Aligned Scaffolds Research Articles

Cell-Free Biomimetic Scaffold with Cartilage Extracellular Matrix-Like Architectures for In Situ Inductive Regeneration of Osteochondral Defects.

The development of a biomimetic scaffold designed to provide a native extracellular matrix (ECM)-like microenvironment is a potential strategy for cartilage repair. The ECM in native articular cartilage is structurally composed of three different architectural zones, i.e., horizontally aligned, randomly arranged, and vertically aligned collagen fibers. However, the effects of scaffolds with these three different ECM-like architectures on in vivo cartilage regeneration are not clear. In this study, we aim to systematically investigate and compare their in situ inductive regenerative efficacy on cartilage defects. ECM-mimetic silk fibroin scaffolds with horizontally aligned, vertically aligned, and random pore architectures are fabricated using the controlled directional freezing technique. All of these scaffolds exhibit similar pore area, swelling ratio, and in vitro degradation behavior. Nevertheless, the aligned scaffolds have a higher pore aspect ratio and hydrophilicity, and increase the proliferation of bone marrow-derived mesenchymal stem cells (BMSCs) in vitro. When implanted into rabbit osteochondral defects, the scaffold with vertically aligned pore architectures provides a more cell-favorable microenvironment conducive to endogenous BMSCs than other scaffolds and supports the simultaneous regeneration of cartilage and subchondral bone. These findings indicate that scaffolds with vertically aligned ECM-like architectures serve as an effective cell-free and growth factor-free scaffold for enhanced endogenous osteochondral regeneration.

Elastomeric Electrospun Scaffolds of a Biodegradable Aliphatic Copolyester Containing PEG-Like Sequences for Dynamic Culture of Human Endothelial Cells.

In the field of artificial prostheses for damaged vessel replacement, polymeric scaffolds showing the right combination of mechanical performance, biocompatibility, and biodegradability are still demanded. In the present work, poly(butylene-co-triethylene trans-1,4-cyclohexanedicarboxylate), a biodegradable random aliphatic copolyester, has been synthesized and electrospun in form of aligned and random fibers properly designed for vascular applications. The obtained materials were analyzed through tensile and dynamic-mechanical tests, the latter performed under conditions simulating the mechanical contraction of vascular tissue. Furthermore, the in vitro biological characterization, in terms of hemocompatibility and cytocompatibility in static and dynamic conditions, was also carried out. The mechanical properties of the investigated scaffolds fit within the range of physiological properties for medium- and small-caliber blood vessels, and the aligned scaffolds displayed a strain-stiffening behavior typical of the blood vessels. Furthermore, all the produced scaffolds showed constant storage and loss moduli in the investigated timeframe (24 h), demonstrating the stability of the scaffolds under the applied conditions of mechanical deformation. The biological characterization highlighted that the mats showed high hemocompatibility and low probability of thrombus formation; finally, the cytocompatibility tests demonstrated that cyclic stretch of electrospun fibers increased endothelial cell activity and proliferation, in particular on aligned scaffolds.

Co-culture methods to study neuronal function and disease.

Co-culture methods to study neuronal function and disease.

Tanshinone IIA-loaded aligned microfibers facilitate stem cell recruitment and capillary formation by inducing M2 macrophage polarization

Direct implantation of cell-free scaffolds capable of promoting tissue regeneration by manipulating immune responses has proven to be a promising therapeutic strategy for regenerative medicine. Here, we developed aligned microfiber scaffolds with sustained release of tanshinone ⅡA (Tan ⅡA) to modulate macrophages phenotypic transition, which subsequently promoted stem cell recruitment and capillary formation. Aligned microfibers scaffolds loaded with 1μM Tan ⅡA (AF-1) significantly down-regulated the expression of proinflammatory genes and proteins, while they upregulated anti-inflammatory genes and proteins, in RAW 264.7 macrophages. Conditioned medium collected from macrophages cultured on AF-1 scaffolds enhanced bone marrow-derived mesenchymal stem cell (BMSC) proliferation and migration, and also regulated their multiple biological functions as evidenced by RNA-Seq assays. Moreover, the conditioned medium also promoted human umbilical vein endothelial cell (HUVEC) proliferation, migration, and tube formation. Enhancement of endogenous stem cell recruitment and vascularization by regulating macrophage phenotype transition was further confirmed by utilizing rat subcutaneous implantation of the scaffolds. These results support the use of drug-loaded aligned microfiber scaffolds to enable immune modulation to stimulate stem cell recruitment and vascularization, which could potentially result in successful cell-free, scaffold-guided tissue regeneration.

