Publications by authors named "Aurore Van de Walle"

19 Publications

  • Page 1 of 1

Massive Intracellular Remodeling of CuS Nanomaterials Produces Nontoxic Bioengineered Structures with Preserved Photothermal Potential.

ACS Nano 2021 06 25;15(6):9782-9795. Epub 2021 May 25.

Laboratoire Matière et Systèmes Complexes MSC, UMR 7057, CNRS and University of Paris, 75205, Paris Cedex 13, France.

Despite efforts in producing nanoparticles with tightly controlled designs and specific physicochemical properties, these can undergo massive nano-bio interactions and bioprocessing upon internalization into cells. These transformations can generate adverse biological outcomes and premature loss of functional efficacy. Hence, understanding the intracellular fate of nanoparticles is a necessary prerequisite for their introduction in medicine. Among nanomaterials devoted to theranostics is copper sulfide (CuS), which provides outstanding optical properties along with easy synthesis and low cost. Herein, we performed a long-term multiscale study on the bioprocessing of hollow CuS nanoparticles (CuS NPs) and rattle-like iron oxide [email protected] core-shell hybrids ([email protected] NPs) when inside stem cells and cancer cells, cultured as spheroids. In the spheroids, both CuS NPs and [email protected] NPs are rapidly dismantled into smaller units (day 0 to 3), and hair-like nanostructures are generated (day 9 to 21). This bioprocessing triggers an adaptation of the cellular metabolism to the internalized metals without impacting cell viability, differentiation, or oxidative stress response. Throughout the remodeling, a loss of IONF-derived magnetism is observed, but, surprisingly, the CuS photothermal potential is preserved, as demonstrated by a full characterization of the photothermal conversion across the bioprocessing process. The maintained photothermal efficiency correlated well with synchrotron X-ray absorption spectroscopy measurements, evidencing a similar chemical phase for Cu but not for Fe over time. These findings evidence that the intracellular bioprocessing of CuS nanoparticles can reshape them into bioengineered nanostructures without reducing the photothermal function and therapeutic potential.
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http://dx.doi.org/10.1021/acsnano.1c00567DOI Listing
June 2021

Using Magnetometry to Monitor Cellular Incorporation and Subsequent Biodegradation of Chemically Synthetized Iron Oxide Nanoparticles.

J Vis Exp 2021 02 27(168). Epub 2021 Feb 27.

Université Sorbonne Paris Nord, Laboratory for Vascular Translational Science, LVTS, INSERM, UMR 1148; Services de Biochimie et Médecine Nucléaire, Hôpital Avicenne Assistance Publique-Hôpitaux de Paris;

Magnetic nanoparticles, made of iron oxide, present a peculiar interest for a wide range of biomedical applications for which they are often internalized in cells and then left within. One challenge is to assess their fate in the intracellular environment with reliable and precise methodologies. Herein, we introduce the use of the vibrating sample magnetometer (VSM) to precisely quantify the integrity of magnetic nanoparticles within cells by measuring their magnetic moment. Stem cells are first labeled with two types of magnetic nanoparticles; the nanoparticles have the same core produced via a fast and efficient microwave-based nonaqueous sol gel synthesis and differ in their coating: the commonly used citric acid molecule is compared to polyacrylic acid. The formation of 3D cell-spheroids is then achieved via centrifugation and the magnetic moment of these spheroids is measured at different times with the VSM. The obtained moment is a direct fingerprint of the nanoparticles' integrity, with decreasing values indicative of a nanoparticle degradation. For both nanoparticles, the magnetic moment decreases over culture time revealing their biodegradation. A protective effect of the polyacrylic acid coating is also shown, when compared to citric acid.
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http://dx.doi.org/10.3791/61106DOI Listing
February 2021

Ever-Evolving Identity of Magnetic Nanoparticles within Human Cells: The Interplay of Endosomal Confinement, Degradation, Storage, and Neocrystallization.

Acc Chem Res 2020 10 16;53(10):2212-2224. Epub 2020 Sep 16.

Laboratoire Matière et Systèmes Complexes, MSC, UMR 7057, CNRS & University of Paris, 75205, Paris, Cedex 13, France.

