Aligned Matrices Research Articles

Highly Aligned Ternary Nanofiber Matrices Loaded with MXene Expedite Regeneration of Volumetric Muscle Loss

HighlightsThe aligned ternary nanofibrous matrices composed of poly(lactide-co-ε-caprolactone), collagen, and Ti3C2Tx MXene nanoparticles were fabricated (referred as PCM matrices).The aligned PCM matrices exhibited favorable physicochemical properties and excellent cytocompatibility and myogenic properties, which in turn promoted fast regeneration of volumetric muscle loss in vivo.The Ca2+ binding of MXene nanoparticles activated inducible nitric oxide synthase and serum/glucocorticoid regulated kinase 1-mediated mTOR-AKT pathway to promote myoblast differentiation and maturation.

Structural and functional connectivity reconstruction with CATO - A Connectivity Analysis TOolbox

We describe a Connectivity Analysis TOolbox (CATO) for the reconstruction of structural and functional brain connectivity based on diffusion weighted imaging and resting-state functional MRI data. CATO is a multimodal software package that enables researchers to run end-to-end reconstructions from MRI data to structural and functional connectome maps, customize their analyses and utilize various software packages to preprocess data. Structural and functional connectome maps can be reconstructed with respect to user-defined (sub)cortical atlases providing aligned connectivity matrices for integrative multimodal analyses. We outline the implementation and usage of the structural and functional processing pipelines in CATO. Performance was calibrated with respect to simulated diffusion weighted imaging data from the ITC2015 challenge and test-retest diffusion weighted imaging data and resting-state functional MRI data from the Human Connectome Project. CATO is open-source software distributed under the MIT License and available as a MATLAB toolbox and as a stand-alone application at www.dutchconnectomelab.nl/CATO.

Abstract 15800: Dissecting Cardiomyocyte-Fibroblast Communication During Homeostasis and Cardiac Fibrosis in a Compartmentalized Biomimetic Tissue Model

Introduction: Cardiac fibroblasts (CFs) maintain cardiac structure and function by synthesizing and remodeling the extracellular matrix (ECM) found throughout the heart. Homeostatic maintenance of the ECM required for healthy tissue function depends on CF communication with cardiomyocytes (CMs). During cardiac fibrosis, signals resulting from tissue injury and dysregulated CF-CM communication promote CF differentiation into α-smooth muscle actin + (αSMA) myofibroblasts (MFs), which deposit excess ECM that mechanically impairs heart function. Given the challenges involved in parsing the dynamic and multifactorial heterocellular interactions underlying tissue homeostasis versus fibrosis, we engineered a compartmentalized, bilayer cardiac tissues where iPSC-derived CMs and CFs can be cultured on either side of a shared fibrous ECM to study CM-CF communication. Methods: Dextran vinyl sulfone (DVS) fibers were electrospun onto arrays of microfabricated PDMS wells. Matrix stiffness and alignment were controlled by modulating the speed of the collecting surface and UV light exposure, respectively. iPSC-CMs and primary human CFs were seeded onto opposing sides of the matrix. Immunostaining for αSMA was conducted to assess CF differentiation to MFs. Results: In CF monocultures on matrices varying in alignment and stiffness, MF differentiation was greatest on stiff (17.3 kPa) aligned matrices with exogenous TGFβ1 compared to soft (0.68 kPa) aligned matrices and both soft/stiff random matrices. We also observed increased nuclear YAP expression on stiff/aligned matrices, indicating altered mechanosensing in response to matrix properties. However, the presence of co-cultured CMs inhibited MF differentiation despite exogenous TGFβ1. MF differentiation was also impaired in CFs cultured on glass below matrix-seeded CMs, implying the role of protective paracrine signaling from CMs to CFs. Conclusions: We developed a microfabricated biomimetic coculture platform to examine CM-CF communication driving tissue homeostasis versus fibrosis. Specifically, we find that fibrous mechanical cues modulate YAP-mediated signaling and αSMA expression. Further, CM-CF paracrine signaling mitigates MF differentiation on aligned fibrous matrices.

