Essential Matrix Research Articles

Preparation of Decellularized Biological Scaffolds for 3D Cell Culture.

The biggest challenge of designing and implementing an in vitro study is developing a microenvironment that most closely represents the interactions observed in vivo. Decellularization of tissues and organs has been shown to be an effective method for the removal of potentially immunogenic constituents while preserving essential growth factors and extracellular matrix (ECM ) proteins necessary for proper cell function. Enzymatic digestion of decellularized tissues allows these tissue-specific components to be reconstituted into bioactive hydrogels through a physical crosslinking of collagen. In the following protocol, we describe unique decellularization methods for both dermis and urinary bladder matrix (UBM) derived from porcine tissues. We then provide details for hydrogel formation and subsequent three-dimensional (3D) culture of two cell types: NIH 3T3 fibroblasts and C2C12 myoblasts .

On the Existence of Epipolar Matrices

This paper considers the foundational question of the existence of a fundamental (resp. essential) matrix given m point correspondences in two views. We present a complete answer for the existence of fundamental matrices for any value of m. We disprove the widely held beliefs that fundamental matrices always exist whenever $$m \le 7$$m≤7. At the same time, we prove that they exist unconditionally when $$m \le 5$$m≤5. Under a mild genericity condition, we show that an essential matrix always exists when $$m \le 4$$m≤4. We also characterize the six and seven point configurations in two views for which all matrices satisfying the epipolar constraint have rank at most one.

Improved Feature Matching for Mobile Devices with IMU.

Thanks to the recent diffusion of low-cost high-resolution digital cameras and to the development of mostly automated procedures for image-based 3D reconstruction, the popularity of photogrammetry for environment surveys is constantly increasing in the last years. Automatic feature matching is an important step in order to successfully complete the photogrammetric 3D reconstruction: this step is the fundamental basis for the subsequent estimation of the geometry of the scene. This paper reconsiders the feature matching problem when dealing with smart mobile devices (e.g., when using the standard camera embedded in a smartphone as imaging sensor). More specifically, this paper aims at exploiting the information on camera movements provided by the inertial navigation system (INS) in order to make the feature matching step more robust and, possibly, computationally more efficient. First, a revised version of the affine scale-invariant feature transform (ASIFT) is considered: this version reduces the computational complexity of the original ASIFT, while still ensuring an increase of correct feature matches with respect to the SIFT. Furthermore, a new two-step procedure for the estimation of the essential matrix E (and the camera pose) is proposed in order to increase its estimation robustness and computational efficiency.

Elastins from patients with Williams–Beuren syndrome and healthy individuals differ on the molecular level

Williams-Beuren syndrome (WBS) is a congenital disorder, which involves the heterozygous deletion of the elastin gene and other genes on chromosome 7. Clinical symptoms that are associated with hemizygosity of the essential extracellular matrix protein elastin include premature aging of the skin and supravalvular aortic stenosis. However, only little is known about the molecular basis of structural abnormalities in the connective tissue of WBS patients. Therefore, for the first time this study aimed to systematically characterize and compare the structure and amount of elastin present in skin and aortic tissue from WBS patients and healthy individuals. Elastin fibers were isolated from tissue biopsies, and it was found that skin of WBS patients contains significantly less elastin compared to skin of healthy individuals. Scanning electron microscopy and mass spectrometric measurements combined with bioinformatics data analysis were used to investigate the molecular-level structure of elastin. Scanning electron microscopy revealed clear differences between WBS and healthy elastin. With respect to the molecular-level structure, it was found that the proline hydroxylation degree differed between WBS and healthy elastin, while the tropoelastin isoform appeared to be the same. In terms of cross-linking, no differences in the content of the tetrafunctional cross-links desmosine and isodesmosine were found between WBS and healthy elastin. However, principal component analysis revealed differences between enzymatic digests of elastin from healthy probands and WBS patients, which indicates differing susceptibility toward enzymatic cleavage. Overall, the study contributes to a better understanding of the correlation between genotypic and elastin-related phenotypic features of WBS patients. © 2016 Wiley Periodicals, Inc.