Regeneration of the peripheral nerve via multifunctional electrospun scaffolds.

Over the last two decades, electrospun scaffolds have proved to be advantageous in the field of nerve tissue regeneration by connecting the cavity among the proximal and distal nerve stumps growth cones and leading to functional recovery after injury. Multifunctional nanofibrous structure of these scaffolds provides enormous potential by combining the advantages of nano-scale topography, and biological science. In these structures, selecting the appropriate materials, designing an optimized structure, modifying the surface to enhance biological functions and neurotrophic factors loading, and native cell-like stem cells should be considered as the essential factors. In this systematic review paper, the fabrication methods for the preparation of aligned nanofibrous scaffolds in yarn or conduit architecture are reviewed. Subsequently, the utilized polymeric materials, including natural, synthetic and blend are presented. Finally, their surface modification techniques, as well as, the recent advances and outcomes of the scaffolds, both in vitro and in vivo, are reviewed and discussed.

Electrospun Aligned Coaxial Nanofibrous Scaffold for Cardiac Repair.

Cardiovascular diseases (CVDs) are one of the leading causes of mortality worldwide and a number one killer in the USA. Cell-based approaches to treat CVDs have only shown modest improvement due to poor survival, retention, and engraftment of the transplanted cells in the ischemic myocardium. Recently, tissue engineering and the use of 3D scaffolds for culturing and delivering stem cells for ischemic heart disease are gaining rapid potential. Here, we describe a protocol for the fabrication of aligned coaxial nanofibrous scaffold comprising of a polycaprolactone (PCL) core and gelatin shell. Furthermore, we describe a detailed protocol for the efficient seeding and maintenance of human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) on these nanofibrous scaffolds, which could have a potential application in the generation of functional "cardiac patch" for myocardial repair applications as well as an in vitro 3D cardiac tissue model to evaluate the efficacy of cardiovascular drugs and cardiac toxicities.

Anisotropically Conductive Biodegradable Scaffold with Coaxially Aligned Carbon Nanotubes for Directional Regeneration of Peripheral Nerves.

Fascicular rearrangement of an injured peripheral nerve requires reconnection of nerve sprouts from anterior and Büngner bands from distal sides of the lesion, failing to which leads to inefficient regeneration of the injured nerve. However, existing neural scaffolds have limited neuroregeneration efficiency because of either the lack of alignment of fibers and a conductive second phase, leading to compromised electrical conductivity, or the lack of extracellular matrix components and in vivo validation. The present study reports a biocompatible, multiwall carbon nanotube (MWCNT)-reinforced, anisotropically conductive, electrospun, aligned nanofibrous scaffold, ensuring maximal peripheral nerve regeneration. Electrospinning parameters were modulated to deposit random and parallel fibers in separate scaffolds for comparative analysis on the effect of fiber alignment on regeneration. Both types of scaffolds were reinforced with MWCNTs to impart electrical conductivity. Nonreinforced scaffolds were nonconductive. In this comparative study, MWCNT-reinforced, aligned scaffolds showed better tensile property with increased conductivity along the direction of alignment, thereby ensuring an escalated neural-regeneration rate. Collectively, in vitro studies established the scaffolds to be highly biocompatible, promoting cell growth and proliferation. With 85% more anisotropic conductivity in the direction of the alignment and the degradation kinetics tuned to the regeneration regime, the MWCNT-reinforced, aligned scaffold efficiently healed injured sciatic nerves in rats within 30 days. Rigorous revivification of the tissue was due to coordinated Wallerian degeneration and expedited guided axonal regeneration. Structural and functional analysis of nerves in vivo showed the aligned, MWCNT-reinforced scaffold to be very efficient in peripheral sciatic nerve regeneration. This study notes the efficacy of the coaxially aligned, MWCNT-reinforced neural scaffold, with a capability of establishing remarkable advancement in the field of peripheral neural regeneration.