Abstract 15136: Matrix Mechanics Regulate Engineered Myocardial Microtissue Organization and Contractility

Introduction: The mechanical function of the myocardium is dictated by cardiomyocytes (CMs) and the surrounding fibrous extracellular matrix (ECM). Due to limitations in existing approaches for engineering myocardial tissue, how CMs sense and respond to mechanical microenvironmental changes remains understudied. Here, we established a method for creating arrays of cardiac microtissues with highly tunable mechanical features by integrating synthetic, fibrous matrices and iPSC-CMs. Through real-time contractility readouts enabled, we explored how biomechanical cues influence engineered cardiac tissue assembly and function. Methods: Arrayed cardiac microtissues were generated by selectively photo-crosslinking electrospun dextran vinyl sulfone fiber matrices onto pairs of microfabricated PDMS cantilevers. Subsequently, iPSC-CMs were patterned onto matrices using a microfabricated seeding mask. Matrix and cantilever stiffnesses were tuned by adjusting photoinitiator concentrations and cantilever geometry, respectively. Results: In tissues contracting against soft cantilevers (0.41 N/m), tissue contractile stress decreased with increasing matrix stiffness (E = 0.68 to 17.3 kPa). We also observed an increase in contractile stress on soft and aligned matrices, compared to non-aligned matrices. However, tissues contracting against stiffer cantilevers (1.2 N/m) exerted similar contractile stresses independent of matrix alignment. This may be explained by the enhanced alignment of myofibrils in non-aligned matrices contracting against stiffer posts. The size of vinculin-rich cell-ECM adhesion decreased on stiff matrices. Vinculin also localized to z-discs forming organized costameres on soft matrices. Tissue response to mavacamten suggests that β-cardiac myosins are required for costamere formation and myofibril maturation. Conclusions: We developed a new microfabrication strategy to create tunable cardiac microtissues and examined how biomechanical cues affect tissue assembly and function. We found that matrix stiffness, matrix alignment, and tissue constraint distinctly affect iPSC-CM function and structure, potentially due to differential influences on costamere formation and myofibrillar assembly.

Microengineering 3D Collagen Hydrogels with Long-Range Fiber Alignment.

Aligned collagen I (COL1) fibers guide tumor cell motility, influence endothelial cell morphology, control stem cell differentiation, and are a hallmark of cardiac and musculoskeletal tissues. To study cell response to aligned microenvironments in vitro, several protocols have been developed to generate COL1 matrices with defined fiber alignment, including magnetic, mechanical, cell-based, and microfluidic methods. Of these, microfluidic approaches offer advanced capabilities such as accurate control over fluid flows and the cellular microenvironment. However, the microfluidic approaches to generate aligned COL1 matrices for advanced in vitro culture platforms have been limited to thin "mats" (<40 µm in thickness) of COL1 fibers that extend over distances less than 500 µm and are not conducive to 3D cell culture applications. Here, we present a protocol to fabricate 3D COL1 matrices (130-250 µm in thickness) with millimeter-scale regions of defined fiber alignment in a microfluidic device. This platform provides advanced cell culture capabilities to model structured tissue microenvironments by providing direct access to the micro-engineered matrix for cell culture.

Membrane supported poly(butylene adipate-co-terephthalate) nanofibrous matrices as cardiac patch: Effect of basement membrane for the fiber deposition and cellular behavior

Electrospun nanofibrous matrices are convenient biomaterials that mimic extracellular matrices for adhesion, migration, proliferation, and differentiation of cells. The aim of this study is to optimize the electrospinning parameters for poly(butylene adipate-co-terephthalate) (PBAT) nanofiber production by the response surface methodology (RSM) and to develop a suitable material design for the usability of PBAT in cardiac tissue engineering. Therefore, electrospun PBAT nanofibrous matrices collected on solvent-casted polycaprolactone (PCL) or PBAT basement membranes at optimized conditions. The attachment and proliferation behavior of the H9C2 rat cardiomyoblasts investigated on different PBAT and PCL surface features as a model cell line. For this purpose, neat PBAT and PCL films have been used comparatively with both random (R-PBAT) and aligned PBAT (A-PBAT) nanofibers coated films. The effect of polymer concentration, flow rate, applied voltage, and needle tip-connector distance on fiber diameter and alignment was examined in the electrospinning process and optimum processing parameters were determined by RSM. The PBAT nanofibers were spun on basement membranes with 10% (w/v) polymer concentration, 1 mL/h volumetric flow rate, 2000 rpm collector rotation velocity (for aligned ones), 15 kV applied voltage, and 20 cm needle tip-collector distance. The average diameter of random (R-PBAT) and aligned (A-PBAT) nanofibers was calculated as 555 ± 126 nm and 417 ± 137 nm. The mechanical test results showed that the alignment topography increased the elastic modulus of PBAT nanofibers compared to random matrices. The alignment of fibers found as 91% and 75% within the ± 10° range for A-PBAT/SC-PBAT and A-PBAT/SC-PCL, respectively. These findings showed that usage of PCL, instead of PBAT, as basement membrane decrease the alignment of deposited nanofibers. A 7-day cell culture study conducted with H9C2 cells seeded samples to investigate the influence of these differences on cell behavior. The results indicated that the alignment of fibers provides a suitable topography to proliferate and spread in myocyte morphology for H9C2 cells especially compared to neat films. Cellular behavior and nanofiber deposition have been affected by the usage of various basement membrane polymers. These findings demonstrated that the usage of basement membrane as support material provides the required thickness and mechanical properties to the aligned PBAT nanofiber matrices, and this double layer structure might be a promising candidate for cardiac tissue engineering with further studies.