Improved algorithm to estimate the rotation angle between two images by using the two‐point correspondence pairs

An algorithm that can compute the rotation angle between two images is proposed, independently of the translation angle by using only two-point correspondence pairs. A parameterised solution vector can be calculated from the null-space of a matrix constructed of two-point correspondence pairs, and then the rotation angle is evaluated from this vector by using the relation between the elements of the essential matrix. Once the rotation angle is determined, the translation angle can be computed by using the Ackermann steering constraint, and then the translation distance can be determined. In the experiments, the proposed algorithm estimated the rotation angle and the translation distance better than did previous algorithms.

Microfluidic spinning of fibronectin fibers via silk-induced cross-linking

Event Abstract Back to Event Microfluidic spinning of fibronectin fibers via silk-induced cross-linking Matthew Jacobsen1*, Joyce Y. Wong1 and Michael L. Smith1 1 Boston University, Department of Biomedical Engineering, United States Introduction: Fibronectin (Fn) is an essential extracellular matrix protein that is assembled from its monomeric form in solution in the cytosol to an insoluble fibrous network as it is extruded by the cell into the extracellular space. This polymerization process is not well understood and difficult to recapitulate in a laboratory environment, even using alternate means such as electrospinning. Successful reconstitution methods for Fn fibers outside of the tissue environment rely on surface tension as a driving polymerization force[1],[2]. The limitations of this method are the length scale and quantity of Fn that can be obtained, such that scalability is low. In this work, we present a microfluidic technique that takes advantage of the polymerization method of silk fibroin (SF) in order to cross-link Fn into a continuous spin to improve fiber yields while not altering the biological effects of Fn. Materials and Methods: A tiered microfluidic device (Figure 1) designed for the spinning of pure SF solutions was adapted by modifying the spinning solution[3]. Concentrated solutions of SF (>70 mg/ml) and Fn (>3 mg/ml) were mixed at various ratios to create a range of protein densities in extruded fibers. Flow through the microfluidic device induces elongation of protein chains leading to hydrogen bonding into hierarchical secondary structures. Extrusion into an organic solvent dehydrates the assembled protein fiber to finalize assembly of protein strands in a mature fiber. Fibers were tested for cellular interactivity and structural effects and compared to Fn[1] and SF properties[3], respectively. Results and Discussion: Continuous extrusion was observed for fibers with mass ratios of Fn:SF ranging from 1:1600 to 1:10. Fiber diameters decreased proportionately from the low Fn:SF ratio (30-50 micron) to the high ratio (1-5 micron), shown in Figure 2. Fiber diameters of Fn fibers extruded using the surface tension methodologies range from hundreds of nanometers to 1-3 microns, demonstrating the shift from a predominantly silk-like fiber morphology to that closer resembling pure Fn fibers. Functional degrees of cellular adherence are observed in the spun fibers shown in Figure 3 even at the minimal level of Fn incorporation (1:1600, fluorescently labeled – red), which is not a capability of SF fibers alone. FTIR results demonstrate that the interaction of the SF and Fn solutions occurs at the secondary structure level. This is supported by the result obtained where SF solution was diluted to deprive it of sufficient protein strands to form physical cross-links and, therefore, no fiber. When Fn is added at minimal concentrations, protein fiber production resumes. Conclusion: These results demonstrate that this method for spinning Fn fibers allows for the creation of a physically cross-linked fiber with composite properties that vary depending on the mixing ratio of the spinning solution. This method provides a means to create large quantities of Fn fibers using a simple microfluidic approach that previously could not be achieved with existing methods.

3D surface reconstruction of retinal vascular structures

We propose in this paper, a three-dimensional surface reconstruction of a retinal vascular network from a pair of 2D retinal images. Our approach attempts to address the above challenges by incorporating an epipolar geometry estimation and adaptive surface modelling in a 3D reconstruction, using three steps: segmentation, 3D skeleton reconstruction and 3D surface modelling of vascular structures. The intrinsic calibration matrices are found via the solution of simplified Kruppa equations. A simple essential matrix based on a self-calibration method has been used for the fundus camera-eye system. The used method has eventually produced vessel surfaces that could be fit for various applications, such as applications for computational fluid dynamics simulations and applications for real-time virtual interventional.

