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Tailoring the collagen film structural properties via direct laser crosslinking of star-shaped polylactide for robust scaffold formation

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Abstract

Application of restructured collagen-based biomaterials is generally restricted by their poor mechanical properties, which ideally must be close to those of a tissue being repaired. Here, we present an approach to the formation of a robust biomaterial using laser-induced curing of a photosensitive star-shaped polylactide. The created collagen-based structures demonstrated an increase in the Young's modulus by more than an order of magnitude with introduction of reinforcing patterns (from 0.15 ± 0.02 MPa for the untreated collagen to 51.2 ± 5.6 MPa for the reinforced collagen). It was shown that the geometrical configuration of the created reinforcing pattern affected the scaffold's mechanical properties only in the case of a relatively high laser radiation power density, when the effect of accumulated thermomechanical stresses in the photocured regions was significant. Photo-crosslinking of polylactide did not compromise the scaffold's cytotoxicity and provided fluorescent regions in the collagen matrix, that create a potential for noninvasive monitoring of such materials' biodegradation kinetics in vivo. VIEW PDF: https://www.sciencedirect.com/science/article/pii/S0928493118340116

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... Functionalized with (meth)acrylate groups star low molecular weight polylactides have been used for stereolithography also by other groups where two-photon polymerization (2PP) technique was applied for crosslinking [70][71][72][73][74][75]. Star-shaped methacrylate-terminated oligo(d,l-lactide)s with M n = 2800 were prepared, and it was demonstrated that oligomer synthesis and their functionalization can be carried out in the same reactor [71]. ...
... Similarly, fabricated scaffolds (2PP technique) were also used for supporting of Schwann cells growth and thus, as neural scaffolds in nerve repair [70]. Laser-induced crosslinked star-shaped methacrylate-terminated oligo(d,l-lactide)s (M n = 2400) were used as a reinforcement of collagen materials [74,75]. The material exhibited improved resistance to biodegradation, while the direct multipotent stromal cell growth during their culture was observed. ...
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... It should be noted that collagen materials not only provide scaffolds with great biocompatibility but also enhance regeneration of various tissues that defines their popularity in the regenerative medicine [34,35]. The implantation of a collagen sponge with MSCs into a defect of the patellar tendon showed that, 12 weeks post-implantation, the biomechanical strength of the neo-tendon was twice as high as that of the control, but 50% lower than that of the normal tendon. ...
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... Compared to Matrigel and gels from decellularized tissues, synthetic hydrogel systems have a defined composition, which can be easily controlled and tuned to provide the required properties. [35][36][37][38][39] This defined composition is essential when fabricating a standardized organoid-based system for drug testing, because it can be validated and avoids the influence of additional microenvironmental factors on experimental data. ...
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A two-photon laser femtosecond crosslinking process at the wavelength of 525 nm was studied in a human donor cornea in the presence of riboflavin using two-photon optical microscopy and nanoindentation. It was shown that such an approach results in efficient crosslinking of the corneal collagen and a significant (three-fold) increase in the Young's modulus of the corneal structure. Application of a femtosecond laser with the wavelength of 525 nm allows a drastic enhancement of efficiency in the presence of riboflavin on human corneas and a 50-fold reduction of the laser treatment duration in comparison with the use of a femtosecond laser with the wavelength of 760 nm. We relate this effect to a significant growth in the coefficient of two-photon absorption due to the laser wavelength falling within the edge of the photoinitiator (riboflavin) absorption band. Our studies on a donor human cornea demonstrate the good potential for the clinical application of a femtosecond laser with the wavelength of 525 nm for increasing the cornea rigidity using the two-photon laser femtosecond crosslinking technique.
