Journal of Tissue Engineering and Regenerative Medicine最新文献

Peripheral neuropathy is painful and can cause a considerable decline in quality of life. Surgery and autograft are the current approaches and clinical standards for restoring function after nerve damage. However, they usually result in unacceptable clinical results, so we need modern peripheral nerve defect treatment approaches. Tissue engineering techniques have been developed as a promising approach, but there are some considerations for translational application. Clinical application of novel tissue engineering methods is related to combining the appropriate cell and scaffold type to introduce safe and efficient bioscaffolds. Efficient nerve regeneration occurs by mimicking the extracellular matrix and combining topographical, biochemical, mechanical, and conductive signs via different cells, biomolecules, and polymers. In brief, ideal engineered biomaterial scaffolds will have to cover all characteristics of nerve tissue, such as nerve number, myelin, and axon thickness. Nerve regeneration has a highly sensitive response to its surrounding microenvironment. For designing a suitable construct, matching the regenerative potential of the autograft as the golden standard is essential. This review article examines the newest advancements in peripheral nerve tissue engineering. Specifically, the discussion will focus on incorporating innovative cues, biological modification, biomaterials, techniques, and concepts in this area of research.

Wound healing is an intricate process involving multiple cells and distinct phases, presenting challenges for comprehensive investigations. Currently available treatments for wounds have limited capacity to fully restore tissue and often require significant investments of time in the form of repetitive dressing changes and/or reapplications. This article presents a novel study that aims to enhance wound healing by developing biomaterial scaffolds using Medpor®, a porous polyethylene implant, as a model scaffold. The study incorporates electrospun poly(e-caprolactone) (PCL) fibers and a protein mixture (PM) containing collagen IV and laminin onto the Medpor® scaffolds. To evaluate the impact of these implants on wound healing, a unique splinted wound model in mice is employed. The wounds were evaluated for closure, inflammation, collagen deposition, angiogenesis, epithelialization, and proliferation. The results show that wounds treated with Medpor® + PCL + PM implants demonstrate accelerated closure rates, improved epithelialization, and enhanced angiogenesis compared to other implant groups. However, there were no significant differences observed in collagen deposition and inflammatory response among the implant groups. This study provides valuable insights into the potential benefits of incorporating PCL fibers and a PM onto scaffolds to enhance wound healing. Furthermore, the developed splinted wound model with integrated implants offers a promising platform for future studies on implant efficacy and the advancement of innovative wound healing strategies.

The challenge in developing tissue-engineered bones (TEBs) for clinical applications lies in the constraints associated with the source and availability of autologous mesenchymal stem cells (MSCs) derived from the bone marrow, which creates a bottleneck. While allogeneic MSCs have shown promise in TEB applications, their ability to promote bone growth is notably diminished because of the inflammatory reaction at the transplant site and the inherent immune response triggered by allogeneic MSCs. Hence, there is a pressing need to develop methods that enhance the osteogenic differentiation of allogeneic MSCs during transplantation. Previous studies have found that IL-17 is a key proinflammatory factor in initiating inflammation and cascade amplification in the early stages of an inflammatory response, and proinflammatory cytokines such as TNF-α and IL-17 can inhibit the osteogenic differentiation of MSCs in an immune environment. In this study, MSCs expressing HVEM were successfully constructed by viral transfection and further reconfirmed that IL-17 can inhibit the in vivo and in vitro osteogenesis of allogeneic MSCs through in vitro experiments and mouse calvarial bone defect (diameter about 3 mm) model, while MSCs that express herpesvirus-entry mediator (HVEM) exhibit the capacity to suppress immune responses and sustain strong osteogenic potential. We further pointed out that the mechanism by which HVEM promotes the osteogenesis of allogeneic MSCs is related to its inhibition of the IκB kinase (IKK)-NF-κB signaling pathway activated by IL-17 in the immune environment, which can significantly inhibit the ubiquitination and degradation of β-catenin in MSCs induced by the IKK-NF-κB pathway, upregulate the expression of β-catenin, and promote bone formation. Hence, this research provides an initial connection between the Wnt/β-catenin signaling pathway and the IKK-NF-κB pathway during allogeneic MSC transplantation, offering new avenues for investigation and establishing a theoretical foundation for the potential use of HVEM-expressing MSCs in clinical treatments for bone defects.