In vivo biocompatibility study of degradable homo- versus multiblock copolymers and their (micro)structure compared to an established biomaterial.

Copolyetheresterurethane (PDC) is a biodegradable, shape-memory biomaterial, which has been shown to be of low toxicity and pro-angiogenic in vitro. In the present study we examined the in vivo compatibility of PDC as a compression molded film and as electrospun scaffolds and its well established constituent, the homopolymer poly(p-dioxanone) (PPDO), which were compared with the clinically used poly[(vinylidene fluoride)-co-hexafluoropropene] (PVDF) as reference material. The materials were implanted in the subcutaneous tissue of mice and the host responses were analyzed histologically 7 and 28 days after implantation.All materials induced a foreign body response (FRB) including the induction of foreign body giant cells and a peripheral fibrous capsule. PDC, PPDO and PVDF films showed no signs of degradation after 28 days. PDC films showed a significantly reduced associated macrophage layer and fibrous capsule on their surface. Few fragments of PDC and PPDO scaffolds were present at the implantation site, while PVDF scaffolds were still present in large amounts at day 28. Especially aligned electrospun PDC scaffold induced a significantly thinner fibrous and a slightly reduced inflammatory response after 28 days of implantation. In addition, only PDC aligned fibrous scaffold structures induced a significant increase in angiogenesis.In summary, PDC films outperformed PPDO and PVDF films in terms of compatibility, especially in capsule and macrophage layer thickness. Through microstructuring of PDC and PPDO into scaffolds an almost complete degradation was observed after 28 days, while their respective films remained almost unchanged. However, the capsule thickness of all scaffolds was comparable to the films after 28 days. Finally, the parallel arrangement of PDC fibers enabled a strong enhancement of angiogenesis within the scaffold. Hence, material chemistries influence overall compatibility in vivo, while angiogenesis could be influenced more strongly by microstructural parameters than chemical ones.

Electrospinning of Eudragit RS100 for Nerve Tissue Engineering Scaffold

Electrospinning technique is widely investigated in medical applications such as tissue engineering scaffolds, wound dressing and drug delivery. In this study, the aligned nanofiber scaffold of Eudragit RS100 was successfully fabricated via electrospinning technique for nerve tissue engineering scaffold. The diameter distribution and degree of alignment of Eudragit RS100 nanofiber scaffold were observed by scanning electron microspore (SEM). The chemical and crystalline structure of Eudragit RS100 nanofiber scaffold were analyzed using Fourier transform infrared spectroscopy (FTIR) and Powder X-ray diffactometer (PXRD). Cell culture studies using rat Schwann cells were determined to evaluate cell proliferation cell alignment and morphology. The results implied that the diameter of fiber was in the nanometer region. The Eudragit RS100 nanofiber scaffold were in an amorphous form and its chemical structure was not destructive after the electrospinning process. The Eudragit RS100 nanofiber scaffold showed biocompatibility with rat Schwann cells and growing parallel to the aligned fibers. In conclusion, the Eudragit RS100 nanofiber scaffold may have the ability to apply to nerve tissue engineering scaffold.

Delivery of Human Stromal Vascular Fraction Cells on Nanofibrillar Scaffolds for Treatment of Peripheral Arterial Disease.