Article
Collagen has shown promise as a bioink for extrusion-based bioprinting, but further development of new collagen bioink formulations is necessary to improve their printability. Screening these formulations by measuring print accuracy is a costly and time consuming process. We hypothesized that rheological properties of the bioink before, during, and/or after gelation can be used to predict printability. In this study, we investigated the effects of riboflavin photocrosslinking and pH on type I collagen bioink rheology before, during, and after gelation and directly correlated these findings to the printability of each bioink formulation. From the riboflavin crosslinking study, results showed that riboflavin crosslinking increased the storage moduli of collagen bioinks, but the degree of improvement was less pronounced at higher collagen concentrations. Dots printed with collagen bioinks with riboflavin crosslinking exhibited smaller dot footprint areas than those printed with collagen bioinks without riboflavin crosslinking. From the pH study, results showed that gelation kinetics and final gel moduli were highly pH dependent and both exhibited maxima around pH 8. The shape fidelity of printed lines was highest at pH 8–9.5. The effect of riboflavin crosslinking and pH on cell viability was assessed using bovine chondrocytes. Cell viability in collagen gels was found to decrease after blue light activated riboflavin crosslinking but was not affected by pH. Correlations between rheological parameters and printability showed that the modulus associated with the bioink immediately after extrusion and before deposition was the best predictor of bioink printability. These findings will allow for the more rapid screening of collagen bioink formulations.
Article
Hyaluronic acid and poly(ethylene glycol) derivatives attract considerable attention as precursors for tissue engineering. In this paper photocuring of biocompatible hyaluronic acid-glycidyl methacrylate (HAGM) and poly(ethylene glycol) diacrylate (PEG-DA) aqueous solutions, using flavin mononucleotide (FMN) as an endogenous photoinitiator, has been studied. The required threshold concentrations of initial macromolecules in water for the strengthening (increase of the Young’s modulus) of irradiated hydrogels have been determined as 57 wt% for 2D cross-linking of PEG-DA compositions and 16 wt% for 3D cross-linking of HAGM compositions. These concentrations are in a good agreement with correspondent values derived from the percolation theory for 2D and 3D lattices. It has been demonstrated that cross-linking proceeds predominantly by the radical mechanism and does not require co-initiators. Hydrogel scaffolds with specific and predetermined architectonics for biocompatibility and biomechanical studies have been produced by photopolymerizable micromolding.
Article
The photoinitiators used in light mediated hydrogelation have been limited due to cytotoxicity and solubility issues. Here we report the use of riboflavin and flavin mononucleotide as nontoxic, naturally occurring photoinitiators for the thiol-ene hydrogelation of functionalised poly(ethylene glycol). High Resolution Magic Angle Spinning (HRMAS) NMR spectroscopy and oscillatory rheometry were used to probe the efficiency of these initiators for this hydrogel system. The gelation and reaction progression could easily be manipulated using different initiator concentrations and light intensities.
Article
Recently, a three-dimensional (3D) bioprinting process for obtaining a cell-laden structure has been widely applied because of its ability to fabricate biomimetic complex structures embedded with and without cells. To successfully obtain a cell-laden porous block, the cell-delivering vehicle, bioink, is one of the significant factors. Until now, various biocompatible hydrogels (synthetic and natural biopolymers) have been utilized in the cell-printing process, but a bioink satisfying both biocompatibility and print-ability requirements to achieve porous structure with reasonable mechanical strength has not been issued. Here, we propose a printing strategy with optimal condition including a safe cross-linking procedure for obtaining a 3D porous cell-block composed of a biocompatible collagen-bioink and genipin, a cross-linking agent. To obtain the optimal processing conditions, we modified the 3D printing machine and selected an optimal cross-linking condition (~1 mM and 1 h) of genipin solution. To show the feasibility of the process, 3D pore-interconnected cell-laden constructs were manufactured using osteoblast-like-cells (MG63) and human adipose stem cells (hASCs). Under these processing conditions, a macroscale 3D collagen-based cell-block of 21 × 21 × 12 mm³ and over 95% cell-viability was obtained. In vitro biological testing of the cell-laden 3D porous structure showed that the embedded cells were sufficiently viable, and their proliferation was significantly higher; the cells also exhibited increased osteogenic activities compared to the conventional alginate-based bioink (control). The results indicated the fabrication process using the collagen-bioink would be an innovative platform to design highly biocompatible and mechanically stable cell-blocks.
