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Sample records for 3d collagen gels

  1. Engineering 3D Cellularized Collagen Gels for Vascular Tissue Regeneration.

    PubMed

    Meghezi, Sébastien; Seifu, Dawit G; Bono, Nina; Unsworth, Larry; Mequanint, Kibret; Mantovani, Diego

    2015-06-16

    Synthetic materials are known to initiate clinical complications such as inflammation, stenosis, and infections when implanted as vascular substitutes. Collagen has been extensively used for a wide range of biomedical applications and is considered a valid alternative to synthetic materials due to its inherent biocompatibility (i.e., low antigenicity, inflammation, and cytotoxic responses). However, the limited mechanical properties and the related low hand-ability of collagen gels have hampered their use as scaffold materials for vascular tissue engineering. Therefore, the rationale behind this work was first to engineer cellularized collagen gels into a tubular-shaped geometry and second to enhance smooth muscle cells driven reorganization of collagen matrix to obtain tissues stiff enough to be handled. The strategy described here is based on the direct assembling of collagen and smooth muscle cells (construct) in a 3D cylindrical geometry with the use of a molding technique. This process requires a maturation period, during which the constructs are cultured in a bioreactor under static conditions (without applied external dynamic mechanical constraints) for 1 or 2 weeks. The "static bioreactor" provides a monitored and controlled sterile environment (pH, temperature, gas exchange, nutrient supply and waste removal) to the constructs. During culture period, thickness measurements were performed to evaluate the cells-driven remodeling of the collagen matrix, and glucose consumption and lactate production rates were measured to monitor the cells metabolic activity. Finally, mechanical and viscoelastic properties were assessed for the resulting tubular constructs. To this end, specific protocols and a focused know-how (manipulation, gripping, working in hydrated environment, and so on) were developed to characterize the engineered tissues.

  2. In the beginning there were soft collagen-cell gels: towards better 3D connective tissue models?

    PubMed

    Brown, Robert A

    2013-10-01

    In the 40 years since Elsdale and Bard's analysis of fibroblast culture in collagen gels we have moved far beyond the concept that such 3D fibril network systems are better models than monolayer cultures. This review analyses key aspects of that progression of models, against a background of what exactly each model system tries to mimic. This story tracks our increasing understanding of fibroblast responses to soft collagen gels, in particularly their cytoskeletal contraction, migration and integrin attachment. The focus on fibroblast mechano-function has generated models designed to directly measure the overall force generated by fibroblast populations, their reaction to external loads and the role of the matrix structure. Key steps along this evolution of 3D collagen models have been designed to mimic normal skin, wound repair, tissue morphogenesis and remodelling, growth and contracture during scarring/fibrosis. As new models are developed to understand cell-mechanical function in connective tissues the collagen material has become progressively more important, now being engineered to mimic more complex aspects of native extracellular matrix structure. These have included collagen fibril density, alignment and hierarchical structure, controlling material stiffness and anisotropy. But of these, tissue-like collagen density is key in that it contributes to control of the others. It is concluded that across this 40 year window major progress has been made towards establishing a family of 3D experimental collagen tissue-models, suitable to investigate normal and pathological fibroblast mechano-functions.

  3. The role of phosphoinositide 3-kinases in neutrophil migration in 3D collagen gels.

    PubMed

    Martin, Kayleigh J S; Muessel, Michelle J; Pullar, Christine E; Willars, Gary B; Wardlaw, Andrew J

    2015-01-01

    The entry of neutrophils into tissue has been well characterised; however the fate of these cells once inside the tissue microenvironment is not fully understood. A variety of signal transduction pathways including those involving class I PI3 Kinases have been suggested to be involved in neutrophil migration. This study aims to determine the involvement of PI3 Kinases in chemokinetic and chemotactic neutrophil migration in response to CXCL8 and GM-CSF in a three-dimensional collagen gel, as a model of tissue. Using a three-dimensional collagen assay chemokinetic and chemotactic migration induced by CXCL8 was inhibited with the pan PI3 Kinase inhibitor wortmannin. Analysis of the specific Class I PI3 Kinase catalytic isoforms alpha, delta and gamma using the inhibitors PIK-75, PIK-294 and AS-605240 respectively indicated differential roles in CXCL8-induced neutrophil migration. PIK-294 inhibited both chemokinetic and chemotactic CXCL8-induced migration. AS-605240 markedly reduced CXCL8 induced chemokinetic migration but had no effect on CXCL8 induced chemotactic migration. In contrast PIK-75 inhibited chemotactic migration but not chemokinetic migration. At optimal concentrations of GM-CSF the inhibitors had no effect on the percentage of neutrophil migration in comparison to the control however at suboptimal concentrations wortmannin, AS-605240 and PIK-294 inhibited chemokinesis. This study suggests that PI3 Kinase is necessary for CXCL8 induced migration in a 3D tissue environment but that chemokinetic and chemotactic migration may be controlled by different isoforms with gamma shown to be important in chemokinesis and alpha important in chemotaxis. Neutrophil migration in response to suboptimal concentrations of GM-CSF is dependent on PI3 Kinase, particularly the gamma and delta catalytic isoforms.

  4. Tensin 2 modulates cell contractility in 3D collagen gels through the RhoGAP DLC1.

    PubMed

    Clark, Katherine; Howe, Jonathan D; Pullar, Christine E; Green, J Angelo; Artym, Vira V; Yamada, Kenneth M; Critchley, David R

    2010-03-01

    Cytoskeletal proteins of the tensin family couple integrins to the actin cytoskeleton. They are found in both focal adhesions and the fibrillar adhesions formed between cells and the fibronectin matrix. There are four tensin genes which encode three large (approximately 200 kDa) tensin isoforms (tensin 1, 2, 3) and one short isoform (cten). However, the subcellular localization and function of the individual isoforms is poorly understood. Using human foreskin fibroblasts (HFFs), and imaging on both fixed and live cells, we show that GFP-tensin 2 is enriched in dynamic focal adhesions at the leading edge of the cell, whereas GFP-tensin 3 translocates rearward, and is enriched in fibrillar adhesions. To investigate the possible role of tensins in cell-matrix remodeling, we used siRNAs to knockdown each tensin isoform. We discovered that tensin 2 knockdown significantly reduced the ability of HFFs to contract 3D collagen gels, whilst no effect on fibronectin fibrillogenesis was observed. This inhibition of collagen gel contraction was associated with a substantial reduction in Rho activity, and it was reversed by depletion of DLC1, a RhoGAP that binds to tensin in focal adhesions. These findings suggest that focal adhesion-localized tensin 2 negatively regulates DLC1 to permit Rho-mediated actomyosin contraction and remodeling of collagen fibers.

  5. Toward single cell traction microscopy within 3D collagen matrices

    SciTech Connect

    Hall, Matthew S.; Long, Rong; Feng, Xinzeng; Huang, YuLing; Hui, Chung-Yuen; Wu, Mingming

    2013-10-01

    Mechanical interaction between the cell and its extracellular matrix (ECM) regulates cellular behaviors, including proliferation, differentiation, adhesion, and migration. Cells require the three-dimensional (3D) architectural support of the ECM to perform physiologically realistic functions. However, current understanding of cell–ECM and cell–cell mechanical interactions is largely derived from 2D cell traction force microscopy, in which cells are cultured on a flat substrate. 3D cell traction microscopy is emerging for mapping traction fields of single animal cells embedded in either synthetic or natively derived fibrous gels. We discuss here the development of 3D cell traction microscopy, its current limitations, and perspectives on the future of this technology. Emphasis is placed on strategies for applying 3D cell traction microscopy to individual tumor cell migration within collagen gels. - Highlights: • Review of the current state of the art in 3D cell traction force microscopy. • Bulk and micro-characterization of remodelable fibrous collagen gels. • Strategies for performing 3D cell traction microscopy within collagen gels.

  6. Towards Single Cell Traction Microscopy within 3D Collagen Matrices

    PubMed Central

    Hall, Matthew S.; Long, Rong; Feng, Xinzeng; Huang, YuLing; Hui, Chung-Yuen; Wu, Mingming

    2013-01-01

    Mechanical interaction between the cell and its extracellular matrix (ECM) regulates cellular behaviors, including proliferation, differentiation, adhesion, and migration. Cells require the three dimensional (3D) architectural support of the ECM to perform physiologically realistic functions. However, current understanding of cell-ECM and cell-cell mechanical interactions is largely derived from 2D cell traction force microscopy, in which cells are cultured on a flat substrate. 3D cell traction microscopy is emerging for mapping traction fields of single animal cells embedded in either synthetic or natively derived fibrous gels. We discuss here the development of 3D cell traction microscopy, its current limitations, and perspectives on the future of this technology. Emphasis is placed on strategies for applying 3D cell traction microscopy to individual tumor cells migration within collagen gels. PMID:23806281

  7. Uncertainty in 3D gel dosimetry

    NASA Astrophysics Data System (ADS)

    De Deene, Yves; Jirasek, Andrew

    2015-01-01

    Three-dimensional (3D) gel dosimetry has a unique role to play in safeguarding conformal radiotherapy treatments as the technique can cover the full treatment chain and provides the radiation oncologist with the integrated dose distribution in 3D. It can also be applied to benchmark new treatment strategies such as image guided and tracking radiotherapy techniques. A major obstacle that has hindered the wider dissemination of gel dosimetry in radiotherapy centres is a lack of confidence in the reliability of the measured dose distribution. Uncertainties in 3D dosimeters are attributed to both dosimeter properties and scanning performance. In polymer gel dosimetry with MRI readout, discrepancies in dose response of large polymer gel dosimeters versus small calibration phantoms have been reported which can lead to significant inaccuracies in the dose maps. The sources of error in polymer gel dosimetry with MRI readout are well understood and it has been demonstrated that with a carefully designed scanning protocol, the overall uncertainty in absolute dose that can currently be obtained falls within 5% on an individual voxel basis, for a minimum voxel size of 5 mm3. However, several research groups have chosen to use polymer gel dosimetry in a relative manner by normalizing the dose distribution towards an internal reference dose within the gel dosimeter phantom. 3D dosimetry with optical scanning has also been mostly applied in a relative way, although in principle absolute calibration is possible. As the optical absorption in 3D dosimeters is less dependent on temperature it can be expected that the achievable accuracy is higher with optical CT. The precision in optical scanning of 3D dosimeters depends to a large extend on the performance of the detector. 3D dosimetry with X-ray CT readout is a low contrast imaging modality for polymer gel dosimetry. Sources of error in x-ray CT polymer gel dosimetry (XCT) are currently under investigation and include inherent

  8. Inhibitory effect of quercetin on epithelial to mesenchymal transition in SK-MEL-28 human melanoma cells defined by in vitro analysis on 3D collagen gels

    PubMed Central

    Patel, Dhairya H; Sharma, Neeti

    2016-01-01

    Considering the emerging concept of complementary and alternative medicine under the paucity of effective treatment for melanoma, we aimed to understand the effect of quercetin (Qu) on collagen I-induced epithelial–mesenchymal transition (EMT) in melanoma cells. To investigate the effect of Qu in melanoma cells, we used multiple methods, including real-time reverse transcription polymerase chain reaction, migration assay, and wound healing assay. We found that EMT was altered by Qu in melanoma cells. Qu-treated cells exhibited decreased migration and invasion activities. Mechanistically, a high expression of epithelial markers and a decrease in the expression of mesenchymal markers were found to be associated with reversal of EMT in melanoma cells. Time-dependent apoptosis was observed in Qu-treated melanoma cells, which was further confirmed by the upregulation in the protein levels of Caspase 3, a proapoptotic marker. Thus, our findings suggest Qu as a promising dietary compound under the new complementary and alternative medicine category of therapeutic drugs in the chemoprevention of melanoma. PMID:27799792

  9. Vinculin is required for cell polarization, migration, and extracellular matrix remodeling in 3D collagen.

    PubMed

    Thievessen, Ingo; Fakhri, Nikta; Steinwachs, Julian; Kraus, Viola; McIsaac, R Scott; Gao, Liang; Chen, Bi-Chang; Baird, Michelle A; Davidson, Michael W; Betzig, Eric; Oldenbourg, Rudolf; Waterman, Clare M; Fabry, Ben

    2015-11-01

    Vinculin is filamentous (F)-actin-binding protein enriched in integrin-based adhesions to the extracellular matrix (ECM). Whereas studies in 2-dimensional (2D) tissue culture models have suggested that vinculin negatively regulates cell migration by promoting cytoskeleton-ECM coupling to strengthen and stabilize adhesions, its role in regulating cell migration in more physiologic, 3-dimensional (3D) environments is unclear. To address the role of vinculin in 3D cell migration, we analyzed the morphodynamics, migration, and ECM remodeling of primary murine embryonic fibroblasts (MEFs) with cre/loxP-mediated vinculin gene disruption in 3D collagen I cultures. We found that vinculin promoted 3D cell migration by increasing directional persistence. Vinculin was necessary for persistent cell protrusion, cell elongation, and stable cell orientation in 3D collagen, but was dispensable for lamellipodia formation, suggesting that vinculin-mediated cell adhesion to the ECM is needed to convert actin-based cell protrusion into persistent cell shape change and migration. Consistent with this finding, vinculin was necessary for efficient traction force generation in 3D collagen without affecting myosin II activity and promoted 3D collagen fiber alignment and macroscopical gel contraction. Our results suggest that vinculin promotes directionally persistent cell migration and tension-dependent ECM remodeling in complex 3D environments by increasing cell-ECM adhesion and traction force generation.

  10. Microstructural and Mechanical Differences Between Digested Collagen-Fibrin Co-Gels and Pure Collagen and Fibrin Gels

    PubMed Central

    Lai, Victor K.; Frey, Christina R.; Kerandi, Allan M.; Lake, Spencer P.; Tranquillo, Robert T.; Barocas, Victor H.

    2012-01-01

    Collagen and fibrin are important extra-cellular matrix (ECM) components in the body, providing structural integrity to various tissues. These biopolymers are also common scaffolds used in tissue engineering. This study investigated how co-gelation of collagen and fibrin affected the properties of each individual protein network. Collagen-fibrin co-gels were cast and subsequently digested using either plasmin or collagenase; the microstructure and mechanical behavior of the resulting networks were then compared with respective pure collagen or fibrin gels of the same protein concentration. The morphologies of the collagen networks were further analyzed via 3-D network reconstruction from confocal image z-stacks. Both collagen and fibrin exhibited a decrease in mean fiber diameter when formed in the co-gels compared to the pure gels; this microstructural change was accompanied by increased failure strain and decreased tangent modulus for both collagen and fibrin following selected digestion of the co-gels. In addition, analysis of the reconstructed collagen networks indicated presence of very long fibers and clustering of fibrils, resulting in very high connectivities for collagen networks formed in co-gels. PMID:22828381

  11. 3D gel printing for soft-matter systems innovation

    NASA Astrophysics Data System (ADS)

    Furukawa, Hidemitsu; Kawakami, Masaru; Gong, Jin; Makino, Masato; Kabir, M. Hasnat; Saito, Azusa

    2015-04-01

    In the past decade, several high-strength gels have been developed, especially from Japan. These gels are expected to use as a kind of new engineering materials in the fields of industry and medical as substitutes to polyester fibers, which are materials of artificial blood vessels. We consider if various gel materials including such high-strength gels are 3D-printable, many new soft and wet systems will be developed since the most intricate shape gels can be printed regardless of the quite softness and brittleness of gels. Recently we have tried to develop an optical 3D gel printer to realize the free-form formation of gel materials. We named this apparatus Easy Realizer of Soft and Wet Industrial Materials (SWIM-ER). The SWIM-ER will be applied to print bespoke artificial organs, including artificial blood vessels, which will be possibly used for both surgery trainings and actual surgery. The SWIM-ER can print one of the world strongest gels, called Double-Network (DN) gels, by using UV irradiation through an optical fiber. Now we also are developing another type of 3D gel printer for foods, named E-Chef. We believe these new 3D gel printers will broaden the applications of soft-matter gels.

  12. Heterogeneous force network in 3D cellularized collagen networks.

    PubMed

    Liang, Long; Jones, Christopher; Chen, Shaohua; Sun, Bo; Jiao, Yang

    2016-10-25

    Collagen networks play an important role in coordinating and regulating collective cellular dynamics via a number of signaling pathways. Here, we investigate the transmission of forces generated by contractile cells in 3D collagen-I networks. Specifically, the graph (bond-node) representations of collagen networks with collagen concentrations of 1, 2 and 4 mg ml(-1) are derived from confocal microscopy data and used to model the network microstructure. Cell contraction is modeled by applying correlated displacements at specific nodes of the network, representing the focal adhesion sites. A nonlinear elastic model is employed to characterize the mechanical behavior of individual fiber bundles including strain hardening during stretching and buckling under compression. A force-based relaxation method is employed to obtain equilibrium network configurations under cell contraction. We find that for all collagen concentrations, the majority of the forces are carried by a small number of heterogeneous force chains emitted from the contracting cells, which is qualitatively consistent with our experimental observations. The force chains consist of fiber segments that either possess a high degree of alignment before cell contraction or are aligned due to fiber reorientation induced by cell contraction. The decay of the forces along the force chains is significantly slower than the decay of radially averaged forces in the system, suggesting that the fibreous nature of biopolymer network structure can support long-range force transmission. The force chains emerge even at very small cell contractions, and the number of force chains increases with increasing cell contraction. At large cell contractions, the fibers close to the cell surface are in the nonlinear regime, and the nonlinear region is localized in a small neighborhood of the cell. In addition, the number of force chains increases with increasing collagen concentration, due to the larger number of focal adhesion sites

  13. Heterogeneous force network in 3D cellularized collagen networks

    NASA Astrophysics Data System (ADS)

    Liang, Long; Jones, Christopher; Chen, Shaohua; Sun, Bo; Jiao, Yang

    2016-12-01

    Collagen networks play an important role in coordinating and regulating collective cellular dynamics via a number of signaling pathways. Here, we investigate the transmission of forces generated by contractile cells in 3D collagen-I networks. Specifically, the graph (bond-node) representations of collagen networks with collagen concentrations of 1, 2 and 4 mg ml-1 are derived from confocal microscopy data and used to model the network microstructure. Cell contraction is modeled by applying correlated displacements at specific nodes of the network, representing the focal adhesion sites. A nonlinear elastic model is employed to characterize the mechanical behavior of individual fiber bundles including strain hardening during stretching and buckling under compression. A force-based relaxation method is employed to obtain equilibrium network configurations under cell contraction. We find that for all collagen concentrations, the majority of the forces are carried by a small number of heterogeneous force chains emitted from the contracting cells, which is qualitatively consistent with our experimental observations. The force chains consist of fiber segments that either possess a high degree of alignment before cell contraction or are aligned due to fiber reorientation induced by cell contraction. The decay of the forces along the force chains is significantly slower than the decay of radially averaged forces in the system, suggesting that the fibreous nature of biopolymer network structure can support long-range force transmission. The force chains emerge even at very small cell contractions, and the number of force chains increases with increasing cell contraction. At large cell contractions, the fibers close to the cell surface are in the nonlinear regime, and the nonlinear region is localized in a small neighborhood of the cell. In addition, the number of force chains increases with increasing collagen concentration, due to the larger number of focal adhesion sites

  14. A novel ultrathin collagen nanolayer assembly for 3-D microtissue engineering: Layer-by-layer collagen deposition for long-term stable microfluidic hepatocyte culture.

    PubMed

    McCarty, William J; Usta, O Berk; Luitje, Martha; Bale, Shyam Sundhar; Bhushan, Abhinav; Hegde, Manjunath; Golberg, Inna; Jindal, Rohit; Yarmush, Martin L

    2014-03-01

    The creation of stable hepatocyte cultures using cell-matrix interactions has proven difficult in microdevices due to dimensional constraints limiting the utility of classic tissue culture techniques that involve the use of hydrogels such as the collagen "double gel" or "overlay". To translate the collagen overlay technique into microdevices, we modified collagen using succinylation and methylation reactions to create polyanionic and polycationic collagen solutions, and deposited them layer-by-layer to create ultrathin collagen nanolayers on hepatocytes. These ultrathin collagen layers covered hepatocytes in microdevices and 1) maintained cell morphology, viability, and polarity, 2) induced bile canalicular formation and actin reorganization, and 3) maintained albumin and urea secretions and CYP activity similar to those observed in hepatocytes in collagen double gel hepatocytes in plate cultures. Beyond the immediate applications of this technique to create stable, in vitro microfluidic hepatocyte cultures for drug toxicity testing, this technique is generally applicable as a thin biomaterial for other 3D microtissues.

  15. Breast epithelial tissue morphology is affected in 3D cultures by species-specific collagen-based extracellular matrix.

    PubMed

    Dhimolea, Eugen; Soto, Ana M; Sonnenschein, Carlos

    2012-11-01

    Collagen-based gels have been widely used to determine the factors that regulate branching morphogenesis in the mammary gland. The patterns of biomechanical gradients and collagen reorganization influence the shape and orientation of epithelial structures in three-dimensional (3D) conditions. We explored in greater detail whether collagen type I fibers with distinct biomechanical and fiber-assembling properties, isolated from either bovine or rat tail tendon, differentially affected the epithelial phenotype in a tissue culture model of the human breast. Rat tail collagen fibers were densely packed into significantly longer and thicker bundles compared to those of the bovine type (average fascicle length 7.35 and 2.29 μm, respectively; p = 0.0001), indicating increased fiber alignment and biomechanical enablement in the former. MCF10A epithelial cells formed elaborated branched tubular structures in bovine but only nonbranched ducts and acini in rat tail collagen matrices. Ductal branching in bovine collagen was associated with interactions between neighboring structures mediated through packed collagen fibers; these fiber-mediated interactions were absent in rat tail collagen gels. Normal breast fibroblasts increased the final size and number of ducts only in rat tail collagen gels while not affecting branching. Our results suggest that the species of origin of collagen used in organotypic cultures may influence epithelial differentiation into alveolar or ductal structures and the patterns of epithelial branching. These observations underscore the importance of considering the species of origin and fiber alignment properties of collagen when engineering branching organs in 3D matrices and interpreting their role in the tissue phenotype.

  16. 3D jet printer of edible gels for food creation

    NASA Astrophysics Data System (ADS)

    Serizawa, Ryo; Shitara, Mariko; Gong, Jin; Makino, Masato; Kabir, M. Hasnat; Furukawa, Hidemitsu

    2014-03-01

    In recent years, aging is progressing in Japan. Elderly people can't swallow the food well. So, the need of soft food is increasing greatly with the aging of the population. There are so few satisfying foods for the elderly to enjoy a meal. An equipment of printing soft food gives the elderly a big dream and is promising. In this study, we aim at developing a 3D edible gel printer in order to make soft food for the elderly. We made a prototype of the 3D edible gel printer. The printer consists of syringe pump and dispenser. The syringe pump extrudes the solution. The dispenser allows to model threedimensional objects. We use agar solution as the ink to carry out the printing. Agar's gelation deeply depends on temperature. Therefore temperature control of the solution is important to mold optimal shapes because the physical crosslinking network of agar's solution is instable. We succeeded in making the gels and plate-shape gel using the 3D edible gel printer. Further more, in order to increase the gelation speed agar's solution, we changed the dispenser and the printing test is being done now. 4 kinds of soft food prepared from agar and gelatin were printed by the 3D edible gel printer. The compression tests of the printed soft food samples were done and their hardness is measured because the hardness is one of very important factors which influence the food texture greatly. In the future, the viscosity of the agar solution or other food ink should be adjusted to suitable for printing.

  17. On the reliability of 3D gel dosimetry

    NASA Astrophysics Data System (ADS)

    De Deene, Y.; Vandecasteele, J.

    2013-06-01

    Gel dosimetry has a unique role to play in safeguarding conformal radiotherapy treatments as it covers the whole treatment chain and provides the radiation oncologist with the integrated dose distribution in 3D. A major obstacle that has hindered the wider dissemination of polymer gel dosimetry in radiotherapy centres is the lack of confidence in the reliability of the measured dose. Discrepancies in dose response of small versus large polymer gel dosimeters have been reported and although several hypothesis for these discrepancies have been postulated, the actual contribution of these error sources to the overall inaccuracy of the dose maps has not been determined. Several gel dosimetry research groups have chosen to use an internal calibration of gel dosimeters. In this study, the inter-and intra-batch reproducibility of the current state-of-the-art 3D gel dosimeters has been assessed. It is demonstrated that with a carefully designed scanning set-up, the overall accuracy that can be obtained with an independent calibration is well within 5% of all pixels.

  18. Discoidin domain receptor 2 regulates the adhesion of fibroblasts to 3D collagen matrices.

    PubMed

    Kim, Daehwan; You, Eunae; Min, Na Young; Lee, Kwang-Ho; Kim, Hyoung Kyu; Rhee, Sangmyung

    2013-05-01

    The collagen matrix constitutes the primary extracellular matrix (ECM) portion of mammalian connective tissues in which the interaction of the cell and the surrounding collagen fibers has a significant impact on cell and tissue physiology, including morphogenesis, development and motility. Discoidin domain receptors (DDR1 and DDR2) have been identified as the receptor tyrosine kinases that are activated upon collagen binding. However, there is a lack of evidence regarding the effect of DDRs on the mechanical interaction between fibroblasts and ECM. In this study, we demonstrated that one of the major phosphotyrosine proteins in human fibroblasts during 3D collagen matrix polymerization is DDR2. Treatment of fibroblasts in 3D collagen matrices with platelet-derived growth factor (PDFG) has been shown to increase DDR2 phosphorylation. Silencing of DDR2 with siRNA in fibroblasts significantly reduced the number of dendritic extensions regardless of whether cells were cultured in the collagen or fibronectin 3D matrices. Decreasing dendritic extensions in DDR2-silenced cells has also been shown to decrease the ability of fibroblast entanglement to collagen fibrils in 3D collagen matrices. Finally, we also showed that the silencing of DDR2 decreased the cell migration in 3D nested collagen matrices but had no effect on 3D floating matrix contraction. Collectively, these results suggest that DDR2 functioning is required for the membrane dynamics to control the mechanical attachment of fibroblasts to the 3D collagen matrices in an integrin-independent manner.

  19. GEM printer: 3D gel printer for free shaping of functional gel engineering materials

    NASA Astrophysics Data System (ADS)

    Furukawa, Hidemitsu; Muroi, Hisato; Yamamoto, Kouki; Serizawa, Ryo; Gong, Jin

    2013-04-01

    In the past decade, several high-strength gels have been developed. These gels are expected to use as a kind of new engineering materials in the fields of industry and medical as substitutes to polyester fibers, which are materials of artificial blood vessels. The gels have both low surface friction and well permeability due to a large amount of water absorbed in the gels, which are superiority of the gels compering to the polyester fibers. It is, however, difficult for gels to be forked structure or cavity structure by using cutting or mold. Consequently, it is necessary to develop the additive manufacturing device to synthesize and mode freely gels at the same time. Here we try to develop an optical 3D gel printer that enables gels to be shaped precisely and freely. For the free forming of high-strength gels, the 1st gels are ground to particles and mixed with 2nd pregel solution, and the mixed solution is gelled by the irradiation of UV laser beam through an optical fiber. The use of the optical fiber makes one-point UV irradiation possible. Since the optical fiber is controlled by 3D-CAD, the precise and free molding in XYZ directions is easily realized. We successfully synthesized tough gels using the gel printer.

  20. Free forming of the gel by 3D gel printer SWIM-ER

    NASA Astrophysics Data System (ADS)

    Okada, Koji; Tase, Taishi; Saito, Azusa; Makino, Masato; Gong, Jin; Kawakami, Masaru; Furukawa, Hidemitsu

    2015-04-01

    Gels, soft and wet materials, have unique properties such as material permeability, biocompatibility and low friction, which are hardly found in hard and dry materials. These superior characteristics of hydrogels promise to expand the medical applications. In recent years, the optical 3D gel printer named SWIM-ER (Soft and Wet Industrial - Easy Realizer) was developed by our team in order to fabricate tough gels with free form. We are aiming to create artificial blood vessel of the gel material by 3D gel printer. Artificial blood vessel is expected to be used for vascular surgery practice. The artificial blood vessel made by 3D gel printer can be create to free form on the basis of the biological data of the patient. Therefore, we believe it is possible to contribute to increasing the success rate and safety of vascular surgery by creating artificial blood vessel with 3D gel printer. The modeling method of SWIM-ER is as follow. Pregel solution is polymerized by one-point UV irradiation with optical fiber. The irradiation area is controlled by computer program, so that exact 3D free forming is realized. In this study, synthesis conditions are re-examined in order to improve the degree of freedom of fabrication. The dimensional accuracy in height direction is improved by increasing the cross linker concentration. We examined the relationship of resolution to the pitch and UV irradiation time in order to improve the modeling accuracy.

  1. Optically characterizing collagen gels made with different cell types

    NASA Astrophysics Data System (ADS)

    Levitz, David; Choudhury, Niloy; Vartanian, Keri; Hinds, Monica T.; Hanson, Stephen R.; Jacques, Steven L.

    2009-02-01

    The ability of optical imaging techniques such as optical coherence tomography (OCT) to non-destructively characterize tissue-engineered constructs has generated enormous interest recently. Collagen gels are 3D structures that represent a simple common model of many engineered tissues that contain 2 primary scatterers: collagen and cells. We are testing the ability of OCT data to characterize the remodeling of such collagen-based constructs by 3 different types of cells: vascular smooth muscle cells (SMCs), endothelial cells (ECs), and osteoblasts (OBs). Collagen gels were prepared with SMCs, ECs, and OBs with a seeding density of 1×106 cells/ml; additionally, acellular controls were also prepared. The disk-shaped constructs were allowed to remodel in the incubator for 5 days, with OCT imaging occurring on days 1 and 5. From the OCT data, the attenuation and reflectivity were evaluated by fitting the data to a theoretical model that relates the tissue optical properties (scattering coefficient and anisotropy factor) and imaging conditions to the OCT signal. The degree of gel compaction was determined from the volume of the culture medium that feeds the constructs. We found that gel compaction (relative to the acellular control) occurred in the SMC constructs, but not in the OB or EC constructs. The optical property data showed that at day 5 the SMC constructs had an overall higher reflectivity (lower g) relative to day 1, whereas there was no obvious change in reflectivity of the EC, OB constructs and acellular controls relative to day 1. Moreover, there was a difference in the attenuation of the OB constructs on day 5 relative to day 1, but not in the other constructs. The apparent decrease in anisotropy observed in the SMC constructs, but not in the OB and EC constructs and acellular controls, suggests that OCT is sensitive to the remodeling of the collagen matrix that accompanies gel compaction, and can offer highly localized information on the construct

  2. 3D Gel Map of Arabidopsis Complex I

    PubMed Central

    Peters, Katrin; Belt, Katharina; Braun, Hans-Peter

    2013-01-01

    Complex I has a unique structure in plants and includes extra subunits. Here, we present a novel study to define its protein constituents. Mitochondria were isolated from Arabidopsis thaliana cell cultures, leaves, and roots. Subunits of complex I were resolved by 3D blue-native (BN)/SDS/SDS-PAGE and identified by mass spectrometry. Overall, 55 distinct proteins were found, seven of which occur in pairs of isoforms. We present evidence that Arabidopsis complex I consists of 49 distinct types of subunits, 40 of which represent homologs of bovine complex I. The nine other subunits represent special proteins absent in the animal linage of eukaryotes, most prominently a group of subunits related to bacterial gamma-type carbonic anhydrases. A GelMap http://www.gelmap.de/arabidopsis-3d-complex-i/ is presented for promoting future complex I research in Arabidopsis thaliana. PMID:23761796

  3. Microfluidic vascular channels in gels using commercial 3D printers

    NASA Astrophysics Data System (ADS)

    Selvaganapathy, P. Ravi; Attalla, Rana

    2016-03-01

    This paper details the development of a three dimensional (3D) printing system with a modified microfluidic printhead used for the generation of complex vascular tissue scaffolds. The print-head features an integrated coaxial nozzle that allows the fabrication of hollow, calcium-polymerized alginate tubes that can easily be patterned using 3Dbioprinting techniques. This microfluidic design allows the incorporation of a wide range of scaffold materials as well as biological constituents such as cells, growth factors, and ECM material. With this setup, gel constructs with embedded arrays of hollow channels can be created and used as a potential substitute for blood vessel networks.

  4. Establishment of gel materials with different mechanical properties by 3D gel printer SWIM-ER

    NASA Astrophysics Data System (ADS)

    Ota, Takafumi; Tase, Taishi; Okada, Koji; Saito, Azusa; Takamatsu, Kyuuichiro; Kawakami, Masaru; Furukawa, Hidemitsu

    2016-04-01

    A 3D printer is a device which can directly produce objects whose shape is the same as the original 3D digital data. Hydrogels have unique properties such as high water content, low frictional properties, biocompatibility, material permeability and high transparency, which are rare in hard and dry materials. These superior characteristics of gels promise useful medical applications. We have been working on the development of a 3D gel printer, SWIM-ER (Soft and Wet Industrial - Easy Realizer), which can make models of organs and artificial blood vessels with gel material. However, 3D printing has a problem: the mechanical properties of the printed object vary depending on printing conditions, and this matter was investigated with SWIM-ER. In the past, we found that mechanical properties of 3D gel objects depend on the deposition orientation in SWIM-ER. In this study, gels were printed with different laser scanning speeds. The mechanical properties of these gels were investigated by compression tests, water content measurements and SMILS (Scanning Microscopic Light Scattering).

  5. A novel ultrathin collagen nanolayer assembly for 3-D microtissue engineering: Layer-by-layer collagen deposition for long-term stable microfluidic hepatocyte culture

    PubMed Central

    McCarty, William J.; Usta, O. Berk; Luitje, Martha; Bale, Shyam Sundhar; Bhushan, Abhinav; Hegde, Manjunath; Golberg, Inna; Jindal, Rohit; Yarmush, Martin L.

    2014-01-01

    The creation of stable hepatocyte cultures using cell-matrix interactions has proven difficult in microdevices due to dimensional constraints limiting the utility of classic tissue culture techniques that involve the use of hydrogels such as the collagen “double gel” or “overlay”. To translate the collagen overlay technique into microdevices, we modified collagen using succinylation and methylation reactions to create polyanionic and polycationic collagen solutions, and deposited them layer-by-layer to create ultrathin collagen nanolayers on hepatocytes. These ultrathin collagen layers covered hepatocytes in microdevices and 1) maintained cell morphology, viability, and polarity, 2) induced bile canalicular formation and actin reorganization, and 3) maintained albumin and urea secretions and CYP activity similar to those observed in hepatocytes in collagen double gel hepatocytes in plate cultures. Beyond the immediate applications of this technique to create stable, in vitro microfluidic hepatocyte cultures for drug toxicity testing, this technique is generally applicable as a thin biomaterial for other 3D microtissues. PMID:24932459

  6. Enhancement of neurite outgrowth in neuron cancer stem cells by growth on 3-D collagen scaffolds

    SciTech Connect

    Chen, Chih-Hao; Kuo, Shyh Ming; Liu, Guei-Sheung; Chen, Wan-Nan U.; Chuang, Chin-Wen; Liu, Li-Feng

    2012-11-09

    Highlights: Black-Right-Pointing-Pointer Neuron cancer stem cells (NCSCs) behave high multiply of growth on collagen scaffold. Black-Right-Pointing-Pointer Enhancement of NCSCs neurite outgrowth on porous collagen scaffold. Black-Right-Pointing-Pointer 3-D collagen culture of NCSCs shows an advance differentiation than 2-D culture. -- Abstract: Collagen is one component of the extracellular matrix that has been widely used for constructive remodeling to facilitate cell growth and differentiation. The 3-D distribution and growth of cells within the porous scaffold suggest a clinical significance for nerve tissue engineering. In the current study, we investigated proliferation and differentiation of neuron cancer stem cells (NCSCs) on a 3-D porous collagen scaffold that mimics the natural extracellular matrix. We first generated green fluorescence protein (GFP) expressing NCSCs using a lentiviral system to instantly monitor the transitions of morphological changes during growth on the 3-D scaffold. We found that proliferation of GFP-NCSCs increased, and a single cell mass rapidly grew with unrestricted expansion between days 3 and 9 in culture. Moreover, immunostaining with neuronal nuclei (NeuN) revealed that NCSCs grown on the 3-D collagen scaffold significantly enhanced neurite outgrowth. Our findings confirmed that the 80 {mu}m porous collagen scaffold could enhance attachment, viability and differentiation of the cancer neural stem cells. This result could provide a new application for nerve tissue engineering and nerve regeneration.

  7. Highlighting the impact of aging on type I collagen: label-free investigation using confocal reflectance microscopy and diffuse reflectance spectroscopy in 3D matrix model.

    PubMed

    Guilbert, Marie; Roig, Blandine; Terryn, Christine; Garnotel, Roselyne; Jeannesson, Pierre; Sockalingum, Ganesh D; Manfait, Michel; Perraut, François; Dinten, Jean-Marc; Koenig, Anne; Piot, Olivier

    2016-02-23

    During aging, alterations of extracellular matrix proteins contribute to various pathological phenotypes. Among these alterations, type I collagen cross-linking and associated glycation products accumulation over time detrimentally affects its physico-chemical properties, leading to alterations of tissue biomechanical stability. Here, different-age collagen 3D matrices using non-destructive and label-free biophotonic techniques were analysed to highlight the impact of collagen I aging on 3D constructs, at macroscopic and microscopic levels. Matrices were prepared with collagens extracted from tail tendons of rats (newborns, young and old adults) to be within the physiological aging process. The data of diffuse reflectance spectroscopy reveal that aging leads to an inhibition of fibril assembly and a resulting decrease of gel density. Investigations by confocal reflectance microscopy highlight poor-fibrillar structures in oldest collagen networks most likely related to the glycation products accumulation. Complementarily, an infrared analysis brings out marked spectral variations in the Amide I profile, specific of the peptidic bond conformation and for carbohydrates vibrations as function of collagen-age. Interestingly, we also highlight an unexpected behavior for newborn collagen, exhibiting poorly-organized networks and microscopic features close to the oldest collagen. These results demonstrate that changes in collagen optical properties are relevant for investigating the incidence of aging in 3D matrix models.

  8. Highlighting the impact of aging on type I collagen: label-free investigation using confocal reflectance microscopy and diffuse reflectance spectroscopy in 3D matrix model

    PubMed Central

    Terryn, Christine; Garnotel, Roselyne; Jeannesson, Pierre; Sockalingum, Ganesh D.; Manfait, Michel; Perraut, François; Dinten, Jean-Marc; Koenig, Anne; Piot, Olivier

    2016-01-01

    During aging, alterations of extracellular matrix proteins contribute to various pathological phenotypes. Among these alterations, type I collagen cross-linking and associated glycation products accumulation over time detrimentally affects its physico-chemical properties, leading to alterations of tissue biomechanical stability. Here, different-age collagen 3D matrices using non-destructive and label-free biophotonic techniques were analysed to highlight the impact of collagen I aging on 3D constructs, at macroscopic and microscopic levels. Matrices were prepared with collagens extracted from tail tendons of rats (newborns, young and old adults) to be within the physiological aging process. The data of diffuse reflectance spectroscopy reveal that aging leads to an inhibition of fibril assembly and a resulting decrease of gel density. Investigations by confocal reflectance microscopy highlight poor-fibrillar structures in oldest collagen networks most likely related to the glycation products accumulation. Complementarily, an infrared analysis brings out marked spectral variations in the Amide I profile, specific of the peptidic bond conformation and for carbohydrates vibrations as function of collagen-age. Interestingly, we also highlight an unexpected behavior for newborn collagen, exhibiting poorly-organized networks and microscopic features close to the oldest collagen. These results demonstrate that changes in collagen optical properties are relevant for investigating the incidence of aging in 3D matrix models. PMID:26885896

  9. Multiphoton crosslinking for biocompatible 3D printing of type I collagen.

    PubMed

    Bell, Alex; Kofron, Matthew; Nistor, Vasile

    2015-09-03

    Multiphoton fabrication is a powerful technique for three-dimensional (3D) printing of structures at the microscale. Many polymers and proteins have been successfully structured and patterned using this method. Type I collagen comprises a large part of the extracellular matrix for most tissue types and is a widely used cellular scaffold material for tissue engineering. Current methods for creating collagen tissue scaffolds do not allow control of local geometry on a cellular scale. This means the environment experienced by cells may be made up of the native material but unrelated to native cellular-scale structure. In this study, we present a novel method to allow multiphoton crosslinking of type I collagen with flavin mononucleotide photosensitizer. The method detailed allows full 3D printing of crosslinked structures made from unmodified type I collagen and uses only demonstrated biocompatible materials. Resolution of 1 μm for both standing lines and high-aspect ratio gaps between structures is demonstrated and complex 3D structures are fabricated. This study demonstrates a means for 3D printing with one of the most widely used tissue scaffold materials. High-resolution, 3D control of the fabrication of collagen scaffolds will facilitate higher fidelity recreation of the native extracellular environment for engineered tissues.

  10. Synthesis of highly interconnected 3D scaffold from Arothron stellatus skin collagen for tissue engineering application.

    PubMed

    Ramanathan, Giriprasath; Singaravelu, Sivakumar; Raja, M D; Sivagnanam, Uma Tiruchirapalli

    2015-11-01

    The substrate which is avidly used for tissue engineering applications should have good mechanical and biocompatible properties, and all these parameters are often considered as essential for dermal reformation. Highly interconnected three dimensional (3D) wound dressing material with enhanced structural integrity was synthesized from Arothron stellatus fish skin (AsFS) collagen for tissue engineering applications. The synthesized 3D collagen sponge (COL-SPG) was further characterized by different physicochemical methods. The scanning electron microscopy analysis of the material demonstrated that well interconnected pores with homogeneous microstructure on the surface aids higher swelling index and that the material also possessed good mechanical properties with a Young's modulus of 0.89±0.2 MPa. Biocompatibility of the 3D COL-SPG showed 92% growth for both NIH 3T3 fibroblasts and keratinocytes. Overall, the study revealed that synthesized 3D COL-SPG from fish skin will act as a promising wound dressing in skin tissue engineering.

  11. Fabrication of high-density collagen fibril matrix gels by renaturation of triple-helix collagen from gelatin.

    PubMed

    Ohyabu, Yoshimi; Yunoki, Shunji; Hatayama, Hirosuke; Teranishi, Yoshikazu

    2013-11-01

    Collagen-based 3-D hydrogels often lack sufficient mechanical strength for tissue engineering. We developed a method for fabrication of high-density collagen fibril matrix (CFM) gels from concentrated solutions of uncleaved gelatin (UCG). Denatured random-coil UCG exhibited more rapid and efficient renaturation into collagen triple-helix than cleaved gelatin (CG) over a broad range of setting temperatures. The UCG solution formed opaque gels with high-density reconstituted collagen fibrils at 28-32 °C and transparent gels similar to CG at <25 °C. The unique gelation properties of UCG enabled the encapsulation of cultured cells in CFM of high solid volume (>5%) and elasticity (1.28 ± 0.15 kPa at 5% and 4.82 ± 0.38 kPa at 8%) with minimal cell loss. The elastic modulus of these gels was higher than that of conventional CFM containing 0.5% collagen. High-strength CFM may provide more durable hydrogels for tissue engineering and regenerative medicine.

  12. Physical and chemical modifications of collagen gels: impact on diffusion.

    PubMed

    Erikson, Arne; Andersen, Hilde Nortvedt; Naess, Stine Nalum; Sikorski, Pawel; Davies, Catharina de Lange

    2008-02-01

    The extracellular matrix (ECM) represents a major barrier for delivery of therapeutic drugs, and the transport is determined by the ECM composition, structure, and distribution. Because of the high interstitial fluid pressure in tumors, diffusion becomes the main transport mechanism through ECM. The purpose of this work was to study the impact of the structure of the collagen network on diffusion, by studying to what extent the orientation and chemical modification of the collagen network influenced diffusion. Collagen gels with a concentration of 0.2-2.0% that is comparable with the amount of collagen in the tumor ECM were used as a model system for ECM. Collagen gels were aligned in a low-strength magnetic field and geometrical confinement, and chemically modified by adding decorin or hyaluronan. Diffusion of dextran 2-MDa molecules in the collagen gels was measured using fluorescence recovery after photobleaching. Alignment of the collagen fibers in our gels was found to have no impact on the diffusion coefficient. Adding decorin reduced the diameter of the collagen fibers, but no effect on diffusion was observed. Hyaluronan also reduced the fiber diameter, and high concentration of hyaluronan (2.5 mg/ml) increased the diffusion coefficient. The results indicate that the structure of the collagen network is not a major factor in determining the diffusion through the ECM. Rather, increasing the concentration of collagen was found to reduce the diffusion coefficient. Concentration of the collagen network is more important than the structure in determining the diffusion coefficient.

  13. Structural and micromechanical characterization of type I collagen gels.

    PubMed

    Latinovic, Olga; Hough, Lawrence A; Daniel Ou-Yang, H

    2010-02-10

    In this paper we report a study where we use a novel optical tweezers technique to measure the local viscoelastic properties of type I collagen solutions spanning the sol-to-gel transition. We use phase contrast optical microscopy to reveal dense and sparse regions of the rigid fibril networks, and find that the spatial variations in the mechanical properties of the collagen gels closely follow the structural properties. Within the dense phase of the connected network in the gel samples, there are regions that exhibit drastically different viscoelastic properties. Within the sparse regions of the gel samples, no evidence of elasticity is found. In type I collagen gels, we find a high degree of structural inhomogeneity. The inhomogeneity in the structural properties of collagen gels and the corresponding viscoelastic properties provide benchmark measurements for the behavior of desirable biological materials, or tissue equivalents.

  14. 3D printing of composite calcium phosphate and collagen scaffolds for bone regeneration.

    PubMed

    Inzana, Jason A; Olvera, Diana; Fuller, Seth M; Kelly, James P; Graeve, Olivia A; Schwarz, Edward M; Kates, Stephen L; Awad, Hani A

    2014-04-01

    Low temperature 3D printing of calcium phosphate scaffolds holds great promise for fabricating synthetic bone graft substitutes with enhanced performance over traditional techniques. Many design parameters, such as the binder solution properties, have yet to be optimized to ensure maximal biocompatibility and osteoconductivity with sufficient mechanical properties. This study tailored the phosphoric acid-based binder solution concentration to 8.75 wt% to maximize cytocompatibility and mechanical strength, with a supplementation of Tween 80 to improve printing. To further enhance the formulation, collagen was dissolved into the binder solution to fabricate collagen-calcium phosphate composites. Reducing the viscosity and surface tension through a physiologic heat treatment and Tween 80, respectively, enabled reliable thermal inkjet printing of the collagen solutions. Supplementing the binder solution with 1-2 wt% collagen significantly improved maximum flexural strength and cell viability. To assess the bone healing performance, we implanted 3D printed scaffolds into a critically sized murine femoral defect for 9 weeks. The implants were confirmed to be osteoconductive, with new bone growth incorporating the degrading scaffold materials. In conclusion, this study demonstrates optimization of material parameters for 3D printed calcium phosphate scaffolds and enhancement of material properties by volumetric collagen incorporation via inkjet printing.

  15. 3D Printing of Composite Calcium Phosphate and Collagen Scaffolds for Bone Regeneration

    PubMed Central

    Inzana, Jason A.; Olvera, Diana; Fuller, Seth M.; Kelly, James P.; Graeve, Olivia A.; Schwarz, Edward M.; Kates, Stephen L.; Awad, Hani A.

    2014-01-01

    Low temperature 3D printing of calcium phosphate scaffolds holds great promise for fabricating synthetic bone graft substitutes with enhanced performance over traditional techniques. Many design parameters, such as the binder solution properties, have yet to be optimized to ensure maximal biocompatibility and osteoconductivity with sufficient mechanical properties. This study tailored the phosphoric acid-based binder solution concentration to 8.75 wt% to maximize cytocompatibility and mechanical strength, with a supplementation of Tween 80 to improve printing. To further enhance the formulation, collagen was dissolved into the binder solution to fabricate collagen-calcium phosphate composites. Reducing the viscosity and surface tension through a physiologic heat treatment and Tween 80, respectively, enabled reliable thermal inkjet printing of the collagen solutions. Supplementing the binder solution with 1–2 wt% collagen significantly improved maximum flexural strength and cell viability. To assess the bone healing performance, we implanted 3D printed scaffolds into a critically sized murine femoral defect for 9 weeks. The implants were confirmed to be osteoconductive, with new bone growth incorporating the degrading scaffold materials. In conclusion, this study demonstrates optimization of material parameters for 3D printed calcium phosphate scaffolds and enhancement of material properties by volumetric collagen incorporation via inkjet printing. PMID:24529628

  16. Microstructural and mechanical differences between digested collagen-fibrin co-gels and pure collagen and fibrin gels.

    PubMed

    Lai, Victor K; Frey, Christina R; Kerandi, Allan M; Lake, Spencer P; Tranquillo, Robert T; Barocas, Victor H

    2012-11-01

    Collagen and fibrin are important extracellular matrix (ECM) components in the body, providing structural integrity to various tissues. These biopolymers are also common scaffolds used in tissue engineering. This study investigated how co-gelation of collagen and fibrin affected the properties of each individual protein network. Collagen-fibrin co-gels were cast and subsequently digested using either plasmin or collagenase; the microstructure and mechanical behavior of the resulting networks were then compared with the respective pure collagen or fibrin gels of the same protein concentration. The morphologies of the collagen networks were further analyzed via three-dimensional network reconstruction from confocal image z-stacks. Both collagen and fibrin exhibited a decrease in mean fiber diameter when formed in co-gels compared with the pure gels. This microstructural change was accompanied by an increased failure strain and decreased tangent modulus for both collagen and fibrin following selective digestion of the co-gels. In addition, analysis of the reconstructed collagen networks indicated the presence of very long fibers and the clustering of fibrils, resulting in very high connectivities for collagen networks formed in co-gels.

  17. Concentric gel system to study the biophysical role of matrix microenvironment on 3D cell migration.

    PubMed

    Kurniawan, Nicholas Agung; Chaudhuri, Parthiv Kant; Lim, Chwee Teck

    2015-04-03

    The ability of cells to migrate is crucial in a wide variety of cell functions throughout life from embryonic development and wound healing to tumor and cancer metastasis. Despite intense research efforts, the basic biochemical and biophysical principles of cell migration are still not fully understood, especially in the physiologically relevant three-dimensional (3D) microenvironments. Here, we describe an in vitro assay designed to allow quantitative examination of 3D cell migration behaviors. The method exploits the cell's mechanosensing ability and propensity to migrate into previously unoccupied extracellular matrix (ECM). We use the invasion of highly invasive breast cancer cells, MDA-MB-231, in collagen gels as a model system. The spread of cell population and the migration dynamics of individual cells over weeks of culture can be monitored using live-cell imaging and analyzed to extract spatiotemporally-resolved data. Furthermore, the method is easily adaptable for diverse extracellular matrices, thus offering a simple yet powerful way to investigate the role of biophysical factors in the microenvironment on cell migration.

  18. Concentric Gel System to Study the Biophysical Role of Matrix Microenvironment on 3D Cell Migration

    PubMed Central

    Kurniawan, Nicholas Agung; Chaudhuri, Parthiv Kant; Lim, Chwee Teck

    2015-01-01

    The ability of cells to migrate is crucial in a wide variety of cell functions throughout life from embryonic development and wound healing to tumor and cancer metastasis. Despite intense research efforts, the basic biochemical and biophysical principles of cell migration are still not fully understood, especially in the physiologically relevant three-dimensional (3D) microenvironments. Here, we describe an in vitro assay designed to allow quantitative examination of 3D cell migration behaviors. The method exploits the cell’s mechanosensing ability and propensity to migrate into previously unoccupied extracellular matrix (ECM). We use the invasion of highly invasive breast cancer cells, MDA-MB-231, in collagen gels as a model system. The spread of cell population and the migration dynamics of individual cells over weeks of culture can be monitored using live-cell imaging and analyzed to extract spatiotemporally-resolved data. Furthermore, the method is easily adaptable for diverse extracellular matrices, thus offering a simple yet powerful way to investigate the role of biophysical factors in the microenvironment on cell migration. PMID:25867104

  19. Hydroxyapatite incorporated into collagen gels for mesenchymal stem cell culture.

    PubMed

    Laydi, F; Rahouadj, R; Cauchois, G; Stoltz, J-F; de Isla, N

    2013-01-01

    Collagen gels could be used as carriers in tissue engineering to improve cell retention and distribution in the defect. In other respect hydroxyapatite could be added to gels to improve mechanical properties and regulate gel contraction. The aim of this work was to analyze the feasibility to incorporate hydroxyapatite into collagen gels and culture mesenchymal stem cells inside it. Human bone marrow mesenchymal stem cells (hMSC-BM) were used in this study. Gels were prepared by mixing rat tail type I collagen, hydroxyapatite microparticles and MSCs. After polymerization gels were kept in culture while gel contraction and mechanical properties were studied. In parallel, cell viability and morphology were analyzed. Gels became free-floating gels contracted from day 3, only in the presence of cells. A linear rapid contraction phase was observed until day 7, then a very slow contraction phase took place. The incorporation of hydroxyapatite improved gel stability and mechanical properties. Cells were randomly distributed on the gel and a few dead cells were observed all over the experiment. This study shows the feasibility and biocompatibility of hydroxyapatite supplemented collagen gels for the culture of mesenchymal stem cells that could be used as scaffolds for cell delivery in osteoarticular regenerative medicine.

  20. Collagen telopeptides (cross-linking sites) play a role in collagen gel lattice contraction

    NASA Technical Reports Server (NTRS)

    Woodley, D. T.; Yamauchi, M.; Wynn, K. C.; Mechanic, G.; Briggaman, R. A.

    1991-01-01

    Solubilized interstitial collagens will form a fibrillar, gel-like lattice when brought to physiologic conditions. In the presence of human dermal fibroblasts the collagen lattice will contract. The rate of contraction can be determined by computer-assisted planemetry. The mechanisms involved in contraction are as yet unknown. Using this system it was found that the rate of contraction was markedly decreased when collagen lacking telopeptides was substituted for native collagen. Histidinohydroxylysinonorleucine (HHL) is a major stable trifunctional collagen cross-link in mature skin that involves a carboxyl terminal, telopeptide site 16c, the sixteenth amino acid residue from the carboxy terminal of the telopeptide region of alpha 1 (I) in type I collagen. Little, if any, HHL was present in native, purified, reconstituted, soluble collagen fibrils from 1% acetic acid-extracted 2-year-old bovine skin. In contrast, HHL cross-links were present (0.22 moles of cross-link per mole of collagen) in lattices of the same collagen contracted by fibroblasts. However, rat tail tendon does not contain HHL cross-links, and collagen lattices made of rat tail tendon collagen are capable of contraction. This suggests that telopeptide sites, and not mature HHL cross-links per se, are essential for fibroblasts to contract collagen lattices. Beta-aminopropionitrile fumarate (BAPN), a potent lathyrogen that perturbs collagen cross-linking by inhibition of lysyl oxidase, also inhibited the rate of lattice cell contraction in lattices composed of native collagen. However, the concentrations of BAPN that were necessary to inhibit the contraction of collagen lattices also inhibited fibroblast growth suggestive of cellular toxicity. In accordance with other studies, we found no inhibition of the rate of lattice contraction when fibronectin-depleted serum was used. Electron microscopy of contracted gels revealed typical collagen fibers with a characteristic axial periodicity. The data

  1. 3D in vitro bioengineered tumors based on collagen I hydrogels

    PubMed Central

    Szot, Christopher S.; Buchanan, Cara F.; Freeman, Joseph W.; Rylander, Marissa N.

    2011-01-01

    Cells cultured within a three-dimensional (3D) in vitro environment have the ability to acquire phenotypes and respond to stimuli analogous to in vivo biological systems. This approach has been utilized in tissue engineering and can also be applied to the development of a physiologically relevant in vitro tumor model. In this study, collagen I hydrogels cultured with MDA-MB-231 human breast cancer cells were bioengineered as a platform for in vitro solid tumor development. The cell–cell and cell-matrix interactions present during in vivo tissue progression were encouraged within the 3D hydrogel architecture, and the biocompatibility of collagen I supported unconfined cellular proliferation. The development of necrosis beyond a depth of ~150–200 μm and the expression of hypoxia-inducible factor (HIF)-1α were demonstrated in the in vitro bioengineered tumors. Oxygen and nutrient diffusion limitations through the collagen I matrix as well as competition for available nutrients resulted in growing levels of intra-cellular hypoxia, quantified by a statistically significant (p < 0.01) upregulation of HIF-1α gene expression. The bioengineered tumors also demonstrated promising angiogenic potential with a statistically significant (p < 0.001) upregulation of vascular endothelial growth factor (VEGF)-A gene expression. In addition, comparable gene expression analysis demonstrated a statistically significant increase of HIF-1α (p < 0.05) and VEGF-A (p < 0.001) by MDA-MB-231 cells cultured in the 3D collagen I hydrogels compared to cells cultured in a monolayer on two-dimensional tissue culture polystyrene. The results presented in this study demonstrate the capacity of collagen I hydrogels to facilitate the development of 3D in vitro bioengineered tumors that are representative of the pre-vascularized stages of in vivo solid tumor progression. PMID:21782234

  2. Effects of Decorin Proteoglycan on Fibrillogenesis, Ultrastructure, and Mechanics of Type I Collagen Gels

    PubMed Central

    Reese, Shawn P.; Underwood, Clayton J.; Weiss, Jeffrey A.

    2013-01-01

    The proteoglycan decorin is known to affect both the fibrillogenesis and the resulting ultrastructure of in vitro polymerized collagen gels. However, little is known about its effects on mechanical properties. In this study, 3D collagen gels were polymerized into tensile test specimens in the presence of decorin proteoglycan, decorin core protein, or dermatan sulfate (DS). Collagen fibrillogenesis, ultrastructure, and mechanical properties were then quantified using a turbidity assay, 2 forms of microscopy (SEM and confocal), and tensile testing. The presence of decorin proteoglycan or core protein decreased the rate and ultimate turbidity during fibrillogenesis and decreased the number of fibril aggregates (fibers) compared to control gels. The addition of decorin and core protein increased the linear modulus by a factor of 2 compared to controls, while the addition of DS reduced the linear modulus by a factor of 3. Adding decorin after fibrillogenesis had no effect, suggesting that decorin must be present during fibrillogenesis to increase the mechanical properties of the resulting gels. These results show that the inclusion of decorin proteoglycan during fibrillogenesis of Type I collagen increases the modulus and tensile strength of resulting collagen gels. The increase in mechanical properties when polymerization occurs in the presence of the decorin proteoglycan is due to a reduction in the aggregation of fibrils into larger order structures such as fibers and fiber bundles. PMID:23608680

  3. Modeling the transparent shape memory gels by 3D printer Acculas

    NASA Astrophysics Data System (ADS)

    Kumagai, Hiroaki; Arai, Masanori; Gong, Jin; Sakai, Kazuyuki; Kawakami, Masaru; Furukawa, Hidemitsu

    2016-04-01

    In our group, highly transparent shape memory gels were successfully synthesized for the first time in the world. These gels have the high strength of 3MPs modulus even with the water content of 40wt% water and high transparency. We consider that these highly transparent and high strength gels can be applied to the optical devices such as intraocular-lenses and optical fibers. In previous research by our group, attempts were made to manufacture the gel intraocular-lenses using highly transparent shape memory gels. However, it was too difficult to print the intraocular-lens finely enough. Here, we focus on a 3D printer, which can produce objects of irregular shape. 3D printers generally we fused deposition modeling (FDM), a stereo lithography apparatus (SLA) and selective laser sintering (SLS). Because highly transparent shape memory gels are gelled by light irradiation, we used 3D printer with stereo lithography apparatus (SLA). In this study, we found the refractive index of highly transparent shape memory gels depend on monomer concentration, and does not depend on the cross-linker or initiator concentration. Furthermore, the cross-linker and initiator concentration can change the gelation progression rate. As a result, we have developed highly transparent shape memory gels, which can have a range of refractive indexes, and we defined the optimal conditions that can be modeling in the 3D printer by changing the cross-linker and initiator concentration. With these discoveries we were able to produce a gel intraocular-lens replica.

  4. A Tunable 3D Nanostructured Conductive Gel Framework Electrode for High-Performance Lithium Ion Batteries.

    PubMed

    Shi, Ye; Zhang, Jun; Bruck, Andrea M; Zhang, Yiman; Li, Jing; Stach, Eric A; Takeuchi, Kenneth J; Marschilok, Amy C; Takeuchi, Esther S; Yu, Guihua

    2017-03-22

    This study develops a tunable 3D nanostructured conductive gel framework as both binder and conductive framework for lithium ion batteries. A 3D nanostructured gel framework with continuous electron pathways can provide hierarchical pores for ion transport and form uniform coatings on each active particle against aggregation. The hybrid gel electrodes based on a polypyrrole gel framework and Fe3 O4 nanoparticles as a model system in this study demonstrate the best rate performance, the highest achieved mass ratio of active materials, and the highest achieved specific capacities when considering total electrode mass, compared to current literature. This 3D nanostructured gel-based framework represents a powerful platform for various electrochemically active materials to enable the next-generation high-energy batteries.

  5. Engineering multi-layered skeletal muscle tissue by using 3D microgrooved collagen scaffolds.

    PubMed

    Chen, Shangwu; Nakamoto, Tomoko; Kawazoe, Naoki; Chen, Guoping

    2015-12-01

    Preparation of three-dimensional (3D) micropatterned porous scaffolds remains a great challenge for engineering of highly organized tissues such as skeletal muscle tissue and cardiac tissue. Two-dimensional (2D) micropatterned surfaces with periodic features (several nanometers to less than 100 μm) are commonly used to guide the alignment of muscle myoblasts and myotubes and lead to formation of pre-patterned cell sheets. However, cell sheets from 2D patterned surfaces have limited thickness, and harvesting the cell sheets for implantation is inconvenient and can lead to less alignment of myotubes. 3D micropatterned scaffolds can promote cell alignment and muscle tissue formation. In this study, we developed a novel type of 3D porous collagen scaffolds with concave microgrooves that mimic muscle basement membrane to engineer skeletal muscle tissue. Highly aligned and multi-layered muscle bundle tissues were engineered by controlling the size of microgrooves and cell seeding concentration. Myoblasts in the engineered muscle tissue were well-aligned and had high expression of myosin heavy chain and synthesis of muscle extracellular matrix. The microgrooved collagen scaffolds could be used to engineer organized multi-layered muscle tissue for implantation to repair/restore the function of diseased tissues or be used to investigate the cell-cell interaction in 3D microscale topography.

  6. Quantitative characterization of developing collagen gels using optical coherence tomography

    NASA Astrophysics Data System (ADS)

    Levitz, David; Hinds, Monica T.; Choudhury, Niloy; Tran, Noi T.; Hanson, Stephen R.; Jacques, Steven L.

    2010-03-01

    Nondestructive optical imaging methods such as optical coherence tomography (OCT) have been proposed for characterizing engineered tissues such as collagen gels. In our study, OCT was used to image collagen gels with different seeding densities of smooth muscle cells (SMCs), including acellular gels, over a five-day period during which the gels contracted and became turbid with increased optical scattering. The gels were characterized quantitatively by their optical properties, specified by analysis of OCT data using a theoretical model. At 6 h, seeded cell density and scattering coefficient (μs) were correlated, with μs equal to 10.8 cm-1/(106 cells/mL). Seeded cell density and the scattering anisotropy (g) were uncorrelated. Over five days, the reflectivity in SMC gels gradually doubled with little change in optical attenuation, which indicated a decrease in g that increased backscatter, but only a small drop in μs. At five days, a subpopulation of sites on the gel showed substantially higher reflectivity (approximately a tenfold increase from the first 24 h). In summary, the increased turbidity of SMC gels that develops over time is due to a change in the structure of collagen, which affects g, and not simply due to a change in number density of collagen fibers due to contraction.

  7. Basic radiological characteristics of a non-scattering gel dosimeter for 3D dosimetry

    NASA Astrophysics Data System (ADS)

    Chang, Kyung Hwan; Ji, Yunseo; Lee, Suk; Kim, Kwang Hyeon; Yang, Dae Sik; Lee, Jung Ae; Park, Young Je; Yoon, Won Sup; Kim, Chul Yong; Cao, Yuanjie; Cho, Samju

    2016-12-01

    We used a spectrophotometer to compare the dosimetric properties of two non-scattering (radiochromic) gel dosimeters: a non-scattering gel dosimeter developed in-house and a PRESAGE™ gel dosimeter. We evaluated the dosimetric characteristics, including spectral absorption, dose linearity, reproducibility, and dose rate dependency of the two gel dosimeters. The non-scattering gel and the PRESAGE™ gel dosimeters showed peak sensitivity at wavelengths of 600 nm and 630 nm, respectively. Over a range of doses the best dose linearities of the non-scattering and the PRESAGE™ gel dosimeters resulted in R2 values of 0.99 at wavelengths of 600 nm and 630 nm, respectively. The reproducibility and dose-rate dependence of each of the two gel dosimeters were within the range of ±3 %. Our results revealed that the peak sensitivities of the two radiochromic gel dosimeters were significantly different; the in-house non-scattering gel dosimeter demonstrated peak sensitivity at a wavelength of 600 nm while the PRESAGE™ gel dosimeter had peak sensitivity at a wavelength of 630 nm. We confirmed that for 3D gel dosimetry, the in-house non-scattering gel dosimeter had a more stable dose response compared with a commercial non-scattering gel dosimeter.

  8. Planar Gradient Diffusion System to Investigate Chemotaxis in a 3D Collagen Matrix.

    PubMed

    Stout, David A; Toyjanova, Jennet; Franck, Christian

    2015-06-12

    The importance of cell migration can be seen through the development of human life. When cells migrate, they generate forces and transfer these forces to their surrounding area, leading to cell movement and migration. In order to understand the mechanisms that can alter and/or affect cell migration, one can study these forces. In theory, understanding the fundamental mechanisms and forces underlying cell migration holds the promise of effective approaches for treating diseases and promoting cellular transplantation. Unfortunately, modern chemotaxis chambers that have been developed are usually restricted to two dimensions (2D) and have complex diffusion gradients that make the experiment difficult to interpret. To this end, we have developed, and describe in this paper, a direct-viewing chamber for chemotaxis studies, which allows one to overcome modern chemotaxis chamber obstacles able to measure cell forces and specific concentration within the chamber in a 3D environment to study cell 3D migration. More compelling, this approach allows one to successfully model diffusion through 3D collagen matrices and calculate the coefficient of diffusion of a chemoattractant through multiple different concentrations of collagen, while keeping the system simple and user friendly for traction force microscopy (TFM) and digital volume correlation (DVC) analysis.

  9. Regeneration of chronic tympanic membrane perforation using 3D collagen with topical umbilical cord serum.

    PubMed

    Jang, Chul Ho; Cho, Yong Beom; Yeo, MyungGu; Lee, Hyeongjin; Min, Eun Jung; Lee, Byung Hhwa; Kim, Geun Hyung

    2013-11-01

    Chronic tympanic membrane (TM) perforation is one of the most common otology complications. Current surgical management of TM perforation includes myringoplasty and tympanoplasty. The purpose of this study was to evaluate the efficacy and feasibility of three dimensional (3D) porous collagen scaffolds with topically applied human umbilical cord serum (UCS) for the regeneration of chronic TM perforation in guinea pigs. To achieve this goal, we fabricated porous 3D collagen scaffolds (avg. strut diameter of 236 ± 51 μm, avg. pore size of 382 ± 67 μm, and a porosity of 96%) by using a 3 axis robot dispensing and low temperature plate systems. Guinea pigs were used in a model of chronic TM perforation. In the experimental group (n=10), 3D collagen scaffold was placed on the perforation and topically applied of UCS every other day for a period of 8 days. The control group ears (n=10) were treated with paper discs and phosphate buffered saline (PBS) only using the same regimen. Healing time, acoustic-mechanical properties, and morphological analysis were performed by otoendoscopy, auditory brainstem response (ABR), single-point laser Doppler vibrometer (LDV), optical coherence tomography (OCT), and light microscopic evaluation. The closure of the TM perforation was achieved in 100% of the experimental group vs. 43% of the control group, and this difference was statistically significant (p=0.034). The ABR threshold at all frequencies of the experimental group was significantly recovered to the normal level compared to the control group. TM vibration velocity in the experimental group recovered similar to the normal control level. The difference is very small and they are not statistically significant below 1 kHz (p=0.074). By OCT and light microscopic examination, regenerated TM of the experimental group showed thickened fibrous and mucosal layer. In contrast, the control group showed absence of fibrous layer like a dimeric TM.

  10. Strain-enhanced stress relaxation impacts nonlinear elasticity in collagen gels

    PubMed Central

    Nam, Sungmin; Hu, Kenneth H.; Chaudhuri, Ovijit

    2016-01-01

    The extracellular matrix (ECM) is a complex assembly of structural proteins that provides physical support and biochemical signaling to cells in tissues. The mechanical properties of the ECM have been found to play a key role in regulating cell behaviors such as differentiation and malignancy. Gels formed from ECM protein biopolymers such as collagen or fibrin are commonly used for 3D cell culture models of tissue. One of the most striking features of these gels is that they exhibit nonlinear elasticity, undergoing strain stiffening. However, these gels are also viscoelastic and exhibit stress relaxation, with the resistance of the gel to a deformation relaxing over time. Recent studies have suggested that cells sense and respond to both nonlinear elasticity and viscoelasticity of ECM, yet little is known about the connection between nonlinear elasticity and viscoelasticity. Here, we report that, as strain is increased, not only do biopolymer gels stiffen but they also exhibit faster stress relaxation, reducing the timescale over which elastic energy is dissipated. This effect is not universal to all biological gels and is mediated through weak cross-links. Mechanistically, computational modeling and atomic force microscopy (AFM) indicate that strain-enhanced stress relaxation of collagen gels arises from force-dependent unbinding of weak bonds between collagen fibers. The broader effect of strain-enhanced stress relaxation is to rapidly diminish strain stiffening over time. These results reveal the interplay between nonlinear elasticity and viscoelasticity in collagen gels, and highlight the complexity of the ECM mechanics that are likely sensed through cellular mechanotransduction. PMID:27140623

  11. Microfabricated electrospun collagen membranes for 3-D cancer models and drug screening applications

    PubMed Central

    Hartman, Olga; Zhang, Chu; Adams, Elizabeth L.; Farach-Carson, Mary C.; Petrelli, Nicholas J.; Chase, Bruce D.; Rabolt, John F.

    2009-01-01

    Invasive epithelial tumors form from cells that are released from their natural basement membrane and form 3-D structures that interact with each other and with the microenvironment of the stromal tissues around the tumor, which often contains collagen. Cancer cells, growing as monolayers on tissue culture plastic, do not reflect many of the properties of whole tumors. This shortcoming limits their ability to serve as models for testing of pharmacologically active compounds, including those that are being tested as anti-neoplastics. This work seeks to create new 3-D cellular materials possessing properties similar to those in native tissues surrounding cancers, specifically electrospun micro- and nanofibrous collagen scaffolds that support tumor growth in 3-D. We hypothesize that a 3-D culture system will provide a better replica of tumor growth in a native environment, and thus better report the bioactivity of anti-neoplastic agents. In addition, we optimized conditions, and identified physical characteristics that support growth of the highly invasive, prostate cancer bone metastatic cell line C4-2B on these matrices for use in anti-cancer drug studies. The effects of matrix porosity, fiber diameter, elasticity and surface roughness on growth of cancer cells were evaluated. Data indicates that while cells attach and grow well on both nano- and microfibrous electrospun membranes, the microfibrous membrane represented a better approximation of the tumor microenvironemt. It was also observed that C4-2B non-adherent cells migrated through the depth of two electrospun membranes and formed colonies resembling tumors on day 3. An apoptosis study revealed that cells on electrospun substrates were more resistant to both anti-neoplastic agents, docetaxel (DOC) and camptothecin (CAM), compared to the cells grown on standard collagen-coated tissue culture polystyrene (TCP). Growth, survival, and apoptosis were measured, as well as the differences in the apoptotic

  12. Development of bioartificial myocardium by electrostimulation of 3D collagen scaffolds seeded with stem cells.

    PubMed

    Haneef, Kanwal; Lila, Nermine; Benadda, Samira; Legrand, Fabien; Carpentier, Alain; Chachques, Juan C

    2012-06-05

    Electrostimulation (ES) can be defined as a safe physical method to induce stem cell differentiation. The aim of this study is to evaluate the effectiveness of ES on bone marrow mesenchymal stem cells (BMSCs) seeded in collagen scaffolds in terms of proliferation and differentiation into cardiomyocytes. BMSCs were isolated from Wistar rats and seeded into 3D collagen type 1 templates measuring 25 × 25 × 6 mm. Bipolar in vitro ES was performed during 21 days. Electrical impedance and cell proliferation were measured. Expression of cardiac markers was assessed by immunocytochemistry. Viscoelasticity of collagen matrix was evaluated. Electrical impedance assessments showed a low resistance of 234±41 Ohms which indicates good electrical conductivity of collagen matrix. Cell proliferation at 570 nm as significantly increased in ES groups after seven day (ES 0.129±0.03 vs non-stimulated control matrix 0.06±0.01, P=0.002) and after 21 days, (ES 0.22±0.04 vs control 0.13±0.01, P=0.01). Immunocytoche mistry of BMSCs after 21 days ES showed positive staining of cardiac markers, troponin I, connexin 43, sarcomeric alpha-actinin, slow myosin, fast myosin and desmin. Staining for BMSCs marker CD29 after 21 days was negative. Electrostimulation of cell-seeded collagen matrix changed stem cell morphology and biochemical characteristics, increasing the expression of cardiac markers. Thus, MSC-derived differentiated cells by electrostimulation grafted in biological scaffolds might result in a convenient tissue engineering source for myocardial diseases.