Cell therapy for treatment of peripheral arterial disease (PAD) is a promising approach but is limited by poor cell survival when cells are delivered using saline. The objective of this study was to examine the feasibility of aligned nanofibrillar scaffolds as a vehicle for the delivery of human stromal vascular fraction (SVF), and then to assess the efficacy of the cell-seeded scaffolds in a murine model of PAD. Flow cytometric analysis was performed to characterize the phenotype of SVF cells from freshly isolated lipoaspirate, as well as after attachment onto aligned nanofibrillar scaffolds. Flow cytometry results demonstrated that the SVF consisted of 33.1 ± 9.6% CD45+ cells, a small fraction of CD45–/CD31+ (4.5 ± 3.1%) and 45.4 ± 20.0% of CD45–/CD31–/CD34+ cells. Although the subpopulations of SVF did not change significantly after attachment to the aligned nanofibrillar scaffolds, protein secretion of vascular endothelial growth factor (VEGF) significantly increased by six-fold, compared to SVF cultured in suspension. Importantly, when SVF-seeded scaffolds were transplanted into immunodeficient mice with induced hindlimb ischemia, the cell-seeded scaffolds induced a significant higher mean perfusion ratio after 14 days, compared to cells delivered using saline. Together, these results show that aligned nanofibrillar scaffolds promoted cellular attachment, enhanced the secretion of VEGF from attached SVF cells, and their implantation with attached SVF cells stimulated blood perfusion recovery. These findings have important therapeutic implications for the treatment of PAD using SVF.

Engineered dual-scale poly (ε-caprolactone) scaffolds using 3D printing and rotational electrospinning for bone tissue regeneration

Large bone defects due to trauma or disease present a significant clinical challenge with limited efficacy of current therapies. A key aim is to develop biomimetic scaffolds that reflect the native tissue structure with 3D printing being an important enabling technology. However, the incorporation of multiple length scales and anisotropic features, mimicking the native architecture, is difficult with current processes. In this study, we propose a simple and versatile hybrid printing process using a screw-assisted additive manufacturing technique combined with rotational electrospinning to fabricate dual-scale anisotropic scaffolds. 3D microscale porous polycaprolactone (PCL) structures with highly aligned nanoscale fibres were successfully produced layer-by-layer. The scaffolds were morphological, mechanical and biological characterised. Human adipose-derived stem cells (hADSCs) were seeded on the hybrid scaffold to evaluate the effects of structural and anisotropic topographic cues on cell attachment, proliferation and osteogenesis differentiation. Results show that the 3D printed microscale structures have uniform and well-defined geometries and the alignment of nanoscale electrospun fibres increases by increasing the electrospinning rotational velocity. Mechanical results show that there is no significant difference between 3D printed scaffolds with or without electrospun meshes. In vitro results show higher cell seeding efficiency and proliferation in dual-scale scaffolds with high density electrospun meshes. A more stretched and elongated cell morphology was observed in aligned nanofibre scaffolds showing higher anisotropic cytoskeletal organization than 3D printed PCL scaffolds without electrospun meshes. The dual-scale scaffolds present improved overall osteogenic markers expressions (COL-1, ALP and OCN). However, no statistical difference between normalised osteogenic marker expressions were observed between dual-scale scaffolds and 3D printed scaffolds. This might be attributed to the poor bioactivity of the substrate material, PCL, suggesting topographical cues might not be sufficient to stimulate cell fate towards to an osteogenic linage. The results suggest that the proposed fabrication strategy is a promising approach for the design of novel bone scaffolds to modulate cell fates by integrating the topographic cue reported in this paper with biochemical cues associated to the use of more bioactive materials.

Remote triggering of TGF-β/Smad2/3 signaling in human adipose stem cells laden on magnetic scaffolds synergistically promotes tenogenic commitment