Considerable knowledge has been acquired in inorganic nanoparticles' synthesis and nanoparticles' potential use in biomedical applications. Among different materials, iron oxide nanoparticles remain unrivaled for several reasons. Not only do they respond to multiple physical stimuli (e.g., magnetism, light) and exert multifunctional therapeutic and diagnostic actions but also they are biocompatible and integrate endogenous iron-related metabolic pathways. With the aim to optimize the use of (magnetic) iron oxide nanoparticles in biomedicine, different biophysical phenomena have been recently identified and studied. Among them, the concept of a "nanoparticle's identity" is of particular importance. Nanoparticles' identities evolve in distinct biological environments and over different periods of time. In this Account, we focus on the remodeling of magnetic nanoparticles' identities following their journey inside cells. For instance, nanoparticles' functions, such as heat generation or magnetic resonance imaging, can be highly impacted by endosomal confinement. Structural degradation of nanoparticles was also evidenced and quantified and correlates with the loss of magnetic nanoparticle properties. Remarkably, in human stem cells, the nonmagnetic products of nanoparticles' degradation could be subsequently reassembled into neosynthesized, endogenous magnetic nanoparticles. This stunning occurrence might account for the natural presence of magnetic particles in human organs, especially the brain. However, mechanistic details and the implication of such phenomena in homeostasis and disease have yet to be completely unraveled.This Account aims to assess the short- and long-term transformations of magnetic iron oxide nanoparticles in living cells, particularly focusing on human stem cells. Precisely, we herein overview the multiple and ever-evolving chemical, physical, and biological magnetic nanoparticles' identities and emphasize the remarkable intracellular fate of these nanoparticles.
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http://dx.doi.org/10.1021/acs.accounts.0c00355DOI Listing
October 2020

Flow with variable pulse frequencies accelerates vascular recellularization and remodeling of a human bioscaffold.

J Biomed Mater Res A 2021 01 16;109(1):92-103. Epub 2020 Jun 16.

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, Gainesville, Florida, USA.

Despite significant advances in vascular tissue engineering, the ideal graft has not yet been developed and autologous vessels remain the gold standard substitutes for small diameter bypass procedures. Here, we explore the use of a flow field with variable pulse frequencies over the regeneration of an ex vivo-derived human scaffold as vascular graft. Briefly, human umbilical veins were decellularized and used as scaffold for cellular repopulation with human smooth muscle cells (SMC) and endothelial cells (EC). Over graft development, the variable flow, which mimics the real-time cardiac output of an individual performing daily activities (e.g., resting vs. exercising), was implemented and compared to the commonly used constant pulse frequency. Results show marked differences on SMC and EC function, with changes at the molecular level reflecting on tissue scales. First, variable frequencies significantly increased SMC proliferation rate and glycosaminoglycan production. These results can be tied with the SMC gene expression that indicates a synthetic phenotype, with a significant downregulation of myosin heavy chain. Additionally and quite remarkably, the variable flow frequencies motivated the re-endothelialization of the grafts, with a quiescent-like structure observed after 10 days of conditioning, contrasting with the low surface coverage and unaligned EC observed under constant frequency (CF). Besides, the overall biomechanics of the generated grafts (conditioned with both pulsed and CFs) evidence a significant remodeling after 55 days of culture, depicted by high burst pressure and Young's modulus. These last results demonstrate the positive recellularization and remodeling of a human-derived scaffold toward an arterial vessel.
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http://dx.doi.org/10.1002/jbm.a.37009DOI Listing
January 2021

Endosomal Confinement of Gold Nanospheres, Nanorods, and Nanoraspberries Governs Their Photothermal Identity and Is Beneficial for Cancer Cell Therapy.

Adv Biosyst 2020 04 9;4(4):e1900284. Epub 2020 Feb 9.

Laboratoire Matière et Systèmes, Complexes MSC, UMR 7057, CNRS & University Paris Diderot, 75205, Paris, Cedex 13, France.

