherited bleeding disorder caused by mutations in the F8 or F9 gene, leading to a deficiency or dysfunction of coagulation factors VIII or IX. While current treatments, such as factor replacement, extended half-life factors, and gene therapy, have improved patient outcomes, they have limitations such as immunogenicity, transient transgene expression, and the requirement for high vector doses. Gene editing for hemophilia is an emerging approach that aims to provide a permanent cure by editing the mutated gene precisely or targeted integration of coagulation factor cDNA into the host genome for stable expression. This approach involves the use of programmable nucleases (CRISPR/Cas9, TALENs, ZFNs) that induce double-stranded DNA breaks at specific sites, allowing precise correction or targeted transgene integration. This review covers the various editing tools and strategies used for precise gene editing in hemophilia, including approaches such as HDR, NHEJ, base editing, prime editing, ex vivo gene editing in iPSCs, and recent LNP-based CRISPR delivery methods for precise editing.

Introduction: Bone tissue engineering requires scaffolds that mimic the native extracellular matrix and provide sustained delivery of osteoinductive factors. This study focuses on developing a multifunctional scaffold using a green electrospinning process to combine the biocompatibility of silk fibroin (SF) with a non-viral gene delivery system for sustained expression of Bone Morphogenetic Protein 2 (BMP2).

Methods: A green electrospinning technique, using an aqueous SF and polyethylene oxide (PEO) solution, was employed to fabricate nanofibrous scaffolds, eliminating the use of harsh organic solvents. Polyethylenimine (PEI) modified liposomes (Lipo^PEI) were used to encapsulate a BMP2-encoding plasmid (pDNA _Bmp2 ). These gene-loaded nanoparticles were incorporated into the SF-PEO nanofibers. The resulting scaffolds were characterized for morphology (SEM), structure (FTIR, XRD), and drug release kinetics. Biological performance was evaluated by assessing cell viability (MTT assay), cell attachment (SEM), gene transfection efficiency (confocal microscopy), and osteogenic differentiation (alkaline phosphatase (ALP) activity, Alizarin Red S staining) using bone marrow mesenchymal stem cells (BMSCs).

Results: Physicochemical characterization confirmed the successful formation of uniform pDNA_Bmp2@Lipo^PEI nanocomplexes with a particle size of approximately 266 nm and a positive surface charge of +16.9 mV. These nanocomplexes were homogeneously incorporated into smooth, bead-free SF-PEO nanofibers with average diameters ranging from 460 to 541 nm. The composite scaffold demonstrated a highly sustained release of pDNA_Bmp2 over 14 days. In vitro studies using rat bone marrow-derived mesenchymal stem cells (BMSCs) revealed that the scaffold possesses excellent biocompatibility, promoting robust cell adhesion, spreading, and proliferation. Furthermore, the gene-loaded scaffold successfully mediated the transfection of BMSCs, leading to significant upregulation of osteogenic markers, including alkaline phosphatase (ALP) activity and extensive calcium mineral deposition over 21 days.

Discussion: The novel composite scaffold combines the structural advantages of SF with a sustained BMP2 gene delivery system, showing remarkable potential to promote osteogenic differentiation. This work presents a promising, environmentally friendly, and effective platform for bone tissue engineering and regenerative medicine.

Objective: To compare the biomechanical properties of the cement-screw technique with the metal block extension stem technique in reconstructing Anderson Orthopaedic Research Institute (AORI) type 2 tibial defects in total knee arthroplasty using the finite element method, in order to provide a basis for clinical selection.

Methods: Non-inclusive 5-mm and 10-mm depth AORI type 2 tibial defects were modeled using the finite element method. The cement-screw technique and the metal block extension stem technique were applied for reconstruction, resulting in a total of four sets of finite element models. Each group of models was tested under two loads: horizontal walking and descending stairs. The von Mises stress distributions in the tibia, prosthesis, and bone defect regions, as well as the peaks of micromotion at the prosthesis-tibia interface, were measured.