Dynamic Heterochromatin States in Anisotropic Nuclei of Cells on Aligned Nanofibers.

The cancer cell nucleus deforms as it invades the interstitial spaces in tissues and the tumor microenvironment. While alteration of the chromatin structure in a deformed nucleus is expected and documented, the chromatin structure in the nuclei of cells on aligned matrices has not been elucidated. In this work we elucidate the spatiotemporal organization of heterochromatin in the elongated nuclei of cells on aligned nanofibers with stimulated emission depletion nanoscopy and fluorescence correlation spectroscopy. We show that the anisotropy of nuclei is sufficient to drive H3K9me3-heterochromatin alterations, with enhanced H3K9me3 nanocluster compaction and aggregation states that otherwise are indistinguishable from diffraction-limited microscopy. We interrogated the higher-order heterochromatin structures within major chromatin compartments in anisotropic nuclei and discovered a wider spatial dispersion of nanodomain clusters in the nucleoplasm and condensed larger nanoclusters near the periphery and pericentromeric heterochromatin. Upon examining the spatiotemporal dynamics of heterochromatin in anisotropic nuclei, we observed reduced mobility of the constitutive heterochromatin mark H3K9me3 and the associated heterochromatin protein 1 (HP1α) at the nucleoplasm and periphery regions, correlating with increased viscosity and changes in gene expression. Since heterochromatin remodeling is crucial to genome integrity, our results reveal an unconventional H3K9me3 heterochromatin distribution, providing cues to an altered chromatin state due to perturbations of the nuclei in aligned fiber configurations.

SenSE: A Toolkit for Semantic Change Exploration via Word Embedding Alignment

Lexical Semantic Change (LSC) detection, also known as Semantic Shift, is the process of identifying and characterizing variations in language usage across different scenarios such as time and domain. It allows us to track the evolution of word senses, as well as to understand the difference between the language used in distinct communities. LSC detection is often done by applying a distance measure over vectors of two aligned word embedding matrices. In this demonstration, we present SenSE, an interactive semantic shift exploration toolkit that provides visualization and explanation of lexical semantic change for an input pair of text sources. Our system focuses on showing how the different alignment strategies may affect the output of an LSC model as well as on explaining semantic change based on the neighbors of a chosen target word, while also extracting examples of sentences where these semantic deviations appear. The system runs as a web application (available at http://sense.mgruppi.me), allowing the audience to interact by configuring the alignment strategies while visualizing the results in a web browser.

ORTHOSKIM: In silico sequence capture from genomic and transcriptomic libraries for phylogenomic and barcoding applications.

Low-coverage whole genome shotgun sequencing (or genome skimming) has emerged as a cost-effective method for acquiring genomic data in nonmodel organisms. This method provides sequence information on chloroplast genome (cpDNA), mitochondrial genome (mtDNA) and nuclear ribosomal regions (rDNA), which are over-represented within cells. However, numerous bioinformatic challenges remain to accurately and rapidly obtain such data in organisms with complex genomic structures and rearrangements, in particular for mtDNA in plants or for cpDNA in some plant families. Here we introduce the pipeline ORTHOSKIM, which performs in silico capture of targeted sequences from genomic and transcriptomic libraries without assembling whole organelle genomes. ORTHOSKIM proceeds in three steps: (i) global sequence assembly, (ii) mapping against reference sequences and (iii) target sequence extraction; importantly it also includes a range of quality control tests. Different modes are implemented to capture both coding and noncoding regions of cpDNA, mtDNA and rDNA sequences, along with predefined nuclear sequences (e.g., ultraconserved elements) or collections of single-copy orthologue genes. Moreover, aligned DNA matrices are produced for phylogenetic reconstructions, by performing multiple alignments of the captured sequences. While ORTHOSKIM is suitable for any eukaryote, a case study is presented here, using 114 genome-skimming libraries and four RNA sequencing libraries obtained for two plant families, Primulaceae and Ericaceae, the latter being a well-known problematic family for cpDNA assemblies. ORTHOSKIM recovered with high success rates cpDNA, mtDNA and rDNA sequences, well suited to accurately infer evolutionary relationships within these families. ORTHOSKIM is released under a GPL-3 licence and is available at: https://github.com/cpouchon/ORTHOSKIM.