  13. 3D scanning of internal structure in gel engineering materials with visual scanning microscopic light scattering

    NASA Astrophysics Data System (ADS)

    Watanabe, Yosuke; Gong, Jing; Masato, Makino; Kabir, M. Hasnat; Furukawa, Hidemitsu

    2014-04-01

    The 3D printing technology, causing much attention from the beginning of 2013, will be possibly an alternative method to fabricate the biological soft tissues. Recently our group of Yamagata University has developed the world-first 3D Gel Printer to fabricate the complicated gel-materials with high-strength and biocompatibility. However, there are no 3D scanners that collect the data from the internal structure of complicated gel objects such as eye lens. It means that a new system for scanning the internal structure is needed now. In this study, firstly, we have tried to investigate the gel network of synthetic and biological gel with scanning microscopic light scattering (SMILS). We calculated the Young's modulus of synthetic gels with the SMILS and with the tensile test, and precisely compared the results between them. The temperature dependences of the inside structure and the transparency are observed in the pig crystalline lens. The quantitative analysis indicates the importance of the internal structure of real object. Secondary, we show the new system named Gel-scanner that can provide the 2-dimentional data of the internal structure. From examining our findings, the scanning of internal structure will enable us to expect physical properties of the real object. We convince that the gelscanner will play major role in the various fields.

  14. Performance of a commercial optical CT scanner and polymer gel dosimeters for 3-D dose verification.

    PubMed

    Xu, Y; Wuu, Cheng-Shie; Maryanski, Marek J

    2004-11-01

    Performance analysis of a commercial three-dimensional (3-D) dose mapping system based on optical CT scanning of polymer gels is presented. The system consists of BANG 3 polymer gels (MGS Research, Inc., Madison, CT), OCTOPUS laser CT scanner (MGS Research, Inc., Madison, CT), and an in-house developed software for optical CT image reconstruction and 3-D dose distribution comparison between the gel, film measurements and the radiation therapy treatment plans. Various sources of image noise (digitization, electronic, optical, and mechanical) generated by the scanner as well as optical uniformity of the polymer gel are analyzed. The performance of the scanner is further evaluated in terms of the reproducibility of the data acquisition process, the uncertainties at different levels of reconstructed optical density per unit length and the effects of scanning parameters. It is demonstrated that for BANG 3 gel phantoms held in cylindrical plastic containers, the relative dose distribution can be reproduced by the scanner with an overall uncertainty of about 3% within approximately 75% of the radius of the container. In regions located closer to the container wall, however, the scanner generates erroneous optical density values that arise from the reflection and refraction of the laser rays at the interface between the gel and the container. The analysis of the accuracy of the polymer gel dosimeter is exemplified by the comparison of the gel/OCT-derived dose distributions with those from film measurements and a commercial treatment planning system (Cadplan, Varian Corporation, Palo Alto, CA) for a 6 cm x 6 cm single field of 6 MV x rays and a 3-D conformal radiotherapy (3DCRT) plan. The gel measurements agree with the treatment plans and the film measurements within the "3%-or-2 mm" criterion throughout the usable, artifact-free central region of the gel volume. Discrepancies among the three data sets are analyzed.

  15. Nonlinear Optical Macroscopic Assessment of 3-D Corneal Collagen Organization and Axial Biomechanics

    PubMed Central

    Winkler, Moritz; Chai, Dongyul; Kriling, Shelsea; Nien, Chyong Jy; Brown, Donald J.; Jester, Bryan; Juhasz, Tibor

    2011-01-01

    Purpose. To characterize and quantify the collagen fiber (lamellar) organization of human corneas in three dimensions by using nonlinear optical high-resolution macroscopy (NLO-HRMac) and to correlate these findings with mechanical data obtained by indentation testing of corneal flaps. Methods. Twelve corneas from 10 donors were studied. Vibratome sections, 200 μm thick, from five donor eyes were cut along the vertical meridian from limbus to limbus (arc length, 12 mm). Backscattered second harmonic–generated (SHG) NLO signals from these sections were collected as a series of overlapping 3-D images, which were concatenated to form a single 3-D mosaic (pixel resolution: 0.44 μm lateral, 2 μm axial). Collagen fiber intertwining was quantified by determining branching point density as a function of stromal depth. Mechanical testing was performed on corneal flaps from seven additional eyes. Corneas were cut into three layers (anterior, middle, and posterior) using a femtosecond surgical laser system and underwent indentation testing to determine the elastic modulus for each layer. Results. The 3-D reconstructions revealed complex collagen fiber branching patterns in the anterior cornea, with fibers extending from the anterior limiting lamina (ALL, Bowman's layer), intertwining with deeper fibers and reinserting back to the ALL, forming bow spring–like structures. Measured branching-point density was four times higher in the anterior third of the cornea than in the posterior third and decreased logarithmically with increasing distance from the ALL. Indentation testing showed an eightfold increase in elastic modulus in the anterior stroma. Conclusions. The axial gradient in lamellar intertwining appears to be associated with an axial gradient in the effective elastic modulus of the cornea, suggesting that collagen fiber intertwining and formation of bow spring–like structures provide structural support similar to cross-beams in bridges and large-scale structures

  16. Tracking immune-related cell responses to drug delivery microparticles in 3D dense collagen matrix.

    PubMed

    Obarzanek-Fojt, Magdalena; Curdy, Catherine; Loggia, Nicoletta; Di Lena, Fabio; Grieder, Kathrin; Bitar, Malak; Wick, Peter

    2016-10-01

    Beyond the therapeutic purpose, the impact of drug delivery microparticles on the local tissue and inflammatory responses remains to be further elucidated specifically for reactions mediated by the host immune cells. Such immediate and prolonged reactions may adversely influence the release efficacy and intended therapeutic pathway. The lack of suitable in vitro platforms limits our ability to gain insight into the nature of immune responses at a single cell level. In order to establish an in vitro 3D system mimicking the connective host tissue counterpart, we utilized reproducible, compressed, rat-tail collagen polymerized matrices. THP1 cells (human acute monocytic leukaemia cells) differentiated into macrophage-like cells were chosen as cell model and their functionality was retained in the dense rat-tail collagen matrix. Placebo microparticles were later combined in the immune cell seeded system during collagen polymerization and secreted pro-inflammatory factors: TNFα and IL-8 were used as immune response readout (ELISA). Our data showed an elevated TNFα and IL-8 secretion by macrophage THP1 cells indicating that Placebo microparticles trigger certain immune cell responses under 3D in vivo like conditions. Furthermore, we have shown that the system is sensitive to measure the differences in THP1 macrophage pro-inflammatory responses to Active Pharmaceutical Ingredient (API) microparticles with different API release kinetics. We have successfully developed a tissue-like, advanced, in vitro system enabling selective "readouts" of specific responses of immune-related cells. Such system may provide the basis of an advanced toolbox enabling systemic evaluation and prediction of in vivo microparticle reactions on human immune-related cells.

  17. The Rho family GEF Asef2 regulates cell migration in three dimensional (3D) collagen matrices through myosin II.

    PubMed

    Jean, Léolène; Yang, Lijie; Majumdar, Devi; Gao, Yandong; Shi, Mingjian; Brewer, Bryson M; Li, Deyu; Webb, Donna J

    2014-01-01

    Cell migration is fundamental to a variety of physiological processes, including tissue development, homeostasis, and regeneration. Migration has been extensively studied with cells on 2-dimensional (2D) substrates, but much less is known about cell migration in 3D environments. Tissues and organs are 3D, which is the native environment of cells in vivo, pointing to a need to understand migration and the mechanisms that regulate it in 3D environments. To investigate cell migration in 3D environments, we developed microfluidic devices that afford a controlled, reproducible platform for generating 3D matrices. Using these devices, we show that the Rho family guanine nucleotide exchange factor (GEF) Asef2 inhibits cell migration in 3D type I collagen (collagen I) matrices. Treatment of cells with the myosin II (MyoII) inhibitor blebbistatin abolished the decrease in migration by Asef2. Moreover, Asef2 enhanced MyoII activity as shown by increased phosphorylation of serine 19 (S19). Furthermore, Asef2 increased activation of Rac, which is a Rho family small GTPase, in 3D collagen I matrices. Inhibition of Rac activity by treatment with the Rac-specific inhibitor NSC23766 abrogated the Asef2-promoted increase in S19 MyoII phosphorylation. Thus, our results indicate that Asef2 regulates cell migration in 3D collagen I matrices through a Rac-MyoII-dependent mechanism.

  18. An in vitro 3D model using collagen coated gelatin nanofibers for studying breast cancer metastasis.

    PubMed

    Janani, G; Pillai, Mamatha M; Selvakumar, R; Bhattacharyya, Amitava; Sabarinath, C

    2017-02-07

    The study of breast cancer metastasis is limited due to poor knowledge of molecular progression of breast tumor and varied heterogeneity. For a better understanding of tumor metastasis, a reliable 3D in vitro model bridging the gap between 2D cultures and in vivo animal model studies is essential. Our study is focused on two key points: (i) designing a 3D microenvironment for studying metastasis and (ii) simulating the metastasis milieu by inducing epithelial to mesenchymal transition (EMT) and mesenchymal to epithelial transition (MET). An electrospun gelatin nanofiber matrix (EGNF) was fabricated using electrospinning and further dip coated with different concentrations of collagen to obtain surface complexity and mechanical properties, similar to connective tissues. Nanofiber matrices were physically characterized by Fourier transform infrared spectroscopy (FTIR), atomic force microscopy (AFM), and field-emission scanning electron microscopy (FESEM). The FTIR, AFM, and FESEM results indicated the crosslinking and confirmed the presence of pores in the nanofiber matrices. Comparative studies on biocompatibility, cell attachment, and the proliferation of MCF-7 cells on EGNF and collagen coated gelatin nanofibrous matrix (CCGM) revealed higher cellular attachment and proliferation in CCGM. CCGM with human metastatic breast cancer cell line (MCF-7) was taken to study breast cancer metastasis using estrogen (induces EMT) and progesterone (induces MET) hormones for 24 h. Quantitative real-time PCR was used for quantifying the expression of metastasis related genes, and fluorescence microscopy for verifying the invasion of cells to the matrices. The expression of E-cadherin and matrix metalloproteinase 2 (MMP 2) confirmed the occurrence of EMT and MET. Live cell imaging and cellular attachment showed significant increase of cellular invasion in crosslinked 0.15% CCGM that serves as a suitable non-toxic, biocompatible, and affordable scaffold for studying breast cancer

  19. 3D composites based on the blends of chitosan and collagen with the addition of hyaluronic acid.

    PubMed

    Sionkowska, Alina; Kaczmarek, Beata; Lewandowska, Katarzyna; Grabska, Sylwia; Pokrywczyńska, Marta; Kloskowski, Tomasz; Drewa, Tomasz

    2016-08-01

    3D porous composites based on blends of chitosan, collagen and hyaluronic acid were obtained through the lyophilization process. Mechanical properties, swelling behavior and thermal stability of the blends were studied. Moreover, SEM images were taken and the structure of the blends was studied. Biological properties of the materials obtained were investigated by analyzing of proliferation rate of fibroblast cells incubated with biomaterial extract using MTT assay (3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide). The results showed that the properties of 3D composites based on the blends of chitosan and collagen were altered after the addition 1%, 2% and 5% of hyaluronic acid. Mechanical properties and thermal stability of chitosan/collagen blends were improved in the presence of hyaluronic acid in the composite. New 3D materials based on the blends of chitosan, collagen and hyaluronic acid were non-toxic and did not significantly affect cell morphology.

  20. Saos-2 cell-mediated mineralization on collagen gels: Effect of densification and bioglass incorporation.

    PubMed

    Liu, Gengbo; Pastakia, Meet; Fenn, Michael B; Kishore, Vipuil

    2016-05-01

    Plastic compression is a collagen densification process that has been widely used for the development of mechanically robust collagen-based materials. Incorporation of bioglass within plastically compressed collagen gels has been shown to mimic the microstructural properties of native bone and enhance in vitro cell-mediated mineralization. The current study seeks to decouple the effects of collagen densification and bioglass incorporation to understand the interplay between collagen packing density and presence of bioglass on cell-mediated mineralization. Saos-2 cell-mediated mineralization was assessed as a measure of the osteoconductivity of four different collagen gels: (1) uncompressed collagen gel (UC), (2) bioglass incorporated uncompressed collagen gel (UC + BG), (3) plastically compressed collagen gel (PC), and (4) bioglass incorporated plastically compressed collagen gel (PC + BG). The results indicated that collagen densification enhanced mineralization as shown by SEM, increased alkaline phosphatase activity and produced significantly higher amounts of mineralized nodules on PC gels compared to UC gels. Further, the amount of nodule formation on PC gels was significantly higher compared to UC + BG gels indicating that increase in matrix stiffness due to collagen densification had a greater effect on cell-mediated mineralization compared to bioglass incorporation into loosely packed UC gels. Incorporation of bioglass into PC gels further enhanced mineralization as evidenced by significantly larger nodule size and higher amount of mineralization on PC + BG gels compared to PC gels. In conclusion, collagen densification via plastic compression improves the osteoconductivity of collagen gels. Further, incorporation of bioglass within PC gels has an additive effect and further enhances the osteoconductivity of collagen gels.

  1. Fluorescein diacetate for determination of cell viability in 3D fibroblast-collagen-GAG constructs.

    PubMed

    Powell, Heather M; Armour, Alexis D; Boyce, Steven T

    2011-01-01

    Quantification of cell viability and distribution within engineered tissues currently relies on representative histology, phenotypic assays, and destructive assays of viability. To evaluate uniformity of cell density throughout 3D collagen scaffolds prior to in vivo use, a nondestructive, field assessment of cell viability is advantageous. Here, we describe a field measure of cell viability in lyophilized collagen-glycosaminoglycan (C-GAG) scaffolds in vitro using fluorescein diacetate (FdA). Fibroblast-C-GAG constructs are stained 1 day after cellular inoculation using 0.04 mg/ml FdA followed by exposure to 366 nm UV light. Construct fluorescence quantified using Metamorph image analysis is correlated with inoculation density, MTT values, and histology of corresponding biopsies. Construct fluorescence correlates significantly with inoculation density (p  <  0.001) and MTT values (p  <  0.001) of biopsies collected immediately after FdA staining. No toxicity is detected in the constructs, as measured by MTT assay before and after the FdA assay at different time points; normal in vitro histology is demonstrated for the FdA-exposed constructs. In conclusion, measurement of intracellular fluorescence with FdA allows for the early, comprehensive measurement of cellular distributions and viability in engineered tissue.

  2. Fibrillogenesis from nanosurfaces: multiphoton imaging and stereological analysis of collagen 3D self-assembly dynamics.

    PubMed

    Bancelin, Stéphane; Decencière, Etienne; Machairas, Vaïa; Albert, Claire; Coradin, Thibaud; Schanne-Klein, Marie-Claire; Aimé, Carole

    2014-09-21

    The assembly of proteins into fibrillar structures is an important process that concerns different biological contexts, including molecular medicine and functional biomaterials. Engineering of hybrid biomaterials can advantageously provide synergetic interactions of the biopolymers with an inorganic component to ensure specific supramolecular organization and dynamics. To this aim, we designed hybrid systems associating collagen and surface-functionalized silica particles and we built a new strategy to investigate fibrillogenesis processes in such multicomponents systems, working at the crossroads of chemistry, physics and mathematics. The self-assembly process was investigated by bimodal multiphoton imaging coupling second harmonic generation (SHG) and 2 photon excited fluorescence (2PEF). The in-depth spatial characterization of the system was further achieved using the three-dimensional analysis of the SHG/2PEF data via mathematical morphology processing. Quantitation of collagen distribution around particles offers strong evidence that the chemically induced confinement of the protein on the silica nanosurfaces has a key influence on the spatial extension of fibrillogenesis. This new approach is unique in the information it can provide on 3D dynamic hybrid systems and may be extended to other associations of fibrillar molecules with optically responsive nano-objects.

  3. Topographical guidance of 3D tumor cell migration at an interface of collagen densities.

    PubMed

    Bordeleau, Francois; Tang, Lauren N; Reinhart-King, Cynthia A

    2013-12-01

    During cancer progression, metastatic cells leave the primary tumor and invade into the fibrous extracellular matrix (ECM) within the surrounding stroma. This ECM network is highly heterogeneous, and interest in understanding how this network can affect cell behavior has increased in the past several decades. However, replicating this heterogeneity has proven challenging. Here, we designed and utilized a method to create a well-defined interface between two distinct regions of high- and low-density collagen gels to mimic the heterogeneities in density found in the tumor stroma. We show that cells will invade preferentially from the high-density side into the low-density side. We also demonstrate that the net cell migration is a function of the density of the collagen in which the cells are embedded, and the difference in density between the two regions has minimal effect on cell net displacement and distance travelled. Our data further indicate that a low-to-high density interface promotes directional migration and induces formation of focal adhesion on the interface surface. Together, the current results demonstrate how ECM heterogeneities, in the form of interfacial boundaries, can affect cell migration.

  4. Advanced 3D Ni(OH)2/CNT Gel Composite Electrodes for Supercapacitors

    NASA Astrophysics Data System (ADS)

    Cheng, Hanlin; Duong, Hai Minh

    2015-03-01

    In order to enhance the performance of supercapacitors, advanced 3D Porous CNT/Ni(OH)2 gel composite electrodes are developed in this work. Compared with previously reported graphene gel supercapacitors, our electrodes using 1D CNTs have smaller diffusion resistance due to a shorter ion transport path. The developed 3D xerogel composite electrodes demonstrate the success of a careful engineered guest/host materials interface. Initially, the CNT gels are coated on the nickel foam to form a 3D scaffold, which serves as a microscopic electrical conductive network. Then Ni(OH)2 are incorporated using a traditional electrodeposition method. In this work, two types of the 3D CNT-coated nickel foams are investigated. The gels can be used directly as hydrogels or dried in air to form xerogels. Both hydrogels and xerogels present 3D tangled CNT networks. It shows that the hydrogel composite electrodes with unbundled CNTs, though presenting high capacitances of 1400 F/g at low discharge rate, possess lower capacitances at higher discharge rate and a poor cycling performance of less than 23% retention. In contrast, the xerogel composite electrodes can overcome these limitations in terms of a satisfied discharge performance of 1200 F/g and a good cycling retention more than 85% due to a stronger Ni(OH)2/CNT interface. The CNT bundles in the xerogel electrodes formed during the drying process can give a flat surface with small curvature, which facilitate the Ni(OH)2 nucleation and growth. Thanks for the support from the A star R-265-000-424-305.

  5. Mechanical Behavior of Collagen-Fibrin Co-Gels Reflects Transition From Series to Parallel Interactions With Increasing Collagen Content

    PubMed Central

    Lai, Victor K.; Lake, Spencer P.; Frey, Christina R.; Tranquillo, Robert T.; Barocas, Victor H.

    2013-01-01

    Fibrin and collagen, biopolymers occurring naturally in the body, are commonly-used biomaterials as scaffolds for tissue engineering. How collagen and fibrin interact to confer macroscopic mechanical properties in collagen-fibrin composite systems remains poorly understood. In this study, we formulated collagen-fibrin co-gels at different collagen-to-fibrin ratios to observe changes in overall mechanical behavior and microstructure. A modeling framework of a two-network system was developed by modifying our micro-scale model, considering two forms of interaction between the networks: (a) two interpenetrating but non-interacting networks (“parallel”), and (b) a single network consisting of randomly alternating collagen and fibrin fibrils (“series”). Mechanical testing of our gels show that collagen-fibrin co-gels exhibit intermediate properties (UTS, strain at failure, tangent modulus) compared to those of pure collagen and fibrin. Comparison with model predictions show that the parallel and series model cases provide upper and lower bounds respectively for the experimental data, suggesting that a combination of such interactions exist between collagen and fibrin in co-gels. A transition from the series model to the parallel model occurs with increasing collagen content, with the series model best describing predominantly fibrin co-gels, and the parallel model best describing predominantly collagen co-gels. PMID:22482659

  6. 3D bioprinting of BMSC-laden methacrylamide gelatin scaffolds with CBD-BMP2-collagen microfibers.

    PubMed

    Du, Mingchun; Chen, Bing; Meng, Qingyuan; Liu, Sumei; Zheng, Xiongfei; Zhang, Cheng; Wang, Heran; Li, Hongyi; Wang, Nuo; Dai, Jianwu

    2015-12-18

    Three-dimensional (3D) bioprinting combines biomaterials, cells and functional components into complex living tissues. Herein, we assembled function-control modules into cell-laden scaffolds using 3D bioprinting. A customized 3D printer was able to tune the microstructure of printed bone mesenchymal stem cell (BMSC)-laden methacrylamide gelatin scaffolds at the micrometer scale. For example, the pore size was adjusted to 282 ± 32 μm and 363 ± 60 μm. To match the requirements of the printing nozzle, collagen microfibers with a length of 22 ± 13 μm were prepared with a high-speed crusher. Collagen microfibers bound bone morphogenetic protein 2 (BMP2) with a collagen binding domain (CBD) as differentiation-control module, from which BMP2 was able to be controllably released. The differentiation behaviors of BMSCs in the printed scaffolds were compared in three microenvironments: samples without CBD-BMP2-collagen microfibers in the growth medium, samples without microfibers in the osteogenic medium and samples with microfibers in the growth medium. The results indicated that BMSCs showed high cell viability (>90%) during printing; CBD-BMP2-collagen microfibers induced BMSC differentiation into osteocytes within 14 days more efficiently than the osteogenic medium. Our studies suggest that these function-control modules are attractive biomaterials and have potential applications in 3D bioprinting.

  7. An extracellular-matrix-specific GEF-GAP interaction regulates Rho GTPase crosstalk for 3D collagen migration.

    PubMed

    Kutys, Matthew L; Yamada, Kenneth M

    2014-09-01

    Rho-family GTPases govern distinct types of cell migration on different extracellular matrix proteins in tissue culture or three-dimensional (3D) matrices. We searched for mechanisms selectively regulating 3D cell migration in different matrix environments and discovered a form of Cdc42-RhoA crosstalk governing cell migration through a specific pair of GTPase activator and inhibitor molecules. We first identified βPix, a guanine nucleotide exchange factor (GEF), as a specific regulator of migration in 3D collagen using an affinity-precipitation-based GEF screen. Knockdown of βPix specifically blocks cell migration in fibrillar collagen microenvironments, leading to hyperactive cellular protrusion accompanied by increased collagen matrix contraction. Live FRET imaging and RNAi knockdown linked this βPix knockdown phenotype to loss of polarized Cdc42 but not Rac1 activity, accompanied by enhanced, de-localized RhoA activity. Mechanistically, collagen phospho-regulates βPix, leading to its association with srGAP1, a GTPase-activating protein (GAP), needed to suppress RhoA activity. Our results reveal a matrix-specific pathway controlling migration involving a GEF-GAP interaction of βPix with srGAP1 that is critical for maintaining suppressive crosstalk between Cdc42 and RhoA during 3D collagen migration.

  8. Extracellular Matrix Fibronectin Stimulates the Self-Assembly of Microtissues on Native Collagen Gels

    PubMed Central

    Sevilla, Carlos A.; Dalecki, Diane

    2010-01-01

    Fibronectin is an adhesive glycoprotein that is polymerized into extracellular matrices via a tightly regulated, cell-dependent process. Here, we demonstrate that fibronectin matrix polymerization induces the self-assembly of multicellular structures in vitro, termed tissue bodies. Fibronectin-null mouse embryonic fibroblasts adherent to compliant gels of polymerized type I collagen failed to spread or proliferate. In contrast, addition of fibronectin to collagen-adherent fibronectin-null mouse embryonic fibroblasts resulted in a dose-dependent increase in cell number, and induced the formation of three-dimensional (3D) multicellular structures that remained adherent and well-spread on the native collagen substrate. An extensive fibrillar fibronectin matrix formed throughout the microtissue. Blocking fibronectin matrix polymerization inhibited both cell proliferation and microtissue formation, demonstrating the importance of fibronectin fibrillogenesis in triggering cellular self-organization. Cell proliferation, tissue body formation, and tissue body shape were dependent on both fibronectin and collagen concentrations, suggesting that the relative proportion of collagen and fibronectin fibrils polymerized into the extracellular matrix influences the extent of cell proliferation and the final shape of microtissues. These data demonstrate a novel role for cell-mediated fibronectin fibrillogenesis in the formation and vertical assembly of microtissues, and provide a novel approach for engineering complex tissue architecture. PMID:20673131

  9. Rapid 3D Printing of Multifunctional Calcium Alginate Gel Pipes using Coaxial Jet Extruder

    NASA Astrophysics Data System (ADS)

    Rykaczewski, Konrad; Damle, Viraj

    2014-11-01

    Calcium alginate (CA) forms when solution containing sodium alginate (SA) comes in contact with a CaCl2 solution. The resulting gel is biocompatible as well as edible and is used in production of bio-scaffolds, artificial plant seeds, and edible substances. In the latter application, referred to in the culinary world as ``spherification,'' flavored liquids are mixed with the SA and dripped into CaCl2 solution to form gel encapsulated flavored ``marbles.'' Previously, crude 3D printing of CA structures has been achieved by stacking of such flavored liquid filled marbles. In turn, solid CA rods have been fabricated by properly mixing flow of the two solutions using a microfluidic device. Here we show that by using two circular cross-section coaxial nozzles to produce coaxial jets of the SA and CaCl2 solutions, liquid filled CA micro-to-mili scale gel pipes can be produced at speeds around ~ 150 mm/s. Such extrusion rate is compatible with most commercially available 3D printers, facilitating adoption of the CA pipe coaxial jet extruder. Here, the impact of inner and outer liquid properties and flow speeds on the gel pipe extrusion process is discussed. KR acknowledges startup funding from ASU.

  10. Collagen esterification enhances the function and survival of pancreatic β cells in 2D and 3D culture systems

    SciTech Connect

    Ko, Jae Hyung; Kim, Yang Hee; Jeong, Seong Hee; Lee, Song; Park, Si-Nae; Shim, In Kyong; Kim, Song Cheol

    2015-08-07

    Collagen, one of the most important components of the extracellular matrix (ECM), may play a role in the survival of pancreatic islet cells. In addition, chemical modifications that change the collagen charge profile to a net positive charge by esterification have been shown to increase the adhesion and proliferation of various cell types. The purpose of this study was to characterize and compare the effects of native collagen (NC) and esterified collagen (EC) on β cell function and survival. After isolation by the collagenase digestion technique, rat islets were cultured with NC and EC in 2 dimensional (2D) and 3 dimensional (3D) environments for a long-term duration in vitro. The cells were assessed for islet adhesion, morphology, viability, glucose-induced insulin secretion, and mRNA expression of glucose metabolism-related genes, and visualized by scanning electron microscopy (SEM). Islet cells attached tightly in the NC group, but islet cell viability was similar in both the NC and EC groups. Glucose-stimulated insulin secretion was higher in the EC group than in the NC group in both 2D and 3D culture. Furthermore, the mRNA expression levels of glucokinase in the EC group were higher than those in the NC group and were associated with glucose metabolism and insulin secretion. Finally, SEM observation confirmed that islets had more intact component cells on EC sponges than on NC sponges. These results indicate that modification of collagen may offer opportunities to improve function and viability of islet cells. - Highlights: • We changed the collagen charge profile to a net positive charge by esterification. • Islets cultured on esterified collagen improved survival in both 2D and 3D culture. • Islets cultured on esterified collagen enhanced glucose-stimulated insulin release. • High levels of glucokinase mRNA may be associated with increased insulin release.

  11. Molecular transport in collagenous tissues measured by gel electrophoresis.

    PubMed

    Hunckler, Michael D; Tilley, Jennifer M R; Roeder, Ryan K

    2015-11-26

    Molecular transport in tissues is important for drug delivery, nutrient supply, waste removal, cell signaling, and detecting tissue degeneration. Therefore, the objective of this study was to investigate gel electrophoresis as a simple method to measure molecular transport in collagenous tissues. The electrophoretic mobility of charged molecules in tissue samples was measured from relative differences in the velocity of a cationic dye passing through an agarose gel in the absence and presence of a tissue section embedded within the gel. Differences in electrophoretic mobility were measured for the transport of a molecule through different tissues and tissue anisotropy, or the transport of different sized molecules through the same tissue. Tissue samples included tendon and fibrocartilage from the proximal (tensile) and distal (compressive) regions of the bovine flexor tendon, respectively, and bovine articular cartilage. The measured electrophoretic mobility was greatest in the compressive region of the tendon (fibrocartilage), followed by the tensile region of tendon, and lowest in articular cartilage, reflecting differences in the composition and organization of the tissues. The anisotropy of tendon was measured by greater electrophoretic mobility parallel compared with perpendicular to the predominate collagen fiber orientation. Electrophoretic mobility also decreased with increased molecular size, as expected. Therefore, the results of this study suggest that gel electrophoresis may be a useful method to measure differences in molecular transport within various tissues, including the effects of tissue type, tissue anisotropy, and molecular size.

  12. Strategies for Directing the Structure and Function of 3D Collagen Biomaterials across Length Scales

    PubMed Central

    Walters, Brandan D.; Stegemann, Jan P.

    2013-01-01

    Collagen type I is a widely used natural biomaterial that has found utility in a variety of biological and medical applications. Its well characterized structure and role as an extracellular matrix protein make it a highly relevant material for controlling cell function and mimicking tissue properties. Collagen type I is abundant in a number of tissues, and can be isolated as a purified protein. This review focuses on hydrogel biomaterials made by reconstituting collagen type I from a solubilized form, with an emphasis on in vitro studies in which collagen structure can be controlled. The hierarchical structure of collagen from the nanoscale to the macroscale is described, with an emphasis on how structure is related to function across scales. Methods of reconstituting collagen into hydrogel materials are presented, including molding of macroscopic constructs, creation of microscale modules, and electrospinning of nanoscale fibers. The modification of collagen biomaterials to achieve desired structures and functions is also addressed, with particular emphasis on mechanical control of collagen structure, creation of collagen composite materials, and crosslinking of collagenous matrices. Biomaterials scientists have made remarkable progress in rationally designing collagen-based biomaterials and in applying them to both the study of biology and for therapeutic benefit. This broad review illustrates recent examples of techniques used to control collagen structure, and to thereby direct its biological and mechanical functions. PMID:24012608

  13. Flexible Fabrication of Shape-Controlled Collagen Building Blocks for Self-Assembly of 3D Microtissues.

    PubMed

    Zhang, Xu; Meng, Zhaoxu; Ma, Jingyun; Shi, Yang; Xu, Hui; Lykkemark, Simon; Qin, Jianhua

    2015-08-12

    Creating artificial tissue-like structures that possess the functionality, specificity, and architecture of native tissues remains a big challenge. A new and straightforward strategy for generating shape-controlled collagen building blocks with a well-defined architecture is presented, which can be used for self-assembly of complex 3D microtissues. Collagen blocks with tunable geometries are controllably produced and released via a membrane-templated microdevice. The formation of functional microtissues by embedding tissue-specific cells into collagen blocks with expression of specific proteins is described. The spontaneous self-assembly of cell-laden collagen blocks into organized tissue constructs with predetermined configurations is demonstrated, which are largely driven by the synergistic effects of cell-cell and cell-matrix interactions. This new strategy would open up new avenues for the study of tissue/organ morphogenesis, and tissue engineering applications.

  14. Bio-printing of collagen and VEGF-releasing fibrin gel scaffolds for neural stem cell culture.

    PubMed

    Lee, Yeong-Bae; Polio, Samuel; Lee, Wonhye; Dai, Guohao; Menon, Lata; Carroll, Rona S; Yoo, Seung-Schik

    2010-06-01

    Time-released delivery of soluble growth factors (GFs) in engineered hydrogel tissue constructs promotes the migration and proliferation of embedded cells, which is an important factor for designing scaffolds that ultimately aim for neural tissue regeneration. We report a tissue engineering technique to print murine neural stem cells (C17.2), collagen hydrogel, and GF (vascular endothelial growth factor: VEGF)-releasing fibrin gel to construct an artificial neural tissue. We examined the morphological changes of the printed C17.2 cells embedded in the collagen and its migration toward the fibrin gel. The cells showed high viability (92.89+/-2.32%) after printing, which was equivalent to that of manually-plated cells. C17.2 cells printed within 1mm from the border of VEGF-releasing fibrin gel showed GF-induced changes in their morphology. The cells printed in this range also migrated toward the fibrin gel, with the total migration distance of 102.4+/-76.1microm over 3days. The cells in the control samples (fibrin without the VEGF or VEGF printed directly in collagen) neither proliferated nor migrated. The results demonstrated that bio-printing of VEGF-containing fibrin gel supported sustained release of the GF in the collagen scaffold. The presented method can be gainfully used in the development of three-dimensional (3D) artificial tissue assays and neural tissue regeneration applications.

  15. The history and principles of chemical dosimetry for 3-D radiation fields: gels, polymers and plastics.

    PubMed

    Doran, Simon J

    2009-03-01

    Over recent decades, modern protocols of external beam radiotherapy have been developed that involve very steep dose gradients and are thus extremely sensitive to errors in treatment delivery. A recent credentialling study by the Radiological Physics Center at the MD Anderson Cancer Center (Texas, USA) has noted potentially significant inaccuracies in test treatments at a variety of institutions. 3-D radiation dosimetry (often referred to as "gel dosimetry") may have an important role in commissioning new treatment protocols, to help prevent this type of error. This article discusses the various techniques of 3-D radiation dosimetry, with a focus on the types of radiosensitive samples used and on the optical computed tomography readout technique.

  16. Focusing optics of a parallel beam CCD optical tomography apparatus for 3D radiation gel dosimetry.

    PubMed

    Krstajić, Nikola; Doran, Simon J

    2006-04-21

    Optical tomography of gel dosimeters is a promising and cost-effective avenue for quality control of radiotherapy treatments such as intensity-modulated radiotherapy (IMRT). Systems based on a laser coupled to a photodiode have so far shown the best results within the context of optical scanning of radiosensitive gels, but are very slow ( approximately 9 min per slice) and poorly suited to measurements that require many slices. Here, we describe a fast, three-dimensional (3D) optical computed tomography (optical-CT) apparatus, based on a broad, collimated beam, obtained from a high power LED and detected by a charged coupled detector (CCD). The main advantages of such a system are (i) an acquisition speed approximately two orders of magnitude higher than a laser-based system when 3D data are required, and (ii) a greater simplicity of design. This paper advances our previous work by introducing a new design of focusing optics, which take information from a suitably positioned focal plane and project an image onto the CCD. An analysis of the ray optics is presented, which explains the roles of telecentricity, focusing, acceptance angle and depth-of-field (DOF) in the formation of projections. A discussion of the approximation involved in measuring the line integrals required for filtered backprojection reconstruction is given. Experimental results demonstrate (i) the effect on projections of changing the position of the focal plane of the apparatus, (ii) how to measure the acceptance angle of the optics, and (iii) the ability of the new scanner to image both absorbing and scattering gel phantoms. The quality of reconstructed images is very promising and suggests that the new apparatus may be useful in a clinical setting for fast and accurate 3D dosimetry.

  17. Microstructure, rheological and wound healing properties of collagen-based gel from cuttlefish skin.

    PubMed

    Jridi, Mourad; Bardaa, Sana; Moalla, Dorsaf; Rebaii, Tarak; Souissi, Nabil; Sahnoun, Zouheir; Nasri, Moncef

    2015-01-01

    Collagen-based biomaterials are of the utmost importance for tissue engineering and regenerative medicine. The aims of the present investigation were to evaluate structural and rheological properties of collagen-based gel obtained from cuttlefish skin, and to investigate its ability to enhance wound healing. Scanning electron microscopy of resulted gel showed a dense fibrillar microstructure with high interconnection network with a smaller pore size. In addition, the rheological characterization of collagen gel showed an excellent reversibility, when subjected to a temperature variation. Moreover, in the wound-healing study, topical application of collagen based gel increased significantly the percentage of wound closure over a period of 12 days, when compared to the untreated and CICAFLORA(®)-treated groups. Wound-healing activity of collagen gel was confirmed by histopathology study. Thus, cuttlefish collagen based gel might be useful as a wound healing agent.

  18. Improved MAGIC gel for higher sensitivity and elemental tissue equivalent 3D dosimetry

    SciTech Connect

    Zhu Xuping; Reese, Timothy G.; Crowley, Elizabeth M.; El Fakhri, Georges

    2010-01-15

    Purpose: Polymer-based gel dosimeter (MAGIC type) is a preferable phantom material for PET range verification of proton beam therapy. However, improvement in elemental tissue equivalency (specifically O/C ratio) is very desirable to ensure realistic time-activity measurements. Methods: Glucose and urea was added to the original MAGIC formulation to adjust the O/C ratio. The dose responses of the new formulations were tested with MRI transverse relaxation rate (R2) measurements. Results: The new ingredients improved not only the elemental composition but also the sensitivity of the MAGIC gel. The O/C ratios of our new gels agree with that of soft tissue within 1%. The slopes of dose response curves were 1.6-2.7 times larger with glucose. The melting point also increased by 5 deg. C. Further addition of urea resulted in a similar slope but with an increased intercept and a decreased melting point. Conclusions: Our improved MAGIC gel formulations have higher sensitivity and better elemental tissue equivalency for 3D dosimetry applications involving nuclear reactions.

  19. Polarized Raman anisotropic response of collagen in tendon: towards 3D orientation mapping of collagen in tissues.

    PubMed

    Galvis, Leonardo; Dunlop, John W C; Duda, Georg; Fratzl, Peter; Masic, Admir

    2013-01-01

    In this study, polarized Raman spectroscopy (PRS) was used to characterize the anisotropic response of the amide I band of collagen as a basis for evaluating three-dimensional collagen fibril orientation in tissues. Firstly, the response was investigated theoretically by applying classical Raman theory to collagen-like peptide crystal structures. The theoretical methodology was then tested experimentally, by measuring amide I intensity anisotropy in rat tail as a function of the orientation of the incident laser polarization. For the theoretical study, several collagen-like triple-helical peptide crystal structures obtained from the Protein Data Bank were rotated "in plane" and "out of plane" to evaluate the role of molecular orientation on the intensity of the amide I band. Collagen-like peptides exhibit a sinusoidal anisotropic response when rotated "in plane" with respect to the polarized incident laser. Maximal intensity was obtained when the polarization of the incident light is perpendicular to the molecule and minimal when parallel. In the case of "out of plane" rotation of the molecular structure a decreased anisotropic response was observed, becoming completely isotropic when the structure was perpendicular to the plane of observation. The theoretical Raman response of collagen was compared to that of alpha helical protein fragments. In contrast to collagen, alpha helices have a maximal signal when incident light is parallel to the molecule and minimal when perpendicular. For out-of-plane molecular orientations alpha-helix structures display a decreased average intensity. Results obtained from experiments on rat tail tendon are in excellent agreement with the theoretical predictions, thus demonstrating the high potential of PRS for experimental evaluation of the three-dimensional orientation of collagen fibers in biological tissues.

  20. Type I collagen gel protects murine fibrosarcoma L929 cells from TNFα-induced cell death

    SciTech Connect

    Wang, Hong-Ju; He, Wen-Qi; Chen, Ling; Liu, Wei-Wei; Xu, Qian; Xia, Ming-Yu; Hayashi, Toshihiko; Fujisaki, Hitomi; Hattori, Shunji; Tashiro, Shin-ichi; Onodera, Satoshi; Ikejima, Takashi

    2015-02-20

    Murine fibrosarcoma L929 cells have been used to test efficacy of proinflammatory cytokine TNFα. In the present study, we reported on protective effect of type I collagen gel used as L929 cell culture. L929 cell grew and proliferated well on collagen gel. However, the L929 cells exhibited cobblestone-like morphology which was much different from the spread fusiform shape when cultured on conventional cell dishes as well as the cells tended to aggregate. On conventional cell culture dishes, the cells treated with TNFα became round in shape and eventually died in a necroptotic manner. The cells cultured on collagen gel, however, were completely unaffected. TNFα treatment was reported to induce autophagy in L929 cells on the plastic dish, and therefore we investigated the effect of collagen gel on induction of autophagy. The results indicated that autophagy induced by TNFα treatment was much reduced when the cells were cultured on collagen gel. In conclusion, type I collagen gel protected L929 cell from TNFα-induced cell death. - Highlights: • Collagen gel culture changed the morphology of L929 cells. • L929 cell cultured on collagen gel were resistant to TNFα-induced cell death. • Collagen gel culture inhibited TNFα-induced autophagy in L929 cells.

  1. Study of a non-diffusing radiochromic gel dosimeter for 3D radiation dose imaging

    NASA Astrophysics Data System (ADS)

    Marsden, Craig Michael

    2000-12-01

    This thesis investigates the potential of a new radiation gel dosimeter, based on nitro-blue tetrazolium (NBTZ) suspended in a gelatin mold. Unlike all Fricke based gel dosimeters this dosimeter does not suffer from diffusive loss of image stability. Images are obtained by an optical tomography method. Nitro blue tetrazolium is a common biological indicator that when irradiated in an aqueous medium undergoes reduction to a highly colored formazan, which has an absorbance maximum at 525nm. Tetrazolium is water soluble while the formazan product is insoluble. The formazan product sticks to the gelatin matrix and the dose image is maintained for three months. Methods to maximize the sensitivity of the system were evaluated. It was found that a chemical detergent, Triton X-100, in combination with sodium formate, increased the dosimeter sensitivity significantly. An initial G-value of formazan production for a dosimeter composed of 1mM NBTZ, gelatin, and water was on the order of 0.2. The addition of Triton and formate produced a G-value in excess of 5.0. The effects of NBTZ, triton, formate, and gel concentration were all investigated. All the gels provided linear dose vs. absorbance plots for doses from 0 to >100 Gy. It was determined that gel concentration had minimal if any effect on sensitivity. Sensitivity increased slightly with increasing NBTZ concentration. Triton and formate individually and together provided moderate to large increases in dosimeter sensitivity. The dosimeter described in this work can provide stable 3D radiation dose images for all modalities of radiation therapy equipment. Methods to increase sensitivity are developed and discussed.

  2. Control of dense collagen gel scaffolds for tissue engineering through measurement and modelling of hydraulic permeability

    NASA Astrophysics Data System (ADS)

    Serpooshan, Vahid

    Among various natural biopolymers, type I collagen gels have demonstrated the highest potential as biomimetic scaffolds for tissue engineering (TE). However, the successful application of collagen gels requires a greater understanding of the relationship between their microstructure and physical-mechanical properties. Therefore, a precise method to modulate collagen gel microstructure in order to attain optimal scaffold properties for diverse biomedical applications is necessary. This dissertation describes a new approach to produce collagen gels with defined microstructures, quantified by hydraulic permeability ( k), in order to optimize scaffold properties for TE applications. It was hypothesized that the measurement of k can be used to study the role of microstructure in collagen gel properties, as well as cell function and cell-scaffold interactions. Applying increasing levels of plastic compression (PC) to the highly hydrated collagen gels resulted in an increase in collagen fibrillar density, reduced Happel model derived k values, increased gel stiffness, promoted MSC metabolic activity, osteogenic differentiation, and mineral deposition, while cell-induced gel contraction diminished. Thus, collagen gels with lower k and higher stiffness values exhibited greater potential for bone tissue engineering. Correlating between collagen gel microstructure, k, and fibroblast function within collagen gels indicated that increasing the level of PC yielded a reduction in pore size and an increase in fibril bundle diameter. Decrease in k values resulted in a decrease in gel contraction and an increase in cell metabolic activity. An increase in cell density accelerated contraction. Therefore, fibroblast function within collagen gels can be optimised by a balance between the microstructure, k, and cell seeding density. Developing a micromechanical model to measure experimental k of collagen gels during confined compression revealed the formation of a dense collagen lamella

  3. The enhancement of cancer stem cell properties of MCF-7 cells in 3D collagen scaffolds for modeling of cancer and anti-cancer drugs.

    PubMed

    Chen, Lei; Xiao, Zhifeng; Meng, Yue; Zhao, Yannan; Han, Jin; Su, Guannan; Chen, Bing; Dai, Jianwu

    2012-02-01

    Three-dimensional (3D) culture could partially simulate in vivo conditions. In this work, we developed a 3D collagen scaffold to investigate cellular properties of MCF-7 cells. The porous scaffolds not only induced the diversification of cell morphologies but also extended cell proliferation. The expression of pro-angiogenic growth factors and the transcriptions of matrix metalloproteinases (MMPs) were significantly increased in cells cultured in 3D collagen scaffolds. In addition, 3D collagen scaffolds could generate a cell population with the properties of cancer stem cells (CSCs). The upregulation of EMT markers and the downregulation of the epithelial cell marker were observed in cells cultured in collagen scaffolds. The expression of stem cell markers, including OCT4A and SOX2, and breast cancer stem cell signatures, including SOX4, JAG1 and CD49F, was significantly unregulated in 3D collagen scaffolds. The proportion of cells with CSC-like CD44(+)/CD24(-/low) phenotype was notably increased. High-level expression of CSC-associated properties of MCF-7 cells cultured in 3D was further confirmed by high tumorigenicity in vivo. Moreover, xenografts with 3D cells formed larger tumors. The properties of MCF-7 cells in 3D may have partially simulated their in vivo behaviors. Thus, 3D collagen scaffolds might provide a useful platform for anti-cancer therapeutics and CSC research.

  4. A method for the quantification of the pressure dependent 3D collagen configuration in the arterial adventitia.

    PubMed

    Schrauwen, J T C; Vilanova, A; Rezakhaniha, R; Stergiopulos, N; van de Vosse, F N; Bovendeerd, P H M

    2012-11-01

    Collagen plays an important role in the response of the arterial wall to mechanical loading and presumably has a load-bearing function preventing overdistension. Collagen configuration is important for understanding this role, in particular in mathematical models of arterial wall mechanics. In this study a new method is presented to image and quantify this configuration. Collagen in the arterial adventitia is stained with CNA35, and imaged in situ at high resolution with confocal microscopy at luminal pressures from 0 to 140mm Hg. The images are processed with a new automatic approach, utilizing techniques intended for MRI-DTI data. Collagen configuration is quantified through three parameters: the waviness, the transmural angle and the helical angle. The method is demonstrated for the case of carotid arteries of the white New Zealand rabbit. The waviness indicated a gradual straightening between 40 and 80mm Hg. The transmural angle was about zero indicating that the fibers stayed within an axial-circumferential plane at all pressures. The helical angle was characterized by a symmetrical distribution around the axial direction, indicating a double symmetrical helix. The method is the first to combine high resolution imaging with a new automatic image processing approach to quantify the 3D configuration of collagen in the adventitia as a function of pressure.

  5. Open-Source-Based 3D Printing of Thin Silica Gel Layers in Planar Chromatography.

    PubMed

    Fichou, Dimitri; Morlock, Gertrud E

    2017-02-07

    On the basis of open-source packages, 3D printing of thin silica gel layers is demonstrated as proof-of-principle for use in planar chromatography. A slurry doser was designed to replace the plastic extruder of an open-source Prusa i3 printer. The optimal parameters for 3D printing of layers were studied, and the planar chromatographic separations on these printed layers were successfully demonstrated with a mixture of dyes. The layer printing process was fast. For printing a 0.2 mm layer on a 10 cm × 10 cm format, it took less than 5 min. It was affordable, i.e., the running costs for producing such a plate were less than 0.25 Euro and the investment costs for the modified hardware were 630 Euro. This approach demonstrated not only the potential of the 3D printing environment in planar chromatography but also opened new avenues and new perspectives for tailor-made plates, not only with regard to layer materials and their combinations (gradient plates) but also with regard to different layer shapes and patterns. As such an example, separations on a printed plane layer were compared with those obtained from a printed channeled layer. For the latter, 40 channels were printed in parallel on a 10 cm × 10 cm format for the separation of 40 samples. For producing such a channeled plate, the running costs were below 0.04 Euro and the printing process took only 2 min. All modifications of the device and software were released open-source to encourage reuse and improvements and to stimulate the users to contribute to this technology. By this proof-of-principle, another asset was demonstrated to be integrated into the Office Chromatography concept, in which all relevant steps for online miniaturized planar chromatography are performed by a single device.

  6. 3D bioprinting of GelMA scaffolds triggers mineral deposition by primary human osteoblasts.

    PubMed

    McBeth, Christine; Lauer, Jasmin; Ottersbach, Michael; Campbell, Jennifer; Sharon, Andre; Sauer-Budge, Alexis F

    2017-01-10

    Due to its relatively low level of antigenicity and high durability, titanium has successfully been used as the major material for biological implants. However, because the typical interface between titanium and tissue precludes adequate transmission of load into the surrounding bone, over time, load-bearing implants tend to loosen and revision surgeries are required. Osseointegration of titanium implants requires presentation of both biological and mechanical cues that promote attachment of and trigger mineral deposition by osteoblasts. While many factors contribute to differentiation, the relative importance of the various cues is unclear. To substantially improve osseointegration of titanium implants, we generated a gelatin methacryloyl (GelMA) scaffold, using an extrusion-based 3D bioprinter, which can be directly printed on and grafted to the titanium implant surface. We demonstrate that this scaffold is able to trigger mineral deposition of both MG63 osteoblasts and primary normal human osteoblasts in the absence of any exogenous osteogenic factors. Films of the same formulation failed to promote mineral deposition suggesting that the three dimensional scaffold was able to tip the balance in favor of differentiation despite other potentially unfavorable differentiation cues of the material. We further show that these GelMA lattices can be directly grafted to titanium alloy and are secure in vitro over a period of seven weeks. When grafted within a groove system, the GelMA hydrogel is protected from shearing forces in a marrow implantation model. This prepares the way for osteogenic coatings to be directly manufactured on the implant surface and packaged for surgery.

  7. 3D bioprinting of GelMA scaffolds triggers mineral deposition by primary human osteoblasts.

    PubMed

    McBeth, Christine; Lauer, Jasmin; Ottersbach, Michael; Campbell, Jennifer; Sharon, Andre; Sauer-Budge, Alexis

    2016-12-14

    Due to its relatively low level of antigenicity and high durability, titanium has successfully been used as the major material for biological implants. However, because the typical interface between titanium and tissue precludes adequate transmission of load into the surrounding bone, over time, load-bearing implants tend to loosen and revision surgeries are required. Osseointegration of titanium implants requires presentation of both biological and mechanical cues that promote attachment of and trigger mineral deposition by osteoblasts. While many factors contribute to differentiation, the relative importance of the various cues is unclear. To substantially improve osseointegration of titanium implants, we generated a gelatin methacryloyl (GelMA) scaffold, using an extrusion-based 3D bioprinter, which can be directly printed on and grafted to the titanium implant surface. We demonstrate that this scaffold is able to trigger mineral deposition of both MG63 osteoblasts and normal human primary osteoblasts in the absence of any exogenous osteogenic factors. Films of the same formulation failed to promote mineral deposition suggesting that the three dimensional scaffold was able to tip the balance in favor of differentiation despite other potentially unfavorable differentiation cues of the material. We further show that these GelMA lattices can be directly grafted to titanium alloy and are secure in vitro over a period of seven weeks. When grafted within a groove system, the GelMA hydrogel is protected from shearing forces in a marrow implantation model. This prepares the way for osteogenic coatings to be directly manufactured on the implant surface and packaged for surgery.

  8. Properties and modification of porous 3-D collagen/hydroxyapatite composites.

    PubMed

    Sionkowska, A; Kozłowska, J

    2013-01-01

    A freeze drying technique was used to form porous three-dimensional collagen matrixes modified by the addition of a variable amount of nano-hydroxyapatite. For chemical cross-linking EDC/NHS were used. Physical cross-linking was achieved by dehydrothermal treatment. Mechanical properties, morphology, dissolution, porosity, density, enzymatic degradation and swelling properties of materials have been studied after cross-linking. The density of scaffolds and its compressive modulus increased with an increasing amount of hydroxyapatite and collagen concentration in the composite scaffold, while the swelling ratio and porosity decreased. The studied scaffolds dissolved slowly in PBS solution. DHT cross-linked collagen matrices showed a much faster degradation rate after exposure to collagenase than the EDC cross-linked samples.

  9. [Experimental study of the collagen matrix for increase the gums using a 3D-modeling].

    PubMed

    Baulin, I M; Badalyan, V A; Ryakhovsky, A N

    2015-01-01

    In an experimental study on mini-pigs demonstrated that the use of collagen matrix Mucograft open method leads to the formation of mature connective tissue around the implants, more pronounced after 70 days, and the width of attached mucosa already 45th day (from 4.4 ± 0.3 to 7.7 ± 0.5 mm) is comparable to that of free gingival graft. Three-dimensional computer modeling of jaws experimental animals showed the soft tissue augmentation by 0.8 ± 0.1 cm3 after use of collagen matrix Mucograft and 1.1 ± 0.12 cm3 after free gingival graft.

  10. Microfluidic assay of endothelial cell migration in 3D interpenetrating polymer semi-network HA-Collagen hydrogel.

    PubMed

    Jeong, Gi Seok; Kwon, Gu Han; Kang, Ah Ran; Jung, Bo Young; Park, Yongdoo; Chung, Seok; Lee, Sang-Hoon

    2011-08-01

    Cell migration through the extracellular matrix (ECM) is one of the key features for physiological and pathological processes such as angiogenesis, cancer metastasis, and wound healing. In particular, the quantitative assay of endothelial cell migration under the well-defined three dimensional (3D) microenvironment is important to analyze the angiogenesis mechanism. In this study, we report a microfluidic assay of endothelial cell sprouting and migration into an interpenetrating polymer semi-network HA-Collagen (SIPNs CH) hydrogel as ECM providing an enhanced in vivo mimicking 3D microenvironment to cells. The microfluidic chip could provide a well-controlled gradient of growth factor to cells, whereas the hydrogel could mimic a well-defined 3D microenvironment in vivo. (In addition/Furthermore, the microfluidic chip gives a well-controlled gradient of growth factor to cells) For this reason, three types of hydrogel, composed of semi-interpenetrating networks of collagen and hyaluronic acid were prepared, and firstly we proved the role of the hydrogel in endothelial cell migration. The diffusion property and swelling ratio of the hydrogel were characterized. It modulated the migration of endothelial cells in quantified manner, also being influenced by additional synthesis of Matrix metalloproteinase(MMP)-sensitive remodeling peptides and Arginine-glycine-lycinee (RGD) cell adhesion peptides. We successfully established a novel cell migration platform by changing major determinants such as ECM material under biochemical synthesis and under growth factor gradients in a microfluidic manner.

  11. Condensed cellular seeded collagen gel as an improved biomaterial for tissue engineering of articular cartilage.

    PubMed

    Mueller-Rath, Ralf; Gavénis, Karsten; Andereya, Stefan; Mumme, Torsten; Albrand, Monique; Stoffel, Marcus; Weichert, Dieter; Schneider, Ulrich

    2010-01-01

    Three-dimensional autologous chondrocyte implantation based on collagen gel as matrix scaffold has become a clinically applied treatment for focal defects of articular cartilage. However, the low biomechanical properties of collagen gel makes intraoperative handling difficult and creates the risk of early damages to the vulnerable implant. The aim of the study was to create a stabilized form of collagen gel and to evaluate its biomechanical and biochemical properties.Collagen type-I gel was seeded with human articular chondrocytes. 20 samples were subject to condensation which was achieved mechanically by compression and filtration. Control samples were left uncondensed. From both types of gels 10 samples were used for initial biomechanical evaluation by means of unconfined compression and 10 samples were cultivated under standard conditions in vitro. Following cultivation the samples were evaluated by conventional histology and immunohistochemistry. The proliferation rate was calculated and matrix gene expression was quantified by real-time PCR.The biomechanical tests revealed a higher force carrying capacity of the condensed specimens. Strain rate dependency and relaxation was seen in both types of collagen gel representing viscoelastic material properties. Cells embedded within the condensed collagen gel were able to produce extracellular matrix proteins and showed proliferation.Condensed collagen gel represents a mechanically improved type of biomaterial which is suitable for three-dimensional autologous chondrocyte implantation.

  12. Influence of collagen gel on the orientation of epithelial cell polarity: follicle formation from isolated thyroid cells and from preformed monolayers

    PubMed Central

    1981-01-01

    The influence of collagen gels on the orientation of the polarity of epithelial thyroid cells in culture was studied under four different conditions. (a) Isolated cells cultured on the surface of a collagen gel formed a monolayer. The apical pole was in contact with the culture medium and the basal membrane was attached to the substratum. (b) Isolated cells embedded inside the gel organized within 8 into follicles. The basal pole was in contact with collagen and the apical pole was oriented towards the interior of the follicular lumen. (c) Cells were first organized into floating vesicles, structures in which the apical surface is in contact with the culture medium, and the vesicles were embedded inside the collagen gel. After 3 d, cell polarity was inverted, the apical pole being oriented towards the cavity encompassed by cells. Vesicles had been transformed into follicles. (d) Monolayers formed on collagen gels as in a were overlaid with a second layer of collagen, which was polymerized in contact with the apical cell surface. A disorganization of the continuous pavement occurred within 24 h; cells attached to the upper layer of collagen and reorganized into follicles in the collagen sandwich within 4-8 d. A similar process occurred when the monolayer was grown on plastic and overlaid with collagen, or grown on collagen and covered with small pieces of glass cover slips. No reorganization was observed between two glass surfaces. In conclusion, first, a basal pole was always formed in the area of contact between the cell membrane and an adhesive surface and, second, the interaction of a preformed apical pole with an adhesive surface was not compatible with the stability of this domain of the plasma membrane. The interaction of the cell membrane with extracellular components having adhesive properties appears to be a determinant factor in the orientation and stabilization of epithelial cell polarity. PMID:7298715

  13. Epidermal growth factor improves the migration and contractility of aged fibroblasts cultured on 3D collagen matrices.

    PubMed

    Kim, Daehwan; Kim, So Young; Mun, Seog Kyun; Rhee, Sangmyung; Kim, Beom Joon

    2015-04-01

    Epidermal growth factor (EGF) plays a critical role in fibroblasts by stimulating the production of collagen and supports cell renewal through the interaction between keratinocytes and fibroblasts. It is well known that the contractile activity of fibroblasts is required for the remodeling of the extracellular matrix (ECM), which contributes to skin elasticity. However, the role of EGF in the contraction of aged fibroblasts under 3-dimensional (3D) culture conditions is not yet fully understood. In the present study, we demonstrated that young fibroblasts spread and proliferated more rapidly than aged fibroblasts under 2-dimensional (2D) culture conditions. Cell migration assay using a nested collagen matrix revealed that the migration of young fibroblasts was also greater than that of aged fibroblasts under 3D culture conditions. However, the addition of recombinant human EGF (rhEGF) resulted in the enhanced migration of aged fibroblasts; the migration rate was similar to that of the young fibroblasts. The aged fibroblasts showed decreased cluster formation compared with the young fibroblasts on the collagen matrix, which was improved by the addition of rhEGF. Furthermore, cell contraction assay revealed that the basal contractility of the aged fibroblasts was lower than that of the young fibroblasts; however, following treatment with rhEGF, the contractility was restored to levels similar or even higher to those of the young fibroblasts. Taken together, our results suggest that rhEGF is a potential renewal agent that acts to improve the migration and contraction of aged fibroblasts more efficiently than young fibroblasts under 3D culture conditions; thus, EGF may have valuable regenerative effects on aged skin.

  14. Human platelet lysate improves human cord blood derived ECFC survival and vasculogenesis in three dimensional (3D) collagen matrices.

    PubMed

    Kim, Hyojin; Prasain, Nutan; Vemula, Sasidhar; Ferkowicz, Michael J; Yoshimoto, Momoko; Voytik-Harbin, Sherry L; Yoder, Mervin C

    2015-09-01

    Human cord blood (CB) is enriched in circulating endothelial colony forming cells (ECFCs) that display high proliferative potential and in vivo vessel forming ability. Since diminished ECFC survival is known to dampen the vasculogenic response in vivo, we tested how long implanted ECFC survive and generate vessels in three-dimensional (3D) type I collagen matrices in vitro and in vivo. We hypothesized that human platelet lysate (HPL) would promote cell survival and enhance vasculogenesis in the 3D collagen matrices. We report that the percentage of ECFC co-cultured with HPL that were alive was significantly enhanced on days 1 and 3 post-matrix formation, compared to ECFC alone containing matrices. Also, co-culture of ECFC with HPL displayed significantly more vasculogenic activity compared to ECFC alone and expressed significantly more pro-survival molecules (pAkt, p-Bad and Bcl-xL) in the 3D collagen matrices in vitro. Treatment with Akt1 inhibitor (A-674563), Akt2 inhibitor (CCT128930) and Bcl-xL inhibitor (ABT-263/Navitoclax) significantly decreased the cell survival and vasculogenesis of ECFC co-cultured with or without HPL and implicated activation of the Akt1 pathway as the critical mediator of the HPL effect on ECFC in vitro. A significantly greater average vessel number and total vascular area of human CD31(+) vessels were present in implants containing ECFC and HPL, compared to the ECFC alone implants in vivo. We conclude that implantation of ECFC with HPL in vivo promotes vasculogenesis and augments blood vessel formation via diminishing apoptosis of the implanted ECFC.

  15. Sol-gel assisted fabrication of collagen hydrolysate composite scaffold: a novel therapeutic alternative to the traditional collagen scaffold.

    PubMed

    Ramadass, Satiesh Kumar; Perumal, Sathiamurthi; Gopinath, Arun; Nisal, Anuya; Subramanian, Saravanan; Madhan, Balaraman

    2014-09-10

    Collagen is one of the most widely used biomaterial for various biomedical applications. In this Research Article, we present a novel approach of using collagen hydrolysate, smaller fragments of collagen, as an alternative to traditionally used collagen scaffold. Collagen hydrolysate composite scaffold (CHCS) was fabricated with sol-gel transition procedure using tetraethoxysilane as the silica precursor. CHCS exhibits porous morphology with pore sizes varying between 380 and 780 μm. Incorporation of silica conferred CHCS with controlled biodegradation and better water uptake capacity. Notably, 3T3 fibroblast proliferation was seen to be significantly better under CHCS treatment when compared to treatment with collagen scaffold. Additionally, CHCS showed excellent antimicrobial activity against the wound pathogens Staphylococcus aureus, Bacillus subtilis, and Escherichia coli due to the inherited antimicrobial activity of collagen hydrolysate. In vivo wound healing experiments with full thickness excision wounds in rat model demonstrated that wounds treated with CHCS showed accelerated healing when compared to wounds treated with collagen scaffold. These findings indicate that the CHCS scaffold from collagen fragments would be an effective and affordable alternative to the traditionally used collagen structural biomaterials.

  16. 3D porous sol-gel matrix incorporated microdevice for effective large volume cell sample pretreatment.

    PubMed

    Lee, Chan Joo; Jung, Jae Hwan; Seo, Tae Seok

    2012-06-05

    In this study, we demonstrated an effective sample pretreatment microdevice that could perform the capture, purification, and release of pathogenic bacteria with a large-volume sample and at a high speed and high-capture yield. We integrated a sol-gel matrix into the microdevice which forms three-dimensional (3D) micropores for the cell solution to pass through and provides a large surface area for the immobilization of antibodies to capture the target Staphylococcus aureus (S. aureus) cells. The antibody was linked to the surface of the sol-gel via a photocleavable linker, allowing the cell-captured antibody moiety to be released by UV irradiation. In addition to the optimization of the antibody immobilization and UV cleavage processes, the cell-capture efficiency was maximized by controlling the sample flow rate with a pumping scheme (3 steps, 5 steps: 3 steps with one flutter step, 7 steps: 3 steps with two flutter steps) and the pumping time (100, 200, and 300 ms). A quantitative capture analysis was performed by targeting a specific gene site of protein A of S. aureus in real-time PCR (RT-PCR). While the 3-step process with an actuation time of 100 ms showed the fastest flow rate (1 mL sample processing time in 10 min), the pumping scheme with the 7-step process and the 300 ms actuation time revealed the highest cell-capture efficiency. A limit of detection study with the 7-step and the 300 ms pumping scheme demonstrated that 100 cells per 100 μL were detected with a 70% yield, and even a single cell could be analyzed via on-chip sample preparation. Thus, our novel sol-gel based microdevice was proven more cost-effective, simple, and efficient in terms of its sample pretreatment ability compared to the use of a conventional 2D flat microdevice. This proposed sample pretreatment device can be further incorporated to an analytical functional unit to realize a micrototal analysis system (μTAS) with sample-in-answer-out capability in the fields of biomedical

  17. Modulating the Rigidity and Mineralization of Collagen Gels Using Poly(Lactic-Co-Glycolic Acid) Microparticles

    PubMed Central

    DeVolder, Ross J.; Kim, Il Won; Kim, Eun-Suk

    2012-01-01

    Extensive efforts have been made to prepare osteoconductive collagen gels for the regeneration of normal bone and the pathological examination of diseased bone; however, collagen gels are often plagued by limited controllability of their rigidity and mineral deposition. This study reports a simple but efficient strategy that tunes the mechanical properties of, and apatite formation in, collagen gels by incorporating hydrolyzable poly(lactic-co-glycolic acid) (PLGA) microparticles within the gels. The PLGA microparticles are associated with the collagen fibrils and increased both the gel's elasticity and rigidity while minimally influencing its permeability. As compared with pure collagen gels, the PLGA microparticle-filled collagen gels, termed PLGA-Col hydrogels, significantly enhanced the deposition of apatite-like minerals within the gels when incubated in simulated body fluid or encapsulated with mesenchymal stem cells (MSCs) undergoing osteogenic differentiation. Finally, PLGA-Col hydrogels mineralized by differentiated MSCs led to an enhanced formation of bone-like tissues within the hydrogels. Overall, the PLGA-Col hydrogel system developed in this study will serve to improve the quality of osteoconductive matrices for both fundamental and clinical studies that are relevant to bone repair, regeneration, and pathogenesis. PMID:22480235

  18. Multiphoton microscopy of engineered dermal substitutes: assessment of 3D collagen matrix remodeling induced by fibroblasts contraction

    NASA Astrophysics Data System (ADS)

    Pena, A.-M.; Olive, C.; Michelet, J.-F.; Galey, J.-B.; Fagot, D.; Leroy, F.; Martin, J.-L.; Colonna, A.; Schanne-Klein, M.-C.

    2010-02-01

    One of the main functions of dermal fibroblasts is the generation of mechanical forces within their surrounding extracellular matrix. Investigating molecules that could modulate fibroblast contraction and act as potent anti aging ingredients requires the development of three-dimensional in situ imaging methodologies for dermal substitute analysis. Here we use multiphoton microscopy in order to investigate the fibroblast-induced collagen matrix reorganization in engineered dermal tissue and to evaluate the effect of Y27632, a RhoA kinase inhibitor on dermal substitutes contraction. We observe that collagen fibrils rearrange around fibroblast with increasing density in control samples, whereas collagen fibrils show no remodeling in the samples containing the RhoA kinase inhibitor. Moreover, when the culture medium containing the inhibitor was replaced with a control medium, the dermal substitutes presented the same 3D reorganization as the control samples, which indicates that the inhibitory effects are reversible. In conclusion, our study demonstrates the relevance of multiphoton microscopy to visualize three-dimensional remodeling of the matrix induced by fibroblast contraction.

  19. Effect of papain-based gel on type I collagen - spectroscopy applied for microstructural analysis

    PubMed Central

    Júnior, Zenildo Santos Silva; Botta, Sergio Brossi; Ana, Patricia Aparecida; França, Cristiane Miranda; Fernandes, Kristianne Porta Santos; Mesquita-Ferrari, Raquel Agnelli; Deana, Alessandro; Bussadori, Sandra Kalil

    2015-01-01

    Considering the improvement of biomaterials that facilitate atraumatic restorative techniques in dentistry, a papain-based gel can be used in the chemomechanical removal of decayed dental tissue. However, there is no information regarding the influence of this gel on the structure of sound collagen. The aim of the present study was to investigate the adsorption of a papain-based gel (PapacarieTM) to collagen and determine collagen integrity after treatment. A pilot study was first performed with 10 samples of type I collagen membrane obtained from bovine Achilles deep tendon to compare the influence of hydration (Milli-Q water) on infrared bands of collagen. In a further experiment, 10 samples of type I collagen membrane were used to evaluate the effects of PapacarieTM on the collagen microstructure. All analyses were performed using the attenuated total reflectance technique of Fourier transform infrared (ATR-FTIR). The results demonstrated that the application of PapacarieTM does not lead to the degradation of collagen and this product can be safely used in minimally invasive dentistry. As the integrity of sound collagen is preserved after the application of the papain-based gel, this product is indicated for the selective removal of infected dentin, leaving the affected dentin intact and capable of re-mineralization. PMID:26101184

  20. Development of gel materials with high transparency and mechanical strength for use with a 3D gel printer SWIM-ER

    NASA Astrophysics Data System (ADS)

    Tase, Taishi; Okada, Koji; Takamatsu, Kyuichiro; Saito, Azusa; Kawakami, Masaru; Furukawa, Hidemitsu

    2016-04-01

    Medical doctors use artificial blood vessels and organ models, which are usually made of plastic, to explain operations to students, or patients awaiting treatment. However, there are some problems such as the high cost of making the model and there is not a realistic feel because the model is hard. These problems can be solved using soft and wet material for instance gel. Gels are materials with unique properties such as transparency, biocompatibility, and low friction. In recent years, high strength gel has been developed and is expected to be applied in medical fields in the future. Artificial models of gel can be produced by 3D gel printers. Our group has been developing a 3D gel printer with 1mm precision in printing, but the shape, size and mechanical strength are not sufficient for medical models. In this study, we overcome these problems and make a gel model which is transparent, mechanically strong with a fine shape. The strength and molding accuracy is improved by changing and preparing the cross linker and ultraviolet absorber. We conducted mechanical and molding tests to confirm that the gel material properties improved.

  1. Matrix density alters zyxin phosphorylation, which limits peripheral process formation and extension in endothelial cells invading 3D collagen matrices.

    PubMed

    Abbey, Colette A; Bayless, Kayla J

    2014-09-01

    This study was designed to determine the optimal conditions required for known pro-angiogenic stimuli to elicit successful endothelial sprouting responses. We used an established, quantifiable model of endothelial cell (EC) sprout initiation where ECs were tested for invasion in low (1 mg/mL) and high density (5 mg/mL) 3D collagen matrices. Sphingosine 1-phosphate (S1P) alone, or S1P combined with stromal derived factor-1α (SDF) and phorbol ester (TPA), elicited robust sprouting responses. The ability of these factors to stimulate sprouting was more effective in higher density collagen matrices. S1P stimulation resulted in a significant increase in invasion distance, and with the exception of treatment groups containing phorbol ester, invasion distance was longer in 1mg/mL compared to 5mg/mL collagen matrices. Closer examination of cell morphology revealed that increasing matrix density and supplementing with SDF and TPA enhanced the formation of multicellular structures more closely resembling capillaries. TPA enhanced the frequency and size of lumen formation and correlated with a robust increase in phosphorylation of p42/p44 Erk kinase, while S1P and SDF did not. Also, a higher number of significantly longer extended processes formed in 5mg/mL compared to 1mg/mL collagen matrices. Because collagen matrices at higher density have been reported to be stiffer, we tested for changes in the mechanosensitive protein, zyxin. Interestingly, zyxin phosphorylation levels inversely correlated with matrix density, while levels of total zyxin did not change significantly. Immunofluorescence and localization studies revealed that total zyxin was distributed evenly throughout invading structures, while phosphorylated zyxin was slightly more intense in extended peripheral processes. Silencing zyxin expression increased extended process length and number of processes, while increasing zyxin levels decreased extended process length. Altogether these data indicate that ECs

  2. Fiber based optical tweezers for simultaneous in situ force exertion and measurements in a 3D polyacrylamide gel compartment.