Persistent air leaks caused by thoracic surgery, physical trauma, or spontaneous pneumothoraces are a cause of patient morbidity with need for extended chest tube durations and surgical interventions. Current treatment measures involve mechanical closure of air leaks in the compromised pleura. Organ and membrane decellularisation offers a broad range of biomimetic scaffolds of allogeneic and xenogeneic origins, exhibiting innate tissue-specific characteristics. We explored a physicochemical method for decellularising porcine pleural membranes (PPM) as potential tissue-engineered surrogates for lung tissue repair. Decellularised PPM (dPPM) was characterised with histology, quantitative assays, mechanical testing, and sterility evaluation. Cytotoxicity and recellularisation assays assessed biocompatibility of decellularised PPM (dPPM). Haematoxylin and Eosin (H&E) staining showed an evident reduction in stained nuclei in the dPPM, confirmed with nuclear staining and analysis ( ^∗∗∗∗p < 0.0001). Sulphated glycosaminoglycans (sGAG) and collagen histology demonstrated minimal disruption to the gross structural assembly of core extracellular matrix (ECM) in dPPM. Confocal imaging demonstrated realignment of ECM fibres in dPPM against native control. Quantitative analysis defined a significant change in the angular distribution ( ^∗∗∗∗p < 0.0001) and coherence ( ^∗∗∗p < 0.001) of fibre orientations in dPPM versus native ECM. DNA quantification indicated ≥85% reduction in native nuclear dsDNA in dPPM ( ^∗∗p < 0.01). Collagen and sGAG quantification indicated reductions of both ( ^∗∗p < 0.01). dPPM displayed increased membrane thickness ( ^∗∗∗p < 0.001). However, Young’s modulus (459.67 ± 10.36 kPa) and ultimate tensile strength (4036.22 ± 155.1 kPa) of dPPM were comparable with those of native controls at (465.82 ± 10.51 kPa) and (3912.9 ± 247.42 kPa), respectively. In vitro cytotoxicity and scaffold biocompatibility assays demonstrated robust human mesothelial cell line (MeT-5A) attachment and viability. DNA quantification in reseeded dPPM with MeT-5A cells exhibited significant increase in DNA content at day 7 ( ^∗∗p < 0.01) and day 15 ( ^∗∗∗∗p < 0.0001) against unseeded dPPM. Here, we define a decellularisation protocol for porcine pleura that represents a step forward in their potential tissue engineering applications as bioscaffolds.

Tissue engineering is a promising approach for the production of small-diameter vascular grafts; however, there are limited data directly comparing the suitability of applicable cell types for vessel biofabrication. Here, we investigated the potential of adult smooth muscle cells (SMCs), placental mesenchymal stem cells (MSCs), placental endothelial colony-forming cells (ECFCs), and a combination of MSCs and ECFCs on highly porous biocompatible poly(ɛ-caprolactone) (PCL) scaffolds produced via melt electrowriting (MEW) for the biofabrication of tissue-engineered vascular grafts (TEVGs). Cellular attachment, proliferation, and deposition of essential extracellular matrix (ECM) components were analysed in vitro over four weeks. TEVGs cultured with MSCs accumulated the highest levels of collagenous components within a dense ECM, while SMCs and the coculture were more sparsely populated, ascertained via histological and immunofluorescence imaging, and biochemical assessment. Scanning electron microscopy (SEM) enabled visualisation of morphological differences in cell attachment and growth, with MSCs and SMCs infiltrating and covering scaffolds completely within the 28-day culture period. Coverage and matrix deposition by ECFCs was limited. However, ECFCs lined the ECM formed by MSCs in coculture, visualised via immunostaining. Thus, of cells investigated, placental MSCs were identified as the preferred cell source for the fabrication of tissue-engineered constructs, exhibiting extensive population of porous polymer scaffolds and production of ECM components; with the inclusion of ECFCs for luminal endothelialisation, an encouraging outcome warranting further consideration in future studies. In combination, these findings represent a substantial step toward the development of the next generation of small-diameter vascular grafts in the management of cardiovascular disease.