Results: In the AORI type 2 tibial defect model, the cement-screw group, when reconstructing 5 mm and 10 mm defects under horizontal walking and descending stairs loads, exhibited higher maximum tibial stress (5 mm: 78.0-140 MPa; 10 mm: 80.9-151 MPa), proximal tibial defect area stress (5 mm: 11.3-25.3 MPa; 10 mm: 10.8-24.1 MPa), and peak micromotion values (5 mm: 9.90-26.99 μm; 10 mm: 11.94-31.98 μm) compared to the metal block extension stem group (tibial stress 5 mm: 73.2-130 MPa, 10 mm: 66.6-118 MPa; defect area stress 5 mm: 7.83-16.3 MPa, 10 mm: 8.54-18.8 MPa; peak micromotion 5 mm: 6.56-14.72 μm, 10 mm: 8.92-24.09 μm). However, prosthesis stresses were lower in the cement-screw group (5 mm: 87.1-183 MPa; 10 mm: 60.2-158 MPa) than in the metal block extension stem group (5 mm: 101-194 MPa, 10 mm: 92.7-167 MPa). Under horizontal walking loading, the two techniques showed no superiority of one over the other in terms of the von Mises stresses and the peaks of micromotion; however, under descending stairs loads, the maximum stress in the tibia of the cement-screw group with a 10-mm defect exceeded 150 MPa (151 MPa), indicating a potential fracture risk, and the peaks of micromotion was smaller in all models.

Conclusion: The findings of this study indicate that the cement-screw technique is more cost-effective and convenient for repairing 5-mm defects and is appropriate for reconstruction of this size. However, when the bone defect reaches 10 mm, the cement-screw technique may elevate the risk of fracture, and thus, safety concerns must be taken into account. In contrast, the metal block extension stem technique offers a better balance between effectiveness and safety, making it the preferred option for defects of this size.

Background: Metabolic syndrome is a pathological state, frequently associated with type 2 diabetes, which is marked by abdominal obesity, impaired insulin action, hypertension, and vascular wall changes. Similar to humans, horses can suffer from equine metabolic syndrome. A representative in vitro vascular wall model is needed to study its pathophysiology and develop novel treatments for both human and equine patients.

Methods: In this study, scaffolds manufactured via digital light processing (DLP) exploiting an acrylate-endcapped urethane-based polymer precursor with a polyethylene glycol backbone (AUP2PEG) were coated with collagen or gelatin derivatives. Their cell-interactive properties were evaluated using equine mesenchymal stromal cells (MSC) and endothelial cells (EC). Coating was performed using either UV-induced photopolymerization of gelatin methacryloyl (GelMA) on the surface of the DLP-printed scaffold or physisorption of type I atelocollagen.

Results: The GelMA coating formed a thin, uniform layer on the scaffold surface and improved the cytocompatibility of DLP-printed AUP2PEG-based scaffolds for EC and MSC. Furthermore, they permitted MSC trilineage differentiation. To mimic the endothelial damage occurring in metabolic syndrome conditions, the GelMA-coated AUP2PEG scaffolds were incubated in high glucose culture conditions. Short-term cell culture in these conditions significantly reduced the number of viable EC. In contrast, the short-term culture of MSC in these conditions did not result in a similarly deleterious impact on cell viability.

Conclusion: In conclusion, GelMA-coated DLP-printed AUP2PEG scaffolds facilitate the growth of EC and MSC. Furthermore, exposing EC cultured on the developed scaffolds to hyperglycemic culture conditions negatively affects the viability of EC, comparable to what is observed in two-dimensional culture conditions.

Current clinical practice has extensively validated the efficacy of left ventricular assist devices (LVADs) in managing end-stage heart failure. A persistent challenge across all ventricular assist systems involves achieving optimal biocompatibility at the critical interface between the LVAD outlet and myocardial tissue. In this study, femtosecond laser processing (FLP) technology was utilized to engineer microtextured surfaces with controlled geometric parameters on the titanium alloy surface. The experimental design systematically assessed surface morphology and compositional variations for four distinct patterns (circular, triangular, square, hexagonal) and three texture depths (10, 20, 40 μm). FLP demonstrated favourable microstructural fabrication quality, producing defined pattern boundaries with minimal thermal impact on adjacent regions. While all textured surfaces exhibited characteristic periodic processing marks at their bases, increased texture depth correlated with progressive roughness amplification in these basal regions. Elemental analysis revealed that oxygen enrichment specifically along texture peripheries compared to untextured surfaces. Cellular early response studies demonstrated that surface texturing significantly enhanced cardiac fibroblasts adhesion on titanium substrates while concurrently modifying fibroblast growth patterns. Quantitative analysis identified 20 μm as the optimal texture depth for cellular proliferation and adhesion, outperforming both shallower (10 μm) and deeper (40 μm) configurations. Geometric comparisons indicated that square patterns induced the best pronounced pro-proliferative effects, followed by hexagonal patterns. Mechanistic observations suggest that surface micro-roughness facilitates initial cell adhesion, with subsequent proliferation biodynamics being governed by topographical guidance effects. These findings establish clear structure-function relationships between engineered surface parameters and biological responses, providing significant insights for LVAD surface treatment and optimization.