Ageing is associated with reduction of mechanically-induced activation of Smad2/3P signaling in articular cartilage

Mechanical signals control key cellular processes in articular cartilage. Previously we have shown that mechanical compression is an important ALK5/Smad2/3P activator in cartilage explants. However, age-related changes in the cartilage are known to affect tissue mechanosensitivity and also ALK5/Smad2/3P signaling. We have investigated whether ageing of cartilage is associated with an altered response to mechanical compression. Articular cartilage explants of two different age groups (young-6-36 months old, aged-6 - 13 years old) were subjected to dynamic mechanical compression with 3 MPa (physiological) or 12 MPa (excessive) load. Subsequently, essential cartilage extracellular matrix (ECM) components and tissue growth factors gene expression was measured in young and aged cartilage by QPCR. Furthermore, the ability of young and aged cartilage, to activate the Smad2/3P signaling in response to compression was analyzed and compared. This was done by immunohistochemical (IH) Smad2P detection and Smad3-responsive gene expression analysis. Aged cartilage showed a highly reduced capacity for mechanically-mediated activation of Smad2/3P signaling when compared to young cartilage. Compression of aged cartilage, induced collagen type II (Col2a1) and fibronectin (Fn1) expression to a far lesser extent than in young cartilage. Additionally, in aged cartilage no mechanically mediated up-regulation of bone morphogenetic protein 2 (Bmp2) and connective tissue growth factor (Ctgf) was observed. We identified age-related changes in cellular responses to mechanical stimulation of articular cartilage. We propose that these changes might be associated with age-related alterations in cartilage functioning and can underlie mechanisms for development of age-related cartilage diseases like osteoarthritis (OA).

Porcine kidneys as a source of ECM scaffold for kidney regeneration.

To produce and examine decellularized kidney scaffolds from porcine as a platform for kidney regeneration research. Porcine kidneys were decellularized with sodium dodecyl sulfate solution and Triton X-100 after the blood was rinsed. Then the renal ECM scaffolds were examined for vascular imaging, histology to investigate the vascular patency, degree of decellularization. Renal ECM scaffolds of porcine kidneys were successfully produced. Decellularized renal scaffolds retained intact microarchitecture including the renal vasculature and essential extracellular matrix components. We have developed an excellent decellularization method that can be used in large organs. These scaffolds maintain their basic components, and show intact vasculature system. This represents a step toward development of a transplantable organ using tissue engineering techniques.

The transcription factors SOX9 and SOX5/SOX6 cooperate genome-wide through super-enhancers to drive chondrogenesis.

SOX9 is a transcriptional activator required for chondrogenesis, and SOX5 and SOX6 are closely related DNA-binding proteins that critically enhance its function. We use here genome-wide approaches to gain novel insights into the full spectrum of the target genes and modes of action of this chondrogenic trio. Using the RCS cell line as a faithful model for proliferating/early prehypertrophic growth plate chondrocytes, we uncover that SOX6 and SOX9 bind thousands of genomic sites, frequently and most efficiently near each other. SOX9 recognizes pairs of inverted SOX motifs, whereas SOX6 favors pairs of tandem SOX motifs. The SOX proteins primarily target enhancers. While binding to a small fraction of typical enhancers, they bind multiple sites on almost all super-enhancers (SEs) present in RCS cells. These SEs are predominantly linked to cartilage-specific genes. The SOX proteins effectively work together to activate these SEs and are required for in vivo expression of their associated genes. These genes encode key regulatory factors, including the SOX trio proteins, and all essential cartilage extracellular matrix components. Chst11, Fgfr3, Runx2 and Runx3 are among many other newly identified SOX trio targets. SOX9 and SOX5/SOX6 thus cooperate genome-wide, primarily through SEs, to implement the growth plate chondrocyte differentiation program.