    PubMed

    Ti, Chaoyang; Thomas, Gawain M; Ren, Yundong; Zhang, Rui; Wen, Qi; Liu, Yuxiang

    2015-07-01

    Optical tweezers play an important role in biological applications. However, it is difficult for traditional optical tweezers based on objective lenses to work in a three-dimensional (3D) solid far away from the substrate. In this work, we develop a fiber based optical trapping system, namely inclined dual fiber optical tweezers, that can simultaneously apply and measure forces both in water and in a 3D polyacrylamide gel matrix. In addition, we demonstrate in situ, non-invasive characterization of local mechanical properties of polyacrylamide gel by measurements on an embedded bead. The fiber optical tweezers measurements agree well with those of atomic force microscopy (AFM). The inclined dual fiber optical tweezers provide a promising and versatile tool for cell mechanics study in 3D environments.

  3. Microencapsulation of Neuroblastoma Cells and Mesenchymal Stromal Cells in Collagen Microspheres: A 3D Model for Cancer Cell Niche Study

    PubMed Central

    Yeung, Pan; Sin, Hoi Shun; Chan, Shing; Chan, Godfrey Chi Fung; Chan, Barbara Pui

    2015-01-01

    There is a growing trend for researchers to use in vitro 3D models in cancer studies, as they can better recapitulate the complex in vivo situation. And the fact that the progression and development of tumor are closely associated to its stromal microenvironment has been increasingly recognized. The establishment of such tumor supportive niche is vital in understanding tumor progress and metastasis. The mesenchymal origin of many cells residing in the cancer niche provides the rationale to include MSCs in mimicking the niche in neuroblastoma. Here we co-encapsulate and co-culture NBCs and MSCs in a 3D in vitro model and investigate the morphology, growth kinetics and matrix remodeling in the reconstituted stromal environment. Results showed that the incorporation of MSCs in the model lead to accelerated growth of cancer cells as well as recapitulation of at least partially the tumor microenvironment in vivo. The current study therefore demonstrates the feasibility for the collagen microsphere to act as a 3D in vitro cancer model for various topics in cancer studies. PMID:26657086

  4. Growth of Normal Mouse Vaginal Epithelial Cells in and on Collagen Gels

    NASA Astrophysics Data System (ADS)

    Iguchi, Taisen; Uchima, Francis-Dean A.; Ostrander, Patricia L.; Bern, Howard A.

    1983-06-01

    Sustained growth in primary culture of vaginal epithelial cells from ovariectomized adult BALB/cCrg1 mice embedded within or seeded on collagen gel matrix was achieved in a serum-free medium composed of Ham's F-12 medium/Dulbecco's modified Eagle's medium, 1:1 (vol/vol), supplemented with insulin, bovine serum albumin fraction V, epidermal growth factor, cholera toxin, and transferrin. Three-dimensional growth of vaginal epithelial cells occurred inside the collagen gel matrix. Cell numbers increased 4- to 8-fold in collagen gel and about 4-fold on collagen gel after 9-10 days in culture. The effect of 17β -estradiol (0.00018-180 nM in gel or 0.018-180 nM on gel) and diethylstilbestrol (DES; 0.0186-186 nM in gel) on the growth of vaginal epithelial cells was examined. The addition of estrogen did not enhance the growth of vaginal epithelial cells during this time period either in the complete medium or in a suboptimal medium. Cultures on floating collagen gels in the serum-free medium are composed of 1-3 cell layers with superficial cornification. Estrogen does not appear to be a direct mitogen for vaginal epithelial cells, at least in this system.

  5. Rotation-based technique for the rapid densification of tubular collagen gel scaffolds.

    PubMed

    Loy, Caroline; Lainé, Audrey; Mantovani, Diego

    2016-12-01

    Type I collagen gel is often used as a tubular scaffold because of its easy molding properties as well as its biocompatibility, low immunogenicity and ability to be remodelled by cells. However, its highly hydrated structure contributes to its weak mechanical properties and reduces its ability to be handled, which is important in tubular tissue engineering. Although cell-driven remodelling of collagen matrices is known to reinforce their mechanical properties, this process can take weeks. This study introduces a novel, simple, and rapid technique using a rotational bioreactor to expel water and densify collagen under sterile conditions to generate denser and stronger collagen gel scaffolds. This process produces a dense tubular-shaped collagen gel which, compared to standard collagen gel scaffolds, shows a decreased wall thickness and a four-fold increase in collagen concentration. A denser collagen fiber network observed by immunofluorescence staining and mechanical characterisation shows a twenty-fold increase in the elastic modulus of the dense constructs which maintain cell viability inside the scaffold. Moreover, by simply modifying the scaffold mold, customised shapes and sizes can be obtained to provide a wide range of applications, including complex tubular geometries and multi-layered scaffolds for the culture of various cell types and tissues.

  6. Optical CT scanner for in-air readout of gels for external radiation beam 3D dosimetry.

    PubMed

    Ramm, Daniel; Rutten, Thomas P; Shepherd, Justin; Bezak, Eva

    2012-06-21

    Optical CT scanners for a 3D readout of externally irradiated radiosensitive hydrogels currently require the use of a refractive index (RI) matching liquid bath to obtain suitable optical ray paths through the gel sample to the detector. The requirement for a RI matching liquid bath has been negated by the design of a plastic cylindrical gel container that provides parallel beam geometry through the gel sample for the majority of the projection. The design method can be used for various hydrogels. Preliminary test results for the prototype laser beam scanner with ferrous xylenol-orange gel show geometric distortion of 0.2 mm maximum, spatial resolution limited to beam spot size of about 0.4 mm and 0.8% noise (1 SD) for a uniform irradiation. Reconstruction of a star pattern irradiated through the cylinder walls demonstrates the suitability for external beam applications. The extremely simple and cost-effective construction of this optical CT scanner, together with the simplicity of scanning gel samples without RI matching fluid increases the feasibility of using 3D gel dosimetry for clinical external beam dose verifications.

  7. Toward 3D graphene oxide gels based adsorbents for high-efficient water treatment via the promotion of biopolymers.

    PubMed

    Cheng, Chong Sage; Deng, Jie; Lei, Bei; He, Ai; Zhang, Xiang; Ma, Lang; Li, Shuang; Zhao, Changsheng

    2013-12-15

    Recent studies showed that graphene oxide (GO) presented high adsorption capacities to various water contaminants. However, the needed centrifugation after adsorption and the potential biological toxicity of GO restricted its applications in wastewater treatment. In this study, a facile method is provided by using biopolymers to mediate and synthesize 3D GO based gels. The obtained hybrid gels present well-defined and interconnected 3D porous network, which allows the adsorbate molecules to diffuse easily into the adsorbent. The adsorption experiments indicate that the obtained porous GO-biopolymer gels can efficiently remove cationic dyes and heavy metal ions from wastewater. Methylene blue (MB) and methyl violet (MV), two cationic dyes, are chosen as model adsorbates to investigate the adsorption capability and desorption ratio; meanwhile, the influence of contacting time, initial concentration, and pH value on the adsorption capacity of the prepared GO-biopolymer gels are also studied. The GO-biopolymer gels displayed an adsorption capacity as high as 1100 mg/g for MB dye and 1350 mg/g for MV dye, respectively. Furthermore, the adsorption kinetics and isotherms of the MB were studied in details. The experimental data of MB adsorption fitted well with the pseudo-second-order kinetic model and the Langmuir isotherm, and the results indicated that the adsorption process was controlled by the intraparticle diffusion. Moreover, the adsorption data revealed that the porous GO-biopolymer gels showed good selective adsorbability to cationic dyes and metal ions.

  8. Diffusion and convection in collagen gels: implications for transport in the tumor interstitium.

    PubMed Central

    Ramanujan, Saroja; Pluen, Alain; McKee, Trevor D; Brown, Edward B; Boucher, Yves; Jain, Rakesh K

    2002-01-01

    Diffusion coefficients of tracer molecules in collagen type I gels prepared from 0-4.5% w/v solutions were measured by fluorescence recovery after photobleaching. When adjusted to account for in vivo tortuosity, diffusion coefficients in gels matched previous measurements in four human tumor xenografts with equivalent collagen concentrations. In contrast, hyaluronan solutions hindered diffusion to a lesser extent when prepared at concentrations equivalent to those reported in these tumors. Collagen permeability, determined from flow through gels under hydrostatic pressure, was compared with predictions obtained from application of the Brinkman effective medium model to diffusion data. Permeability predictions matched experimental results at low concentrations, but underestimated measured values at high concentrations. Permeability measurements in gels did not match previous measurements in tumors. Visualization of gels by transmission electron microscopy and light microscopy revealed networks of long collagen fibers at lower concentrations along with shorter fibers at high concentrations. Negligible assembly was detected in collagen solutions pregelation. However, diffusion was similarly hindered in pre and postgelation samples. Comparison of diffusion and convection data in these gels and tumors suggests that collagen may obstruct diffusion more than convection in tumors. These findings have significant implications for drug delivery in tumors and for tissue engineering applications. PMID:12202388

  9. A novel collagen gel-based measurement technique for quantitation of cell contraction force.

    PubMed

    Jin, Tianrong; Li, Li; Siow, Richard C M; Liu, Kuo-Kang

    2015-05-06

    Cell contraction force plays an important role in wound healing, inflammation,angiogenesis and metastasis. This study describes a novel method to quantify single cell contraction force in vitro using human aortic adventitial fibroblasts embedded in a collagen gel. The technique is based on a depth sensing nano-indentation tester to measure the thickness and elasticity of collagen gels containing stimulated fibroblasts and a microscopy imaging system to estimate the gel area. In parallel, a simple theoretical model has been developed to calculate cell contraction force based on the measured parameters. Histamine (100 mM) was used to stimulate fibroblast contraction while the myosin light chain kinase inhibitor ML-7 (25 mM) was used to inhibit cell contraction. The collagen matrix used in the model provides a physiological environment for fibroblast contraction studies. Measurement of changes in collagen gel elasticity and thickness arising from histamine treatments provides a novel convenient technique to measure cell contraction force within a collagen matrix. This study demonstrates that histamine can elicit a significant increase in contraction force of fibroblasts embedded in collagen,while the Young's modulus of the gel decreases due to the gel degradation.

  10. A novel collagen gel-based measurement technique for quantitation of cell contraction force

    PubMed Central

    Jin, Tianrong; Li, Li; Siow, Richard C. M.; Liu, Kuo-Kang

    2015-01-01

    Cell contraction force plays an important role in wound healing, inflammation, angiogenesis and metastasis. This study describes a novel method to quantify single cell contraction force in vitro using human aortic adventitial fibroblasts embedded in a collagen gel. The technique is based on a depth sensing nano-indentation tester to measure the thickness and elasticity of collagen gels containing stimulated fibroblasts and a microscopy imaging system to estimate the gel area. In parallel, a simple theoretical model has been developed to calculate cell contraction force based on the measured parameters. Histamine (100 µM) was used to stimulate fibroblast contraction while the myosin light chain kinase inhibitor ML-7 (25 µM) was used to inhibit cell contraction. The collagen matrix used in the model provides a physiological environment for fibroblast contraction studies. Measurement of changes in collagen gel elasticity and thickness arising from histamine treatments provides a novel convenient technique to measure cell contraction force within a collagen matrix. This study demonstrates that histamine can elicit a significant increase in contraction force of fibroblasts embedded in collagen, while the Young's modulus of the gel decreases due to the gel degradation. PMID:25977960

  11. Fibroblast morphogenesis on 3D collagen matrices: the balance between cell clustering and cell migration.

    PubMed

    da Rocha-Azevedo, Bruno; Grinnell, Frederick

    2013-10-01

    Fibroblast clusters have been observed in tissues under a variety of circumstances: in fibrosis and scar, in the formation of hair follicle dermal papilla, and as part of the general process of mesenchymal condensation that takes place during development. Cell clustering has been shown to depend on features of the extracellular matrix, growth factor environment, and mechanisms to stabilize cell-cell interactions. In vitro studies have shown that increasing the potential for cell-cell adhesion relative to cell-substrate adhesion promotes cell clustering. Experimental models to study fibroblast clustering have utilized centrifugation, hanging drops, and substrata with poorly adhesive, soft and mechanically unstable properties. In this review, we summarize work on a new, highly tractable, cell clustering research model in which human fibroblasts are incubated on the surfaces of collagen matrices. Fibroblast clustering occurs under procontractile growth factor conditions (e.g., serum or the serum lipid agonist lysophosphatidic acid) but not under promigratory growth factor conditions (e.g., platelet-derived growth factor) and can be reversed by switching growth factor environments. Cell contraction plays a dual role in clustering to bring cells closer together and to stimulate cells to organize fibronectin into a fibrillar matrix. Binding of fibroblasts to a shared fibronectin fibrillar matrix stabilizes clusters, and fragmentation of the fibrillar matrix occurs when growth factor conditions are switched to promote cell dispersal.

  12. Effect of gravity and diffusion interface proximity on the morphology of collagen gels.

    PubMed

    Roedersheimer, M T; Bateman, T A; Simske, S J

    1997-11-01

    Collagen solutions (0.25% w/v) were polymerized in microgravity (STS-77, 10 days) along with simultaneous ground controls. Assembly conditions were achieved by the passage of buffer ions across a dialysis membrane into a reaction chamber containing the dissolved collagen. The gels were analyzed macroscopically and microscopically to assess the influence of gravity and the oriented diffusion of buffer ions on the resulting product. Double-blind rankings based on visual observation of the gels established that all of the flight gels (n = 8) were more uniform in appearance than all of the ground gels (n = 6). Photography using side illumination of the gels revealed the more granular appearance of the ground gels relative to the highly uniform appearance of the flight gels. Scanning electron microscopy established this difference at the microscopic level. Proximity to the dialysis interface and the presence or absence of gravity were both found to control the porosity and uniformity of the matrix.

  13. Collagen/heparin sulfate scaffolds fabricated by a 3D bioprinter improved mechanical properties and neurological function after spinal cord injury in rats.

    PubMed

    Chen, Chong; Zhao, Ming-Liang; Zhang, Ren-Kun; Lu, Gang; Zhao, Chang-Yu; Fu, Feng; Sun, Hong-Tao; Zhang, Sai; Tu, Yue; Li, Xiao-Hong

    2017-01-25

    Effective treatments promoting axonal regeneration and functional recovery for spinal cord injury (SCI) are still in the early stages of development. Most approaches have been focused on providing supportive substrates for guiding neurons and overcoming the physical and chemical barriers to healing that arise after SCI. Although collagen has become a promising natural substrate with good compatibility, its low mechanical properties restrict its potential applications. The mechanical properties mainly rely on the composition and pore structure of scaffolds. For the composition of a scaffold, we used heparin sulfate to react with collagen by crosslinking. For the structure, we adopted a three-dimensional (3D) printing technology to fabricate a scaffold with a uniform pore distributions. We observed that the internal structure of the scaffold printed with a 3D bioprinter was regular and porous. We also found that both the compression modulus and strengths of the scaffold were significantly enhanced by the collagen/heparin sulfate composition compared to a collagen scaffold. Meanwhile, the collagen/heparin sulfate scaffold presented good biocompatibility when it was co-cultured with neural stem cells in vitro. We also demonstrated that heparin sulfate modification significantly improved bFGF immobilization and absorption to the collagen by examining the release kinetics of bFGF from scaffolds. Two months after implantating the scaffold into transection lesions in T10 of the spinal cord in rats, the collagen/heparin sulfate group demonstrated significant recovery of locomotor function and according to electrophysiological examinations. Parallel to functional recovery, collagen/heparin sulfate treatment further ameliorated the pathological process and markedly increased the number of neurofilament (NF) positive cells compared to collagen treatment alone. These data suggested that a collagen/heparin sulfate scaffold fabricated by a 3D bioprinter could enhance the

  14. Thickness sensing of hMSCs on collagen gel directs stem cell fate

    SciTech Connect

    Leong, Wen Shing; Tay, Chor Yong; Yu, Haiyang; Li, Ang; Wu, Shu Cheng; Duc, Duong-Hong; Lim, Chwee Teck; Tan, Lay Poh

    2010-10-15

    Research highlights: {yields} hMSCs appeared to sense thin collagen gel (130 {mu}m) with higher effective modulus as compared to thick gel (1440 {mu}m). {yields} Control of collagen gel thickness can modulate cellular behavior, even stem cell fate (neuronal vs. Quiescent). {yields} Distinct cellular behavior of hMSCs on thin and thick collagen gel suggests long range interaction of hMSCs with collagen gel. -- Abstract: Mechanically compliant substrate provides crucial biomechanical cues for multipotent stem cells to regulate cellular fates such as differentiation, proliferation and maintenance of their phenotype. Effective modulus of which cells sense is not only determined by intrinsic mechanical properties of the substrate, but also the thickness of substrate. From our study, it was found that interference from underlying rigid support at hundreds of microns away could induce significant cellular response. Human mesenchymal stem cells (hMSCs) were cultured on compliant biological gel, collagen type I, of different thickness but identical ECM composition and local stiffness. The cells sensed the thin gel (130 {mu}m) as having a higher effective modulus than the thick gel (1440 {mu}m) and this was reflected in their changes in morphology, actin fibers structure, proliferation and tissue specific gene expression. Commitment into neuronal lineage was observed on the thin gel only. Conversely, the thick gel (1440 {mu}m) was found to act like a substrate with lower effective modulus that inhibited actin fiber polymerization. Stem cells on the thick substrate did not express tissue specific genes and remained at their quiescent state. This study highlighted the need to consider not only the local modulus but also the thickness of biopolymer gel coating during modulation of cellular responses.

  15. Migration and Proliferative Activity of Mesenchymal Stem Cells in 3D Polylactide Scaffolds Depends on Cell Seeding Technique and Collagen Modification.

    PubMed

    Rodina, A V; Tenchurin, T Kh; Saprykin, V P; Shepelev, A D; Mamagulashvili, V G; Grigor'ev, T E; Lukanina, K I; Orekhov, A S; Moskaleva, E Yu; Chvalun, S N

    2016-11-01

    We analyzed viability of mesenchymal stem cells seeded by static and dynamic methods to highly porous fibrous 3D poly-L-lactide scaffolds with similar physical and chemical properties, but different spatial organization modified with collagen. Standard collagen coating promoted protein adsorption on the scaffold surface and improved adhesive properties of 100 μ-thick scaffolds. Modification of 600-μ scaffolds with collagen under pressure increased proliferative activity of mesenchymal stem cells seeded under static and dynamic (delivery of 100,000 cells in 10 ml medium in a perfusion system at a rate of 1 ml/min) conditions by 47 and 648%, respectively (measured after 120-h culturing by MTT test). Dynamic conditions provide more uniform distribution of collagen on scaffold fibers and promote cell penetration into 3D poly-L-lactide scaffolds with thickness >600 μ.

  16. Roles of epithelial cell-derived periostin in TGF-beta activation, collagen production, and collagen gel elasticity in asthma.

    PubMed

    Sidhu, Sukhvinder S; Yuan, Shaopeng; Innes, Anh L; Kerr, Sheena; Woodruff, Prescott G; Hou, Lydia; Muller, Susan J; Fahy, John V

    2010-08-10

    Periostin is considered to be a matricellular protein with expression typically confined to cells of mesenchymal origin. Here, by using in situ hybridization, we show that periostin is specifically up-regulated in bronchial epithelial cells of asthmatic subjects, and in vitro, we show that periostin protein is basally secreted by airway epithelial cells in response to IL-13 to influence epithelial cell function, epithelial-mesenchymal interactions, and extracellular matrix organization. In primary human bronchial epithelial cells stimulated with periostin and epithelial cells overexpressing periostin, we reveal a function for periostin in stimulating the TGF-beta signaling pathway in a mechanism involving matrix metalloproteinases 2 and 9. Furthermore, conditioned medium from the epithelial cells overexpressing periostin caused TGF-beta-dependent secretion of type 1 collagen by airway fibroblasts. In addition, mixing recombinant periostin with type 1 collagen in solution caused a dramatic increase in the elastic modulus of the collagen gel, indicating that periostin alters collagen fibrillogenesis or cross-linking and leads to stiffening of the matrix. Epithelial cell-derived periostin in asthma has roles in TGF-beta activation and collagen gel elasticity in asthma.

  17. STRUCTURAL MECHANISM FOR ALTERATION OF COLLAGEN GEL MECHANICS BY GLUTARALDEHYDE CROSSLINKING

    PubMed Central

    Chandran, Preethi L.; Paik, David C.; Holmes, Jeffrey W.

    2013-01-01

    Soft collagenous tissues that are loaded in vivo undergo crosslinking during aging and wound healing. Bio-prosthetic tissues implanted in vivo are also commonly crosslinked with glutaraldehyde. While crosslinking changes the mechanical properties of the tissue, the nature of the mechanical changes and the underlying microstructural mechanism is poorly understood. In this study, a combined mechanical, biochemical and simulation approach was employed to identify the microstructural mechanism by which crosslinking alters mechanical properties. The model collagenous tissue used was an anisotropic cell-compacted collagen gel, and the model crosslinking agent was monomeric glutaraldehyde. The collagen gels were incrementally crosslinked by either increasing the glutaraldehyde concentration or by increasing the crosslinking time. In biaxial loading experiments, increased crosslinking produced: (1) decreased strain response to a small equibiaxial preload, with little change in response to subsequent loading, and (2) decreased coupling between the fiber and cross-fiber direction. The mechanical trend was found to be better described by the lysine consumption data than by the shrinkage temperature. The biaxial loading of incrementally-crosslinked collagen gels was simulated computationally with a previously published network model. Crosslinking was represented by increased fibril stiffness or by increased resistance to fibril rotation. Only the latter produced mechanical trends similar to that observed experimentally. Representing crosslinking as increased fibril stiffness did not reproduce the decreased coupling between the fiber and cross-fiber directions. The study concludes that the mechanical changes in crosslinked collagen gels are caused by the microstructural mechanism of increased resistance to fibril rotation. PMID:22775003

  18. Properties of collagen gels cross-linked by N-hydroxysuccinimide activated adipic acid deriviate.

    PubMed

    Duan, Lian; Liu, Wentao; Tian, Zhenhua; Li, Conghu; Li, Guoying

    2014-08-01

    In order to improve the properties of collagen gel, N-hydroxysuccinimide activated adipic acid derivative (NHS-AA) was introduced into the formation of collagen fibrils. NHS-AA with different [NHS-AA]/[NH2] ratios (0.1-1.5, calculated by [ester group] of NHS-AA and [NH2] of lysine and hydroxylysine residues of collagen) was added after, simultaneously with or before the formation of collagen fibrils (abbreviated CAF, CSF and CBF, respectively) to obtain different collagen gels. With the same dose of NHS-AA, the cross-linking degree for CAF was lower than those for CSF and CBF. The formation of collagen fibrils was restrained by NHS-AA for CSF and CBF while that for CAF was unaffected. When the dose of NHS-AA increased from 0.1 to 1.5, the water contents of CSF and CBF increased while that of CAF had no obvious change. With lower dose of NHS-AA (0.1), CAF possessed higher value of G' (87.3Pa) and the best thermal stability (47.6°C). As the ratio of [NHS-AA]/[NH2] increased to 1.5, CSF had the maximum value of G' (288.8Pa) and CAF had the best thermal stability (52.9°C). These results showed collagen gels with different properties could be prepared by adding NHS-AA with different adding sequence and dose.

  19. Mechanosensing of cells in 3D gel matrices based on natural and synthetic materials.

    PubMed

    Shan, Jieling; Chi, Qingjia; Wang, Hongbing; Huang, Qiping; Yang, Li; Yu, Guanglei; Zou, Xiaobing

    2014-11-01

    Cells in vivo typically are found in 3D matrices, the mechanical stiffness of which is important to the cell and tissue-scale biological processes. Although it is well characterized that as to how cells sense matrix stiffness in 2D substrates, the scenario in 3D matrices needs to be explored. Thus, materials that can mimic native 3D environments and possess wide, physiologically relevant elasticity are highly desirable. Natural polymer-based materials and synthetic hydrogels could provide an better 3D platforms to investigate the mechano-response of cells with stiffness comparable to their native environments. However, the limited stiffness range together with interdependence of matrix stiffness and adhesive ligand density are inherent in many kinds of materials, and hinder efforts to demonstrate the true effects contributed by matrix stiffness. These problems have been addressed by the recently emerging exquisitely designed materials based on native matrix components, designer matrices, and synthetic polymers. In this review, a variety of materials with a wide stiffness range that mimic the mechanical environment of native 3D matrices and the independent affection of stiffness for cellular behavior and tissue-level processes are discussed.

  20. Fabrication and characterization of gels with integrated channels using 3D printing with microfluidic nozzle for tissue engineering applications.

    PubMed

    Attalla, R; Ling, C; Selvaganapathy, P

    2016-02-01

    The lack of a simple and effective method to integrate vascular network with engineered scaffolds and tissue constructs remains one of the biggest challenges in true 3D tissue engineering. Here, we detail the use of a commercially available, low-cost, open-source 3D printer modified with a microfluidic print-head in order to develop a method for the generation of instantly perfusable vascular network integrated with gel scaffolds seeded with cells. The print-head features an integrated coaxial nozzle that allows the fabrication of hollow, calcium-polymerized alginate tubes that can be easily patterned using 3D printing techniques. The diameter of the hollow channel can be precisely controlled and varied between 500 μm - 2 mm by changing applied flow rates or print-head speed. These channels are integrated into gel layers with a thickness of 800 μm - 2.5 mm. The structural rigidity of these constructs allows the fabrication of multi-layered structures without causing the collapse of hollow channels in lower layers. The 3D printing method was fully characterized at a range of operating speeds (0-40 m/min) and corresponding flow rates (1-30 mL/min) were identified to produce precise definition. This microfluidic design also allows the incorporation of a wide range of scaffold materials as well as biological constituents such as cells, growth factors, and ECM material. Media perfusion of the channels causes a significant viability increase in the bulk of cell-laden structures over the long-term. With this setup, gel constructs with embedded arrays of hollow channels can be created and used as a potential substitute for blood vessel networks.

  1. Molecular weight specific impact of soluble and immobilized hyaluronan on CD44 expressing melanoma cells in 3D collagen matrices.

    PubMed

    Sapudom, Jiranuwat; Ullm, Franziska; Martin, Steve; Kalbitzer, Liv; Naab, Johanna; Möller, Stephanie; Schnabelrauch, Matthias; Anderegg, Ulf; Schmidt, Stephan; Pompe, Tilo

    2017-03-01

    Hyaluronan (HA) and its principal receptor CD44 are known to be involved in regulating tumor cell dissemination and metastasis. The direct correlation of CD44-HA interaction on proliferation and invasion of tumor cells in dependence on the molecular weight and the presentation form of HA is not fully understood because of lack of appropriate matrix models. To address this issue, we reconstituted 3D collagen (Coll I) matrices and functionalized them with HA of molecular weight of 30-50kDa (low molecular weight; LMW-HA) and 500-750kDa (high molecular weight; HMW-HA). A post-modification strategy was applied to covalently immobilize HA to reconstituted fibrillar Coll I matrices, resulting in a non-altered Coll I network microstructure and stable immobilization over days. Functionalized Coll I matrices were characterized regarding topological and mechanical characteristics as well as HA amount using confocal laser scanning microscopy, colloidal probe force spectroscopy and quantitative Alcian blue assay, respectively. To elucidate HA dependent tumor cell behavior, BRO melanoma cell lines with and without CD44 receptor expression were used for in vitro cell experiments. We demonstrated that only soluble LMW-HA promoted cell proliferation in a CD44 dependent manner, while HMW-HA and immobilized LMW-HA did not. Furthermore, an enhanced cell invasion was found only for immobilized LMW-HA. Both findings correlated with a very strong and specific adhesive interaction of LMW-HA and CD44+ cells quantified in single cell adhesion measurements using soft colloidal force spectroscopy. Overall, our results introduce an in vitro biomaterials model allowing to test presentation mode and molecular weight specificity of HA in a 3D fibrillar matrix thus mimicking important in vivo features of tumor microenvironments.

  2. Developing a Procedure for the Characterization of Mechanical Properties of Collagen Gels

    NASA Astrophysics Data System (ADS)

    Chambers, Christopher; Lovelady, Heather; Matthews, Garrett

    2011-03-01

    The characterization of bulk mechanical properties of type I collagen gels is critical to understanding the role of collagen in the extracellular matrix (ECM), and developing biocompatible devices for use in the human body. Understanding the mechanical properties of the gel state of collagen can lead to the ability to adjust these properties for multiple uses. Here, we examined the Young's modulus of the synthesized gels. This project used a microrheological approach to discover these properties. Gels were first formed using a known process and magnetic microspheres were embedded in the gel prior to formation. An optical microscope was fitted with a magnetic chamber used to drive the embedded beads in two modes, an oscillatory motion and a pulse motion. Tracking software was modified and used to analyze the motion of the beads recorded with a CCD camera on the microscope. These techniques should be sufficient to obtain a reliable value for the Young's modulus of collagen gels, as well as other similar materials. This work was supported by NSF REU program (award No DMR-1004873).

  3. Elastic moduli of collagen gels can be predicted from two-dimensional confocal microscopy.

    PubMed

    Yang, Ya-Li; Leone, Lindsay M; Kaufman, Laura J

    2009-10-07

    We quantitatively compare data obtained from imaging two-dimensional slices of three-dimensional unlabeled and fluorescently labeled collagen gels with confocal reflectance microscopy (CRM) and/or confocal fluorescence microscopy (CFM). Different network structures are obtained by assembling the gels over a range of concentrations at various temperatures. Comparison between CRM and CFM shows that the techniques are not equally sensitive to details of network structure, with CFM displaying higher fidelity in imaging fibers parallel to the optical axis. Comparison of CRM of plain and labeled collagen gels shows that labeling itself induces changes in gel structure, chiefly through inhibition of fibril bundling. Despite these differences, image analyses carried out on two-dimensional CFM and CRM slices of collagen gels reveal identical trends in structural parameters as a function of collagen concentration and gelation temperature. Fibril diameter approximated from either CRM or CFM is in good accord with that determined via electron microscopy. Two-dimensional CRM images are used to show that semiflexible polymer theory can relate network structural properties to elastic modulus successfully. For networks containing bundled fibrils, it is shown that average structural diameter, rather than fibril diameter, is the length scale that sets the magnitude of the gel elastic modulus.

  4. Non-enzymatic glycation of chondrocyte-seeded collagen gels for cartilage tissue engineering.

    PubMed

    Roy, Rani; Boskey, Adele L; Bonassar, Lawrence J

    2008-11-01

    Collagen glycated with ribose (250 mM) in solution (pre-glycation) and as a gel (post-glycation) was seeded with chondrocytes and the effects of glycation on chondrocyte matrix assembly in culture were determined. Pre-glycation enhanced GAG accumulation significantly over controls at both 2 and 4 weeks (p < 0.05), although at both time points there were no statistical differences in cell number between pre-glycated and control gels. The increased proteoglycan accumulation was shown to be in part due to significantly increased GAG retention by the pre-glycated constructs (p < 0.05). Total collagen content in these pre-glycated gels was also significantly higher than unglycated gels at 4 weeks (p < 0.05). With post-glycation of collagen gels, chondrocyte number and GAG accumulation were all significantly lower than controls (p < 0.05). Post-glycation also inhibited GAG retention by the constructs (p < 0.05). Given these results, pre-glycation may be an improved processing method for collagen gels for tissue engineering techniques.

  5. Small-Field Measurements of 3D Polymer Gel Dosimeters through Optical Computed Tomography

    PubMed Central

    Shih, Cheng-Ting; Lee, Yao-Ting; Wu, Shin-Hua; Yao, Chun-Hsu; Hsieh, Bor-Tsung

    2016-01-01

    With advances in therapeutic instruments and techniques, three-dimensional dose delivery has been widely used in radiotherapy. The verification of dose distribution in a small field becomes critical because of the obvious dose gradient within the field. The study investigates the dose distributions of various field sizes by using NIPAM polymer gel dosimeter. The dosimeter consists of 5% gelatin, 5% monomers, 3% cross linkers, and 5 mM THPC. After irradiation, a 24 to 96 hour delay was applied, and the gel dosimeters were read by a cone beam optical computed tomography (optical CT) scanner. The dose distributions measured by the NIPAM gel dosimeter were compared to the outputs of the treatment planning system using gamma evaluation. For the criteria of 3%/3 mm, the pass rates for 5 × 5, 3 × 3, 2 × 2, 1 × 1, and 0.5 × 0.5 cm2 were as high as 91.7%, 90.7%, 88.2%, 74.8%, and 37.3%, respectively. For the criteria of 5%/5 mm, the gamma pass rates of the 5 × 5, 3 × 3, and 2 × 2 cm2 fields were over 99%. The NIPAM gel dosimeter provides high chemical stability. With cone-beam optical CT readouts, the NIPAM polymer gel dosimeter has potential for clinical dose verification of small-field irradiation. PMID:26974434

  6. Pore structure and dielectric behaviour of the 3D collagen-DAC scaffolds designed for nerve tissue repair.

    PubMed

    Pietrucha, Krystyna; Marzec, Ewa; Kudzin, Marcin

    2016-11-01

    The design and selection of a suitable scaffold with well-defined pores size distribution and dielectric properties are critical features for neural tissue engineering. In this study we use mercury porosimetry and the dielectric spectroscopy in the alpha-dispersion region of the electric field to determine the microarchitecture and activation energy of collagen (Col) modified by 2,3 dialdehyde cellulose (DAC). The scaffold was synthesized in three steps: (i) preparation of DAC by oxidation of cellulose, (ii) construction of a 3D Col sponge-shape or film, (iii) cross-linkage of the Col samples using DAC. The activation energy needed to break the bonds formed by water in the Col-DAC composite is approximately 2 times lower than that in the unmodified Col. In addition, the magnitude of conductivity for modified Col at 70°C is approximately 40% lower than that recorded for the unmodified Col. The largest fraction, of which at least 70% of the total pore volume comprises the sponge, is occupied by pores ranging from 20 to 100μm in size. The knowledge on the dielectric behaviour and microstructure of the Col-DAC scaffold may prove relevant to neural tissue engineering focused on the regeneration of the nervous system.

  7. Integration of 3D Printed and Micropatterned Polycaprolactone Scaffolds for Guidance of Oriented Collagenous Tissue Formation In vivo

    PubMed Central

    Pilipchuk, Sophia P; Monje, Alberto; Jiao, Yizu; Hao, Jie; Kruger, Laura; Flanagan, Colleen L; Hollister, Scott J

    2016-01-01

    Scaffold design incorporating multi-scale cues for clinically-relevant, aligned tissue regeneration has potential to improve structural and functional integrity of multi-tissue interfaces. The objective of this pre-clinical study was to develop poly(ε-caprolactone) (PCL) scaffolds with mesoscale and microscale architectural cues specific to human ligament progenitor cells and assess their ability to form aligned bone-ligament-cementum complexes in vivo. PCL scaffolds were designed to integrate a 3D printed bone region with a micropatterned PCL thin film consisting of grooved pillars. The patterned film region was seeded with human ligament cells, fibroblasts transduced with BMP-7 genes seeded within the bone region, and a tooth dentin segment positioned on the ligament region prior to subcutaneous implantation into a murine model. Results indicated increased tissue alignment in vivo using micropatterned PCL films, compared to random-porous PCL. At 6 weeks, 30um groove depth significantly enhanced oriented collagen fiber thickness, overall cell alignment, and nuclear elongation relative to 10um groove depth. This study demonstrates for the first time that scaffolds with combined hierarchical mesoscale and microscale features can align cells in vivo for oral tissue repair with potential for improving the regenerative response of other bone-ligament complexes. PMID:26820240

  8. Integration of 3D Printed and Micropatterned Polycaprolactone Scaffolds for Guidance of Oriented Collagenous Tissue Formation In Vivo.

    PubMed

    Pilipchuk, Sophia P; Monje, Alberto; Jiao, Yizu; Hao, Jie; Kruger, Laura; Flanagan, Colleen L; Hollister, Scott J; Giannobile, William V

    2016-03-01

    Scaffold design incorporating multiscale cues for clinically relevant, aligned tissue regeneration has potential to improve structural and functional integrity of multitissue interfaces. The objective of this preclinical study is to develop poly(ε-caprolactone) (PCL) scaffolds with mesoscale and microscale architectural cues specific to human ligament progenitor cells and assess their ability to form aligned bone-ligament-cementum complexes in vivo. PCL scaffolds are designed to integrate a 3D printed bone region with a micropatterned PCL thin film consisting of grooved pillars. The patterned film region is seeded with human ligament cells, fibroblasts transduced with bone morphogenetic protein-7 genes seeded within the bone region, and a tooth dentin segment positioned on the ligament region prior to subcutaneous implantation into a murine model. Results indicate increased tissue alignment in vivo using micropatterned PCL films, compared to random-porous PCL. At week 6, 30 μm groove depth significantly enhances oriented collagen fiber thickness, overall cell alignment, and nuclear elongation relative to 10 μm groove depth. This study demonstrates for the first time that scaffolds with combined hierarchical mesoscale and microscale features can align cells in vivo for oral tissue repair with potential for improving the regenerative response of other bone-ligament complexes.

  9. Accelerated craniofacial bone regeneration through dense collagen gel scaffolds seeded with dental pulp stem cells

    PubMed Central

    Chamieh, Frédéric; Collignon, Anne-Margaux; Coyac, Benjamin R.; Lesieur, Julie; Ribes, Sandy; Sadoine, Jérémy; Llorens, Annie; Nicoletti, Antonino; Letourneur, Didier; Colombier, Marie-Laure; Nazhat, Showan N.; Bouchard, Philippe; Chaussain, Catherine; Rochefort, Gael Y.

    2016-01-01

    Therapies using mesenchymal stem cell (MSC) seeded scaffolds may be applicable to various fields of regenerative medicine, including craniomaxillofacial surgery. Plastic compression of collagen scaffolds seeded with MSC has been shown to enhance the osteogenic differentiation of MSC as it increases the collagen fibrillary density. The aim of the present study was to evaluate the osteogenic effects of dense collagen gel scaffolds seeded with mesenchymal dental pulp stem cells (DPSC) on bone regeneration in a rat critical-size calvarial defect model. Two symmetrical full-thickness defects were created (5 mm diameter) and filled with either a rat DPSC-containing dense collagen gel scaffold (n = 15), or an acellular scaffold (n = 15). Animals were imaged in vivo by microcomputer tomography (Micro-CT) once a week during 5 weeks, whereas some animals were sacrificed each week for histology and histomorphometry analysis. Bone mineral density and bone micro-architectural parameters were significantly increased when DPSC-seeded scaffolds were used. Histological and histomorphometrical data also revealed significant increases in fibrous connective and mineralized tissue volume when DPSC-seeded scaffolds were used, associated with expression of type I collagen, osteoblast-associated alkaline phosphatase and osteoclastic-related tartrate-resistant acid phosphatase. Results demonstrate the potential of DPSC-loaded-dense collagen gel scaffolds to benefit of bone healing process. PMID:27934940

  10. Accelerated craniofacial bone regeneration through dense collagen gel scaffolds seeded with dental pulp stem cells

    NASA Astrophysics Data System (ADS)

    Chamieh, Frédéric; Collignon, Anne-Margaux; Coyac, Benjamin R.; Lesieur, Julie; Ribes, Sandy; Sadoine, Jérémy; Llorens, Annie; Nicoletti, Antonino; Letourneur, Didier; Colombier, Marie-Laure; Nazhat, Showan N.; Bouchard, Philippe; Chaussain, Catherine; Rochefort, Gael Y.

    2016-12-01

    Therapies using mesenchymal stem cell (MSC) seeded scaffolds may be applicable to various fields of regenerative medicine, including craniomaxillofacial surgery. Plastic compression of collagen scaffolds seeded with MSC has been shown to enhance the osteogenic differentiation of MSC as it increases the collagen fibrillary density. The aim of the present study was to evaluate the osteogenic effects of dense collagen gel scaffolds seeded with mesenchymal dental pulp stem cells (DPSC) on bone regeneration in a rat critical-size calvarial defect model. Two symmetrical full-thickness defects were created (5 mm diameter) and filled with either a rat DPSC-containing dense collagen gel scaffold (n = 15), or an acellular scaffold (n = 15). Animals were imaged in vivo by microcomputer tomography (Micro-CT) once a week during 5 weeks, whereas some animals were sacrificed each week for histology and histomorphometry analysis. Bone mineral density and bone micro-architectural parameters were significantly increased when DPSC-seeded scaffolds were used. Histological and histomorphometrical data also revealed significant increases in fibrous connective and mineralized tissue volume when DPSC-seeded scaffolds were used, associated with expression of type I collagen, osteoblast-associated alkaline phosphatase and osteoclastic-related tartrate-resistant acid phosphatase. Results demonstrate the potential of DPSC-loaded-dense collagen gel scaffolds to benefit of bone healing process.

  11. Gamma Knife 3-D dose distribution near the area of tissue inhomogeneities by normoxic gel dosimetry

    SciTech Connect

    Isbakan, Fatih; Uelgen, Yekta; Bilge, Hatice; Ozen, Zeynep; Agus, Onur; Buyuksarac, Bora

    2007-05-15

    The accuracy of the Leksell GammaPlan registered , the dose planning system of the Gamma Knife Model-B, was evaluated near tissue inhomogeneities, using the gel dosimetry method. The lack of electronic equilibrium around the small-diameter gamma beams can cause dose calculation errors in the neighborhood of an air-tissue interface. An experiment was designed to investigate the effects of inhomogeneity near the paranosal sinuses cavities. The homogeneous phantom was a spherical glass balloon of 16 cm diameter, filled with MAGIC gel; i.e., the normoxic polymer gel. Two hollow PVC balls of 2 cm radius, filled with N{sub 2} gas, represented the air cavities inside the inhomogeneous phantom. For dose calibration purposes, 100 ml gel-containing vials were irradiated at predefined doses, and then scanned in a MR unit. Linearity was observed between the delivered dose and the reciprocal of the T2 relaxation time constant of the gel. Dose distributions are the results of a single shot of irradiation, obtained by collimating all 201 cobalt sources to a known target in the phantom. Both phantoms were irradiated at the same dose level at the same coordinates. Stereotactic frames and fiducial markers were attached to the phantoms prior to MR scanning. The dose distribution predicted by the Gamma Knife planning system was compared with that of the gel dosimetry. As expected, for the homogeneous phantom the isodose diameters measured by the gel dosimetry and the GammaPlan registered differed by 5% at most. However, with the inhomogeneous phantom, the dose maps in the axial, coronal and sagittal planes were spatially different. The diameters of the 50% isodose curves differed 43% in the X axis and 32% in the Y axis for the Z=90 mm axial plane; by 44% in the X axis and 24% in the Z axis for the Y=90 mm coronal plane; and by 32% in the Z axis and 42% in the Y axis for the X=92 mm sagittal plane. The lack of ability of the GammaPlan registered to predict the rapid dose fall off, due

  12. On the development of a VIPAR(nd) radiotherapy 3D polymer gel dosimeter.

    PubMed

    Kozicki, Marek; Jaszczak, Malwina; Maras, Piotr; Dudek, Mariusz; Cłapa, Marian

    2017-02-07

    This work presents an improvement of the VIPAR(nd) ('nd' stands for 'normoxic, double', or VIP) polymer gel dosimeter. The gel composition was altered by increasing the concentration of the monomeric components, N-vinylpyrrolidone (NVP) and N,N'-methylenebisacrylamide (MBA), in co-solvent solutions. The optimal composition (VIPAR(CT), where 'CT' stands for computed tomography, or VIC) comprised: 17% NVP, 8% MBA, 12% t-BuOH, 7.5% gelatine, 0.007% ascorbic acid, 0.0008% CuSO4  ×  5H2O and 0.02% hydroquinone. The following characteristics of VIC were achieved: (i) linear dose range of 0.9(_)30 Gy, (ii) saturation for radiation doses of over 50 Gy, (iii) threshold dose of about 0.5 Gy, (iv) dose sensitivity of 0.171 Gy(-1) s(-1), which is roughly 2.2 times higher than that of VIP (for nuclear magnetic resonance measurements). It was also found that VIC is dose- rate-independent, and its dose response does not alter if the radiation source is changed from electrons to photons for external beam radiotherapy. The gel responded similarly to irradiation with small changes in radiation energy but was sensitive to larger energy changes. The VIC gel retained temporal stability from 20 h until at least 10 d after irradiation, whereas spatial stability was retained from 20 h until at least 6 d after irradiation. The scheme adopted for VIC manufacturing yields repeatable gels in terms of radiation dose response. The VIC was also shown to perform better than VIP using x-ray computed tomography as a readout method; the dose sensitivity of VIC (0.397 HU Gy(-1)) was 1.5 times higher than that of VIP. Also, the dose resolution of VIC was better than that of VIP in the whole dose range examined.

  13. On the development of a VIPARnd radiotherapy 3D polymer gel dosimeter

    NASA Astrophysics Data System (ADS)

    Kozicki, Marek; Jaszczak, Malwina; Maras, Piotr; Dudek, Mariusz; Cłapa, Marian

    2017-02-01

    This work presents an improvement of the VIPARnd (‘nd’ stands for ‘normoxic, double’, or VIP) polymer gel dosimeter. The gel composition was altered by increasing the concentration of the monomeric components, N-vinylpyrrolidone (NVP) and N,N‧-methylenebisacrylamide (MBA), in co-solvent solutions. The optimal composition (VIPARCT, where ‘CT’ stands for computed tomography, or VIC) comprised: 17% NVP, 8% MBA, 12% t-BuOH, 7.5% gelatine, 0.007% ascorbic acid, 0.0008% CuSO4  ×  5H2O and 0.02% hydroquinone. The following characteristics of VIC were achieved: (i) linear dose range of 0.9_30 Gy, (ii) saturation for radiation doses of over 50 Gy, (iii) threshold dose of about 0.5 Gy, (iv) dose sensitivity of 0.171 Gy-1 s-1, which is roughly 2.2 times higher than that of VIP (for nuclear magnetic resonance measurements). It was also found that VIC is dose- rate-independent, and its dose response does not alter if the radiation source is changed from electrons to photons for external beam radiotherapy. The gel responded similarly to irradiation with small changes in radiation energy but was sensitive to larger energy changes. The VIC gel retained temporal stability from 20 h until at least 10 d after irradiation, whereas spatial stability was retained from 20 h until at least 6 d after irradiation. The scheme adopted for VIC manufacturing yields repeatable gels in terms of radiation dose response. The VIC was also shown to perform better than VIP using x-ray computed tomography as a readout method; the dose sensitivity of VIC (0.397 HU Gy-1) was 1.5 times higher than that of VIP. Also, the dose resolution of VIC was better than that of VIP in the whole dose range examined.

  14. Fabrication of endothelialized tube in collagen gel as starting point for self-developing capillary-like network to construct three-dimensional organs in vitro.

    PubMed

    Takei, Takayuki; Sakai, Shinji; Ono, Tsutomu; Ijima, Hiroyuki; Kawakami, Koei

    2006-09-05

    A possible strategy for creating three-dimensional (3D) tissue-engineered organs in vitro with similar volumes to the primary organs is to develop a capillary network throughout the constructs to provide sufficient oxygenation and nutrition to the cells composing them. Here, we propose a novel approach for the creation of a capillary-like network in vitro, based on the spontaneous tube-forming activity of vascular endothelial cells (ECs) in collagen gel. We fabricated a linear tube of 500 microm in diameter, the inner surface of which was filled with bovine carotid artery vascular endothelial cells (BECs), in type I collagen gel as a starting point for the formation of a capillary-like network. The BECs exposed to a medium containing vascular endothelial growth factor (VEGF) migrated into the ambient gel around the tube. After 2 weeks of VEGF exposure, the distance of the migration into the ambient gel in the radial direction of the tube reached approximately 800 microm. Cross-sections of capillary-like structures composed of the migrating BECs, with a lumen-like interior space, were observed in slices of the gel around the tube stained with hematoxylin-eosin (H&E). These results demonstrate that this approach using a pre-established tube, which is composed of ECs, as a starting point for a self-developing capillary-like network is potentially useful for constructing 3D organs in vitro.

  15. Integrin {beta}1-dependent invasive migration of irradiation-tolerant human lung adenocarcinoma cells in 3D collagen matrix

    SciTech Connect

    Ishihara, Seiichiro; Haga, Hisashi; Yasuda, Motoaki; Mizutani, Takeomi; Kawabata, Kazushige; Shirato, Hiroki; Nishioka, Takeshi

    2010-06-04

    Radiotherapy is one of the effective therapies used for treating various malignant tumors. However, the emergence of tolerant cells after irradiation remains problematic due to their high metastatic ability, sometimes indicative of poor prognosis. In this study, we showed that subcloned human lung adenocarcinoma cells (A549P-3) that are irradiation-tolerant indicate high invasive activity in vitro, and exhibit an integrin {beta}1 activity-dependent migratory pattern. In collagen gel overlay assay, majority of the A549P-3 cells displayed round morphology and low migration activity, whereas a considerable number of A549P-3IR cells surviving irradiation displayed a spindle morphology and high migration rate. Blocking integrin {beta}1 activity reduced the migration rate of A549P-3IR cells and altered the cell morphology allowing them to assume a round shape. These results suggest that the A549P-3 cells surviving irradiation acquire a highly invasive integrin {beta}1-dependent phenotype, and integrin {beta}1 might be a potentially effective therapeutic target in combination with radiotherapy.

  16. Synthesis of glycosaminoglycans in differently loaded regions of collagen gels seeded with valvular interstitial cells.

    PubMed

    Gupta, Vishal; Werdenberg, Jennifer A; Blevins, Tracy L; Grande-Allen, K Jane

    2007-01-01

    Cells respond to changes in mechanical strains by varying their production of extracellular matrix macromolecules. Because differences in strain patterns between mitral valve leaflets and chordae tendineae have been linked to different quantities and types of glycosaminoglycans (GAGs), we investigated the effects of various strain conditions on GAG synthesis by valvular interstitial cells (VICs) using an in vitro 3-dimensional tissue-engineering model. VICs from leaflets or chordae were seeded within collagen gels and subjected to uniaxial or biaxial static tension for 1 week. GAGs synthesized within the collagen gels and secreted into the surrounding medium were analyzed using fluorophore-assisted carbohydrate electrophoresis. In constrained conditions, more 4-sulfated GAGs were retained within the collagen gel, whereas more hyaluronan was secreted into the surrounding medium. Selected GAG classes were found in significantly different proportions in collagen gels seeded with leaflet cells versus chordal cells. The only significant difference between uniaxial and biaxial regions was found for 6-sulfated GAGs in the gels seeded with chordal cells (p<0.05). This study suggests how mechanical loading may influence GAG production and localization in the remodeling of the mitral valve and has design implications for engineered tissues.

  17. Photophysical and photochemical processes in 3D self-assembled gels as confined microenvironments.

    PubMed

    Pérez-Ruiz, Raúl; Díaz Díaz, David

    2015-07-14

    Numerous challenging transformations take place in nature with high efficiency within confined and compartmented environments. This has inspired scientists to develop spatially micro- and nanoreactors by 'bottom-up' approaches in order to improve different processes in comparison to solution, in terms of kinetics, selectivity or processability. In this respect, investigation of photophysical and photochemical processes in soft gel materials has recently emerged as a new and promising research field oriented towards expanding their applications in important areas such as photovoltaics, photocatalysis and phototherapy. Herein, we summarize the few examples dealing with intragel photo-induced physical and chemical processes involving embedded reactants that do not participate in the assembly of the gel network.

  18. Effect of ultrasonication on the fibril-formation and gel properties of collagen from grass carp skin.

    PubMed

    Jiang, Ying; Wang, Haibo; Deng, Mingxia; Wang, Zhongwen; Zhang, Juntao; Wang, Haiyin; Zhang, Hanjun

    2016-02-01

    Controlling the fibril-formation process of collagen in vitro to fabricate novel biomaterials is a new area in the field of collagen research. This study aimed to determine the effect of ultrasonication on collagen fibril formation and the properties of the resulting collagen gels. Native collagen, extracted from the skin of grass carp, self-assembled under ultrasonic conditions (at different ultrasonic power and duration). The self-assembly kinetics, fibrillar morphology, and physical and cell growth-promoting properties of the collagen gels were analyzed and compared. The results showed that the self-assembly rate of collagen was increased by ultrasonication at the nucleation stage. The resulting fibrils exhibited smaller diameters and D-periodicity lengths than that of the untreated collagen samples (p<0.05). The viscoelasticity and textural properties of collagen gels also changed after ultrasonication at the nucleation stage. Texture profile analysis and cell proliferation assays showed that ultrasonication produced softer collagen gel colloids, which were more suitable for cell proliferation than the untreated collagen gels.

  19. Assessment of angiogenesis in osseointegration of a silica-collagen biomaterial using 3D-nano-CT.

    PubMed

    Alt, Volker; Kögelmaier, Daniela Vera; Lips, Katrin S; Witt, Vera; Pacholke, Sabine; Heiss, Christian; Kampschulte, Marian; Heinemann, Sascha; Hanke, Thomas; Thormann, Ulrich; Schnettler, Reinhard; Langheinrich, Alexander C

    2011-10-01

    Bony integration of biomaterials is a complex process in which angiogenesis plays a crucial role. We evaluated micro- and nano-CT imaging to demonstrate and quantify neovascularization in bony integration of a biomaterial and to give an image based estimation for the needed resolution for imaging angiogenesis in an animal model of femora defect healing. In 8 rats 5mm full-size defects were created at the left femur that was filled with silica-collagen bone substitute material and internally fixed with plate osteosynthesis. After 6 weeks the femora were infused in situ with Microfil, harvested and scanned for micro-CT (9 μm)(3) and nano-CT (3 μm)(3) imaging. Using those 3D images, the newly formed blood vessels in the area of the biomaterial were assessed and the total vascular volume fraction, the volume of the bone substitute material and the volume of the bone defect were quantitatively characterized. Results were complemented by histology. Differences were statistically assessed using (ANOVA). High-resolution nano-CT demonstrated new blood vessel formation surrounding the biomaterial in all animals at capillary level. Immunohistochemistry confirmed the newly formed blood vessels surrounding the bone substitute material. The mean vascular volume fraction (VVF) around the implant was calculated to be 3.01 ± 0.4%. The VVF was inversely correlated with the volume of the bone substitute material (r=0.8) but not with the dimension of the fracture zone (r=0.3). Nano-CT imaging is feasible for quantitative analysis of angiogenesis during bony integration of biomaterials and a promising tool in this context for the future.

  20. Fibrin binds to collagen and provides a bridge for αVβ3 integrin-dependent contraction of collagen gels.

    PubMed

    Reyhani, Vahid; Seddigh, Pegah; Guss, Bengt; Gustafsson, Renata; Rask, Lars; Rubin, Kristofer

    2014-08-15

    The functional significance of fibrin deposits typically seen in inflammatory lesions, carcinomas and in healing wounds is not fully understood. In the present study, we demonstrate that fibrinogen/fibrin specifically bound to native Col I (collagen type I) and used the Col I fibre network as a base to provide a functional interface matrix that connects cells to the Col I fibres through αVβ3 integrins. This allowed murine myoblast C2C12 cells to contract the collagenous composite gel via αVβ3 integrin. We show that fibrinogen specifically bound to immobilized native Col I at the site known to bind matrix metalloproteinase-1, discoidin domain receptor-2 and fibronectin, and that binding had no effect on Col I fibrillation. A specific competitive inhibitor blocking the Col-I-binding site for fibrinogen abolished the organization of fibrin into discernable fibrils, as well as the C2C12-mediated contraction of Col I gels. Our data show that fibrin can function as a linkage protein between Col I fibres and cells, and suggest that fibrin at inflammatory sites indirectly connects αVβ3 integrins to Col I fibres and thereby promotes cell-mediated contraction of collagenous tissue structures.

  1. A 3D in situ cell counter reveals that breast tumor cell (MDA-MB-231) proliferation rate is reduced by the collagen matrix density

    PubMed Central

    Bunaciu, Rodica P.; Yen, Andrew; Wu, Mingming

    2015-01-01

    Many cell types require the biophysical and biochemical cues within the 3D extracellular matrix (ECM) to exhibit their true physiologically relevant behavior. As a result, cell culture platforms have been evolving from traditional 2D petri-dish plates into 3D biomatrices, and there is a need for developing analytic tools to characterize 3D cell culture. The existing cell counting method, using a hemocytometer or coulter counter, requires that cells are suspended in fluids prior to counting. This poses a challenge for 3D cell culture as cells are embedded in a 3D biomatrix. We use a facile 3D cell counting method that overcomes this limitation and allows for in situ cell counting in a 3D cell culture using equipment that is commonly available in a biology lab. Using a breast tumor cell line, MDA-MB-231, as a model system, we demonstrated that MDA-MB-231 cells (1) grow slower within a 3D collagen matrix than on a 2D substrate for an extended growth time (a week) with a comparable, initial cell-to-cell distance, (2) their cell growth rate decreases with the increase of collagen concentration, showing a linear growth rate rather than an exponential growth rate. Further work using flow cytometry showed that the observed growth rate reduction was consistent with the retardation of the transition to S (synthesis) phase in the cell cycle. This work demonstrates the validity of the 3D cell counting method and the importance of cell-ECM interactions in cell proliferation. PMID:25683564

  2. Mathematical Modeling of Uniaxial Mechanical Properties of Collagen Gel Scaffolds for Vascular Tissue Engineering

    PubMed Central

    Irastorza, Ramiro M.; Drouin, Bernard; Blangino, Eugenia; Mantovani, Diego

    2015-01-01

    Small diameter tissue-engineered arteries improve their mechanical and functional properties when they are mechanically stimulated. Applying a suitable stress and/or strain with or without a cycle to the scaffolds and cells during the culturing process resides in our ability to generate a suitable mechanical model. Collagen gel is one of the most used scaffolds in vascular tissue engineering, mainly because it is the principal constituent of the extracellular matrix for vascular cells in human. The mechanical modeling of such a material is not a trivial task, mainly for its viscoelastic nature. Computational and experimental methods for developing a suitable model for collagen gels are of primary importance for the field. In this research, we focused on mechanical properties of collagen gels under unconfined compression. First, mechanical viscoelastic models are discussed and framed in the control system theory. Second, models are fitted using system identification. Several models are evaluated and two nonlinear models are proposed: Mooney-Rivlin inspired and Hammerstein models. The results suggest that Mooney-Rivlin and Hammerstein models succeed in describing the mechanical behavior of collagen gels for cyclic tests on scaffolds (with best fitting parameters 58.3% and 75.8%, resp.). When Akaike criterion is used, the best is the Mooney-Rivlin inspired model. PMID:25834840

  3. Mathematical modeling of uniaxial mechanical properties of collagen gel scaffolds for vascular tissue engineering.

    PubMed

    Irastorza, Ramiro M; Drouin, Bernard; Blangino, Eugenia; Mantovani, Diego

    2015-01-01

    Small diameter tissue-engineered arteries improve their mechanical and functional properties when they are mechanically stimulated. Applying a suitable stress and/or strain with or without a cycle to the scaffolds and cells during the culturing process resides in our ability to generate a suitable mechanical model. Collagen gel is one of the most used scaffolds in vascular tissue engineering, mainly because it is the principal constituent of the extracellular matrix for vascular cells in human. The mechanical modeling of such a material is not a trivial task, mainly for its viscoelastic nature. Computational and experimental methods for developing a suitable model for collagen gels are of primary importance for the field. In this research, we focused on mechanical properties of collagen gels under unconfined compression. First, mechanical viscoelastic models are discussed and framed in the control system theory. Second, models are fitted using system identification. Several models are evaluated and two nonlinear models are proposed: Mooney-Rivlin inspired and Hammerstein models. The results suggest that Mooney-Rivlin and Hammerstein models succeed in describing the mechanical behavior of collagen gels for cyclic tests on scaffolds (with best fitting parameters 58.3% and 75.8%, resp.). When Akaike criterion is used, the best is the Mooney-Rivlin inspired model.

  4. Fabrication of Compositionally and Topographically Complex Robust Tissue Forms by 3D-Electrochemical Compaction of Collagen

    PubMed Central

    Younesi, Mousa; Islam, Anowarul; Kishore, Vipuil; Panit, Stefi; Akkus, Ozan

    2015-01-01

    Collagen solutions are phase-transformed to mechanically robust shell structures with curviplanar topographies using electrochemically induced pH gradients. The process enables rapid layer-by-layer deposition of collagen-rich mixtures over the entire field simultaneously to obtain compositionally diverse multilayered structures. In-plane tensile strength and modulus of the electrocompacted collagen sheet samples were 5200 -fold and 2300 -fold greater than that of uncompacted collagen samples. Out of plane compression tests showed 27 -fold and fold increase in compressive stress and 46 -fold increase in compressive modulus compared to uncompacted collagen sheets. Cells proliferated 4.9 times faster, and cellular area spread was 2.7 times greater on compacted collagen sheets. Electrocompaction also resulted in 2.9 times greater focal adhesion area than on regular collagen hydrogel. The reported improvements in the cell-matrix interactions with electrocompaction would serve to expedite the population of electrocompacted collagen scaffolds by cells. The capacity of the method to fabricate nonlinear curved topographies with compositional heterogeneous layers is demonstrated by sequential deposition of collagenhydroxyapatite layer over a collagen layer. The complex curved topography of the nasal structure is replicated by the electrochemical compaction method. The presented electrochemical compaction process is an enabling modality which holds significant promise for reconstruction of a wide spectrum of topographically complex systems such as joint surfaces, craniofacial defects, ears, nose or urogenital forms. PMID:26069162

  5. Strategy to Achieve Highly Porous/Biocompatible Macroscale Cell Blocks, Using a Collagen/Genipin-bioink and an Optimal 3D Printing Process.

    PubMed

    Kim, Yong Bok; Lee, Hyeongjin; Kim, Geun Hyung

    2016-11-30

    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 a porous structure with reasonable mechanical strength has not been issued. Here, we propose a printing strategy with optimal conditions 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 mm(3) 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.

  6. The feasibility assessment of radiation dose of movement 3D NIPAM gel by magnetic resonance imaging

    NASA Astrophysics Data System (ADS)

    Hsieh, Chih-Ming; Leung, Joseph Hang; Ng, Yu-Bun; Cheng, Chih-Wu; Sun, Jung-Chang; Lin, Ping-Chin; Hsieh, Bor-Tsung

    2015-11-01

    NIPAM dosimeter is widely accepted and recommended for its 3D distribution and accuracy in dose absorption. Up to the moment, most research works on dose measurement are based on a fixed irradiation target without the consideration of the effect from physiological motion. We present a study to construct a respiratory motion simulating patient anatomical and dosimetry model for the study of dosimetic effect of organ motion. The dose on fixed and motion targets was measured by MRI after a dose adminstration of 1, 2, 5, 8, and 10 Gy from linear accelerator. Comparison of two situations is made. The average sensitivity of fixed NIPAM was 0.1356 s-1/Gy with linearity R2=0.998. The average sensitivity of movement NIPAM was 0.1366 s-1/Gy with linearity R2=0.998 both having only 0.001 of the sensitivity difference. The difference between the two based on dose rate dependency, position and depth was not significant. There was thus no apparent impact on NIPAM dosimeter from physiological motion. The high sensitivity, linearity and stability of NIPAM dosimeter proved to be an ideal apparatus in the dose measurement in these circumstances.

  7. Injectable, high-density collagen gels for annulus fibrosus repair: An in vitro rat tail model.

    PubMed

    Borde, Brandon; Grunert, Peter; Härtl, Roger; Bonassar, Lawrence J

    2015-08-01

    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 cross-linked with riboflavin at 0.03 mM, 0.07 mM, or 0.10 mM. These cross-linked, high-density collagen gels maintained their 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 fourfold 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 the delivery of biological agents such as cells or growth factors that may aid in the repair of the annulus fibrosus.

  8. Injectable, High Density Collagen Gels for Annulus Fibrosus Repair: An In Vitro Rat Tail Model

    PubMed Central

    Borde, Brandon; Grunert, Peter; Härtl, Roger; Bonassar, Lawrence J.

    2014-01-01

    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. PMID:25504661

  9. Sustained Delivery of Bioactive GDNF from Collagen and Alginate-Based Cell-Encapsulating Gel Promoted Photoreceptor Survival in an Inherited Retinal Degeneration Model

    PubMed Central

    Chan, Barbara P.; Lo, Amy C. Y.

    2016-01-01

    Encapsulated-cell therapy (ECT) is an attractive approach for continuously delivering freshly synthesized therapeutics to treat sight-threatening posterior eye diseases, circumventing repeated invasive intravitreal injections and improving local drug availability clinically. Composite collagen-alginate (CAC) scaffold contains an interpenetrating network that integrates the physical and biological merits of its constituents, including biocompatibility, mild gelling properties and availability. However, CAC ECT properties and performance in the eye are not well-understood. Previously, we reported a cultured 3D CAC system that supported the growth of GDNF-secreting HEK293 cells with sustainable GDNF delivery. Here, the system was further developed into an intravitreally injectable gel with 1x104 or 2x105 cells encapsulated in 2mg/ml type I collagen and 1% alginate. Gels with lower alginate concentration yielded higher initial cell viability but faster spheroid formation while increasing initial cell density encouraged cell growth. Continuous GDNF delivery was detected in culture and in healthy rat eyes for at least 14 days. The gels were well-tolerated with no host tissue attachment and contained living cell colonies. Most importantly, gel-implanted in dystrophic Royal College of Surgeons rat eyes for 28 days retained photoreceptors while those containing higher initial cell number yielded better photoreceptor survival. CAC ECT gels offers flexible system design and is a potential treatment option for posterior eye diseases. PMID:27441692

  10. Production and in vitro characterization of 3D porous scaffolds made of magnesium carbonate apatite (MCA)/anionic collagen using a biomimetic approach.

    PubMed

    Sader, Marcia S; Martins, Virginia C A; Gomez, Santiago; LeGeros, Racquel Z; Soares, Gloria A

    2013-10-01

    3D porous scaffolds are relevant biomaterials to bone engineering as they can be used as templates to tissue reconstruction. The aim of the present study was to produce and characterize in vitro 3D magnesium-carbonate apatite/collagen (MCA/col) scaffolds. They were prepared by using biomimetic approach, followed by cross-linking with 0.25% glutaraldehyde solution (GA) and liofilization. Results obtained with Fourier-transform infrared spectroscopy (FT-IR) confirmed the type-B carbonate substitution, while by X-ray diffraction (XRD), a crystallite size of ~10nm was obtained. Optical and electron microscopy showed that the cylindrical samples exhibited an open-porous morphology, with apatite nanocrystals precipitated on collagen fibrils. The cross-linked 3D scaffolds showed integrity when immersed in culture medium up to 14 days. Also, the immersion of such samples into an acid buffer solution, to mimic the osteoclastic resorption environment, promotes the release of important ions for bone repair, such as calcium, phosphorus and magnesium. Bone cells (SaOs2) adhered, and proliferated on the 3D composite scaffolds, showing that synthesis and the cross-linking processes did not induce cytotoxicity.

  11. Human pancreatic stellate cells modulate 3D collagen alignment to promote the migration of pancreatic ductal adenocarcinoma cells.

    PubMed

    Drifka, Cole R; Loeffler, Agnes G; Esquibel, Corinne R; Weber, Sharon M; Eliceiri, Kevin W; Kao, W John

    2016-12-01

    A hallmark of pancreatic ductal adenocarcinoma (PDAC) is the ability for cancer cells to aggressively infiltrate and navigate through a dense stroma during the metastatic process. Key features of the PDAC stroma include an abundant population of activated pancreatic stellate cells (PSCs) and highly aligned collagen fibers; however, important questions remain regarding how collagen becomes aligned and what the biological manifestations are. To better understand how PSCs, aligned collagen, and PDAC cells might cooperate during the transition to invasion, we utilized a microchannel-based in vitro tumor model and advanced imaging technologies to recreate and examine in vivo-like heterotypic interactions. We found that PSCs participate in a collaborative process with cancer cells by orchestrating the alignment of collagen fibers that, in turn, are permissive to enhanced cell migration. Additionally, direct contact between PSCs, collagen, and PDAC cells is critical to invasion and co-migration of both cell types. This suggests PSCs may accompany and assist in navigating PDAC cells through the stromal terrain. Together, our data provides a new role for PSCs in stimulating the metastatic process and underscores the importance of collagen alignment in cancer progression.

  12. 3-D ultrastructure and collagen composition of healthy and overloaded human tendon: evidence of tenocyte and matrix buckling

    PubMed Central

    Pingel, Jessica; Lu, Yinhui; Starborg, Tobias; Fredberg, Ulrich; Langberg, Henning; Nedergaard, Anders; Weis, MaryAnn; Eyre, David; Kjaer, Michael; Kadler, Karl E

    2014-01-01

    Achilles tendinopathies display focal tissue thickening with pain and ultrasonography changes. Whilst complete rupture might be expected to induce changes in tissue organization and protein composition, little is known about the consequences of non-rupture-associated tendinopathies, especially with regards to changes in the content of collagen type I and III (the major collagens in tendon), and changes in tendon fibroblast (tenocyte) shape and organization of the extracellular matrix (ECM). To gain new insights, we took biopsies from the tendinopathic region and flanking healthy region of Achilles tendons of six individuals with clinically diagnosed tendinopathy who had no evidence of cholesterol, uric acid and amyloid accumulation. Biochemical analyses of collagen III/I ratio were performed on all six individuals, and electron microscope analysis using transmission electron microscopy and serial block face-scanning electron microscopy were made on two individuals. In the tendinopathic regions, compared with the flanking healthy tissue, we observed: (i) an increase in the ratio of collagen III : I proteins; (ii) buckling of the collagen fascicles in the ECM; (iii) buckling of tenocytes and their nuclei; and (iv) an increase in the ratio of small-diameter : large-diameter collagen fibrils. In summary, load-induced non-rupture tendinopathy in humans is associated with localized biochemical changes, a shift from large-to small-diameter fibrils, buckling of the tendon ECM, and buckling of the cells and their nuclei. PMID:24571576

  13. In vitro bone formation by mesenchymal stem cells with 3D collagen/β-TCP composite scaffold.

    PubMed

    Todo, Mitsugu; Arahira, Takaaki

    2013-01-01

    Recent years, various kinds of natural polymers and bioceramics has been used to develop porous scaffolds for bone tissue engineering. Among of them, collagen guarantees good biological conditions, and β-tricalcium phosphate (β-TCP) possesses good oseteoconductivity, cellular adhesion, accelerated differentiation and mechanical property. In this study, rat bone marrow mesenchymal stem cells (rMSC) were cultured in β-TCP/collagen composite scaffolds up to 28 days in order to assess the time-dependent behavior of the extracellular matrix formation and the mechanical performance of the scaffold-cell system. The cell number and ALP activity were evaluated using a spectrophotometric plate reader. Gene expression of osteogenesis was analyzed using the real-time PCR reactions. Compression tests were also conducted periodically by using a conventional testing machine to evaluate the elastic modulus. The increasing behaviors of cell number and ALP activity in the composite scaffold were much better than in the collagen scaffold. The gene expression of osteocalcin and collagen type-I in collagen/β-TCP scaffold was higher than that of the collagen scaffold. The compressive modulus also increased up to 28 days. These results clearly showed that the distribution of micro β-TCP particles is very effective to increase the elastic modulus and promote cell growth.

  14. Evaluation of 3D printed PCL/PLGA/β-TCP versus collagen membranes for guided bone regeneration in a beagle implant model.