Article
Unlabelled: Natural biomaterials such as collagen show promise in tissue engineering applications due to their inherent bioactivity. The main limitation of collagen is its low mechanical strength and somewhat unpredictable and rapid degradation rate; however, combining collagen with another material, such as chitosan, can reinforce the scaffold mechanically and may improve the rate of degradation. Additionally, the high cost and the risk of prion transmission associated with mammal-derived collagen has prompted research into alternative sources such as marine-origin collagen. In this context, the overall goal of this study was to determine if the incorporation of chitosan into collagen scaffolds could improve the mechanical and biological properties of the scaffold. In addition the study assessed if collagen, derived from salmon skin (marine), can provide an alternative to collagen derived from bovine tendon (mammal) for tissue engineering applications. Scaffold architecture and mechanical properties were assessed as well as their ability to support mesenchymal stem cell growth and differentiation. Overall, the addition of chitosan to bovine and salmon skin-derived collagen scaffolds improved the mechanical properties, increasing the compressive strength, swelling ratio and prolonged the degradation rate. Mesenchymal stem cell (MSC) attachment and proliferation was most improved on the bovine-derived collagen scaffold containing a 75:25 ratio of collagen:chitosan, and when MSC osteogenic and chondrogenic potential on the scaffold was assessed, a significant increase in calcium production (p<0.001) and sulfated glycosaminoglycan (sGAG) production (p<0.001) was observed respectively. Regardless of chitosan content, the bovine-derived collagen scaffolds out-performed the salmon skin-derived collagen scaffolds, displaying a larger pore size and higher percentage porosity, more regular architecture, higher compressive modulus, a greater capacity for water uptake and allowed for more MSC proliferation and differentiation. This versatile scaffold incorporating the marine biomaterial chitosan show great potential as appropriate platforms for promoting orthopaedic tissue repair while the use of salmon skin-derived collagen may be more suitable in the repair of soft tissues such as skin. Statement of significance: Collagen is commonly used in tissue engineering due to its biocompatibility; however, it has low mechanical strength and an unpredictable degradation rate. In addition, high cost and risk of prion transmission associated with mammalian-derived collagen has prompted research into alternative collagen sources, namely, marine-derived collagen. In this study, scaffolds made from salmon-skin collagen were compared to the more commonly used bovine-derived collagen with a focus on orthopaedic applications. To improve the mechanical properties of these scaffolds, another marine biomaterial, chitosan, was added to produce scaffolds with increased mechanical stability. The collagen-chitosan composites were also shown to support mesenchymal stem cell differentiation towards both bone and cartilage tissue. This multi-functional scaffold therefore has potential in both bone and cartilage regeneration applications.
Article
Aim: To assess the properties of 3D biodegradable scaffolds fabricated from novel star-shaped poly(D,L-lactide) (SSL) materials for bone tissue regeneration. Materials & methods: The SSL polymer was synthesized using an optimized synthetic procedure and applied for scaffold fabrication by the two-photon polymerization technique. The osteogenic differentiation was controlled using human adipose-derived stem cells cultured for 28 days. The SSL scaffolds with or without murine MSCs were implanted into the cranial bone of C57/Bl6 mice. Results: The SSL scaffolds supported differentiation of human adipose-derived stem cells toward the osteogenic lineage in vitro. The SSL scaffolds with murine MSCs enhanced the mineralized tissue formation. Conclusion: The SSL scaffolds provide a beneficial microenvironment for the osteogenic MSCs' differentiation in vitro and support de novo bone formation in vivo.
Article
Type I collagen is a versatile biomaterial that is widely used in medical applications due to its weak antigenicity, robust biocompatibility, and its ability to be modified for a wide array of applications. As such, collagen has become a major component of many tissue engineering scaffolds, drug delivery platforms, and substrates for in vitro cell culture. In these applications, collagen constructs are fabricated to recapitulate a diverse set of conditions. Collagen fibrils can be aligned during or post-fabrication, cross-linked via numerous techniques, polymerized to create various fibril sizes and densities, and copolymerized into a wide array of composite scaffolds. Here, we review approaches that have been used to tune collagen to better recapitulate physiological environments for use in tissue engineering applications and studies of basic cell behavior. We discuss techniques to control fibril alignment, methods for cross-linking collagen constructs to modulate stiffness, and composite collagen constructs to better mimic physiological extracellular matrix.