THE EFFECT OF SURFACE HETEROGENEITY ON WETTABILITY OF POROUS THREE DIMENSIONAL (3-D) SCAFFOLDS OF POLY(3-HYDROXYBUTYRIC ACID) (PHB) AND POLY(3-HYDROXYBUTYRIC-CO-3-HYDROXYVALERIC ACID) (PHBV)

Biopolymers such as poly(3-hydroxybutyric acid) (PHB) and poly(3-hydroxybutyric-co-3-hydroxyvaleric acid) (PHBV) are preferred ingredients for the manufacture of materials especially in tissue engineering and regenerative medicine application because they are biocompatible, biodegradable and do not possess any environmental problems associated with their production, fabrication, use and disposal. For that reason, studies have been initiated to produce a 3-D biomimetic scaffolds with an improved thickness greater than 1 mm as an alternative material to synthetic polymers in tissue engineering application. The purpose of this study is to evaluate the physico-chemical surface properties of these polymeric 3-D structures and its corresponding thin films prior to be used as biomimetic materials. To measure the surface free energy (wettability), a Cassie-Baxter contact angle correction for heterogeneous wetting of two liquids (water and methylene iodide) were utilized. To verify its wettability, surface free energies were used to estimate the interfacial energy, work of adhesion and spreading coefficient. The results indicate that the calculated Cassie-Baxter contact angle correction method have proven to exhibit one dominant factor that cause a huge deviation from the true contact angles of its corresponding thin films which is surface heterogeneity. This material characteristic would possibly give a side effect (e.g., low cell attachment and proliferation) against cell-biomaterial affinity as highly hydrophobic material are most likely to be unfavorable for the absorption of essential extracellular matrix proteins (compounds that give a signal to cell to attach on the solid surface).

Ontogeny

The ontogeny of the anthropology consists in returning to the history of the discipline, of its formation. It involves studying its institutional and intellectual development.The philosophical anthropology is an essential matrix of the current anthropology; its study allows us to understand the historic shape of intellectualization of the concepts and the work of definition which notably begins in the XVI century with Montaigne and in the XVIII century with Kant. As a consequence, the philosophy was the armature of the ontogeny of the anthropology and allowed to propose a universal frame of analysis on the world and men.

Fibrinogen Regulates MMP‐9 Processing of VEGF‐induced Angiogenesis and Tumor Growth in Osteosarcoma

PurposeOsteosarcoma (OGS) is the most common primary malignant bone tumor in children and adolescents. Traditionally, fibrin, the essential structural matrix protein of a blood clot, is thought to enhance tumor adhesion and provide a matrix to support tumor growth. Evidence strongly supports that major angiogenic factors, such as vascular endothelial growth factor (VEGF) and matrix metallopeptidase 9 (MMP‐9), can bind fibrin in a concentration‐dependent manner. Therefore, we designed experiments to determine the effects of fibrin on OGS tumor angiogenesis by interacting with VEGF and MMP‐9.MethodsAggressive murine OGS cells are intratibially injected into Balb/C mice. OGS establishment is determined by radiographic analysis, micro‐CT, histology, and angiography.ResultsOpposed to traditional beliefs surrounding fibrin's role in tumor establishment, this work has shown that the loss of fibrin promotes the proliferation and vascularity of OGS, therefore promoting tumor establishment and disease progression.imageTherefore, we have confirmed that fibrin does play a role in tumor angiogenesis through its interaction with VEGF, however we have also shown for the first time that MMP‐9 is a novel intermediary factor for this interaction.ConclusionsIn this study, we have demonstrated that fibrin can inhibit tumor angiogenesis by inhibiting MMP‐9 activity and by sequestering VEGF that is cleaved by MMP‐9. This finding demonstrates a novel synergistic relationship between fibrin and MMP‐9 in the progression of OGS, while also identifying new therapeutic targets.

A hierarchical method for coupling analysis of design services in distributed collaborative design environment

Design is a kind of highly complex activities with a large number of coupling relationships existing among essential design factors and design resources. This paper investigates the approach and mechanism of distributed design resources integration and binding in collaborative design process based on service-oriented architecture. A SORCER-based collaborative design environment is presented. An essential design factors matrix is designed as the binding mechanism to drive distributed design resources integration process during the design tasks execution time. Then, a coupling analysis method for design service modelling and execution is proposed. In order to build a multi-resolution coupling model in the local and global size, an essential design factors binding matrix and an essential design factors matrix group are adopted to express the internal relationships between design services. A hybrid approach, based on the partitioning operation, clustering operation and the genetic algorithm, are used to solve the coupling sets for global essential design factors.

