Alves A.L.Costa-Gouveia J.Vieira de Castro J.Sotelo C.G....
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查看更多>>摘要：? 2022The host immunologic response to a specific material is a critical aspect when considering it for clinical implementation. Collagen and gelatin extracted from marine sources have been proposed as biomaterials for tissue engineering applications, but there is a lack of information in the literature about their immunogenicity. In this work, we evaluated the immune response to collagen and/or gelatin from blue shark and codfish, previously extracted and characterized. After endotoxin evaluation, bone marrow-derived macrophages were exposed to the materials and a panel of pro- and anti-inflammatory cytokines were evaluated both for protein quantification and gene expression. Then, the impact of those materials in the host was evaluated through peritoneal injection in C57BL/6 mice. The results suggested shark collagen as the less immunogenic material, inducing low expression of pro-inflammatory cytokines as well as inducible nitric oxide synthase (encoded by Nos2) and high expression of Arginase 1 (encoded by Arg1). Although shark gelatin appeared to be the material with higher pro-inflammatory expression, it also presents a high expression of IL-10 (anti-inflammatory cytokine) and Arginase (both markers for M2-like macrophages). When injected in the peritoneal cavity of mice, our materials demonstrated a transient recruitment of neutrophil, being almost non-existent after 24 hours of injection. Based on these findings, the studied collagenous materials can be considered interesting biomaterial candidates for regenerative medicine as they may induce an activation of the M2-like macrophage population, which is involved in suppressing the inflammatory processes promoting tissue remodeling. Statement of significance: Marine-origin biomaterials are emerging in the biomedical arena, namely the ones based in marine-derived collagen/gelatin proposed as cell templates for tissue regeneration. Nevertheless, although the major cause of implant rejection in clinical practice is the host's negative immune response, there is a lack of information in the literature about the immunological impact of these marine collagenous materials. This work aims to contribute with knowledge about the immunologic response to collagen/gelatin extracted from blue shark and codfish skins. The results demonstrated that despite some differences observed, all the materials can induce a macrophage phenotype related with anti-inflammation resolution and then act as immuno-modulators and anti-inflammatory inducible materials.
查看更多>>摘要：? 2022 Acta Materialia Inc.The clinical success of Toll-like receptor (TLR) agonists is based on their capacity to efficiently mobilize both innate and adaptive immunity. However, rapid distribution of TLR agonists into the systemic circulation may result in systemic cytokine storms. Telratolimod (Tel) is a TLR 7/8 agonist whose structure has a hydrophobic long chain that helps to prolong its release. Despite this, the phase I study of Tel showed cytokine release syndromes in 3/35 patients. Herein, we designed an injectable phase transition gel (PGE) that served as a superior drug depot for fatty acid-modified drugs. PGE further minimized the systemic drug exposure of Tel and the possible cytokine storms. In vivo studies demonstrated that Tel@PGE facilitated the recruitment of effector CD8+ T lymphocytes (T cells) and the polarization of myeloid-derived suppressor cells (MDSCs) and immunosuppressive M2-like macrophages to tumoricidal antigen-presenting cells. The reshaping of the tumor microenvironment (TME) by Tel@PGE elicited systematic immune responses to significantly prevent B16F10 or 4T-1 tumor postoperative recurrence and metastasis. Therefore, this platform of Tel is expected to provide a clinically available option for effective postoperative combined therapy. Statement of significance: A series of prodrugs or conjugates containing hydrophobic blocks were designed to achieve sustained release at the injection site by reducing the water solubility. However, this strategy sometimes failed short of expectations. Thus, we constructed a biocompatible and biodegradable injectable phase transition gel (PGE) with superior release properties that can be injected subcutaneously into the surgery site. In the long-lasting treatment, the melanoma and breast cancer immunotherapeutic effect significantly enhanced and the risk of cancer metastasis and relapse was reduced. Crucially, for some immune agonists, a superior release control can significantly reduce adverse effects which was decisive for the availability of the drugs.