Selective Cross-Subject Transfer Learning Based on Riemannian Tangent Space for Motor Imagery Brain-Computer Interface.

A motor imagery (MI) brain-computer interface (BCI) plays an important role in the neurological rehabilitation training for stroke patients. Electroencephalogram (EEG)-based MI BCI has high temporal resolution, which is convenient for real-time BCI control. Therefore, we focus on EEG-based MI BCI in this paper. The identification of MI EEG signals is always quite challenging. Due to high inter-session/subject variability, each subject should spend long and tedious calibration time in collecting amounts of labeled samples for a subject-specific model. To cope with this problem, we present a supervised selective cross-subject transfer learning (sSCSTL) approach which simultaneously makes use of the labeled samples from target and source subjects based on Riemannian tangent space. Since the covariance matrices representing the multi-channel EEG signals belong to the smooth Riemannian manifold, we perform the Riemannian alignment to make the covariance matrices from different subjects close to each other. Then, all aligned covariance matrices are converted into the Riemannian tangent space features to train a classifier in the Euclidean space. To investigate the role of unlabeled samples, we further propose semi-supervised and unsupervised versions which utilize the total samples and unlabeled samples from target subject, respectively. Sequential forward floating search (SFFS) method is executed for source selection. All our proposed algorithms transfer the labeled samples from most suitable source subjects into the feature space of target subject. Experimental results on two publicly available MI datasets demonstrated that our algorithms outperformed several state-of-the-art algorithms using small number of the labeled samples from target subject, especially for good target subjects.

In-vitro effectiveness of poly-β-alanine reinforced poly(3-hydroxybutyrate) fibrous scaffolds for skeletal muscle regeneration

In skeletal muscle tissue engineering, success has not been achieved yet, since the properties of the tissue cannot be fully mimicked. The aim of this study is to investigate the potential use of poly-3-hydroxybutyrate (P3HB)/poly-β-alanine (PBA) fibrous tissue scaffolds with piezoelectric properties for skeletal muscle regeneration. Random and aligned P3HB/PBA (5:1) fibrous matrices were prepared by electrospinning with average diameters of 951 ± 153 nm and 891 ± 247 nm, respectively. X-ray diffraction (XRD) analysis showed that PBA reinforcement and aligned orientation of fibers reduced the crystallinity and brittleness of P3HB matrix. While tensile strength and elastic modulus of random fibrous matrices were determined as 3.9 ± 1.0 MPa and 86.2 ± 10.6 MPa, respectively, in the case of aligned fibers they increased to 8.5 ± 1.8 MPa and 378.2 ± 4.2 MPa, respectively. Aligned matrices exhibited a soft and an elastic behaviour with ~70% elongation in similar to the natural tissue. For the first time, d33 piezoelectric modulus of P3HB/PBA matrices were measured as 5 pC/N and 5.3 pC/N, for random and aligned matrices, respectively. Cell culture studies were performed with C2C12 myoblastic cell line. Both of random and aligned P3HB/PBA fibrous matrices supported attachment and proliferation of myoblasts, but cells cultured on aligned fibers formed regular and thick myofibril structures similar to the native muscle tissue. Reverse transcription polymerase chain reaction (RT-qPCR) analysis indicated that MyoD gene was expressed in the cells cultured on both fiber orientation, however, on the aligned fibers significant increase was determined in Myogenin and Myosin Heavy Chain (MHC) gene expressions, which indicate functional tubular structures. The results of RT-qPCR analysis were also supported with immunohistochemistry for myogenic markers. These in vitro studies have shown that piezoelectric P3HB/PBA aligned fibrous scaffolds can successfully mimic skeletal muscle tissue with its superior chemical, morphological, mechanical, and electroactive properties.