Injuries affecting load bearing tendon tissues are a significant clinical burden and efficient treatments are still unmet. Tackling tendon regeneration, tissue engineering strategies aim to develop functional substitutes that recreate native tendon milieu. Tendon mimetic scaffolds capable of remote magnetic responsiveness and functionalized magnetic nanoparticles (MNPs) targeting cellular mechanosensitive receptors are potential instructive tools to mediate mechanotransduction in guiding tenogenic responses. In this work, we combine magnetically responsive scaffolds and targeted Activin A type II receptor in human adipose stem cells (hASCs), under alternating magnetic field (AMF), to synergistically facilitate external control over signal transduction. The combination of remote triggering TGF-β/Smad2/3 using MNPs tagged hASCs, through magnetically actuated scaffolds, stimulates overall expression of tendon related genes and the deposition of tendon related proteins, in comparison to non-stimulated conditions. Moreover, the phosphorylation of Smad2/3 proteins and their nuclear co-localization was also more evident. Overall, biophysical stimuli resulting from magnetic scaffolds and magnetically triggered cells under AMF stimulation modulate the mechanosensing response of hASCs towards tenogenesis, holding therapeutic promise. Statement of SignificanceThe concept of magnetically-assisted tissue engineering may assist the development of innovative solutions to treat tendon disorders upon remote control of biological processes as cell migration or differentiation. Herein, we originally combine a fibrous aligned superparamagnetic scaffold, based on a biodegradable polymeric blend of starch and poly-ɛ-caprolactone incorporating magnetic nanoparticles (MNPs), and human adipose stem cells (hASCs) labelled with MNPs functionalized with anti-activin receptor type IIA (ActRIIA). Constructs were stimulated using alternating magnetic field (AMF), to activate the ActRIIA and subsequent induction of TGF-β signaling, through Smad2/3 phosphorylation cascade, enhancing the expression of tendon-related markers. Altogether, these findings contribute with powerful bio-magnetic approaches to activate key tenogenic pathways, envisioning future translation of magnetic biomaterials into regenerative platforms for tendon repair.

Tissue Ingrowth Markedly Reduces Mechanical Anisotropy and Stiffness in Fibre Direction of Highly Aligned Electrospun Polyurethane Scaffolds.

The lack of long-term patency of synthetic vascular grafts currently available on the market has directed research towards improving the performance of small diameter grafts. Improved radial compliance matching and tissue ingrowth into the graft scaffold are amongst the main goals for an ideal vascular graft. Biostable polyurethane scaffolds were manufactured by electrospinning and implanted in subcutaneous and circulatory positions in the rat for 7, 14 and 28 days. Scaffold morphology, tissue ingrowth, and mechanical properties of the scaffolds were assessed before implantation and after retrieval. Tissue ingrowth after 24 days was 96.5 ± 2.3% in the subcutaneous implants and 77.8 ± 5.4% in the circulatory implants. Over the 24 days implantation, the elastic modulus at 12% strain decreased by 59% in direction of the fibre alignment whereas it increased by 1379% transverse to the fibre alignment of the highly aligned scaffold of the subcutaneous implants. The lesser aligned scaffold of the circulatory graft implants exhibited an increase of the elastic modulus at 12% strain by 77% in circumferential direction. Based on the observations, it is proposed that the mechanism underlying the softening of the highly aligned scaffold in the predominant fibre direction is associated with scaffold compaction and local displacement of fibres by the newly formed tissue. The stiffening of the scaffold, observed transverse to highly aligned fibres and for more a random fibre distribution, represents the actual mechanical contribution of the tissue that developed in the scaffold.

Combining Electrospinning and Electrospraying to Prepare a Biomimetic Neural Scaffold with Synergistic Cues of Topography and Electrotransduction.

The nerve tissue consists of aligned fibrous nerve bundles, in which neurons communicate and transmit information through electrical signals. Hence, biocompatibility, oriented fibrous structure, and electrical conductivity are key factors for the biomimetic design of nerve scaffolds. Herein, we built a technical platform to combine electrospinning and electrospraying for preparing a biomimetic scaffold with conductivity and aligned fibrous structure. The highly aligned polycaprolactone (PCL) microfibrous scaffolds with co-sprayed collagen and conductive polypyrrole nanoparticles (PPy NPs) showed good bioactivity, supplying a platform for exploring the effects of topographical guidance, fiber conductivity, and its mediated external electrical signals on neurogenesis. The results revealed that collagen-coated highly aligned PCL microfibrous scaffold induced PC12 cells oriented and elongated along the direction of fibers. In addition, the improved conductivity of PPy-coated aligned fibers and its mediated external electrical stimulation collectively contributed to the functional expression, including elongation, gene expression, and protein expression, of PC12 cells. We further demonstrated the potential mechanism where the fiber conductivity and its mediated external electrical signals resulted in the upregulation of voltage-gated calcium channel, leading to the influx of Ca2+, thereby activating intracellular signaling cascades, ultimately enhancing neurogenesis. This approach provides a strategy to design aligned fibrillary scaffolds with bioactive adhesion domains and electroconductivity for neural regeneration.