    PubMed

    Won, J-Y; Park, C-Y; Bae, J-H; Ahn, G; Kim, C; Lim, D-H; Cho, D-W; Yun, W-S; Shim, J-H; Huh, J-B

    2016-10-07

    Here, we compared 3D-printed polycaprolactone/poly(lactic-co-glycolic acid)/β-tricalcium phosphate (PCL/PLGA/β-TCP) membranes with the widely used collagen membranes for guided bone regeneration (GBR) in beagle implant models. For mechanical property comparison in dry and wet conditions and cytocompatibility determination, we analyzed the rate and pattern of cell proliferation of seeded fibroblasts and preosteoblasts using the cell counting kit-8 assay and scanning electron microscopy. Osteogenic differentiation was verified using alizarin red S staining. At 8 weeks following implantation in vivo using beagle dogs, computed tomography and histological analyses were performed after sacrifice. Cell proliferation rates in vitro indicated that early cell attachment was higher in collagen than in PCL/PLGA/β-TCP membranes; however, the difference subsided by day 7. Similar outcomes were found for osteogenic differentiation, with approximately 2.5 times greater staining in collagen than PCL/PLGA/β-TCP, but without significant difference by day 14. In vivo, bone regeneration in the defect area, represented by new bone formation and bone-to-implant contact, paralleled those associated with collagen membranes. However, tensile testing revealed that whereas the PCL/PLGA/β-TCP membrane mechanical properties were conserved in both wet and dry states, the tensile property of collagen was reduced by 99% under wet conditions. Our results demonstrate in vitro and in vivo that PCL/PLGA/β-TCP membranes have similar levels of biocompatibility and bone regeneration as collagen membranes. In particular, considering that GBR is always applied to a wet environment (e.g. blood, saliva), we demonstrated that PCL/PLGA/β-TCP membranes maintained their form more reliably than collagen membranes in a wet setting, confirming their appropriateness as a GBR membrane.

  15. Crack Propagation Versus Fiber Alignment in Collagen Gels: Experiments and Multiscale Simulation

    PubMed Central

    Vanderheiden, Sarah M.; Hadi, Mohammad F.; Barocas, V. H.

    2015-01-01

    It is well known that the organization of the fibers constituting a collagenous tissue can affect its failure behavior. Less clear is how that effect can be described computationally so as to predict the failure of a native or engineered tissue under the complex loading conditions that can occur in vivo. Toward the goal of a general predictive strategy, we applied our multiscale model of collagen gel mechanics to the failure of a double-notched gel under tension, comparing the results for aligned and isotropic samples. In both computational and laboratory experiments, we found that the aligned gels were more likely to fail by connecting the two notches than the isotropic gels. For example, when the initial notches were 30% of the sample width (normalized tip-to-edge distance = 0.7), the normalized tip-to-tip distance at which the transition occurred from between-notch failure to across-sample failure shifted from 0.6 to 1.0. When the model predictions for the type of failure event (between the two notches versus across the sample width) were compared to the experimental results, the two were found to be strongly covariant by Fisher’s exact test (p < 0.05) for both the aligned and isotropic gels with no fitting parameters. Although the double-notch system is idealized, and the collagen gel system is simpler than a true tissue, it presents a simple model system for studying failure of anisotropic tissues in a controlled setting. The success of the computational model suggests that the multiscale approach, in which the structural complexity is incorporated via changes in the model networks rather than via changes to a constitutive equation, has the potential to predict tissue failure under a wide range of conditions. PMID:26355475

  16. Expression of osteoblastic phenotype in periodontal ligament fibroblasts cultured in three-dimensional collagen gel

    PubMed Central

    ALVES, Luciana Bastos; MARIGUELA, Viviane Casagrande; GRISI, Márcio Fernando de Moraes; de SOUZA, Sérgio Luiz Scaombatti; NOVAES, Arthur Belém; TABA, Mário; de OLIVEIRA, Paulo Tambasco; PALIOTO, Daniela Bazan

    2015-01-01

    Objective : To investigate the influence of a three-dimensional cell culture model on the expression of osteoblastic phenotype in human periodontal ligament fibroblast (hPDLF) cultures. Material and Methods : hPDLF were seeded on bi-dimensional (2D) and three-dimensional (3D) collagen type I (experimental groups) and and on a plastic coverslip (control) for up to 14 days. Cell viability and alkaline phosphatase (ALP) activity were performed. Also, cell morphology and immunolabeling for alkaline phosphatase (ALP) and osteopontin (OPN) were assessed by epifluorescence and confocal microscopy. The expression of osteogenic markers, including alkaline phosphatase, osteopontin, osteocalcin (OC), collagen I (COL I) and runt-related transcription factor 2 (RUNX2), were analyzed using real-time polymerase chain reaction (RT-PCR). Mineralized bone-like nodule formation was visualized by microscopy and calcium content was assessed quantitatively by alizarin red assay. Results : Experimental cultures produced an increase in cell proliferation. Immunolabeling for OPN and ALP in hPDLF were increased and ALP activity was inhibited by three-dimensional conditions. OPN and RUNX2 gene expression was significantly higher on 3D culture when compared with control surface. Moreover, ALP and COL I gene expression were significantly higher in three-dimensional collagen than in 2D cultures at 7 days. However, at 14 days, 3D cultures exhibited ALP and COL I gene expression significantly lower than the control, and the COL I gene expression was also significantly lower in 3D than in 2D cultures. Significant calcium mineralization was detected and quantified by alizarin red assay, and calcified nodule formation was not affected by tridimensionality. Conclusion : This study suggests that the 3D cultures are able to support hPDLF proliferation and favor the differentiation and mineralized matrix formation, which may be a potential periodontal regenerative therapy. PMID:26018313

  17. Physics of soft hyaluronic acid-collagen type II double network gels

    NASA Astrophysics Data System (ADS)

    Morozova, Svetlana; Muthukumar, Murugappan

    2015-03-01

    Many biological hydrogels are made up of multiple interpenetrating, charged components. We study the swelling, elastic diffusion, mechanical, and optical behaviors of 100 mol% ionizable hyaluronic acid (HA) and collagen type II fiber networks. Dilute, 0.05-0.5 wt% hyaluronic acid networks are extremely sensitive to solution salt concentration, but are stable at pH above 2. When swelled in 0.1M NaCl, single-network hyaluronic acid gels follow scaling laws relevant to high salt semidilute solutions; the elastic shear modulus G' and diffusion constant D scale with the volume fraction ϕ as G' ~ϕ 9 / 4 and D ~ϕ 3 / 4 , respectively. With the addition of a collagen fiber network, we find that the hyaluronic acid network swells to suspend the rigid collagen fibers, providing extra strength to the hydrogel. Results on swelling equilibria, elasticity, and collective diffusion on these double network hydrogels will be presented.

  18. Factors affecting morphogenesis of rabbit gallbladder epithelial cells cultured in collagen gels.

    PubMed

    Mori, M; Miyazaki, K

    2000-05-01

    Although peptide growth factors play an important role in the morphogenesis of gallbladder, little is known about how they effect the morphogenesis of gallbladder epithelial cells. Rabbit gallbladder epithelial cells (RGEC) were isolated and cultured in monolayer or collagen gels. Epidermal growth factor (EGF), hepatocyte growth factor (HGF), epimorphin, transforming growth factor-beta 1 (TGF-beta 1), and fibroblast-conditioned medium (FCM) were added to the cultured cells to clarify the effects of these peptides and FCM on morphogenesis of RGEC. RGEC suspended in collagen gels form spherical cysts with morphologic polarity. EGF, HGF, epimorphin, and FCM promoted cyst maturation by accelerating the proliferation and aggregation of clear, polarized vesicles. In contrast, TGF-beta 1 markedly inhibited DNA synthesis in both monolayer and collagen gel cultures and promoted formation of branching structures in collagen gels. Furthermore, in the presence of EGF, TGF-beta 1 induced a drastic change in morphogenesis, with the formation of branching networks that showed cell-cell contact only at sites where branches touched. RGEC-forming multicellular cysts did not express vimentin but expressed significant amounts of cytokeratin and regained junctional complexes. In contrast, TGF-beta 1-treated cells strongly expressed vimentin along with branching structures and showed decreases in cytokeratin expression and junctional complexes. Thus, TGF-beta 1 induces a mesenchyme-like cell shape accompanied by cytoskeletal molecular changes, with loss of both epithelial polarization and junctional complexes. These results suggest that the morphogenetic program of RGEC is likely to be determined by the interaction of these peptides and the timing of their presence.

  19. Leishmania amazonensis promastigotes in 3D Collagen I culture: an in vitro physiological environment for the study of extracellular matrix and host cell interactions

    PubMed Central

    Rodrigues, Juliany C.F.; Viana, Nathan B.; Pontes, Bruno; Pereira, Camila F.A.; Silva-Filho, Fernando C.

    2014-01-01

    Leishmania amazonensis is the causative agent of American cutaneous leishmaniasis, an important neglected tropical disease. Once Leishmania amazonensis is inoculated into the human host, promastigotes are exposed to the extracellular matrix (ECM) of the dermis. However, little is known about the interaction between the ECM and Leishmania promastigotes. In this study we established L. amazonensis promastigote culture in a three-dimensional (3D) environment mainly composed of Collagen I (COL I). This 3D culture recreates in vitro some aspects of the human host infection site, enabling the study of the interaction mechanisms of L. amazonensis with the host ECM. Promastigotes exhibited “freeze and run” migration in the 3D COL I matrix, which is completely different from the conventional in vitro swimming mode of migration. Moreover, L. amazonensis promastigotes were able to invade, migrate inside, and remodel the 3D COL I matrix. Promastigote trans-matrix invasion and the freeze and run migration mode were also observed when macrophages were present in the matrix. At least two classes of proteases, metallo- and cysteine proteases, are involved in the 3D COL I matrix degradation caused by Leishmania. Treatment with a mixture of protease inhibitors significantly reduced promastigote invasion and migration through this matrix. Together our results demonstrate that L. amazonensis promastigotes release proteases and actively remodel their 3D environment, facilitating their migration. This raises the possibility that promastigotes actively interact with their 3D environment during the search for their cellular “home”—macrophages. Supporting this hypothesis, promastigotes migrated faster than macrophages in a novel 3D co-culture model. PMID:24765565

  20. Leishmania amazonensis promastigotes in 3D Collagen I culture: an in vitro physiological environment for the study of extracellular matrix and host cell interactions.

    PubMed

    Petropolis, Debora B; Rodrigues, Juliany C F; Viana, Nathan B; Pontes, Bruno; Pereira, Camila F A; Silva-Filho, Fernando C

    2014-01-01

    Leishmania amazonensis is the causative agent of American cutaneous leishmaniasis, an important neglected tropical disease. Once Leishmania amazonensis is inoculated into the human host, promastigotes are exposed to the extracellular matrix (ECM) of the dermis. However, little is known about the interaction between the ECM and Leishmania promastigotes. In this study we established L. amazonensis promastigote culture in a three-dimensional (3D) environment mainly composed of Collagen I (COL I). This 3D culture recreates in vitro some aspects of the human host infection site, enabling the study of the interaction mechanisms of L. amazonensis with the host ECM. Promastigotes exhibited "freeze and run" migration in the 3D COL I matrix, which is completely different from the conventional in vitro swimming mode of migration. Moreover, L. amazonensis promastigotes were able to invade, migrate inside, and remodel the 3D COL I matrix. Promastigote trans-matrix invasion and the freeze and run migration mode were also observed when macrophages were present in the matrix. At least two classes of proteases, metallo- and cysteine proteases, are involved in the 3D COL I matrix degradation caused by Leishmania. Treatment with a mixture of protease inhibitors significantly reduced promastigote invasion and migration through this matrix. Together our results demonstrate that L. amazonensis promastigotes release proteases and actively remodel their 3D environment, facilitating their migration. This raises the possibility that promastigotes actively interact with their 3D environment during the search for their cellular "home"-macrophages. Supporting this hypothesis, promastigotes migrated faster than macrophages in a novel 3D co-culture model.

  1. Extracellular Protease Inhibition Alters the Phenotype of Chondrogenically Differentiating Human Mesenchymal Stem Cells (MSCs) in 3D Collagen Microspheres

    PubMed Central

    Han, Sejin; Li, Yuk Yin; Chan, Barbara Pui

    2016-01-01

    Matrix remodeling of cells is highly regulated by proteases and their inhibitors. Nevertheless, how would the chondrogenesis of mesenchymal stem cells (MSCs) be affected, when the balance of the matrix remodeling is disturbed by inhibiting matrix proteases, is incompletely known. Using a previously developed collagen microencapsulation platform, we investigated whether exposing chondrogenically differentiating MSCs to intracellular and extracellular protease inhibitors will affect the extracellular matrix remodeling and hence the outcomes of chondrogenesis. Results showed that inhibition of matrix proteases particularly the extracellular ones favors the phenotype of fibrocartilage rather than hyaline cartilage in chondrogenically differentiating hMSCs by upregulating type I collagen protein deposition and type II collagen gene expression without significantly altering the hypertrophic markers at gene level. This study suggests the potential of manipulating extracellular proteases to alter the outcomes of hMSC chondrogenesis, contributing to future development of differentiation protocols for fibrocartilage tissues for intervertebral disc and meniscus tissue engineering. PMID:26760956

  2. Multiphoton microscopy of engineered dermal substitutes: assessment of 3-D collagen matrix remodeling induced by fibroblast contraction

    NASA Astrophysics Data System (ADS)

    Pena, Ana-Maria; Fagot, Dominique; Olive, Christian; Michelet, Jean-François; Galey, Jean-Baptiste; Leroy, Frédéric; Beaurepaire, Emmanuel; Martin, Jean-Louis; Colonna, Anne; Schanne-Klein, Marie-Claire

    2010-09-01

    Dermal fibroblasts are responsible for the generation of mechanical forces within their surrounding extracellular matrix and can be potentially targeted by anti-aging ingredients. Investigation of the modulation of fibroblast contraction by these ingredients requires the implementation of three-dimensional in situ imaging methodologies. We use multiphoton microscopy to visualize unstained engineered dermal tissue by combining second-harmonic generation that reveals specifically fibrillar collagen and two-photon excited fluorescence from endogenous cellular chromophores. We study the fibroblast-induced reorganization of the collagen matrix and quantitatively evaluate the effect of Y-27632, a RhoA-kinase inhibitor, on dermal substitute contraction. We observe that collagen fibrils rearrange around fibroblasts with increasing density in control samples, whereas collagen fibrils show no remodeling in the samples containing the RhoA-kinase inhibitor. Moreover, we show that the inhibitory effects are reversible. Our study demonstrates the relevance of multiphoton microscopy to visualize three-dimensional remodeling of the extracellular matrix induced by fibroblast contraction or other processes.

  3. The 3-D collagen structure of equine articular cartilage, characterized using variable-angle-of-incidence polarization-sensitive optical coherence tomography

    NASA Astrophysics Data System (ADS)

    Ugryumova, Nadya; Gangnus, Sergei V.; Matcher, Stephen J.

    2005-08-01

    Polarization-sensitive optical coherence tomography has been used to spatially map the birefringence of equine articular cartilage. Images obtained in the vicinity of visible osteoarthritic lesions display a characteristic disruption of the regular birefringence bands shown by normal cartilage. We also note that significant (e.g. ×2) variations in the apparent birefringence of samples taken from young (18 month) animals that otherwise appear visually homogeneous are found over spatial scales of a few millimeters. We suggest that whilst some of this variation may be due to changes in the intrinsic birefringence of the tissue, the 3-D orientation of the collagen fibers relative to the plane of the joint surface should also be taken into account. We propose a method based on multiple angles of illumination to determine the polar angle of the collagen fibers.

  4. Protocol and cell responses in three-dimensional conductive collagen gel scaffolds with conductive polymer nanofibres for tissue regeneration.

    PubMed

    Sirivisoot, Sirinrath; Pareta, Rajesh; Harrison, Benjamin S

    2014-02-06

    It has been established that nerves and skeletal muscles respond and communicate via electrical signals. In regenerative medicine, there is current emphasis on using conductive nanomaterials to enhance electrical conduction through tissue-engineered scaffolds to increase cell differentiation and tissue regeneration. We investigated the role of chemically synthesized polyaniline (PANI) and poly(3,4-ethylenedioxythiophene) (PEDOT) conductive polymer nanofibres for conductive gels. To mimic a naturally derived extracellular matrix for cell growth, type I collagen gels were reconstituted with conductive polymer nanofibres and cells. Cell viability and proliferation of PC-12 cells and human skeletal muscle cells on these three-dimensional conductive collagen gels were evaluated in vitro. PANI and PEDOT nanofibres were found to be cytocompatible with both cell types and the best results (i.e. cell growth and gel electrical conductivity) were obtained with a low concentration (0.5 wt%) of PANI. After 7 days of culture in the conductive gels, the densities of both cell types were similar and comparable to collagen positive controls. Moreover, PC-12 cells were found to differentiate in the conductive hydrogels without the addition of nerve growth factor or electrical stimulation better than collagen control. Importantly, electrical conductivity of the three-dimensional gel scaffolds increased by more than 400% compared with control. The increased conductivity and injectability of the cell-laden collagen gels to injury sites in order to create an electrically conductive extracellular matrix makes these biomaterials very conducive for the regeneration of tissues.

  5. Gel-spinning of mimetic collagen and collagen/nano-carbon fibers: Understanding multi-scale influences on molecular ordering and fibril alignment.

    PubMed

    Green, Emily C; Zhang, Yiying; Li, Heng; Minus, Marilyn L

    2017-01-01

    Synthetic gel-spun collagen and collagen/nano-carbon fibers were found to exhibit structural mimicry comparable to native tendons. X-ray scattering and microscopy analyses are used to characterize the molecular and fibrillar alignment in the synthetic fibers, where D-banding is observed throughout the spun fibers - consistent with native collagen. For the composite collagen/nano-carbon fibers, the morphology and dispersion quality of the nano-carbons within was found to play a significant role in influencing collagen molecular ordering and fibril alignment. Fibrillar and molecular alignment was also better preserved during elongation of the composites as compared to the control collagen fibers. These results show the structural influence of a rigid inclusion on the collagen fibril structure. Both dry- and wet-state tensile testing were performed on the collagen fibers, and these results show behavior comparable to the native materials. Dry-state tests also reveal interfacial interaction between the nano-fillers and the collagen fibrils through theoretical analysis. Wet-state tensile testing indicates the structure-property behavior of the mimetic hierarchical structure within the synthetic fibers.

  6. The endogenous fluorescence of fibroblast in collagen gels as indicator of stiffness of the extracellular matrix

    NASA Astrophysics Data System (ADS)

    Padilla-Martinez, J. P.; Ortega-Martinez, A.; Franco, W.

    2016-03-01

    The stiffness or rigidity of the extracellular matrix (ECM) regulates cell response. Established mechanical tests to measure stiffness, such as indentation and tensile tests, are invasive and destructive to the sample. Endogenous or native molecules to cells and ECM components, like tryptophan and cross-links of collagen, display fluorescence upon irradiation with ultraviolet light. Most likely, the concentration of these endogenous fluorophores changes as the stiffness of the ECM changes. In this work we investigate the endogenous fluorescence of collagen gels containing fibroblasts as a non-invasive non-destructive method to measure stiffness of the ECM. Human fibroblast cells were cultured in three-dimensional gels of type I collagen (50,000 cells/ml). This construct is a simple model of tissue contraction. During contraction, changes in the excitation-emission matrix (a fluorescence map in the 240-520/290-530 nm range) of constructs were measured with a spectrofluoremeter, and changes in stiffness were measured with a standard indentation test over 16 days. Results show that a progressive increase in fluorescence of the 290/340 nm excitation-emission pair correlates with a progressive increase in stiffness (r=0.9, α=0.5). The fluorescence of this excitation-emission pair is ascribed to tryptophan and variations in the fluorescence of this pair correlate with cellular proliferation. In this tissue model, the endogenous functional fluorescence of proliferating fibroblast cells is a biomechanical marker of stiffness of the ECM.

  7. A modified collagen gel enhances healing outcome in a preclinical swine model of excisional wounds.

    PubMed

    Elgharably, Haytham; Roy, Sashwati; Khanna, Savita; Abas, Motaz; Dasghatak, Piya; Das, Amitava; Mohammed, Kareem; Sen, Chandan K

    2013-01-01

    Collagen-based dressings are of great interest in wound care. However, evidence supporting their mechanism of action is scanty. This work provides first results from a preclinical swine model of excisional wounds, elucidating the mechanism of action of a modified collagen gel (MCG) dressing. Following wounding, wound-edge tissue was collected at specific time intervals (3, 7, 14, and 21 days postwounding). On day 7, histological analysis showed significant increase in the length of rete ridges, suggesting improved biomechanical properties of the healing wound tissue. Rapid and transient mounting of inflammation is necessary for efficient healing. MCG significantly accelerated neutrophil and macrophage recruitment to the wound site on day 3 and day 7 with successful resolution of inflammation on day 21. MCG induced monocyte chemotactic protein-1 expression in neutrophil-like human promyelocytic leukemia-60 cells in vitro. In vivo, MCG-treated wound tissue displayed elevated vascular endothelial growth factor expression. Consistently, MCG-treated wounds displayed significantly higher abundance of endothelial cells with increased blood flow to the wound area indicating improved vascularization. This observation was explained by the finding that MCG enhanced proliferation of wound-site endothelial cells. In MCG-treated wound tissue, Masson's trichrome and picrosirius red staining showed higher abundance of collagen and increased collagen type I:III ratio. This work presents first evidence from a preclinical setting explaining how a collagen-based dressing may improve wound closure by targeting multiple key mechanisms. The current findings warrant additional studies to determine whether the responses to the MCG are different from other collagen-based products used in clinical setting.

  8. 2D protrusion but not motility predicts growth factor-induced cancer cell migration in 3D collagen.

    PubMed

    Meyer, Aaron S; Hughes-Alford, Shannon K; Kay, Jennifer E; Castillo, Amalchi; Wells, Alan; Gertler, Frank B; Lauffenburger, Douglas A

    2012-06-11

    Growth factor-induced migration is a critical step in the dissemination and metastasis of solid tumors. Although differences in properties characterizing cell migration on two-dimensional (2D) substrata versus within three-dimensional (3D) matrices have been noted for particular growth factor stimuli, the 2D approach remains in more common use as an efficient surrogate, especially for high-throughput experiments. We therefore were motivated to investigate which migration properties measured in various 2D assays might be reflective of 3D migratory behavioral responses. We used human triple-negative breast cancer lines stimulated by a panel of receptor tyrosine kinase ligands relevant to mammary carcinoma progression. Whereas 2D migration properties did not correlate well with 3D behavior across multiple growth factors, we found that increased membrane protrusion elicited by growth factor stimulation did relate robustly to enhanced 3D migration properties of the MDA-MB-231 and MDA-MB-157 lines. Interestingly, we observed this to be a more reliable relationship than cognate receptor expression or activation levels across these and two additional mammary tumor lines.

  9. The effect of pulsed HIFU on the porosity and permeability of collagen gels: An in vitro study

    NASA Astrophysics Data System (ADS)

    Vipulanandan, Geethanjali; O'Neill, Brian E.

    2012-10-01

    Pulsed HIFU is hypothesized to alter permeability of the extracellular matrix by altering the collagen network. In this study, the ability of HIFU to disrupt the extracellular matrix, particularly Type I collagen, in vitro, was investigated in order to enhance the drug delivery to highly collagenous tumors. This was tested in vitro in two ways, first using dye penetration, and second, by confocal reflection microscopy. Based on the analyses, it was concluded that there was at least a three-fold increase in porosity of the collagen gels after HIFU treatment.

  10. Optical laser scanning of a leucodye micelle gel: preliminary results of a 3D dose verification of an IMRT treatment for a brain tumor

    NASA Astrophysics Data System (ADS)

    Vandecasteele, J.; De Deene, Y.

    2013-06-01

    In the present study an in-house developed leucodye micelle gel was used in combination with an in-house developed optical laser scanner for the 3D dose verification of an IMRT treatment of a pituitary adenoma. In an initial prospective study, a gel measured depth dose distribution of a square 6 MV photon beam was compared with an ion chamber measurement. In a second experiment, the gel and scanner were used to verify a clinical dose distribution on a recently installed linear accelerator. The calibration procedure is identified as the major source of dose deviations.

  11. Contraction-induced Mmp13 and -14 expression by goat articular chondrocytes in collagen type I but not type II gels.

    PubMed

    Berendsen, Agnes D; Vonk, Lucienne A; Zandieh-Doulabi, Behrouz; Everts, Vincent; Bank, Ruud A

    2012-10-01

    Collagen gels are promising scaffolds to prepare an implant for cartilage repair but several parameters, such as collagen concentration and composition as well as cell density, should be carefully considered, as they are reported to affect phenotypic aspects of chondrocytes. In this study we investigated whether the presence of collagen type I or II in gel lattices affects matrix contraction and relative gene expression levels of matrix proteins, MMPs and the subsequent degradation of collagen by goat articular chondrocytes. Only floating collagen I gels, and not those attached or composed of type II collagen, contracted during a culture period of 12 days. This coincided with an upregulation of both Mmp13 and -14 gene expression, whereas Mmp1 expression was not affected. The release of hydroxyproline in the culture medium, indicating matrix degradation, was increased five-fold in contracted collagen I gels compared to collagen II gels without contraction. Furthermore, blocking contraction of collagen I gels by cytochalasin B inhibited Mmp13 and -14 expression and the release of hydroxyproline. The expression of cartilage-specific ECM genes was decreased in contracted collagen I gels, with increased numbers of cells with an elongated morphology, suggesting that matrix contraction induces dedifferentiation of chondrocytes into fibroblast-like cells. We conclude that the collagen composition of the gels affects matrix contraction by articular chondrocytes and that matrix contraction induces an increased Mmp13 and -14 expression as well as matrix degradation.

  12. Longitudinal, 3D Imaging of Collagen Remodeling in Murine Hypertrophic Scars In Vivo using Polarization-sensitive Optical Frequency Domain Imaging

    PubMed Central

    Lo, William C. Y.; Villiger, Martin; Golberg, Alexander; Broelsch, G. Felix; Khan, Saiqa; Lian, Christine G.; Austen, William G.; Yarmush, Martin; Bouma, Brett E.

    2016-01-01

    Hypertrophic scars (HTS), frequently seen after traumatic injuries and surgery, remain a major clinical challenge due to the limited success of existing therapies. A significant obstacle to understanding HTS etiology is the lack of tools to monitor scar remodeling longitudinally and non-invasively. We present an in vivo, label-free technique using polarization-sensitive optical frequency domain imaging (PS-OFDI) for the 3D, longitudinal assessment of collagen remodeling in murine HTS. In this study, HTS was induced with a mechanical tension device for 4 to 10 days on incisional wounds and imaged up to one month after device removal; an excisional HTS model was also imaged at 6 months after injury to investigate deeper and more mature scars. We showed that local retardation (LR) and degree of polarization (DOP) provide a robust signature for HTS. Compared to normal skin with heterogeneous LR and low DOP, HTS was characterized by an initially low LR, which increased as collagen fibers remodeled, and a persistently high DOP. This study demonstrates that PS-OFDI offers a powerful tool to gain significant biological insights into HTS remodeling by enabling longitudinal assessment of collagen in vivo, which is critical to elucidating HTS etiology and developing more effective HTS therapies. PMID:26763427

  13. Longitudinal, 3D Imaging of Collagen Remodeling in Murine Hypertrophic Scars In Vivo Using Polarization-Sensitive Optical Frequency Domain Imaging.

    PubMed

    Lo, William C Y; Villiger, Martin; Golberg, Alexander; Broelsch, G Felix; Khan, Saiqa; Lian, Christine G; Austen, William G; Yarmush, Martin; Bouma, Brett E

    2016-01-01

    Hypertrophic scars (HTS), frequently seen after traumatic injuries and surgery, remain a major clinical challenge because of the limited success of existing therapies. A significant obstacle to understanding HTS etiology is the lack of tools to monitor scar remodeling longitudinally and noninvasively. We present an in vivo, label-free technique using polarization-sensitive optical frequency domain imaging for the 3D, longitudinal assessment of collagen remodeling in murine HTS. In this study, HTS was induced with a mechanical tension device for 4-10 days on incisional wounds and imaged up to 1 month after device removal; an excisional HTS model was also imaged at 6 months after injury to investigate deeper and more mature scars. We showed that local retardation and degree of polarization provide a robust signature for HTS. Compared with normal skin with heterogeneous local retardation and low degree of polarization, HTS was characterized by an initially low local retardation, which increased as collagen fibers remodeled, and a persistently high degree of polarization. This study demonstrates that polarization-sensitive optical frequency domain imaging offers a powerful tool to gain significant biological insights into HTS remodeling by enabling longitudinal assessment of collagen in vivo, which is critical to elucidating HTS etiology and developing more effective HTS therapies.

  14. A modified collagen gel dressing promotes angiogenesis in a preclinical swine model of chronic ischemic wounds.

    PubMed

    Elgharably, Haytham; Ganesh, Kasturi; Dickerson, Jennifer; Khanna, Savita; Abas, Motaz; Ghatak, Piya Das; Dixit, Sriteja; Bergdall, Valerie; Roy, Sashwati; Sen, Chandan K

    2014-01-01

    We recently performed proteomic characterization of a modified collagen gel (MCG) dressing and reported promising effects of the gel in healing full-thickness excisional wounds. In this work, we test the translational relevance of our aforesaid findings by testing the dressing in a swine model of chronic ischemic wounds recently reported by our laboratory. Full-thickness excisional wounds were established in the center of bipedicle ischemic skin flaps on the backs of animals. Ischemia was verified by laser Doppler imaging, and MCG was applied to the test group of wounds. Seven days post wounding, macrophage recruitment to the wound was significantly higher in MCG-treated ischemic wounds. In vitro, MCG up-regulated expression of Mrc-1 (a reparative M2 macrophage marker) and induced the expression of anti-inflammatory cytokine interleukin (IL)-10 and of fibroblast growth factor-basic (β-FGF). An increased expression of CCR2, an M2 macrophage marker, was noted in the macrophages from MCG treated wounds. Furthermore, analyses of wound tissues 7 days post wounding showed up-regulation of transforming growth factor-β, vascular endothelial growth factor, von Willebrand's factor, and collagen type I expression in MCG-treated ischemic wounds. At 21 days post wounding, MCG-treated ischemic wounds displayed higher abundance of proliferating endothelial cells that formed mature vascular structures and increased blood flow to the wound. Fibroblast count was markedly higher in MCG-treated ischemic wound-edge tissue. In addition, MCG-treated wound-edge tissues displayed higher abundance of mature collagen with increased collagen type I : III deposition. Taken together, MCG helped mount a more robust inflammatory response that resolved in a timely manner, followed by an enhanced proliferative phase, angiogenic outcome, and postwound tissue remodeling. Findings of the current study warrant clinical testing of MCG in a setting of ischemic chronic wounds.

  15. Riboflavin crosslinked high-density collagen gel for the repair of annular defects in intervertebral discs: An in vivo study.

    PubMed

    Grunert, Peter; Borde, Brandon H; Towne, Sara B; Moriguchi, Yu; Hudson, Katherine D; Bonassar, Lawrence J; Härtl, Roger

    2015-10-01

    Open annular defects compromise the ability of the annulus fibrosus to contain nuclear tissue in the disc space, and therefore lead to disc herniation with subsequent degenerative changes to the entire intervertebral disc. This study reports the use of riboflavin crosslinked high-density collagen gel for the repair of annular defects in a needle-punctured rat-tail model. High-density collagen has increased stiffness and greater hydraulic permeability than conventional low-density gels; riboflavin crosslinking further increases these properties. This study found that treating annular defects with crosslinked high-density collagen inhibited the progression of disc degeneration over 18 weeks compared to untreated control discs. Histological sections of FITC-labeled collagen gel revealed an early tight attachment to host annular tissue. The gel was subsequently infiltrated by host fibroblasts which remodeled it into a fibrous cap that bridged the outer disrupted annular fibers and partially repaired the defect. This repair tissue enhanced retention of nucleus pulposus tissue, maintained physiological disc hydration, and preserved hydraulic permeability, according to MRI, histological, and mechanical assessments. Degenerative changes were partially reversed in treated discs, as indicated by an increase in nucleus pulposus size and hydration between weeks 5 and 18. The collagen gel appeared to work as an instant sealant and by enhancing the intrinsic healing capabilities of the host tissue.

  16. Maturation of human embryonic stem cell-derived cardiomyocytes (hESC-CMs) in 3D collagen matrix: Effects of niche cell supplementation and mechanical stimulation.

    PubMed

    Zhang, W; Kong, C W; Tong, M H; Chooi, W H; Huang, N; Li, R A; Chan, B P

    2017-02-01

    Cardiomyocytes derived from human embryonic stem cells (hESC-CMs) are regarded as a promising source for regenerative medicine, drug testing and disease modeling. Nevertheless, cardiomyocytes are immature in terms of their contractile structure, metabolism and electrophysiological properties. Here, we fabricate cardiac muscle strips by encapsulating hESC-CMs in collagen-based biomaterials. Supplementation of niche cells at 3% to the number of hESC-CMs enhance the maturation of the hESC-CMs in 3D tissue matrix. The benefits of adding mesenchymal stem cells (MSCs) are comparable to that of adding fibroblasts. These two cell types demonstrate similar effects in promoting the compaction and cell spreading, as well as expression of maturation markers at both gene and protein levels. Mechanical loading, particularly cyclic stretch, produces engineered cardiac tissues with higher maturity in terms of twitch force, elastic modulus, sarcomere length and molecular signature, when comparing to static stretch or non-stretched controls. The current study demonstrates that the application of niche cells and mechanical stretch both stimulate the maturation of hESC-CMs in 3D architecture. Our results therefore suggest that this 3D model can be used for in vitro cardiac maturation study.

  17. SU-E-T-105: Development of 3D Dose Verification System for Volumetric Modulated Arc Therapy Using Improved Polyacrylamide-Based Gel Dosimeter

    SciTech Connect

    Ono, K; Fujimoto, S; Akagi, Y; Hirokawa, Y; Hayashi, S; Miyazawa, M

    2014-06-01

    Purpose: The aim of this dosimetric study was to develop 3D dose verification system for volumetric modulated arc therapy (VMAT) using polyacrylamide-based gel (PAGAT) dosimeter improved the sensitivity by magnesium chloride (MgCl{sub 2}). Methods: PAGAT gel containing MgCl{sub 2} as a sensitizer was prepared in this study. Methacrylic-acid-based gel (MAGAT) was also prepared to compare the dosimetric characteristics with PAGAT gel. The cylindrical glass vials (4 cm diameter, 12 cm length) filled with each polymer gel were irradiated with 6 MV photon beam using Novalis Tx linear accelerator (Varian/BrainLAB). The irradiated polymer gel dosimeters were scanned with Signa 1.5 T MRI system (GE), and dose calibration curves were obtained using T{sub 2} relaxation rate (R{sub 2} = 1/T{sub 2}). Dose rate (100-600 MU min{sup −1}) and fractionation (1-8 fractions) were varied. In addition, a cubic acrylic phantom (10 × 10 × 10 cm{sup 3}) filled with improved PAGAT gel inserted into the IMRT phantom (IBA) was irradiated with VMAT (RapidArc). C-shape structure was used for the VMAT planning by the Varian Eclipse treatment planning system (TPS). The dose comparison of TPS and measurements with the polymer gel dosimeter was accomplished by the gamma index analysis, overlaying the dose profiles for a set of data on selected planes using in-house developed software. Results: Dose rate and fractionation dependence of improved PAGAT gel were smaller than MAGAT gel. A high similarity was found by overlaying the dose profiles measured with improved PAGAT gel dosimeter and the TPS dose, and the mean pass rate of the gamma index analysis using 3%/3 mm criteria was achieved 90% on orthogonal planes for VMAT using improved PAGAT gel dosimeter. Conclusion: In-house developed 3D dose verification system using improved polyacrylamide-based gel dosimeter had a potential as an effective tool for VMAT QA.

  18. Ectopic bone formation in rapidly fabricated acellular injectable dense collagen-Bioglass hybrid scaffolds via gel aspiration-ejection.

    PubMed

    Miri, Amir K; Muja, Naser; Kamranpour, Neysan O; Lepry, William C; Boccaccini, Aldo R; Clarke, Susan A; Nazhat, Showan N

    2016-04-01

    Gel aspiration-ejection (GAE) has recently been introduced as an effective technique for the rapid production of injectable dense collagen (IDC) gel scaffolds with tunable collagen fibrillar densities (CFDs) and microstructures. Herein, a GAE system was applied for the advanced production and delivery of IDC and IDC-Bioglass(®) (IDC-BG) hybrid gel scaffolds for potential bone tissue engineering applications. The efficacy of GAE in generating mineralizable IDC-BG gels (from an initial 75-25 collagen-BG ratio) produced through needle gauge numbers 8G (3.4 mm diameter and 6 wt% CFD) and 14G (1.6 mm diameter and 14 wt% CFD) was investigated. Second harmonic generation (SHG) imaging of as-made gels revealed an increase in collagen fibril alignment with needle gauge number. In vitro mineralization of IDC-BG gels was confirmed where carbonated hydroxyapatite was detected as early as day 1 in simulated body fluid, which progressively increased up to day 14. In vivo mineralization of, and host response to, acellular IDC and IDC-BG gel scaffolds were further investigated following subcutaneous injection in adult rats. Mineralization, neovascularization and cell infiltration into the scaffolds was enhanced by the addition of BG and at day 21 post injection, there was evidence of remodelling of granulation tissue into woven bone-like tissue in IDC-BG. SHG imaging of explanted scaffolds indicated collagen fibril remodelling through cell infiltration and mineralization over time. In sum, the results suggest that IDC-BG hybrid gels have osteoinductive properties and potentially offer a novel therapeutic approach for procedures requiring the injectable delivery of a malleable and dynamic bone graft that mineralizes under physiological conditions.

  19. A novel rat tail collagen type-I gel for the cultivation of human articular chondrocytes in low cell density.

    PubMed

    Muller-Rath, R; Gavénis, K; Andereya, S; Mumme, T; Schmidt-Rohlfing, B; Schneider, U

    2007-12-01

    Collagen type-I matrix systems have gained growing importance as a cartilage repair device. However, most of the established matrix systems use collagen type-I of bovine origin seeded in high cell densities. Here we present a novel collagen type-I gel system made of rat tail collagen for the cultivation of human chondrocytes in low cell densities. Rat tail collagen type-I gel (CaReS, Arthro Kinetics, Esslingen, Germany) was seeded with human passage 2 chondrocytes in different cell densities to evaluate the optimal cell number. In vitro, the proliferation factor of low density cultures was more than threefold higher compared with high density cultures. After 6 weeks of in vitro cultivation, freshly prepared chondrocytes with an initial cell density of 2x10(5) cells/mL showed a proliferation factor of 33. A cell density of 2x10(5) cells/mL was chosen for in vitro and in vivo cultivation using the common nude mouse model as an in vivo system. Chondrocytes stayed viable as a Live/Dead fluorescence assay and TUNEL staining revealed. During in vitro cultivation, passage 0 cells partly dedifferentiated morphologically. In vivo, passage 0 cells maintained the chondrocyte phenotype and demonstrated an increased synthesis of collagen type-II protein and gene expression compared to passage 2 cells. Passage 2 cells did not redifferentiate in vivo. Cultivating a cell-seeded collagen gel of bovine origin as a control (AtelocollagenTM, Koken, Tokyo, Japan) did not lead to superior results with regard to cell morphology, col-II protein production and col-II gene expression. With the CaReS collagen gel system the best quality of repair tissue was obtained by seeding freshly isolated chondrocytes.

  20. Hybrid Macro-Porous Titanium Ornamented by Degradable 3D Gel/nHA Micro-Scaffolds for Bone Tissue Regeneration

    PubMed Central

    Yin, Bo; Ma, Pei; Chen, Jun; Wang, Hai; Wu, Gui; Li, Bo; Li, Qiang; Huang, Zhifeng; Qiu, Guixing; Wu, Zhihong

    2016-01-01

    Porous titanium is a kind of promising material for bone substitution, while its bio-inert property results in demand of modifications to improve the osteointegration capacity. In this study, gelatin (Gel) and nano-hydroxyapatite (nHA) were used to construct 3D micro-scaffolds in the pores of porous titanium in the ratios of Gel:nHA = 1:0, Gel:nHA = 1:1, and Gel:nHA = 1:3, respectively. Cell attachment and proliferation, and gene and protein expression levels of osteogenic markers were evaluated in MC3T3-E1 cells, followed by bone regeneration assessment in a rabbit radius defect model. All hybrid scaffolds with different composition ratio were found to have significant promotional effects in cell adhesion, proliferation and differentiation, in which the group with Gel:nHA = 1:1 showed the best performance in vitro, as well as the most bone regeneration volume in vivo. This 3D micro-scaffolds modification may be an innovative method for porous titanium ornamentation and shows potential application values in clinic. PMID:27092492

  1. Mechanical and structural contribution of non-fibrillar matrix in uniaxial tension: a collagen-agarose co-gel model.

    PubMed

    Lake, Spencer P; Barocas, Victor H

    2011-07-01

    The mechanical role of non-fibrillar matrix and the nature of its interaction with the collagen network in soft tissues remain poorly understood, in part because of the lack of a simple experimental model system to quantify these interactions. This study's objective was to examine mechanical and structural properties of collagen-agarose co-gels, utilized as a simplified model system, to understand better the relationships between the collagen network and non-fibrillar matrix. We hypothesized that the presence of agarose would have a pronounced effect on microstructural reorganization and mechanical behavior. Samples fabricated from gel solutions containing 1.0 mg/mL collagen and 0, 0.125, or 0.25% w/v agarose were evaluated via scanning electron microscopy, incremental tensile stress-relaxation tests, and polarized light imaging. While the incorporation of agarose did not dramatically alter collagen network morphology, agarose led to concentration-dependent changes in mechanical and structural properties. Specifically, resistance of co-gels to volume change corresponded with differences in fiber reorientation and elastic/viscoelastic mechanics. Results demonstrate strong relationships between tissue properties and offer insight into behavior of tissues of varying Poisson's ratio and fiber kinematics. Results also suggest that non-fibrillar material may have significant effects on properties of artificial and native tissues even in tension, which is generally assumed to be collagen dominated.

  2. In vitro apatite forming ability of type I collagen hydrogels containing bioactive glass and silica sol-gel particles.

    PubMed

    Eglin, David; Maalheem, Sonia; Livage, Jacques; Coradin, Thibaud

    2006-02-01

    Type I collagen hydrogel containing bioactive glass (CaO-P2O5-SiO2) and silica sol-gel micrometric particles were prepared and their in vitroapatite-forming ability in simulated body fluid assessed. X-ray diffraction and scanning electron microscopy analysis showed that bioactive glass particles entrapment in collagen matrix did not inhibit calcium phosphate formation and induced morphology variations on the crystalline phase precipitated on the hydrogel surface. The silica--collagen hydrogel composite precipitated calcium phosphate whereas silica particles and collagen hydrogel alone did not, indicating a possible synergetic effect between collagen and silica on the apatite-forming ability. Mechanisms of calcium phosphate precipitation and its relevance to biomaterial development are discussed.

  3. Preparation and evaluation of mesalamine collagen in situ rectal gel: a novel therapeutic approach for treating ulcerative colitis.

    PubMed

    Ramadass, Satiesh Kumar; Perumal, Sathiamurthi; Jabaris, Sugin Lal; Madhan, Balaraman

    2013-01-23

    Ulcerative colitis (UC) is a chronic inflammatory disease that primarily affects the colonic mucosa. Mesalamine had been established as a first line drug for treating mild to moderate UC. A continued availability of the drug for treatment of damaged tissues remains a great challenge today. In the present study, a novel mesalamine collagen in situ gel has been prepared using type I collagen, which is pH/temperature sensitive. This hydrogel undergoes sol-gel transition under physiological pH and temperature which was confirmed by rheological studies. The in vitro release profile demonstrated sustained release of mesalamine over a period of 12h. The in vivo efficacy of the in situ gel was performed using dextran sodium sulphate induced ulcerative colitis model in BALB/c mice. The clinical parameters such as, body weight changes, rectal bleeding and stool consistency were evaluated. In addition, the histopathological investigation was conducted to assess severity of mucosal damage and inflammation infiltrate. There was a significant reduction in rectal bleeding and mucosal damage score for collagen-mesalamine in situ gel group compared to the reference group. Apart from releasing mesalamine in controlled manner, the strategy of administering mesalamine through collagen in situ gel facilitates regeneration of damaged mucosa resulting in a synergistic effect for the treatment of ulcerative colitis.

  4. 3D printing of soft and wet systems benefit from hard-to-soft transition of transparent shape memory gels (presentation video)

    NASA Astrophysics Data System (ADS)

    Furukawa, Hidemitsu; Gong, Jin; Makino, Masato; Kabir, Md. Hasnat

    2014-04-01

    Recently we successfully developed novel transparent shape memory gels. The SMG memorize their original shapes during the gelation process. In the room temperature, the SMG are elastic and show plasticity (yielding) under deformation. However when heated above about 50˚C, the SMG induce hard-to-soft transition and go back to their original shapes automatically. We focus on new soft and wet systems made of the SMG by 3-D printing technology.

  5. Epidermal growth factor promotes a mesenchymal over an amoeboid motility of MDA-MB-231 cells embedded within a 3D collagen matrix

    NASA Astrophysics Data System (ADS)

    Geum, Dongil T.; Kim, Beum Jun; Chang, Audrey E.; Hall, Matthew S.; Wu, Mingming

    2016-01-01

    The receptor of epidermal growth factor (EGFR) critically regulates tumor cell invasion and is a potent therapeutic target for treatment of many types of cancers, including carcinomas and glioblastomas. It is known that EGF regulates cell motility when tumor cells are embedded within a 3D biomatrix. However, roles of EGF in modulating tumor cell motility phenotype are largely unknown. In this article, we report that EGF promotes a mesenchymal over an amoeboid motility phenotype using a malignant breast tumor cell line, MDA-MB-231, embedded within a 3D collagen matrix. Amoeboid cells are rounded in shape, while mesenchymal cells are elongated, and their migrations are governed by a distinctly different set of biomolecules. Using single cell tracking analysis, we also show that EGF promotes cell dissemination through a significant increase in cell persistence along with a moderate increase of speed. The increase of persistence is correlated with the increase of the percentage of the mesenchymal cells within the population. Our work reveals a novel role of microenvironmental cue, EGF, in modulating heterogeneity and plasticity of tumor cell motility phenotype. In addition, it suggests a potential visual cue for diagnosing invasive states of breast cancer cells. This work can be easily extended beyond breast cancer cells.

  6. Effects of DS-modified agarose gels on neurite extension in 3D scaffold through mechanisms other than changing the pore radius of the gels.

    PubMed

    Peng, Jin; Pan, Qian; Zhang, Wei; Yang, Hao; Zhou, Xue; Jiang, Hua

    2014-07-01

    Dermatan sulfate is widely distributed as glycosaminoglycan side chains of proteoglycans, which are the main components of glial scar and inhibit neurite regeneration after nerve injury. However its role in the inhibiting process is not clear. Understanding neurite extension in three-dimensional scaffolds is critical for neural tissue engineering. This study used agarose gels modified with dermatan sulfate as the three-dimensional culture scaffold. We explored structure-function relationship between the three-dimensional scaffold and neurite extension and examined the role of dermatan sulfate on neurite extension in the three-dimensional scaffold. A range of agarose concentrations was used to generate varied gel physical structures and the corresponding neurite extension of embryonic day (E9) chick dorsal root ganglia was examined. We measured gel stiffness and gel pore size to determine whether dermatan sulfate changed the gels' conformation. As gel concentration increased, neurite length and gel pore size decreased, and gel stiffness increased. At 1.00 and 1.25% (wt/vol) concentrations, dermatan sulfates both immobilized with agarose gels and dissolved in culture medium inhibit neurite extension. While at 1.50 and 1.75% (wt/vol) concentrations, only immobilized dermatan sulfate worked. Immobilized dermatan sulfate could modify molecular shape of agarose gels, decrease gel pore size statistically, but did not influence gel stiffness. We have proved that the decrease of gel pore size is insufficient to inhibit neurite extension. These results indicate that dermatan sulfate inhibits neurite extension not through forming a mechanical barrier. Maybe its interaction with neuron membrane is the key factor in neurite extension.

  7. Preparation of a collagen/polymer hybrid gel designed for tissue membranes. Part I: controlling the polymer-collagen cross-linking process using an ethanol/water co-solvent.

    PubMed

    Nam, Kwangwoo; Kimura, Tsuyoshi; Funamoto, Seiichi; Kishida, Akio

    2010-02-01

    The drawback with collagen/2-methacryloyloxyethyl phosphorylcholine (MPC) polymer hybrid gels (collagen/phospholipid polymer hybrid gels) prepared in alkaline morpholinoethane sulfonic acid (MES) aqueous solution is that the cross-linking rate between the polymer and the collagen is low. To solve this problem, ethanol has been adopted as the reaction solvent, to prevent 1-ethyl-3-(3-dimethylaminopropyl)-1-carbodiimide hydrochloride (EDC) hydrolysis. Alterations in the ethanol mole concentration changed the cross-linking rate between the MPC polymer and the collagen gel. Prevention of EDC hydrolysis is clearly observed; protonation of carboxyl groups implies that the ratio of ethanol to water should be controlled. The polymer shows signs of penetration into the collagen gel layer, thus forming a totally homogeneous phase gel. This affects the mechanical strength of the collagen gel, making the gel much stiffer and brittle with an increase in the swelling ratio, as compared with that prepared in MES buffer. However, it is possible to obtain a collagen/phospholipid polymer hybrid gel with a high polymer portion and the cross-linking rate can be successfully controlled.

  8. Development of multifunctional collagen scaffolds directed by collagen mimetic peptides

    NASA Astrophysics Data System (ADS)

    Wang, Yi-Lan (Allen)

    Collagen is widely used for soft tissue replacement and tissue engineering scaffold. Functionalized collagen may offer new and improved applications for collagen-based biomaterials. But passively adsorbed molecules readily diffuse out from collagen matrix, and conventional chemical reactions on collagen are difficult to control and may compromise the biochemical feature of natural collagen. Hence, the aim of this dissertation is to develop a new physical collagen modification method through the non-covalent immobilization of collagen mimetic peptides (CMPs) and CMP derivatives on collagen scaffolds, thereby evading the drawbacks of passive and chemical modifications. Most of the research on CMPs over the past three decades has focused on synthesizing CMPs and understanding the effects of amino acid sequence on the peptide structural stability. Although few attempts have been made to develop biomaterials based on pure CMP, CMP has never used in complex with natural collagen. We demonstrate that CMPs with varying chain lengths have strong propensity to associate with natural 2-D and 3-D collagen substrates. We also show that CMPs can recognize and bind to reconstituted type I collagen fibers as well as collagens of ex vivo human liver tissue. The practical use of CMPs conjugated with linear and multi-arm poly(ethylene glycol)s allows to control cell organization in 2-D collagen substrates. Our cell adhesion studies suggest that under certain conditions (e.g. high incubation temperature, small CMP size), the bound CMP derivatives can be released from the collagen matrix, which may provide new opportunities for manipulating cell behavior especially by dynamically controlling the amount of signaling molecules in the collagen matrix. Polyanionic charged CMP was synthesized to modulate tubulogenesis of endothelial cells by attracting VEGF with 3-D collagen gel and a new PEG hydrogel using bifunctional CMP conjugates was synthesized as physico-chemical crosslinkers for

  9. Effects of tensile and compressive strains on response of a chondrocytic cell line embedded in type I collagen gel.

    PubMed

    Hirano, Yuji; Ishiguro, Naoki; Sokabe, Masahiro; Takigawa, Masaharu; Naruse, Keiji

    2008-01-20

    Tensile and compressive strains are commonly used in mechanobiological models. Here we report on the development of a novel three-dimensional cell-culture method, which allows both tensile and compressive loads to be applied. Preliminary results were obtained using HCS2/8 chondrocytic cells embedded in type I collagen gel. This construct was subjected to either 16% tension or 14% compression. Confocal laser scanning microscopy showed that both tension and compression caused significant cell deformation. The collagen gel-embedded HCS2/8 cells were subjected to static tension, dynamic tension, static compression or dynamic compression for 24h. Dynamic compression led to significantly decreased 5-bromo-2'-deoxyuridine incorporation compared with the control group. PCR analysis revealed upregulation of type II collagen caused by dynamic tension, upregulation of aggrecan caused by static compression, and downregulation of type II collagen and aggrecan caused by dynamic compression. Nitric oxide production was significantly increased by static tension and static compression compared with the control group. Our experimental system effectively applied several types of strain to HCS2/8 cells embedded in collagen gel. Our results suggest that the mode of mechanical strain affects the response of HCS2/8 cells.

  10. SU-E-CAMPUS-T-05: Validation of High-Resolution 3D Patient QA for Proton Pencil Beam Scanning and IMPT by Polymer Gel Dosimetry

    SciTech Connect

    Cardin, A; Avery, S; Ding, X; Kassaee, A; Lin, L; Maryanski, M

    2014-06-15

    Purpose: Validation of high-resolution 3D patient QA for proton pencil beam scanning and IMPT by polymer gel dosimetry. Methods: Four BANG3Pro polymer gel dosimeters (manufactured by MGS Research Inc, Madison, CT) were used for patient QA at the Robert's Proton Therapy Center (RPTC, Philadelphia, PA). All dosimeters were sealed in identical thin-wall Pyrex glass spheres. Each dosimeter contained a set of markers for 3D registration purposes. The dosimeters were mounted in a consistent and reproducible manner using a custom build holder. Two proton pencil beam scanning plans were designed using Varian Eclipse™ treatment planning system: 1) A two-field intensity modulated proton therapy (IMPT) plan and 2) one single field uniform dose (SFUD) plan. The IMPT fields were evaluated as a composite plan and individual fields, the SFUD plan was delivered as a single field plan.Laser CT scanning was performed using the manufacturer's OCTOPUS-IQ axial transmission laser CT scanner using a 1 mm slice thickness. 3D registration, analysis, and OD/cm to absorbed dose calibrations were perfomed using DICOM RT-Dose and CT files, and software developed by the manufacturer. 3D delta index, a metric equivalent to the gamma tool, was used for dose comparison. Results: Very good agreement with single IMPT fields and with SFUD was obtained. Composite IMPT fields had a less satisfactory agreement. The single fields had 3D delta index passing rates (3% dose difference, 3 mm DTA) of 98.98% and 94.91%. The composite 3D delta index passing rate was 80.80%. The SFUD passing rate was 93.77%. Required shifts of the dose distributions were less than 4 mm. Conclusion: A formulation of the BANG3Pro polymer gel dosimeter, suitable for 3D QA of proton patient plans is established and validated. Likewise, the mailed QA analysis service provided by the manufacturer is a practical option when required resources are unavailable. We fully disclose that the subject of this research regards a production

  11. TU-C-BRE-04: 3D Gel Dosimetry Using ViewRay On-Board MR Scanner: A Feasibility Study

    SciTech Connect

    Zhang, L; Du, D; Green, O; Rodriguez, V; Wooten, H; Xiao, Z; Yang, D; Hu, Y; Li, H

    2014-06-15

    Purpose: MR based 3D gel has been proposed for radiation therapy dosimetry. However, access to MR scanner has been one of the limiting factors for its wide acceptance. Recent commercialization of an on-board MR-IGRT device (ViewRay) may render the availability issue less of a concern. This work reports our attempts to simulate MR based dose measurement accuracy on ViewRay using three different gels. Methods: A spherical BANG gel dosimeter was purchased from MGS Research. Cylindrical MAGIC gel and Fricke gel were fabricated in-house according to published recipes. After irradiation, BANG and MAGIC were imaged using a dual-echo spin echo sequence for T2 measurement on a Philips 1.5T MR scanner, while Fricke gel was imaged using multiple spin echo sequences. Difference between MR measured and TPS calculated dose was defined as noise. The noise power spectrum was calculated and then simulated for the 0.35 T magnetic field associated with ViewRay. The estimated noise was then added to TG-119 test cases to simulate measured dose distributions. Simulated measurements were evaluated against TPS calculated doses using gamma analysis. Results: Given same gel, sequence and coil setup, with a FOV of 180×90×90 mm3, resolution of 3×3×3 mm3, and scanning time of 30 minutes, the simulated measured dose distribution using BANG would have a gamma passing rate greater than 90% (3%/3mm and absolute). With a FOV 180×90×90 mm3, resolution of 4×4×5 mm3, and scanning time of 45 minutes, the simulated measuremened dose distribution would have a gamma passing rate greater than 97%. MAGIC exhibited similar performance while Fricke gel was inferior due to much higher noise. Conclusions: The simulation results demonstrated that it may be feasible to use MAGIC and BANG gels for 3D dose verification using ViewRay low-field on-board MRI scanner.

  12. Tracer diffusion in a polymer gel: simulations of static and dynamic 3D networks using spherical boundary conditions.

    PubMed

    Kamerlin, Natasha; Elvingson, Christer

    2016-11-30

    We have investigated an alternative to the standard periodic boundary conditions for simulating the diffusion of tracer particles in a polymer gel by performing Brownian dynamics simulations using spherical boundary conditions. The gel network is constructed by randomly distributing tetravalent cross-linking nodes and connecting nearest pairs. The final gel structure is characterised by the radial distribution functions, chain lengths and end-to-end distances, and the pore size distribution. We have looked at the diffusion of tracer particles with a wide range of sizes, diffusing in both static and dynamic networks of two different volume fractions. It is quantitatively shown that the dynamical effect of the network becomes more important in facilitating the diffusional transport for larger particle sizes, and that one obtains a finite diffusion also for particle sizes well above the maximum in the pore size distribution.

  13. Deformation simulation of cells seeded on a collagen-GAG scaffold in a flow perfusion bioreactor using a sequential 3D CFD-elastostatics model.

    PubMed

    Jungreuthmayer, C; Jaasma, M J; Al-Munajjed, A A; Zanghellini, J; Kelly, D J; O'Brien, F J

    2009-05-01

    Tissue-engineered bone shows promise in meeting the huge demand for bone grafts caused by up to 4 million bone replacement procedures per year, worldwide. State-of-the-art bone tissue engineering strategies use flow perfusion bioreactors to apply biophysical stimuli to cells seeded on scaffolds and to grow tissue suitable for implantation into the patient's body. The aim of this study was to quantify the deformation of cells seeded on a collagen-GAG scaffold which was perfused by culture medium inside a flow perfusion bioreactor. Using a microCT scan of an unseeded collagen-GAG scaffold, a sequential 3D CFD-deformation model was developed. The wall shear stress and the hydrostatic wall pressure acting on the cells were computed through the use of a CFD simulation and fed into a linear elastostatics model in order to calculate the deformation of the cells. The model used numerically seeded cells of two common morphologies where cells are either attached flatly on the scaffold wall or bridging two struts of the scaffold. Our study showed that the displacement of the cells is primarily determined by the cell morphology. Although cells of both attachment profiles were subjected to the same mechanical load, cells bridging two struts experienced a deformation up to 500 times higher than cells only attached to one strut. As the scaffold's pore size determines both the mechanical load and the type of attachment, the design of an optimal scaffold must take into account the interplay of these two features and requires a design process that optimizes both parameters at the same time.

  14. A fibril-based structural constitutive theory reveals the dominant role of network characteristics on the mechanical behavior of fibroblast-compacted collagen gels.

    PubMed

    Feng, Zhonggang; Ishiguro, Yuki; Fujita, Kyohei; Kosawada, Tadashi; Nakamura, Takao; Sato, Daisuke; Kitajima, Tatsuo; Umezu, Mitsuo

    2015-10-01

    In this paper, we present a general, fibril-based structural constitutive theory which accounts for three material aspects of crosslinked filamentous materials: the single fibrillar force response, the fibrillar network model, and the effects of alterations to the fibrillar network. In the case of the single fibrillar response, we develop a formula that covers the entropic and enthalpic deformation regions, and introduce the relaxation phase to explain the observed force decay after crosslink breakage. For the filamentous network model, we characterize the constituent element of the fibrillar network in terms its end-to-end distance vector and its contour length, then decompose the vector orientation into an isotropic random term and a specific alignment, paving the way for an expanded formalism from principal deformation to general 3D deformation; and, more important, we define a critical core quantity over which macroscale mechanical characteristics can be integrated: the ratio of the initial end-to-end distance to the contour length (and its probability function). For network alterations, we quantitatively treat changes in constituent elements and relate these changes to the alteration of network characteristics. Singular in its physical rigor and clarity, this constitutive theory can reproduce and predict a wide range of nonlinear mechanical behavior in materials composed of a crosslinked filamentous network, including: stress relaxation (with dual relaxation coefficients as typically observed in soft tissues); hysteresis with decreasing maximum stress under serial cyclic loading; strain-stiffening under uniaxial tension; the rupture point of the structure as a whole; various effects of biaxial tensile loading; strain-stiffening under simple shearing; the so-called "negative normal stress" phenomenon; and enthalpic elastic behaviors of the constituent element. Applied to compacted collagen gels, the theory demonstrates that collagen fibrils behave as enthalpic

  15. Autocrine secretion of osteopontin by vascular smooth muscle cells regulates their adhesion to collagen gels.

    PubMed Central

    Weintraub, A. S.; Giachelli, C. M.; Krauss, R. S.; Almeida, M.; Taubman, M. B.

    1996-01-01

    Osteopontin (OPN) is a secreted protein postulated to facilitate vascular smooth muscle cell (VSMC) adhesion and migration. Rat aortic VSMC lines were isolated after infection with recombinant retroviruses harboring OPN sense and antisense constructs. All lines grew normally in monolayer culture. On three-dimensional collagen gels, normal VSMCs and lines containing sense constructs (n=15) or empty vector (n=10) attached to gel and invaded the matrix. Four of five antisense clones did not adhere or invade. Antisense clones had lower OPN levels after stimulation with angiotensin II than sense clones or clones containing the empty vector (antisense, 257+/-102 ng/ml; sense, 473+/-104; vector, 434+/-66). Non-adhering antisense clones had lower mean OPN levels after angiotensin II stimulation (161+/-47 ng/ml) than sense or antisense lines with normal adhesion (486+/-63 ng/ml). The ability to adhere correlated with OPN levels >250 ng/ml. Adhesion and invasion were fully restored with addition of 100 to 200 ng/ml of exogenous OPN and were inhibited in normal VSMCs by incubation with 1 microgram/ml anti-OPN antibody. The autocrine secretion of OPN appears to play an important role in VSMC adhesion, spreading, and invasion. Images Figure 2 Figure 3 Figure 5 Figure 6 Figure 7 PMID:8686750

  16. Construction of a 3D rGO-collagen hybrid scaffold for enhancement of the neural differentiation of mesenchymal stem cells

    NASA Astrophysics Data System (ADS)

    Guo, Weibo; Wang, Shu; Yu, Xin; Qiu, Jichuan; Li, Jianhua; Tang, Wei; Li, Zhou; Mou, Xiaoning; Liu, Hong; Wang, Zhonglin

    2016-01-01

    The cell-material interface is one of the most important considerations in designing a high-performance tissue engineering scaffold because the surface of the scaffold can determine the fate of stem cells. A conductive surface is required for a scaffold to direct stem cells toward neural differentiation. However, most conductive polymers are toxic and not amenable to biological degradation, which restricts the design of neural tissue engineering scaffolds. In this study, we used a bioactive three-dimensional (3D) porcine acellular dermal matrix (PADM), which is mainly composed of type I collagen, as a basic material and successfully assembled a layer of reduced graphene oxide (rGO) nanosheets on the surface of the PADM channels to obtain a porous 3D, biodegradable, conductive and biocompatible PADM-rGO hybrid neural tissue engineering scaffold. Compared with the PADM scaffold, assembling the rGO into the scaffold did not induce a significant change in the microstructure but endowed the PADM-rGO hybrid scaffold with good conductivity. A comparison of the neural differentiation of rat bone-marrow-derived mesenchymal stem cells (MSCs) was performed by culturing the MSCs on PADM and PADM-rGO scaffolds in neuronal culture medium, followed by the determination of gene expression and immunofluorescence staining. The results of both the gene expression and protein level assessments suggest that the rGO-assembled PADM scaffold may promote the differentiation of MSCs into neuronal cells with higher protein and gene expression levels after 7 days under neural differentiation conditions. This study demonstrated that the PADM-rGO hybrid scaffold is a promising scaffold for neural tissue engineering; this scaffold can not only support the growth of MSCs at a high proliferation rate but also enhance the differentiation of MSCs into neural cells.The cell-material interface is one of the most important considerations in designing a high-performance tissue engineering scaffold

  17. Sol-gel processed mupirocin silica microspheres loaded collagen scaffold: a synergistic bio-composite for wound healing.

    PubMed

    Perumal, Sathiamurthi; Ramadass, Satiesh kumar; Madhan, Balaraman

    2014-02-14

    Development of a bio-composite using synergistic combination is a promising strategy to address various pathological manifestations of acute and chronic wounds. In the present work, we have combined three materials viz., mupirocin as an antimicrobial drug, sol-gel processed silica microsphere as drug carrier for sustained delivery of drug and collagen, an established wound healer as scaffold. The mupirocin-loaded silica microspheres (Mu-SM) and Mu-SM loaded collagen scaffold were characterized for surface morphology, entrapment efficiency and distribution homogeneity, in vitro drug release, water uptake capacity, cell proliferation and antibacterial activity. In vivo wound healing efficacy of the bio-composite was experimented using full thickness excision wound model in Wistar albino rats. The Mu-SM incorporated collagen scaffold showed good in vitro characteristics in terms of better water uptake, sustained drug availability and antimicrobial activity. The wound closure analysis revealed that the complete epithelialisation was observed at 14.2 ± 0.44 days for Mu-SM loaded collagen, whereas this was 17.4 ± 0.44 days and 20.6 ± 0.54 days for collagen and control groups, respectively. Consequently, the synergistic strategy of combining mupirocin-loaded silica microspheres and collagen as a Mu-SM loaded collagen dressing material would be an ideal biomaterial for the treatment of surface wounds, burns and foot ulcers.

  18. Synthesis of chitin nanofibers, MWCNTs and MnO2 nanoflakes 3D porous network flexible gel-film for high supercapacitive performance electrodes

    NASA Astrophysics Data System (ADS)

    Liu, Shengnan; Li, Dagang

    2017-03-01

    As the porous structure and conductivity result in improvement of electrochemical properties, the chitin nanofibers (ChNFs), multi-walled carbon nanotubes (MWCNTs) and MnO2 (manganese dioxide) nanoflakes 3D porous network core-shell structure gel-film was fabricated for flexible free-standing supercapacitor electrodes. The electrodes were characterized by various techniques and the results demonstrate that the as-synthesized ChNFs/MWCNTs/MnO2 gel-film electrodes exhibits excellent supercapacitive behaviours. The ChNFs/MWCNTs/MnO2 gel-film electrode shows a high capacitance of 295.2 mF/cm2 at 0.1 mA/cm2 in 1 M Na2SO4 aqueous electrolyte because of its 3D porous structure. Furthermore, the electrodes also showed surprising cycling stability for 5000 cycles with retention rate up to 157.14% at 1 mA/cm2. The data presents great promise in the application of high-performance flexible supercapacitors with the low cost, light-weight and excellent cycling ability.

  19. Cell-generated forces influence the viability, metabolism and mechanical properties of fibroblast-seeded collagen gel constructs.

    PubMed

    Berry, Catherine C; Shelton, Julia C; Lee, David A

    2009-01-01

    The aim of this study was to investigate the influence of the endogenous forces generated by fibroblast-mediated contraction, using four individual collagen gel models that differed with respect to the ability of the cells to contract the gel. Human neonatal dermal fibroblasts were seeded in type I collagen and the gels were cast in a racetrack-shaped mould containing a removable central island. Two of the models were mechanically stressed (20 mm and 10 mm), as complete contraction was prevented by the presence of a central island. The central island was removed in the third model (released) and the final model was cast in a Petri dish and detached, allowing full multi-axial contraction (SR). Cell viability was maintained in the 10 mm, released and SR models over a 6 day culture period but localized regions of cell death were evident in the 20 mm model. Cell and collagen alignment was developed in the 20 mm and 10 mm models and to a lesser extent in the released model, but was absent in the SR model. Cell proliferation and collagen synthesis was lower in the 20 mm model compared to the other systems and there was evidence of enhanced matrix metalloproteinase production. The mechanical properties of the 20 mm model system were inferior to the 10 mm and released systems. The 10 mm model system induced a high level of cell and matrix orientation and may, therefore, represent the best option for tissue-engineered ligament repair involving an orientated fibroblast-seeded collagen gel.

  20. Influence on the physicochemical properties of fish collagen gels using self-assembly and simultaneous cross-linking with the N-hydroxysuccinimide adipic acid derivative.

    PubMed

    Shen, Lirui; Tian, Zhenhua; Liu, Wentao; Li, Guoying

    2015-06-01

    Collagen gels from Southern catfish (Silurus meridionalis Chen) skins were prepared via the self-assembly of collagen molecules and simultaneous cross-linking with the N-hydroxysuccinimide adipic acid derivative (NHS-AA). The doses of NHS-AA were converted to [NHS-AA]/[NH2] ratios (0.025-1.6, calculated by the [active ester group] of NHS-AA and [ε-NH2] of lysine and hydroxylysine residues of collagen). When the ratio < 0.05, collagen gels were formed by collagen molecule self-assembly, resulting in the opalescent appearance of collagen gels and the characteristic D-periodicity of partial collagen fibrils, the collagen gel ([NHS-AA]/[NH2] = 0.05) displayed a small increase in denaturation temperature (Td, 42.8 °C), remaining weight (12.59%), specific water content (SWC 233.7) and elastic modulus (G' 128.4 Pa) compared with uncross-linked collagen gel (39.1 °C, 9.12%, 222.4 and 85.4 Pa, respectively). As the ratio > 0.05, disappearance of D-periodicity and a gradual change in appearance from opalescent to transparent suggested that the inhibition of NHS-AA in the self-assembly of collagen molecules was more obvious. As a result, the collagen gel ([NHS-AA]/[NH2] = 0.2) had the lowest Td (35.8 °C), remaining weight (7.96%), SWC (130.9) and G' (31.9 Pa). When the ratio was 1.6, the collagen molecule self-assembly was markedly suppressed and the formation of collagen gel was predominantly via the covalent cross-linking bonds which led to the transparent appearance, and the maximum values of Td (47.0 °C), remaining weight (45.92%) and G' (420.7 Pa) of collagen gel. These results indicated that collagen gels with different properties can be prepared using different NHS-AA doses.

  1. Intestinal epithelial organoids fuse to form self-organizing tubes in floating collagen gels.

    PubMed

    Sachs, Norman; Tsukamoto, Yoshiyuki; Kujala, Pekka; Peters, Peter J; Clevers, Hans

    2017-03-15

    Multiple recent examples highlight how stem cells can self-organize in vitro to establish organoids that closely resemble their in vivo counterparts. Single Lgr5(+) mouse intestinal stem cells can be cultured under defined conditions forming ever-expanding epithelial organoids that retain cell polarization, cell type diversity and anatomical organization of the in vivo epithelium. Although exhibiting a remarkable level of self-organization, the so called 'mini-guts' have a closed cystic structure of microscopic size. Here, we describe a simple protocol to generate macroscopic intestinal tubes from small cystic organoids. Embedding proliferating organoids within a contracting floating collagen gel allows them to align and fuse to generate macroscopic hollow structures ('tubes') that are lined with a simple epithelium containing all major cell types (including functional stem cells) of the small intestine. Cells lining the central contiguous lumen closely resemble the epithelial cells on luminal villi in vivo, whereas buds that protrude from the main tube into the surrounding matrix closely resemble crypts. Thus, the remarkable self-organizing properties of Lgr5(+) stem cells extend beyond the level of the microscopic cystic organoid to the next, macroscopic, level of tube formation.