Gold nanoparticles can act as photothermal agents to generate local tumor heating and subsequent depletion upon laser exposure. Herein, photothermal heating of four gold nanoparticles and the resulting induced cancer cell death are systematically assessed, within extra- or intracellular localizations. Two state-of-the-art gold nanorods are compared with small nanospheres (single-core) and nanoraspberries (multicore). Heat generation is measured in water dispersion and in cancer cells, using lasers at wavelengths of 680, 808, and 1064 nm, covering the entire range used in photothermal therapy, defined as near infrared first (NIR-I) and second (NIR-II) windows, with NIR-II offering more tissue penetration. When dispersed in water, gold nanospheres provide no significant heating, gold nanorods are efficient in NIR-I, and only gold nanoraspberries are still heating in NIR-II. However, in cells, due to endosomal confinement, all nanoparticles present an absorption red-shift translating visible and NIR-I absorbing nanoparticles into effective NIR-I and NIR-II nanoheaters, respectively. The gold nanorods then become competitive with the multicore nanoparticles (nanoraspberries) in NIR-II. Similarly, once in cells, gold nanospheres can be envisaged for NIR-I heating. Remarkably, nanoraspberries are efficient nanoheaters, whatever the laser applied, and the extra- versus intra-cellular localization demonstrates treatment versatility.
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http://dx.doi.org/10.1002/adbi.201900284DOI Listing
April 2020

Versatile iron cobalt nanoparticles for theranostics.

Nat Biomed Eng 2020 03;4(3):252-253

Laboratoire Matière et Systemes Complexes, UMR 7057, CNRS and University Paris Diderot, Paris, France.

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http://dx.doi.org/10.1038/s41551-020-0532-yDOI Listing
March 2020

Janus Magnetic-Plasmonic Nanoparticles for Magnetically Guided and Thermally Activated Cancer Therapy.

Small 2020 03 20;16(11):e1904960. Epub 2020 Feb 20.

Laboratoire Matière et Systèmes Complexes (MSC), UMR 7057, CNRS and Université Paris Diderot, 75205, Paris cedex 13, France.

Progress of thermal tumor therapies and their translation into clinical practice are limited by insufficient nanoparticle concentration to release therapeutic heating at the tumor site after systemic administration. Herein, the use of Janus magneto-plasmonic nanoparticles, made of gold nanostars and iron oxide nanospheres, as efficient therapeutic nanoheaters whose on-site delivery can be improved by magnetic targeting, is proposed. Single and combined magneto- and photo-thermal heating properties of Janus nanoparticles render them as compelling heating elements, depending on the nanoparticle dose, magnetic lobe size, and milieu conditions. In cancer cells, a much more effective effect is observed for photothermia compared to magnetic hyperthermia, while combination of the two modalities into a magneto-photothermal treatment results in a synergistic cytotoxic effect in vitro. The high potential of the Janus nanoparticles for magnetic guiding confirms them to be excellent nanostructures for in vivo magnetically enhanced photothermal therapy, leading to efficient tumor growth inhibition.
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http://dx.doi.org/10.1002/smll.201904960DOI Listing
March 2020

Sequential adaptation of perfusion and transport conditions significantly improves vascular construct recellularization and biomechanics.

J Tissue Eng Regen Med 2020 03 6;14(3):510-520. Epub 2020 Feb 6.

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, Gainesville, FL.

Recellularization of ex vivo-derived scaffolds remains a significant hurdle primarily due to the scaffolds subcellular pore size that restricts initial cell seeding to the scaffolds periphery and inhibits migration over time. With the aim to improve cell migration, repopulation, and graft mechanics, the effects of a four-step culture approach were assessed. Using an ex vivo-derived vein as a model scaffold, human smooth muscle cells were first seeded onto its ablumen (Step 1: 3 hr) and an aggressive 0-100% nutrient gradient (lumenal flow under hypotensive pressure) was created to initiate cell migration across the scaffold (Step 2: Day 0 to 19). The effects of a prolonged aggressive nutrient gradient created by this single lumenal flow was then compared with a dual flow (lumenal and ablumenal) in Step 3 (Day 20 to 30). Analyses showed that a single lumenal flow maintained for 30 days resulted in a higher proportion of cells migrating across the scaffold toward the vessel lumen (nutrient source), with improved distribution. In Step 4 (Day 31 to 45), the transition from hypotensive pressure (12/8 mmHg) to normotensive (arterial-like) pressure (120/80 mmHg) was assessed. It demonstrated that recellularized scaffolds exposed to arterial pressures have increased glycosaminoglycan deposition, physiological modulus, and Young's modulus. By using this stepwise conditioning, the challenging recellularization of a vein-based scaffold and its positive remodeling toward arterial biomechanics were obtained.
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http://dx.doi.org/10.1002/term.3015DOI Listing
March 2020

Transformation Cycle of Magnetosomes in Human Stem Cells: From Degradation to Biosynthesis of Magnetic Nanoparticles Anew.