Human mesenchymal stem cells (hMSCs) are multipotent cells that differentiate into adipocytes, chondrocytes, and osteoblasts. Owing to their differentiation potential, hMSCs are among the cells most frequently used for therapeutic applications in tissue engineering and regenerative medicine. However, the number of cells obtained through isolation alone is insufficient for hMSC-based therapies and basic research, which necessitates in vitro expansion. Conventionally, this is often performed on rigid surfaces such as tissue culture plates (TCPs). However, during in vitro expansion, hMSCs lose their proliferative ability and multilineage differentiation potential, rendering them unsuitable for clinical use. Although multiple approaches have been attempted to maintain hMSC stemness during prolonged expansion, finding a suitable culture system remains an unmet need. Recently, a few research groups have shown that hMSCs maintain their stemness over long passages when cultured on soft substrates. In addition, it has been shown that hMSCs cultured on soft substrates have more condensed chromatin and lower levels of histone acetylation compared to those cultured on stiff substrates. Furthermore, it has also been shown that condensing/decondensing chromatin by deacetylation/acetylation can delay replicative senescence in hMSCs during long-term expansion on TCPs. However, the mechanism by which chromatin condensation/decondensation influences nuclear morphology and DNA damage, which are strongly related to the onset of senescence, remains unknown. To answer this question, we cultured hMSCs for long duration in the presence of epigenetic modifiers, histone acetyltransferase inhibitor (HATi), which promotes chromatin condensation by preventing histone acetylation, and histone deacetylase inhibitor (HDACi), which promotes chromatin decondensation, and investigated their effects on various nuclear markers related to senescence. We found that consistent acetylation causes severe nuclear abnormalities, whereas chromatin condensation by deacetylation helps to safeguard the nucleus from damage caused by in vitro expansion.

The aim of the study was to investigate the therapeutic potential of exosomes secreted by endothelial cells (EC-exos) on steroid-induced osteonecrosis of femoral head (SNFH). First, we successfully obtained EC-exos through differential centrifugation. Then, the effects of EC-exos on mouse embryo osteoblast precursor (MC3T3-E1) cells under high concentration of dexamethasone (Dex) were analysed in vitro, which included cell migration, viability, and apoptosis. In vivo, a SNFH rat model was successfully established and treated with EC-exos. Micro-computed tomography (micro-CT) and haematoxylin and eosin (H&E) were used to observe femoral trabeculae. Our in vitro results showed that EC-exos improved cell viability and migration of osteoblasts and reduced the apoptotic effect of high concentration of Dex on osteoblasts in vitro. Phosphoinositide 3-kinase (PI3K)/Akt/Bcl-2 signalling pathway was activated in MC3T3-E1 cells under the response to EC-exos. In vivo, increased bone volume per tissue volume (BV/TV) (p = 0.031), trabecular thickness (Tb.Th) (p = 0.020), and decreased separation (Tb.Sp) (p = 0.040) were observed in SNFH rats treated with EC-exos. H&E staining revealed fewer empty lacunae and pyknotic osteocytes in trabeculae. The expression of Bcl-2 and Akt in EC-exos group was significantly increased in trabeculae tissue. Overall, our finding indicated that EC-exos could attenuate SNFH by inhibiting osteoblast apoptosis via the PI3K/Akt/Bcl-2 pathway.

Saethre-Chotzen syndrome (SCS) is one of the most prevalent craniosynostosis, caused by a loss-of-function mutation in the TWIST-1 gene, with current treatment options relying on major invasive transcranial surgery. TWIST-1 haploinsufficient osteogenic progenitor cells exhibit increased osteogenic differentiation potential due to an upregulation of the transmembrane tyrosine kinase receptor, C-ROS-1, a TWIST-1 target gene known to promote bone formation. The present study assessed the efficacy of suppressing C-ROS-1 activity using a known chemical inhibitor to C-ROS-1, crizotinib, to halt premature coronal suture fusion in a preclinical mouse model of SCS. Crizotinib (1 μM, 2 μM, or 4 μM) was administered locally over the calvaria of Twist-1^del/+ heterozygous mice prior to coronal suture fusion using either a nonresorbable collagen sponge (quick drug release) or a resorbable sodium carboxymethylcellulose microdisk (slow sustained release). Coronal suture fusion rates and bone parameters were determined by μCT imaging and histomorphometric analysis of calvaria postcoronal suture fusion. Results demonstrated a dose-dependent increase in the efficacy of crizotinib to maintain coronal suture patency, with no adverse effects to brain, kidney, liver, and spleen tissue, or blood cell parameters. Moreover, crizotinib delivered on microdisks resulted in a greater efficacy at a lower concentration to reduce bone formation at the coronal suture sites compared to sponges. However, the bone inhibitory effects were found to be diminished by over time following cessation of treatment. Our findings lay the foundation for the development of a pharmacological nonsurgical, targeted approach to temporarily maintain open coronal sutures in SCS patients. This study could potentially be used to develop similar therapeutic strategies to treat different syndromic craniosynostosis conditions caused by known genetic mutations.