Article
Chitosan/collagen (Chit/Col) blends have demonstrated great potential for use in tissue engineering (TE) applications. However, there exists a lack of detailed study on the influence of important design parameters (i.e, component ratio or crosslinking methods) on the essential properties of the scaffolds (morphology, mechanical stiffness, swelling, degradation and cytotoxicity). This work entailed a systematic study of these essential properties of three Chit/Col compositions, covering a wide range of component ratios and using different crosslinking methods. Our results showed the possibility of tailoring these properties by changing component ratios, since different interactions occurred between Chit/Col: samples with Chit-enriched compositions showed a hydrogen-bonding type complex (HC), whereas a self-crosslinking phenomenon was induced in Col-enriched scaffolds. Additionally, material and biological properties of the resultant matrices were further adjusted and tuned by changing crosslinking conditions. In such way, we obtained a wide range of scaffolds whose properties were tailored to meet specific needs of TE applications. Copyright © 2015 Elsevier Ltd. All rights reserved.
Article
A herniated intervertebral disc often causes back pain when disc tissue is displaced through a damaged annulus fibrosus. Currently the only methods available for annulus fibrosus repair involve mechanical closure of defect, which does little to address biological healing in the damaged tissue. Collagen hydrogels are injectable and have been used to repair annulus defects in vivo. In this study, high-density collagen hydrogels at 5, 10 and 15 mg/ml were used to repair defects made to intact rat caudal intervertebral discs in vitro. A group of gels at 15 mg/ml were also crosslinked with riboflavin at 0.03 mM, 0.07 mM or 0.10 mM. These crosslinked, high-density collagen gels maintained presence in the defect under loading and contributed positively to the mechanical response of damaged discs. Discs exhibited increases to 95% of undamaged effective equilibrium and instantaneous moduli as well as up to four fold decreases in effective hydraulic permeability from the damaged discs. These data suggest that high density collagen gels may be effective at restoring mechanical function of injured discs as well as potential vehicles for delivery of biological agents such as cells or growth factors that may aid in the repair of the annulus fibrosus. This article is protected by copyright. All rights reserved.
Article
Although collagen with outstanding biocompatibility has promising application in corneal tissue engineering, the mechanical properties of collagen-based scaffolds, especially suture retention strength, must be further improved to satisfy the requirements of clinical applications. This paper describes a toughness reinforced collagen-based membrane using silk fibroin. The collagen-silk fibroin membranes based on collagen(silk fibroin(w/w) ratios of 100:5, 100:10 and 100:20) were prepared by using silk fibroin and cross-linking by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC). These membranes were analyzed by scanning electron microscopy (SEM) and their optical property and NaCl and tryptophan diffusivity had been tested. The water content was found to be dependent on the content of silk fibroin, and CS10 membrane (loading 10 wt % of silk fibroin) performed the optimal mechanical properties. Also the suture experiments have proved CS10 has high suture retention strength, which can be sutured in rabbit eyes integrally. Moreover, the composite membrane proved good biocompatibility for the proliferation of HCECs (human corneal epithelial cells) in vitro. Lamellar keratoplasty shows that CS10 membrane promoted complete epithelialisation in 35 ± 5 days, and their transparency is restored quickly in the first month. Corneal rejection reaction, neovascularization and keratoconus are not observed. The composite films show potential for use in the field of corneal tissue engineering.