Engineering complex orthopaedic tissues via strategic biomimicry.

The primary current challenge in regenerative engineering resides in the simultaneous formation of more than one type of tissue, as well as their functional assembly into complex tissues or organ systems. Tissue-tissue synchrony is especially important in the musculoskeletal system, wherein overall organ function is enabled by the seamless integration of bone with soft tissues such as ligament, tendon, or cartilage, as well as the integration of muscle with tendon. Therefore, in lieu of a traditional single-tissue system (e.g., bone, ligament), composite tissue scaffold designs for the regeneration of functional connective tissue units (e.g., bone-ligament-bone) are being actively investigated. Closely related is the effort to re-establish tissue-tissue interfaces, which is essential for joining these tissue building blocks and facilitating host integration. Much of the research at the forefront of the field has centered on bioinspired stratified or gradient scaffold designs which aim to recapitulate the structural and compositional inhomogeneity inherent across distinct tissue regions. As such, given the complexity of these musculoskeletal tissue units, the key question is how to identify the most relevant parameters for recapitulating the native structure-function relationships in the scaffold design. Therefore, the focus of this review, in addition to presenting the state-of-the-art in complex scaffold design, is to explore how strategic biomimicry can be applied in engineering tissue connectivity. The objective of strategic biomimicry is to avoid over-engineering by establishing what needs to be learned from nature and defining the essential matrix characteristics that must be reproduced in scaffold design. Application of this engineering strategy for the regeneration of the most common musculoskeletal tissue units (e.g., bone-ligament-bone, muscle-tendon-bone, cartilage-bone) will be discussed in this review. It is anticipated that these exciting efforts will enable integrative and functional repair of soft tissue injuries, and moreover, lay the foundation for the development of composite tissue systems and ultimately, total limb or joint regeneration.

A global approach for image orientation using Lie algebraic rotation averaging and convex L∞ minimisation

Abstract. In this paper we present a new global image orientation approach for a set of multiple overlapping images with given homologous point tuples which is based on a two-step procedure. The approach is independent on initial values, robust with respect to outliers and yields the global minimum solution under relatively mild constraints. The first step of the approach consists of the estimation of global rotation parameters by averaging relative rotation estimates for image pairs (these are determined from the homologous points via the essential matrix in a pre-processing step). For the averaging we make use of algebraic group theory in which rotations, as part of the special orthogonal group SO(3), form a Lie group with a Riemannian manifold structure. This allows for a mapping to the local Euclidean tangent space of SO(3), the Lie algebra. In this space the redundancy of relative orientations is used to compute an average of the absolute rotation for each image and furthermore to detect and eliminate outliers. In the second step translation parameters and the object coordinates of the homologous points are estimated within a convex L∞ optimisation, in which the rotation parameters are kept fixed. As an optional third step the results can be used as initial values for a final bundle adjustment that does not suffer from bad initialisation and quickly converges to a globally optimal solution. We investigate our approach for global image orientation based on synthetic data. The results are compared to a robust least squares bundle adjustment. In this way we show that our approach is independent of initial values and more robust against outliers than a conventional bundle adjustment.

A high-accuracy monocular self-calibration method based on the essential matrix and bundle adjustment

Accurate camera calibration is necessary in photogrammetry systems. While traditional calibration methods need precise calibration pattern, this paper introduces a high-accuracy self-calibration method that doesn’t need any metric information of the scene. The proposed method combines epipolar geometry and bundle adjustment, and both intrinsic and extrinsic parameters are obtained. Synthetic data as well as real data have been used to test the method, and the simulation result shows that the mean absolute relative calibration errors of the focal lengths and the principal point are about 4.5e-4 and 8e-4, respectively, under zero-mean Gaussian noise with 0.1 pixels standard deviation. Compared with existing calibration methods, the proposed method is cost-effective and simple with comparable accuracy.