  2. Role of xenogenous bovine platelet gel embedded within collagen implant on tendon healing: an in vitro and in vivo study

    PubMed Central

    Oryan, Ahmad; Meimandi-Parizi, Abdolhamid; Maffulli, Nicola

    2015-01-01

    Surgical reconstruction of large Achilles tendon defects is demanding. Platelet concentrates may be useful to favor healing in such conditions. The characteristics of bovine platelet-gel embedded within a collagen-implant were determined in vitro, and its healing efficacy was examined in a large Achilles tendon defect in rabbits. Two cm of the left Achilles tendon of 60 rabbits were excised, and the animals were randomly assigned to control (no implant), collagen-implant, or bovine-platelet-gel-collagen-implant groups. The tendon edges were maintained aligned using a Kessler suture. No implant was inserted in the control group. In the two other groups, a collagen-implant or bovine-platelet-gel-collagen-implant was inserted in the defect. The bioelectricity and serum platelet-derived growth factor levels were measured weekly and at 60 days post injury, respectively. After euthanasia at 60 days post injury, the tendons were tested at macroscopic, microscopic, and ultrastructural levels, and their dry matter and biomechanical performances were also assessed. Another 60 rabbits were assigned to receive no implant, a collagen-implant, or a bovine-platelet-gel-collagen-implant, euthanized at 10, 20, 30, and 40 days post injury, and their tendons were evaluated grossly and histologically to determine host-graft interactions. Compared to the control and collagen-implant, treatment with bovine-platelet-gel-collagen-implant improved tissue bioelectricity and serum platelet-derived growth factor levels, and increased cell proliferation, differentiation, and maturation. It also increased number, diameter, and density of the collagen fibrils, alignment and maturation of the collagen fibrils and fibers, biomechanical properties and dry matter content of the injured tendons at 60 days post injury. The bovine-platelet-gel-collagen-implant also increased biodegradability, biocompatibility, and tissue incorporation behavior of the implant compared to the collagen-implant alone

  3. 3D-printed polylactic acid supports for enhanced ionization efficiency in desorption electrospray mass spectrometry analysis of liquid and gel samples.

    PubMed

    Elviri, Lisa; Foresti, Ruben; Bianchera, Annalisa; Silvestri, Marco; Bettini, Ruggero

    2016-08-01

    The potential of 3D printing technology was here exploited to prepare tailored polylactic acid (PLA) supports for desorption electrospray ionization (DESI) experiments. PLA rough solid supports presenting wells of different shape (i.e. cylindrical, cubic and hemispherical cavities) were designed to accommodate samples of different physical state. The potentials of such supports in terms of sample loading capacity, sensitivity, signal stability were tested by analysing a peptide (i.e. insulin) and an aminoglycoside antibiotic (i.e. gentamicin sulphate) from solution and a chitosan-based gel. The results obtained were compared with those obtained by using a traditional polytetrafluoroethylene (PTFE) support and discussed. By using PLA support on the flat side, signal intensity improved almost twice with respect to PTFE support, whereas with spherical wells a five times improved signal sensitivity and good stability (RSD<6%) were obtained for the analysis of two model molecules. Limits of detection were in the 3-10nM range and linearity was demonstrated for both analytes in the 0.05-0.5μM range for semi-quantitative or quantitative purposes. The use of a well and the set-up of optimal source parameters allowed the analysis of samples in a gel state with good precision (RSD<10%) and accuracy (86±6-102±9%), otherwise difficult to analyse on a flat smooth surface. These findings are of great interest and stimulus to exploit the advantages of 3D printing technology for the development of devices for a DESI source, presenting different shapes or configuration as a function of the sample types.

  4. Maintenance of a bone collagen phenotype by osteoblast-like cells in 3D periodic porous titanium (Ti-6Al-4 V) structures fabricated by selective electron beam melting.

    PubMed

    Hrabe, Nikolas W; Heinl, Peter; Bordia, Rajendra K; Körner, Carolin; Fernandes, Russell J

    2013-01-01

    Regular 3D periodic porous Ti-6Al-4 V structures were fabricated by the selective electron beam melting method (EBM) over a range of relative densities (0.17-0.40) and pore sizes (500-1500 µm). Structures were seeded with human osteoblast-like cells (SAOS-2) and cultured for four weeks. Cells multiplied within these structures and extracellular matrix collagen content increased. Type I and type V collagens typically synthesized by osteoblasts were deposited in the newly formed matrix with time in culture. High magnification scanning electron microscopy revealed cells attached to surfaces on the interior of the structures with an increasingly fibrous matrix. The in-vitro results demonstrate that the novel EBM-processed porous structures, designed to address the effect of stress-shielding, are conducive to osteoblast attachment, proliferation and deposition of a collagenous matrix characteristic of bone.

  5. 3D Viscoelastic traction force microscopy.

    PubMed

    Toyjanova, Jennet; Hannen, Erin; Bar-Kochba, Eyal; Darling, Eric M; Henann, David L; Franck, Christian

    2014-10-28

    Native cell-material interactions occur on materials differing in their structural composition, chemistry, and physical compliance. While the last two decades have shown the importance of traction forces during cell-material interactions, they have been almost exclusively presented on purely elastic in vitro materials. Yet, most bodily tissue materials exhibit some level of viscoelasticity, which could play an important role in how cells sense and transduce tractions. To expand the realm of cell traction measurements and to encompass all materials from elastic to viscoelastic, this paper presents a general, and comprehensive approach for quantifying 3D cell tractions in viscoelastic materials. This methodology includes the experimental characterization of the time-dependent material properties for any viscoelastic material with the subsequent mathematical implementation of the determined material model into a 3D traction force microscopy (3D TFM) framework. Utilizing this new 3D viscoelastic TFM (3D VTFM) approach, we quantify the influence of viscosity on the overall material traction calculations and quantify the error associated with omitting time-dependent material effects, as is the case for all other TFM formulations. We anticipate that the 3D VTFM technique will open up new avenues of cell-material investigations on even more physiologically relevant time-dependent materials including collagen and fibrin gels.

  6. In situ cell-matrix mechanics in tendon fascicles and seeded collagen gels: implications for the multiscale design of biomaterials.

    PubMed

    Duncan, Neil A; Bruehlmann, Sabina B; Hunter, Christopher J; Shao, Xinxin; Kelly, Elizabeth J

    2014-01-01

    Designing biomaterials to mimic and function within the complex mechanobiological conditions of connective tissues requires a detailed understanding of the micromechanical environment of the cell. The objective of our study was to measure the in situ cell-matrix strains from applied tension in both tendon fascicles and cell-seeded type I collagen scaffolds using laser scanning confocal microscopy techniques. Tendon fascicles and collagen gels were fluorescently labelled to simultaneously visualise the extracellular matrix and cell nuclei under applied tensile strains of 5%. There were significant differences observed in the micromechanics at the cell-matrix scale suggesting that the type I collagen scaffold did not replicate the pattern of native tendon strains. In particular, although the overall in situ tensile strains in the matrix were quite similar (∼2.5%) between the tendon fascicles and the collagen scaffolds, there were significant differences at the cell-matrix boundary with visible shear across cell nuclei of >1 μm measured in native tendon which was not observed at all in the collagen scaffolds. Similarly, there was significant non-uniformity of intercellular strains with relative sliding observed between cell rows in tendon which again was not observed in the collagen scaffolds where the strain environment was much more uniform. If the native micromechanical environment is not replicated in biomaterial scaffolds, then the cells may receive incorrect or mixed mechanical signals which could affect their biosynthetic response to mechanical load in tissue engineering applications. This study highlights the importance of considering the microscale mechanics in the design of biomaterial scaffolds and the need to incorporate such features in computational models of connective tissues.

  7. Preparation and characterization of malonic acid cross-linked chitosan and collagen 3D scaffolds: an approach on non-covalent interactions.

    PubMed

    Mitra, Tapas; Sailakshmi, G; Gnanamani, A; Mandal, A B

    2012-05-01

    The present study emphasizes the influence of non-covalent interactions on the mechanical and thermal properties of the scaffolds of chitosan/collagen origin. Malonic acid (MA), a bifuncitonal diacid was chosen to offer non-covalent cross-linking. Three dimensional scaffolds was prepared using chitosan at 1.0% (w/v) and MA at 0.2% (w/v), similarly collagen 0.5% (w/v) and MA 0.2% (w/v) and characterized. Results on FT-IR, TGA, DSC, SEM and mechanical properties (tensile strength, stiffness, Young's modulus, etc.) assessment demonstrated the existence of non-covalent interaction between MA and chitosan/collagen, which offered flexibility and high strength to the scaffolds suitable for tissue engineering research. Studies using NIH 3T3 fibroblast cells suggested biocompatibility nature of the scaffolds. Docking simulation study further supports the intermolecular hydrogen bonding interactions between MA and chitosan/collagen.

  8. Fast and mild strategy, using superhydrophobic surfaces, to produce collagen/platelet lysate gel beads for skin regeneration.

    PubMed

    Lima, Ana Catarina; Mano, João F; Concheiro, Angel; Alvarez-Lorenzo, Carmen

    2015-02-01

    Platelet lysate (PL) was encapsulated in collagen (Coll) millimetric gel beads, on biomimetic superhydrophobic surfaces, under mild conditions, with the aim of obtaining easy-to-handle formulations able to provide sustained release of multiple growth factors for skin ulcers treatment. The gel particles were prepared with various concentrations of PL incorporating or not stem cells, and tested as freshly prepared or after being freeze-dried or cryopreserved. Coll + PL particles were evaluated regarding degradation in collagenase-rich environment (simulating the aggressive environment of the chronic ulcers), sustained release of total protein, PDGF-BB and VEGF, cell proliferation (using particles as the only source of growth factors), scratch wound recovery and angiogenic capability. Compared to Coll solely particles, incorporation of PL notably enhanced cell proliferation (inside and outside gels) and favored scratch wound recovery and angiogenesis. Moreover, cell-laden gel particles containing PL notably improved cell proliferation and even migration of cells from one particle towards a neighbor one, which led to cell-cell contacts and the spontaneous formation of tissue layers in which the spherical gels were interconnected by the stem cells.

  9. Collagen Gel Contraction by Fibroblasts: The Role of Myosin 2 and Gravity Effects

    NASA Technical Reports Server (NTRS)

    Johnson-Wint, Barbara P.; Malouvier, Alexandre; Holton, Emily

    1996-01-01

    Several lines of evidence suggest that collagen organization by connective tissue cells is sensitive to force. For instance, in flight experiments on rats the collagen fibrils which were produced under weightlessness and which were immediately next to the tendon fibroblasts were shown to be oriented randomly around the cells while the older fibrils right next to these and which were produced under 1 G, were highly organized.

  10. SU-E-T-678: Response Calibration Using Electron Depth-Dose Data for MRI-Based 3D Polymer Gel Dosimetry

    SciTech Connect

    Watanabe, Y; Warmington, L; Gopishankar, N

    2015-06-15

    Purpose: To evaluate a calibration method using the depth-dose data of an electron beam for MRI-based polymer gel dosimetry. Methods: MAGAT was manufactured in-house to fill two 400mL-cylindrical phantoms and nine 22mL-glass vials. Phantom-A was irradiated along the cylinder axis with a 9MeV electron beam of 6 cm x 6 cm field size (FS). Phantom-B was irradiated with a 6MV photon beam of 3 cm x 3 cm FS by a 360-degree arc technique. Eight vials were irradiated in a water-bath to various doses with a 20 cm x 20 cm FS 6MV photon beam. All irradiated phantoms and one un-irradiated vial were scanned with a 3T MRI scanner to obtain the spin-spin relaxation rate (R2) distributions. By comparing the measured R2-to-depth data with the known depth-dose data for Phantom-A, R2-to-dose calibration data were obtained (e-beam method). Another calibration data were obtained from the 9 vials data (9-vial method). We tested two regression equations, i.e., third-order polynomial and tangent functions, and two dose normalization methods, i.e., one-point and two-point methods. Then, these two calibration methods were used to obtain the 3D dose distribution of Phantom-B and evaluated by comparing the measured data with the dose distribution from a treatment planning system. The comparison was made with gamma passing rate (2%/2mm criteria). Results: We did not observe a clear advantage of the e-beam method over the 9-vial method for the 3D dose comparison with the test case. Nevertheless, we found that the e-beam method required a smaller dose scaling for the dose comparison. Furthermore, the tangent function showed better data fitting than the polynomial function with smaller uncertainty of the estimated coefficients. Conclusions: Considering the overall superior performance, we recommend the e-beam method with the tangent function as the regression equation and one-point dose normalization for the MRI-based polymer gel dosimetry.

  11. The use of gel dosimetry to measure the 3D dose distribution of a 90Sr/90Y intravascular brachytherapy seed.

    PubMed

    Massillon-Jl, G; Minniti, R; Mitch, M G; Maryanski, M J; Soares, C G

    2009-03-21

    Absorbed dose distributions in 3D imparted by a single (90)Sr/(90)Y beta particle seed source of the type used for intravascular brachytherapy were investigated. A polymer gel dosimetry medium was used as a dosemeter and phantom, while a special high-resolution laser CT scanner with a spatial resolution of 100 microm in all dimensions was used to quantify the data. We have measured the radial dose function, g(L)(r), observing that g(L)(r) increases to a maximum value and then decreases as the distance from the seed increases. This is in good agreement with previous data obtained with radiochromic film and thermoluminescent dosemeters (TLDs), even if the TLDs underestimate the dose at distances very close to the seed. Contrary to the measurements, g(L)(r) calculated through Monte Carlo simulations and reported previously steadily decreases without a local maximum as a function of the distance from the seed. At distances less than 1.5 mm, differences of more than 20% are observed between the measurements and the Monte Carlo calculations. This difference could be due to a possible underestimation of the energy absorbed into the seed core and encapsulation in the Monte Carlo simulation, as a consequence of the unknown precise chemical composition of the core and its respective density for this seed. The results suggest that g(L)(r) can be measured very close to the seed with a relative uncertainty of about 1% to 2%. The dose distribution is isotropic only at distances greater than or equal to 2 mm from the seed and is almost symmetric, independent of the depth. This study indicates that polymer gel coupled with the special small format laser CT scanner are valid and accurate methods for measuring the dose distribution at distances close to an intravascular brachytherapy seed.

  12. On the correlation between continuum mechanics entities and cell activity in biological soft tissues: assessment of three possible criteria for cell-controlled fibre reorientation in collagen gels and collagenous tissues.

    PubMed

    Kroon, Martin

    2010-05-07

    The biomechanical behaviour of biological cells is of great importance in many physiological processes. One such process is the maintenance of fibrous networks, such as collagenous tissues. The activity of the fibre-producing cells in this type of tissue is very important, and a comprehensive material description needs to incorporate the activity of the cells. In biomechanics, continuum mechanics is often employed to describe deforming solids, and modelling can be much simplified if continuum mechanics entities, such as stress and strain, can be correlated with cell activity. To investigate this, a continuum mechanics framework is employed in which remodelling of a collagen gel is modelled. The remodelling is accomplished by fibroblasts, and the activity of the fibroblasts is linked to the continuum mechanics theory. The constitutive model for the collagen fabric is formulated in terms of a strain energy function, which includes a density function describing the distribution of the collagen fibre orientation. This density function evolves according to an evolution law, where fibroblasts reorient fibres towards the direction of increasing Cauchy stress, elastic deformation, or stiffness. The theoretical framework is applied to experimental results from collagen gels, where gels have undergone remodelling under both biaxial and uniaxial constraint. The analyses indicated that criteria 1 and 2 (Cauchy stress and elastic deformations) are able to predict the collagen fibre distribution after remodelling, whereas criterion 3 (current stiffness) is not. This conclusion is, however, tentative and pertains, strictly speaking, only to fibre remodelling processes, and may not be valid for other types of cell activities.

  13. Analysis of the contraction of fibroblast-collagen gels and the traction force of individual cells by a novel elementary structural model.

    PubMed

    Feng, Z; Wagatsuma, Y; Kobayashi, S; Kosawada, T; Sato, D; Nakamura, T; Kitajima, T; Umezu, M

    2013-01-01

    Based on the experimental data of the contraction ratio of fibroblast-collagen gels with different initial collagen concentrations and cell numbers, we analyzed the traction force exerted by individual cells through a novel elementary structural model. We postulate that the mechanical mechanism of the gel contraction is mainly because that populated cells apply traction force to some of the surrounding collagen fibrils with such proper length potential to be pulled straight so as to be able to sustain the traction force; this traction induce the cells moving closely to each other and consequently compact the fibrillar network; the bending force of the fibrils in turn resists the movement. By employing fiber packing theory for random fibrillar networks and network alteration theory, the bending force of collagen fibrils was deduced. The traction force exerted by individual fibroblasts in the gels was balanced by the bending force and the resistance from interstitial fluid since inertial force can be neglected. The maximum traction force per cell under free floating condition is in the range of 0.27-9.02 nN depending on the initial collagen concentration and populated cell number. The most important outcome of this study is that the traction force of individual cells dynamically varies under different gel conditions, whereas the adhesion force between cell and individual fibrils is relatively converging and stable.

  14. Variable angle-of-incidence polarization-sensitive optical coherence tomography: its use to study the 3D collagen structure of equine articular cartilage

    NASA Astrophysics Data System (ADS)

    Ugryumova, Nadya; Gangnus, Sergei V.; Matcher, Stephen J.

    2006-02-01

    Polarization-sensitive optical coherence tomography has been used to spatially map the birefringence of equine articular cartilage. The polar orientation of the collagen fibers relative to the plane of the joint surface must be taken into account if a quantitative measurement of true birefringence is required. Using a series of images taken at different angles of illumination, we determine the fiber polar angle and true birefringence at one site on a sample of equine cartilage, on the assumption that the fibers lie within the plane of imaging. We propose a more general method based on the extended Jones matrix formalism to determine both the polar and azimuthal orientation of the collagen fibers as well as the true birefringence as functions of depth.

  15. Gelatin-methacrylamide gel loaded with microspheres to deliver GDNF in bilayer collagen conduit promoting sciatic nerve growth.

    PubMed

    Zhuang, Hai; Bu, Shoushan; Hua, Lei; Darabi, Mohammad A; Cao, Xiaojian; Xing, Malcolm

    2016-01-01

    In this study, we fabricated glial cell-line derived neurotrophic factor (GDNF)-loaded microspheres, then seeded the microspheres in gelatin-methacrylamide hydrogel, which was finally integrated with the commercial bilayer collagen membrane (Bio-Gide(®)). The novel composite of nerve conduit was employed to bridge a 10 mm long sciatic nerve defect in a rat. GDNF-loaded gelatin microspheres had a smooth surface with an average diameter of 3.9±1.8 μm. Scanning electron microscopy showed that microspheres were uniformly distributed in both the GelMA gel and the layered structure. Using enzyme-linked immunosorbent assay, in vitro release studies (pH 7.4) of GDNF from microspheres exhibited an initial burst release during the first 3 days (18.0%±1.3%), and then, a prolonged-release profile extended to 32 days. However, in an acidic condition (pH 2.5), the initial release percentage of GDNF was up to 91.2%±0.9% within 4 hours and the cumulative release percentage of GDNF was 99.2%±0.2% at 48 hours. Then the composite conduct was implanted in a 10 mm critical defect gap of sciatic nerve in a rat. We found that the nerve was regenerated in both conduit and autograft (AG) groups. A combination of electrophysiological assessment and histomorphometry analysis of regenerated nerves showed that axonal regeneration and functional recovery in collagen tube filled with GDNF-loaded microspheres (GM + CT) group were similar to AG group (P>0.05). Most myelinated nerves were matured and arranged densely with a uniform structure of myelin in a neat pattern along the long axis in the AG and GM + CT groups, however, regenerated nerve was absent in the BLANK group, left the 10 mm gap empty after resection, and the nerve fiber exhibited a disordered arrangement in the collagen tube group. These results indicated that the hybrid system of bilayer collagen conduit and GDNF-loaded gelatin microspheres combined with gelatin-methacrylamide hydrogels could serve as a new biodegradable

  16. Gelatin-methacrylamide gel loaded with microspheres to deliver GDNF in bilayer collagen conduit promoting sciatic nerve growth

    PubMed Central

    Zhuang, Hai; Bu, Shoushan; Hua, Lei; Darabi, Mohammad A; Cao, Xiaojian; Xing, Malcolm

    2016-01-01

    In this study, we fabricated glial cell-line derived neurotrophic factor (GDNF)-loaded microspheres, then seeded the microspheres in gelatin-methacrylamide hydrogel, which was finally integrated with the commercial bilayer collagen membrane (Bio-Gide®). The novel composite of nerve conduit was employed to bridge a 10 mm long sciatic nerve defect in a rat. GDNF-loaded gelatin microspheres had a smooth surface with an average diameter of 3.9±1.8 μm. Scanning electron microscopy showed that microspheres were uniformly distributed in both the GelMA gel and the layered structure. Using enzyme-linked immunosorbent assay, in vitro release studies (pH 7.4) of GDNF from microspheres exhibited an initial burst release during the first 3 days (18.0%±1.3%), and then, a prolonged-release profile extended to 32 days. However, in an acidic condition (pH 2.5), the initial release percentage of GDNF was up to 91.2%±0.9% within 4 hours and the cumulative release percentage of GDNF was 99.2%±0.2% at 48 hours. Then the composite conduct was implanted in a 10 mm critical defect gap of sciatic nerve in a rat. We found that the nerve was regenerated in both conduit and autograft (AG) groups. A combination of electrophysiological assessment and histomorphometry analysis of regenerated nerves showed that axonal regeneration and functional recovery in collagen tube filled with GDNF-loaded microspheres (GM + CT) group were similar to AG group (P>0.05). Most myelinated nerves were matured and arranged densely with a uniform structure of myelin in a neat pattern along the long axis in the AG and GM + CT groups, however, regenerated nerve was absent in the BLANK group, left the 10 mm gap empty after resection, and the nerve fiber exhibited a disordered arrangement in the collagen tube group. These results indicated that the hybrid system of bilayer collagen conduit and GDNF-loaded gelatin microspheres combined with gelatin-methacrylamide hydrogels could serve as a new biodegradable

  17. Influence of external uniaxial cyclic strain on oriented fibroblast-seeded collagen gels.

    PubMed

    Berry, Catherine C; Shelton, Julia C; Bader, Dan L; Lee, David A

    2003-08-01

    This study investigates the influence of cyclic tensile strain, applied to fully contracted fibroblast-seeded collagen constructs. The constructs were preloaded to either 2 or 10 mN. The preloaded constructs were subsequently subjected to a further 10% cyclic strain (0-10%) at 1 Hz, using a triangular waveform, or were cultured in the preloaded state. In all cases cellular viability was maintained during the conditioning period. Cell proliferation was enhanced by the application of cyclic strain within constructs preloaded to both 2 and 10 mN. Collagen synthesis was enhanced by cyclic strain within constructs preloaded at 2 mN only. The profile of matrix metalloproteinase (MMP) expression, determined by zymography, was broadly similar in constructs preloaded at 2 mN with or without the application of cyclic strain. By contrast, constructs preloaded at 10 mN and subjected to cyclic strain expressed enhanced levels of staining for latent MMP-1, latent MMP-9, and both latent and active MMP-2, when compared with the other conditioning regimens. The structural stiffness of constructs preloaded at 2 mN and subjected to cyclic strain was enhanced compared with control specimens, reflecting the increase in collagen synthesis. By contrast, the initial failure loads for cyclically strained constructs preloaded at 10 mN were reduced, potentially because of enhanced catabolic activity.

  18. Synthesis, development, characterization and effectiveness of bovine pure platelet gel-collagen-polydioxanone bioactive graft on tendon healing.

    PubMed

    Moshiri, Ali; Oryan, Ahmad; Meimandi-Parizi, Abdolhamid

    2015-06-01

    Bovine platelet gel (BPG) is an accessible and cost-effective source of growth factors which may have a value in tendon regenerative medicine. We produced a collagen implant (CI) as a tendon proper, covered it with polydioxanone (PDS) sheath to simulate paratenon and finally embedded the BPG as an active source of growth factor within the bioimplant to test whether BPG would be able to accelerate and enhance tendon regeneration and repair. After in vitro characterization of the bioactive grafts, the grafts were implanted in rabbit large tendon defect model. Untreated tendons and tendons treated with either CI or CI-PDS were served as controls for the CI-PDS-BPG. The animals were investigated clinically, ultrasonographically and haematologically for 120 days. After euthanasia, dry matter content, water uptake and delivery characteristics and also gross morphological, histopathological and scanning electron microscopic features of the healing tendons were assessed. In vitro, the activated platelets in the scaffold, released their growth factors significantly more than the controls. BPG also increased cell viability, and enhanced cellular differentiation, maturation and proliferation inside the CI-PDS compared with the controls. In vivo, the BPG modulated inflammation, increased quality and rate of fibroplasia and produced a remodelled tendon that had significantly higher collagen content and superior collagen fibril and fibre differentiation than controls. Treatment also significantly improved tendon water uptake and delivery characteristics, animals' serum PDGF level, CI-PDS biocompatibility and biodegradability and reduced peritendinous adhesions, muscle fibrosis and atrophy. BPG was effective on tendon healing and CI-PDS-BPG may be a valuable bioscaffold in tendon reconstructive surgery.

  19. Synthesis, development, characterization and effectiveness of bovine pure platelet gel-collagen-polydioxanone bioactive graft on tendon healing

    PubMed Central

    Moshiri, Ali; Oryan, Ahmad; Meimandi-Parizi, Abdolhamid

    2015-01-01

    Bovine platelet gel (BPG) is an accessible and cost-effective source of growth factors which may have a value in tendon regenerative medicine. We produced a collagen implant (CI) as a tendon proper, covered it with polydioxanone (PDS) sheath to simulate paratenon and finally embedded the BPG as an active source of growth factor within the bioimplant to test whether BPG would be able to accelerate and enhance tendon regeneration and repair. After in vitro characterization of the bioactive grafts, the grafts were implanted in rabbit large tendon defect model. Untreated tendons and tendons treated with either CI or CI-PDS were served as controls for the CI-PDS-BPG. The animals were investigated clinically, ultrasonographically and haematologically for 120 days. After euthanasia, dry matter content, water uptake and delivery characteristics and also gross morphological, histopathological and scanning electron microscopic features of the healing tendons were assessed. In vitro, the activated platelets in the scaffold, released their growth factors significantly more than the controls. BPG also increased cell viability, and enhanced cellular differentiation, maturation and proliferation inside the CI-PDS compared with the controls. In vivo, the BPG modulated inflammation, increased quality and rate of fibroplasia and produced a remodelled tendon that had significantly higher collagen content and superior collagen fibril and fibre differentiation than controls. Treatment also significantly improved tendon water uptake and delivery characteristics, animals’ serum PDGF level, CI-PDS biocompatibility and biodegradability and reduced peritendinous adhesions, muscle fibrosis and atrophy. BPG was effective on tendon healing and CI-PDS-BPG may be a valuable bioscaffold in tendon reconstructive surgery. PMID:25702535

  20. Fabrication of type I collagen microcarrier using a microfluidic 3D T-junction device and its application for the quantitative analysis of cell-ECM interactions.

    PubMed

    Yoon, Junghyo; Kim, Jaehoon; Jeong, Hyo Eun; Sudo, Ryo; Park, Myung-Jin; Chung, Seok

    2016-08-26

    We presented a new quantitative analysis for cell and extracellular matrix (ECM) interactions, using cell-coated ECM hydrogel microbeads (hydrobeads) made of type I collagen. The hydrobeads can carry cells as three-dimensional spheroidal forms with an ECM inside, facilitating a direct interaction between the cells and ECM. The cells on hydrobeads do not have a hypoxic core, which opens the possibility for using as a cell microcarrier for bottom-up tissue reconstitution. This technique can utilize various types of cells, even MDA-MB-231 cells, which have weak cell-cell interactions and do not form spheroids in conventional spheroid culture methods. Morphological indices of the cell-coated hydrobead visually present cell-ECM interactions in a quantitative manner.

  1. Fabrication of injectable, cellular, anisotropic collagen tissue equivalents with modular fibrillar densities.

    PubMed

    Marelli, Benedetto; Ghezzi, Chiara E; James-Bhasin, Mark; Nazhat, Showan N

    2015-01-01

    Technological improvements in collagen gel fabrication are highly desirable as they may enable significant advances in the formation of tissue-equivalent biomaterials for regenerative medicine, three-dimensional (3D) in vitro tissue models, and injectable scaffolds for cell and drug delivery applications. Thus, strategies to modulate collagen gel fibrillar density and organization in the mesostructure have been pursued to fabricate collagenous matrices with extracellular matrix-like features. Herein, we introduce a robust and simple method, namely gel aspiration-ejection (GAE), to engineer 3D, anisotropic, cell seeded, injectable dense collagen (I-DC) gels with controllable fibrillar densities, without the use of crosslinking. GAE allows for the hybridization of collagen gels with bioactive agents for increased functionality and supports highly aligned homogenous cell seeding, thus providing I-DC gels with distinct properties when compared to isotropic DC gels of random fibrillar orientation. The hybridization of I-DC with anionic fibroin derived polypeptides resulted in the nucleation of carbonated hydroxyapatite within the aligned nanofibrillar network upon exposure to simulated body fluid, yielding a 3D, anisotropic, mineralized collagen matrix. In addition, I-DC gels accelerated the osteoblastic differentiation of seeded murine mesenchymal stem cells (m-MSCs) when exposed to osteogenic supplements, which resulted in the cell-mediated, bulk mineralization of the osteoid-like gels. In addition, and upon exposure to neuronal transdifferentiation medium, I-DC gels supported and accelerated the differentiation of m-MSCs toward neuronal cells. In conclusion, collagen GAE presents interesting opportunities in a number of fields spanning tissue engineering and regenerative medicine to drug and cell delivery.

  2. Liposomes encapsulating Aloe vera leaf gel extract significantly enhance proliferation and collagen synthesis in human skin cell lines.

    PubMed

    Takahashi, Makoto; Kitamoto, Dai; Asikin, Yonathan; Takara, Kensaku; Wada, Koji

    2009-01-01

    Aloe vela leaf gel extract (AGE) are widely used as cosmetic and pharmaceutical ingredients because of its versatile skin care properties. In order to enhance the bioavailability of AGE, liposomes encapsulating AGE were prepared and examined for their interfacial and biochemical properties. The liposomes prepared from a soybean lecithin (SLP-WHITE, 1.0 wt%) by the Bangham method gave relatively a good trapping efficiency up to the AGE concentration of 0.5 wt%. The stable liposomes were then prepared from 1.0 wt% of SLP-WHITE and different concentrations of AGE by the mechanochemical method using a homogenizer and microfluidizer. The liposomes obtained from 0.25 wt% of AGE were confirmed to be small unilamellar vesicles with a diameter of less than 200 nm, and remained well dispersed for at least two weeks. The obtained liposomes encapsulating AGE were further examined for the effects on proliferation and type I collagen synthesis in normal human neonatal skin fibroblasts, NB1RGB cells. Liposomal AGE clearly showed higher proliferation rate than that of AGE alone. In addition, compared to the control, liposomal AGE significantly increased the collagen synthesis by 23%, while AGE alone showed a small effect. Liposomal AGE was also assayed for the effect on proliferation in normal human epidermal keratinocytes, NHEK(F) cells. Interestingly, liposomal AGE fractions containing 4 and 20 microg/mL of the extract considerably increased the proliferation rate by 77% and 101%, respectively. In contrast, AGE alone fractions containing 4 and 20 microg/mL of the extract increased the rate by 41% and 60%, respectively. Accordingly, the bioavailability and skin care properties of AGE will be significantly enhanced by liposome encapsulation, and the present liposomal AGE should have a great potential as an effective skin care formulation.

  3. Agent-based modeling traction force mediated compaction of cell-populated collagen gels using physically realistic fibril mechanics.

    PubMed

    Reinhardt, James W; Gooch, Keith J

    2014-02-01

    Agent-based modeling was used to model collagen fibrils, composed of a string of nodes serially connected by links that act as Hookean springs. Bending mechanics are implemented as torsional springs that act upon each set of three serially connected nodes as a linear function of angular deflection about the central node. These fibrils were evaluated under conditions that simulated axial extension, simple three-point bending and an end-loaded cantilever. The deformation of fibrils under axial loading varied <0.001% from the analytical solution for linearly elastic fibrils. For fibrils between 100 μm and 200 μm in length experiencing small deflections, differences between simulated deflections and their analytical solutions were <1% for fibrils experiencing three-point bending and <7% for fibrils experiencing cantilever bending. When these new rules for fibril mechanics were introduced into a model that allowed for cross-linking of fibrils to form a network and the application of cell traction force, the fibrous network underwent macroscopic compaction and aligned between cells. Further, fibril density increased between cells to a greater extent than that observed macroscopically and appeared similar to matrical tracks that have been observed experimentally in cell-populated collagen gels. This behavior is consistent with observations in previous versions of the model that did not allow for the physically realistic simulation of fibril mechanics. The significance of the torsional spring constant value was then explored to determine its impact on remodeling of the simulated fibrous network. Although a stronger torsional spring constant reduced the degree of quantitative remodeling that occurred, the inclusion of torsional springs in the model was not necessary for the model to reproduce key qualitative aspects of remodeling, indicating that the presence of Hookean springs is essential for this behavior. These results suggest that traction force mediated matrix

  4. Role of dynamin in elongated cell migration in a 3D matrix.

    PubMed

    Lees, Justin G; Gorgani, Nick N; Ammit, Alaina J; McCluskey, Adam; Robinson, Phillip J; O'Neill, Geraldine M

    2015-03-01

    The use of 3-dimensional (3D) collagen gels has yielded new insights into the migratory behaviour of cancer cells. While the large GTPase dynamin has emerged as an important regulator of cancer cell migration and invasion under 2D conditions, its role in 3D migration is unclear. We have used a potent dynamin modulator, a bis-tyrphostin derivative, Ryngo® 1-23, to investigate the role of dynamin in 3D migration in 3 different cell lines. The compound specifically inhibits persistent, elongated 3D migration in U87MG and SMA-560 cells. Treated U87MG cells adopt a rounded morphology that is not due to apoptosis, loss of matrix metalloprotease activity or inhibition of clathrin-mediated endocytosis. Given that Ryngo 1-23 is known to regulate dynamin oligomerisation and actin dynamics at the leading edge, we analysed actin filament distribution. Ryngo 1-23 induced a switch in actin filament organization in 3D cultures resulting in the generation of multiple short actin-rich microspikes. Correlated with the change in actin filament distribution, cells displayed reduced collagen gel contraction. Since acto-myosin force transmission to the extra-cellular matrix underpins persistent, elongated migration, our results suggest that Ryngo 1-23 modulates this process in 3D migration via dynamin-mediated regulation of acto-myosin force transmission to the extra-cellular matrix.

  5. Gel electrophoretic studies of photochemical cross-linking of type I collagen with brominated 1,8-naphthalimide dyes and visible light

    NASA Astrophysics Data System (ADS)

    Judy, Millard M.; Fuh, L.; Matthews, James Lester; Lewis, David E.; Utecht, Ronald E.

    1994-09-01

    Insoluble Type I collagen from bovine Achilles tendon (Sigma C9879) was suspended in a 3 mM solution of the dye diEd66Br dissolved in Cremophor ELR (BASF) to give a molecular concentration ratio. Fifty-microliter aliquots in 5-mm-diameter wells were exposed to 458 J/cm2 (225 mW/cm2, 1800 sec) of 457.9-nm light from an argon ion laser; similar aliquots with and without dye were kept in the dark to serve as controls. Following pelleting of the collagen by centrifugation and 3x washing in phosphate-buffered saline, aliquots of light-treated and control sample pellets were (1) digested in collagenase (Sigma C9891) or (2) extracted in 0.5 M acetic acid, followed by centrifugative ultrafiltration (10-kd cutoff) in 0.01 M acetic acid. Aliquots of the supernatant of the acid-extracted collagen also were digested in pepsin. Electrophoretic protein migration in 8% to 25% gradient polyacrylamide gels following SDS solubilization disclosed numerous, densely packed, essentially contiguous protein bands. These studies indicate that the dye and light treatment of insoluble Type I collagen (1) results in cross-linking of collagen molecules and (2) does not denature the trimer conformation sufficiently to enable significant digestion by pepsin.

  6. Standardization of the CFU-GM assay: Advantages of plating a fixed number of CD34+ cells in collagen gels.

    PubMed

    Dobo, Irène; Pineau, Danielle; Robillard, Nelly; Geneviève, Frank; Piard, Nicole; Zandecki, Marc; Hermouet, Sylvie

    2003-10-01

    We investigated whether plating a stable amount of CD34(+) cells improves the CFU-GM assay. Data of CFU-GM assays performed with leukaphereses products in two transplant centers using a commercial collagen-based medium and unified CFU-GM scoring criteria were pooled and analyzed according to the numbers of CD34(+) cells plated. A first series of 113 CFU-GM assays was performed with a fixed number of mononuclear cells (i.e., a variable number of CD34(+) cells). In these cultures the CFU-GM/CD34 ratio varied according to the number of CD34(+) cells plated: median CFUGM/CD34 ratios were 1/6.2 to 1/6.6 for grafts containing <2% CD34(+) cells, vs. 1/10.2 for grafts containing > or =2% CD34(+) cells. The median CFU-GM/CD34 ratio also varied depending on pathology: 1/9.3 for multiple myeloma (MM), 1/6.8 for Hodgkin's disease (HD), 1/6.5 for non-Hodgkin lymphoma (NHL), and 1/4.5 for solid tumors (ST). A second series of 95 CFU-GM assays was performed with a fixed number of CD34(+) cells (220/ml). The range of median CFU-GM/CD34 ratios was narrowed to 1/7.0 to 1/5.2, and coefficients of variation for CFU-GM counts decreased by half to 38.1% (NHL), 36.1% (MM), 49.9% (HD), and 22.4% (ST). In addition, CFU-GM scoring was facilitated as the percentages of cultures with >50 CFU/GM/ml decreased from 6.7% to 43.8% when a variable number of CD34(+) cells was plated, to 4.5% to 16.7% when 220 CD34(+) cells/ml were plated. Hence, plating a fixed number of CD34(+) cells in collagen gels improves the CFU-GM assay by eliminating cell number-related variability and reducing pathology-related variability in colony growth.

  7. Controlling sensitivity and stability of ferrous-xylenol orange-gelatin 3D gel dosimeters by doping with phenanthroline-type ligands and glyoxal

    NASA Astrophysics Data System (ADS)

    Penev, Kalin I.; Mequanint, Kibret

    2013-03-01

    The ferrous-xylenol orange-gelatin (FXG) dosimeter is widely used for three-dimensional ionizing radiation field mapping through optical scanning. Upon irradiation, the ferrous iron (Fe2+) is oxidized to ferric iron (Fe3+), which forms an intensely coloured complex with xylenol orange (XO). XO also acts as a diffusion-limiting additive; however, its presence may cause rapid auto-oxidation of Fe2+ during storage and low stability of the dose response. In this work, phenanthroline-type ligands were added to FXG system in a bid to bind the ferrous iron in a stable complex and minimize the rate of the auto-oxidation, whereas glyoxal was used as a chemical cross-linker, aiming to minimize the ferric iron diffusion. It was found that addition of either 1,10-phenanthroline or 5-nitro-1,10-phenanthroline can improve the auto-oxidation behaviour of the gels. However, the initial background absorbance was slightly increased, and the sensitivity of the dosimeters was decreased. Doping with glyoxal led to a moderate decrease of the diffusion only in those gels that also contained a phenanthroline-type ligand, and did not affect the initial dose response. Glyoxal also afforded an extended period of stable background absorbance level after an initial period of bleaching of the gel. Following re-irradiation, most glyoxal-containing dosimeters showed an excellent linearity of the dose response, albeit at a decreased sensitivity. We recommend further testing of FXG dosimeters, doped with phenanthroline-type ligands and glyoxal as a means for controlling the dose response and improving the long-term storage properties of the gels and the potential for dose fractionation.

  8. Performance evaluation of an improved optical computed tomography polymer gel dosimeter system for 3D dose verification of static and dynamic phantom deliveries

    SciTech Connect

    Lopatiuk-Tirpak, O.; Langen, K. M.; Meeks, S. L.; Kupelian, P. A.; Zeidan, O. A.; Maryanski, M. J.

    2008-09-15

    The performance of a next-generation optical computed tomography scanner (OCTOPUS-5X) is characterized in the context of three-dimensional gel dosimetry. Large-volume (2.2 L), muscle-equivalent, radiation-sensitive polymer gel dosimeters (BANG-3) were used. Improvements in scanner design leading to shorter acquisition times are discussed. The spatial resolution, detectable absorbance range, and reproducibility are assessed. An efficient method for calibrating gel dosimeters using the depth-dose relationship is applied, with photon- and electron-based deliveries yielding equivalent results. A procedure involving a preirradiation scan was used to reduce the edge artifacts in reconstructed images, thereby increasing the useful cross-sectional area of the dosimeter by nearly a factor of 2. Dose distributions derived from optical density measurements using the calibration coefficient show good agreement with the treatment planning system simulations and radiographic film measurements. The feasibility of use for motion (four-dimensional) dosimetry is demonstrated on an example comparing dose distributions from static and dynamic delivery of a single-field photon plan. The capability to visualize three-dimensional dose distributions is also illustrated.

  9. Performance evaluation of an improved optical computed tomography polymer gel dosimeter system for 3D dose verification of static and dynamic phantom deliveries.

    PubMed

    Lopatiuk-Tirpak, O; Langen, K M; Meeks, S L; Kupelian, P A; Zeidan, O A; Maryanski, M J

    2008-09-01

    The performance of a next-generation optical computed tomography scanner (OCTOPUS-5X) is characterized in the context of three-dimensional gel dosimetry. Large-volume (2.2 L), muscle-equivalent, radiation-sensitive polymer gel dosimeters (BANG-3) were used. Improvements in scanner design leading to shorter acquisition times are discussed. The spatial resolution, detectable absorbance range, and reproducibility are assessed. An efficient method for calibrating gel dosimeters using the depth-dose relationship is applied, with photon- and electron-based deliveries yielding equivalent results. A procedure involving a preirradiation scan was used to reduce the edge artifacts in reconstructed images, thereby increasing the useful cross-sectional area of the dosimeter by nearly a factor of 2. Dose distributions derived from optical density measurements using the calibration coefficient show good agreement with the treatment planning system simulations and radiographic film measurements. The feasibility of use for motion (four-dimensional) dosimetry is demonstrated on an example comparing dose distributions from static and dynamic delivery of a single-field photon plan. The capability to visualize three-dimensional dose distributions is also illustrated.

  10. Sol-gel synthesis, phase composition, morphological and structural characterization of Ca10(PO4)6(OH)2: XRD, FTIR, SEM, 3D SEM and solid-state NMR studies

    NASA Astrophysics Data System (ADS)

    Kareiva, Simonas; Klimavicius, Vytautas; Momot, Aleksandr; Kausteklis, Jonas; Prichodko, Aleksandra; Dagys, Laurynas; Ivanauskas, Feliksas; Sakirzanovas, Simas; Balevicius, Vytautas; Kareiva, Aivaras

    2016-09-01

    Aqueous sol-gel chemistry route based on ammonium-hydrogen phosphate as the phosphorus precursor, calcium acetate monohydrate as source of calcium ions, and 1,2-ethylendiaminetetraacetic acid (EDTA), or 1,2-diaminocyclohexanetetracetic acid (DCTA), or tartaric acid (TA), or ethylene glycol (EG), or glycerol (GL) as complexing agents have been used to prepare calcium hydroxyapatite (Ca10(PO4)6(OH)2, CHAp). The phase transformations, composition, and structural changes in the polycrystalline samples were studied by infrared spectroscopy (FTIR), X-ray powder diffraction analysis (XRD), and scanning electron microscopy (SEM). The local short-range (nano- and mezo-) scale effects in CHAp were studied using solid-state NMR spectroscopy. The spatial 3D data from the SEM images of CHAp samples obtained by TA, EG and GL sol-gel routes were recovered for the first time to our knowledge.

  11. Collagen Extracted from Persian Gulf Squid Exhibits Anti-Cytotoxic Properties on Apple Pectic Treated Cells: Assessment in an In Vitro Bioassay Model

    PubMed Central

    DELPHI, Ladan; SEPEHRI, Houri; MOTEVASELI, Elaheh; KHORRAMIZADEH, Mohammad Reza

    2016-01-01

    Background: Collagen-based three-dimensional (3D) in vitro systems have been introduced to study the physiological states of cells. As a biomolecule, collagen is usually extracted from terrestrial animals whilst aquatic animals like squid contain large amounts of collagen. Methods: In order to make effective use of marine organisms, we selected Persian Gulf squid in 2015 to extract the required collagen. Then, a 3D culture system based on the extracted collagen was applied to investigate cellular mechanisms in a native microenvironment. The formed collagen gel was used to investigate the growth of MDA-MB-231 breast cancer cells as well as responses to pectic acid. Results: The results revealed that the extracted collagen contained α, ß and γ components with high water holding capacity. This collagen formed a gel-like structure, which could promote the proliferation of MDA-MB-231 breast cancer cells. The MDA-MB-231 cells’ viability in presence of pectic acid, demonstrating the cells’ behavior in a 3D culture system. Conclusion: It seems that the collagen extracted from squid skin has type I collagen properties. It might be used as a substrate in 3D cell culture systems. PMID:27928532

  12. Local 3D matrix microenvironment regulates cell migration through spatiotemporal dynamics of contractility-dependent adhesions

    PubMed Central

    Doyle, Andrew D.; Carvajal, Nicole; Jin, Albert; Matsumoto, Kazue; Yamada, Kenneth M.

    2015-01-01

    The physical properties of two-dimensional (2D) extracellular matrices (ECMs) modulate cell adhesion dynamics and motility, but little is known about the roles of local microenvironmental differences in three-dimensional (3D) ECMs. Here we generate 3D collagen gels of varying matrix microarchitectures to characterize their regulation of 3D adhesion dynamics and cell migration. ECMs containing bundled fibrils demonstrate enhanced local adhesion-scale stiffness and increased adhesion stability through balanced ECM/adhesion coupling, whereas highly pliable reticular matrices promote adhesion retraction. 3D adhesion dynamics are locally regulated by ECM rigidity together with integrin/ECM association and myosin II contractility. Unlike 2D migration, abrogating contractility stalls 3D migration regardless of ECM pore size. We find force is not required for clustering of activated integrins on 3D native collagen fibrils. We propose that efficient 3D migration requires local balancing of contractility with ECM stiffness to stabilize adhesions, which facilitates the detachment of activated integrins from ECM fibrils. PMID:26548801

  13. Local 3D matrix microenvironment regulates cell migration through spatiotemporal dynamics of contractility-dependent adhesions.

    PubMed

    Doyle, Andrew D; Carvajal, Nicole; Jin, Albert; Matsumoto, Kazue; Yamada, Kenneth M

    2015-11-09

    The physical properties of two-dimensional (2D) extracellular matrices (ECMs) modulate cell adhesion dynamics and motility, but little is known about the roles of local microenvironmental differences in three-dimensional (3D) ECMs. Here we generate 3D collagen gels of varying matrix microarchitectures to characterize their regulation of 3D adhesion dynamics and cell migration. ECMs containing bundled fibrils demonstrate enhanced local adhesion-scale stiffness and increased adhesion stability through balanced ECM/adhesion coupling, whereas highly pliable reticular matrices promote adhesion retraction. 3D adhesion dynamics are locally regulated by ECM rigidity together with integrin/ECM association and myosin II contractility. Unlike 2D migration, abrogating contractility stalls 3D migration regardless of ECM pore size. We find force is not required for clustering of activated integrins on 3D native collagen fibrils. We propose that efficient 3D migration requires local balancing of contractility with ECM stiffness to stabilize adhesions, which facilitates the detachment of activated integrins from ECM fibrils.

  14. Local 3D matrix microenvironment regulates cell migration through spatiotemporal dynamics of contractility-dependent adhesions

    NASA Astrophysics Data System (ADS)

    Doyle, Andrew D.; Carvajal, Nicole; Jin, Albert; Matsumoto, Kazue; Yamada, Kenneth M.

    2015-11-01

    The physical properties of two-dimensional (2D) extracellular matrices (ECMs) modulate cell adhesion dynamics and motility, but little is known about the roles of local microenvironmental differences in three-dimensional (3D) ECMs. Here we generate 3D collagen gels of varying matrix microarchitectures to characterize their regulation of 3D adhesion dynamics and cell migration. ECMs containing bundled fibrils demonstrate enhanced local adhesion-scale stiffness and increased adhesion stability through balanced ECM/adhesion coupling, whereas highly pliable reticular matrices promote adhesion retraction. 3D adhesion dynamics are locally regulated by ECM rigidity together with integrin/ECM association and myosin II contractility. Unlike 2D migration, abrogating contractility stalls 3D migration regardless of ECM pore size. We find force is not required for clustering of activated integrins on 3D native collagen fibrils. We propose that efficient 3D migration requires local balancing of contractility with ECM stiffness to stabilize adhesions, which facilitates the detachment of activated integrins from ECM fibrils.

  15. Nonpolarized signaling reveals two distinct modes of 3D cell migration.

    PubMed

    Petrie, Ryan J; Gavara, Núria; Chadwick, Richard S; Yamada, Kenneth M

    2012-04-30

    We search in this paper for context-specific modes of three-dimensional (3D) cell migration using imaging for phosphatidylinositol (3,4,5)-trisphosphate (PIP3) and active Rac1 and Cdc42 in primary fibroblasts migrating within different 3D environments. In 3D collagen, PIP3 and active Rac1 and Cdc42 were targeted to the leading edge, consistent with lamellipodia-based migration. In contrast, elongated cells migrating inside dermal explants and the cell-derived matrix (CDM) formed blunt, cylindrical protrusions, termed lobopodia, and Rac1, Cdc42, and PIP3 signaling was nonpolarized. Reducing RhoA, Rho-associated protein kinase (ROCK), or myosin II activity switched the cells to lamellipodia-based 3D migration. These modes of 3D migration were regulated by matrix physical properties. Specifically, experimentally modifying the elasticity of the CDM or collagen gels established that nonlinear elasticity supported lamellipodia-based migration, whereas linear elasticity switched cells to lobopodia-based migration. Thus, the relative polarization of intracellular signaling identifies two distinct modes of 3D cell migration governed intrinsically by RhoA, ROCK, and myosin II and extrinsically by the elastic behavior of the 3D extracellular matrix.

  16. The influence of particle size and static magnetic fields on the uptake of magnetic nanoparticles into three dimensional cell-seeded collagen gel cultures.

    PubMed

    Lewis, Emily E L; Child, Hannah W; Hursthouse, Andrew; Stirling, David; McCully, Mark; Paterson, David; Mullin, Margaret; Berry, Catherine C

    2015-08-01

    Over recent decades there has been and continues to be major advances in the imaging, diagnosis and potential treatment of medical conditions, by the use of magnetic nanoparticles. However, to date the majority of cell delivery studies employ a traditional 2D monolayer culture. This article aims to determine the ability of various sized magnetic nanoparticles to penetrate and travel through a cell seeded collagen gel model, in the presence or absence of a magnetic field. Three different sized (100, 200, and 500 nm) nanoparticles were employed in the study. The results showed cell viability was unaffected by the presence of nanoparticles over a 24-h test period. The initial uptake of the 100 nm nanoparticle into the collagen gel structure was superior compared to the larger sized nanoparticles under the influence of a magnetic field and incubated for 24 h. Interestingly, it was the 200 nm nanoparticles, which proved to penetrate the gel furthest, under the influence of a magnetic field, during the initial culture stage after 1-h incubation.

  17. A novel injectable thermoresponsive and cytocompatible gel of poly(N-isopropylacrylamide) with layered double hydroxides facilitates siRNA delivery into chondrocytes in 3D culture.

    PubMed

    Yang, Hsiao-yin; van Ee, Renz J; Timmer, Klaas; Craenmehr, Eric G M; Huang, Julie H; Öner, F Cumhur; Dhert, Wouter J A; Kragten, Angela H M; Willems, Nicole; Grinwis, Guy C M; Tryfonidou, Marianna A; Papen-Botterhuis, Nicole E; Creemers, Laura B

    2015-09-01

    Hybrid hydrogels composed of poly(N-isopropylacrylamide) (pNIPAAM) and layered double hydroxides (LDHs) are presented in this study as novel injectable and thermoresponsive materials for siRNA delivery, which could specifically target several negative regulators of tissue homeostasis in cartilaginous tissues. Effectiveness of siRNA transfection using pNIPAAM formulated with either MgAl-LDH or MgFe-LDH platelets was investigated using osteoarthritic chondrocytes. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as an endogenous model gene to evaluate the extent of silencing. No significant adverse effects of pNIPAAM/LDH hydrogels on cell viability were noticed. Cellular uptake of fluorescently labeled siRNA was greatly enhanced (>75%) in pNIPAAM/LDH hydrogel constructs compared to alginate, hyaluronan and fibrin gels, and was absent in pNIPAAM hydrogel without LDH platelets. When using siRNA against GAPDH, 82-98% reduction of gene expression was found in both types of pNIPAAM/LDH hydrogel constructs after 6 days of culturing. In the pNIPAAM/MgAl-LDH hybrid hydrogel, 80-95% of GAPDH enzyme activity was reduced in parallel with gene. Our findings show that the combination of a cytocompatible hydrogel and therapeutic RNA oligonucleotides is feasible. Thus it might hold promise in treating degeneration of cartilaginous tissues by providing supporting scaffolds for cells and interference with locally produced degenerative factors.

  18. Layer-by-layer Collagen Deposition in Microfluidic Devices for Microtissue Stabilization

    PubMed Central

    McCarty, William J.; Prodanov, Ljupcho; Bale, Shyam Sundhar; Bhushan, Abhinav; Jindal, Rohit; Yarmush, Martin L.; Usta, O. Berk

    2016-01-01

    Although microfluidics provides exquisite control of the cellular microenvironment, culturing cells within microfluidic devices can be challenging. 3D culture of cells in collagen type I gels helps to stabilize cell morphology and function, which is necessary for creating microfluidic tissue models in microdevices. Translating traditional 3D culture techniques for tissue culture plates to microfluidic devices is often difficult because of the limited channel dimensions. In this method, we describe a technique for modifying native type I collagen to generate polycationic and polyanionic collagen solutions that can be used with layer-by-layer deposition to create ultrathin collagen assemblies on top of cells cultured in microfluidic devices. These thin collagen layers stabilize cell morphology and function, as shown using primary hepatocytes as an example cell, allowing for the long term culture of microtissues in microfluidic devices. PMID:26485274

  19. Vascularization of Three-Dimensional Collagen Hydrogels Using Ultrasound Standing Wave Fields

    PubMed Central

    Garvin, Kelley A.; Dalecki, Diane; Hocking, Denise C.

    2011-01-01

    The successful fabrication of large, three-dimensional (3D) tissues and organs in vitro requires the rapid development of a vascular network to maintain cell viability and tissue function. In this study, we utilized an application of ultrasound standing wave field (USWF) technology to vascularize 3D, collagen-based hydrogels in vitro. Acoustic radiation forces associated with USWF were used to non-invasively organize human endothelial cells into distinct, multicellular planar bands within 3D collagen gels. The formation and maturation of capillary-like endothelial cell sprouts was monitored over time and compared to sham-exposed collagen constructs which were characterized by a homogeneous cell distribution. USWF10 induced cell banding accelerated the formation and elongation of capillary-like sprouts, promoted collagen fiber alignment, and resulted in the maturation of endothelial cell sprouts into lumen-containing, anastomosing networks found throughout the entire volume of the collagen gel. USWF-induced endothelial cell networks contained large, arteriole-sized lumen areas that branched into smaller, capillary-sized structures indicating the development of vascular tree-like networks. In contrast, sprout formation was delayed in sham-exposed collagen gels, and endothelial cell networks were absent from sham gel centers and failed to develop into the vascular tree-like structures found in USWF-exposed constructs. Our results demonstrate that USWF technology leads to rapid and extensive vascularization of 3D collagen-based engineered tissue and therefore, provides a new strategy to vascularize engineered tissues in vitro. PMID:21924816

  20. Primary cilium mechanotransduction of tensile strain in 3D culture: Finite element analyses of strain amplification caused by tensile strain applied to a primary cilium embedded in a collagen matrix.

    PubMed

    Mathieu, Pattie S; Bodle, Josephine C; Loboa, Elizabeth G

    2014-06-27

    Human adipose-derived stem cells (hASC) exhibit multilineage differentiation potential with lineage specification that is dictated by both the chemical and mechanical stimuli to which they are exposed. We have previously shown that 10% cyclic tensile strain increases hASC osteogenesis and cell-mediated calcium accretion. We have also recently shown that primary cilia are present on hASC and that chemically-induced lineage specification of hASC concurrently results in length and conformation changes of the primary cilia. Further, we have observed cilia length changes in hASC cultured within a collagen I gel in response to 10% cyclic tensile strain. We therefore hypothesize that primary cilia may play a key mechanotransduction role for hASC exposed to tensile strain. The goal of this study was to use finite element analysis (FEA) to determine strains occurring within the ciliary membrane in response to 10% tensile strain applied parallel, or perpendicular, to cilia orientation. To elucidate the mechanical environment experienced by the cilium, several lengths were modeled and evaluated based on cilia lengths measured on hASC grown under varied culture conditions. Principal tensile strains in both hASC and ciliary membranes were calculated using FEA, and the magnitude and location of maximum principal tensile strain determined. We found that maximum principal tensile strain was concentrated at the base of the cilium. In the linear elastic model, applying strain perpendicular to the cilium resulted in maximum strains within the ciliary membrane from 150% to 200%, while applying strain parallel to the cilium resulted in much higher strains, approximately 400%. In the hyperelastic model, applying strain perpendicular to the cilium resulted in maximum strains within the ciliary membrane around 30%, while applying strain parallel to the cilium resulted in much higher strains ranging from 50% to 70%. Interestingly, FEA results indicated that primary cilium length was not

  1. Primary Cilium Mechanotransduction of Tensile Strain in 3D Culture: Finite Element Analyses of Strain Amplification Caused by 10% Tensile Strain Applied to a Primary Cilium Embedded in a Collagen Matrix

    PubMed Central

    Mathieu, Pattie S.; Bodle, Josephine C.; Loboa, Elizabeth G.

    2014-01-01

    Human adipose-derived stem cells (hASC) exhibit multilineage differentiation potential with lineage specification that is dictated by both the chemical and mechanical stimuli to which they are exposed. We have previously shown that 10% cyclic tensile strain increases hASC osteogenesis and cell-mediated calcium accretion. We have also recently shown that primary cilia are present on hASC and that chemically-induced lineage specification of hASC concurrently results in length and conformation changes of the primary cilia. Further, we have observed cilia length changes on hASC cultured within a collagen I gel in response to 10% cyclic tensile strain. We therefore hypothesize that primary cilia may play a key mechanotransduction role for hASC exposed to tensile strain. The goal of this study was to use finite element analysis (FEA) to determine strains occurring within the ciliary membrane in response to 10% tensile strain applied parallel, or perpendicular, to cilia orientation. To elucidate the mechanical environment experienced by the cilium, several lengths were modeled and evaluated based on cilia lengths measured on hASC grown under varied culture conditions. Principal tensile strains in both hASC and ciliary membranes were calculated using FEA, and the magnitude and location of maximum principal tensile strain determined. We found that maximum principal tensile strain was concentrated at the base of the cilium. In the linear elastic model, applying strain perpendicular to the cilium resulted in maximum strains within the ciliary membrane from 150 to 200%, while applying strain parallel to the cilium resulted in much higher strains, approximately 400%. In the hyperelastic model, applying strain perpendicular to the cilium resulted in maximum strains within the ciliary membrane around 30%, while applying strain parallel to the cilium resulted in much higher strains ranging from 50% to 70% . Interestingly, FEA results indicated that primary cilium length was not

  2. Development of a 3D cell printed construct considering angiogenesis for liver tissue engineering.

    PubMed

    Lee, Jin Woo; Choi, Yeong-Jin; Yong, Woon-Jae; Pati, Falguni; Shim, Jin-Hyung; Kang, Kyung Shin; Kang, In-Hye; Park, Jaesung; Cho, Dong-Woo

    2016-01-12

    Several studies have focused on the regeneration of liver tissue in a two-dimensional (2D) planar environment, whereas actual liver tissue is three-dimensional (3D). Cell printing technology has been successfully utilized for building 3D structures; however, the poor mechanical properties of cell-laden hydrogels are a major concern. Here, we demonstrate the printing of a 3D cell-laden construct and its application to liver tissue engineering using 3D cell printing technology through a multi-head tissue/organ building system. Polycaprolactone (PCL) was used as a framework material because of its excellent mechanical properties. Collagen bioink containing three different types of cells-hepatocytes (HCs), human umbilical vein endothelial cells , and human lung fibroblasts--was infused into the canals of a PCL framework to induce the formation of capillary--like networks and liver cell growth. A co-cultured 3D microenvironment of the three types of cells was successfully established and maintained. The vascular formation and functional abilities of HCs (i.e., albumin secretion and urea synthesis) demonstrated that the heterotypic interaction among HCs and nonparenchymal cells increased the survivability and functionality of HCs within the collagen gel. Therefore, our results demonstrate the prospect of using cell printing technology for the creation of heterotypic cellular interaction within a structure for liver tissue engineering.

  3. Using a 3D Culture System to Differentiate Visceral Adipocytes In Vitro.

    PubMed

    Emont, Margo P; Yu, Hui; Jun, Heejin; Hong, Xiaowei; Maganti, Nenita; Stegemann, Jan P; Wu, Jun

    2015-12-01

    It has long been recognized that body fat distribution and regional adiposity play a major role in the control of metabolic homeostasis. However, the ability to study and compare the cell autonomous regulation and response of adipocytes from different fat depots has been hampered by the difficulty of inducing preadipocytes isolated from the visceral depot to differentiate into mature adipocytes in culture. Here, we present an easily created 3-dimensional (3D) culture system that can be used to differentiate preadipocytes from the visceral depot as robustly as those from the sc depot. The cells differentiated in these 3D collagen gels are mature adipocytes that retain depot-specific characteristics, as determined by imaging, gene expression, and functional assays. This 3D culture system therefore allows for study of the development and function of adipocytes from both depots in vitro and may ultimately lead to a greater understanding of site-specific functional differences of adipose tissues to metabolic dysregulation.

  4. Effects of Matrix Alignment and Mechanical Constraints on Cellular Behavior in 3D Engineered Microtissues

    NASA Astrophysics Data System (ADS)

    Bose, Prasenjit; Eyckmans, Jeroen; Chen, Christopher; Reich, Daniel

    The adhesion of cells to the extracellular matrix (ECM) plays a crucial role in a variety of cellular functions. The main building blocks of the ECM are 3D networks of fibrous proteins whose structure and alignments varies with tissue type. However, the impact of ECM alignment on cellular behaviors such as cell adhesion, spreading, extension and mechanics remains poorly understood. We present results on the development of a microtissue-based system that enables control of the structure, orientation, and degree of fibrillar alignment in 3D fibroblast-populated collagen gels. The tissues self-assemble from cell-laden collagen gels placed in micro-fabricated wells containing sets of elastic pillars. The contractile action of the cells leads to controlled alignment of the fibrous collagen, depending on the number and location of the pillars in each well. The pillars are elastic, and are utilized to measure the contractile forces of the microtissues, and by incorporating magnetic material in selected pillars, time-varying forces can be applied to the tissues for dynamic stimulation and measurement of mechanical properties. Results on the effects of varying pillar shape, spacing, location, and stiffness on microtissue organization and contractility will be presented. This work is supported by NSF CMMI-1463011.

  5. Europeana and 3D

    NASA Astrophysics Data System (ADS)

    Pletinckx, D.

    2011-09-01

    The current 3D hype creates a lot of interest in 3D. People go to 3D movies, but are we ready to use 3D in our homes, in our offices, in our communication? Are we ready to deliver real 3D to a general public and use interactive 3D in a meaningful way to enjoy, learn, communicate? The CARARE project is realising this for the moment in the domain of monuments and archaeology, so that real 3D of archaeological sites and European monuments will be available to the general public by 2012. There are several aspects to this endeavour. First of all is the technical aspect of flawlessly delivering 3D content over all platforms and operating systems, without installing software. We have currently a working solution in PDF, but HTML5 will probably be the future. Secondly, there is still little knowledge on how to create 3D learning objects, 3D tourist information or 3D scholarly communication. We are still in a prototype phase when it comes to integrate 3D objects in physical or virtual museums. Nevertheless, Europeana has a tremendous potential as a multi-facetted virtual museum. Finally, 3D has a large potential to act as a hub of information, linking to related 2D imagery, texts, video, sound. We describe how to create such rich, explorable 3D objects that can be used intuitively by the generic Europeana user and what metadata is needed to support the semantic linking.

  6. Macrophage podosomes go 3D.

    PubMed

    Van Goethem, Emeline; Guiet, Romain; Balor, Stéphanie; Charrière, Guillaume M; Poincloux, Renaud; Labrousse, Arnaud; Maridonneau-Parini, Isabelle; Le Cabec, Véronique

    2011-01-01

    Macrophage tissue infiltration is a critical step in the immune response against microorganisms and is also associated with disease progression in chronic inflammation and cancer. Macrophages are constitutively equipped with specialized structures called podosomes dedicated to extracellular matrix (ECM) degradation. We recently reported that these structures play a critical role in trans-matrix mesenchymal migration mode, a protease-dependent mechanism. Podosome molecular components and their ECM-degrading activity have been extensively studied in two dimensions (2D), but yet very little is known about their fate in three-dimensional (3D) environments. Therefore, localization of podosome markers and proteolytic activity were carefully examined in human macrophages performing mesenchymal migration. Using our gelled collagen I 3D matrix model to obligate human macrophages to perform mesenchymal migration, classical podosome markers including talin, paxillin, vinculin, gelsolin, cortactin were found to accumulate at the tip of F-actin-rich cell protrusions together with β1 integrin and CD44 but not β2 integrin. Macrophage proteolytic activity was observed at podosome-like protrusion sites using confocal fluorescence microscopy and electron microscopy. The formation of migration tunnels by macrophages inside the matrix was accomplished by degradation, engulfment and mechanic compaction of the matrix. In addition, videomicroscopy revealed that 3D F-actin-rich protrusions of migrating macrophages were as dynamic as their 2D counterparts. Overall, the specifications of 3D podosomes resembled those of 2D podosome rosettes rather than those of individual podosomes. This observation was further supported by the aspect of 3D podosomes in fibroblasts expressing Hck, a master regulator of podosome rosettes in macrophages. In conclusion, human macrophage podosomes go 3D and take the shape of spherical podosome rosettes when the cells perform mesenchymal migration. This work

  7. 3d-3d correspondence revisited

    DOE PAGES

    Chung, Hee -Joong; Dimofte, Tudor; Gukov, Sergei; ...

    2016-04-21

    In fivebrane compactifications on 3-manifolds, we point out the importance of all flat connections in the proper definition of the effective 3d N = 2 theory. The Lagrangians of some theories with the desired properties can be constructed with the help of homological knot invariants that categorify colored Jones polynomials. Higgsing the full 3d theories constructed this way recovers theories found previously by Dimofte-Gaiotto-Gukov. As a result, we also consider the cutting and gluing of 3-manifolds along smooth boundaries and the role played by all flat connections in this operation.

  8. Biocompatible 3D Matrix with Antimicrobial Properties.

    PubMed

    Ion, Alberto; Andronescu, Ecaterina; Rădulescu, Dragoș; Rădulescu, Marius; Iordache, Florin; Vasile, Bogdan Ștefan; Surdu, Adrian Vasile; Albu, Madalina Georgiana; Maniu, Horia; Chifiriuc, Mariana Carmen; Grumezescu, Alexandru Mihai; Holban, Alina Maria

    2016-01-20

    The aim of this study was to develop, characterize and assess the biological activity of a new regenerative 3D matrix with antimicrobial properties, based on collagen (COLL), hydroxyapatite (HAp), β-cyclodextrin (β-CD) and usnic acid (UA). The prepared 3D matrix was characterized by Scanning Electron Microscopy (SEM), Fourier Transform Infrared Microscopy (FT-IRM), Transmission Electron Microscopy (TEM), and X-ray Diffraction (XRD). In vitro qualitative and quantitative analyses performed on cultured diploid cells demonstrated that the 3D matrix is biocompatible, allowing the normal development and growth of MG-63 osteoblast-like cells and exhibited an antimicrobial effect, especially on the Staphylococcus aureus strain, explained by the particular higher inhibitory activity of usnic acid (UA) against Gram positive bacterial strains. Our data strongly recommend the obtained 3D matrix to be used as a successful alternative for the fabrication of three dimensional (3D) anti-infective regeneration matrix for bone tissue engineering.

  9. Decoupling diffusional from dimensional control of signaling in 3D culture reveals a role for myosin in tubulogenesis

    PubMed Central

    Raghavan, Srivatsan; Shen, Colette J.; Desai, Ravi A.; Sniadecki, Nathan J.; Nelson, Celeste M.; Chen, Christopher S.

    2010-01-01

    We present a novel microfabricated platform to culture cells within arrays of micrometer-scale three-dimensional (3D) extracellular matrix scaffolds (microgels). These microscale cultures eliminate diffusion barriers that are intrinsic to traditional 3D culture systems (macrogels) and enable uniform cytokine stimulation of the entire culture population, as well as allow immunolabeling, imaging and population-based biochemical assays across the relatively coplanar microgels. Examining early signaling associated with hepatocyte growth factor (HGF)-mediated scattering and tubulogenesis of MDCK cells revealed that 3D culture modulates cellular responses both through dimensionality and altered stimulation rates. Comparing responses in 2D culture, microgels and macrogels demonstrated that HGF-induced ERK signaling was driven by the dynamics of stimulation and not by whether cells were in a 2D or 3D environment, and that this ERK signaling was equally important for HGF-induced cell scattering on 2D substrates and tubulogenesis in 3D. By contrast, we discovered a specific HGF-induced increase in myosin expression leading to sustained downregulation of myosin activity that occurred only within 3D contexts and was required for 3D tubulogenesis but not 2D scattering. Interestingly, although absent in cells on collagen-coated plates, downregulation of myosin activity also occurred for cells on collagen gels, but was transient and mediated by a combination of myosin dephosphorylation and enhanced myosin expression. Furthermore, upregulating myosin activity via siRNA targeted to a myosin phosphatase did not attenuate scattering in 2D but did inhibit tubulogenesis in 3D. Together, these results demonstrate that cellular responses to soluble cues in 3D culture are regulated by both rates of stimulation and by matrix dimensionality, and highlight the importance of decoupling these effects to identify early signals relevant to cellular function in 3D environments. PMID:20682635

  10. 3D and Education

    NASA Astrophysics Data System (ADS)

    Meulien Ohlmann, Odile

    2013-02-01

    Today the industry offers a chain of 3D products. Learning to "read" and to "create in 3D" becomes an issue of education of primary importance. 25 years professional experience in France, the United States and Germany, Odile Meulien set up a personal method of initiation to 3D creation that entails the spatial/temporal experience of the holographic visual. She will present some different tools and techniques used for this learning, their advantages and disadvantages, programs and issues of educational policies, constraints and expectations related to the development of new techniques for 3D imaging. Although the creation of display holograms is very much reduced compared to the creation of the 90ies, the holographic concept is spreading in all scientific, social, and artistic activities of our present time. She will also raise many questions: What means 3D? Is it communication? Is it perception? How the seeing and none seeing is interferes? What else has to be taken in consideration to communicate in 3D? How to handle the non visible relations of moving objects with subjects? Does this transform our model of exchange with others? What kind of interaction this has with our everyday life? Then come more practical questions: How to learn creating 3D visualization, to learn 3D grammar, 3D language, 3D thinking? What for? At what level? In which matter? for whom?

  11. Chemotherapeutic efficiency of drugs in vitro: Comparison of doxorubicin exposure in 3D and 2D culture matrices.

    PubMed

    Casey, A; Gargotti, M; Bonnier, F; Byrne, H J

    2016-06-01

    The interest in the use of 3D matrices for in vitro analysis, with a view to increasing the relevance of in vitro studies and reducing the dependence on in vivo studies, has been growing in recent years. Cells grown in a 3D in vitro matrix environment have been reported to exhibit significantly different properties to those in a conventional 2D culture environment. However, comparison of 2D and 3D cell culture models have recently been noted to result in differing responses of cytotoxic assays, without any associated change in viability. The effect was attributed to differing conversion rates and effective concentrations of the resazurin assay in 2D and 3D environments, rather than differences in cellular metabolism. In this study, the efficacy of a chemotherapeutic agent, doxorubicin, is monitored and compared in conventional 2D and 3D collagen gel exposures of immortalized human cervical cells. Viability was monitored with the aid of the Alamar Blue assay and drug internalisation was verified using confocal microscopy. Drug uptake and retention within the collagen matrix was monitored by absorption spectroscopy. The viability studies showed apparent differences between the 2D and 3D culture systems, the differences attributed in part to the physical transition from 2D to a 3D environment causing alterations to dye resazurin uptake and conversion rates. The use of 3D culture matrices has widely been interpreted to result in "reduced" toxicity or cellular "resistance" to the chemotherapeutic agent. The results of this study show that the reduced efficiency of the drug to cells grown in the 3D environment can be accounted for by a sequential reduction of the effective concentration of the test compound and assay. This is due to absorption within the collagen gel inducing a higher uptake of both drug and assay thereby influencing the toxic impact of the drug and conversion rate of resazurin, and. The increased effective surface area of the cell exposed to the drug

  12. Screening for Stromal and Matrix Effects in 3D Microenvironments of Breast Cancer Cells

    NASA Astrophysics Data System (ADS)

    Montanez-Sauri, Sara I.