查看更多>>摘要：? 2022 Acta Materialia Inc.Most dry eye syndromes (DES) are caused by oxidative stress and an overactive inflammatory response, leading to tear deficiency and excessive tear evaporation. Conventional eye drops for DES treatment require high doses and frequent administration due to their insufficient precorneal residence time. To overcome these problems, in this study, we have developed carbonized nanogels (CNGs) via the straightforward pyrolysis of lysine hydrochloride (Lys) to provide a long-lasting eye drop formulation for topical DES therapy. This methodology thermally converts Lys-into nitrogen-doped crosslinked polymers with embedded nanographitic structures, which enable efficient free radical scavenging. The cationic and crosslinked polymeric features of the Lys-CNGs also prolong the precorneal retention time and improve ocular bioavailability. These Lys-CNGs exhibit high biocompatibility with corneal epithelial cells both in vitro and in vivo, indicating their safety as eye drops. In a DES rabbit model, a single dose of Lys-CNGs (50 μg mL?1) can effectively alleviate the signs of DES within 4 days, whereas multiple treatments of 10-fold higher concentration of cyclosporine A are needed to achieve similar therapeutic effects (one dose every 12 h; 500 μg mL?1). The topical administration of Lys-CNGs enable a reduced therapeutic dose and extended dosing interval, thereby demonstrating a superior therapeutic efficacy compared to the commercial cyclosporine A eye drops. These Lys-CNGs, which exhibit significant free radical scavenging, anti-inflammatory activity, high biocompatibility, and a remarkable ocular bioadhesive property, hold great potential as a long-lasting eye drop formulation for the treatment of dry eye disease. Statement of significance: Multifunctional nanobiomaterial-based eye drops can render an ideal pharmaceutical formulation for the treatment of a variety of ocular surface diseases. To our knowledge, this is the first report describing the development of carbonized nanogels as topically administered therapeutics for alleviating dry eye syndrome (DES). We present evidence that the thermal transformation of lysine hydrochloride into carbonized nanogels (Lys-CNGs) endows superior antioxidant, anti-inflammatory, and bioadhesive properties. While a single dose of Lys-CNGs (50 μg mL?1) is sufficient to relieve the symptoms of DES for 4 days, multiple treatments of 10-fold higher concentration of commercially available cyclosporine eye drops are needed to achieve similar therapeutic outcomes (one dose every 12 h; 500 μg mL?1), suggesting an effective and long-lasting ocular carbonized nanomedicine.
查看更多>>摘要：? 2022Thermo-gels based on chitosan crosslinked poly(N-isopropylacrylamide) were developed as alternatives to conventional eye drops for the sustained release of ketotifen fumarate in the treatment of allergic conjunctivitis. The thermo-gelling properties of the base polymer were altered prior to crosslinking with chitosan by incorporation of the hydrophilic and hydrophobic comonomers acrylic acid and methyl methacrylate respectively. Varying amounts of chitosan were incorporated by ionic interaction to produce polyelectrolyte complexes or by carbodiimide chemistry to produce covalently crosslinked networks. The lower critical solution temperature of all the chitosan crosslinked thermo-gels produced was below the surface temperature of the eye. All the chitosan crosslinked thermo-gels were found to have greater than 80% equilibrium water contents following gelation. The method and amount of chitosan incorporation allowed for tailor-ability of material rheologic properties, with full degradation occurring over a one-to-four-day period, and tailorable rates of release of 40–60% of the loaded allergy medication ketotifen fumarate. The chitosan crosslinked thermo-gels were demonstrated to be nontoxic both in vitro and in vivo. It was demonstrated that the synthesized materials could be applied to the inferior fornix of eye, sustaining a multiple day release of ketotifen fumarate, as an alternative to conventional eyedrops. Statement of Significance: Topical eyedrops are the main treatment modality for anterior ocular conditions. However, due to the natural clearance mechanisms of the eye, topical eyedrops are well established to be largely ineffective as a method of drug delivery. Herein, we investigate a method of altering thermo-gel properties of an n-isopropylacrylamide based polymer to enable the incorporation of greater amounts of chitosan by different methods of crosslinking. By controlling the synthesis parameters, final material properties can be tailored to impart ideal spreading, retention on the eye, and the rate of degradation and drug release over several days. This work also focuses on studying the rheological properties of the chitosan crosslinked thermo-gels which has not been investigated previously.