Purpose: This study compares the biomechanical performance of InterTan nails of three lengths (180 mm, 240 mm, and 320 mm) in treating AO/OTA 31A2.3 comminuted intertrochanteric fractures, which are highly unstable and prone to fixation failure. The research question focuses on identifying the nail length that optimizes stress distribution, displacement, and strain to enhance fracture healing and reduce failure risk, thereby providing a theoretical foundation for clinical decision-making.

Methods: Femoral CT images from a healthy 24-year-old male were used to reconstruct cortical and cancellous bone models in Mimics Research 21.0 and Geomagic Wrap 2021. A complete femur and AO/OTA 31A2.3 fracture model were constructed in SolidWorks 2022. InterTan models (180 mm, 240 mm, and 320 mm) were assembled with the fracture model, and finite element analysis (FEA) was performed in Ansys Workbench 18.0 under three loading conditions (standing, walking, and stair descent) to evaluate stress, deformation, and failure risk.

Results: Stress concentrated at the nail-screw junction and proximal aperture, with the 180 mm nail exhibiting the highest stress, the 320 mm nail the lowest, and the 240 mm nail intermediate values. Displacement increased with nail length under dynamic loads, whereas the 180 mm nail minimized displacement during standing. The 240 mm nail showed the lowest strain during standing but the highest during stair descent. Differences in stress and displacement were statistically significant (P < 0.05).

Conclusion: The 320 mm nail optimizes stress distribution, micromotion, and strain, thereby reducing failure risk and promoting healing. These findings align with biological osteosynthesis principles and support personalized treatment strategies.

Introduction: Stress shielding is a major cause of radiological changes in the humeral component, which is commonly evident in cementless stems of reverse shoulder implants. The bulkiness of the humeral stem results in less load being transmitted to the bone, curtailing bone remodeling. Designing an implant with adequate strength and a suitable material that matches the mechanical properties of bone can help prevent the implant migration or loosening, thereby lowering bone resorption.

Methods: Humeral stems with no porosity and varied porosities, such as circular, elliptical, and trabecular architecture, were designed using 316L stainless steel and titanium alloy (Ti6Al4V). Finite element analysis (FEA) was conducted on eight bone-implant assemblies under four loading conditions for cortical and trabecular bones. Weighted mean of von Mises stress and mechanobiology associated with the strain energy densities were studied. This serves as a precursor in predicting the effect of stress shielding.

Results: The titanium implant with trabecular architecture was mechanically close to the intact bone compared to the other varied porosity designs. It also had better load-bearing capacity than the solid stems.

Discussion: These investigations help understand the load-bearing capacity of reverse shoulder humeral stems and ascertain the importance of combining the design and material in enhancing implant stability and longevity.

Introduction: Given the high prevalence of vertebral fractures following radiotherapy in patients with metastatic spine disease, torso muscle segmentation is necessary for biomechanical modeling of vertebral loading, permitting individualized evaluation of fracture risk.

Methods: In this study, we developed and validated a deep-learning model for full volumetric segmentation of the thoracic and abdominal spinal musculature in cancer patients with metastatic spine disease from sparsely annotated clinical CT image data. We obtained CT data for 148 metastatic spine disease patients undergoing radiotherapy treatment, and an external set of randomly selected 30 subjects from the National Lung Screening Trial. We extracted 1924 axial CT images at the midpoint of each vertebral level (T4 to L4) and manually labeled the key extensor and flexor muscles (up to 8 muscles per side) at each level. We trained a 2D nnU-Net deep-learning (DL) model to segment each muscle and, using these sparse annotations, trained the model to segment each muscle's 3D volume per spine. Two experienced radiologists independently and blindly evaluated the anatomical fidelity of the segmentations using a Likert scale, for 1) manual- and 2) DL-segmentation, 3) random test samples from the muscle's 3D volume and 4) an external NLST CT data.