    Breast cancer progression ensures through the acquisition of genetic mutations, the uncontrollable growth of cells, and their progression to invasion. Studies have shown that the surrounding three-dimensional (3D) microenvironment can also influence breast cancer cell progression by controlling the morphology, differentiation, proliferation, and migration of cells. However, most of the currently available in vitro screening platforms are based on the two-dimensional (2D) culture of cells, and do not provide cells with the complex 3D microenvironment that exists in vivo. Therefore, there is a need for more biologically relevant in vitro platforms to help decipher the complexity of the microenvironment and its influence in breast cancer. In this dissertation we present an automated microfluidic platform that allows to efficiently screen for the effect of multiple matrix and stromal microenvironment in 3D cultures of breast cancer cells. Several extracellular matrix (ECM) compositions and stromal cells are included in the 3D microenvironments to examine their influence on breast cancer cell behavior. The screening results suggest that collagen gels with fibronectin might be influencing paracrine signals between breast cancer cells and stromal cells. The ability of the platform to culture and treat cells in 3D microenvironments offers a powerful screening tool for the identification of compounds and interactions using more in vivo-like 3D microenvironments. The identification of these mechanisms will increase our current understanding of breast cancer, and will aid in the identification of potential therapeutics.

  13. 3D Printed Trileaflet Valve Conduits Using Biological Hydrogels and Human Valve Interstitial Cells

    PubMed Central

    Duan, Bin; Kapetanovic, Edi; Hockaday, Laura A.; Butcher, Jonathan T.

    2014-01-01

    Tissue engineering has great potential to provide a functional de novo living valve replacement capable of integration with host tissue and growth. Among various valve conduit fabrication techniques, 3D bioprinting enables deposition of cells and hydrogels into 3D constructs with anatomical geometry and heterogeneous mechanical properties. Successful translation of this approach is however constrained by the dearth of printable and biocompatible hydrogel materials. Furthermore, it is not known how human valve cells respond to these printed environments. In this study, we develop 3D printable formulations of hybrid hydrogels based on methacrylated hyaluronic acid (Me-HA) and methacrylated gelatin (Me-Gel), and utilize them to bioprint heart valve conduits containing encapsulated human aortic valvular interstitial cells (HAVIC). Increasing Me-Gel concentration resulted in lower stiffness and higher viscosity, facilitated cell spreading, and better maintained HAVIC fibroblastic phenotype. Bioprinting accuracy was dependent upon the relative concentrations of Me-Gel and Me-HA, but when optimized enabled the fabrication of a trileaflet valve shape accurate to the original design. HAVIC encapsulated within bioprinted heart valves maintained high viability, and remodeled the initial matrix by depositing collagen and glyosaminoglycans. These findings represent the first rational design of bioprinted trileaflet valve hydrogels that regulate encapsulated human VIC behavior. The use of anatomically accurate living valve scaffolds through bioprinting may accelerate our understanding of physiological valve cell interactions and our progress towards de novo living valve replacements. PMID:24334142

  14. Temperature-sensitive polymer-conjugated IFN-gamma induces the expression of IDO mRNA and activity by fibroblasts populated in collagen gel (FPCG).

    PubMed

    Sarkhosh, Kourosh; Tredget, Edward E; Uludag, Hasan; Kilani, Ruhangiz T; Karami, Ali; Li, Yunyuan; Iwashina, Takashi; Ghahary, Aziz

    2004-10-01

    Indoleamine 2,3-dioxygenase (IDO) is an intracellular tryptophan-catabolizing enzyme possessing various immunosuppressive properties. Here, we report the use of this enzyme to suppress the proliferation of peripheral blood mononuclear cells (PBMC) co-cultured with IDO-expressing fibroblasts of an allogeneic skin substitute in vitro. Fetal foreskin fibroblasts populated within collagen gel (FPCG) were treated with interferon-gamma (IFN-gamma) conjugated with a temperature-sensitive polymer to induce the expression of IDO mRNA and protein. SDS-PAGE showed successful conjugation of IFN-gamma with the temperature-sensitive polymer. Expression of IDO mRNA was evaluated by Northern analysis. IDO enzyme activity was evaluated by the measurement of kynurenine levels. The results of Northern blot analysis showed an induction of IDO mRNA expression when treated with polymer-conjugated IFN-gamma. Kynurenine levels, as a measure of IDO bioactivity, were significantly higher in IFN-gamma-treated fibroblasts than in controls (P < 0.001). In a lasting effect experiment, the expression of IDO mRNA in FPCG treated with polymer-conjugated IFN-gamma was significantly longer than in those treated with free (non-conjugated) IFN-gamma (P < 0.001). IFN-gamma radiolabeling showed a prolonged retention of IFN-gamma within collagen gel in its polymer-conjugated form, compared to its free form. Presence of IDO protein in FPCG was demonstrated by Western analysis even 16 days after removal of the conditioned medium (containing released IFN-gamma). To demonstrate the immunosuppressive effects of IDO on the proliferation of PBMC, IDO-expressing FPCG treated with polymer-conjugated IFN-gamma were co-cultured with PBMC for a period of 5 days. The results showed a significant reduction in proliferation of PBMC co-cultured with IFN-gamma-treated IDO-expressing fibroblasts, compared to those co-cultured with non-IDO-expressing fibroblasts (P < 0.001). The addition of an IDO inhibitor (1-methyl

  15. 3D Imaging.

    ERIC Educational Resources Information Center

    Hastings, S. K.

    2002-01-01

    Discusses 3 D imaging as it relates to digital representations in virtual library collections. Highlights include X-ray computed tomography (X-ray CT); the National Science Foundation (NSF) Digital Library Initiatives; output peripherals; image retrieval systems, including metadata; and applications of 3 D imaging for libraries and museums. (LRW)

  16. Tenogenic Induction of Human MSCs by Anisotropically Aligned Collagen Biotextiles

    PubMed Central

    Younesi, Mousa; Islam, Anowarul; Kishore, Vipuil; Anderson, James M.; Akkus, Ozan

    2015-01-01

    A novel biofabrication modality, electrophoretic compaction with macromolecular alignment, was utilized to make collagen threads that mimic the native tendon’s structure and mechanical properties. A device with kinematic electrodes was designed to fabricate collagen threads in continuous length. For the first time, a 3D-biotextile was woven purely from collagen. Mechanical properties and load-displacement behavior of the biotextile mimicked those of the native tendon while presenting a porosity of 80%. The open pore network facilitated cell seeding across the continuum of the bioscaffold. Mesenchymal stem cells (MSCs) seeded in the woven scaffold underwent tenogenic differentiation in the absence of growth factors and synthesized a matrix that was positive for tenomodulin, COMP and type I collagen. Up-regulation of tenomodulin, a tendon specific marker, was 11.6 ± 3.5 fold, COMP was up-regulated 16.7 ± 5.5 fold, and Col I was up-regulated 6.9 ± 2.7 fold greater on ELAC threads when compared to randomly oriented collagen gels. These results demonstrate that a bioscaffold woven by using collagen threads with densely compacted and anisotropically aligned substrate texture stimulates tenogenesis topographically, rendering the electrochemically aligned collagen as a promising candidate for functional repair of tendons and ligaments. PMID:25750610

  17. Image-based multiscale modeling predicts tissue-level and network-level fiber reorganization in stretched cell-compacted collagen gels

    PubMed Central

    Sander, Edward A.; Stylianopoulos, Triantafyllos; Tranquillo, Robert T.; Barocas, Victor H.

    2009-01-01

    The mechanical environment plays an important role in cell signaling and tissue homeostasis. Unraveling connections between externally applied loads and the cellular response is often confounded by extracellular matrix (ECM) heterogeneity. Image-based multiscale models provide a foundation for examining the fine details of tissue behavior, but they require validation at multiple scales. In this study, we developed a multiscale model that captured the anisotropy and heterogeneity of a cell-compacted collagen gel subjected to an off-axis hold mechanical test and subsequently to biaxial extension. In both the model and experiments, the ECM reorganized in a nonaffine and heterogeneous manner that depended on multiscale interactions between the fiber networks. Simulations predicted that tensile and compressive fiber forces were produced to accommodate macroscopic displacements. Fiber forces in the simulation ranged from −11.3 to 437.7 nN, with a significant fraction of fibers under compression (12.1% during off-axis stretch). The heterogeneous network restructuring predicted by the model serves as an example of how multiscale modeling techniques provide a theoretical framework for understanding relationships between ECM structure and tissue-level mechanical properties and how microscopic fiber rearrangements could lead to mechanotransductive cell signaling. PMID:19805118

  18. TGF-β2 promotes RPE cell invasion into a collagen gel by mediating urokinase-type plasminogen activator (uPA) expression.

    PubMed

    Sugioka, Koji; Kodama, Aya; Okada, Kiyotaka; Iwata, Mihoko; Yoshida, Koji; Kusaka, Shunji; Matsumoto, Chota; Kaji, Hiroshi; Shimomura, Yoshikazu

    2013-10-01

    Transforming growth factor-beta (TGF-β) is one of the main epithelial-mesenchymal transition (EMT)-inducing factors. In general, TGF-β-induced EMT promotes cell migration and invasion. TGF-β also acts as a potent regulator of pericellular proteolysis by regulating the expression and secretion of plasminogen activators. Urokinase-type plasminogen activator (uPA) is a serine protease that binds to its cell surface receptor (uPAR) with high affinity. uPA binding to uPAR stimulates uPAR's interaction with transmembrane proteins, such as integrins, to regulate cytoskeletal reorganization and cell migration, differentiation and proliferation. However, the influence of TGF-β and the uPA/uPAR system on EMT in retinal pigment epithelial (RPE) cells is still unclear. The purpose of this study was to determine the effect of TGF-β2, which is the predominant isoform in the retina, and the uPA/uPAR system on RPE cells. In this study, we first examined the effect of TGF-β2 and/or the inhibitor of uPA (u-PA-STOP(®)) on the proliferation of a human retinal pigment epithelial cell line (ARPE-19 cells). Treatment with TGF-β2 or u-PA-STOP(®) suppressed cell proliferation. Combination treatment of TGF-β2 and u-PA-STOP(®) enhanced cell growth suppression. Furthermore, western blot analysis, fibrin zymography and real-time reverse transcription PCR showed that that TGF-β2 induced EMT in ARPE-19 cells and that the expression of uPA and uPAR expression was up-regulated during EMT. The TGF-β inhibitor SB431542 suppressed TGF-β2-stimulated uPA expression and secretion but did not suppress uPAR expression. Furthermore, we seeded ARPE-19 cells onto Transwell chambers and allowed them to invade the collagen matrix in the presence of TGF-β2 alone or with TGF-β2 and u-PA-STOP(®). TGF-β2 treatment induced ARPE-19 cell invasion into the collagen gel. Treatment with a combination of TGF-β2 and the uPA inhibitor strongly inhibited ARPE-19 cell invasion compared with treatment with

  19. 3D Cell Entrapment as a Function of the Weight Percent of Peptide-Amphiphile Hydrogels

    PubMed Central

    Scott, Carolyn M.; Forster, Colleen L.; Kokkoli, Efrosini

    2015-01-01

    The design of scaffolds which mimic the stiffness, nanofiber structure, and biochemistry of the native extra-cellular matrix (ECM) has been a major objective for the tissue engineering field. Furthermore, mimicking the innate three dimensional (3D) environment of the ECM has been shown to significantly alter cellular response compared to traditional two dimensional (2D) culture. We report the development of a self-assembling, fibronectin-mimetic, peptide-amphiphile nanofiber scaffold for 3D cell culture. To form such a scaffold, 5 mol% of a bioactive PR_g fibronectin-mimetic peptide-amphiphile was mixed with 95 mol% of a diluent peptide-amphiphile (E2) whose purpose was to neutralize electrostatic interactions, increase the gelation kinetics and promote cell survival. Atomic force microscopy verified the fibrilar structure of the gels and the mechanical properties were characterized for various weight percent (wt%) formulations of the 5 mol% PR_g - 95 mol% E2 peptide-amphiphile mixture. The 0.5 wt% formulations had an elastic modulus of 429.0 ± 21.3 Pa while the 1.0 wt% peptide-amphiphile hydrogels had an elastic modulus of 808.6 ± 38.1 Pa. The presence of entrapped cells in the gels decreased the elastic modulus and the decrease was a function of the cell loading. While both formulations supported cell proliferation, the 0.5 wt% gels supported significantly greater NIH3T3/GFP fibroblast cell proliferation throughout the gels than the 1.0 wt% gels. However, compared to the 0.5 wt% formulations, the 1.0 wt% hydrogels promoted greater increase in mRNA expression and production of fibronectin and type IV collagen ECM proteins. This study suggests that this fibronectin-mimetic scaffold holds great promise in the advance of 3D culture applications and cell therapies. PMID:25970351

  20. AE3D

    SciTech Connect

    Spong, Donald A

    2016-06-20

    AE3D solves for the shear Alfven eigenmodes and eigenfrequencies in a torodal magnetic fusion confinement device. The configuration can be either 2D (e.g. tokamak, reversed field pinch) or 3D (e.g. stellarator, helical reversed field pinch, tokamak with ripple). The equations solved are based on a reduced MHD model and sound wave coupling effects are not currently included.

  1. 3-D Seismic Interpretation

    NASA Astrophysics Data System (ADS)

    Moore, Gregory F.

    2009-05-01

    This volume is a brief introduction aimed at those who wish to gain a basic and relatively quick understanding of the interpretation of three-dimensional (3-D) seismic reflection data. The book is well written, clearly illustrated, and easy to follow. Enough elementary mathematics are presented for a basic understanding of seismic methods, but more complex mathematical derivations are avoided. References are listed for readers interested in more advanced explanations. After a brief introduction, the book logically begins with a succinct chapter on modern 3-D seismic data acquisition and processing. Standard 3-D acquisition methods are presented, and an appendix expands on more recent acquisition techniques, such as multiple-azimuth and wide-azimuth acquisition. Although this chapter covers the basics of standard time processing quite well, there is only a single sentence about prestack depth imaging, and anisotropic processing is not mentioned at all, even though both techniques are now becoming standard.

  2. Radiochromic 3D Detectors

    NASA Astrophysics Data System (ADS)

    Oldham, Mark

    2015-01-01

    Radiochromic materials exhibit a colour change when exposed to ionising radiation. Radiochromic film has been used for clinical dosimetry for many years and increasingly so recently, as films of higher sensitivities have become available. The two principle advantages of radiochromic dosimetry include greater tissue equivalence (radiologically) and the lack of requirement for development of the colour change. In a radiochromic material, the colour change arises direct from ionising interactions affecting dye molecules, without requiring any latent chemical, optical or thermal development, with important implications for increased accuracy and convenience. It is only relatively recently however, that 3D radiochromic dosimetry has become possible. In this article we review recent developments and the current state-of-the-art of 3D radiochromic dosimetry, and the potential for a more comprehensive solution for the verification of complex radiation therapy treatments, and 3D dose measurement in general.

  3. Bootstrapping 3D fermions

    DOE PAGES

    Iliesiu, Luca; Kos, Filip; Poland, David; ...

    2016-03-17

    We study the conformal bootstrap for a 4-point function of fermions <ψψψψ> in 3D. We first introduce an embedding formalism for 3D spinors and compute the conformal blocks appearing in fermion 4-point functions. Using these results, we find general bounds on the dimensions of operators appearing in the ψ × ψ OPE, and also on the central charge CT. We observe features in our bounds that coincide with scaling dimensions in the GrossNeveu models at large N. Finally, we also speculate that other features could coincide with a fermionic CFT containing no relevant scalar operators.

  4. Bootstrapping 3D fermions

    SciTech Connect

    Iliesiu, Luca; Kos, Filip; Poland, David; Pufu, Silviu S.; Simmons-Duffin, David; Yacoby, Ran

    2016-03-17

    We study the conformal bootstrap for a 4-point function of fermions <ψψψψ> in 3D. We first introduce an embedding formalism for 3D spinors and compute the conformal blocks appearing in fermion 4-point functions. Using these results, we find general bounds on the dimensions of operators appearing in the ψ × ψ OPE, and also on the central charge CT. We observe features in our bounds that coincide with scaling dimensions in the GrossNeveu models at large N. Finally, we also speculate that other features could coincide with a fermionic CFT containing no relevant scalar operators.

  5. A microfluidic device for 2D to 3D and 3D to 3D cell navigation

    NASA Astrophysics Data System (ADS)

    Shamloo, Amir; Amirifar, Leyla

    2016-01-01

    Microfluidic devices have received wide attention and shown great potential in the field of tissue engineering and regenerative medicine. Investigating cell response to various stimulations is much more accurate and comprehensive with the aid of microfluidic devices. In this study, we introduced a microfluidic device by which the matrix density as a mechanical property and the concentration profile of a biochemical factor as a chemical property could be altered. Our microfluidic device has a cell tank and a cell culture chamber to mimic both 2D to 3D and 3D to 3D migration of three types of cells. Fluid shear stress is negligible on the cells and a stable concentration gradient can be obtained by diffusion. The device was designed by a numerical simulation so that the uniformity of the concentration gradients throughout the cell culture chamber was obtained. Adult neural cells were cultured within this device and they showed different branching and axonal navigation phenotypes within varying nerve growth factor (NGF) concentration profiles. Neural stem cells were also cultured within varying collagen matrix densities while exposed to NGF concentrations and they experienced 3D to 3D collective migration. By generating vascular endothelial growth factor concentration gradients, adult human dermal microvascular endothelial cells also migrated in a 2D to 3D manner and formed a stable lumen within a specific collagen matrix density. It was observed that a minimum absolute concentration and concentration gradient were required to stimulate migration of all types of the cells. This device has the advantage of changing multiple parameters simultaneously and is expected to have wide applicability in cell studies.

  6. Cell-instructive starPEG-heparin-collagen composite matrices.

    PubMed

    Binner, Marcus; Bray, Laura J; Friedrichs, Jens; Freudenberg, Uwe; Tsurkan, Mikhail V; Werner, Carsten

    2017-02-16

    Polymer hydrogels can be readily modulated with regard to their physical properties and functionalized to recapitulate molecular cues of the extracellular matrix (ECM). However, they remain structurally different from the hierarchical supramolecular assemblies of natural ECM. Accordingly, we herein report a set of hydrogel composite materials made from starPEG-peptide conjugates, maleimide-functionalized heparin and collagen type I that combine semisynthetic and ECM-derived components. Collagen fibrillogenesis was controlled by temperature and collagen concentration to form collagen microstructures which were then homogeneously distributed within the 3D composite matrix during hydrogel formation. The collagen-laden hydrogel materials showed a heterogeneous local variation of the stiffness and adhesion ligand density. Composite gels functionalized with growth factors and cell adhesive peptides (RGDSP) supported the growth of embedded human umbilical cord vein endothelial cells (HUVECs) and induced the alignment of embedded bone marrow-derived human mesenchymal stem cells (MSCs) to the collagen microstructures in vitro. The introduced composite hydrogel material is concluded to faithfully mimic cell-instructive features of the ECM.

  7. Microscale mechanisms of agarose-induced disruption of collagen remodeling.

    PubMed

    Ulrich, Theresa A; Lee, Tae Geol; Shon, Hyun Kyong; Moon, Dae Won; Kumar, Sanjay

    2011-08-01

    Cells are strongly influenced by the local structure and mechanics of the extracellular matrix (ECM). We recently showed that adding agarose to soft collagen ECMs can mechanically stiffen these hydrogels by two orders of magnitude while limiting 3D cell motility, which we speculated might derive from agarose-mediated inhibition of collagen fiber deformation and remodeling. Here, we directly address this hypothesis by investigating the effects of agarose on cell-collagen interactions at the microscale. Addition of agarose progressively restricts cell spreading, reduces stress fiber and focal adhesion assembly, and inhibits macroscopic gel compaction. While time-of-flight secondary ion mass spectrometry and scanning electron microscopy fail to reveal agarose-induced alterations in collagen ligand presentation, the latter modality shows that agarose strongly impairs cell-directed assembly of large collagen bundles. Agarose-mediated inhibition of cell spreading and cytoarchitecture can be rescued by β-agarase digestion or by covalently crosslinking the matrix with glutaraldehyde. Based on these results, we argue that cell spreading and motility on collagen requires local matrix stiffening, which can be achieved via cell-mediated fiber remodeling or by chemically crosslinking the fibers. These findings provide new mechanistic insights into the regulatory function of agarose and bear general implications for cell adhesion and motility in fibrous ECMs.

  8. Venus in 3D

    NASA Technical Reports Server (NTRS)

    Plaut, Jeffrey J.

    1993-01-01

    Stereographic images of the surface of Venus which enable geologists to reconstruct the details of the planet's evolution are discussed. The 120-meter resolution of these 3D images make it possible to construct digital topographic maps from which precise measurements can be made of the heights, depths, slopes, and volumes of geologic structures.

  9. 3D photoacoustic imaging

    NASA Astrophysics Data System (ADS)

    Carson, Jeffrey J. L.; Roumeliotis, Michael; Chaudhary, Govind; Stodilka, Robert Z.; Anastasio, Mark A.

    2010-06-01

    Our group has concentrated on development of a 3D photoacoustic imaging system for biomedical imaging research. The technology employs a sparse parallel detection scheme and specialized reconstruction software to obtain 3D optical images using a single laser pulse. With the technology we have been able to capture 3D movies of translating point targets and rotating line targets. The current limitation of our 3D photoacoustic imaging approach is its inability ability to reconstruct complex objects in the field of view. This is primarily due to the relatively small number of projections used to reconstruct objects. However, in many photoacoustic imaging situations, only a few objects may be present in the field of view and these objects may have very high contrast compared to background. That is, the objects have sparse properties. Therefore, our work had two objectives: (i) to utilize mathematical tools to evaluate 3D photoacoustic imaging performance, and (ii) to test image reconstruction algorithms that prefer sparseness in the reconstructed images. Our approach was to utilize singular value decomposition techniques to study the imaging operator of the system and evaluate the complexity of objects that could potentially be reconstructed. We also compared the performance of two image reconstruction algorithms (algebraic reconstruction and l1-norm techniques) at reconstructing objects of increasing sparseness. We observed that for a 15-element detection scheme, the number of measureable singular vectors representative of the imaging operator was consistent with the demonstrated ability to reconstruct point and line targets in the field of view. We also observed that the l1-norm reconstruction technique, which is known to prefer sparseness in reconstructed images, was superior to the algebraic reconstruction technique. Based on these findings, we concluded (i) that singular value decomposition of the imaging operator provides valuable insight into the capabilities of

  10. Feasibility of silica-hybridized collagen hydrogels as three-dimensional cell matrices for hard tissue engineering.

    PubMed

    Yu, Hye-Sun; Lee, Eun-Jung; Seo, Seog-Jin; Knowles, Jonathan C; Kim, Hae-Won

    2015-09-01

    Exploiting hydrogels for the cultivation of stem cells, aiming to provide them with physico-chemical cues suitable for osteogenesis, is a critical demand for bone engineering. Here, we developed hybrid compositions of collagen and silica into hydrogels via a simple sol-gel process. The physico-chemical and mechanical properties, degradation behavior, and bone-bioactivity were characterized in-depth; furthermore, the in vitro mesenchymal stem cell growth and osteogenic differentiation behaviors within the 3D hybrid gel matrices were communicated for the first time. The hydrolyzed and condensed silica phase enabled chemical links with the collagen fibrils to form networked hybrid gels. The hybrid gels showed improved chemical stability and greater resistance to enzymatic degradation. The in vitro apatite-forming ability was enhanced by the hybrid composition. The viscoelastic mechanical properties of the hybrid gels were significantly improved in terms of the deformation resistance to an applied load and the modulus values under a dynamic oscillation. Mesenchymal stem cells adhered well to the hybrid networks and proliferated actively with substantial cytoskeletal extensions within the gel matrices. Of note, the hybrid gels substantially reduced the cell-mediated gel contraction behaviors, possibly due to the stiffer networks and higher resistance to cell-mediated degradation. Furthermore, the osteogenic differentiation of cells, including the expression of bone-associated genes and protein, was significantly upregulated within the hybrid gel matrices. Together with the physico-chemical and mechanical properties, the cellular behaviors observed within 3D gel matrices, being different from the previous approaches reported on 2D substrates, provide new information on the feasibility and usefulness of the silica-collagen system for stem cell culture and tissue engineering of hard tissues.

  11. Collagen gel droplet-embedded culture drug sensitivity testing in squamous cell carcinoma cell lines derived from human oral cancers: Optimal contact concentrations of cisplatin and fluorouracil

    PubMed Central

    Sakuma, Kaname; Tanaka, Akira; Mataga, Izumi

    2016-01-01

    The collagen gel droplet-embedded culture drug sensitivity test (CD-DST) is an anticancer drug sensitivity test that uses a method of three-dimensional culture of extremely small samples, and it is suited to primary cultures of human cancer cells. It is a useful method for oral squamous cell carcinoma (OSCC), in which the cancer tissues available for testing are limited. However, since the optimal contact concentrations of anticancer drugs have yet to be established in OSCC, CD-DST for detecting drug sensitivities of OSCC is currently performed by applying the optimal contact concentrations for stomach cancer. In the present study, squamous carcinoma cell lines from human oral cancer were used to investigate the optimal contact concentrations of cisplatin (CDDP) and fluorouracil (5-FU) during CD-DST for OSCC. CD-DST was performed in 7 squamous cell carcinoma cell lines derived from human oral cancers (Ca9-22, HSC-3, HSC-4, HO-1-N-1, KON, OSC-19 and SAS) using CDDP (0.15, 0.3, 1.25, 2.5, 5.0 and 10.0 µg/ml) and 5-FU (0.4, 0.9, 1.8, 3.8, 7.5, 15.0 and 30.0 µg/ml), and the optimal contact concentrations were calculated from the clinical response rate of OSCC to single-drug treatment and the in vitro efficacy rate curve. The optimal concentrations were 0.5 µg/ml for CDDP and 0.7 µg/ml for 5-FU. The antitumor efficacy of CDDP at this optimal contact concentration in CD-DST was compared to the antitumor efficacy in the nude mouse method. The T/C values, which were calculated as the ratio of the colony volume of the treatment group and the colony volume of the control group, at the optimal contact concentration of CDDP and of the nude mouse method were almost in agreement (P<0.05) and predicted clinical efficacy, indicating that the calculated optimal contact concentration is valid. Therefore, chemotherapy for OSCC based on anticancer drug sensitivity tests offers patients a greater freedom of choice and is likely to assume a greater importance in the selection of

  12. Twin Peaks - 3D

    NASA Technical Reports Server (NTRS)

    1997-01-01

    The two hills in the distance, approximately one to two kilometers away, have been dubbed the 'Twin Peaks' and are of great interest to Pathfinder scientists as objects of future study. 3D glasses are necessary to identify surface detail. The white areas on the left hill, called the 'Ski Run' by scientists, may have been formed by hydrologic processes.

    The IMP is a stereo imaging system with color capability provided by 24 selectable filters -- twelve filters per 'eye.

    Click below to see the left and right views individually. [figure removed for brevity, see original site] Left [figure removed for brevity, see original site] Right

  13. 3D and beyond

    NASA Astrophysics Data System (ADS)

    Fung, Y. C.

    1995-05-01

    This conference on physiology and function covers a wide range of subjects, including the vasculature and blood flow, the flow of gas, water, and blood in the lung, the neurological structure and function, the modeling, and the motion and mechanics of organs. Many technologies are discussed. I believe that the list would include a robotic photographer, to hold the optical equipment in a precisely controlled way to obtain the images for the user. Why are 3D images needed? They are to achieve certain objectives through measurements of some objects. For example, in order to improve performance in sports or beauty of a person, we measure the form, dimensions, appearance, and movements.

  14. 3D Audio System

    NASA Technical Reports Server (NTRS)

    1992-01-01

    Ames Research Center research into virtual reality led to the development of the Convolvotron, a high speed digital audio processing system that delivers three-dimensional sound over headphones. It consists of a two-card set designed for use with a personal computer. The Convolvotron's primary application is presentation of 3D audio signals over headphones. Four independent sound sources are filtered with large time-varying filters that compensate for motion. The perceived location of the sound remains constant. Possible applications are in air traffic control towers or airplane cockpits, hearing and perception research and virtual reality development.

  15. 3D Surgical Simulation

    PubMed Central

    Cevidanes, Lucia; Tucker, Scott; Styner, Martin; Kim, Hyungmin; Chapuis, Jonas; Reyes, Mauricio; Proffit, William; Turvey, Timothy; Jaskolka, Michael

    2009-01-01

    This paper discusses the development of methods for computer-aided jaw surgery. Computer-aided jaw surgery allows us to incorporate the high level of precision necessary for transferring virtual plans into the operating room. We also present a complete computer-aided surgery (CAS) system developed in close collaboration with surgeons. Surgery planning and simulation include construction of 3D surface models from Cone-beam CT (CBCT), dynamic cephalometry, semi-automatic mirroring, interactive cutting of bone and bony segment repositioning. A virtual setup can be used to manufacture positioning splints for intra-operative guidance. The system provides further intra-operative assistance with the help of a computer display showing jaw positions and 3D positioning guides updated in real-time during the surgical procedure. The CAS system aids in dealing with complex cases with benefits for the patient, with surgical practice, and for orthodontic finishing. Advanced software tools for diagnosis and treatment planning allow preparation of detailed operative plans, osteotomy repositioning, bone reconstructions, surgical resident training and assessing the difficulties of the surgical procedures prior to the surgery. CAS has the potential to make the elaboration of the surgical plan a more flexible process, increase the level of detail and accuracy of the plan, yield higher operative precision and control, and enhance documentation of cases. Supported by NIDCR DE017727, and DE018962 PMID:20816308

  16. Martian terrain - 3D

    NASA Technical Reports Server (NTRS)

    1997-01-01

    An area of rocky terrain near the landing site of the Sagan Memorial Station can be seen in this image, taken in stereo by the Imager for Mars Pathfinder (IMP) on Sol 3. 3D glasses are necessary to identify surface detail. This image is part of a 3D 'monster' panorama of the area surrounding the landing site.

    Mars Pathfinder is the second in NASA's Discovery program of low-cost spacecraft with highly focused science goals. The Jet Propulsion Laboratory, Pasadena, CA, developed and manages the Mars Pathfinder mission for NASA's Office of Space Science, Washington, D.C. JPL is an operating division of the California Institute of Technology (Caltech). The Imager for Mars Pathfinder (IMP) was developed by the University of Arizona Lunar and Planetary Laboratory under contract to JPL. Peter Smith is the Principal Investigator.

    Click below to see the left and right views individually. [figure removed for brevity, see original site] Left [figure removed for brevity, see original site] Right

  17. Collagen gels and the 'Bornstein legacy': from a substrate for tissue culture to cell culture systems and biomaterials for tissue regeneration.

    PubMed

    García-Gareta, Elena

    2014-07-01

    As collagen is the main structural component of connective tissues and skin, much effort was made in the past and still today to use it in cell culture applications. Moreover, collagen biomaterials are widely used in tissue regeneration, including the treatment of burns and chronic wounds. The great implications of the research carried out by Bornstein, Ehrmann and Gey on collagen preparations in the 1950s for cell culture and more recently tissue engineering and regeneration are described in this commentary. Specifically, it is explored why the 1958 paper on 'Reconstituted Rat-Tail Collagen Used as Substrate for Tissue Cultures on Coverslips in Maximow Slides and Roller Tubes' by M. B. Bornstein has made an invaluable contribution to the field.

  18. 3D Cell Culture in Alginate Hydrogels

    PubMed Central

    Andersen, Therese; Auk-Emblem, Pia; Dornish, Michael

    2015-01-01

    This review compiles information regarding the use of alginate, and in particular alginate hydrogels, in culturing cells in 3D. Knowledge of alginate chemical structure and functionality are shown to be important parameters in design of alginate-based matrices for cell culture. Gel elasticity as well as hydrogel stability can be impacted by the type of alginate used, its concentration, the choice of gelation technique (ionic or covalent), and divalent cation chosen as the gel inducing ion. The use of peptide-coupled alginate can control cell–matrix interactions. Gelation of alginate with concomitant immobilization of cells can take various forms. Droplets or beads have been utilized since the 1980s for immobilizing cells. Newer matrices such as macroporous scaffolds are now entering the 3D cell culture product market. Finally, delayed gelling, injectable, alginate systems show utility in the translation of in vitro cell culture to in vivo tissue engineering applications. Alginate has a history and a future in 3D cell culture. Historically, cells were encapsulated in alginate droplets cross-linked with calcium for the development of artificial organs. Now, several commercial products based on alginate are being used as 3D cell culture systems that also demonstrate the possibility of replacing or regenerating tissue. PMID:27600217

  19. 3D field harmonics

    SciTech Connect

    Caspi, S.; Helm, M.; Laslett, L.J.

    1991-03-30

    We have developed an harmonic representation for the three dimensional field components within the windings of accelerator magnets. The form by which the field is presented is suitable for interfacing with other codes that make use of the 3D field components (particle tracking and stability). The field components can be calculated with high precision and reduced cup time at any location (r,{theta},z) inside the magnet bore. The same conductor geometry which is used to simulate line currents is also used in CAD with modifications more readily available. It is our hope that the format used here for magnetic fields can be used not only as a means of delivering fields but also as a way by which beam dynamics can suggest correction to the conductor geometry. 5 refs., 70 figs.

  20. 3D co-cultures of keratinocytes and melanocytes and cytoprotective effects on keratinocytes against reactive oxygen species by insect virus-derived protein microcrystals.

    PubMed

    Shimabukuro, Junji; Yamaoka, Ayako; Murata, Ken-Ichi; Kotani, Eiji; Hirano, Tomoko; Nakajima, Yumiko; Matsumoto, Goichi; Mori, Hajime

    2014-09-01

    Stable protein microcrystals called polyhedra are produced by certain insect viruses. Cytokines, such as fibroblast growth factors (FGFs), can be immobilized within polyhedra. Here, we investigated three-dimensional (3D) co-cultures of keratinocytes and melanocytes on collagen gel containing FGF-2 and FGF-7 polyhedra. Melanocytes were observed to reside at the base of the 3D cell culture and melanin was also typically observed in the lower layer. The 3D cell culture model with FGF-2 and FGF-7 polyhedra was a useful in vitro model of the epidermis due to effective melanogenesis, proliferation and differentiation of keratinocytes. FGF-7 polyhedra showed a potent cytoprotective effect when keratinocytes were treated with menadione, which is a generator of reactive oxygen species. The cytoprotective effect was activated by the inositol triphosphate kinase-Akt pathway leading to upregulation of the antioxidant enzymes superoxide dismutase and peroxiredoxin 6.

  1. Hybrid Gel Composed of Native Heart Matrix and Collagen Induces Cardiac Differentiation of Human Embryonic Stem Cells without Supplemental Growth Factors

    PubMed Central

    Duan, Yi; Liu, Zen; O'Neill, John; Wan, Leo Q.; Freytes, Donald O.; Vunjak-Novakovic, Gordana

    2011-01-01

    Our goal was to assess the ability of native heart extracellular matrix (ECM) to direct cardiac differentiation of human embryonic stem cells (hESCs) in vitro. In order to probe the effects of cardiac matrix on hESC differentiation, a series of hydrogels was prepared from decellularized ECM from porcine hearts by mixing ECM and collagen type I at varying ratios. Maturation of cardiac function in embryoid bodies formed from hESCs was documented in terms of spontaneous contractile behavior and the mRNA and protein expression of cardiac markers. Hydrogel with high ECM content (75% ECM, 25% collagen, no supplemental soluble factors) increased the fraction of cells expressing cardiac marker troponin T, when compared with either hydrogel with low ECM content (25% ECM, 75% collagen, no supplemental soluble factors) or collagen hydrogel (100% collagen, with supplemental soluble factors). Furthermore, cardiac maturation was promoted in high-ECM content hydrogels, as evidenced by the striation patterns of cardiac troponin I and by upregulation of Cx43 gene. Consistently, high-ECM content hydrogels improved the contractile function of cardiac cells, as evidenced by increased numbers of contracting cells and increased contraction amplitudes. The ability of native ECM hydrogel to induce cardiac differentiation of hESCs without the addition of soluble factors makes it an attractive biomaterial system for basic studies of cardiac development and potentially for the delivery of therapeutic cells into the heart. PMID:21744185

  2. Intraoral 3D scanner

    NASA Astrophysics Data System (ADS)

    Kühmstedt, Peter; Bräuer-Burchardt, Christian; Munkelt, Christoph; Heinze, Matthias; Palme, Martin; Schmidt, Ingo; Hintersehr, Josef; Notni, Gunther

    2007-09-01

    Here a new set-up of a 3D-scanning system for CAD/CAM in dental industry is proposed. The system is designed for direct scanning of the dental preparations within the mouth. The measuring process is based on phase correlation technique in combination with fast fringe projection in a stereo arrangement. The novelty in the approach is characterized by the following features: A phase correlation between the phase values of the images of two cameras is used for the co-ordinate calculation. This works contrary to the usage of only phase values (phasogrammetry) or classical triangulation (phase values and camera image co-ordinate values) for the determination of the co-ordinates. The main advantage of the method is that the absolute value of the phase at each point does not directly determine the coordinate. Thus errors in the determination of the co-ordinates are prevented. Furthermore, using the epipolar geometry of the stereo-like arrangement the phase unwrapping problem of fringe analysis can be solved. The endoscope like measurement system contains one projection and two camera channels for illumination and observation of the object, respectively. The new system has a measurement field of nearly 25mm × 15mm. The user can measure two or three teeth at one time. So the system can by used for scanning of single tooth up to bridges preparations. In the paper the first realization of the intraoral scanner is described.

  3. 'Diamond' in 3-D

    NASA Technical Reports Server (NTRS)

    2004-01-01

    This 3-D, microscopic imager mosaic of a target area on a rock called 'Diamond Jenness' was taken after NASA's Mars Exploration Rover Opportunity ground into the surface with its rock abrasion tool for a second time.

    Opportunity has bored nearly a dozen holes into the inner walls of 'Endurance Crater.' On sols 177 and 178 (July 23 and July 24, 2004), the rover worked double-duty on Diamond Jenness. Surface debris and the bumpy shape of the rock resulted in a shallow and irregular hole, only about 2 millimeters (0.08 inch) deep. The final depth was not enough to remove all the bumps and leave a neat hole with a smooth floor. This extremely shallow depression was then examined by the rover's alpha particle X-ray spectrometer.

    On Sol 178, Opportunity's 'robotic rodent' dined on Diamond Jenness once again, grinding almost an additional 5 millimeters (about 0.2 inch). The rover then applied its Moessbauer spectrometer to the deepened hole. This double dose of Diamond Jenness enabled the science team to examine the rock at varying layers. Results from those grindings are currently being analyzed.

    The image mosaic is about 6 centimeters (2.4 inches) across.

  4. Prominent rocks - 3D

    NASA Technical Reports Server (NTRS)

    1997-01-01

    Many prominent rocks near the Sagan Memorial Station are featured in this image, taken in stereo by the Imager for Mars Pathfinder (IMP) on Sol 3. 3D glasses are necessary to identify surface detail. Wedge is at lower left; Shark, Half-Dome, and Pumpkin are at center. Flat Top, about four inches high, is at lower right. The horizon in the distance is one to two kilometers away.

    Mars Pathfinder is the second in NASA's Discovery program of low-cost spacecraft with highly focused science goals. The Jet Propulsion Laboratory, Pasadena, CA, developed and manages the Mars Pathfinder mission for NASA's Office of Space Science, Washington, D.C. JPL is an operating division of the California Institute of Technology (Caltech). The Imager for Mars Pathfinder (IMP) was developed by the University of Arizona Lunar and Planetary Laboratory under contract to JPL. Peter Smith is the Principal Investigator.

    Click below to see the left and right views individually. [figure removed for brevity, see original site] Left [figure removed for brevity, see original site] Right

  5. Additive manufacturing of collagen scaffolds by three-dimensional plotting of highly viscous dispersions.

    PubMed

    Lode, Anja; Meyer, Michael; Brüggemeier, Sophie; Paul, Birgit; Baltzer, Hagen; Schröpfer, Michaela; Winkelmann, Claudia; Sonntag, Frank; Gelinsky, Michael

    2016-02-27

    Additive manufacturing (AM) allows the free form fabrication of three-dimensional (3D) structures with distinct external geometry, fitting into a patient-specific defect, and defined internal pore architecture. However, fabrication of predesigned collagen scaffolds using AM-based technologies is challenging due to the low viscosity of collagen solutions, gels or dispersions commonly used for scaffold preparation. In the present study, we have developed a straightforward method which is based on 3D plotting of a highly viscous, high density collagen dispersion. The swollen state of the collagen fibrils at pH 4 enabled the homogenous extrusion of the material, the deposition of uniform strands and finally the construction of 3D scaffolds. Stabilization of the plotted structures was achieved by freeze-drying and chemical crosslinking with the carbodiimide EDC. The scaffolds exhibited high shape and dimensional fidelity and a hierarchical porosity consisting of macropores generated by strand deposition as well as an interconnected microporosity within the strands as result of the freeze-drying process. Cultivation of human mesenchymal stromal cells on the scaffolds, with and without adipogenic or osteogenic stimulation, revealed their cytocompatibility and potential applicability for adipose and bone tissue engineering.

  6. Biomimetic 3D Clusters Using Human Adipose Derived Mesenchymal Stem Cells and Breast Cancer Cells: A Study on Migration and Invasion of Breast Cancer Cells.

    PubMed

    Park, Min Hee; Song, Boa; Hong, Seungpyo; Kim, Sang Heon; Lee, Kangwon

    2016-07-05

    Invasion and metastasis of cancer directly related to human death have been associated with interactions among many different types of cells and three-dimensional (3D) tissue matrices. Precise mechanisms related to cancer invasion and metastasis still remain unknown due to their complexities. Development of tumor microenvironment (TME)-mimicking system could play a key role in understanding cancer environments and in elucidating the relating phenomena and their driving forces. Here we report a facile and novel platform of 3D cancer cell-clusters using human adipose-derived mesenchymal stem cells (hASCs) and breast cancer cells (MDA-MB-231) within a collagen gel matrix to show cancer invasion in the cell and extracellular matrix (ECM). Both clusters A (hASC only) and AC (hASC and MDA-MB-231) exhibited different behaviors and expressions of migration and invasion, as observed by the relating markers such as fibronectin, α-SMA, and CXCR4. hASCs showed a protrusive migration from a cluster center, whereas MDA-MB-231 spread out radially followed by hASC migration. Finally, the effect of matrix was further discussed by varying collagen gel densities. The new biomimetic system of 3D cancer clusters developed here has the potential to be utilized for research on migration and invasion of cancer cells in extracellular matrices.

  7. Multizone Paper Platform for 3D Cell Cultures

    PubMed Central

    Derda, Ratmir; Hong, Estrella; Mwangi, Martin; Mammoto, Akiko; Ingber, Donald E.; Whitesides, George M.

    2011-01-01

    In vitro 3D culture is an important model for tissues in vivo. Cells in different locations of 3D tissues are physiologically different, because they are exposed to different concentrations of oxygen, nutrients, and signaling molecules, and to other environmental factors (temperature, mechanical stress, etc). The majority of high-throughput assays based on 3D cultures, however, can only detect the average behavior of cells in the whole 3D construct. Isolation of cells from specific regions of 3D cultures is possible, but relies on low-throughput techniques such as tissue sectioning and micromanipulation. Based on a procedure reported previously (“cells-in-gels-in-paper” or CiGiP), this paper describes a simple method for culture of arrays of thin planar sections of tissues, either alone or stacked to create more complex 3D tissue structures. This procedure starts with sheets of paper patterned with hydrophobic regions that form 96 hydrophilic zones. Serial spotting of cells suspended in extracellular matrix (ECM) gel onto the patterned paper creates an array of 200 micron-thick slabs of ECM gel (supported mechanically by cellulose fibers) containing cells. Stacking the sheets with zones aligned on top of one another assembles 96 3D multilayer constructs. De-stacking the layers of the 3D culture, by peeling apart the sheets of paper, “sections” all 96 cultures at once. It is, thus, simple to isolate 200-micron-thick cell-containing slabs from each 3D culture in the 96-zone array. Because the 3D cultures are assembled from multiple layers, the number of cells plated initially in each layer determines the spatial distribution of cells in the stacked 3D cultures. This capability made it possible to compare the growth of 3D tumor models of different spatial composition, and to examine the migration of cells in these structures. PMID:21573103

  8. The biocompatibility of titanium in a buffer solution: compared effects of a thin film of TiO2 deposited by MOCVD and of collagen deposited from a gel.

    PubMed

    Popescu, Simona; Demetrescu, Ioana; Sarantopoulos, Christos; Gleizes, Alain N; Iordachescu, Dana

    2007-10-01

    This study aims at evaluating the biocompatibility of titanium surfaces modified according two different ways: (i) deposition of a bio-inert, thin film of rutile TiO(2) by chemical vapour deposition (MOCVD), and (ii) biochemical treatment with collagen gel, in order to obtain a bio-interactive coating. Behind the comparison is the idea that either the bio-inert or the bio-active coating has specific advantages when applied to implant treatment, such as the low price of the collagen treatment for instance. The stability in buffer solution was evaluated by open circuit potential (OCP) for medium time and cyclic voltametry. The OCP stabilized after 5.10(4) min for all the specimens except the collagen treated sample which presented a stable OCP from the first minutes. MOCVD treated samples stabilized to more electropositive values. Numeric results were statistically analysed to obtain the regression equations for long time predictable evolution. The corrosion parameters determined from cyclic curves revealed that the MOCVD treatment is an efficient way to improve corrosion resistance. Human dermal fibroblasts were selected for cell culture tests, taking into account that these cells are present in all bio-interfaces, being the main cellular type of connective tissue. The cells grew on either type of surface without phenotype modification. From the reduction of yellow, water-soluble 3-(4,5-dimethyldiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT cytotoxicity test), MOCVD treated samples offer better viability than mechanically polished Ti and collagen treated samples as well. Cell spreading, as evaluated from microscope images processed by the program Sigma Scan, showed also enhancement upon surface modification. Depending on the experimental conditions, MOCVD deposited TiO(2) exhibits different nanostructures that may influence biological behaviour. The results demonstrate the capacity of integration in simulated physiologic liquids for an implant pretreated by

  9. Sequential assembly of 3D perfusable microfluidic hydrogels.

    PubMed

    He, Jiankang; Zhu, Lin; Liu, Yaxiong; Li, Dichen; Jin, Zhongmin

    2014-11-01

    Bottom-up tissue engineering provides a promising way to recreate complex structural organizations of native organs in artificial constructs by assembling functional repeating modules. However, it is challenging for current bottom-up strategies to simultaneously produce a controllable and immediately perfusable microfluidic network in modularly assembled 3D constructs. Here we presented a bottom-up strategy to produce perfusable microchannels in 3D hydrogels by sequentially assembling microfluidic modules. The effects of agarose-collagen composition on microchannel replication and 3D assembly of hydrogel modules were investigated. The unique property of predefined microchannels in transporting fluids within 3D assemblies was evaluated. Endothelial cells were incorporated into the microfluidic network of 3D hydrogels for dynamic culture in a house-made bioreactor system. The results indicated that the sequential assembly method could produce interconnected 3D predefined microfluidic networks in optimized agarose-collagen hydrogels, which were fully perfusable and successfully functioned as fluid pathways to facilitate the spreading of endothelial cells. We envision that the presented method could be potentially used to engineer 3D vascularized parenchymal constructs by encapsulating primary cells in bulk hydrogels and incorporating endothelial cells in predefined microchannels.

  10. Cancer-associated Fibroblasts Induce a Collagen Cross-link Switch in Tumor Stroma

    PubMed Central

    Pankova, Daniela; Chen, Yulong; Terajima, Masahiko; Schliekelman, Mark J.; Baird, Brandi N.; Fahrenholtz, Monica; Sun, Li; Gill, Bartley J.; Vadakkan, Tegy J.; Kim, Min P.; Ahn, Young-Ho; Roybal, Jonathon D.; Liu, Xin; Parra Cuentas, Edwin Roger; Rodriguez, Jaime; Wistuba, Ignacio I.; Creighton, Chad J.; Gibbons, Don L.; Hicks, John M.; Dickinson, Mary E.; West, Jennifer L.; Grande-Allen, K. Jane; Hanash, Samir M.; Yamauchi, Mitsuo; Kurie, Jonathan M.

    2015-01-01

    Intratumoral collagen cross-links heighten stromal stiffness and stimulate tumor cell invasion, but it is unclear how collagen cross-linking is regulated in epithelial tumors. To address this question, we used KrasLA1 mice, which develop lung adenocarcinomas from somatic activation of a KrasG12D allele. The lung tumors in KrasLA1 mice were highly fibrotic and contained cancer-associated fibroblasts (CAFs) that produced collagen and generated stiffness in collagen gels. In xenograft tumors generated by injection of wild-type mice with lung adenocarcinoma cells alone or in combination with CAFs, the total concentration of collagen cross-links was the same in tumors generated with or without CAFs, but co-injected tumors had higher hydroxylysine aldehyde-derived collagen cross-links (HLCCs) and lower lysine-aldehyde-derived collagen cross-links (LCCs). Therefore, we postulated that an LCC-to-HLCC switch induced by CAFs promotes the migratory and invasive properties of lung adenocarcinoma cells. To test this hypothesis, we created co-culture models in which CAFs are positioned interstitially or peripherally in tumor cell aggregates, mimicking distinct spatial orientations of CAFs in human lung cancer. In both contexts, CAFs enhanced the invasive properties of tumor cells in 3-dimensional (3D) collagen gels. Tumor cell aggregates that attached to CAF networks on a Matrigel surface dissociated and migrated on the networks. Lysyl hydroxylase 2 (PLOD2/LH2), which drives HLCC formation, was expressed in CAFs, and LH2 depletion abrogated the ability of CAFs to promote tumor cell invasion and migration. PMID:26631572

  11. 3D Printed Dry EEG Electrodes

    PubMed Central

    Krachunov, Sammy; Casson, Alexander J.

    2016-01-01

    Electroencephalography (EEG) is a procedure that records brain activity in a non-invasive manner. The cost and size of EEG devices has decreased in recent years, facilitating a growing interest in wearable EEG that can be used out-of-the-lab for a wide range of applications, from epilepsy diagnosis, to stroke rehabilitation, to Brain-Computer Interfaces (BCI). A major obstacle for these emerging applications is the wet electrodes, which are used as part of the EEG setup. These electrodes are attached to the human scalp using a conductive gel, which can be uncomfortable to the subject, causes skin irritation, and some gels have poor long-term stability. A solution to this problem is to use dry electrodes, which do not require conductive gel, but tend to have a higher noise floor. This paper presents a novel methodology for the design and manufacture of such dry electrodes. We manufacture the electrodes using low cost desktop 3D printers and off-the-shelf components for the first time. This allows quick and inexpensive electrode manufacturing and opens the possibility of creating electrodes that are customized for each individual user. Our 3D printed electrodes are compared against standard wet electrodes, and the performance of the proposed electrodes is suitable for BCI applications, despite the presence of additional noise. PMID:27706094

  12. Construction of Mesenchymal Stem Cell–Containing Collagen Gel with a Macrochanneled Polycaprolactone Scaffold and the Flow Perfusion Culturing for Bone Tissue Engineering

    PubMed Central

    Yu, Hye-Sun; Won, Jong-Eun; Jin, Guang-Zhen

    2012-01-01

    Abstract A novel bone tissue-engineering construct was developed by using poly(ɛ-caprolactone) (PCL)-macrochanneled scaffolds combined with stem cell-seeded collagen hydrogels and then applying flow perfusion culture. Rat mesenchymal stem cells (MSCs) were loaded into collagen hydrogels, which were then combined with macrochanneled PCL scaffolds. Collagen hydrogels were demonstrated to provide favorable growth environments for MSCs and to foster proliferation. Cell number determination identified retention of substantially fewer (50–60%) cells when they were seeded directly onto macrochanneled PCL than of cells engineered within collagen hydrogels. Additionally, the cells actively proliferated within the combined scaffold for up to 7 days. MSC-loaded collagen–PCL scaffolds were subsequently cultured under flow perfusion to promote proliferation and osteogenic differentiation. Cells proliferated to levels significantly higher in flow perfusion culture than that under static conditions during 21 days. A quantitative polymerase chain reaction (QPCR) assay revealed significant alterations in the transcription of bone-related genes such as osteopontin (OPN), osteocalcin (OCN), and bone sialoprotein (BSP), such as 8-, 2.5-, and 3-fold induction, respectively, after 10 days of flow perfusion relative to those in static culture. OPN and OCN protein levels, as determined by Western blot, increased under flow perfusion. Cellular mineralization was significantly enhanced by the flow perfusion during 21 and 28 days. Analyses of mechanosensitive gene expression induced by flow perfusion shear stress revealed significant upregulation of c-fos and cyclooxygenase-2 (COX-2) during the initial culture period (3–5 days), suggesting that osteogenic stimulation was possible as a result of mechanical force-driven transduction. These results provide valuable information for the design of a new bone tissue-engineering system by combining stem cell-loaded collagen hydrogels with

  13. Distinct Contributions of Astrocytes and Pericytes to Neuroinflammation Identified in a 3D Human Blood-Brain Barrier on a Chip.

    PubMed

    Herland, Anna; van der Meer, Andries D; FitzGerald, Edward A; Park, Tae-Eun; Sleeboom, Jelle J F; Ingber, Donald E

    2016-01-01

    Neurovascular inflammation is a major contributor to many neurological disorders, but modeling these processes in vitro has proven to be difficult. Here, we microengineered a three-dimensional (3D) model of the human blood-brain barrier (BBB) within a microfluidic chip by creating a cylindrical collagen gel containing a central hollow lumen inside a microchannel, culturing primary human brain microvascular endothelial cells on the gel's inner surface, and flowing medium through the lumen. Studies were carried out with the engineered microvessel containing endothelium in the presence or absence of either primary human brain pericytes beneath the endothelium or primary human brain astrocytes within the surrounding collagen gel to explore the ability of this simplified model to identify distinct contributions of these supporting cells to the neuroinflammatory response. This human 3D BBB-on-a-chip exhibited barrier permeability similar to that observed in other in vitro BBB models created with non-human cells, and when stimulated with the inflammatory trigger, tumor necrosis factor-alpha (TNF-α), different secretion profiles for granulocyte colony-stimulating factor (G-CSF) and interleukin-6 (IL-6) were observed depending on the presence of astrocytes or pericytes. Importantly, the levels of these responses detected in the 3D BBB chip were significantly greater than when the same cells were co-cultured in static Transwell plates. Thus, as G-CSF and IL-6 have been reported to play important roles in neuroprotection and neuroactivation in vivo, this 3D BBB chip potentially offers a new method to study human neurovascular function and inflammation in vitro, and to identify physiological contributions of individual cell types.

  14. A Membrane-Type-1 Matrix Metalloproteinase (MT1-MMP) – Discoidin Domain Receptor 1 Axis Regulates Collagen-Induced Apoptosis in Breast Cancer Cells

    PubMed Central

    Assent, Delphine; Bourgot, Isabelle; Hennuy, Benoît; Geurts, Pierre; Noël, Agnès; Foidart, Jean-Michel; Maquoi, Erik

    2015-01-01

    During tumour dissemination, invading breast carcinoma cells become confronted with a reactive stroma, a type I collagen-rich environment endowed with anti-proliferative and pro-apoptotic properties. To develop metastatic capabilities, tumour cells must acquire the capacity to cope with this novel microenvironment. How cells interact with and respond to their microenvironment during cancer dissemination remains poorly understood. To address the impact of type I collagen on the fate of tumour cells, human breast carcinoma MCF-7 cells were cultured within three-dimensional type I collagen gels (3D COL1). Using this experimental model, we have previously demonstrated that membrane type-1 matrix metalloproteinase (MT1-MMP), a proteinase overexpressed in many aggressive tumours, promotes tumour progression by circumventing the collagen-induced up-regulation of BIK, a pro-apoptotic tumour suppressor, and hence apoptosis. Here we performed a transcriptomic analysis to decipher the molecular mechanisms regulating 3D COL1-induced apoptosis in human breast cancer cells. Control and MT1-MMP expressing MCF-7 cells were cultured on two-dimensional plastic plates or within 3D COL1 and a global transcriptional time-course analysis was performed. Shifting the cells from plastic plates to 3D COL1 activated a complex reprogramming of genes implicated in various biological processes. Bioinformatic analysis revealed a 3D COL1-mediated alteration of key cellular functions including apoptosis, cell proliferation, RNA processing and cytoskeleton remodelling. By using a panel of pharmacological inhibitors, we identified discoidin domain receptor 1 (DDR1), a receptor tyrosine kinase specifically activated by collagen, as the initiator of 3D COL1-induced apoptosis. Our data support the concept that MT1-MMP contributes to the inactivation of the DDR1-BIK signalling axis through the cleavage of collagen fibres and/or the alteration of DDR1 receptor signalling unit, without triggering a

  15. Vinculin Regulates Directionality and Cell Polarity in 2D, 3D Matrix and 3D Microtrack Migration.

    PubMed

    Rahman, Aniqua; Carey, Shawn P; Kraning-Rush, Casey M; Goldblatt, Zachary E; Bordeleau, Francois; Lampi, Marsha C; Lin, Deanna Y; García, Andrés J; Reinhart-King, Cynthia A

    2016-03-09

    During metastasis, cells can use proteolytic activity to form tube-like "microtracks" within the extracellular matrix (ECM). Using these microtracks, cells can migrate unimpeded through the stroma. To investigate the molecular mechanisms of microtrack migration, we developed an in vitro 3D micromolded collagen platform. When in microtracks, cells tend to migrate unidirectionally. Since focal adhesions are the primary mechanism by which cells interact with the ECM, we examined the roles of several focal adhesion molecules in driving unidirectional motion. Vinculin knockdown results in the repeated reversal of migration direction compared with control cells. Tracking the position of the Golgi centroid relative to the position of the nucleus centroid reveals that vinculin knockdown disrupts cell polarity in microtracks. Vinculin also directs migration on 2D substrates and in 3D uniform collagen matrices, indicated by reduced speed, shorter net displacement and decreased directionality in vinculin-deficient cells. In addition, vinculin is necessary for Focal Adhesion Kinase (FAK) activation in 3D as vinculin knockdown results in reduced FAK activation in both 3D uniform collagen matrices and microtracks, but not on 2D substrates, and accordingly, FAK inhibition halts cell migration in 3D microtracks. Together, these data indicate that vinculin plays a key role in polarization during migration.

  16. Imaging of collagen matrix remodeling in three-dimensional space using second harmonic generation and two photon excitation fluorescence

    NASA Astrophysics Data System (ADS)

    Abraham, Thomas; Carthy, Jon; McManus, Bruce

    2009-02-01

    Second harmonic generation (SHG), a nonlinear optical phenomenon, exhibits several in-common characteristics of twophoton excited fluorescence (TPEF) microscopy. These characteristics include identical equipment requirements from experiment to experiment and the intrinsic capability of generating 3-dimensional (D) high resolution images. Structural protein arrays that are highly ordered, such as collagen, produce strong SHG signals without the need for any exogenous label (stain). SHG and TPEF can be used together to provide information on structural rearrangements in 3D space of the collagen matrix associated with various physiological processes. In this study, we used SHG and TPEF to detect cellmediated structural reorganization of the extracellular collagen matrix in 3D space triggered by dimensional changes of embedded fibroblasts. These fibroblasts were cultured in native type I collagen gels and were stimulated to contract for a period of 24 hours. The gels were stained for cell nuclei with Hoechst and for actin with phalloidin conjugated to Alexa Fluor 488. We used non-de-scanned detectors and spectral scanning mode both in the reflection geometry for generating the 3D images and for SHG spectra, respectively. We used a tunable infrared laser with 100-fs pulses at a repetition rate of 80-MHz tuned to 800-nm for Hoechst and Alexa 488 excitations. We employed a broad range of excitation wavelengths (800 to 880-nm) with a scan interval of 10 nm to detect the SHG signal. We found that spectrally clean SHG signal peaked at 414-nm with excitation wavelength of 830-nm. The SHG spectrum has a full width half maximum (FWHM) bandwidth of 6.60-nm, which is consistent with its scaling relation to FWHM bandwidth 100-fs excitation pulses. When stimulated to contract, we found the fibroblasts to be highly elongated as well as interconnected in 2D space, and the collagen matrix, in the form of a visibly clear fibril structure, accumulated around the cells. In the absence of

  17. 3D Spectroscopy in Astronomy

    NASA Astrophysics Data System (ADS)

    Mediavilla, Evencio; Arribas, Santiago; Roth, Martin; Cepa-Nogué, Jordi; Sánchez, Francisco

    2011-09-01

    Preface; Acknowledgements; 1. Introductory review and technical approaches Martin M. Roth; 2. Observational procedures and data reduction James E. H. Turner; 3. 3D Spectroscopy instrumentation M. A. Bershady; 4. Analysis of 3D data Pierre Ferruit; 5. Science motivation for IFS and galactic studies F. Eisenhauer; 6. Extragalactic studies and future IFS science Luis Colina; 7. Tutorials: how to handle 3D spectroscopy data Sebastian F. Sánchez, Begona García-Lorenzo and Arlette Pécontal-Rousset.

  18. Spherical 3D isotropic wavelets

    NASA Astrophysics Data System (ADS)

    Lanusse, F.; Rassat, A.; Starck, J.-L.

    2012-04-01

    Context. Future cosmological surveys will provide 3D large scale structure maps with large sky coverage, for which a 3D spherical Fourier-Bessel (SFB) analysis in spherical coordinates is natural. Wavelets are particularly well-suited to the analysis and denoising of cosmological data, but a spherical 3D isotropic wavelet transform does not currently exist to analyse spherical 3D data. Aims: The aim of this paper is to present a new formalism for a spherical 3D isotropic wavelet, i.e. one based on the SFB decomposition of a 3D field and accompany the formalism with a public code to perform wavelet transforms. Methods: We describe a new 3D isotropic spherical wavelet decomposition based on the undecimated wavelet transform (UWT) described in Starck et al. (2006). We also present a new fast discrete spherical Fourier-Bessel transform (DSFBT) based on both a discrete Bessel transform and the HEALPIX angular pixelisation scheme. We test the 3D wavelet transform and as a toy-application, apply a denoising algorithm in wavelet space to the Virgo large box cosmological simulations and find we can successfully remove noise without much loss to the large scale structure. Results: We have described a new spherical 3D isotropic wavelet transform, ideally suited to analyse and denoise future 3D spherical cosmological surveys, which uses a novel DSFBT. We illustrate its potential use for denoising using a toy model. All the algorithms presented in this paper are available for download as a public code called MRS3D at http://jstarck.free.fr/mrs3d.html

  19. 3D Elevation Program—Virtual USA in 3D

    USGS Publications Warehouse

    Lukas, Vicki; Stoker, J.M.

    2016-04-14

    The U.S. Geological Survey (USGS) 3D Elevation Program (3DEP) uses a laser system called ‘lidar’ (light detection and ranging) to create a virtual reality map of the Nation that is very accurate. 3D maps have many uses with new uses being discovered all the time.  

  20. Human Astrocytes Develop Physiological Morphology and Remain Quiescent in a Novel 3D Matrix

    PubMed Central

    Placone, Amanda F.; McGuiggan, Patricia M.; Bergles, Dwight E.; Guerrero-Cazares, Hugo; Quiñones-Hinojosa, Alfredo; Searson, Peter C.

    2014-01-01

    Astrocytes are the most abundant glial cells in the brain and are responsible for diverse functions, from modulating synapse function to regulating the blood-brain barrier. In vivo, these cells exhibit a star-shaped morphology with multiple radial processes that contact synapses and completely surround brain capillaries. In response to trauma or CNS disease, astrocytes become reactive, a state associated with profound changes in gene expression, including upregulation of intermediate filament proteins, such as glial fibrillary acidic protein (GFAP). The inability to recapitulate the complex structure of astrocytes and maintain their quiescent state in vitro is a major roadblock to further developments in tissue engineering and regenerative medicine. Here, we characterize astrocyte morphology and activation in various hydrogels to assess the feasibility of developing a matrix that mimics key aspects of the native microenvironment. We show that astrocytes seeded in optimized matrix composed of collagen, hyaluronic acid, and matrigel exhibit a star-shaped morphology with radial processes and do not upregulate GFAP expression, hallmarks of quiescent astrocytes in the brain. In these optimized gels, collagen I provides structural support, HA mimics the brain extracellular matrix, and matrigel provides endothelial cell compatibility and was found to minimize GFAP upregulation. This defined 3D microenvironment for maintaining human astrocytes in vitro provides new opportunities for developing improved models of the blood-brain barrier and studying their response to stress signals. PMID:25542801

  1. UV damage of collagen: insights from model collagen peptides.

    PubMed

    Jariashvili, Ketevan; Madhan, Balaraman; Brodsky, Barbara; Kuchava, Ana; Namicheishvili, Louisa; Metreveli, Nunu

    2012-03-01

    Fibrils of Type I collagen in the skin are exposed to ultraviolet (UV) light and there have been claims that collagen photo-degradation leads to wrinkles and may contribute to skin cancers. To understand the effects of UV radiation on collagen, Type I collagen solutions were exposed to the UV-C wavelength of 254 nm for defined lengths of time at 4°C. Circular dichroism (CD) experiments show that irradiation of collagen leads to high loss of triple helical content with a new lower thermal stability peak and SDS-gel electrophoresis indicates breakdown of collagen chains. To better define the effects of UV radiation on the collagen triple-helix, the studies were extended to peptides which model the collagen sequence and conformation. CD studies showed irradiation for days led to lower magnitudes of the triple-helix maximum at 225 nm and lower thermal stabilities for two peptides containing multiple Gly-Pro-Hyp triplets. In contrast, the highest radiation exposure led to little change in the T(m) values of (Gly-Pro-Pro)(10) and (Ala-Hyp-Gly)(10) , although (Gly-Pro-Pro)(10) did show a significant decrease in triple helix intensity. Mass spectroscopy indicated preferential cleavage sites within the peptides, and identification of some of the most susceptible sites of cleavage. The effect of radiation on these well defined peptides gives insight into the sequence and conformational specificity of photo-degradation of collagen.

  2. Heterogeneity of collagens in rabbit cornea: type VI collagen

    SciTech Connect

    Cintron, C.; Hong, B.S.

    1988-05-01

    Normal adult rabbit corneas were digested with 5% pepsin and their collagens extracted with acetic acid. Collagen extracts were fractionated by differential salt precipitation. The 2.5 M NaCl fraction was then redissolved with tris buffer and precipitated with sodium acetate. The precipitate contained a high-molecular-weight disulfide-bonded aggregate which, upon reduction with mercaptoethanol, was converted into three distinct polypeptides having molecular weights between 45 and 66 Kd. These physical characteristics, together with the susceptibility of these polypeptides to collagenase and their amino acid composition, identified the high molecular weight aggregate as type VI collagen. Corneas from neonate rabbits and adult corneas containing 2-week-old scars were organ cultured in the presence of (/sup 14/C) glycine to incorporate radiolabel into collagen. Tissues were digested with 0.02% pepsin and their collagens extracted with formic acid. The total radioactivity of the extracts and tissue residues was determined before the collagens were separated by SDS-polyacrylamide slab gel electrophoresis. Radioactive collagen polypeptides bands were then stained with Coomassie blue, processed for fluorography, and analyzed by densitometry. The results show that: (1) type VI collagen is synthesized by neonate corneas and healing adult corneas; (2) it is not readily solubilized from either corneal tissue by 0.02% pepsin digestion and formic acid extraction; and (3) the proportion of type VI collagen deposited in scar tissue is markedly lower than that found in neonate corneas.

  3. Heterogeneity of collagens in rabbit cornea: type III collagen

    SciTech Connect

    Cintron, C.; Hong, B.S.; Covington, H.I.; Macarak, E.J.

    1988-05-01

    Whole neonate rabbit corneas and adult corneas containing 2-week-old scars were incubated in the presence of (/sup 14/C) glycine. Radiolabeled collagen extracted from the corneas and scar tissue were analyzed by sodium dodecylsulfate/polyacrylamide gel electrophoresis and fluorography to determine the types and relative quantity of collagen polypeptides present and synthesized by these tissues. In addition to other collagen types, type III was found in both neonate cornea and scar tissue from adult cornea, albeit in relatively small quantities. Type III collagen in normal cornea was associated with the residue after pepsin digestion and formic acid extraction of the tissue, and the same type of collagen was extracted from scar tissue after similar treatment. Type III collagen-specific monoclonal antibody bound to developing normal corneas and healing adult tissue sections, as determined by immunofluorescence. Antibody binding was localized to the endothelium and growing Descemet's membrane in fetal and neonate corneas, and restricted to the most posterior region of the corneal scar tissue. Although monoclonal antibody to keratan sulfate, used as a marker for stromal fibroblasts, bound to most of the scar tissue, the antibody failed to bind to the posterior scar tissue positive for type III collagen. We conclude that endothelial cells from fetal and neonate rabbit cornea and endothelium-derived fibroblasts from healing wounds of adult cornea synthesize and deposit type III collagen. Moreover, this collagen appears to be incorporated into the growing Descemet's membrane of normal corneas and narrow posterior portion of the scar tissue.

  4. 3D World Building System

    ScienceCinema

    None

    2016-07-12

    This video provides an overview of the Sandia National Laboratories developed 3-D World Model Building capability that provides users with an immersive, texture rich 3-D model of their environment in minutes using a laptop and color and depth camera.

  5. 3D Buckligami: Digital Matter

    NASA Astrophysics Data System (ADS)

    van Hecke, Martin; de Reus, Koen; Florijn, Bastiaan; Coulais, Corentin

    2014-03-01

    We present a class of elastic structures which exhibit collective buckling in 3D, and create these by a 3D printing/moulding technique. Our structures consist of cubic lattice of anisotropic unit cells, and we show that their mechanical properties are programmable via the orientation of these unit cells.

  6. 3D World Building System

    SciTech Connect

    2013-10-30

    This video provides an overview of the Sandia National Laboratories developed 3-D World Model Building capability that provides users with an immersive, texture rich 3-D model of their environment in minutes using a laptop and color and depth camera.

  7. LLNL-Earth3D

    SciTech Connect

    2013-10-01

    Earth3D is a computer code designed to allow fast calculation of seismic rays and travel times through a 3D model of the Earth. LLNL is using this for earthquake location and global tomography efforts and such codes are of great interest to the Earth Science community.

  8. Market study: 3-D eyetracker

    NASA Technical Reports Server (NTRS)

    1977-01-01

    A market study of a proposed version of a 3-D eyetracker for initial use at NASA's Ames Research Center was made. The commercialization potential of a simplified, less expensive 3-D eyetracker was ascertained. Primary focus on present and potential users of eyetrackers, as well as present and potential manufacturers has provided an effective means of analyzing the prospects for commercialization.

  9. Euro3D Science Conference

    NASA Astrophysics Data System (ADS)

    Walsh, J. R.