Decellularized ovine arteries as small-diameter vascular grafts

Atherosclerosis and its complications still represent the leading cause of death in the developed countries. While autologous blood vessels may be regarded as the best solution for peripheral and coronary bypass, they are unavailable in most patients. Even though tissue engineering techniques are often applied to the development of small-diameter vascular grafts, limiting factors of this approach are represented by the lack of essential extracellular matrix proteins and/or poor biomechanical properties of the scaffolds used. Along these lines, the aim of this study was to develop a decellularization protocol for ovine carotids to be used as suitable small-diameter vascular grafts. Samples were treated either with sodium dodecyl sulphate (SDS) or with Trypsin and Triton X-100; a final nuclease digestion was performed for both protocols. Morphological analyses demonstrate complete removal of nuclei and cellular components in treated vessels, also confirmed by significant reduction in wall thickness and DNA content. Essential extracellular matrix proteins such as collagen, elastin, and fibronectin are well preserved after decellularization. From a mechanical point of view, Trypsin and Triton X-100 treated arteries show elastic modules and compliance comparable to native carotids, whereas the use of SDS makes samples stiffer, with a significant decrease in the compliance mean value and an increase in longitudinal and circumferential Young’s modules. It is demonstrated that the treatment where Trypsin and Triton X-100 are combined guarantees complete decellularization of carotids, with no significant alteration of biomechanical and structural properties, thus preserving a suitable environment for adhesion, proliferation, and migration of cells.

Unique mutation portraits and frequent COL2A1 gene alteration in chondrosarcoma.

Chondrosarcoma is the second most frequent malignant bone tumor. However, the etiological background of chondrosarcomagenesis remains largely unknown, along with details on molecular alterations and potential therapeutic targets. Massively parallel paired-end sequencing of whole genomes of 10 primary chondrosarcomas revealed that the process of accumulation of somatic mutations is homogeneous irrespective of the pathological subtype or the presence of IDH1 mutations, is unique among a range of cancer types, and shares significant commonalities with that of prostate cancer. Clusters of structural alterations localized within a single chromosome were observed in four cases. Combined with targeted resequencing of additional cartilaginous tumor cohorts, we identified somatic alterations of the COL2A1 gene, which encodes an essential extracellular matrix protein in chondroskeletal development, in 19.3% of chondrosarcoma and 31.7% of enchondroma cases. Epigenetic regulators (IDH1 and YEATS2) and an activin/BMP signal component (ACVR2A) were recurrently altered. Furthermore, a novel FN1-ACVR2A fusion transcript was observed in both chondrosarcoma and osteochondromatosis cases. With the characteristic accumulative process of somatic changes as a background, molecular defects in chondrogenesis and aberrant epigenetic control are primarily causative of both benign and malignant cartilaginous tumors.

A novel p. Gly630Ser mutation of COL2A1 in a Chinese family with presentations of Legg-Calvé-Perthes disease or avascular necrosis of the femoral head.

ObjectiveMutations in the type II collagen gene are associated with certain human disorders, collectively termed type II collagenopathies. They include Legg–Calvé–Perthes disease (LCPD) and avascular necrosis of the femoral head (ANFH). These two diseases are skeletal dysplasias, inherited in an autosomal dominant fashion, characterized by groin pain, dislocation of the hip and diminished joint mobility. Coxa vara and elevation of the greater trochanter of the femur comprise the typical phenotype of LCPD, but do not occur in ANFH. Lack of synthesis of type II collagen and structural defects are responsible for the major clinical outcomes, because collagen is the essential matrix protein of all connective tissues. Type II collagen, encoded by the COL2A1 gene, contains N- and C- terminal regions that are cleaved after secretion into the extracellular matrix, and the core area is composed of a triple helical (Gly–X–Y) domain. If the Gly in this specific region is replaced by other amino acids, the structure of type II collagen will be destroyed.MethodForty-five members of a four-generation family were recruited and investigated. Diagnosis was made by independent orthopedic surgeons and radiologists. A mutation of the COL2A1 gene was detected.ResultIn our research, we identify a heterozygous mutation (c.1888 G>A, p. Gly630Ser) in exon 29 of COL2A1 in the Gly–X–Y domain, in a Chinese family affected by LCPD and ANFH. Our findings provide significant clues to the phenotype–genotype relationships in these syndromes and may be helpful in clinical diagnosis. Furthermore, these results should assist further studies of the mechanisms underlying collagen diseases.ConclusionOur data add new variants to the repertoire of COL2A1 mutation resulting in related collagenopathies.