scholarly journals Fabrication of Hollow Structures in Photodegradable Hydrogels Using a Multi-Photon Excitation Process for Blood Vessel Tissue Engineering

Micromachines ◽  
2020 ◽  
Vol 11 (7) ◽  
pp. 679
Author(s):  
Uran Watanabe ◽  
Shinji Sugiura ◽  
Masayuki Kakehata ◽  
Fumiki Yanagawa ◽  
Toshiyuki Takagi ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Blood Vessel ◽  
Microfluidic Device ◽  
Vascular Function ◽  
Laser Scanning ◽  
Crosslinking Reaction ◽  
Excitation Process ◽  
Hollow Structures ◽  
Photon Excitation

Engineered blood vessels generally recapitulate vascular function in vitro and can be utilized in drug discovery as a novel microphysiological system. Recently, various methods to fabricate vascular models in hydrogels have been reported to study the blood vessel functions in vitro; however, in general, it is difficult to fabricate hollow structures with a designed size and structure with a tens of micrometers scale for blood vessel tissue engineering. This study reports a method to fabricate the hollow structures in photodegradable hydrogels prepared in a microfluidic device. An infrared femtosecond pulsed laser, employed to induce photodegradation via multi-photon excitation, was scanned in the hydrogel in a program-controlled manner for fabricating the designed hollow structures. The photodegradable hydrogel was prepared by a crosslinking reaction between an azide-modified gelatin solution and a dibenzocyclooctyl-terminated photocleavable tetra-arm polyethylene glycol crosslinker solution. After assessing the composition of the photodegradable hydrogel in terms of swelling and cell adhesion, the hydrogel prepared in the microfluidic device was processed by laser scanning to fabricate linear and branched hollow structures present in it. We introduced a microsphere suspension into the fabricated structure in photodegradable hydrogels, and confirmed the fabrication of perfusable hollow structures of designed patterns via the multi-photon excitation process.

scholarly journals Sustained VEGF delivery via PLGA nanoparticles promotes vascular growth

2010 ◽  
Vol 298 (6) ◽  
pp. H1959-H1965 ◽  
Author(s):  
Justin S. Golub ◽  
Young-tae Kim ◽  
Craig L. Duvall ◽  
Ravi V. Bellamkonda ◽  
Divya Gupta ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Blood Vessel ◽  
Femoral Artery ◽  
Release Kinetics ◽  
Aortic Ring ◽  
Plga Nanoparticles ◽  
Cardiovascular Medicine ◽  
Vessel Growth ◽  
Vessel Volume

Technologies to increase tissue vascularity are critically important to the fields of tissue engineering and cardiovascular medicine. Currently, limited technologies exist to encourage angiogenesis and arteriogenesis in a controlled manner. In the present study, we describe an injectable controlled release system consisting of VEGF encapsulated in poly(lactic- co-glycolic acid) (PLGA) nanoparticles (NPs). The majority of VEGF was released gradually over 2–4 days from the NPs as determined by an ELISA release kinetics experiment. An in vitro aortic ring bioassay was used to verify the bioactivity of VEGF-NPs compared with empty NPs and no treatment. A mouse femoral artery ischemia model was then used to measure revascularization in VEGF-NP-treated limbs compared with limbs treated with naked VEGF and saline. 129/Sv mice were anesthetized with isoflurane, and a region of the common femoral artery and vein was ligated and excised. Mice were then injected with VEGF-NPs, naked VEGF, or saline. After 4 days, three-dimensional microcomputed tomography angiography was used to quantify vessel growth and morphology. Mice that received VEGF-NP treatment showed a significant increase in total vessel volume and vessel connectivity compared with 5 μg VEGF, 2.5 μg VEGF, and saline treatment (all P < 0.001). When the yield of the fabrication process was taken into account, VEGF-NPs were over an order of magnitude more potent than naked VEGF in increasing blood vessel volume. Differences between the VEGF-NP group and all other groups were even greater when only small-sized vessels under 300 μm diameter were analyzed. In conclusion, sustained VEGF delivery via PLGA NPs shows promise for encouraging blood vessel growth in tissue engineering and cardiovascular medicine applications.