    2004-02-01

    The Euro3D RTN is an EU funded Research Training Network to foster the exploitation of 3D spectroscopy in Europe. 3D spectroscopy is a general term for spectroscopy of an area of the sky and derives its name from its two spatial + one spectral dimensions. There are an increasing number of instruments which use integral field devices to achieve spectroscopy of an area of the sky, either using lens arrays, optical fibres or image slicers, to pack spectra of multiple pixels on the sky (``spaxels'') onto a 2D detector. On account of the large volume of data and the special methods required to reduce and analyse 3D data, there are only a few centres of expertise and these are mostly involved with instrument developments. There is a perceived lack of expertise in 3D spectroscopy spread though the astronomical community and its use in the armoury of the observational astronomer is viewed as being highly specialised. For precisely this reason the Euro3D RTN was proposed to train young researchers in this area and develop user tools to widen the experience with this particular type of data in Europe. The Euro3D RTN is coordinated by Martin M. Roth (Astrophysikalisches Institut Potsdam) and has been running since July 2002. The first Euro3D science conference was held in Cambridge, UK from 22 to 23 May 2003. The main emphasis of the conference was, in keeping with the RTN, to expose the work of the young post-docs who are funded by the RTN. In addition the team members from the eleven European institutes involved in Euro3D also presented instrumental and observational developments. The conference was organized by Andy Bunker and held at the Institute of Astronomy. There were over thirty participants and 26 talks covered the whole range of application of 3D techniques. The science ranged from Galactic planetary nebulae and globular clusters to kinematics of nearby galaxies out to objects at high redshift. Several talks were devoted to reporting recent observations with newly

  10. 3D vision system assessment

    NASA Astrophysics Data System (ADS)

    Pezzaniti, J. Larry; Edmondson, Richard; Vaden, Justin; Hyatt, Bryan; Chenault, David B.; Kingston, David; Geulen, Vanilynmae; Newell, Scott; Pettijohn, Brad

    2009-02-01

    In this paper, we report on the development of a 3D vision system consisting of a flat panel stereoscopic display and auto-converging stereo camera and an assessment of the system's use for robotic driving, manipulation, and surveillance operations. The 3D vision system was integrated onto a Talon Robot and Operator Control Unit (OCU) such that direct comparisons of the performance of a number of test subjects using 2D and 3D vision systems were possible. A number of representative scenarios were developed to determine which tasks benefited most from the added depth perception and to understand when the 3D vision system hindered understanding of the scene. Two tests were conducted at Fort Leonard Wood, MO with noncommissioned officers ranked Staff Sergeant and Sergeant First Class. The scenarios; the test planning, approach and protocols; the data analysis; and the resulting performance assessment of the 3D vision system are reported.

  11. 3D printing in dentistry.

    PubMed

    Dawood, A; Marti Marti, B; Sauret-Jackson, V; Darwood, A

    2015-12-01

    3D printing has been hailed as a disruptive technology which will change manufacturing. Used in aerospace, defence, art and design, 3D printing is becoming a subject of great interest in surgery. The technology has a particular resonance with dentistry, and with advances in 3D imaging and modelling technologies such as cone beam computed tomography and intraoral scanning, and with the relatively long history of the use of CAD CAM technologies in dentistry, it will become of increasing importance. Uses of 3D printing include the production of drill guides for dental implants, the production of physical models for prosthodontics, orthodontics and surgery, the manufacture of dental, craniomaxillofacial and orthopaedic implants, and the fabrication of copings and frameworks for implant and dental restorations. This paper reviews the types of 3D printing technologies available and their various applications in dentistry and in maxillofacial surgery.

  12. PLOT3D user's manual

    NASA Technical Reports Server (NTRS)

    Walatka, Pamela P.; Buning, Pieter G.; Pierce, Larry; Elson, Patricia A.

    1990-01-01

    PLOT3D is a computer graphics program designed to visualize the grids and solutions of computational fluid dynamics. Seventy-four functions are available. Versions are available for many systems. PLOT3D can handle multiple grids with a million or more grid points, and can produce varieties of model renderings, such as wireframe or flat shaded. Output from PLOT3D can be used in animation programs. The first part of this manual is a tutorial that takes the reader, keystroke by keystroke, through a PLOT3D session. The second part of the manual contains reference chapters, including the helpfile, data file formats, advice on changing PLOT3D, and sample command files.

  13. PLOT3D/AMES, APOLLO UNIX VERSION USING GMR3D (WITHOUT TURB3D)

    NASA Technical Reports Server (NTRS)

    Buning, P.

    1994-01-01

    PLOT3D is an interactive graphics program designed to help scientists visualize computational fluid dynamics (CFD) grids and solutions. Today, supercomputers and CFD algorithms can provide scientists with simulations of such highly complex phenomena that obtaining an understanding of the simulations has become a major problem. Tools which help the scientist visualize the simulations can be of tremendous aid. PLOT3D/AMES offers more functions and features, and has been adapted for more types of computers than any other CFD graphics program. Version 3.6b+ is supported for five computers and graphic libraries. Using PLOT3D, CFD physicists can view their computational models from any angle, observing the physics of problems and the quality of solutions. As an aid in designing aircraft, for example, PLOT3D's interactive computer graphics can show vortices, temperature, reverse flow, pressure, and dozens of other characteristics of air flow during flight. As critical areas become obvious, they can easily be studied more closely using a finer grid. PLOT3D is part of a computational fluid dynamics software cycle. First, a program such as 3DGRAPE (ARC-12620) helps the scientist generate computational grids to model an object and its surrounding space. Once the grids have been designed and parameters such as the angle of attack, Mach number, and Reynolds number have been specified, a "flow-solver" program such as INS3D (ARC-11794 or COS-10019) solves the system of equations governing fluid flow, usually on a supercomputer. Grids sometimes have as many as two million points, and the "flow-solver" produces a solution file which contains density, x- y- and z-momentum, and stagnation energy for each grid point. With such a solution file and a grid file containing up to 50 grids as input, PLOT3D can calculate and graphically display any one of 74 functions, including shock waves, surface pressure, velocity vectors, and particle traces. PLOT3D's 74 functions are organized into

  14. PLOT3D/AMES, APOLLO UNIX VERSION USING GMR3D (WITH TURB3D)

    NASA Technical Reports Server (NTRS)

    Buning, P.

    1994-01-01

    PLOT3D is an interactive graphics program designed to help scientists visualize computational fluid dynamics (CFD) grids and solutions. Today, supercomputers and CFD algorithms can provide scientists with simulations of such highly complex phenomena that obtaining an understanding of the simulations has become a major problem. Tools which help the scientist visualize the simulations can be of tremendous aid. PLOT3D/AMES offers more functions and features, and has been adapted for more types of computers than any other CFD graphics program. Version 3.6b+ is supported for five computers and graphic libraries. Using PLOT3D, CFD physicists can view their computational models from any angle, observing the physics of problems and the quality of solutions. As an aid in designing aircraft, for example, PLOT3D's interactive computer graphics can show vortices, temperature, reverse flow, pressure, and dozens of other characteristics of air flow during flight. As critical areas become obvious, they can easily be studied more closely using a finer grid. PLOT3D is part of a computational fluid dynamics software cycle. First, a program such as 3DGRAPE (ARC-12620) helps the scientist generate computational grids to model an object and its surrounding space. Once the grids have been designed and parameters such as the angle of attack, Mach number, and Reynolds number have been specified, a "flow-solver" program such as INS3D (ARC-11794 or COS-10019) solves the system of equations governing fluid flow, usually on a supercomputer. Grids sometimes have as many as two million points, and the "flow-solver" produces a solution file which contains density, x- y- and z-momentum, and stagnation energy for each grid point. With such a solution file and a grid file containing up to 50 grids as input, PLOT3D can calculate and graphically display any one of 74 functions, including shock waves, surface pressure, velocity vectors, and particle traces. PLOT3D's 74 functions are organized into

  15. Unassisted 3D camera calibration

    NASA Astrophysics Data System (ADS)

    Atanassov, Kalin; Ramachandra, Vikas; Nash, James; Goma, Sergio R.

    2012-03-01

    With the rapid growth of 3D technology, 3D image capture has become a critical part of the 3D feature set on mobile phones. 3D image quality is affected by the scene geometry as well as on-the-device processing. An automatic 3D system usually assumes known camera poses accomplished by factory calibration using a special chart. In real life settings, pose parameters estimated by factory calibration can be negatively impacted by movements of the lens barrel due to shaking, focusing, or camera drop. If any of these factors displaces the optical axes of either or both cameras, vertical disparity might exceed the maximum tolerable margin and the 3D user may experience eye strain or headaches. To make 3D capture more practical, one needs to consider unassisted (on arbitrary scenes) calibration. In this paper, we propose an algorithm that relies on detection and matching of keypoints between left and right images. Frames containing erroneous matches, along with frames with insufficiently rich keypoint constellations, are detected and discarded. Roll, pitch yaw , and scale differences between left and right frames are then estimated. The algorithm performance is evaluated in terms of the remaining vertical disparity as compared to the maximum tolerable vertical disparity.

  16. Engineering of In Vitro 3D Capillary Beds by Self-Directed Angiogenic Sprouting

    PubMed Central

    Chan, Juliana M.; Zervantonakis, Ioannis K.; Rimchala, Tharathorn; Polacheck, William J.; Whisler, Jordan; Kamm, Roger D.

    2012-01-01

    In recent years, microfluidic systems have been used to study fundamental aspects of angiogenesis through the patterning of single-layered, linear or geometric vascular channels. In vivo, however, capillaries exist in complex, three-dimensional (3D) networks, and angiogenic sprouting occurs with a degree of unpredictability in all x,y,z planes. The ability to generate capillary beds in vitro that can support thick, biological tissues remains a key challenge to the regeneration of vital organs. Here, we report the engineering of 3D capillary beds in an in vitro microfluidic platform that is comprised of a biocompatible collagen I gel supported by a mechanical framework of alginate beads. The engineered vessels have patent lumens, form robust ∼1.5 mm capillary networks across the devices, and support the perfusion of 1 µm fluorescent beads through them. In addition, the alginate beads offer a modular method to encapsulate and co-culture cells that either promote angiogenesis or require perfusion for cell viability in engineered tissue constructs. This laboratory-constructed vascular supply may be clinically significant for the engineering of capillary beds and higher order biological tissues in a scalable and modular manner. PMID:23226527

  17. Effects of extracellular fiber architecture on cell membrane shear stress in a 3D fibrous matrix.

    PubMed

    Pedersen, John A; Boschetti, Federica; Swartz, Melody A

    2007-01-01

    Interstitial fluid flow has been shown to affect the organization and behavior of cells in 3D environments in vivo and in vitro, yet the forces driving such responses are not clear. Due to the complex architecture of the extracellular matrix (ECM) and the difficulty of measuring fluid flow near cells embedded in it, the levels of shear stress experienced by cells in this environment are typically estimated using bulk-averaged matrix parameters such as hydraulic permeability. While this is useful for estimating average stresses, it cannot yield insight into how local matrix fiber architecture-which is cell-controlled in the immediate pericellular environment-affects the local stresses imposed on the cell surface. To address this, we used computational fluid dynamics to study flow through an idealized mesh constructed of a cubic lattice of fibers simulating a typical in vitro collagen gel. We found that, in such high porosity matrices, the fibers strongly affect the flow fields near the cell, with peak shear stresses up to five times higher than those predicted by the Brinkman equation. We also found that minor remodeling of the fibers near the cell surface had major effects on the shear stress profile on the cell. These findings demonstrate the importance of fiber architecture to the fluid forces on a cell embedded in a 3D matrix, and also show how small modifications in the local ECM can lead to large changes in the mechanical environment of the cell.

  18. Preparation of Collagen-Coated Gels that Maximize In Vitro Myogenesis of Stem Cells by Matching the Lateral Elasticity of In Vivo Muscle

    PubMed Central

    Chaudhuri, Tathagata; Rehfeldt, Florian; Sweeney, H. Lee; Discher, Dennis E.

    2011-01-01

    The physical nature of a cell’s microenvironment – including the elasticity of the surrounding tissue – appears to exert a significant influence on cell morphology, cytoskeleton, and gene expression. We have previously shown that committed muscle cells will develop sarcomeric striations of skeletal muscle myosin II only when the cells are grown on a compliant gel that closely matches the passive compliance of skeletal muscle. We have more recently shown with the same types of elastic gels that mesenchymal stem cells (MSCs) maximally express myogenic genes, even in the absence of tailored soluble factors. Here, we provide detailed methods not only for how we make and nanomechanically characterize hydrogels of muscle-like elasticity, but also how we culture MSCs and characterize their myogenic induction by whole human genome transcript analysis. PMID:20405368

  19. 3D Scan Systems Integration

    DTIC Science & Technology

    2007-11-02

    AGENCY USE ONLY (Leave Blank) 2. REPORT DATE 5 Feb 98 4. TITLE AND SUBTITLE 3D Scan Systems Integration REPORT TYPE AND DATES COVERED...2-89) Prescribed by ANSI Std. Z39-1 298-102 [ EDO QUALITY W3PECTEDI DLA-ARN Final Report for US Defense Logistics Agency on DDFG-T2/P3: 3D...SCAN SYSTEMS INTEGRATION Contract Number SPO100-95-D-1014 Contractor Ohio University Delivery Order # 0001 Delivery Order Title 3D Scan Systems

  20. 3D Printing of Carbon Nanotubes-Based Microsupercapacitors.

    PubMed

    Yu, Wei; Zhou, Han; Li, Ben Q; Ding, Shujiang

    2017-02-08

    A novel 3D printing procedure is presented for fabricating carbon-nanotubes (CNTs)-based microsupercapacitors. The 3D printer uses a CNTs ink slurry with a moderate solid content and prints a stream of continuous droplets. Appropriate control of a heated base is applied to facilitate the solvent removal and adhesion between printed layers and to improve the structure integrity without structure delamination or distortion upon drying. The 3D-printed electrodes for microsupercapacitors are characterized by SEM, laser scanning confocal microscope, and step profiler. Effect of process parameters on 3D printing is also studied. The final solid-state microsupercapacitors are assembled with the printed multilayer CNTs structures and poly(vinyl alcohol)-H3PO4 gel as the interdigitated microelectrodes and electrolyte. The electrochemical performance of 3D printed microsupercapacitors is also tested, showing a significant areal capacitance and excellent cycle stability.

  1. 3D polymer scaffold arrays.

    PubMed

    Simon, Carl G; Yang, Yanyin; Dorsey, Shauna M; Ramalingam, Murugan; Chatterjee, Kaushik

    2011-01-01

    We have developed a combinatorial platform for fabricating tissue scaffold arrays that can be used for screening cell-material interactions. Traditional research involves preparing samples one at a time for characterization and testing. Combinatorial and high-throughput (CHT) methods lower the cost of research by reducing the amount of time and material required for experiments by combining many samples into miniaturized specimens. In order to help accelerate biomaterials research, many new CHT methods have been developed for screening cell-material interactions where materials are presented to cells as a 2D film or surface. However, biomaterials are frequently used to fabricate 3D scaffolds, cells exist in vivo in a 3D environment and cells cultured in a 3D environment in vitro typically behave more physiologically than those cultured on a 2D surface. Thus, we have developed a platform for fabricating tissue scaffold libraries where biomaterials can be presented to cells in a 3D format.

  2. Autofocus for 3D imaging

    NASA Astrophysics Data System (ADS)

    Lee-Elkin, Forest

    2008-04-01

    Three dimensional (3D) autofocus remains a significant challenge for the development of practical 3D multipass radar imaging. The current 2D radar autofocus methods are not readily extendable across sensor passes. We propose a general framework that allows a class of data adaptive solutions for 3D auto-focus across passes with minimal constraints on the scene contents. The key enabling assumption is that portions of the scene are sparse in elevation which reduces the number of free variables and results in a system that is simultaneously solved for scatterer heights and autofocus parameters. The proposed method extends 2-pass interferometric synthetic aperture radar (IFSAR) methods to an arbitrary number of passes allowing the consideration of scattering from multiple height locations. A specific case from the proposed autofocus framework is solved and demonstrates autofocus and coherent multipass 3D estimation across the 8 passes of the "Gotcha Volumetric SAR Data Set" X-Band radar data.

  3. Distinct Contributions of Astrocytes and Pericytes to Neuroinflammation Identified in a 3D Human Blood-Brain Barrier on a Chip

    PubMed Central

    FitzGerald, Edward A.; Park, Tae-Eun; Sleeboom, Jelle J. F.; Ingber, Donald E.

    2016-01-01

    Neurovascular inflammation is a major contributor to many neurological disorders, but modeling these processes in vitro has proven to be difficult. Here, we microengineered a three-dimensional (3D) model of the human blood-brain barrier (BBB) within a microfluidic chip by creating a cylindrical collagen gel containing a central hollow lumen inside a microchannel, culturing primary human brain microvascular endothelial cells on the gel’s inner surface, and flowing medium through the lumen. Studies were carried out with the engineered microvessel containing endothelium in the presence or absence of either primary human brain pericytes beneath the endothelium or primary human brain astrocytes within the surrounding collagen gel to explore the ability of this simplified model to identify distinct contributions of these supporting cells to the neuroinflammatory response. This human 3D BBB-on-a-chip exhibited barrier permeability similar to that observed in other in vitro BBB models created with non-human cells, and when stimulated with the inflammatory trigger, tumor necrosis factor-alpha (TNF-α), different secretion profiles for granulocyte colony-stimulating factor (G-CSF) and interleukin-6 (IL-6) were observed depending on the presence of astrocytes or pericytes. Importantly, the levels of these responses detected in the 3D BBB chip were significantly greater than when the same cells were co-cultured in static Transwell plates. Thus, as G-CSF and IL-6 have been reported to play important roles in neuroprotection and neuroactivation in vivo, this 3D BBB chip potentially offers a new method to study human neurovascular function and inflammation in vitro, and to identify physiological contributions of individual cell types. PMID:26930059

  4. New 3D-Culture Approaches to Study Interactions of Bone Marrow Adipocytes with Metastatic Prostate Cancer Cells.

    PubMed

    Herroon, Mackenzie Katheryn; Diedrich, Jonathan Driscoll; Podgorski, Izabela

    2016-01-01

    Adipocytes are a major component of the bone marrow that can critically affect metastatic progression in bone. Understanding how the marrow fat cells influence growth, behavior, and survival of tumor cells requires utilization of in vitro cell systems that can closely mimic the physiological microenvironment. Herein, we present two new three-dimensional (3D) culture approaches to study adipocyte-tumor cell interactions in vitro. The first is a transwell-based system composed of the marrow-derived adipocytes in 3D collagen I gels and reconstituted basement membrane-overlayed prostate tumor cell spheroids. Tumor cells cultured under these 3D conditions are continuously exposed to adipocyte-derived factors, and their response can be evaluated by morphological and immunohistochemical analyses. We show via immunofluorescence analysis of metabolism-associated proteins that under 3D conditions tumor cells have significantly different metabolic response to adipocytes than tumor cells grown in 2D culture. We also demonstrate that this model allows for incorporation of other cell types, such as bone marrow macrophages, and utilization of dye-quenched collagen substrates for examination of proteolysis-driven responses to adipocyte- and macrophage-derived factors. Our second 3D culture system is designed to study tumor cell invasion toward the adipocytes and the consequent interaction between the two cell types. In this model, marrow adipocytes are separated from the fluorescently labeled tumor cells by a layer of collagen I. At designated time points, adipocytes are stained with BODIPY and confocal z-stacks are taken through the depth of the entire culture to determine the distance traveled between the two cell types over time. We demonstrate that this system can be utilized to study effects of candidate factors on tumor invasion toward the adipocytes. We also show that immunohistochemical analyses can be performed to evaluate the impact of direct interaction of prostate

  5. Combinatorial 3D Mechanical Metamaterials

    NASA Astrophysics Data System (ADS)

    Coulais, Corentin; Teomy, Eial; de Reus, Koen; Shokef, Yair; van Hecke, Martin

    2015-03-01

    We present a class of elastic structures which exhibit 3D-folding motion. Our structures consist of cubic lattices of anisotropic unit cells that can be tiled in a complex combinatorial fashion. We design and 3d-print this complex ordered mechanism, in which we combine elastic hinges and defects to tailor the mechanics of the material. Finally, we use this large design space to encode smart functionalities such as surface patterning and multistability.

  6. Mean deformation metrics for quantifying 3D cell–matrix interactions without requiring information about matrix material properties

    PubMed Central

    Stout, David A.; Bar-Kochba, Eyal; Estrada, Jonathan B.; Toyjanova, Jennet; Kesari, Haneesh; Reichner, Jonathan S.; Franck, Christian

    2016-01-01

    Mechanobiology relates cellular processes to mechanical signals, such as determining the effect of variations in matrix stiffness with cell tractions. Cell traction recorded via traction force microscopy (TFM) commonly takes place on materials such as polyacrylamide- and polyethylene glycol-based gels. Such experiments remain limited in physiological relevance because cells natively migrate within complex tissue microenvironments that are spatially heterogeneous and hierarchical. Yet, TFM requires determination of the matrix constitutive law (stress–strain relationship), which is not always readily available. In addition, the currently achievable displacement resolution limits the accuracy of TFM for relatively small cells. To overcome these limitations, and increase the physiological relevance of in vitro experimental design, we present a new approach and a set of associated biomechanical signatures that are based purely on measurements of the matrix's displacements without requiring any knowledge of its constitutive laws. We show that our mean deformation metrics (MDM) approach can provide significant biophysical information without the need to explicitly determine cell tractions. In the process of demonstrating the use of our MDM approach, we succeeded in expanding the capability of our displacement measurement technique such that it can now measure the 3D deformations around relatively small cells (∼10 micrometers), such as neutrophils. Furthermore, we also report previously unseen deformation patterns generated by motile neutrophils in 3D collagen gels. PMID:26929377

  7. Mean deformation metrics for quantifying 3D cell-matrix interactions without requiring information about matrix material properties.

    PubMed

    Stout, David A; Bar-Kochba, Eyal; Estrada, Jonathan B; Toyjanova, Jennet; Kesari, Haneesh; Reichner, Jonathan S; Franck, Christian

    2016-03-15

    Mechanobiology relates cellular processes to mechanical signals, such as determining the effect of variations in matrix stiffness with cell tractions. Cell traction recorded via traction force microscopy (TFM) commonly takes place on materials such as polyacrylamide- and polyethylene glycol-based gels. Such experiments remain limited in physiological relevance because cells natively migrate within complex tissue microenvironments that are spatially heterogeneous and hierarchical. Yet, TFM requires determination of the matrix constitutive law (stress-strain relationship), which is not always readily available. In addition, the currently achievable displacement resolution limits the accuracy of TFM for relatively small cells. To overcome these limitations, and increase the physiological relevance of in vitro experimental design, we present a new approach and a set of associated biomechanical signatures that are based purely on measurements of the matrix's displacements without requiring any knowledge of its constitutive laws. We show that our mean deformation metrics (MDM) approach can provide significant biophysical information without the need to explicitly determine cell tractions. In the process of demonstrating the use of our MDM approach, we succeeded in expanding the capability of our displacement measurement technique such that it can now measure the 3D deformations around relatively small cells (∼10 micrometers), such as neutrophils. Furthermore, we also report previously unseen deformation patterns generated by motile neutrophils in 3D collagen gels.

  8. 3D visualization of polymer nanostructure

    SciTech Connect

    Werner, James H

    2009-01-01

    Soft materials and structured polymers are extremely useful nanotechnology building blocks. Block copolymers, in particular, have served as 2D masks for nanolithography and 3D scaffolds for photonic crystals, nanoparticle fabrication, and solar cells. F or many of these applications, the precise 3 dimensional structure and the number and type of defects in the polymer is important for ultimate function. However, directly visualizing the 3D structure of a soft material from the nanometer to millimeter length scales is a significant technical challenge. Here, we propose to develop the instrumentation needed for direct 3D structure determination at near nanometer resolution throughout a nearly millimeter-cubed volume of a soft, potentially heterogeneous, material. This new capability will be a valuable research tool for LANL missions in chemistry, materials science, and nanoscience. Our approach to soft materials visualization builds upon exciting developments in super-resolution optical microscopy that have occurred over the past two years. To date, these new, truly revolutionary, imaging methods have been developed and almost exclusively used for biological applications. However, in addition to biological cells, these super-resolution imaging techniques hold extreme promise for direct visualization of many important nanostructured polymers and other heterogeneous chemical systems. Los Alamos has a unique opportunity to lead the development of these super-resolution imaging methods for problems of chemical rather than biological significance. While these optical methods are limited to systems transparent to visible wavelengths, we stress that many important functional chemicals such as polymers, glasses, sol-gels, aerogels, or colloidal assemblies meet this requirement, with specific examples including materials designed for optical communication, manipulation, or light-harvesting Our Research Goals are: (1) Develop the instrumentation necessary for imaging materials

  9. 3D cancer cell migration in a confined matrix

    NASA Astrophysics Data System (ADS)

    Alobaidi, Amani; Sun, Bo

    Cancer cell migration is widely studied in 2D motion, which does not mimic the invasion processes in vivo. More recently, 3D cell migration studies have been performed. The ability of cancer cells to migrate within the extracellular matrix depends on the physical and biochemical features of the extracellular matrix. We present a model of cell motility in confined matrix geometry. The aim of the study is to study cancer migration in collagen matrix, as a soft tissue, to investigate their motility within the confined and surrounding collagen environment. Different collagen concentrations have been used to show the ability of these cancer cells to move through such a complex structure by measuring Cancer cell migration velocity as well as the displacement. Graduate student physics department.

  10. Fully 3D refraction correction dosimetry system.

    PubMed

    Manjappa, Rakesh; Makki, S Sharath; Kumar, Rajesh; Vasu, Ram Mohan; Kanhirodan, Rajan

    2016-02-21

    The irradiation of selective regions in a polymer gel dosimeter results in an increase in optical density and refractive index (RI) at those regions. An optical tomography-based dosimeter depends on rayline path through the dosimeter to estimate and reconstruct the dose distribution. The refraction of light passing through a dose region results in artefacts in the reconstructed images. These refraction errors are dependant on the scanning geometry and collection optics. We developed a fully 3D image reconstruction algorithm, algebraic reconstruction technique-refraction correction (ART-rc) that corrects for the refractive index mismatches present in a gel dosimeter scanner not only at the boundary, but also for any rayline refraction due to multiple dose regions inside the dosimeter. In this study, simulation and experimental studies have been carried out to reconstruct a 3D dose volume using 2D CCD measurements taken for various views. The study also focuses on the effectiveness of using different refractive-index matching media surrounding the gel dosimeter. Since the optical density is assumed to be low for a dosimeter, the filtered backprojection is routinely used for reconstruction. We carry out the reconstructions using conventional algebraic reconstruction (ART) and refractive index corrected ART (ART-rc) algorithms. The reconstructions based on FDK algorithm for cone-beam tomography has also been carried out for comparison. Line scanners and point detectors, are used to obtain reconstructions plane by plane. The rays passing through dose region with a RI mismatch does not reach the detector in the same plane depending on the angle of incidence and RI. In the fully 3D scanning setup using 2D array detectors, light rays that undergo refraction are still collected and hence can still be accounted for in the reconstruction algorithm. It is found that, for the central region of the dosimeter, the usable radius using ART-rc algorithm with water as RI matched

  11. Fully 3D refraction correction dosimetry system

    NASA Astrophysics Data System (ADS)

    Manjappa, Rakesh; Sharath Makki, S.; Kumar, Rajesh; Mohan Vasu, Ram; Kanhirodan, Rajan

    2016-02-01

    The irradiation of selective regions in a polymer gel dosimeter results in an increase in optical density and refractive index (RI) at those regions. An optical tomography-based dosimeter depends on rayline path through the dosimeter to estimate and reconstruct the dose distribution. The refraction of light passing through a dose region results in artefacts in the reconstructed images. These refraction errors are dependant on the scanning geometry and collection optics. We developed a fully 3D image reconstruction algorithm, algebraic reconstruction technique-refraction correction (ART-rc) that corrects for the refractive index mismatches present in a gel dosimeter scanner not only at the boundary, but also for any rayline refraction due to multiple dose regions inside the dosimeter. In this study, simulation and experimental studies have been carried out to reconstruct a 3D dose volume using 2D CCD measurements taken for various views. The study also focuses on the effectiveness of using different refractive-index matching media surrounding the gel dosimeter. Since the optical density is assumed to be low for a dosimeter, the filtered backprojection is routinely used for reconstruction. We carry out the reconstructions using conventional algebraic reconstruction (ART) and refractive index corrected ART (ART-rc) algorithms. The reconstructions based on FDK algorithm for cone-beam tomography has also been carried out for comparison. Line scanners and point detectors, are used to obtain reconstructions plane by plane. The rays passing through dose region with a RI mismatch does not reach the detector in the same plane depending on the angle of incidence and RI. In the fully 3D scanning setup using 2D array detectors, light rays that undergo refraction are still collected and hence can still be accounted for in the reconstruction algorithm. It is found that, for the central region of the dosimeter, the usable radius using ART-rc algorithm with water as RI matched

  12. From 3D view to 3D print

    NASA Astrophysics Data System (ADS)

    Dima, M.; Farisato, G.; Bergomi, M.; Viotto, V.; Magrin, D.; Greggio, D.; Farinato, J.; Marafatto, L.; Ragazzoni, R.; Piazza, D.

    2014-08-01

    In the last few years 3D printing is getting more and more popular and used in many fields going from manufacturing to industrial design, architecture, medical support and aerospace. 3D printing is an evolution of bi-dimensional printing, which allows to obtain a solid object from a 3D model, realized with a 3D modelling software. The final product is obtained using an additive process, in which successive layers of material are laid down one over the other. A 3D printer allows to realize, in a simple way, very complex shapes, which would be quite difficult to be produced with dedicated conventional facilities. Thanks to the fact that the 3D printing is obtained superposing one layer to the others, it doesn't need any particular work flow and it is sufficient to simply draw the model and send it to print. Many different kinds of 3D printers exist based on the technology and material used for layer deposition. A common material used by the toner is ABS plastics, which is a light and rigid thermoplastic polymer, whose peculiar mechanical properties make it diffusely used in several fields, like pipes production and cars interiors manufacturing. I used this technology to create a 1:1 scale model of the telescope which is the hardware core of the space small mission CHEOPS (CHaracterising ExOPlanets Satellite) by ESA, which aims to characterize EXOplanets via transits observations. The telescope has a Ritchey-Chrétien configuration with a 30cm aperture and the launch is foreseen in 2017. In this paper, I present the different phases for the realization of such a model, focusing onto pros and cons of this kind of technology. For example, because of the finite printable volume (10×10×12 inches in the x, y and z directions respectively), it has been necessary to split the largest parts of the instrument in smaller components to be then reassembled and post-processed. A further issue is the resolution of the printed material, which is expressed in terms of layers

  13. Bioengineered collagens

    PubMed Central

    Ramshaw, John AM; Werkmeister, Jerome A; Dumsday, Geoff J

    2014-01-01

    Mammalian collagen has been widely used as a biomedical material. Nevertheless, there are still concerns about the variability between preparations, particularly with the possibility that the products may transmit animal-based diseases. Many groups have examined the possible application of bioengineered mammalian collagens. However, translating laboratory studies into large-scale manufacturing has often proved difficult, although certain yeast and plant systems seem effective. Production of full-length mammalian collagens, with the required secondary modification to give proline hydroxylation, has proved difficult in E. coli. However, recently, a new group of collagens, which have the characteristic triple helical structure of collagen, has been identified in bacteria. These proteins are stable without the need for hydroxyproline and are able to be produced and purified from E. coli in high yield. Initial studies indicate that they would be suitable for biomedical applications. PMID:24717980

  14. Vascular smooth muscle cell response on thin films of collagen.

    PubMed

    Elliott, John T; Woodward, John T; Langenbach, Kurt J; Tona, Alex; Jones, Peter L; Plant, Anne L

    2005-10-01

    Vascular smooth muscle cells (vSMC) cultured on gels of fibrillar type I collagen or denatured collagen (gelatin) comprise a model system that has been widely used for studying the role of the extracellular matrix in vascular diseases such as hypertension, restenosis and athrosclerosis. Despite the wide use of this model system, there are several disadvantages to using collagen gels for cellular studies. These include poor optical characteristics for microscopy, difficulty in verifying that the properties of the preparations are identical from experiment to experiment, heterogeneity within the gels, and difficulty in handling the gels because they are fragile. Previously, we developed an alternative collagen matrix by forming thin films of native fibrillar collagen or denatured collagen on self-assembled monolayers of alkanethiols [Elliott, J.T., Tona, A., Woodward, J., Jones,P., Plant, A., 2003a. Thin films of collagen affect smooth muscle cell morphology. Langmuir 19, 1506-1514.]. These substrates are robust and can be characterized by surface analytical techniques that allow both verification of the reproducibility of the preparation and high-resolution analysis of collagen structure. In addition, they have excellent optical properties that allow more details of the cell-matrix interactions to be observed by microscopy. In this study, we performed a side-by-side structural and functional comparison of collagen gels with thin films of collagen. Our results indicate that vSMC on thin films of collagen are nearly identical to vSMC on thick gels as determined by morphology, proliferation rate, integrin ligation, tenascin-C expression and intracellular signaling events. These results suggest that the features of collagen gels that direct the observed vSMC responses are adequately reconstituted in the thin films of collagen. These thin films will be useful for elucidating the features of the collagen matrix that regulate vSMC response and may be applicable to high

  15. Measurement of cytotoxicity and irritancy potential of sugar-based surfactants on skin-related 3D models.

    PubMed

    Lu, Biao; Miao, Yong; Vigneron, Pascale; Chagnault, Vincent; Grand, Eric; Wadouachi, Anne; Postel, Denis; Pezron, Isabelle; Egles, Christophe; Vayssade, Muriel

    2017-04-01

    Sugar-based surfactants present surface-active properties and relatively low cytotoxicity. They are often considered as safe alternatives to currently used surfactants in cosmetic industries. In this study, four sugar-based surfactants, each with an eight carbon alkyl chain bound to a glucose or a maltose headgroup through an amide linkage, were synthesized and compared to two standard surfactants. The cytotoxic and irritant effects of surfactants were evaluated using two biologically relevant models: 3D dermal model (mouse fibroblasts embedded in collagen gel) and reconstituted human epidermis (RHE, multi-layered human keratinocytes). Results show that three synthesized surfactants possess lower cytotoxicity compared to standard surfactants as demonstrated in the 3D dermal model. Moreover, the IC50s of surfactants against the 3D dermal model are higher than IC50s obtained with the 2D dermal model (monolayer mouse fibroblasts). Both synthesized and standard surfactants show no irritant effects after 48h of topical application on RHE. Throughout the study, we demonstrate the difficulty to link the physico-chemical properties of surfactants and their cytotoxicity in complex models. More importantly, our data suggest that, prior to in vivo tests, a complete understanding of surfactant cytotoxicity or irritancy potential requires a combination of cellular and tissue models.

  16. Extracellular environment contribution to astrogliosis—lessons learned from a tissue engineered 3D model of the glial scar

    PubMed Central

    Rocha, Daniela N.; Ferraz-Nogueira, José P.; Barrias, Cristina C.; Relvas, João B.; Pêgo, Ana P.

    2015-01-01

    Glial scars are widely seen as a (bio)mechanical barrier to central nervous system regeneration. Due to the lack of a screening platform, which could allow in-vitro testing of several variables simultaneously, up to now no comprehensive study has addressed and clarified how different lesion microenvironment properties affect astrogliosis. Using astrocytes cultured in alginate gels and meningeal fibroblast conditioned medium, we have built a simple and reproducible 3D culture system of astrogliosis mimicking many features of the glial scar. Cells in this 3D culture model behave similarly to scar astrocytes, showing changes in gene expression (e.g., GFAP) and increased extra-cellular matrix production (chondroitin 4 sulfate and collagen), inhibiting neuronal outgrowth. This behavior being influenced by the hydrogel network properties. Astrocytic reactivity was found to be dependent on RhoA activity, and targeting RhoA using shRNA-mediated lentivirus reduced astrocytic reactivity. Further, we have shown that chemical inhibition of RhoA with ibuprofen or indirectly targeting RhoA by the induction of extracellular matrix composition modification with chondroitinase ABC, can diminish astrogliosis. Besides presenting the extracellular matrix as a key modulator of astrogliosis, this simple, controlled and reproducible 3D culture system constitutes a good scar-like system and offers great potential in future neurodegenerative mechanism studies, as well as in drug screenings envisaging the development of new therapeutic approaches to minimize the effects of the glial scar in the context of central nervous system disease. PMID:26483632

  17. The structural analysis of three-dimensional fibrous collagen hydrogels by Raman microspectroscopy.

    PubMed

    Hwang, Yu Jer; Lyubovitsky, Julia G

    2013-06-01

    To investigate molecular effects of 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC), EDC/N-hydroxysuccinimide (NHS), glyceraldehyde cross-linking as well as polymerization temperature and concentration on the three-dimensional (3D) collagen hydrogels, we analyzed the structures in situ by Raman microspectroscopy. The increased intensity of the 814 and 936 cm(-1) Raman bands corresponding to the C-C stretch of a protein backbone and a shift in the amide III bands from 1241 cm(-1)/1268 cm(-1) in controls to 1247 cm(-1)/1283 cm(-1) in glyceraldehyde-treated gels indicated changes to the alignment of the collagen molecules, fibrils/fibers and/or changes to the secondary structure on glyceraldehyde treatment. The increased intensity of 1450 cm(-1) band and the appearance of a strong peak at 1468 cm(-1) reflected a change in the motion of lysine/arginine CH2 groups. For the EDC-treated collagen hydrogels, the increased intensity of 823 cm(-1) peak corresponding to the C-C stretch of the protein backbone indicated that EDC also changed the packing of collagen molecules. The 23% decrease in the ratio of 1238 cm(-1) to 1271 cm(-1) amide III band intensities in the EDC-modified samples compared with the controls indicated changes to the alignment of the collagen molecules/fibrils and/or the secondary structure. A change in the motion of lysine/arginine CH2 groups was detected as well. The addition of NHS did not induce additional Raman shifts compared to the effect of EDC alone with the exception of a 1416 cm(-1) band corresponding to a COO(-) stretch. Overall, the Raman spectra suggest that glyceraldehyde affects the collagen states within 3D hydrogels to a greater extent compared to EDC and the effects of temperature and concentration are minimal and/or not detectable.

  18. YouDash3D: exploring stereoscopic 3D gaming for 3D movie theaters

    NASA Astrophysics Data System (ADS)

    Schild, Jonas; Seele, Sven; Masuch, Maic

    2012-03-01

    Along with the success of the digitally revived stereoscopic cinema, events beyond 3D movies become attractive for movie theater operators, i.e. interactive 3D games. In this paper, we present a case that explores possible challenges and solutions for interactive 3D games to be played by a movie theater audience. We analyze the setting and showcase current issues related to lighting and interaction. Our second focus is to provide gameplay mechanics that make special use of stereoscopy, especially depth-based game design. Based on these results, we present YouDash3D, a game prototype that explores public stereoscopic gameplay in a reduced kiosk setup. It features live 3D HD video stream of a professional stereo camera rig rendered in a real-time game scene. We use the effect to place the stereoscopic effigies of players into the digital game. The game showcases how stereoscopic vision can provide for a novel depth-based game mechanic. Projected trigger zones and distributed clusters of the audience video allow for easy adaptation to larger audiences and 3D movie theater gaming.

  19. Speaking Volumes About 3-D

    NASA Technical Reports Server (NTRS)

    2002-01-01

    In 1999, Genex submitted a proposal to Stennis Space Center for a volumetric 3-D display technique that would provide multiple users with a 360-degree perspective to simultaneously view and analyze 3-D data. The futuristic capabilities of the VolumeViewer(R) have offered tremendous benefits to commercial users in the fields of medicine and surgery, air traffic control, pilot training and education, computer-aided design/computer-aided manufacturing, and military/battlefield management. The technology has also helped NASA to better analyze and assess the various data collected by its satellite and spacecraft sensors. Genex capitalized on its success with Stennis by introducing two separate products to the commercial market that incorporate key elements of the 3-D display technology designed under an SBIR contract. The company Rainbow 3D(R) imaging camera is a novel, three-dimensional surface profile measurement system that can obtain a full-frame 3-D image in less than 1 second. The third product is the 360-degree OmniEye(R) video system. Ideal for intrusion detection, surveillance, and situation management, this unique camera system offers a continuous, panoramic view of a scene in real time.

  20. A collagen-poly(lactic acid-co-ɛ-caprolactone) hybrid scaffold for bladder tissue regeneration.

    PubMed

    Engelhardt, Eva-Maria; Micol, Lionel A; Houis, Stephanie; Wurm, Florian M; Hilborn, Jöns; Hubbell, Jeffrey A; Frey, Peter

    2011-06-01

    Scaffold materials should favor cell attachment and proliferation, and provide designable 3D structures with appropriate mechanical strength. Collagen matrices have proven to be beneficial scaffolds for tissue regeneration. However, apart from small intestinal submucosa, they offer a limited mechanical strength even if crosslinking can enhance their mechanical properties. A more cell-friendly way to increase material strength is to combine synthetic polymer meshes with plastic compressed collagen gels. This work describes the potential of plastic compressed collagen-poly(lactic acid-co-ɛ-caprolactone) (PLAC) hybrids as scaffolds for bladder tissue regeneration. Human bladder smooth muscle and urothelial cells were cultured on and inside collagen-PLAC hybrids in vitro. Scaffolds were analyzed by electron microscopy, histology, immunohistochemistry, and AlamarBlue assay. Both cell types proliferated in and on the hybrid, forming dense cell layers on top after two weeks. Furthermore, hybrids were implanted subcutaneously in the backs of nude mice. Host cell infiltration, scaffold degradation, and the presence of the seeded bladder cells were analyzed. Hybrids showed a lower inflammatory reaction in vivo than PLAC meshes alone, and first signs of polymer degradation were visible at six months. Collagen-PLAC hybrids have potential for bladder tissue regeneration, as they show efficient cell seeding, proliferation, and good mechanical properties.

  1. Enhancement of the predicted drug hepatotoxicity in gel entrapped hepatocytes within polysulfone-g-poly (ethylene glycol) modified hollow fiber

    SciTech Connect

    Shen Chong; Zhang Guoliang; Meng Qin

    2010-12-01

    Collagen gel-based 3D cultures of hepatocytes have been proposed for evaluation of drug hepatotoxicity because of their more reliability than traditional monolayer culture. The collagen gel entrapment of hepatocytes in hollow fibers has been proven to well reflect the drug hepatotoxicity in vivo but was limited by adsorption of hydrophobic drugs onto hollow fibers. This study aimed to investigate the impact of hollow fibers on hepatocyte performance and drug hepatotoxicity. Polysulfone-g-poly (ethylene glycol) (PSf-g-PEG) hollow fiber was fabricated and applied for the first time to suppress the drug adsorption. Then, the impact of hollow fibers was evaluated by detecting the hepatotoxicity of eight selected drugs to gel entrapped hepatocytes within PSf and PSf-g-PEG hollow fibers, or without hollow fibers. The hepatocytes in PSf-g-PEG hollow fiber showed the highest sensitivity to drug hepatotoxicity, while those in PSf hollow fiber and cylindrical gel without hollow fiber underestimated the hepatotoxicity due to either drug adsorption or low hepatic functions. Therefore, the 3D culture of gel entrapped hepatocytes within PSf-g-PEG hollow fiber would be a promising tool for investigation of drug hepatotoxicity in vitro.

  2. Enhancement of the predicted drug hepatotoxicity in gel entrapped hepatocytes within polysulfone-g-poly (ethylene glycol) modified hollow fiber.

    PubMed

    Shen, Chong; Zhang, Guoliang; Meng, Qin

    2010-12-01

    Collagen gel-based 3D cultures of hepatocytes have been proposed for evaluation of drug hepatotoxicity because of their more reliability than traditional monolayer culture. The collagen gel entrapment of hepatocytes in hollow fibers has been proven to well reflect the drug hepatotoxicity in vivo but was limited by adsorption of hydrophobic drugs onto hollow fibers. This study aimed to investigate the impact of hollow fibers on hepatocyte performance and drug hepatotoxicity. Polysulfone-g-poly (ethylene glycol) (PSf-g-PEG) hollow fiber was fabricated and applied for the first time to suppress the drug adsorption. Then, the impact of hollow fibers was evaluated by detecting the hepatotoxicity of eight selected drugs to gel entrapped hepatocytes within PSf and PSf-g-PEG hollow fibers, or without hollow fibers. The hepatocytes in PSf-g-PEG hollow fiber showed the highest sensitivity to drug hepatotoxicity, while those in PSf hollow fiber and cylindrical gel without hollow fiber underestimated the hepatotoxicity due to either drug adsorption or low hepatic functions. Therefore, the 3D culture of gel entrapped hepatocytes within PSf-g-PEG hollow fiber would be a promising tool for investigation of drug hepatotoxicity in vitro.

  3. 3D Printed Bionic Nanodevices.

    PubMed

    Kong, Yong Lin; Gupta, Maneesh K; Johnson, Blake N; McAlpine, Michael C

    2016-06-01

    The ability to three-dimensionally interweave biological and functional materials could enable the creation of bionic devices possessing unique and compelling geometries, properties, and functionalities. Indeed, interfacing high performance active devices with biology could impact a variety of fields, including regenerative bioelectronic medicines, smart prosthetics, medical robotics, and human-machine interfaces. Biology, from the molecular scale of DNA and proteins, to the macroscopic scale of tissues and organs, is three-dimensional, often soft and stretchable, and temperature sensitive. This renders most biological platforms incompatible with the fabrication and materials processing methods that have been developed and optimized for functional electronics, which are typically planar, rigid and brittle. A number of strategies have been developed to overcome these dichotomies. One particularly novel approach is the use of extrusion-based multi-material 3D printing, which is an additive manufacturing technology that offers a freeform fabrication strategy. This approach addresses the dichotomies presented above by (1) using 3D printing and imaging for customized, hierarchical, and interwoven device architectures; (2) employing nanotechnology as an enabling route for introducing high performance materials, with the potential for exhibiting properties not found in the bulk; and (3) 3D printing a range of soft and nanoscale materials to enable the integration of a diverse palette of high quality functional nanomaterials with biology. Further, 3D printing is a multi-scale platform, allowing for the incorporation of functional nanoscale inks, the printing of microscale features, and ultimately the creation of macroscale devices. This blending of 3D printing, novel nanomaterial properties, and 'living' platforms may enable next-generation bionic systems. In this review, we highlight this synergistic integration of the unique properties of nanomaterials with the

  4. Petal, terrain & airbags - 3D

    NASA Technical Reports Server (NTRS)

    1997-01-01

    Portions of the lander's deflated airbags and a petal are at the lower area of this image, taken in stereo by the Imager for Mars Pathfinder (IMP) on Sol 3. 3D glasses are necessary to identify surface detail. The metallic object at lower right is part of the lander's low-gain antenna. This image is part of a 3D 'monster

    Click below to see the left and right views individually. [figure removed for brevity, see original site] Left [figure removed for brevity, see original site] Right

  5. 3D Computations and Experiments

    SciTech Connect

    Couch, R; Faux, D; Goto, D; Nikkel, D

    2004-04-05

    This project consists of two activities. Task A, Simulations and Measurements, combines all the material model development and associated numerical work with the materials-oriented experimental activities. The goal of this effort is to provide an improved understanding of dynamic material properties and to provide accurate numerical representations of those properties for use in analysis codes. Task B, ALE3D Development, involves general development activities in the ALE3D code with the focus of improving simulation capabilities for problems of mutual interest to DoD and DOE. Emphasis is on problems involving multi-phase flow, blast loading of structures and system safety/vulnerability studies.

  6. Fabrication of 3D-culture platform with sandwich architecture for preserving liver-specific functions of hepatocytes using 3D bioprinter.

    PubMed

    Arai, Kenichi; Yoshida, Toshiko; Okabe, Motonori; Goto, Mitsuaki; Mir, Tanveer Ahmad; Soko, Chika; Tsukamoto, Yoshinari; Akaike, Toshihiro; Nikaido, Toshio; Zhou, Kaixuan; Nakamura, Makoto

    2016-09-19

    The development of new three-dimensional (3D) cell culture system that maintains the physiologically relevant signals of hepatocytes is essential in drug discovery and tissue engineering research. Conventional two-dimensional (2D) culture yields cell growth, proliferation, and differentiation. However, gene expression and signaling profiles can be different from in vivo environment. Here, we report the fabrication of a 3D culture system using an artificial scaffold and our custom-made inkjet 3D bioprinter as a new strategy for studying liver-specific functions of hepatocytes. We built a 3D culture platform for hepatocytes-attachment and formation of cell monolayer by interacting the galactose chain of galactosylated alginate gel (GA-gel) with asialoglycoprotein receptor (ASGPR) of hepatocytes. The 3D geometrical arrangement of cells was controlled by using 3D bioprinter, and cell polarity was controlled with the galactosylated hydrogels. The fabricated GA-gel was able to successfully promote adhesion of hepatocytes. To observe liver-specific functions and to mimic hepatic cord, an additional parallel layer of hepatocytes was generated using two gel sheets. These results indicated that GA-gel biomimetic matrices can be used as a 3D culture system that could be effective for the engineering of liver tissues. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2016.

  7. 3D printing of natural organic materials by photochemistry

    NASA Astrophysics Data System (ADS)

    Da Silva Gonçalves, Joyce Laura; Valandro, Silvano Rodrigo; Wu, Hsiu-Fen; Lee, Yi-Hsiung; Mettra, Bastien; Monnereau, Cyrille; Schmitt Cavalheiro, Carla Cristina; Pawlicka, Agnieszka; Focsan, Monica; Lin, Chih-Lang; Baldeck, Patrice L.

    2016-03-01

    In previous works, we have used two-photon induced photochemistry to fabricate 3D microstructures based on proteins, anti-bodies, and enzymes for different types of bio-applications. Among them, we can cite collagen lines to guide the movement of living cells, peptide modified GFP biosensing pads to detect Gram positive bacteria, anti-body pads to determine the type of red blood cells, and trypsin columns in a microfluidic channel to obtain a real time biochemical micro-reactor. In this paper, we report for the first time on two-photon 3D microfabrication of DNA material. We also present our preliminary results on using a commercial 3D printer based on a video projector to polymerize slicing layers of gelatine-objects.

  8. The World of 3-D.

    ERIC Educational Resources Information Center

    Mayshark, Robin K.

    1991-01-01

    Students explore three-dimensional properties by creating red and green wall decorations related to Christmas. Students examine why images seem to vibrate when red and green pieces are small and close together. Instructions to conduct the activity and construct 3-D glasses are given. (MDH)

  9. 3D Printing: Exploring Capabilities

    ERIC Educational Resources Information Center

    Samuels, Kyle; Flowers, Jim

    2015-01-01

    As 3D printers become more affordable, schools are using them in increasing numbers. They fit well with the emphasis on product design in technology and engineering education, allowing students to create high-fidelity physical models to see and test different iterations in their product designs. They may also help students to "think in three…

  10. SNL3dFace

    SciTech Connect

    Russ, Trina; Koch, Mark; Koudelka, Melissa; Peters, Ralph; Little, Charles; Boehnen, Chris; Peters, Tanya

    2007-07-20

    This software distribution contains MATLAB and C++ code to enable identity verification using 3D images that may or may not contain a texture component. The code is organized to support system performance testing and system capability demonstration through the proper configuration of the available user interface. Using specific algorithm parameters the face recognition system has been demonstrated to achieve a 96.6% verification rate (Pd) at 0.001 false alarm rate. The system computes robust facial features of a 3D normalized face using Principal Component Analysis (PCA) and Fisher Linear Discriminant Analysis (FLDA). A 3D normalized face is obtained by alighning each face, represented by a set of XYZ coordinated, to a scaled reference face using the Iterative Closest Point (ICP) algorithm. The scaled reference face is then deformed to the input face using an iterative framework with parameters that control the deformed surface regulation an rate of deformation. A variety of options are available to control the information that is encoded by the PCA. Such options include the XYZ coordinates, the difference of each XYZ coordinates from the reference, the Z coordinate, the intensity/texture values, etc. In addition to PCA/FLDA feature projection this software supports feature matching to obtain similarity matrices for performance analysis. In addition, this software supports visualization of the STL, MRD, 2D normalized, and PCA synthetic representations in a 3D environment.

  11. Making Inexpensive 3-D Models

    ERIC Educational Resources Information Center

    Manos, Harry

    2016-01-01

    Visual aids are important to student learning, and they help make the teacher's job easier. Keeping with the "TPT" theme of "The Art, Craft, and Science of Physics Teaching," the purpose of this article is to show how teachers, lacking equipment and funds, can construct a durable 3-D model reference frame and a model gravity…

  12. 3D printing facilitated scaffold-free tissue unit fabrication.

    PubMed

    Tan, Yu; Richards, Dylan J; Trusk, Thomas C; Visconti, Richard P; Yost, Michael J; Kindy, Mark S; Drake, Christopher J; Argraves, William Scott; Markwald, Roger R; Mei, Ying

    2014-06-01

    Tissue spheroids hold great potential in tissue engineering as building blocks to assemble into functional tissues. To date, agarose molds have been extensively used to facilitate fusion process of tissue spheroids. As a molding material, agarose typically requires low temperature plates for gelation and/or heated dispenser units. Here, we proposed and developed an alginate-based, direct 3D mold-printing technology: 3D printing microdroplets of alginate solution into biocompatible, bio-inert alginate hydrogel molds for the fabrication of scaffold-free tissue engineering constructs. Specifically, we developed a 3D printing technology to deposit microdroplets of alginate solution on calcium containing substrates in a layer-by-layer fashion to prepare ring-shaped 3D hydrogel molds. Tissue spheroids composed of 50% endothelial cells and 50% smooth muscle cells were robotically placed into the 3D printed alginate molds using a 3D printer, and were found to rapidly fuse into toroid-shaped tissue units. Histological and immunofluorescence analysis indicated that the cells secreted collagen type I playing a critical role in promoting cell-cell adhesion, tissue formation and maturation.

  13. 3D Printing Facilitated Scaffold-free Tissue Unit Fabrication

    PubMed Central

    Tan, Yu; Richards, Dylan J.; Trusk, Thomas C.; Visconti, Richard P.; Yost, Michael J.; Kindy, Mark S.; Drake, Christopher J.; Argraves, William Scott; Markwald, Roger R.; Mei, Ying

    2014-01-01

    Tissue spheroids hold great potential in tissue engineering as building blocks to assemble into functional tissues. To date, agarose molds have been extensively used to facilitate fusion process of tissue spheroids. As a molding material, agarose typically requires low temperature plates for gelation and/or heated dispenser units. Here, we proposed and developed an alginate-based, direct 3D mold-printing technology: 3D printing micro-droplets of alginate solution into biocompatible, bio-inert alginate hydrogel molds for the fabrication of scaffold-free tissue engineering constructs. Specifically, we developed a 3D printing technology to deposit micro-droplets of alginate solution on calcium containing substrates in a layer-by-layer fashion to prepare ring-shaped 3D hydrogel molds. Tissue spheroids composed of 50% endothelial cells and 50% smooth muscle cells were robotically placed into the 3D printed alginate molds using a 3D printer, and were found to rapidly fuse into toroid-shaped tissue units. Histological and immunofluorescence analysis indicated that the cells secreted collagen type I playing a critical role in promoting cell-cell adhesion, tissue formation and maturation. PMID:24717646

  14. TACO3D. 3-D Finite Element Heat Transfer Code

    SciTech Connect

    Mason, W.E.

    1992-03-04

    TACO3D is a three-dimensional, finite-element program for heat transfer analysis. An extension of the two-dimensional TACO program, it can perform linear and nonlinear analyses and can be used to solve either transient or steady-state problems. The program accepts time-dependent or temperature-dependent material properties, and materials may be isotropic or orthotropic. A variety of time-dependent and temperature-dependent boundary conditions and loadings are available including temperature, flux, convection, and radiation boundary conditions and internal heat generation. Additional specialized features treat enclosure radiation, bulk nodes, and master/slave internal surface conditions (e.g., contact resistance). Data input via a free-field format is provided. A user subprogram feature allows for any type of functional representation of any independent variable. A profile (bandwidth) minimization option is available. The code is limited to implicit time integration for transient solutions. TACO3D has no general mesh generation capability. Rows of evenly-spaced nodes and rows of sequential elements may be generated, but the program relies on separate mesh generators for complex zoning. TACO3D does not have the ability to calculate view factors internally. Graphical representation of data in the form of time history and spatial plots is provided through links to the POSTACO and GRAPE postprocessor codes.

  15. Mandibular Cartilage Collagen Network Nanostructure

    PubMed Central

    Vanden Berg-Foels, Wendy S.

    2015-01-01

    Background Mandibular condyle cartilage (MCC) has a unique structure among articular cartilages; however, little is known about its nanoscale collagen network architecture, hampering design of regeneration therapies and rigorous evaluation of regeneration experiment outcomes in preclinical research. Helium ion microscopy is a novel technology with a long depth of field that is uniquely suited to imaging open 3D collagen networks at multiple scales without obscuring conductive coatings. Objective The objective of this research was to image, at the micro- and nanoscales, the depth-dependent MCC collagen network architecture. Design MCC was collected from New Zealand white rabbits. Images of MCC zones were acquired using helium ion, transmission electron, and light microscopy. Network fibril and canal diameters were measured. Results For the first time, the MCC was visualized as a 3D collagen fibril structure at the nanoscale, the length scale of network assembly. Fibril diameters ranged from 7 to 110 nm and varied by zone. The articular surface was composed of a fine mesh that was woven through thin layers of larger fibrils. The fibrous zone was composed of approximately orthogonal lamellae of aligned fibrils. Fibrocyte processes surrounded collagen bundles forming extracellular compartments. The proliferative, mature, and hypertrophic zones were composed of a branched network that was progressively remodeled to accommodate chondrocyte hypertrophy. Osteoid fibrils were woven around osteoblast cytoplasmic processes to create numerous canals similar in size to canaliculi of mature bone. Conclusion This multiscale investigation advances our foundational understanding of the complex, layered 3D architecture of the MCC collagen network. PMID:27375843

  16. Forensic 3D scene reconstruction

    NASA Astrophysics Data System (ADS)

    Little, Charles Q.; Small, Daniel E.; Peters, Ralph R.; Rigdon, J. B.

    2000-05-01

    Traditionally law enforcement agencies have relied on basic measurement and imaging tools, such as tape measures and cameras, in recording a crime scene. A disadvantage of these methods is that they are slow and cumbersome. The development of a portable system that can rapidly record a crime scene with current camera imaging, 3D geometric surface maps, and contribute quantitative measurements such as accurate relative positioning of crime scene objects, would be an asset to law enforcement agents in collecting and recording significant forensic data. The purpose of this project is to develop a fieldable prototype of a fast, accurate, 3D measurement and imaging system that would support law enforcement agents to quickly document and accurately record a crime scene.

  17. 3D Printed Robotic Hand

    NASA Technical Reports Server (NTRS)

    Pizarro, Yaritzmar Rosario; Schuler, Jason M.; Lippitt, Thomas C.

    2013-01-01

    Dexterous robotic hands are changing the way robots and humans interact and use common tools. Unfortunately, the complexity of the joints and actuations drive up the manufacturing cost. Some cutting edge and commercially available rapid prototyping machines now have the ability to print multiple materials and even combine these materials in the same job. A 3D model of a robotic hand was designed using Creo Parametric 2.0. Combining "hard" and "soft" materials, the model was printed on the Object Connex350 3D printer with the purpose of resembling as much as possible the human appearance and mobility of a real hand while needing no assembly. After printing the prototype, strings where installed as actuators to test mobility. Based on printing materials, the manufacturing cost of the hand was $167, significantly lower than other robotic hands without the actuators since they have more complex assembly processes.

  18. Comparing swimsuits in 3D.

    PubMed

    van Geer, Erik; Molenbroek, Johan; Schreven, Sander; deVoogd-Claessen, Lenneke; Toussaint, Huib

    2012-01-01

    In competitive swimming, suits have become more important. These suits influence friction, pressure and wave drag. Friction drag is related to the surface properties whereas both pressure and wave drag are greatly influenced by body shape. To find a relationship between the body shape and the drag, the anthropometry of several world class female swimmers wearing different suits was accurately defined using a 3D scanner and traditional measuring methods. The 3D scans delivered more detailed information about the body shape. On the same day the swimmers did performance tests in the water with the tested suits. Afterwards the result of the performance tests and the differences found in body shape was analyzed to determine the deformation caused by a swimsuit and its effect on the swimming performance. Although the amount of data is limited because of the few test subjects, there is an indication that the deformation of the body influences the swimming performance.

  19. Forensic 3D Scene Reconstruction

    SciTech Connect

    LITTLE,CHARLES Q.; PETERS,RALPH R.; RIGDON,J. BRIAN; SMALL,DANIEL E.

    1999-10-12

    Traditionally law enforcement agencies have relied on basic measurement and imaging tools, such as tape measures and cameras, in recording a crime scene. A disadvantage of these methods is that they are slow and cumbersome. The development of a portable system that can rapidly record a crime scene with current camera imaging, 3D geometric surface maps, and contribute quantitative measurements such as accurate relative positioning of crime scene objects, would be an asset to law enforcement agents in collecting and recording significant forensic data. The purpose of this project is to develop a feasible prototype of a fast, accurate, 3D measurement and imaging system that would support law enforcement agents to quickly document and accurately record a crime scene.

  20. 3D-graphite structure

    SciTech Connect

    Belenkov, E. A. Ali-Pasha, V. A.

    2011-01-15

    The structure of clusters of some new carbon 3D-graphite phases have been calculated using the molecular-mechanics methods. It is established that 3D-graphite polytypes {alpha}{sub 1,1}, {alpha}{sub 1,3}, {alpha}{sub 1,5}, {alpha}{sub 2,1}, {alpha}{sub 2,3}, {alpha}{sub 3,1}, {beta}{sub 1,2}, {beta}{sub 1,4}, {beta}{sub 1,6}, {beta}{sub 2,1}, and {beta}{sub 3,2} consist of sp{sup 2}-hybridized atoms, have hexagonal unit cells, and differ in regards to the structure of layers and order of their alternation. A possible way to experimentally synthesize new carbon phases is proposed: the polymerization and carbonization of hydrocarbon molecules.

  1. Differential effects of MAPK pathway inhibitors on migration and invasiveness of BRAF(V600E) mutant thyroid cancer cells in 2D and 3D culture.

    PubMed

    Ingeson-Carlsson, Camilla; Martinez-Monleon, Angela; Nilsson, Mikael

    2015-11-01

    Tumor microenvironment influences targeted drug therapy. In this study we compared drug responses to RAF and MEK inhibitors on tumor cell migration in 2D and 3D culture of BRAF(V600E) mutant cell lines derived from human papillary (BCPAP) and anaplastic (SW1736) thyroid carcinomas. Scratch wounding was compared to a double-layered collagen gel model developed for analysis of directed tumor cell invasion during prolonged culture. In BCPAP both PLX4720 and U0126 inhibited growth and migration in 2D and decreased tumor cell survival in 3D. In SW1736 drugs had no effect on migration in 2D but decreased invasion in 3D, however this related to reduced growth. Dual inhibition of BRAF(V600E) and MEK reduced but did not prevent SW1736 invasion although rebound phosphorylation of ERK in response to PLX4720 was blocked by U0126. These findings indicate that anti-tumor drug effects in vitro differ depending on culture conditions (2D vs. 3D) and that the invasive features of anaplastic thyroid cancer depend on non-MEK mechanism(s).

  2. [Real time 3D echocardiography

    NASA Technical Reports Server (NTRS)

    Bauer, F.; Shiota, T.; Thomas, J. D.

    2001-01-01

    Three-dimensional representation of the heart is an old concern. Usually, 3D reconstruction of the cardiac mass is made by successive acquisition of 2D sections, the spatial localisation and orientation of which require complex guiding systems. More recently, the concept of volumetric acquisition has been introduced. A matricial emitter-receiver probe complex with parallel data processing provides instantaneous of a pyramidal 64 degrees x 64 degrees volume. The image is restituted in real time and is composed of 3 planes (planes B and C) which can be displaced in all spatial directions at any time during acquisition. The flexibility of this system of acquisition allows volume and mass measurement with greater accuracy and reproducibility, limiting inter-observer variability. Free navigation of the planes of investigation allows reconstruction for qualitative and quantitative analysis of valvular heart disease and other pathologies. Although real time 3D echocardiography is ready for clinical usage, some improvements are still necessary to improve its conviviality. Then real time 3D echocardiography could be the essential tool for understanding, diagnosis and management of patients.

  3. GPU-Accelerated Denoising in 3D (GD3D)

    SciTech Connect

    2013-10-01

    The raw computational power GPU Accelerators enables fast denoising of 3D MR images using bilateral filtering, anisotropic diffusion, and non-local means. This software addresses two facets of this promising application: what tuning is necessary to achieve optimal performance on a modern GPU? And what parameters yield the best denoising results in practice? To answer the first question, the software performs an autotuning step to empirically determine optimal memory blocking on the GPU. To answer the second, it performs a sweep of algorithm parameters to determine the combination that best reduces the mean squared error relative to a noiseless reference image.

  4. Magmatic Systems in 3-D

    NASA Astrophysics Data System (ADS)

    Kent, G. M.; Harding, A. J.; Babcock, J. M.; Orcutt, J. A.; Bazin, S.; Singh, S.; Detrick, R. S.; Canales, J. P.; Carbotte, S. M.; Diebold, J.

    2002-12-01

    Multichannel seismic (MCS) images of crustal magma chambers are ideal targets for advanced visualization techniques. In the mid-ocean ridge environment, reflections originating at the melt-lens are well separated from other reflection boundaries, such as the seafloor, layer 2A and Moho, which enables the effective use of transparency filters. 3-D visualization of seismic reflectivity falls into two broad categories: volume and surface rendering. Volumetric-based visualization is an extremely powerful approach for the rapid exploration of very dense 3-D datasets. These 3-D datasets are divided into volume elements or voxels, which are individually color coded depending on the assigned datum value; the user can define an opacity filter to reject plotting certain voxels. This transparency allows the user to peer into the data volume, enabling an easy identification of patterns or relationships that might have geologic merit. Multiple image volumes can be co-registered to look at correlations between two different data types (e.g., amplitude variation with offsets studies), in a manner analogous to draping attributes onto a surface. In contrast, surface visualization of seismic reflectivity usually involves producing "fence" diagrams of 2-D seismic profiles that are complemented with seafloor topography, along with point class data, draped lines and vectors (e.g. fault scarps, earthquake locations and plate-motions). The overlying seafloor can be made partially transparent or see-through, enabling 3-D correlations between seafloor structure and seismic reflectivity. Exploration of 3-D datasets requires additional thought when constructing and manipulating these complex objects. As numbers of visual objects grow in a particular scene, there is a tendency to mask overlapping objects; this clutter can be managed through the effective use of total or partial transparency (i.e., alpha-channel). In this way, the co-variation between different datasets can be investigated

  5. Nanoimaging of Focal Adhesion Dynamics in 3D

    PubMed Central

    Chiu, Chi-Li; Aguilar, Jose S.; Tsai, Connie Y.; Wu, GuiKai; Gratton, Enrico; Digman, Michelle A.

    2014-01-01

    Organization and dynamics of focal adhesion proteins have been well characterized in cells grown on two-dimensional (2D) cell culture surfaces. However, much less is known about the dynamic association of these proteins in the 3D microenvironment. Limited imaging technologies capable of measuring protein interactions in real time and space for cells grown in 3D is a major impediment in understanding how proteins function under different environmental cues. In this study, we applied the nano-scale precise imaging by rapid beam oscillation (nSPIRO) technique and combined the scaning-fluorescence correlation spectroscopy (sFCS) and the number and molecular brightness (N&B) methods to investigate paxillin and actin dynamics at focal adhesions in 3D. Both MDA-MB-231 cells and U2OS cells produce elongated protrusions with high intensity regions of paxillin in cell grown in 3D collagen matrices. Using sFCS we found higher percentage of slow diffusing proteins at these focal spots, suggesting assembling/disassembling processes. In addition, the N&B analysis shows paxillin aggregated predominantly at these focal contacts which are next to collagen fibers. At those sites, actin showed slower apparent diffusion rate, which indicated that actin is either polymerizing or binding to the scaffolds in these locals. Our findings demonstrate that by multiplexing these techniques we have the ability to spatially and temporally quantify focal adhesion assembly and disassembly in 3D space and allow the understanding tumor cell invasion in a more complex relevant environment. PMID:24959851

  6. In vitro evaluation of a fibrin gel antibiotic delivery system containing mesenchymal stem cells and vancomycin alginate beads for treating bone infections and facilitating bone formation.

    PubMed

    Hou, Tianyong; Xu, Jianzhong; Li, Qiang; Feng, Jianghua; Zen, Ling

    2008-07-01

    Bone infection and defects are two major problems that occur in the course of treating posttraumatic open bone fractures and osteomyelitis for which local antibiotic delivery is efficacious. Further, hemostasis is an essential treatment after removal of infected bones. Herein we report a new antibiotics delivery system made of vancomycin alginate beads embedded in a fibrin gel (Vanco-AB-FG) to treat bone infections, with the addition of bone marrow-derived mesenchymal stem cells (BMMSCs) seeded in the fibrin gel to promote bone formation. The proliferation of BMMSCs was measured under different conditions of three-dimensional (3D) gel or monolayer, with or without Vanco-AB; cells were labeled by enhanced green fluorescence protein, and their morphology and distribution were observed. The alkaline phosphatase (ALP) activity, real-time RT-PCR, and von Kossa staining were used for determining the osteogenic differentiation of BMMSCs. The concentrations of vancomycin resulting from the antibiotic delivery were determined; the antibiotic activity was evaluated by an assay with standard Staphylococcus aureus (ATCC 25923) as a biological target. The results showed that for Vanco-AB-FG, vancomycin concentrations remained above the breakpoint sensitivity for 22 days. The 3D culture within the gel and the addition of Vanco-AB affected the cell behavior. The morphology of BMMSCs within the 3D gel was different from that in monolayer. The proliferation of the cells within the 3D gel was lower than that in monolayer in early stage, but in later stage the number of BMMSCs in Vanco-AB-FG was similar to that in monolayer. The ALP activity was higher in the 3D gel, and the addition of Vanco-AB slightly increased ALP activity. The osteogenic gene expression levels of ALP, osteopontin, and alpha1 chain of collagen I were higher in the 3D gel than those in monolayer, and additional Vanco-AB could also increase their expression. The von Kossa staining showed that the deposition of

  7. Peptide Directed 3D Assembly of Nanoparticles through Biomolecular Interaction

    NASA Astrophysics Data System (ADS)

    Kaur, Prerna

    The current challenge of the 'bottom up' process is the programmed self-assembly of nanoscale building blocks into complex and larger-scale superstructures with unique properties that can be integrated as components in solar cells, microelectronics, meta materials, catalysis, and sensors. Recent trends in the complexity of device design demand the fabrication of three-dimensional (3D) superstructures from multi-nanomaterial components in precise configurations. Bio mimetic assembly is an emerging technique for building hybrid materials because living organisms are efficient, inexpensive, and environmentally benign material generators, allowing low temperature fabrication. Using this approach, a novel peptide-directed nanomaterial assembly technology based on bio molecular interaction of streptavidin and biotin is presented for assembling nanomaterials with peptides for the construction of 3D peptide-inorganic superlattices with defined 3D shape. We took advantage of robust natural collagen triple-helix peptides and used them as nanowire building blocks for 3D peptide-gold nanoparticles superlattice generation. The type of 3D peptide superlattice assembly with hybrid NP building blocks described herein shows potential for the fabrication of complex functional device which demands precise long-range arrangement and periodicity of NPs.

  8. Microfibrous β-TCP/collagen scaffolds mimic woven bone in structure and composition.

    PubMed

    Zhang, Shen; Zhang, Xin; Cai, Qing; Wang, Bo; Deng, Xuliang; Yang, Xiaoping

    2010-12-01

    Woven bone, as the initial form of bone tissue, is always found in developing and repairing bone. It is thought of as a temporary scaffold for the deposition of osteogenic cells and the laying down of lamellar bone. Thus, we hypothesize that a matrix which resembles the architecture and components of woven bone can provide an osteoblastic microenvironment for bone cell growth and new bone formation. In this study, woven-bone-like beta-tricalcium phosphate (β-TCP)/collagen scaffolds were fabricated by sol-gel electrospinning and impregnating methods. Optimization studies on sol-gel synthesis and electrospinning process were conducted respectively to prepare pure β-TCP fibers with dimensions close to mineralized collagen fibrils in woven bone. The collagen-coating layer prepared by impregnation had an adhesive role that held the β-TCP fibers together, and resulted in rapid degradation and matrix mineralization in in vitro tests. MG63 osteoblast-like cells seeded on the resultant scaffolds showed three-dimensional (3D) morphologies, and merged into multicellular layers after 7 days culture. Cytotoxicity test further revealed that extracts from the resultant scaffolds could promote the proliferation of MG63 cells. Therefore, the woven-bone-like matrix that we constructed favored the attachment and proliferation of MG63 cells in three dimensions. It has great potential ability to shorten the time of formation of new bone.

  9. Interactive 3D Mars Visualization

    NASA Technical Reports Server (NTRS)

    Powell, Mark W.