An updated infra-familial classification of Sapindaceae based on targeted enrichment data.

PremiseThe economically important, cosmopolitan soapberry family (Sapindaceae) comprises ca. 1900 species in 144 genera. Since the seminal work of Radlkofer, several authors have attempted to overcome challenges presented by the family’s complex infra‐familial classification. With the advent of molecular systematics, revisions of the various proposed groupings have provided significant momentum, but we still lack a formal classification system rooted in an evolutionary framework.MethodsNuclear DNA sequence data were generated for 123 genera (86%) of Sapindaceae using target sequence capture with the Angiosperms353 universal probe set. HybPiper was used to produce aligned DNA matrices. Phylogenetic inferences were obtained using coalescence‐based and concatenated methods. The clades recovered are discussed in light of both benchmark studies to identify synapomorphies and distributional evidence to underpin an updated infra‐familial classification.Key ResultsCoalescence‐based and concatenated phylogenetic trees had identical topologies and node support, except for the placement of Melicoccus bijugatus Jacq. Twenty‐one clades were recovered, which serve as the basis for a revised infra‐familial classification.ConclusionsTwenty tribes are recognized in four subfamilies: two tribes in Hippocastanoideae, two in Dodonaeoideae, and 16 in Sapindoideae (no tribes are recognized in the monotypic subfamily Xanthoceratoideae). Within Sapindoideae, six new tribes are described: Blomieae Buerki & Callm.; Guindilieae Buerki, Callm. & Acev.‐Rodr.; Haplocoeleae Buerki & Callm.; Stadmanieae Buerki & Callm.; Tristiropsideae Buerki & Callm.; and Ungnadieae Buerki & Callm. This updated classification provides a backbone for further research and conservation efforts on this family.

Directional cues in the tumor microenvironment due to cell contraction against aligned collagen fibers

It is well established that collagen alignment in the breast tumor microenvironment provides biophysical cues to drive disease progression. Numerous mechanistic studies have demonstrated that tumor cell behavior is driven by the architecture and stiffness of the collagen matrix. However, the mechanical properties within a 3D collagen microenvironment, particularly at the scale of the cell, remain poorly defined. To investigate cell-scale mechanical cues with respect to local collagen architecture, we employed a combination of intravital imaging of the mammary tumor microenvironment and a 3D collagen gel system with both acellular pNIPAAm microspheres and MDA-MB-231 breast carcinoma cells. Within the in vivo tumor microenvironment, the displacement of collagen fiber was identified in response to tumor cells migrating through the stromal matrix. To further investigate cell-scale stiffness in aligned fiber architectures and the propagation of cell-induced fiber deformations, precise control of collagen architecture was coupled with innovative methodology to measure mechanical properties of the collagen fiber network. This method revealed up to a 35-fold difference in directional cell-scale stiffness resulting from contraction against aligned fibers. Furthermore, the local anisotropy of the matrix dramatically altered the rate at which contractility-induced fiber displacements decayed over distance. Together, our results reveal mechanical properties in aligned matrices that provide dramatically different cues to the cell in perpendicular directions. These findings are supported by the mechanosensing behavior of tumor cells and have important implications for cell-cell communication within the tissue microenvironment. Statement of significanceIt is widely appreciated that the architecture of the extracellular matrix impacts cellular behavior in normal and disease states. Numerous studies have determined the fundamental role of collagen matrix architecture on cellular mechanosensing, but effectively quantifying anisotropic mechanical properties of the collagen matrix at the cell-scale remains challenging. Here, we developed innovative methodology to discover that collagen alignment results in a 35-fold difference in cell-scale stiffness and alters contractile force transmission through the fiber network. Furthermore, we identified bias in cell response along the axis of alignment, where local stiffness is highest. Overall, our results define cell-scale stiffness and fiber deformations due to collagen architecture that may instruct cell communication within a broad range of tissue microenvironments.

Biomimetic electroconductive nanofibrous matrices for skeletal muscle regenerative engineering.