ACS Nano 2020 02 16;14(2):1406-1417. Epub 2020 Jan 16.

Laboratoire Matière et Systèmes, Complexes MSC, UMR 7057, CNRS and University of Paris , 75205 , Paris Cedex 13 , France.

The nanoparticles produced by magnetotactic bacteria, called magnetosomes, are made of a magnetite core with high levels of crystallinity surrounded by a lipid bilayer. This organized structure has been developed during the course of evolution of these organisms to adapt to their specific habitat and is assumed to resist degradation and to be able to withstand the demanding biological environment. Herein, we investigated magnetosomes' structural fate upon internalization in human stem cells using magnetic and photothermal measurements, electron microscopy, and X-ray absorption spectroscopy. All measurements first converge to the demonstration that intracellular magnetosomes can experience an important biodegradation, with up to 70% of their initial content degraded, which is associated with the progressive storage of the released iron in the ferritin protein. It correlates with an extensive magnetite to ferrihydrite phase transition. The ionic species delivered by this degradation could then be used by the cells to biosynthesize magnetic nanoparticles anew. In this case, cell magnetism first decreased with magnetosomes being dissolved, but then cells remagnetized entirely, evidencing the neo-synthesis of biogenic magnetic nanoparticles. Bacteria-made biogenic magnetosomes can thus be totally remodeled by human stem cells, into human cells-made magnetic nanoparticles.
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http://dx.doi.org/10.1021/acsnano.9b08061DOI Listing
February 2020

[Human stem cells can neo-biosynthesize magnetic nanoparticles after degrading man-made nanoparticles].

Med Sci (Paris) 2019 Oct 18;35(10):725-727. Epub 2019 Oct 18.

Laboratoire matière et systèmes complexes, CNRS UMR 7057, Université Paris Diderot, 10 rue Alice Domon et Léonie Duquet, 75205 Paris Cedex 13, France.

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http://dx.doi.org/10.1051/medsci/2019157DOI Listing
October 2019

Impact of magnetic nanoparticle surface coating on their long-term intracellular biodegradation in stem cells.

Nanoscale 2019 Sep 27;11(35):16488-16498. Epub 2019 Aug 27.

Laboratoire Matière et Systèmes, Complexes MSC, UMR 7057, CNRS & University Paris Diderot, 75205, Paris Cedex 13, France.

Magnetic nanoparticles (MNPs) internalized within stem cells have paved the way for remote magnetic cell manipulation and imaging in regenerative medicine. A full understanding of their interactions with stem cells and of their fate in the intracellular environment is then required, in particular with respect to their surface coatings. Here, we investigated the biological interactions of MNPs composed of an identical magnetic core but coated with different molecules: phosphonoacetic acid, polyethylene glycol phosphonic carboxylic acid, caffeic acid, citric acid, and polyacrylic acid. These coatings vary in the nature of the chelating function, the number of binding sites, and the presence or absence of a polymer. The nanoparticle magnetism was systematically used as an indicator of their internalization within human stem cells and of their structural long-term biodegradation in a 3D stem cell spheroid model. Overall, we evidence that the coating impacts the aggregation status of the nanoparticles and subsequently their uptake within stem cells, but it has little effect on their intracellular degradation. Only a high number of chelating functions (polyacrylic acid) had a significant protective effect. Interestingly, when the nanoparticles aggregated prior to cellular internalization, less degradation was also observed. Finally, for all coatings, a robust dose-dependent intracellular degradation rate was demonstrated, with higher doses of internalized nanoparticles leading to a lower degradation extent.
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http://dx.doi.org/10.1039/c9nr05624fDOI Listing
September 2019

Biosynthesis of magnetic nanoparticles from nano-degradation products revealed in human stem cells.