Article
Degradation behaviors of both the unplasticized and plasticized poly(vinyl chloride) (UPVC and PPVC) under an artificial accelerating aging condition were extensively studied. The dependence of mechanical properties, average molecular weight (MW), and surface morphology of the earlier PVC on aging time was investigated by tensile tests, gel penetrate chromatogram (GPC), and scanning electron microscope (SEM), respectively. Fourier transform infrared and ultraviolet (UV)–visible spectroscopy were used to evaluate the probable formation of both the oxygen‐containing groups and the conjugated sequences during aging. The results reveal that UPVC is much easier to be degraded than PPVC under the same testing conditions. The irradiated surface is detected to change from an even topology into a rough topology initially, and then follows the appearance of many voids even cracks in the SEM morphology. During the aging process, oxygen‐containing groups and conjugation of PVC molecular chains around the cracks are observed, and noticeably increase with aging time. However, visible difference of the corresponding MWs of PVC before and after aging is not detected. Moreover, a novel degradation mechanism nearly related to the formation of microvoids and microcracks based on the cohesion energy of groups along PVC molecular chains is developed and semiquantitatively discussed. It is detected that the formation of microvoids and microcracks is attributed to both the thermodynamic changes of PVC backbone during the aging and the aggregation of oxygen‐containing groups with relatively large volumes. © 2011 Wiley Periodicals, Inc. J Appl Polym Sci, 2012
Article
Fabricating materials with tailored mechanical properties is a challenge and crucial for their successful application in a variety of fields such as tissue engineering. Here collagen and riboflavin were used to create hydrogels with controlled mechanical properties mimicking those of soft tissues (e.g. liver). Collagen-based hydrogels were obtained using a two-step gelation method. Firstly a physical gelation step (i.e. modulation of temperature and pH) was used to fix a specific shape; then photo-initiated cross-links were formed to increase the stiffness. Specifically the chemical cross-linking step was initiated with UV (ultra-violet) radiation to obtain riboflavin derivatised radical polymerization of collagen chains. Cylindrical shaped samples with controlled dimensions were fabricated, and then tested using compressive loading. We show that the compressive elastic modulus of collagen-based hydrogels can be tuned between 0.9 and 3.6 kPa by changing collagen concentration, irradiation with UV in the presence of riboflavin and freeze-drying.
Article
Collagen (Col) hydrogels have poor physicochemical and mechanical properties and are susceptible to substantial shrinkage during cell culture, which limits their potential applications in hard tissue engineering. Here, we developed novel nanocomposite hydrogels made of collagen and mesoporous bioactive glass nanoparticle (mBGn) with surface amination, and addressed the effects of mBGn addition (Col: mBG = 2:1, 1:1 and 1:2) and its surface amination on the physicochemical and mechanical properties of the hydrogels. The amination of mBGn was shown to enable chemical bonding with collagen molecules. As a result, the nanocomposite hydrogels exhibited a significantly improved physicochemical and mechanical stability. The hydrolytic and enzymatic degradation of the Col-mBGn hydrogels were slowed down due to the incorporation of mBGn and its surface amination. Mechanical properties of the hydrogels, specifically the resistance to loading as well as the stiffness significantly increased with addition of mBGn and its aminated form, as assessed by a dynamical mechanical analysis. Mesenchymal stem cells cultivated within the Col-mBGn hydrogels were highly viable with enhanced cytoskeletal extensions due to the addition of surface aminated mBGn. While the Col hydrogel showed extensive shrinkage (down to ∼20% of initial size) during a few days of culture, the mBGn-added hydrogel exhibited substantially reduced shrinkage, and the aminated mBGn-added hydrogel had no observable shrinkage over 21 days. Results demonstrated the effective roles of the aminated mBGn in significantly improving the physicochemical and mechanical properties of Col hydrogel that is ultimately favorable for the applications in stem cell culture for bone tissue engineering.
Article
Accelerated UV degradation conditions were imposed on different epoxy resin composites reinforced with carbon fibre, 3D parabeam glass, and E-glass. X-ray micro computed tomography (XμCT) was used to evaluate the changes in the composites’ inner structure as a result of the treatment. Structural damage factors such as density changes, reinforcement filler damage, homogeneity, cracks and microcracks were examined. XμCT 3D images, 2D reconstructed slices and void calculations showed cracks in all composites with different shapes and volume in response to the UV degradation conditions. 3D-glass reinforced epoxy composites showed the least amount of damage in comparison with other composites. E-glass suffered the most damage upon UV degradation. Most of the structural damage in both 3D-glass and E-glass composites occurred in the resin matrix in the near-surface region, while internal damage was present in the form of delamination in the carbon fibre composite.