The regeneration of the muscles of the rotator cuff represents a grand challenge in musculoskeletal regenerative engineering. Several types of matrices have been proposed for skeletal muscle regeneration. However, biomimetic matrices to promote muscle regeneration and mimic native muscle tissue have not been successfully engineered. Besides topographical cues, an electrical stimulus may serve as a critical cue to improve interactions between materials and cells in scenarios fostering muscle regeneration. In this in vitro study, we engineered a novel stimuli-responsive conductive nanocomposite matrix, and studied its ability to regulate muscle cell adhesion, proliferation, and differentiation. Electroconductive nanocomposite matrices demonstrated tunable conductivity and biocompatibility. Under the optimum concentration of conductive material, the matrices facilitated muscle cell adhesion, proliferation, and differentiation. Importantly, conductive aligned fibrous matrices were effective in promoting myoblast differentiation by upregulation of myogenic markers. The results demonstrated promising potential of aligned conductive fibrous matrices for skeletal muscle regenerative engineering.

Functional comparison of beating cardiomyocytes differentiated from umbilical cord-derived mesenchymal/stromal stem cells and human foreskin-derived induced pluripotent stem cells.

In recent years, stem cell-based therapies shown to have promising effects on the clinical management of ischemic heart disease. Moreover, stem cells differentiation into cardiomyocytes (CMs) can overcome the cell source limitations. The current research involves the isolation and expansion of mesenchymal stem cells (MSCs) and induced pluripotent stem cells (iPSCs), their differentiation into CMs and subsequent construction of tissue-engineered myocardium supported by random and aligned polycaprolactone (PCL) nanofibrous matrices (av. dia: 350-850 nm). Umbilical cord matrix (UCM)-derived MSCs were isolated successfully by routine enzymatic digestion and a nonenzymatic explant culture method and characterized by their morphology, differentiation into different lineages, and surface marker expression. Treatment of UCM-derived MSCs with 5-azacytidine (5 μM) induced their differentiation into putative cardiac cells, as revealed by the expression of cardiac-specific troponin T (cTnT), smooth muscles actin, myogenin (MYOG), smoothelin, cardiac α-actin genes and cTnT, α-actinin proteins by RT-PCR and immunocytochemistry, respectively. However, no beating cells were observed in differentiated MSCs. On the other hand, adult human foreskin-derived iPSCs cultured on Matrigel™-coated aligned PCL nanofibrous matrices showed anisotropic behavior along the PCL nanofibers and, upon differentiation, expressed cardiac-specific cTnT (23.34 vs. 32.55%) proteins and showed more synchronized beating than those differentiated on Matrigel™-coated tissue culture coated polystyrene surfaces. Moreover, aligned PCL nanofibers are able to promote cells orientation parallel to the fibers, thus providing an effective way to control anisotropic nature under in vitro condition.

Aligned fibrous decellularized cell derived matrices for mesenchymal stem cell amplification.

Biochemical and biophysical stimuli of stem cell niches finely regulate the self-renewal/differentiation equilibrium. Replicating this in vitro is technically challenging, making the control of stem cell functions difficult. Cell derived matrices capture certain aspect of niches that influence fate decisions. Here, aligned fibrous matrices synthesized by MC3T3 cells were produced and the role of matrix orientation and stiffness on the maintenance of stem cell characteristics and adipo- or osteo-genic differentiation of murine mesenchymal stem cells (mMSCs) was investigated. Decellularized matrices promoted mMSC proliferation. Fibrillar alignment and matrix stiffness work in concert in defining cell fate. Soft matrices preserve stemness, whereas stiff ones, in presence of biochemical supplements, promptly induce differentiation. Matrix alignment impacts the homogeneity of the cell population, that is, soft aligned matrices ameliorate the spontaneous adipogenic differentiation, whereas stiff aligned matrices reduce cross-differentiation. We infer that mechanical signaling is a dominant factor in mMSC fate decision and the matrix alignment contributes to produce a more homogeneous environment, which results in a uniform response of cells to biophysical environment. Matrix thus produced can be obtained in vitro in a facile and consistent manner and can be used for homogeneous stem cell amplification or for mechanotransduction-related studies.