Proc Natl Acad Sci U S A 2019 03 13;116(10):4044-4053. Epub 2019 Feb 13.

Laboratoire Matière et Systèmes Complexes, UMR 7057, CNRS and University Paris Diderot, 75205 Paris Cedex 13, France;

While magnetic nanoparticles offer exciting possibilities for stem cell imaging or tissue bioengineering, their long-term intracellular fate remains to be fully documented. Besides, it appears that magnetic nanoparticles can occur naturally in human cells, but their origin and potentially endogenous synthesis still need further understanding. In an effort to explore the life cycle of magnetic nanoparticles, we investigated their transformations upon internalization in mesenchymal stem cells and as a function of the cells' differentiation status (undifferentiated, or undergoing adipogenesis, osteogenesis, and chondrogenesis). Using magnetism as a fingerprint of the transformation process, we evidenced an important degradation of the nanoparticles during chondrogenesis. For the other pathways, stem cells were remarkably "remagnetized" after degradation of nanoparticles. This remagnetization phenomenon is the direct demonstration of a possible neosynthesis of magnetic nanoparticles and could lay some foundation to understand the presence of magnetic crystals in human cells. The neosynthesis was shown to take place within the endosomes and to involve the H-subunit of ferritin. Moreover, it appeared to be the key process to avoid long-term cytotoxicity (impact on differentiation) related to high doses of magnetic nanoparticles within stem cells.
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http://dx.doi.org/10.1073/pnas.1816792116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6410821PMC
March 2019

Role of growth factors and oxygen to limit hypertrophy and impact of high magnetic nanoparticles dose during stem cell chondrogenesis.

Comput Struct Biotechnol J 2018 30;16:532-542. Epub 2018 Oct 30.

Laboratoire Matière et Systèmes Complexes (MSC), UMR 7057 CNRS, University Paris Diderot, 75205 Paris cedex 13, France.

Due to an unmet clinical need of curative treatments for osteoarthritic patients, tissue engineering strategies that propose the development of cartilage tissue replacements from stem cells have emerged. Some of these strategies are based on the internalization of magnetic nanoparticles into stem cells to then initiate the chondrogenesis via magnetic compaction. A major difficulty is to drive the chondrogenic differentiation of the cells such as they produce an extracellular matrix free of hypertrophic collagen. An additional difficulty has to be overcome when nanoparticles are used, knowing that a high dose of nanoparticles can limit the chondrogenesis. We here propose a gene-based analysis of the effects of chemical factors (growth factors, hypoxia) on the chondrogenic differentiation of human mesenchymal stem cells both with and without nanoparticles. We focus on the synthesis of two of the most important constituents present in the cartilaginous extracellular matrix (Collagen II and Aggrecan) and on the expression of collagen X, the signature of hypertrophic cartilage, in order to provide a quantitative index of the type of cartilage produced (i.e. hyaline, hypertrophic). We demonstrate that by applying specific environmental conditions, gene expression can be directed toward the production of hyaline cartilage, with limited hypertrophy. Besides, a combination of the growth factors IGF-1, TGF-β3, with a hypoxic conditioning remarkably reduced the impact of high nanoparticles concentration.
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http://dx.doi.org/10.1016/j.csbj.2018.10.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6260287PMC
October 2018

Human Perinatal-Derived Biomaterials.

Adv Healthc Mater 2017 Sep 7;6(18). Epub 2017 Aug 7.

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, JG-56 Biomedical Sciences Building, P.O. Box 116131, Gainesville, FL, 32611-6131, USA.

Human perinatal tissues have been used for over a century as allogeneic biomaterials. Due to their advantageous properties including angiogenecity, anti-inflammation, anti-microbial, and immune privilege, these tissues are being utilized for novel applications across wide-ranging medical disciplines. Given continued clinical success, increased adoption of perinatal tissues as a disruptive technology platform has allowed for significant penetration into the multi-billion dollar biologics market. Here, we review current progress and future applications of perinatal biomaterials, as well as associated regulatory issues.
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http://dx.doi.org/10.1002/adhm.201700345DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5733692PMC
September 2017

3D Magnetic Stem Cell Aggregation and Bioreactor Maturation for Cartilage Regeneration.

J Vis Exp 2017 04 27(122). Epub 2017 Apr 27.