Article
Purpose: To evaluate the effect of collagen cross-linking induced by genipin in porcine sclera. Methods: Porcine cadaver eyes were treated with genipin at concentrations (by w/v) of 0.01%, 0.03%, 0.1%, 0.3%, 1.0% for 15 and 30 min. Riboflavin/ultraviolet A(UVA)-treated and untreated samples were used as controls. After treatment, scleral strips of 4.0 × 10.0 mm were cut. Twenty-four hours later, the stress-strain parameters of the strips were measured using a biomaterial microtester. The stress and Young’s modulus at 8% strain were evaluated. Results: Compared with untreated groups, after treatment with genipin for 15 min, the stress was increased by 66–246%, depending on the concentration of genipin. As for the 30-min groups, the stress was 171–444% higher than that of the control. The difference of the Young’s modulus between genipin 15-min groups, except the 0.01% groups (p = 0.095), also had statistical significance (p < 0.05). The Young’s modulus had significant difference between the untreated group and the genipin 30-min groups (all p < 0.05). Of 0.3% genipin for 15 min or 0.01% genipin for 30 min had a similar stress-strain curve with those of eyes treated with the riboflavin/UVA group. The sclera exhibited a bluish colour which became deeper with increase concentration and cross-linking time. Conclusions: Collagen cross-linking induced by genipin could increase the biomechanical strength in porcine sclera. The effect depends on the concentration and treatment time of genipin.
Article
The aim of this study is to characterize and compare damage processes in carbon/epoxy laminates submitted to either isothermal ageing or thermal cycling in neutral (vacuum or nitrogen) and oxidative (air) atmospheres. During a thermal cycling test performed in an oxidative atmosphere like air, there is a coupling effect between matrix oxidation, occurring at the highest temperatures of the cycle, and matrix cracking due to thermo-mechanical ply stresses induced by the prevented differential expansions of the plies. In order to separate those two damage mechanisms, a model of oxidation is used to determine the experimental conditions of an isothermal test ‘equivalent’ to a thermal cycling one, in terms of mass loss due to oxidation. After having checked this equivalence, a quantitative analysis of the damages induced by the two types of tests, is carried out. It is shown that the oxidation of a laminate in isothermal conditions results in damages, which concern only the skin of the sample. On the contrary, the coupling of such damage mechanisms with cyclic stresses in thermal cycling accelerates the damage processes and especially the matrix crack propagation from the surfaces to the core of the laminate.
Article
The matrix oxidation of composite materials involves a weight loss and a density increase and as a consequence, a shrinkage of the skin layer of the material. To simulate this behaviour, we chose a kinetic model of radical chain oxidation coupled with the equation of oxygen diffusion. This model predicts the concentration profile of oxidation products, the weight loss profile and the shrinkage profile in a thick part. When the stress field induced by shrinkage reaches a critical value (the ultimate stress of the oxidized polymer), a “spontaneous” cracking appears in the skin layer of thick parts. In order to predict these critical conditions, we determined the mechanical properties of the oxidized layer of thick parts by studying the ageing of matrix thin films for which oxidation is not controlled by the oxygen diffusion.