Pre-innervated tissue-engineered muscle promotes a pro-regenerative microenvironment following volumetric muscle loss

Volumetric muscle loss (VML) is the traumatic or surgical loss of skeletal muscle beyond the inherent regenerative capacity of the body, generally leading to severe functional deficit. Formation of appropriate somato-motor innervations remains one of the biggest challenges for both autologous grafts as well as tissue-engineered muscle constructs. We aim to address this challenge by developing pre-innervated tissue-engineered muscle comprised of long aligned networks of spinal motor neurons and skeletal myocytes on aligned nanofibrous scaffolds. Motor neurons led to enhanced differentiation and maturation of skeletal myocytes in vitro. These pre-innervated tissue-engineered muscle constructs when implanted in a rat VML model significantly increased satellite cell density, neuromuscular junction maintenance, graft revascularization, and muscle volume over three weeks as compared to myocyte-only constructs and nanofiber scaffolds alone. These pro-regenerative effects may enhance functional neuromuscular regeneration following VML, thereby improving the levels of functional recovery following these devastating injuries.

Aligned graphene/silk fibroin conductive fibrous scaffolds for guiding neurite outgrowth in rat spinal cord neurons.

Graphene, as a highly conducting material, incorporated into silk fibroin (SF) substrates is promising to fabricate an electroactive flexible scaffold toward neural tissue engineering. It is well known that aligned morphology could promote cell adhesion and directional growth. The purpose of this study was to develop aligned conductive scaffolds made of graphene and SF (G/SF) by electrospinning technique for neural tissue engineering applications. The physicochemical characterization of scaffolds revealed that the mechanical and electrochemical property of aligned G/SF scaffolds continually raised with the increasing contents of graphene (A0% G/SF, A1% G/SF, A2% G/SF, and A3% G/SF), but the mechanical property descended when the graphene concentration reached to 4% (the A4% G/SF group). The results of the cell experiment in vitro indicated that all the aligned G/SF scaffolds were no neurotoxic to primary cultured spinal cord neurons. In addition, the neurite elongation in all aligned groups was significantly enhanced by the upregulation of Netrin-1 expression compared to them in the control group. Thus, A3% G/SF scaffolds not only possessed the optimal property based on the mechanical and electrochemical performances but also displayed the beneficial capability to neurite outgrowth, which might perform a suitable candidate to successfully scaffold electrically active tissues during neural regeneration or engineering.

Corneal stromal regeneration by hybrid oriented poly (ε-caprolactone)/lyophilized silk fibroin electrospun scaffold

Applying biological macromolecule like silk fibroin (SF) is a promising material for corneal tissue engineering. However, designing an appropriate tissue-like construct to compensate the shortages of traditional routes are still challenging. SF besides possessing biocompatibility and transparency, the biomaterial should be mechanically strong. In the present study, a hybrid scaffold composed of poly-ε-caprolactone (PCL)-silk fibroin (SF) is fabricated through electro spinning technique. The aligned and non-aligned PCL-SF scaffolds with various weight ratios are fabricated. The results reveal that the addition of SF yields the scaffolds with more uniform and aligned structure. The ultimate tensile strength and Young's modulus of aligned and non-aligned PCL-SF (60:40 and 50:50) fibers are in an acceptable range for cornea applications. It is noteworthy that the aligned PCL-SF (60:40 and 50:50) scaffolds have more transparency, hydrophilicity, water uptake, and in vitro degradation rate than the other scaffolds. The cell compatibility results show that human stromal keratocyte cells are attached and proliferated on the aligned and non-aligned PCL-SF scaffolds. The overall results recommend that PCL-SF (60:40 and 50:50) scaffolds have a great potential for human corneal stromal regeneration.