查看更多>>摘要：? 2022 Acta Materialia Inc.Chitosan oligosaccharide-stearic acid-Valyl-Valyl-Valine/1-2-Dioleoyl-sn-glycero-3-phosphoethanolamine (CSO-SA-VVV5:2/DOPE) nanomicelles were rationally designed and developed for topical drug delivery to the posterior segment of the eye. The new ligand of VVV selected by computer-aided design exhibited better peptide transporter 1 active targeting in human conjunctival epithelial cells (HConEpiC) than other ligands mentioned in this project. The classic membrane fusion lipid of DOPE was indicated to facilitate the stability and lysosomal escape of the mixed nanomicelles. F?rster Resonance Energy Transfer was used to investigate the integrity of mixed nanomicelles in HConEpiC after passing through cell monolayer as well as in ocular tissues after topical administration. The results indicated that mixed nanomicelles could keep more intact micellar structure than CSO-SA nanomicelles in transit. These findings suggested that CSO-SA-VVV5:2/DOPE nanomicelles could overcome multiple ocular barriers and offer an efficient strategy for drug delivery from ocular surface to the posterior segment of the eye. Statement of significance: Ocular drug delivery systems face multiple physiological barriers in delivering drugs to the posterior segment of the eye by topical administration. In this study, new ligand of Valyl-Valyl-Valine was selected with computer-aided design for active targeting to peptide transporter 1 and anchored onto nanomicelles. Chitosan oligosaccharide-stearic acid- Valyl-Valyl-Valine/1-2-Dioleoyl-sn-glycero-3-phosphoethanolamine nanomicelles were rational designed. The mixed nanomicelles exhibited better active targeting ability and lysosomal escape. Nanomicellar integrity analysis with fluorescence resonance energy transfer technique demonstrated that mixed nanomicelles significantly enhanced cell permeability and exhibited more intact micellar structure in transit. These results suggested that the mixed nanomicelle eye drops have the potential to deliver drugs from ocular surface to the posterior segment of the eye.
查看更多>>摘要：? 2022Hydrogels are widely used as substrates to investigate interactions between cells and their microenvironment as they mimic many attributes of the extracellular matrix. The stiffness of hydrogels is an important property that is known to regulate cell behavior. Beside stiffness, cells also respond to structural cues such as mesh size. However, since the mesh size of hydrogel is intrinsically coupled to its stiffness, its role in regulating cell behavior has never been independently investigated. Here, we report a hydrogel system whose mesh size and stiffness can be independently controlled. Cell behavior, including spreading, migration, and formation of focal adhesions is significantly altered on hydrogels with different mesh sizes but with the same stiffness. At the transcriptional level, hydrogel mesh size affects cellular mechanotransduction by regulating nuclear translocation of yes-associated protein. These findings demonstrate that the mesh size of a hydrogel plays an important role in cell-substrate interactions. Statement of significance: Hydrogels are ideal platforms with which to investigate interactions between cells and their microenvironment as they mimic many physical properties of the extracellular matrix. However, the mesh size of hydrogels is intrinsically coupled to their stiffness, making it challenging to investigate the contribution of mesh size to cell behavior. In this work, we use hydrogel-on-glass substrates with defined thicknesses whose stiffness and mesh size can be independently tuned. We use these substrates to isolate the effects of mesh size on cell behavior, including attachment, spreading, migration, focal adhesion formation and YAP localization in the nucleus. Our results show that mesh size has significant, yet often overlooked, effects, on cell behavior, and contribute to a further understanding of cell-substrate interactions.