Article
Cell-encapsulating hydrogels used in regenerative medicine are designed to undergo a rapid liquid-to-solid phase transition in the presence of cells and tissues so as to maximize crosslinking and minimize cell toxicity. Light-activated free-radical crosslinking (photopolymerization) is of particular interest in this regard because it can provide rapid reaction rates that result in uniform hydrogel properties with excellent temporal and spatial control features. Among the many initiator systems available for photopolymerization, only a few have been identified as suitable for cell-based hydrogel formation owing to their water solubility, crosslinking properties and non-toxic reaction conditions. In this study, three long-wave ultraviolet (UV) light-activtied photoinitiators (PIs) were comparatively tested in terms of cytotoxicity, crosslinking efficiency and crosslinking kinetics of cell-encapsulating hydrogels. The hydrogels were photopolymerized from poly(ethylene glycol) (PEG) diacrylate or PEG–fibrinogen precursors using Irgacure® PIs I2959, I184 and I651, as well as with a chemical initiator/accelerator (APS/TEMED). The study specifically evaluated the PI type, PI concentration and UV light intensity, and how these affected the mechanical properties of the hydrogel (i.e. maximum storage modulus), the crosslinking reaction times and the reaction’s cytotoxicity to encapsulated cells. Only two initiators (I2959 and I184) were identified as being suitable for achieving both high cell viability and efficient crosslinking of the cell-encapsulating hydrogels during the photopolymerization reaction. Optimization of PI concentration or irradiation intensity was particularly important for achieving maximum mechanical properties; a sub-optimal choice of PI concentration or irradiation intensity resulted in a substantial reduction in hydrogel modulus.
Article
Collagen is the most abundant protein in animals. This fibrous, structural protein comprises a right-handed bundle of three parallel, left-handed polyproline II-type helices. Much progress has been made in elucidating the structure of collagen triple helices and the physicochemical basis for their stability. New evidence demonstrates that stereoelectronic effects and preorganization play a key role in that stability. The fibrillar structure of type I collagen-the prototypical collagen fibril-has been revealed in detail. Artificial collagen fibrils that display some properties of natural collagen fibrils are now accessible using chemical synthesis and self-assembly. A rapidly emerging understanding of the mechanical and structural properties of native collagen fibrils will guide further development of artificial collagenous materials for biomedicine and nanotechnology.
Article
During the last two decades molecular genetic and cell mechanisms of proliferation and differentiation of mammalian stem cells have been intensively studied in leading laboratories all over the world. Studies in this field are very important both for basic science and for the development of promising cell therapy technologies. Embryonic stem cells represent a unique experimental model for the investigation of basic principles of mammalian cell differentiation and development. Using this model, important data on similarity in genetic programs during embryonic development and embryonic stem cells differentiation have been obtained. These include basically similar consequent expression of transcription factors, cell receptors, tissue specific proteins, and ion channels. Lines of embryonic stem cells are widely used for the investigation of gene functions in ontogenesis as well as in adult organisms (using gene-knockout strategy). This review deals with different pathways of mammalian (including human) embryonic stem cells differentiation. It considers the main approaches to directed differentiation of these cells in vitro: use of feeder cells, growth factors, and other chemical compounds and also genetic modification. Some examples of application of embryonic stem cells derivatives for cell therapy of some pathological conditions are discussed.
Article
A method to impose and measure a one dimensional strain field via confined compression of a tissue-equivalent and measure the resulting cell and collagen fibril alignment was developed Strain was determined locally by the displacement of polystyrene beads dispersed and entrapped within the network of collagen fibrils along with the cells, and it was correlated to the spatial variation of collagen network birefringence and concentration. Alignment of fibroblasts and smooth muscle cells was determined based on the long axis of elongated cells. Cell and collagen network alignment were observed normal to the direction of compression after a step strain and increased monotonically up to 50% strain. These results were independent of time after straining over 24 hr despite continued cell motility after responding instantly to the step strain with a change in alignment by deforming/convecting with the strained network. Since the time course of cell alignment followed that of strain and not stress which, due to the viscoelastic fluid-like nature of the network relaxes completely within the observation period, these results imply cell alignment in a compacting tissue-equivalent is due to fibril alignment associated with anisotropic network strain. Estimation of a contact guidance sensitivity parameter indicates that both cell types align to a greater extent than the surrounding fibrils.
Using a Collagen/Genipin-bioink and an Optimal 3D Printing Process
  • Y B Kim
  • H Lee
  • G H Kim
Y.B. Kim, H. Lee, G.H. Kim, Strategy to Achieve Highly Porous/Biocompatible Macroscale Cell Blocks, Using a Collagen/Genipin-bioink and an Optimal 3D Printing Process, ACS Appl. Mater. Interfaces. 8 (2016) 32230-32240. doi:10.1021/acsami.6b11669.