Beneficial effects of biodelivery of brain-derived neurotrophic factor and gold nanoparticles from functionalized electrospun PLGA scaffold for nerve tissue engineering

Despite of the pivotal role of Schwann cells (SCs) in peripheral nerve regeneration, lack of an available source has prompted researches for Schwann-like cells transdifferentiation. This study suggests an effective method for tuning the surface of aligned poly (lactic-co-glycolic acid) (PLGA) nanofibers to enhance attachment and proliferation of h-ADSCs on the scaffold through coating of laminin. Following the characterization of biofunctionalized PLGA, brain-derived neurotropic factor (BDNF) and gold nanoparticles (AuNPs) were encapsulated in chitosan nanoparticles (CSNPs), added into laminin solution, and coated on the surface of aligned PLGA scaffold. The release behavior of BDNF and AuNPs from scaffold was evaluated by Bradford assay and inductive coupled plasma optical emission spectrometry (ICP-OES) technique, respectively. Afterwards, experimental groups were investigated for potential of Schwann cell differentiation using immunocytochemical staining and real-time RT-PCR technique. Results of MTT assay showed the significantly higher proliferation of h-ADSCs on laminin-functionalized scaffold compared to PLGA scaffold (p < 0.05). Additionally, the presence of BDNF and AuNPs on scaffold significantly improved the expression of SCs markers as compared to the control group (p < 0.05). Therefore, use of biofunctionalized PLGA nanofibers can be a promising strategy for inducing the differentiation of h-ADSCs into SCs for nerve tissue engineering.

Remodeling of aligned fibrous extracellular matrix by encapsulated cells under mechanical stretching

Extracellular matrix (ECM) remodeling is essential for the development and functions of connective tissues (e.g., heart, muscle and the periodontal ligament), and entails the highly anisotropic response of cells and their organized ECM molecules to mechanical stimulation. However, the nature of how cells remodel their surrounding ECM under mechanical stimulation remains elusive. Here, we encapsulated human periodontal ligament stem cells (hPDLSCs) within an aligned rat collagen scaffold labeled with fluorescein isothiocyanate (FITC) and applied mechanical stimulation on the scaffold using magnetic stretching. Through tracking the FITC-labeled rat collagen scaffold and the newly secreted human type I collagen, we studied the effect of magnetic stretching on the mechanism of aligned ECM remodeling by the encapsulated cells. We found that the aligned topography combined with magnetic stretching could significantly promote initial ECM degradation and new ECM secretion: expression of matrix metalloproteinases 1 and 9 is increased markedly, and the elastic modulus of the stretched scaffold (75 kPa) is significantly higher than that of the random scaffold (50 kPa). The data support a model whereby the cells remodel their surrounding ECM under continuous stretching through degradation and then secretion of new ECM to integrate with the aligned ECM and maintain tissue function. Our study offers a valuable basis for future optimized design of biomaterial scaffolds for clinical translation. Statement of significanceExtracellular matrix (ECM) remodeling is essential for the development and functions of connective tissues. However, the nature of how cells remodel their surrounding aligned ECM under mechanical stimulation remains elusive. Herein, we developed a method to reveal the remodeling of aligned rat collagen scaffold by the encapsulated human periodontal ligament stem cells (hPDLSCs) using fluorescence imaging. We found that the aligned topography combined with magnetic stretching could significantly promote initial ECM degradation and new ECM secretion: the expression of matrix metalloproteinase 1 and 9 are significantly higher, and the elastic modulus increases from 50 kPa to 75 kPa as compared to the random collagen scaffold encapsulating hPDLSCs. Our study holds great potential in optimization of bio-scaffold design for clinical translation.