查看更多>>摘要：? 2022Osteochondral lesion potentially causes a variety of joint degenerative diseases if it cannot be treated effectively and timely. Microfracture as the conservative surgical choice achieves limited results for the larger defect whereas cartilage patches trigger integrated instability and cartilage fibrosis. To tackle aforementioned issues, here we explore to fabricate an integrated osteochondral scaffold for synergetic regeneration of cartilage and subchondral bone in one system. On the macro level, we fabricated three integrated scaffolds with distinct channel patterns of Non-channel, Consecutive-channel and Inconsecutive-channel via Selective Laser Sintering (SLS). On the micro level, both cartilage zone and subchondral bone zone of integrated scaffold were made of small polycaprolactone (PCL) microspheres and large PCL microspheres, respectively. Our findings showed that Inconsecutive-channel scaffolds possessed integrated hierarchical structure, adaptable compression strength, gradient interconnected porosity. Cartilage zone presented a dense phase for the inhibition of vessel invasion while subchondral bone zone generated a porous phase for the ingrowth of bone and vessel. Both cartilage regeneration and subchondral bone remodeling in the group of Inconsecutive-channel scaffolds have been demonstrated by histological evaluation and immunofluorescence staining in vivo. Consequently, our current work not only achieves an effective and regenerative microsphere scaffold for osteochondral reconstruction, but also provides a feasible methodology to recover injured joint through integrated design with diverse hierarchy. Statement of significance: Recovery of osteochondral lesion highly depends on hierarchical architecture and tunable vascularization in distinct zones. We therefore design a special integrated osteochondral scaffold with inconsecutive channel structure and vascularized modulation. The channel pattern impacts on mechanical strength and the infiltration of bone marrow, and eventually triggers synergetic repair of osteochondral defect. The cartilage zone of integrated scaffolds consisted of small PCL microspheres forms a dense phase for physical restriction of vascularized infiltration whereas the subchondral bone zone made of large PCL microspheres generates porous trabecula-like structure for promoting vascularization. Consequently, the current work indicates both mechanical adaptation and regional vascularized modulation play a pivotal role on osteochondral repair.
查看更多>>摘要：? 2022 Acta Materialia Inc.Bio-inspired nanostructures have demonstrated highly efficient mechano-bactericidal performances with no risk of bacterial resistance; however, they are prone to become contaminated with the killed bacterial debris. Herein, a biocompatible mechano-bactericidal nanopatterned surface with salt-responsive bacterial releasing behavior is developed by grafting salt-responsive polyzwitterionic (polyDVBAPS) brushes on a bio-inspired nanopattern surface. Benefiting from the salt-triggered configuration change of the grafted polymer brushes, this dual-functional surface shows high mechano-bactericidal efficiency in water (low ionic strength condition), while the dead bacterial residuals can be easily lifted by the extended polymer chains and removed from the surface in 1 M NaCl solution (high ionic strength conditions). Notably, this functionalized nanopatterned surface shows selective biocidal activity between bacterial cells sand eukaryotic cells. The biocompatibility with red blood cells (RBCs) and mammalian cells was tested in vitro. The histocompatibility and prevention of perioperative contamination activity were verified by in vivo evaluation in a rat subcutaneous implant model. This nanopatterned surface with bacterial killing and releasing activities may open new avenues for designing bio-inspired mechano-bactericidal platforms with long-term efficacy, thus presenting a facile alternative in combating perioperative-related bacterial infection. Statement of significance: Bioinspired nanostructured surfaces with noticeable mechano-bactericidal activity showed great potential in moderating drug-resistance. However, the nanopatterned surfaces are prone to be contaminated by the killed bacterial debris and compromised the bactericidal performance. In this study, we provide a dual-functional antibacterial conception with both mechano-bactericidal and bacterial releasing performances not requiring external chemical bactericidal agents. Additionally, this functionalized antibacterial surface also shows selective biocidal activity between bacteria and eukaryotic cells, and the excellent biocompatibility was tested in vitro and in vivo. The new concept for the functionalized mechano-bactericidal surface here illustrated presents a facile antibiotic-free alternative in combating perioperative related bacterial infection in practical application.