3D electrohydrodynamic printing of highly aligned dual-core graphene composite matrices

The aim of this study was to develop an EHD printing method to fabricate graphene-loaded polycaprolactone (PCL)/polyethylene oxide (PEO) dual-core matrices. Graphene was incorporated in shell PCL components, while gelatin and dopamine hydrochloride (DAH) were encapsulated in two PEO cores to enhance biocompatibility of graphene-loaded matrices. Furthermore, the effect of PEO concentration on dual-core fiber formation was evaluated. The influence of process parameters (applied voltage, inner flow rate, outer flow rate and X-Y-Z collector stage speed) on dual-core fiber morphology was evaluated. Our findings show graphene-loaded structures to possess two inner cores and increasing graphene content yields matrices with smoother surfaces, causing a slight reduction in their contact angle behavior. Furthermore, the addition of graphene to matrices results in reduced elasticity. DAH release from matrices comprising various graphene concentrations showed no significant difference and drug release mechanism was diffusion based. In vitro biological tests indicate resulting graphene-loaded dual-core matrices exhibit good biocompatibility and also improve PC12 cell migration. The findings suggest matrices to have potential applications in nerve restoration and regeneration.

Highly aligned and geometrically structured poly(glycerol sebacate)-polyethylene oxide composite fiber matrices towards bioscaffolding applications.

The biocompatible and biodegradable polymer poly(glycerol sebacate), or PGS, is a rubber-like material that finds use in several biomedical applications. PGS is often cast into a mold to form desired structures; alternatively, blending PGS with other reinforcing polymers produces viscous solutions that can be spun into non-woven fibrous scaffolds. For tissue scaffolding applications, ordered fibrous matrices are advantageous and have been shown to promote cell orientation and proliferation by contact guidance, providing topographical cues for the seeded cells. The development of techniques for easily producing aligned fibrous matrices is therefore a priority. PGS nanofibers have been fabricated successfully using electrospinning techniques. For producing PGS microfibers, we introduce the electro-less STRAND (Substrate Translation and Rotation for Aligned Nanofiber Deposition) process as an alternative to electrospinning. STRAND provides superior control of fiber properties including diameter, alignment, spacing, and therefore deposition density by mechanically drawing polymer fibers from solution. The goal in using this method is the simple production of aligned PGS fiber matrices for retinal tissue scaffolding.

Fiber alignment drives changes in architectural and mechanical features in collagen matrices.

Aligned collagen architecture is a characteristic feature of the tumor extracellular matrix (ECM) and has been shown to facilitate cancer metastasis using 3D in vitro models. Additional features of the ECM, such as pore size and stiffness, have also been shown to influence cellular behavior and are implicated in cancer progression. While there are several methods to produce aligned matrices to study the effect on cell behavior in vitro, it is unclear how the alignment itself may alter these other important features of the matrix. In this study, we have generated aligned collagen matrices and characterized their pore sizes and mechanical properties at the micro- and macro-scale. Our results indicate that collagen alignment can alter pore-size of matrices depending on the polymerization temperature of the collagen. Furthermore, alignment does not affect the macro-scale stiffness but alters the micro-scale stiffness in a temperature independent manner. Overall, these results describe the manifestation of confounding variables that arise due to alignment and the importance of fully characterizing biomaterials at both micro- and macro-scales.

Stromal PTEN Regulates Extracellular Matrix Organization in the Mammary Gland

The organization of the extracellular matrix has a profound impact on cancer development and progression. The matrix becomes aligned throughout tumor progression, providing “highways” for tumor cell invasion. Aligned matrix is associated with breast density and is a negative prognostic factor in several cancers; however, the underlying mechanisms regulating this reorganization remain poorly understood. Deletion of the tumor suppressor Pten in the stroma was previously shown to promote extracellular matrix expansion and tumor progression. However, it was unknown if PTEN also regulated matrix organization. To address this question, a murine model with fibroblast-specific Pten deletion was used to examine how PTEN regulates matrix remodeling. Using second harmonic generation microscopy, Pten deletion was found to promote collagen alignment parallel to the mammary duct in the normal gland and further remodeling perpendicular to the tumor edge in tumor-bearing mice. Increased alignment was observed with Pten deletion in vitro using fibroblast-derived matrices. PTEN loss was associated with fibroblast activation and increased cellular contractility, as determined by traction force microscopy. Inhibition of contractility abrogated the increased matrix alignment observed with PTEN loss. Murine mammary adenocarcinoma cells cultured on aligned matrices derived from Pten−/− fibroblasts migrated faster than on matrices from wild-type fibroblasts. Combined, these data demonstrate that PTEN loss in fibroblasts promotes extracellular matrix deposition and alignment independently from cancer cell presence, and this reorganization regulates cancer cell behavior. Importantly, stromal PTEN negatively correlated with collagen alignment and high mammographic density in human breast tissue, suggesting parallel function for PTEN in patients.