Results: The DL method achieved comparable performance to manual segmentations with a mean Dice score above 0.769. Mann-Whitney test analysis showed that the radiologist ratings of DL-generated muscle segmentations were noninferior to the manual segmentation for each muscle.

Discussion: Demonstrating excellent performance for rapid, high-anatomical fidelity 3D segmentation of the main flexor, extensor, and stabilizing thoracolumbar muscles, the DL model from clinical CT scans, this development holds significant potential for reducing the manual effort required to generate individualized musculoskeletal models in cancer patients.

Ovarian tissue cryopreservation and transplantation (OTCT) is an important fertility preservation method for female cancer patients; however, its efficacy is limited by post-transplantation ischemia-reperfusion injury, leading to oxidative stress, apoptosis, and fibrosis that impair ovarian reserve and graft function. Mesenchymal stem cell-conditioned medium shows therapeutic potential through paracrine actions, but clinical use is restricted by relatively limited antioxidant capacity and delivery challenges. To address this, an antioxidant-enriched hydrogel (PG-gel) was developed from N-acetylcysteine-modified gelatin and poly (ethylene glycol) succinimidyl succinate, loaded with conditioned medium from umbilical cord mesenchymal stem cells (CM-UCMSCs). This study evaluated the efficacy of PG-gel in protecting vitrified-thawed ovarian tissue during in vitro culture. The CM-UCMSCs-loaded PG-gel significantly suppressed intracellular reactive oxygen species generation. The PG + CM-UCMSCs group showed markedly reduced follicle loss, improved follicle morphology, decreased collagen deposition, lower apoptosis (fewer TUNEL-positive cells and reduced caspase-3 expression), diminished oxidative damage (lower 8-OHdG), and enhanced glucose consumption compared with the other culture groups. Transcriptomic analysis revealed downregulation of apoptosis-related genes (e.g., Ddit3, Trib3 and Hmox1) and upregulation of mitochondrial metabolism genes (e.g., Mt-atp8, Mt-nd1 and Mt-cyb). In conclusion, the PG + CM-UCMSCs system provided comprehensive protection to cryopreserved ovarian tissue by mitigating oxidative stress, fibrosis, and apoptosis, likely through regulation of apoptotic signaling and enhancement of mitochondrial energy metabolism, thereby offering a promising strategy to improve OTCT outcomes.

Total joint arthroplasty consistently alleviates pain and improves function in patients with end-stage joint disease. Nevertheless, up to 10% of hip and 20% of knee recipients remain dissatisfied after surgery, and registry data indicate that approximately one in five implants requires revision within 25 years, most commonly due to aseptic loosening, mechanical instability, or periprosthetic joint infection. Conventional postoperative surveillance relies on intermittent clinic visits and imaging, leaving a critical blind spot in our understanding of implant performance during daily activities. To address this gap, research has turned to fully implantable smart prostheses, such as hip and knee implants, embedded with sensors and low-power wireless telemetry that enable real-time monitoring of in vivo conditions. This review traces the evolution from early instrumented prototypes to the first commercially available smart knee; outlines enabling technologies, including sensing, communication, powering, and system integration; and summarizes clinical applications and early human data across this development continuum. Smart implants capture objective in vivo parameters that are not accessible to routine follow-up, including joint loads, range of motion, spatiotemporal gait metrics, and temperature, thereby enabling orthopedic phenotyping through dynamic, longitudinal digital representations of recovery trajectories and complication patterns. Fully implantable smart prostheses have the potential to shift arthroplasty toward continuous remote monitoring and proactive, precision follow-up care. Coupled with robust clinical decision-support systems and rigorous long-term evaluation, these technologies may usher in a new era of intelligent joint arthroplasty, with the potential to improve outcomes and extend implant longevity.