查看更多>>摘要：? 2022 The AuthorsLung implantable devices have been widely adopted as mechanical interventions for a wide variety of pulmonary pathologies. Despite successful initial treatment, long-term efficacy can often be impacted by fibrotic or granulation tissue formation at the implant sites. This study aimed to explore the lung-device interface by identifying the adhered proteome on lung devices explanted from patients with severe emphysema. In this study, scanning electron microscopy is used to visualize the adhesion of cells and proteins to silicone and nitinol surfaces of explanted endobronchial valves. By applying high-resolution mass-spectrometry, the surface proteome of eight explanted valves is characterized, identifying 263 unique protein species to be mutually adsorbed on the valves. This subset is subjected to gene enrichment analysis, matched with known databases and further validated using immunohistochemistry. Enrichment analyses reveal dominant clusters of functionally-related ontology terms associated with coagulation, pattern recognition receptor signaling, immune responses, cytoskeleton organization, cell adhesion and migration. Matching results show that extracellular matrix proteins and damage-associated molecular patterns are cardinal in the formation of the surface proteome. This is the first study investigating the composition of the adhered proteome on explanted lung devices, setting the groundwork for hypothesis generation and further exploration. Statement of significance: This is the first study investigating the composition of the adhered proteome on explanted lung devices. Lung implantable devices have been widely adopted as mechanical interventions for pulmonary pathologies. Despite successful initial treatment, long-term efficacy can often be impacted by fibrotic or granulation tissue formation around the implant sites. We identified the adhered proteome on explanted lung devices using several techniques. We identified 263 unique protein species to be mutually adsorbed on explanted lung devices. Pathway analyses revealed that these proteins are associated with coagulation, pattern recognition receptor signaling, immune responses, cytoskeleton organization, cell adhesion and migration. Furthermore, we identified that especially extracellular matrix proteins and damage-associated molecular patterns were cardinal in the formation of the surface proteome.
查看更多>>摘要：? 2022Peripheral nerve injury causes severe loss of motor and sensory functions, consequently increasing morbidity in affected patients. An autogenous nerve graft is considered the current gold standard for reconstructing nerve defects and recovering lost neurological functions; however, there are certain limitations to this method, such as limited donor nerve supply. With advances in regenerative medicine, recent research has focused on the fabrication of tissue-engineered nerve grafts as promising alternatives to the autogenous nerve grafts. In this study, we designed a nerve guidance conduit using an electrospun poly(lactide-co-ε-caprolactone) (PLCL) membrane with a visible light-crosslinked gelatin hydrogel. The PLCL nanoporous membrane with permeability served as a flexible and non-collapsible epineurium for the nerve conduit; the inner-aligned gelatin hydrogel paths were fabricated via 3D printing and a photocrosslinking system. The resultant gelatin hydrogel with microgrooved surface pattern was established as a conducting guidance path for the effective regeneration of axons and served as a reservoir that can incorporate and release bioactive molecules. From in vivo performance tests using a rat sciatic nerve defect model, our PLCL/gelatin conduit demonstrated successful axonal regeneration, remyelination capacities and facilitated functional recovery. Hence, the PLCL/gelatin conduit developed in this study is a promising substitute for autogenous nerve grafts. Statement of significance: Nerve guidance conduits (NGCs) are developed as promising recovery techniques for bridging peripheral nerve defects. However, there are still technological limitations including differences in the structures and components between natural peripheral nerve and NGCs. In this study, we designed a NGC composed of an electrospun poly(lactide-co-ε-caprolactone) (PLCL) membrane and 3D printed inner gelatin hydrogel to serve as a flexible and non-collapsible epineurium and a conducting guidance path, respectively, to mimic the fascicular structure of the peripheral nerve. In particular, in vitro cell tests clearly showed that gelatin hydrogel could guide the cells and function as a reservoir that incorporate and release nerve growth factor. From in vivo performance tests, our regenerative conduit successfully led to axonal regeneration with effective functional recovery.
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Elsevier