Laboratoire Matière et Systèmes Complexes (MSC), UMR 7057 CNRS and University Paris Diderot;

Cartilage engineering remains a challenge due to the difficulties in creating an in vitro functional implant similar to the native tissue. An approach recently explored for the development of autologous replacements involves the differentiation of stem cells into chondrocytes. To initiate this chondrogenesis, a degree of compaction of the stem cells is required; hence, we demonstrated the feasibility of magnetically condensing cells, both within thick scaffolds and scaffold-free, using miniaturized magnetic field sources as cell attractors. This magnetic approach was also used to guide aggregate fusion and to build scaffold-free, organized, three-dimensional (3D) tissues several millimeters in size. In addition to having an enhanced size, the tissue formed by magnetic-driven fusion presented a significant increase in the expression of collagen II, and a similar trend was observed for aggrecan expression. As the native cartilage was subjected to forces that influenced its 3D structure, dynamic maturation was also performed. A bioreactor that provides mechanical stimuli was used to culture the magnetically seeded scaffolds over a 21-day period. Bioreactor maturation largely improved chondrogenesis into the cellularized scaffolds; the extracellular matrix obtained under these conditions was rich in collagen II and aggrecan. This work outlines the innovative potential of magnetic condensation of labeled stem cells and dynamic maturation in a bioreactor for improved chondrogenic differentiation, both scaffold-free and within polysaccharide scaffolds.
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http://dx.doi.org/10.3791/55221DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5565124PMC
April 2017

The consequence of biologic graft processing on blood interface biocompatibility and mechanics.

Cardiovasc Eng Technol 2015 Sep;6(3):303-13

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, PO Box 116131, 1275 Center Drive, Gainesville, FL 32611.

Processing ex vivo derived tissues to reduce immunogenicity is an effective approach to create biologically complex materials for vascular reconstruction. Due to the sensitivity of small diameter vascular grafts to occlusive events, the effect of graft processing on critical parameters for graft patency, such as peripheral cell adhesion and wall mechanics, requires detailed analysis. Isolated human umbilical vein sections were used as model allogenic vascular scaffolds that were processed with either: 1. sodium dodecyl sulfate (SDS), 2. ethanol/acetone (EtAc), or 3. glutaraldehyde (Glu). Changes in material mechanics were assessed via uniaxial tensile testing. Peripheral cell adhesion to the opaque grafting material was evaluated using an innovative flow chamber that allows direct observation of the blood-graft interface under physiological shear conditions. All treatments modified the grafts tensile strain and stiffness properties, with physiological modulus values decreasing from Glu 240±12 kPa to SDS 210±6 kPa and EtAc 140±3 kPa, P<.001. Relative to glutaraldehyde treatments, neutrophil adhesion to the decellularized grafts increased, with no statistical difference observed between SDS or EtAc treatments. Early platelet adhesion (% surface coverage) showed no statistical difference between the three treatments; however, quantification of platelet aggregates was significantly higher on SDS scaffolds compared to EtAc or Glu. Tissue processing strategies applied to the umbilical vein scaffold were shown to modify structural mechanics and cell adhesion properties, with the EtAc treatment reducing thrombotic events relative to SDS treated samples. This approach allows time and cost effective prescreening of clinically relevant grafting materials to assess initial cell reactivity.
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http://dx.doi.org/10.1007/s13239-015-0221-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4548972PMC
September 2015

In vitro method for real-time, direct observation of cell-vascular graft interactions under simulated blood flow.

Tissue Eng Part C Methods 2014 Feb 24;20(2):116-28. Epub 2013 Aug 24.

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida , Gainesville, Florida.