Collagen Structure and Stability
  • R R T Shoulders
R.R.T. Shoulders M. D., Collagen Structure and Stability, Annu. Rev. Biochem. 78 (2009) 929-958. doi:10.1146/annurev.biochem.77.032207.120833.
  • T V Chirila
  • D G Harkin
T. V. Chirila, D.G. Harkin, Biomaterials and regenerative medicine in ophthalmology: Second edition, Woodhead Publishing, 2016. doi:10.1016/C2014-0-01443-8.
Compressed collagen gel: a novel scaffold for human bladder cells
  • E M Engelhardt
  • E Stegberg
  • R A Brown
  • J A Hubbell
  • F M Wurm
  • M Adam
  • P Frey
E.M. Engelhardt, E. Stegberg, R.A. Brown, J.A. Hubbell, F.M. Wurm, M. Adam, P. Frey, Compressed collagen gel: a novel scaffold for human bladder cells, J. Tissue Eng. Regenerat. Med. 4 (2010) 123-130, https://doi.org/10.1002/term.
  • Photobiol
  • Chem
Photobiol. A Chem. 341 (2017) 108-114. doi:10.1016/j.jphotochem.2017.03.026.
Novel biodegradable star-shaped polylactide scaffolds for bone regeneration fabricated by two-photon polymerization
  • Bagratashvili
Bagratashvili, Novel biodegradable star-shaped polylactide scaffolds for bone regeneration fabricated by two-photon polymerization, Nanomedicine. 11 (2016) 1041-1053. doi:10.2217/nnm-2015-0022.
  • T V Chirila
  • D G Harkin
T.V. Chirila, D.G. Harkin, Biomaterials and Regenerative Medicine in Ophthalmology, second ed., Woodhead Publishing, 2016, https://doi.org/10.1016/ C2014-0-01443-8.
  • M C Lafarie-Frenot
  • S Rouquié
  • N Q Ho
  • V Bellenger
  • K N Comparison Of Damage
  • Bardakova
M.C. Lafarie-Frenot, S. Rouquié, N.Q. Ho, V. Bellenger, Comparison of damage K.N. Bardakova, et al. Materials Science & Engineering C 107 (2020) 110300
Grebenik obtained her PhD in Biophysics at Macquarie University in Australia, followed by post-doctoral research in tissue engineering at Sechenov University in Russia. Her research interests are mainly focused on engineering nervous
  • Dr
  • Ekaterina
Dr. Ekaterina Grebenik obtained her PhD in Biophysics at Macquarie University in Australia, followed by post-doctoral research in tissue engineering at Sechenov University in Russia. Her research interests are mainly focused on engineering nervous, bone and epithelial tissues.
he gained his PhD degree, and in 2010 -DSc degree. He was engaged in scientific, medical and educational activities at the Department of Urology
  • Dr
  • German
Dr. German Krupinov graduated from the Sechenov Moscow Medical Academy in 1997. In 1999, he graduated from residency in urology. In 1999, he gained his PhD degree, and in 2010 -DSc degree. He was engaged in scientific, medical and educational activities at the Department of Urology. In 2010, he became a professor of the Department of Urology. Since 2018, he has been working as a professor of the Institute for urology and reproductive health. His research interests cover the diagnostics and minimally invasive treatment for prostate diseases.
Since 2018, he has been working as a Director. His research interests cover the development of novel biodegradable biocompatible materials and approaches to their structurization (inc. laser-based technologies)
  • Dr
  • Peter
Dr. Peter Timashev graduated from the Moscow State University of Fine Chemical Technologies in 2002. In 2006, he gained his PhD degree at the Karpov Institute of Physical Chemistry, and in 2016 -DSc degree. In 2016, he became a Head of the Department for Advanced Biomaterials at the Institute for Regenerative Medicine (Sechenov University, Russia). Since 2018, he has been working as a Director. His research interests cover the development of novel biodegradable biocompatible materials and approaches to their structurization (inc. laser-based technologies), 3D bioprinting, and clinical translation of tissue engineering.