Efficient mesenchymal stem cell conversion to corneal keratocytes by the use of specific biomaterial and Cell Rev™ mSC diffKera media

Background & Aim Corneal blindness is a prevalent cause of vision loss, affecting approximately 4.9 million people worldwide (Flaxman et al, 2017). The demand for donor replacement corneas often exceeds the supply, requiring alternative treatment options. To achieve corneal function while minimizing the risks of donor rejection, we propose an autologous biomimetic cornea whereby patient-derived mesenchymal stem cells (MSCs) are converted to corneal-like cells on scaffolds that closely resemble the structure of the human cornea. We hypothesize that the effectiveness of cell differentiation depends on the biomaterials used and the cultured media composition. The more closely the environment resembles that of the human cornea, the higher the likelihood that a greater proportion of MSCs will differentiate into corneal keratocytes. Methods, Results & Conclusion Here, we investigated MSC differentiation into corneal keratocytes using various medium formulations on our electrospun scaffolds. Polycaprolactone-bovine dermis type I atelocollagen (PCL-atelocollagen) and polycaprolactone (PCL) solutions were electrospun to obtain the aligned nanofibrous scaffolds. MSCs were cultured on the scaffolds for 12 days in 3 different types of media: the expansion media described by Lynch & Ahearne (2007); keratocyte differentiation media used in the same study; and our Cell Rev™ mSC diffKera media. Immunofluorescence staining for keratocyte markers such as aldehyde dehydrogenase 1 and lumican was conducted on cells fixed on the scaffolds at days 6 and 12. The results confirmed that a greater proportion of MSCs differentiated into keratocytes in Cell Rev™ mSC diffKera media compared with the media used by Lynch & Ahearne (Figure 1). In addition, electrospun PCL-atelocollagen scaffolds are likely to support keratocyte differentiation better than electrospun PCL scaffolds (control) because corneal stromal collagen fibrils comprise mainly type I collagen. These findings demonstrate that the use of our electrospun PCL-atelocollagen scaffolds and Cell Rev™ mSC diffKera formulation effectively enhances MSC differentiation into keratocytes, and represents a novel step for the creation of artificial corneas. Corneal blindness is a prevalent cause of vision loss, affecting approximately 4.9 million people worldwide (Flaxman et al, 2017). The demand for donor replacement corneas often exceeds the supply, requiring alternative treatment options. To achieve corneal function while minimizing the risks of donor rejection, we propose an autologous biomimetic cornea whereby patient-derived mesenchymal stem cells (MSCs) are converted to corneal-like cells on scaffolds that closely resemble the structure of the human cornea. We hypothesize that the effectiveness of cell differentiation depends on the biomaterials used and the cultured media composition. The more closely the environment resembles that of the human cornea, the higher the likelihood that a greater proportion of MSCs will differentiate into corneal keratocytes. Here, we investigated MSC differentiation into corneal keratocytes using various medium formulations on our electrospun scaffolds. Polycaprolactone-bovine dermis type I atelocollagen (PCL-atelocollagen) and polycaprolactone (PCL) solutions were electrospun to obtain the aligned nanofibrous scaffolds. MSCs were cultured on the scaffolds for 12 days in 3 different types of media: the expansion media described by Lynch & Ahearne (2007); keratocyte differentiation media used in the same study; and our Cell Rev™ mSC diffKera media. Immunofluorescence staining for keratocyte markers such as aldehyde dehydrogenase 1 and lumican was conducted on cells fixed on the scaffolds at days 6 and 12. The results confirmed that a greater proportion of MSCs differentiated into keratocytes in Cell Rev™ mSC diffKera media compared with the media used by Lynch & Ahearne (Figure 1). In addition, electrospun PCL-atelocollagen scaffolds are likely to support keratocyte differentiation better than electrospun PCL scaffolds (control) because corneal stromal collagen fibrils comprise mainly type I collagen. These findings demonstrate that the use of our electrospun PCL-atelocollagen scaffolds and Cell Rev™ mSC diffKera formulation effectively enhances MSC differentiation into keratocytes, and represents a novel step for the creation of artificial corneas.