    2012-01-01

    The Interactive 3D Mars Visualization system provides high-performance, immersive visualization of satellite and surface vehicle imagery of Mars. The software can be used in mission operations to provide the most accurate position information for the Mars rovers to date. When integrated into the mission data pipeline, this system allows mission planners to view the location of the rover on Mars to 0.01-meter accuracy with respect to satellite imagery, with dynamic updates to incorporate the latest position information. Given this information so early in the planning process, rover drivers are able to plan more accurate drive activities for the rover than ever before, increasing the execution of science activities significantly. Scientifically, this 3D mapping information puts all of the science analyses to date into geologic context on a daily basis instead of weeks or months, as was the norm prior to this contribution. This allows the science planners to judge the efficacy of their previously executed science observations much more efficiently, and achieve greater science return as a result. The Interactive 3D Mars surface view is a Mars terrain browsing software interface that encompasses the entire region of exploration for a Mars surface exploration mission. The view is interactive, allowing the user to pan in any direction by clicking and dragging, or to zoom in or out by scrolling the mouse or touchpad. This set currently includes tools for selecting a point of interest, and a ruler tool for displaying the distance between and positions of two points of interest. The mapping information can be harvested and shared through ubiquitous online mapping tools like Google Mars, NASA WorldWind, and Worldwide Telescope.

  10. 3D Nanostructuring of Semiconductors

    NASA Astrophysics Data System (ADS)

    Blick, Robert

    2000-03-01

    Modern semiconductor technology allows to machine devices on the nanometer scale. I will discuss the current limits of the fabrication processes, which enable the definition of single electron transistors with dimensions down to 8 nm. In addition to the conventional 2D patterning and structuring of semiconductors, I will demonstrate how to apply 3D nanostructuring techniques to build freely suspended single-crystal beams with lateral dimension down to 20 nm. In transport measurements in the temperature range from 30 mK up to 100 K these nano-crystals are characterized regarding their electronic as well as their mechanical properties. Moreover, I will present possible applications of these devices.

  11. What Lies Ahead (3-D)

    NASA Technical Reports Server (NTRS)

    2004-01-01

    This 3-D cylindrical-perspective mosaic taken by the navigation camera on the Mars Exploration Rover Spirit on sol 82 shows the view south of the large crater dubbed 'Bonneville.' The rover will travel toward the Columbia Hills, seen here at the upper left. The rock dubbed 'Mazatzal' and the hole the rover drilled in to it can be seen at the lower left. The rover's position is referred to as 'Site 22, Position 32.' This image was geometrically corrected to make the horizon appear flat.

  12. Making Inexpensive 3-D Models

    NASA Astrophysics Data System (ADS)

    Manos, Harry

    2016-03-01

    Visual aids are important to student learning, and they help make the teacher's job easier. Keeping with the TPT theme of "The Art, Craft, and Science of Physics Teaching," the purpose of this article is to show how teachers, lacking equipment and funds, can construct a durable 3-D model reference frame and a model gravity well tailored to specific class lessons. Most of the supplies are readily available in the home or at school: rubbing alcohol, a rag, two colors of spray paint, art brushes, and masking tape. The cost of these supplies, if you don't have them, is less than 20.

  13. A Clean Adirondack (3-D)

    NASA Technical Reports Server (NTRS)

    2004-01-01

    This is a 3-D anaglyph showing a microscopic image taken of an area measuring 3 centimeters (1.2 inches) across on the rock called Adirondack. The image was taken at Gusev Crater on the 33rd day of the Mars Exploration Rover Spirit's journey (Feb. 5, 2004), after the rover used its rock abrasion tool brush to clean the surface of the rock. Dust, which was pushed off to the side during cleaning, can still be seen to the left and in low areas of the rock.

  14. 3D Printed Shelby Cobra

    SciTech Connect

    Love, Lonnie

    2015-01-09

    ORNL's newly printed 3D Shelby Cobra was showcased at the 2015 NAIAS in Detroit. This "laboratory on wheels" uses the Shelby Cobra design, celebrating the 50th anniversary of this model and honoring the first vehicle to be voted a national monument. The Shelby was printed at the Department of Energy’s Manufacturing Demonstration Facility at ORNL using the BAAM (Big Area Additive Manufacturing) machine and is intended as a “plug-n-play” laboratory on wheels. The Shelby will allow research and development of integrated components to be tested and enhanced in real time, improving the use of sustainable, digital manufacturing solutions in the automotive industry.

  15. From 1D to 3D - macroscopic nanowire aerogel monoliths

    NASA Astrophysics Data System (ADS)

    Cheng, Wei; Rechberger, Felix; Niederberger, Markus

    2016-07-01

    Here we present a strategy to assemble one-dimensional nanostructures into a three-dimensional architecture with macroscopic size. With the assistance of centrifugation, we successfully gel ultrathin W18O49 nanowires with diameters of 1 to 2 nm and aspect ratios larger than 100 into 3D networks, which are transformed into monolithic aerogels by supercritical drying.Here we present a strategy to assemble one-dimensional nanostructures into a three-dimensional architecture with macroscopic size. With the assistance of centrifugation, we successfully gel ultrathin W18O49 nanowires with diameters of 1 to 2 nm and aspect ratios larger than 100 into 3D networks, which are transformed into monolithic aerogels by supercritical drying. Electronic supplementary information (ESI) available: Experimental details, SEM and TEM images, and digital photographs. See DOI: 10.1039/c6nr04429h

  16. The inhibition of tube formation in a collagen-fibrinogen, three-dimensional gel by cleaved kininogen (HKa) and HK domain 5 (D5) is dependent on Src family kinases

    SciTech Connect

    Liu Yuchuan; Sainz, Irma M.; Wu Yi; Pixley, Robin; Espinola, Ricardo G.; Hassan, Sarmina; Khan, Mohammad M.; Colman, Robert W.

    2008-02-15

    Cleaved high molecular weight kininogen (HKa), as well as its domain 5 (D5), inhibits migration and proliferation induced by angiogenic factors and induces apoptosis in vitro. To study its effect on tube formation we utilized a collagen-fibrinogen, three-dimensional gel, an in vitro model of angiogenesis. HKa, GST-D5 and D5 had a similar inhibitory effect of tube length by 90 {+-} 4.5%, 86 {+-} 5.5% and 77 {+-} 12.9%, respectively. D5-derived synthetic peptides: G440-H455 H475-H485 and G486-K502 inhibited tube length by 51 {+-} 3.7%, 54 {+-} 3.8% and 77 {+-} 1.7%, respectively. By a comparison of its inhibitory potency and its sequences, a functional sequence of HKa was defined to G486-G496. PP2, a Src family kinase inhibitor, prevented tube formation in a dose-dependent manner (100-400 nM), but PP3 at 5 {mu}M, an inactive analogue of PP2, did not. HKa and D5 inhibited Src 416 phosphorylation by 62 {+-} 12.3% and 83 {+-} 6.1%, respectively. The C-terminal Src kinase (Csk) inhibits Src kinase activity. Using a siRNA to Csk, expression of Csk was down-regulated by 86 {+-} 7.0%, which significantly increased tube length by 27 {+-} 5.8%. The addition of HKa and D5 completely blocked this effect. We further showed that HKa inhibited Src family kinase activity by disrupting the complex of uPAR, {alpha}v{beta}3 integrin and Src. Our results indicate that the anti-angiogenic effect of HKa and D5 is mediated at least in part through Src family kinases and identify a potential novel target for therapeutic inhibition of neovascularization in cancer and inflammatory arthritis.

  17. Positional Awareness Map 3D (PAM3D)

    NASA Technical Reports Server (NTRS)

    Hoffman, Monica; Allen, Earl L.; Yount, John W.; Norcross, April Louise

    2012-01-01

    The Western Aeronautical Test Range of the National Aeronautics and Space Administration s Dryden Flight Research Center needed to address the aging software and hardware of its current situational awareness display application, the Global Real-Time Interactive Map (GRIM). GRIM was initially developed in the late 1980s and executes on older PC architectures using a Linux operating system that is no longer supported. Additionally, the software is difficult to maintain due to its complexity and loss of developer knowledge. It was decided that a replacement application must be developed or acquired in the near future. The replacement must provide the functionality of the original system, the ability to monitor test flight vehicles in real-time, and add improvements such as high resolution imagery and true 3-dimensional capability. This paper will discuss the process of determining the best approach to replace GRIM, and the functionality and capabilities of the first release of the Positional Awareness Map 3D.

  18. Quantitative Multiscale Cell Imaging in Controlled 3D Microenvironments

    PubMed Central

    Welf, Erik S.; Driscoll, Meghan K.; Dean, Kevin M.; Schäfer, Claudia; Chu, Jun; Davidson, Michael W.; Lin, Michael Z.; Danuser, Gaudenz; Fiolka, Reto

    2016-01-01

    The microenvironment determines cell behavior, but the underlying molecular mechanisms are poorly understood because quantitative studies of cell signaling and behavior have been challenging due to insufficient spatial and/or temporal resolution and limitations on microenvironmental control. Here we introduce microenvironmental selective plane illumination microscopy (meSPIM) for imaging and quantification of intracellular signaling and submicrometer cellular structures as well as large-scale cell morphological and environmental features. We demonstrate the utility of this approach by showing that the mechanical properties of the microenvironment regulate the transition of melanoma cells from actin-driven protrusion to blebbing, and we present tools to quantify how cells manipulate individual collagen fibers. We leverage the nearly isotropic resolution of meSPIM to quantify the local concentration of actin and phosphatidylinositol 3-kinase signaling on the surfaces of cells deep within 3D collagen matrices and track the many small membrane protrusions that appear in these more physiologically relevant environments. PMID:26906741

  19. 3D printed bionic ears.

    PubMed

    Mannoor, Manu S; Jiang, Ziwen; James, Teena; Kong, Yong Lin; Malatesta, Karen A; Soboyejo, Winston O; Verma, Naveen; Gracias, David H; McAlpine, Michael C

    2013-06-12

    The ability to three-dimensionally interweave biological tissue with functional electronics could enable the creation of bionic organs possessing enhanced functionalities over their human counterparts. Conventional electronic devices are inherently two-dimensional, preventing seamless multidimensional integration with synthetic biology, as the processes and materials are very different. Here, we present a novel strategy for overcoming these difficulties via additive manufacturing of biological cells with structural and nanoparticle derived electronic elements. As a proof of concept, we generated a bionic ear via 3D printing of a cell-seeded hydrogel matrix in the anatomic geometry of a human ear, along with an intertwined conducting polymer consisting of infused silver nanoparticles. This allowed for in vitro culturing of cartilage tissue around an inductive coil antenna in the ear, which subsequently enables readout of inductively-coupled signals from cochlea-shaped electrodes. The printed ear exhibits enhanced auditory sensing for radio frequency reception, and complementary left and right ears can listen to stereo audio music. Overall, our approach suggests a means to intricately merge biologic and nanoelectronic functionalities via 3D printing.

  20. 3D Printable Graphene Composite

    PubMed Central

    Wei, Xiaojun; Li, Dong; Jiang, Wei; Gu, Zheming; Wang, Xiaojuan; Zhang, Zengxing; Sun, Zhengzong

    2015-01-01

    In human being’s history, both the Iron Age and Silicon Age thrived after a matured massive processing technology was developed. Graphene is the most recent superior material which could potentially initialize another new material Age. However, while being exploited to its full extent, conventional processing methods fail to provide a link to today’s personalization tide. New technology should be ushered in. Three-dimensional (3D) printing fills the missing linkage between graphene materials and the digital mainstream. Their alliance could generate additional stream to push the graphene revolution into a new phase. Here we demonstrate for the first time, a graphene composite, with a graphene loading up to 5.6 wt%, can be 3D printable into computer-designed models. The composite’s linear thermal coefficient is below 75 ppm·°C−1 from room temperature to its glass transition temperature (Tg), which is crucial to build minute thermal stress during the printing process. PMID:26153673

  1. 3D Printed Bionic Ears

    PubMed Central

    Mannoor, Manu S.; Jiang, Ziwen; James, Teena; Kong, Yong Lin; Malatesta, Karen A.; Soboyejo, Winston O.; Verma, Naveen; Gracias, David H.; McAlpine, Michael C.

    2013-01-01

    The ability to three-dimensionally interweave biological tissue with functional electronics could enable the creation of bionic organs possessing enhanced functionalities over their human counterparts. Conventional electronic devices are inherently two-dimensional, preventing seamless multidimensional integration with synthetic biology, as the processes and materials are very different. Here, we present a novel strategy for overcoming these difficulties via additive manufacturing of biological cells with structural and nanoparticle derived electronic elements. As a proof of concept, we generated a bionic ear via 3D printing of a cell-seeded hydrogel matrix in the precise anatomic geometry of a human ear, along with an intertwined conducting polymer consisting of infused silver nanoparticles. This allowed for in vitro culturing of cartilage tissue around an inductive coil antenna in the ear, which subsequently enables readout of inductively-coupled signals from cochlea-shaped electrodes. The printed ear exhibits enhanced auditory sensing for radio frequency reception, and complementary left and right ears can listen to stereo audio music. Overall, our approach suggests a means to intricately merge biologic and nanoelectronic functionalities via 3D printing. PMID:23635097

  2. Martian terrain & airbags - 3D

    NASA Technical Reports Server (NTRS)

    1997-01-01

    Portions of the lander's deflated airbags and a petal are at lower left in this image, taken in stereo by the Imager for Mars Pathfinder (IMP) on Sol 3. 3D glasses are necessary to identify surface detail. This image is part of a 3D 'monster' panorama of the area surrounding the landing site.

    Mars Pathfinder is the second in NASA's Discovery program of low-cost spacecraft with highly focused science goals. The Jet Propulsion Laboratory, Pasadena, CA, developed and manages the Mars Pathfinder mission for NASA's Office of Space Science, Washington, D.C. JPL is an operating division of the California Institute of Technology (Caltech). The Imager for Mars Pathfinder (IMP) was developed by the University of Arizona Lunar and Planetary Laboratory under contract to JPL. Peter Smith is the Principal Investigator.

    Click below to see the left and right views individually. [figure removed for brevity, see original site] Left [figure removed for brevity, see original site] Right

  3. Martian terrain & airbags - 3D

    NASA Technical Reports Server (NTRS)

    1997-01-01

    Portions of the lander's deflated airbags and a petal are at the lower area of this image, taken in stereo by the Imager for Mars Pathfinder (IMP) on Sol 3. 3D glasses are necessary to identify surface detail. This image is part of a 3D 'monster' panorama of the area surrounding the landing site.

    Mars Pathfinder is the second in NASA's Discovery program of low-cost spacecraft with highly focused science goals. The Jet Propulsion Laboratory, Pasadena, CA, developed and manages the Mars Pathfinder mission for NASA's Office of Space Science, Washington, D.C. JPL is an operating division of the California Institute of Technology (Caltech). The Imager for Mars Pathfinder (IMP) was developed by the University of Arizona Lunar and Planetary Laboratory under contract to JPL. Peter Smith is the Principal Investigator.

    Click below to see the left and right views individually. [figure removed for brevity, see original site] Left [figure removed for brevity, see original site] Right

  4. 3D structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Dougherty, William M.; Goodwin, Paul C.

    2011-03-01

    Three-dimensional structured illumination microscopy achieves double the lateral and axial resolution of wide-field microscopy, using conventional fluorescent dyes, proteins and sample preparation techniques. A three-dimensional interference-fringe pattern excites the fluorescence, filling in the "missing cone" of the wide field optical transfer function, thereby enabling axial (z) discrimination. The pattern acts as a spatial carrier frequency that mixes with the higher spatial frequency components of the image, which usually succumb to the diffraction limit. The fluorescence image encodes the high frequency content as a down-mixed, moiré-like pattern. A series of images is required, wherein the 3D pattern is shifted and rotated, providing down-mixed data for a system of linear equations. Super-resolution is obtained by solving these equations. The speed with which the image series can be obtained can be a problem for the microscopy of living cells. Challenges include pattern-switching speeds, optical efficiency, wavefront quality and fringe contrast, fringe pitch optimization, and polarization issues. We will review some recent developments in 3D-SIM hardware with the goal of super-resolved z-stacks of motile cells.

  5. Individual versus collective fibroblast spreading and migration: regulation by matrix composition in 3D culture.

    PubMed

    Miron-Mendoza, Miguel; Lin, Xihui; Ma, Lisha; Ririe, Peter; Petroll, W Matthew

    2012-06-01

    Extracellular matrix (ECM) supplies both physical and chemical signals to cells and provides a substrate through which fibroblasts migrate during wound repair. To directly assess how ECM composition regulates this process, we used a nested 3D matrix model in which cell-populated collagen buttons were embedded in cell-free collagen or fibrin matrices. Time-lapse microscopy was used to record the dynamic pattern of cell migration into the outer matrices, and 3D confocal imaging was used to assess cell connectivity and cytoskeletal organization. Corneal fibroblasts stimulated with PDGF migrated more rapidly into collagen as compared to fibrin. In addition, the pattern of fibroblast migration into fibrin and collagen ECMs was strikingly different. Corneal fibroblasts migrating into collagen matrices developed dendritic processes and moved independently, whereas cells migrating into fibrin matrices had a more fusiform morphology and formed an interconnected meshwork. A similar pattern was observed when using dermal fibroblasts, suggesting that this response is not unique to corneal cells. We next cultured corneal fibroblasts within and on top of standard collagen and fibrin matrices to assess the impact of ECM composition on the cell spreading response. Similar differences in cell morphology and connectivity were observed – cells remained separated on collagen but coalesced into clusters on fibrin. Cadherin was localized to junctions between interconnected cells, whereas fibronectin was present both between cells and at the tips of extending cell processes. Cells on fibrin matrices also developed more prominent stress fibers than those on collagen matrices. Importantly, these spreading and migration patterns were consistently observed on both rigid and compliant substrates, thus differences in ECM mechanical stiffness were not the underlying cause. Overall, these results demonstrate for the first time that ECM protein composition alone (collagen vs. fibrin) can induce

  6. Collagenous colitis.

    PubMed Central

    Kingham, J G; Levison, D A; Morson, B C; Dawson, A M

    1986-01-01

    Clinical and pathological aspects of six patients with collagenous colitis are presented. These patients have been observed for between four and 15 years and the evolution of the condition is documented in three (cases 1, 3 and 5). Management and possible pathogenetic mechanisms of this enigmatic condition are discussed. Images Fig. 1 Fig. 2 PMID:3699567

  7. Collagenous gastritis.

    PubMed

    Jin, Xiaoyi; Koike, Tomoyuki; Chiba, Takashi; Kondo, Yutaka; Ara, Nobuyuki; Uno, Kaname; Asano, Naoki; Iijima, Katsunori; Imatani, Akira; Watanabe, Mika; Shirane, Akio; Shimosegawa, Tooru

    2013-09-01

    In the present paper, we report a case of rare collagenous gastritis. The patient was a 25-year-old man who had experienced nausea, abdominal distention and epigastralgia since 2005. Esophagogastroduodenoscopy (EGD) carried out at initial examination by the patient's local doctor revealed an extensively discolored depression from the upper gastric body to the lower gastric body, mainly including the greater curvature, accompanied by residual mucosa with multiple islands and nodularity with a cobblestone appearance. Initial biopsies sampled from the nodules and accompanying atrophic mucosa were diagnosed as chronic gastritis. In August, 2011, the patient was referred to Tohoku University Hospital for observation and treatment. EGD at our hospital showed the same findings as those by the patient's local doctor. Pathological findings included a membranous collagen band in the superficial layer area of the gastric mucosa, which led to a diagnosis of collagenous gastritis. Collagenous gastritis is an extremely rare disease, but it is important to recognize its characteristic endoscopic findings to make a diagnosis.

  8. Myosin IIA dependent retrograde flow drives 3D cell migration.

    PubMed

    Shih, Wenting; Yamada, Soichiro

    2010-04-21

    Epithelial cell migration is an essential part of embryogenesis and tissue regeneration, yet their migration is least understood. Using our three-dimensional (3D) motility analysis, migrating epithelial cells formed an atypical polarized cell shape with the nucleus leading the cell front and a contractile cell rear. Migrating epithelial cells exerted traction forces to deform both the anterior and posterior extracellular matrix toward the cell body. The cell leading edge exhibited a myosin II-dependent retrograde flow with the magnitude and direction consistent with surrounding network deformation. Interestingly, on a two-dimensional substrate, myosin IIA-deficient cells migrated faster than wild-type cells, but in a 3D gel, these myosin IIA-deficient cells were unpolarized and immobile. In contrast, the migration rates of myosin IIB-deficient cells were similar to wild-type cells. Therefore, myosin IIA, not myosin IIB, is required for 3D epithelial cell migration.

  9. A software tool for 3D dose verification and analysis

    NASA Astrophysics Data System (ADS)

    Sa'd, M. Al; Graham, J.; Liney, G. P.

    2013-06-01

    The main recent developments in radiotherapy have focused on improved treatment techniques in order to generate further significant improvements in patient prognosis. There is now an internationally recognised need to improve 3D verification of highly conformal radiotherapy treatments. This is because of the very high dose gradients used in modern treatment techniques, which can result in a small error in the spatial dose distribution leading to a serious complication. In order to gain the full benefits of using 3D dosimetric technologies (such as gel dosimetry), it is vital to use 3D evaluation methods and algorithms. We present in this paper a software solution that provides a comprehensive 3D dose evaluation and analysis. The software is applied to gel dosimetry, which is based on magnetic resonance imaging (MRI) as a read-out method. The software can also be used to compare any two dose distributions, such as two distributions planned using different methods of treatment planning systems, or different dose calculation algorithms.

  10. 3D Printing of Graphene Aerogels.

    PubMed

    Zhang, Qiangqiang; Zhang, Feng; Medarametla, Sai Pradeep; Li, Hui; Zhou, Chi; Lin, Dong

    2016-04-06

    3D printing of a graphene aerogel with true 3D overhang structures is highlighted. The aerogel is fabricated by combining drop-on-demand 3D printing and freeze casting. The water-based GO ink is ejected and freeze-cast into designed 3D structures. The lightweight (<10 mg cm(-3) ) 3D printed graphene aerogel presents superelastic and high electrical conduction.

  11. Quasi 3D dispersion experiment

    NASA Astrophysics Data System (ADS)

    Bakucz, P.

    2003-04-01

    This paper studies the problem of tracer dispersion in a coloured fluid flowing through a two-phase 3D rough channel-system in a 40 cm*40 cm plexi-container filled by homogen glass fractions and colourless fluid. The unstable interface between the driving coloured fluid and the colourless fluid develops viscous fingers with a fractal structure at high capillary number. Five two-dimensional fractal fronts have been observed at the same time using four cameras along the vertical side-walls and using one camera located above the plexi-container. In possession of five fronts the spatial concentration contours are determined using statistical models. The concentration contours are self-affine fractal curves with a fractal dimension D=2.19. This result is valid for disperison at high Péclet numbers.

  12. ShowMe3D

    SciTech Connect

    Sinclair, Michael B

    2012-01-05

    ShowMe3D is a data visualization graphical user interface specifically designed for use with hyperspectral image obtained from the Hyperspectral Confocal Microscope. The program allows the user to select and display any single image from a three dimensional hyperspectral image stack. By moving a slider control, the user can easily move between images of the stack. The user can zoom into any region of the image. The user can select any pixel or region from the displayed image and display the fluorescence spectrum associated with that pixel or region. The user can define up to 3 spectral filters to apply to the hyperspectral image and view the image as it would appear from a filter-based confocal microscope. The user can also obtain statistics such as intensity average and variance from selected regions.

  13. 3D Printed Shelby Cobra

    ScienceCinema

    Love, Lonnie

    2016-11-02

    ORNL's newly printed 3D Shelby Cobra was showcased at the 2015 NAIAS in Detroit. This "laboratory on wheels" uses the Shelby Cobra design, celebrating the 50th anniversary of this model and honoring the first vehicle to be voted a national monument. The Shelby was printed at the Department of Energy’s Manufacturing Demonstration Facility at ORNL using the BAAM (Big Area Additive Manufacturing) machine and is intended as a “plug-n-play” laboratory on wheels. The Shelby will allow research and development of integrated components to be tested and enhanced in real time, improving the use of sustainable, digital manufacturing solutions in the automotive industry.

  14. Supernova Remnant in 3-D

    NASA Technical Reports Server (NTRS)

    2009-01-01

    wavelengths. Since the amount of the wavelength shift is related to the speed of motion, one can determine how fast the debris are moving in either direction. Because Cas A is the result of an explosion, the stellar debris is expanding radially outwards from the explosion center. Using simple geometry, the scientists were able to construct a 3-D model using all of this information. A program called 3-D Slicer modified for astronomical use by the Astronomical Medicine Project at Harvard University in Cambridge, Mass. was used to display and manipulate the 3-D model. Commercial software was then used to create the 3-D fly-through.

    The blue filaments defining the blast wave were not mapped using the Doppler effect because they emit a different kind of light synchrotron radiation that does not emit light at discrete wavelengths, but rather in a broad continuum. The blue filaments are only a representation of the actual filaments observed at the blast wave.

    This visualization shows that there are two main components to this supernova remnant: a spherical component in the outer parts of the remnant and a flattened (disk-like) component in the inner region. The spherical component consists of the outer layer of the star that exploded, probably made of helium and carbon. These layers drove a spherical blast wave into the diffuse gas surrounding the star. The flattened component that astronomers were unable to map into 3-D prior to these Spitzer observations consists of the inner layers of the star. It is made from various heavier elements, not all shown in the visualization, such as oxygen, neon, silicon, sulphur, argon and iron.

    High-velocity plumes, or jets, of this material are shooting out from the explosion in the plane of the disk-like component mentioned above. Plumes of silicon appear in the northeast and southwest, while those of iron are seen in the southeast and north. These jets were already known and Doppler velocity measurements have been made for these

  15. MO-F-CAMPUS-I-04: Magnetic Resonance Imaging of An in Vitro 3D Tumor Model

    SciTech Connect

    Veiga, C; Long, T; Siow, B; Loizidou, M; Royle, G; Ricketts, K

    2015-06-15

    Purpose: To investigate the use of an in vitro 3D tumor model (tumoroid) as a bio-phantom for repetitive and sequential magnetic resonance imaging (MRI) studies. Methods: The tissue engineered tumoroid comprised an artificial cancer mass (ACM) containing 30 million HT29 cancer cells seeded in a collagen type I matrix, whose density was increased by plastic compression (dry/wet weight=40%). The ACM was embedded in an uncompressed collagen gel that mimicked the tumor stroma, and the tumoroid was incubated for 24h before imaging. Images were acquired using the 1T ICON™ (Bruker Corporation, Billerica, MA) MRI scanner. T1 maps were calculated using an IR-RARE sequence (TE=12ms, TR=10000ms, 7 inversion times), while for T2 maps a MSME technique (TR=6000ms, 16 echoes) was used. T1 and T2 fittings were performed using a pixel-wise approach to produce relaxometric parametric maps. Results: The images acquired and corresponding T1 and T2 maps indicate contrast between the ACM and the stroma. T1 was 2500 and 2800ms, while T2 was 520 and 760ms, for the ACM and stroma respectively. The ACM construct was not homogenous and internal features were visible, which can be explained by local gradients of cell and/or collagen density. The viability of the cells was confirmed via confocal microscopy for several days after the imaging session, demonstrating the suitability of the tumoroid for sequential imaging studies. Conclusions: We have engineered a tumor model compatible with repetitive and sequential MRI. We found T1 and T2 contrast between the ACM and stroma using a pre-clinical MRI scanner. The model, which enables controllable cell and matrix densities, has potential for a wide range of applications in radiotherapy, such as to study tumor progression and to validate imaging biomarkers. Further work is necessary to understand the mechanisms behind the contrast achieved, and to correlate findings with biology and histology data.

  16. Collective epithelial cell invasion overcomes mechanical barriers of collagenous extracellular matrix by a narrow tube-like geometry and MMP14-dependent local softening†

    PubMed Central

    Alcaraz, Jordi; Mori, Hidetoshi; Ghajar, Cyrus M.; Brownfield, Doug; Galgoczy, Roland; Bissell, Mina J.

    2013-01-01

    Collective cell invasion (CCI) through interstitial collagenous extracellular matrix (ECM) is crucial to the initial stages of branching morphogenesis, and a hallmark of tissue repair and dissemination of certain tumors. The collagenous ECM acts as a mechanical barrier against CCI. However, the physical nature of this barrier and how it is overcome by cells remains incompletely understood. To address these questions, we performed theoretical and experimental analysis of mammary epithelial branching morphogenesis in 3D type I collagen (collagen-I) gels. We found that the mechanical resistance of collagen-I is largely due to its elastic rather than its viscous properties. We also identified two strategies utilized by mammary epithelial cells that can independently minimize ECM mechanical resistance during CCI. First, cells adopt a narrow tube-like geometry during invasion, which minimizes the elastic opposition from the ECM as revealed by theoretical modeling of the most frequent invasive shapes and sizes. Second, the stiffness of the collagenous ECM is reduced at invasive fronts due to its degradation by matrix metalloproteinases (MMPs), as indicated by direct measurements of collagen-I microelasticity by atomic force microscopy. Molecular techniques further specified that the membrane-bound MMP14 mediates degradation of collagen-I at invasive fronts. Thus, our findings reveal that MMP14 is necessary to efficiently reduce the physical restraints imposed by collagen-I during branching morphogenesis, and help our overall understanding of how forces are balanced between cells and their surrounding ECM to maintain collective geometry and mechanical stability during CCI. PMID:21993836

  17. Structure of collagen-glycosaminoglycan matrix and the influence to its integrity and stability.

    PubMed

    Bi, Yuying; Patra, Prabir; Faezipour, Miad

    2014-01-01

    Glycosaminoglycan (GAG) is a chain-like disaccharide that is linked to polypeptide core to connect two collagen fibrils/fibers and provide the intermolecular force in Collagen-GAG matrix (C-G matrix). Thus, the distribution of GAG in C-G matrix contributes to the integrity and mechanical properties of the matrix and related tissue. This paper analyzes the transverse isotropic distribution of GAG in C-G matrix. The angle of GAGs related to collagen fibrils is used as parameters to qualify the GAGs isotropic characteristic in both 3D and 2D rendering. Statistical results included that over one third of GAGs were perpendicular directed to collagen fibril with symmetrical distribution for both 3D matrix and 2D plane cross through collagen fibrils. The three factors tested in this paper: collagen radius, collagen distribution, and GAGs density, were not statistically significant for the strength of Collagen-GAG matrix in 3D rendering. However in 2D rendering, a significant factor found was the radius of collagen in matrix for the GAGs directed to orthogonal plane of Collagen-GAG matrix. Between two cross-section selected from Collagen-GAG matrix model, the plane cross through collagen fibrils was symmetrically distributed but the total percentage of perpendicular directed GAG was deducted by decreasing collagen radius. There were some symmetry features of GAGs angle distribution in selected 2D plane that passed through space between collagen fibrils, but most models showed multiple peaks in GAGs angle distribution. With less GAGs directed to perpendicular of collagen fibril, strength in collagen cross-section weakened. Collagen distribution was also a factor that influences GAGs angle distribution in 2D rendering. True hexagonal collagen packaging is reported in this paper to have less strength at collagen cross-section compared to quasi-hexagonal collagen arrangement. In this work focus is on GAGs matrix within the collagen and its relevance to anisotropy.

  18. Composite alginate gels for tunable cellular microenvironment mechanics

    PubMed Central

    Khavari, Adele; Nydén, Magnus; Weitz, David A.; Ehrlicher, Allen J.

    2016-01-01

    The mechanics of the cellular microenvironment can be as critical as biochemistry in directing cell behavior. Many commonly utilized materials derived from extra-cellular-matrix create excellent scaffolds for cell growth, however, evaluating the relative mechanical and biochemical effects independently in 3D environments has been difficult in frequently used biopolymer matrices. Here we present 3D sodium alginate hydrogel microenvironments over a physiological range of stiffness (E = 1.85 to 5.29 kPa), with and without RGD binding sites or collagen fibers. We use confocal microscopy to measure the growth of multi-cellular aggregates (MCAs), of increasing metastatic potential in different elastic moduli of hydrogels, with and without binding factors. We find that the hydrogel stiffness regulates the growth and morphology of these cell clusters; MCAs grow larger and faster in the more rigid environments similar to cancerous breast tissue (E = 4–12 kPa) as compared to healthy tissue (E = 0.4–2 kpa). Adding binding factors from collagen and RGD peptides increases growth rates, and change maximum MCA sizes. These findings demonstrate the utility of these independently tunable mechanical/biochemistry gels, and that mechanical confinement in stiffer microenvironments may increase cell proliferation. PMID:27484403

  19. Composite alginate gels for tunable cellular microenvironment mechanics

    NASA Astrophysics Data System (ADS)

    Khavari, Adele; Nydén, Magnus; Weitz, David A.; Ehrlicher, Allen J.

    2016-08-01

    The mechanics of the cellular microenvironment can be as critical as biochemistry in directing cell behavior. Many commonly utilized materials derived from extra-cellular-matrix create excellent scaffolds for cell growth, however, evaluating the relative mechanical and biochemical effects independently in 3D environments has been difficult in frequently used biopolymer matrices. Here we present 3D sodium alginate hydrogel microenvironments over a physiological range of stiffness (E = 1.85 to 5.29 kPa), with and without RGD binding sites or collagen fibers. We use confocal microscopy to measure the growth of multi-cellular aggregates (MCAs), of increasing metastatic potential in different elastic moduli of hydrogels, with and without binding factors. We find that the hydrogel stiffness regulates the growth and morphology of these cell clusters; MCAs grow larger and faster in the more rigid environments similar to cancerous breast tissue (E = 4–12 kPa) as compared to healthy tissue (E = 0.4–2 kpa). Adding binding factors from collagen and RGD peptides increases growth rates, and change maximum MCA sizes. These findings demonstrate the utility of these independently tunable mechanical/biochemistry gels, and that mechanical confinement in stiffer microenvironments may increase cell proliferation.

  20. 3D Kitaev spin liquids

    NASA Astrophysics Data System (ADS)

    Hermanns, Maria

    The Kitaev honeycomb model has become one of the archetypal spin models exhibiting topological phases of matter, where the magnetic moments fractionalize into Majorana fermions interacting with a Z2 gauge field. In this talk, we discuss generalizations of this model to three-dimensional lattice structures. Our main focus is the metallic state that the emergent Majorana fermions form. In particular, we discuss the relation of the nature of this Majorana metal to the details of the underlying lattice structure. Besides (almost) conventional metals with a Majorana Fermi surface, one also finds various realizations of Dirac semi-metals, where the gapless modes form Fermi lines or even Weyl nodes. We introduce a general classification of these gapless quantum spin liquids using projective symmetry analysis. Furthermore, we briefly outline why these Majorana metals in 3D Kitaev systems provide an even richer variety of Dirac and Weyl phases than possible for electronic matter and comment on possible experimental signatures. Work done in collaboration with Kevin O'Brien and Simon Trebst.

  1. 3D multiplexed immunoplasmonics microscopy

    NASA Astrophysics Data System (ADS)

    Bergeron, Éric; Patskovsky, Sergiy; Rioux, David; Meunier, Michel

    2016-07-01

    Selective labelling, identification and spatial distribution of cell surface biomarkers can provide important clinical information, such as distinction between healthy and diseased cells, evolution of a disease and selection of the optimal patient-specific treatment. Immunofluorescence is the gold standard for efficient detection of biomarkers expressed by cells. However, antibodies (Abs) conjugated to fluorescent dyes remain limited by their photobleaching, high sensitivity to the environment, low light intensity, and wide absorption and emission spectra. Immunoplasmonics is a novel microscopy method based on the visualization of Abs-functionalized plasmonic nanoparticles (fNPs) targeting cell surface biomarkers. Tunable fNPs should provide higher multiplexing capacity than immunofluorescence since NPs are photostable over time, strongly scatter light at their plasmon peak wavelengths and can be easily functionalized. In this article, we experimentally demonstrate accurate multiplexed detection based on the immunoplasmonics approach. First, we achieve the selective labelling of three targeted cell surface biomarkers (cluster of differentiation 44 (CD44), epidermal growth factor receptor (EGFR) and voltage-gated K+ channel subunit KV1.1) on human cancer CD44+ EGFR+ KV1.1+ MDA-MB-231 cells and reference CD44- EGFR- KV1.1+ 661W cells. The labelling efficiency with three stable specific immunoplasmonics labels (functionalized silver nanospheres (CD44-AgNSs), gold (Au) NSs (EGFR-AuNSs) and Au nanorods (KV1.1-AuNRs)) detected by reflected light microscopy (RLM) is similar to the one with immunofluorescence. Second, we introduce an improved method for 3D localization and spectral identification of fNPs based on fast z-scanning by RLM with three spectral filters corresponding to the plasmon peak wavelengths of the immunoplasmonics labels in the cellular environment (500 nm for 80 nm AgNSs, 580 nm for 100 nm AuNSs and 700 nm for 40 nm × 92 nm AuNRs). Third, the developed

  2. Crowdsourcing Based 3d Modeling

    NASA Astrophysics Data System (ADS)

    Somogyi, A.; Barsi, A.; Molnar, B.; Lovas, T.

    2016-06-01

    Web-based photo albums that support organizing and viewing the users' images are widely used. These services provide a convenient solution for storing, editing and sharing images. In many cases, the users attach geotags to the images in order to enable using them e.g. in location based applications on social networks. Our paper discusses a procedure that collects open access images from a site frequently visited by tourists. Geotagged pictures showing the image of a sight or tourist attraction are selected and processed in photogrammetric processing software that produces the 3D model of the captured object. For the particular investigation we selected three attractions in Budapest. To assess the geometrical accuracy, we used laser scanner and DSLR as well as smart phone photography to derive reference values to enable verifying the spatial model obtained from the web-album images. The investigation shows how detailed and accurate models could be derived applying photogrammetric processing software, simply by using images of the community, without visiting the site.

  3. Drug-releasing nano-engineered titanium implants: therapeutic efficacy in 3D cell culture model, controlled release and stability.

    PubMed

    Gulati, Karan; Kogawa, Masakazu; Prideaux, Matthew; Findlay, David M; Atkins, Gerald J; Losic, Dusan

    2016-12-01

    There is an ongoing demand for new approaches for treating localized bone pathologies. Here we propose a new strategy for treatment of such conditions, via local delivery of hormones/drugs to the trauma site using drug releasing nano-engineered implants. The proposed implants were prepared in the form of small Ti wires/needles with a nano-engineered oxide layer composed of array of titania nanotubes (TNTs). TNTs implants were inserted into a 3D collagen gel matrix containing human osteoblast-like, and the results confirmed cell migration onto the implants and their attachment and spread. To investigate therapeutic efficacy, TNTs/Ti wires loaded with parathyroid hormone (PTH), an approved anabolic therapeutic for the treatment of severe bone fractures, were inserted into 3D gels containing osteoblast-like cells. Gene expression studies revealed a suppression of SOST (sclerostin) and an increase in RANKL (receptor activator of nuclear factor kappa-B ligand) mRNA expression, confirming the release of PTH from TNTs at concentrations sufficient to alter cell function. The performance of the TNTs wire implants using an example of a drug needed at relatively higher concentrations, the anti-inflammatory drug indomethacin, is also demonstrated. Finally, the mechanical stability of the prepared implants was tested by their insertion into bovine trabecular bone cores ex vivo followed by retrieval, which confirmed the robustness of the TNT structures. This study provides proof of principle for the suitability of the TNT/Ti wire implants for localized bone therapy, which can be customized to cater for specific therapeutic requirements.

  4. Probing multiscale mechanics of collagen with optical tweezers

    NASA Astrophysics Data System (ADS)

    Shayegan, Marjan; Rezaei, Naghmeh; Lam, Norman H.; Altindal, Tuba; Wieczorek, Andrew; Forde, Nancy R.

    2013-09-01

    How the molecular structure of the structural, extracellular matrix protein collagen correlates with its mechanical properties at different hierarchical structural levels is not known. We demonstrate the utility of optical tweezers to probe collagen's mechanical response throughout its assembly hierarchy, from single molecule force-extension measurements through microrheology measurements on solutions of collagen molecules, collagen fibrillar gels and gelatin. These experiments enable the determination of collagen's flexibility, mechanics, and timescales and strengths of interaction at different levels of hierarchy, information critical to developing models of how collagen's physiological function and stability are influenced by its chemical composition. By investigating how the viscoelastic properties of collagen are affected by the presence of telopeptides, protein domains that strongly influence fibril formation, we demonstrate that these play a role in conferring transient elasticity to collagen solutions.

  5. SU-E-T-231: Cross-Validation of 3D Gamma Comparison Tools

    SciTech Connect

    Alexander, KM; Jechel, C; Pinter, C; Lasso, A; Fichtinger, G; Salomons, G; Schreiner, LJ

    2015-06-15

    Purpose: Moving the computational analysis for 3D gel dosimetry into the 3D Slicer (www.slicer.org) environment has made gel dosimetry more clinically accessible. To ensure accuracy, we cross-validate the 3D gamma comparison module in 3D Slicer with an independently developed algorithm using simulated and measured dose distributions. Methods: Two reference dose distributions were generated using the Varian Eclipse treatment planning system. The first distribution consisted of a four-field box irradiation delivered to a plastic water phantom and the second, a VMAT plan delivered to a gel dosimeter phantom. The first reference distribution was modified within Eclipse to create an evaluated dose distribution by spatially shifting one field by 3mm, increasing the monitor units of the second field, applying a dynamic wedge for the third field, and leaving the fourth field unchanged. The VMAT plan was delivered to a gel dosimeter and the evaluated dose in the gel was calculated from optical CT measurements. Results from the gamma comparison tool built into the SlicerRT toolbox were compared to results from our in-house gamma algorithm implemented in Matlab (via MatlabBridge in 3D Slicer). The effects of noise, resolution and the exchange of reference and evaluated designations on the gamma comparison were also examined. Results: Perfect agreement was found between the gamma results obtained using the SlicerRT tool and our Matlab implementation for both the four-field box and gel datasets. The behaviour of the SlicerRT comparison with respect to changes in noise, resolution and the role of the reference and evaluated dose distributions was consistent with previous findings. Conclusion: Two independently developed gamma comparison tools have been cross-validated and found to be identical. As we transition our gel dosimetry analysis from Matlab to 3D Slicer, this validation serves as an important test towards ensuring the consistency of dose comparisons using the 3D Slicer

  6. Computational model of mesenchymal migration in 3D under chemotaxis

    PubMed Central

    Ribeiro, F. O.; Gómez-Benito, M. J.; Folgado, J.; Fernandes, P. R.; García-Aznar, J. M.

    2017-01-01

    Abstract Cell chemotaxis is an important characteristic of cellular migration, which takes part in crucial aspects of life and development. In this work, we propose a novel in silico model of mesenchymal 3D migration with competing protrusions under a chemotactic gradient. Based on recent experimental observations, we identify three main stages that can regulate mesenchymal chemotaxis: chemosensing, dendritic protrusion dynamics and cell–matrix interactions. Therefore, each of these features is considered as a different module of the main regulatory computational algorithm. The numerical model was particularized for the case of fibroblast chemotaxis under a PDGF-bb gradient. Fibroblasts migration was simulated embedded in two different 3D matrices – collagen and fibrin – and under several PDGF-bb concentrations. Validation of the model results was provided through qualitative and quantitative comparison with in vitro studies. Our numerical predictions of cell trajectories and speeds were within the measured in vitro ranges in both collagen and fibrin matrices. Although in fibrin, the migration speed of fibroblasts is very low, because fibrin is a stiffer and more entangling matrix. Testing PDGF-bb concentrations, we noticed that an increment of this factor produces a speed increment. At 1 ng mL−1 a speed peak is reached after which the migration speed diminishes again. Moreover, we observed that fibrin exerts a dampening behavior on migration, significantly affecting the migration efficiency. PMID:27336322

  7. Computational model of mesenchymal migration in 3D under chemotaxis.

    PubMed

    Ribeiro, F O; Gómez-Benito, M J; Folgado, J; Fernandes, P R; García-Aznar, J M

    2017-01-01

    Cell chemotaxis is an important characteristic of cellular migration, which takes part in crucial aspects of life and development. In this work, we propose a novel in silico model of mesenchymal 3D migration with competing protrusions under a chemotactic gradient. Based on recent experimental observations, we identify three main stages that can regulate mesenchymal chemotaxis: chemosensing, dendritic protrusion dynamics and cell-matrix interactions. Therefore, each of these features is considered as a different module of the main regulatory computational algorithm. The numerical model was particularized for the case of fibroblast chemotaxis under a PDGF-bb gradient. Fibroblasts migration was simulated embedded in two different 3D matrices - collagen and fibrin - and under several PDGF-bb concentrations. Validation of the model results was provided through qualitative and quantitative comparison with in vitro studies. Our numerical predictions of cell trajectories and speeds were within the measured in vitro ranges in both collagen and fibrin matrices. Although in fibrin, the migration speed of fibroblasts is very low, because fibrin is a stiffer and more entangling matrix. Testing PDGF-bb concentrations, we noticed that an increment of this factor produces a speed increment. At 1 ng mL(-1) a speed peak is reached after which the migration speed diminishes again. Moreover, we observed that fibrin exerts a dampening behavior on migration, significantly affecting the migration efficiency.

  8. [3D emulation of epicardium dynamic mapping].

    PubMed

    Lu, Jun; Yang, Cui-Wei; Fang, Zu-Xiang

    2005-03-01

    In order to realize epicardium dynamic mapping of the whole atria, 3-D graphics are drawn with OpenGL. Some source codes are introduced in the paper to explain how to produce, read, and manipulate 3-D model data.

  9. An interactive multiview 3D display system

    NASA Astrophysics Data System (ADS)

    Zhang, Zhaoxing; Geng, Zheng; Zhang, Mei; Dong, Hui

    2013-03-01

    The progresses in 3D display systems and user interaction technologies will help more effective 3D visualization of 3D information. They yield a realistic representation of 3D objects and simplifies our understanding to the complexity of 3D objects and spatial relationship among them. In this paper, we describe an autostereoscopic multiview 3D display system with capability of real-time user interaction. Design principle of this autostereoscopic multiview 3D display system is presented, together with the details of its hardware/software architecture. A prototype is built and tested based upon multi-projectors and horizontal optical anisotropic display structure. Experimental results illustrate the effectiveness of this novel 3D display and user interaction system.

  10. Laser Based 3D Volumetric Display System

    DTIC Science & Technology

    1993-03-01

    Literature, Costa Mesa, CA July 1983. 3. "A Real Time Autostereoscopic Multiplanar 3D Display System", Rodney Don Williams, Felix Garcia, Jr., Texas...8217 .- NUMBERS LASER BASED 3D VOLUMETRIC DISPLAY SYSTEM PR: CD13 0. AUTHOR(S) PE: N/AWIU: DN303151 P. Soltan, J. Trias, W. Robinson, W. Dahlke 7...laser generated 3D volumetric images on a rotating double helix, (where the 3D displays are computer controlled for group viewing with the naked eye

  11. True 3d Images and Their Applications

    NASA Astrophysics Data System (ADS)

    Wang, Z.; wang@hzgeospace., zheng.

    2012-07-01

    A true 3D image is a geo-referenced image. Besides having its radiometric information, it also has true 3Dground coordinates XYZ for every pixels of it. For a true 3D image, especially a true 3D oblique image, it has true 3D coordinates not only for building roofs and/or open grounds, but also for all other visible objects on the ground, such as visible building walls/windows and even trees. The true 3D image breaks the 2D barrier of the traditional orthophotos by introducing the third dimension (elevation) into the image. From a true 3D image, for example, people will not only be able to read a building's location (XY), but also its height (Z). true 3D images will fundamentally change, if not revolutionize, the way people display, look, extract, use, and represent the geospatial information from imagery. In many areas, true 3D images can make profound impacts on the ways of how geospatial information is represented, how true 3D ground modeling is performed, and how the real world scenes are presented. This paper first gives a definition and description of a true 3D image and followed by a brief review of what key advancements of geospatial technologies have made the creation of true 3D images possible. Next, the paper introduces what a true 3D image is made of. Then, the paper discusses some possible contributions and impacts the true 3D images can make to geospatial information fields. At the end, the paper presents a list of the benefits of having and using true 3D images and the applications of true 3D images in a couple of 3D city modeling projects.

  12. A 3D tension bioreactor platform to study the interplay between ECM stiffness and tumor phenotype.

    PubMed

    Cassereau, Luke; Miroshnikova, Yekaterina A; Ou, Guanqing; Lakins, Johnathon; Weaver, Valerie M

    2015-01-10

    Extracellular matrix (ECM) structure, composition, and stiffness have profound effects on tissue development and pathologies such as cardiovascular disease and cancer. Accordingly, a variety of synthetic hydrogel systems have been designed to study the impact of ECM composition, density, mechanics, and topography on cell and tissue phenotype. However, these synthetic systems fail to accurately recapitulate the biological properties and structure of the native tissue ECM. Natural three dimensional (3D) ECM hydrogels, such as collagen or hyaluronic acid, feature many of the chemical and physical properties of tissue, yet, these systems have limitations including the inability to independently control biophysical properties such as stiffness and pore size. Here, we present a 3D tension bioreactor system that permits precise mechanical tuning of collagen hydrogel stiffness, while maintaining consistent composition and pore size. We achieve this by mechanically loading collagen hydrogels covalently-conjugated to a polydimethylsiloxane (PDMS) membrane to induce hydrogel stiffening. We validated the biological application of this system with oncogenically transformed mammary epithelial cell organoids embedded in a 3D collagen I hydrogel, either uniformly stiffened or calibrated to create a gradient of ECM stiffening, to visually demonstrate the impact of ECM stiffening on transformation and tumor cell invasion. As such, this bioreactor presents the first tunable 3D natural hydrogel system that is capable of independently assessing the role of ECM stiffness on tissue phenotype.

  13. In vitro analysis of chemotactic leukocyte migration in 3D environments.

    PubMed

    Sixt, Michael; Lämmermann, Tim

    2011-01-01

    Cell migration on two-dimensional (2D) substrates follows entirely different rules than cell migration in three-dimensional (3D) environments. This is especially relevant for leukocytes that are able to migrate in the absence of adhesion receptors within the confined geometry of artificial 3D extracellular matrix scaffolds and within the interstitial space in vivo. Here, we describe in detail a simple and economical protocol to visualize dendritic cell migration in 3D collagen scaffolds along chemotactic gradients. This method can be adapted to other cell types and may serve as a physiologically relevant paradigm for the directed locomotion of most amoeboid cells.

  14. 3D Printing and Its Urologic Applications

    PubMed Central

    Soliman, Youssef; Feibus, Allison H; Baum, Neil

    2015-01-01

    3D printing is the development of 3D objects via an additive process in which successive layers of material are applied under computer control. This article discusses 3D printing, with an emphasis on its historical context and its potential use in the field of urology. PMID:26028997

  15. Teaching Geography with 3-D Visualization Technology

    ERIC Educational Resources Information Center

    Anthamatten, Peter; Ziegler, Susy S.

    2006-01-01

    Technology that helps students view images in three dimensions (3-D) can support a broad range of learning styles. "Geo-Wall systems" are visualization tools that allow scientists, teachers, and students to project stereographic images and view them in 3-D. We developed and presented 3-D visualization exercises in several undergraduate courses.…

  16. Expanding Geometry Understanding with 3D Printing

    ERIC Educational Resources Information Center

    Cochran, Jill A.; Cochran, Zane; Laney, Kendra; Dean, Mandi

    2016-01-01

    With the rise of personal desktop 3D printing, a wide spectrum of educational opportunities has become available for educators to leverage this technology in their classrooms. Until recently, the ability to create physical 3D models was well beyond the scope, skill, and budget of many schools. However, since desktop 3D printers have become readily…

  17. Beowulf 3D: a case study

    NASA Astrophysics Data System (ADS)

    Engle, Rob

    2008-02-01

    This paper discusses the creative and technical challenges encountered during the production of "Beowulf 3D," director Robert Zemeckis' adaptation of the Old English epic poem and the first film to be simultaneously released in IMAX 3D and digital 3D formats.

  18. 3D Flow Visualization Using Texture Advection

    NASA Technical Reports Server (NTRS)

    Kao, David; Zhang, Bing; Kim, Kwansik; Pang, Alex; Moran, Pat (Technical Monitor)

    2001-01-01

    Texture advection is an effective tool for animating and investigating 2D flows. In this paper, we discuss how this technique can be extended to 3D flows. In particular, we examine the use of 3D and 4D textures on 3D synthetic and computational fluid dynamics flow fields.

  19. Hydrogel-based reinforcement of 3D bioprinted constructs

    PubMed Central

    Levato, R; Peiffer, Q C; de Ruijter, M; Hennink, W E; Vermonden, T; Malda, J

    2016-01-01

    Progress within the field of biofabrication is hindered by a lack of suitable hydrogel formulations. Here, we present a novel approach based on a hybrid printing technique to create cellularized 3D printed constructs. The hybrid bioprinting strategy combines a reinforcing gel for mechanical support with a bioink to provide a cytocompatible environment. In comparison with thermoplastics such as є-polycaprolactone, the hydrogel-based reinforcing gel platform enables printing at cell-friendly temperatures, targets the bioprinting of softer tissues and allows for improved control over degradation kinetics. We prepared amphiphilic macromonomers based on poloxamer that form hydrolysable, covalently cross-linked polymer networks. Dissolved at a concentration of 28.6%w/w in water, it functions as reinforcing gel, while a 5%w/w gelatin-methacryloyl based gel is utilized as bioink. This strategy allows for the creation of complex structures, where the bioink provides a cytocompatible environment for encapsulated cells. Cell viability of equine chondrocytes encapsulated within printed constructs remained largely unaffected by the printing process. The versatility of the system is further demonstrated by the ability to tune the stiffness of printed constructs between 138 and 263 kPa, as well as to tailor the degradation kinetics of the reinforcing gel from several weeks up to more than a year. PMID:27431861

  20. 3D-printed microfluidic chips with patterned, cell-laden hydrogel constructs.

    PubMed

    Knowlton, Stephanie; Yu, Chu Hsiang; Ersoy, Fulya; Emadi, Sharareh; Khademhosseini, Ali; Tasoglu, Savas

    2016-06-20

    Three-dimensional (3D) printing offers potential to fabricate high-throughput and low-cost fabrication of microfluidic devices as a promising alternative to traditional techniques which enables efficient design iterations in the development stage. In this study, we demonstrate a single-step fabrication of a 3D transparent microfluidic chip using two alternative techniques: a stereolithography-based desktop 3D printer and a two-step fabrication using an industrial 3D printer based on polyjet technology. This method, compared to conventional fabrication using relatively expensive materials and labor-intensive processes, presents a low-cost, rapid prototyping technique to print functional 3D microfluidic chips. We enhance the capabilities of 3D-printed microfluidic devices by coupling 3D cell encapsulation and spatial patterning within photocrosslinkable gelatin methacryloyl (GelMA). The platform presented here serves as a 3D culture environment for long-term cell culture and growth. Furthermore, we have demonstrated the ability to print complex 3D microfluidic channels to create predictable and controllable fluid flow regimes. Here, we demonstrate the novel use of 3D-printed microfluidic chips as controllable 3D cell culture environments, advancing the applicability of 3D printing to engineering physiological systems for future applications in bioengineering.

  1. 3D Printing of Ultratough Polyion Complex Hydrogels.

    PubMed

    Zhu, Fengbo; Cheng, Libo; Yin, Jun; Wu, Zi Liang; Qian, Jin; Fu, Jianzhong; Zheng, Qiang

    2016-11-16

    Polyion complex (PIC) hydrogels have been proposed as promising engineered soft materials due to their high toughness and good processability. In this work, we reported manufacturing of complex structures with tough PIC hydrogels based on three-dimensional (3D) printing technology. The strategy relies on the distinct strength of ionic bonding in PIC hydrogels at different stages of printing. In concentrated saline solution, PIC forms viscous solution, which can be directly extruded out of a nozzle into water, where dialyzing out of salt and counterions results in sol-gel transition to form tough physical PIC gel with intricate structures. The printability of PIC solutions was systematically investigated by adjusting the PIC material formula and printing parameters in which proper viscosity and gelation rate were found to be key factors for successful 3D printing. Uniaxial tensile tests were performed to printed single fibers and multilayer grids, both exhibiting distinct yet controllable strength and toughness. More complex 3D structures with negative Poisson's ratio, gradient grid, and material anisotropy were constructed as well, demonstrating the flexible printability of PIC hydrogels. The methodology and capability here provide a versatile platform to fabricate complex structures with tough PIC hydrogels, which should broaden the use of such materials in applications such as biomedical devices and artificial tissues.

  2. 3-D Perspective Pasadena, California

    NASA Technical Reports Server (NTRS)

    2000-01-01

    This perspective view shows the western part of the city of Pasadena, California, looking north towards the San Gabriel Mountains. Portions of the cities of Altadena and La Canada, Flintridge are also shown. The image was created from three datasets: the Shuttle Radar Topography Mission (SRTM) supplied the elevation data; Landsat data from November 11, 1986 provided the land surface color (not the sky) and U.S. Geological Survey digital aerial photography provides the image detail. The Rose Bowl, surrounded by a golf course, is the circular feature at the bottom center of the image. The Jet Propulsion Laboratory is the cluster of large buildings north of the Rose Bowl at the base of the mountains. A large landfill, Scholl Canyon, is the smooth area in the lower left corner of the scene. This image shows the power of combining data from different sources to create planning tools to study problems that affect large urban areas. In addition to the well-known earthquake hazards, Southern California is affected by a natural cycle of fire and mudflows. Wildfires strip the mountains of vegetation, increasing the hazards from flooding and mudflows for several years afterwards. Data such as shown on this image can be used to predict both how wildfires will spread over the terrain and also how mudflows will be channeled down the canyons. The Shuttle Radar Topography Mission (SRTM), launched on February 11, 2000, uses the same radar instrument that comprised the Spaceborne Imaging Radar-C/X-Band Synthetic Aperture Radar (SIR-C/X-SAR) that flew twice on the Space Shuttle Endeavour in 1994. The mission was designed to collect three dimensional measurements of the Earth's surface. To collect the 3-D data, engineers added a 60-meter-long (200-foot) mast, an additional C-band imaging antenna and improved tracking and navigation devices. The mission is a cooperative project between the National Aeronautics and Space Administration (NASA), the National Imagery and Mapping Agency

  3. 3D CARS image reconstruction and pattern recognition on SHG images

    NASA Astrophysics Data System (ADS)

    Medyukhina, Anna; Vogler, Nadine; Latka, Ines; Dietzek, Benjamin; Cicchi, Riccardo; Pavone, Francesco S.; Popp, Jürgen

    2012-06-01

    Nonlinear optical imaging techniques based e.g. on coherent anti-Stokes Raman scattering (CARS) or second-harmonic generation (SHG) show great potential for in-vivo investigations of tissue. While the microspectroscopic imaging tools are established, automized data evaluation, i.e. image pattern recognition and automized image classification, of nonlinear optical images still bares great possibilities for future developments towards an objective clinical diagnosis. This contribution details the capability of nonlinear microscopy for both 3D visualization of human tissues and automated discrimination between healthy and diseased patterns using ex-vivo human skin samples. By means of CARS image alignment we show how to obtain a quasi-3D model of a skin biopsy, which allows us to trace the tissue structure in different projections. Furthermore, the potential of automated pattern and organization recognition to distinguish between healthy and keloidal skin tissue is discussed. A first classification algorithm employs the intrinsic geometrical features of collagen, which can be efficiently visualized by SHG microscopy. The shape of the collagen pattern allows conclusions about the physiological state of the skin, as the typical wavy collagen structure of healthy skin is disturbed e.g. in keloid formation. Based on the different collagen patterns a quantitative score characterizing the collagen waviness - and hence reflecting the physiological state of the tissue - is obtained. Further, two additional scoring methods for collagen organization, respectively based on a statistical analysis of the mutual organization of fibers and on FFT, are presented.

  4. Modeling Extracellular Matrix Reorganization in 3D Environments

    PubMed Central

    Harjanto, Dewi; Zaman, Muhammad H.

    2013-01-01

    Extracellular matrix (ECM) remodeling is a key physiological process that occurs in a number of contexts, including cell migration, and is especially important for cellular form and function in three-dimensional (3D) matrices. However, there have been few attempts to computationally model how cells modify their environment in a manner that accounts for both cellular properties and the architecture of the surrounding ECM. To this end, we have developed and validated a novel model to simulate matrix remodeling that explicitly defines cells in a 3D collagenous matrix. In our simulation, cells can degrade, deposit, or pull on local fibers, depending on the fiber density around each cell. The cells can also move within the 3D matrix. Different cell phenotypes can be modeled by varying key cellular parameters. Using the model we have studied how two model cancer cell lines, of differing invasiveness, modify matrices with varying fiber density in their vicinity by tracking the metric of fraction of matrix occupied by fibers. Our results quantitatively demonstrate that in low density environments, cells deposit more collagen to uniformly increase fibril fraction. On the other hand, in higher density environments, the less invasive model cell line reduced the fibril fraction as compared to the highly invasive phenotype. These results show good qualitative and quantitative agreement with existing experimental literature. Our simulation is therefore able to function as a novel platform to provide new insights into the clinically relevant and physiologically critical process of matrix remodeling by helping identify critical parameters that dictate cellular behavior in complex native-like environments. PMID:23341900

  5. Case study: Beauty and the Beast 3D: benefits of 3D viewing for 2D to 3D conversion

    NASA Astrophysics Data System (ADS)

    Handy Turner, Tara

    2010-02-01

    From the earliest stages of the Beauty and the Beast 3D conversion project, the advantages of accurate desk-side 3D viewing was evident. While designing and testing the 2D to 3D conversion process, the engineering team at Walt Disney Animation Studios proposed a 3D viewing configuration that not only allowed artists to "compose" stereoscopic 3D but also improved efficiency by allowing artists to instantly detect which image features were essential to the stereoscopic appeal of a shot and which features had minimal or even negative impact. At a time when few commercial 3D monitors were available and few software packages provided 3D desk-side output, the team designed their own prototype devices and collaborated with vendors to create a "3D composing" workstation. This paper outlines the display technologies explored, final choices made for Beauty and the Beast 3D, wish-lists for future development and a few rules of thumb for composing compelling 2D to 3D conversions.

  6. Mini 3D for shallow gas reconnaissance

    SciTech Connect

    Vallieres, T. des; Enns, D.; Kuehn, H.; Parron, D.; Lafet, Y.; Van Hulle, D.

    1996-12-31

    The Mini 3D project was undertaken by TOTAL and ELF with the support of CEPM (Comite d`Etudes Petrolieres et Marines) to define an economical method of obtaining 3D seismic HR data for shallow gas assessment. An experimental 3D survey was carried out with classical site survey techniques in the North Sea. From these data 19 simulations, were produced to compare different acquisition geometries ranging from dual, 600 m long cables to a single receiver. Results show that short offset, low fold and very simple streamer positioning are sufficient to give a reliable 3D image of gas charged bodies. The 3D data allow a much more accurate risk delineation than 2D HR data. Moreover on financial grounds Mini-3D is comparable in cost to a classical HR 2D survey. In view of these results, such HR 3D should now be the standard for shallow gas surveying.

  7. In situ gelation properties of a collagen-genipin sol with a potential for the treatment of gastrointestinal ulcers.

    PubMed

    Narita, Takefumi; Yunoki, Shunji; Ohyabu, Yoshimi; Yahagi, Naohisa; Uraoka, Toshio

    2016-01-01

    We investigated the potential of collagen-genipin sols as biomaterials for treating artificial ulcers following endoscopic submucosal dissection. Collagen sol viscosity increased with condensation, allowing retention on tilted ulcers before gelation and resulting in collagen gel deposition on whole ulcers. The 1.44% collagen sols containing genipin as a crosslinker retained sol fluidity at 23°C for >20 min, facilitating endoscopic use. Collagen sols formed gel depositions on artificial ulcers in response to body temperature, and high temperature responsiveness of gelation because of increased neutral phosphate buffer concentration allowed for thick gel deposition on tilted ulcers. Finally, histological observations showed infiltration of gels into submucosal layers. Taken together, the present data show that genipin-induced crosslinking significantly improves the mechanical properties of collagen gels even at low genipin concentrations of 0.2-1 mM, warranting the use of in situ gelling collagen-genipin sols for endoscopic treatments of gastrointestinal ulcers.

  8. X-ray phase nanotomography resolves the 3D human bone ultrastructure.

    PubMed

    Langer, Max; Pacureanu, Alexandra; Suhonen, Heikki; Grimal, Quentin; Cloetens, Peter; Peyrin, Françoise

    2012-01-01

    Bone strength and failure are increasingly thought to be due to ultrastructural properties, such as the morphology of the lacuno-canalicular network, the collagen fiber orientation and the mineralization on the nanoscale. However, these properties have not been studied in 3D so far. Here we report the investigation of the human bone ultrastructure with X-ray phase nanotomography, which now provides the required sensitivity, spatial resolution and field of view. The 3D organization of the lacuno-canalicular network is studied in detail over several cells in osteonal and interstitial tissue. Nanoscale density variations are revealed and show that the cement line separating these tissues is hypermineralized. Finally, we show that the collagen fibers are organized as a twisted plywood structure in 3D.

  9. 3D Cultivation Techniques for Primary Human Hepatocytes

    PubMed Central

    Bachmann, Anastasia; Moll, Matthias; Gottwald, Eric; Nies, Cordula; Zantl, Roman; Wagner, Helga; Burkhardt, Britta; Sánchez, Juan J. Martínez; Ladurner, Ruth; Thasler, Wolfgang; Damm, Georg; Nussler, Andreas K.

    2015-01-01

    One of the main challenges in drug development is the prediction of in vivo toxicity based on in vitro data. The standard cultivation system for primary human hepatocytes is based on monolayer cultures, even if it is known that these conditions result in a loss of hepatocyte morphology and of liver-specific functions, such as drug-metabolizing enzymes and transporters. As it has been demonstrated that hepatocytes embedded between two sheets of collagen maintain their function, various hydrogels and scaffolds for the 3D cultivation of hepatocytes have been developed. To further improve or maintain hepatic functions, 3D cultivation has been combined with perfusion. In this manuscript, we discuss the benefits and drawbacks of different 3D microfluidic devices. For most systems that are currently available, the main issues are the requirement of large cell numbers, the low throughput, and expensive equipment, which render these devices unattractive for research and the drug-developing industry. A higher acceptance of these devices could be achieved by their simplification and their compatibility with high-throughput, as both aspects are of major importance for a user-friendly device. PMID:27600213

  10. Microfluidic titer plate for stratified 3D cell culture.

    PubMed

    Trietsch, Sebastiaan J; Israëls, Guido D; Joore, Jos; Hankemeier, Thomas; Vulto, Paul

    2013-09-21

    Human tissues and organs are inherently heterogeneous. Their functionality is determined by the interplay between different cell types, their secondary architecture, vascular system and gradients of signaling molecules and metabolites. Here we propose a stratified 3D cell culture platform, in which adjacent lanes of gels and liquids are patterned by phaseguides to capture this tissue heterogeneity. We demonstrate 3D cell culture of HepG2 hepatocytes under continuous perfusion, a rifampicin toxicity assay and co-culture with fibroblasts. 4T1 breast cancer cells are used to demonstrate invasion and aggregation models. The platform is incorporated in a microtiter plate format that renders it fully compatible with automation and high-content screening equipment. The extended functionality, ease of handling and full compatibility to standard equipment is an important step towards adoption of Organ-on-a-Chip technology for screening in an industrial setting.

  11. Phenotypic changes in proliferation, differentiation, and migration of chondrocytes: 3D in vitro models for joint wound healing.

    PubMed

    Tsai, Yu-Hui; Chen, Chun-Wei; Lai, Wen-Fu T; Tang, Ja-Reng; Deng, Win-Ping; Yeh, Shauh-Der; Chung, Andrew; Zuo, Chun S; Bowley, John F

    2010-03-01

    We aim to establish a 3D model of cartilage wound healing, and explore the involvement of chondrocytes in its repair. To characterize chondrocyte involvement in wound healing, an in vitro 3D model composed of chondrocyte mixing with either type II/I collagen or type I collagen matrix was established. The "defects" measuring 5 mm in diameter were made on each collagen matrix-chondrocyte construct to mimic in vivo cartilage defects. The effects of basic fibroblast growth factor (bFGF) on chondrocytes migration and differentiation were studied. The migration and Glucosaminoglycan (GAG) synthesis of chondrocytes in the defect areas were observed by microscopy after Alcian-blue staining. In the presence of bFGF, GAG expression increased significantly when chondrocytes were cultured in type II/I collagen matrix compared to type I collagen matrix. However, mild GAG accumulation was also found when cells were cultured in either type I or type II/I collagens without bFGF. In a 3D model of cartilage wound healing, bFGF promote chondrocyte proliferation, migration and differentiation in the presence of type II/I collagen matrix, and showed potential to regulate wound healing. These wound healing models may provide feasible methods to explore various drugs prior to human trials.

  12. 3-D Technology Approaches for Biological Ecologies

    NASA Astrophysics Data System (ADS)

    Liu, Liyu; Austin, Robert; U. S-China Physical-Oncology Sciences Alliance (PS-OA) Team

    Constructing three dimensional (3-D) landscapes is an inevitable issue in deep study of biological ecologies, because in whatever scales in nature, all of the ecosystems are composed by complex 3-D environments and biological behaviors. Just imagine if a 3-D technology could help complex ecosystems be built easily and mimic in vivo microenvironment realistically with flexible environmental controls, it will be a fantastic and powerful thrust to assist researchers for explorations. For years, we have been utilizing and developing different technologies for constructing 3-D micro landscapes for biophysics studies in in vitro. Here, I will review our past efforts, including probing cancer cell invasiveness with 3-D silicon based Tepuis, constructing 3-D microenvironment for cell invasion and metastasis through polydimethylsiloxane (PDMS) soft lithography, as well as explorations of optimized stenting positions for coronary bifurcation disease with 3-D wax printing and the latest home designed 3-D bio-printer. Although 3-D technologies is currently considered not mature enough for arbitrary 3-D micro-ecological models with easy design and fabrication, I hope through my talk, the audiences will be able to sense its significance and predictable breakthroughs in the near future. This work was supported by the State Key Development Program for Basic Research of China (Grant No. 2013CB837200), the National Natural Science Foundation of China (Grant No. 11474345) and the Beijing Natural Science Foundation (Grant No. 7154221).

  13. Second harmonic generation quantitative measurements on collagen fibrils through correlation to electron microscopy

    NASA Astrophysics Data System (ADS)

    Bancelin, S.; Aimé, C.; Gusachenko, I.; Kowalczuk, L.; Latour, G.; Coradin, T.; Schanne-Klein, M.-C.

    2015-03-01

    Type I collagen is a major structural protein in mammals that shows highly structured macromolecular organizations specific to each tissue. This biopolymer is synthesized as triple helices, which self-assemble into fibrils (Ø =10-300 nm) and further form various 3D organization. In recent years, Second Harmonic Generation (SHG) microscopy has emerged as a powerful technique to probe in situ the fibrillar collagenous network within tissues. However, this optical technique cannot resolve most of the fibrils and is a coherent process, which has impeded quantitative measurements of the fibril diameter so far. In this study, we correlated SHG microscopy with Transmission Electron Microscopy to determine the sensitivity of SHG microscopy and to calibrate SHG signals as a function of the fibril diameter in reconstructed collagen gels. To that end, we synthetized isolated fibrils with various diameters and successfully imaged the very same fibrils with both techniques, down to 30 nm diameter. We observed that SHG signals scaled as the fourth power of the fibril diameter, as expected from analytical and numerical calculations. This calibration was then applied to diabetic rat cornea in which we successfully recovered the diameter of hyperglycemia-induced fibrils in the Descemet's membrane without having to resolve them. Finally we derived the first hyperpolarizability from a single collagen triple helix which validates the bottom-up approach used to calculate the non-linear response at the fibrillar scale and denotes a parallel alignment of triple helices within the fibrils. These results represent a major step towards quantitative SHG imaging of nm-sized collagen fibrils.

  14. 3D change detection - Approaches and applications

    NASA Astrophysics Data System (ADS)

    Qin, Rongjun; Tian, Jiaojiao; Reinartz, Peter

    2016-12-01

    Due to the unprecedented technology development of sensors, platforms and algorithms for 3D data acquisition and generation, 3D spaceborne, airborne and close-range data, in the form of image based, Light Detection and Ranging (LiDAR) based point clouds, Digital Elevation Models (DEM) and 3D city models, become more accessible than ever before. Change detection (CD) or time-series data analysis in 3D has gained great attention due to its capability of providing volumetric dynamics to facilitate more applications and provide more accurate results. The state-of-the-art CD revi