While acute wounds typically progress through the phases of wound healing, chronic wounds often stall in the inflammatory phase due to elevated levels of matrix metalloproteinases (MMPs) and proinflammatory cytokines. Dysregulated expression of MMPs can result in the breakdown of extracellular matrix (ECM) formed during the wound healing process, resulting in stalled wounds. Native collagen-based wound dressings offer a potential wound management option to sequester excess MMPs and support cellular interactions that allow wound progression through the natural healing process. Herein, we utilized commercially available ECM matrices, two derived from porcine small intestinal submucosa (PCMP, 2 layers; PCMP-XT, 5 layers) and one derived from propria submucosa (ovine forestomach matrix, OFM, 1 layer), to demonstrate the impact of processing methodologies (e.g., layering and crosslinking) on functional characteristics needed for the management of chronic wounds. Grafts were evaluated for structural composition using scanning electron microscopy and histology, ability to reduce MMPs using fluorometric assays, and durability in an in vitro degradation chronic wound model. Both intact (nondegraded) and partially degraded grafts were assessed for their ability to serve as a functional cell scaffold using primary human fibroblasts. Grafts differed in matrix substructure and composition. While all grafts demonstrated attenuation of MMP activity, PCMP and PCMP-XT showed larger reductions of MMP levels. OFM rapidly degraded in the in vitro degradation model (<3 hours), while PCMP and PCMP-XT were significantly more durable (>7 days). The ability of PCMP and PCMP-XT to serve as scaffolds for cellular attachment was not impacted by degradation in vitro. Three ECM grafts with varying structural and functional characteristics exhibited differential durability when degraded in a simulated chronic wound model. Those that withstood rapid degradation maintained their ability to function as a scaffold to support attachment and proliferation of fibroblasts, a cell type important for wound healing.

Introduction. The decellularized human amniotic membrane (dHAM) emerges as a viable 3D scaffold for organ repair and replacement using a tissue engineering strategy. Glutaraldehyde (GTA) and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) can increase the biomechanical properties of dHAM. However, the crosslinking process is associated with biochemical changes and residual toxic materials, dampening the biocompatibility of the dHAM. From a histologic point of view, each side of the amniotic membrane is biologically different. While the dHAM basement membrane side is rich in growth factors, the stromal side of the dHAM contains more connective tissue matrix (e.g., collagen fibers) which supports its biomechanical properties. Biocompatibility and biomechanical properties are two important challenges in the field of materials science. In this study, for the first time, the stromal and basement membrane side are cross-linked with GTA and EDC, respectively, to optimize the biocompatibility of the treated dHAM while sparing the GTA-mediated biomechanical improvements. Methods. Crosslinking was carried out on dHAM in three groups: EDC, GTA and bilateral treatment with EDC&GTA. Mechanical resistance, degradability, and crosslinking measurements were performed on treated dHAM. The viability of mesenchymal stem cells (MSCs) on the scaffolds was evaluated by the MTT assay. The expression levels of surface markers and images of the MSCs were thoroughly studied. Results. The results obtained showed that bilateral treatment of dHAM with EDC and GTA increased mechanical resistance. Similarly, the evaluation of surface markers revealed that bilaterally treated dHAM sustains the stemness and viability of MSCs at a level equal to that achieved with EDC alone. The SEM images indicated that the MSCs maintained adhesion on EDC&GTA-cross-linked dHAM. Conclusion. The current study explores a pioneering treatment of dHAM, a material long recognized for its regenerative properties, in a novel context. This research delves into the utilization of dHAM cross-linked with EDC&GTA, demonstrating its optimized efficacy in tissue engineering. The enhanced crosslinking technique significantly alters the membrane’s properties, amplifying its durability and therapeutic potential. In this novel bilateral treatment strategy (EDC and GTA), improving mechanical properties by GTA on the stromal surface and maintaining the biocompatibility of EDC on the side of the basement membrane of dHAM had been attained together. By investigating the handling and impact of this cross-linked membrane, this study unveils a new approach in leveraging a well-known material through an innovative process, revolutionizing its application in wound care.