In the development of engineered vascular grafts, assessing the material's interactive properties with peripheral blood cells and its capacity to endothelialize are important for predicting in vivo graft behavior. Current in vitro techniques used for characterizing cell adhesion at the surface of engineered scaffolds under flow only facilitate a terminal quantification of cell/surface interactions. Here, we present the design of an innovative flow chamber for real-time analysis of blood-biomaterial interactions under controllable hemodynamic conditions. Decellularized human umbilical veins (dHUV) were used as model vascular allografts to characterize platelet, leukocyte, and endothelial cell (EC) adhesion dynamics. Confluent EC monolayers adhered to the lumenal surface of the grafting material were flow conditioned to resist arterial shear stress levels (up to 24 dynes/cm(2)) over a 48 h period, and shown to maintain viability over the 1 week assessment period. The basement membrane was imaged while whole blood/neutrophil suspensions were perfused across the HUV surface to quantify cell accumulation. This novel method facilitates live visualization of dynamic events, including cell adhesion, migration, and morphological adaptation at the blood-graft interface on opaque materials, and it can be used for preliminary assessment of clinically relevant biomaterials before implantation.
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http://dx.doi.org/10.1089/ten.TEC.2012.0771DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3910478PMC
February 2014

Preimplantation processing of ex vivo-derived vascular biomaterials: effects on peripheral cell adhesion.

J Biomed Mater Res A 2013 Jan 24;101(1):123-31. Epub 2012 Jul 24.

J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, Gainesville, Florida 32611, USA.

The use of ex vivo-derived scaffolds as vascular conduits has shown to be a clinically valid approach to repair or bypass occluded vessels. Implantation of allogeneic tissue grafts requires careful processing to lower immunogenicity and prevent bacterial infection. However, the mechanical/chemical treatments used to prepare biological scaffolds can result in significant alterations to the native structure and surface chemistry, which can affect in vivo performance. Of particular importance for vascular grafts are binding interactions between the implanted biomaterial and host cells from the circulation and adjacent vasculature. Here we present a comparison of four strategies used to decellularize allogeneic human umbilical vein (HUV) scaffolds: ethanol/acetone, sodium chloride, sodium dodecyl sulfate (SDS), or Triton X-100. Scanning electron microscopy revealed that all four techniques achieved removal of native cells from both the lumenal and ablumenal surfaces of HUV grafts. Platelets and promyelocytic HL-60 cells showed preferential binding on the more loosely structured ablumenal surface, although low surface coverage was observed overall by peripheral blood cells. Vascular endothelial cell adhesion was highest on HUV decellularized using ethanol/acetone, and significantly higher than on SDS-processed grafts (p = 0.016). Primary cells showed high viability on the lumenal surface regardless of decellularization technique (over 95% in all cases). These results demonstrate the critical effects of various chemical processing strategies on the adhesive properties of ex vivo-derived vascular grafts. Careful application-specific consideration is warranted when selecting a processing strategy that minimizes innate responses (e.g. thrombosis, inflammation) that are often deleterious to graft survival.
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http://dx.doi.org/10.1002/jbm.a.34308DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3505264PMC
January 2013

The role of fibrinogen spacing and patch size on platelet adhesion under flow.

Acta Biomater 2012 Nov 20;8(11):4080-91. Epub 2012 Jul 20.

University of Oklahoma Bioengineering Center, University of Oklahoma, Norman, OK 73019, USA.

Platelet adhesion to the vessel wall during vascular injury is mediated by platelet glycoproteins binding to their respective ligands on the vascular wall. In this study we investigated the roles that ligand patch spacing and size play in regulating platelet interactions with fibrinogen under hemodynamic flow conditions. To regulate the size and distance between patches of fibrinogen we developed a photolithography-based technique to fabricate patterns of proteins surrounded by a protein-repellant layer of poly(ethylene glycol). We demonstrate that when mepacrine labeled whole blood is perfused at a shear rate of 100 s ⁻¹ over substrates patterned with micron-sized wide lines of fibrinogen, platelets selectively adhere to the areas of patterned fibrinogen. Using fluorescent and scanning electron microscopy we demonstrate that the degree of platelet coverage (3-35%) and the ability of platelet aggregates to grow laterally are dependent upon the distance (6-30 μm) between parallel lines of fibrinogen. We also report on the effects of fibrinogen patch size on platelet adhesion by varying the size of the protein patch (2-20 μm) available for adhesion, demonstrating that the downstream length of the ligand patch is a critical parameter in platelet adhesion under flow. We expect that these results and protein patterning surfaces will be useful in understanding the spatial and temporal dynamics of platelet adhesion under physiologic flow, and in the development of novel platelet adhesion assays.
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http://dx.doi.org/10.1016/j.actbio.2012.07.013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3462277PMC
November 2012
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