A Biological 3D Printer for the Preparation of Tissue Engineering Micro-Channel Scaffold

2015 ◽  
Vol 645-646 ◽  
pp. 1290-1297 ◽  
Author(s):  
Ya Nan Zhang ◽  
Yuan Yuan Liu ◽  
Yu Li ◽  
Shuai Li ◽  
Qing Xi Hu
Keyword(s):  
Tissue Engineering ◽  
3D Printing ◽  
Laser Scanning ◽  
Inner Core ◽  
Vascular Supply ◽  
Hollow Fibers ◽  
Dead Cell ◽  
Layer By Layer ◽  
3D Scaffold

The clinical applications of tissue engineering are still limited by the lack of a functional vascular supply in tissue-engineered constructs. In order to improve the pre-vascularization of tissue-engineered scaffold during in vitro culture, in this study, based on three-dimensional (3D) printing technology, using the crosslinking effect of coaxial fluids (sodium alginate and CaCl2) to prepare vessel-like hollow gel fibers, then layer by layer overlapping into 3D scaffold. The biological 3D printing platform was successfully developed and a coaxial nozzle module was introduced to generate a CaCl2-in-Alginate coaxial microfluidic. The inner core diameters of the prepared hollow gel fibers were 220~380 micrometers. In addition, the influence of materials concentration and dispensing rates on hollow fiber dimension were investigated, the cell-encapsulated in the printed hollow fibers was realized and the viability of endothelial cells (ECs) was studied with Laser scanning confocal microscopy (LSCM) and Live-Dead cell staining. The 3D scaffold built by hollow fibers could improve the phenomenon of diffusion constrain and enhance the survival rate of those ECs growing at a greater depth in the construct. This study provides a new theoretical basis for the vascularization of bone scaffold.

scholarly journals Two-Photon Calcium Imaging of Network Activity in XFP-Expressing Neurons in the Mouse

2007 ◽  
Vol 97 (4) ◽  
pp. 3118-3125 ◽  
Author(s):  
Jennifer M. Wilson ◽  
Daniel A. Dombeck ◽  
Manuel Díaz-Ríos ◽  
Ronald M. Harris-Warrick ◽  
Robert M. Brownstone
Keyword(s):  
Spinal Cord ◽  
Laser Scanning ◽  
Fluorescent Protein ◽  
Laser Scanning Microscopy ◽  
Network Activity ◽  
Spatiotemporal Pattern ◽  
Two Photon ◽  
Physiological Investigation ◽  
Photon Excitation

Fluorescent protein (XFP) expression in postnatal neurons allows the anatomical and physiological investigation of identified subpopulations of interneurons with established techniques. However, the spatiotemporal pattern of activity of these XFP neurons within a network and their role in the functional output of the network are more challenging issues to investigate. Here we apply two-photon excitation laser scanning microscopy to mouse spinal cord locomotor networks and present the methodology by which calcium activity can be recorded in XFP-expressing neurons. Such activity can be studied both in relation to neighboring non-XFP neurons in a spinal cord slice preparation and in relation to functional locomotor output monitored by ventral root activity in the intact in vitro spinal cord. Thus the network properties and functional correlates with locomotion of identified populations of interneurons can be studied simultaneously.

scholarly journals Long-term viability of coronary artery smooth muscle cells on poly( l -lactide- co -ϵ-caprolactone) nanofibrous scaffold indicates its potential for blood vessel tissue engineering

2008 ◽  
Vol 5 (26) ◽  
pp. 1109-1118 ◽  
Author(s):  
Yixiang Dong ◽  
Thomas Yong ◽  
Susan Liao ◽  
Casey K Chan ◽  
S Ramakrishna
Keyword(s):  
Tissue Engineering ◽  
Smooth Muscle ◽  
Coronary Artery ◽  
Blood Vessel ◽  
Smooth Muscle Cells ◽  
Material Selection ◽  
Muscle Cells ◽  
Cell Viability Assay

Biodegradable polymer nanofibres have been extensively studied as cell culture scaffolds in tissue engineering. However, long-term in vitro studies of cell–nanofibre interactions were rarely reported and successful organ regeneration using tissue engineering techniques may take months (e.g. blood vessel tissue engineering). Understanding the long-term interaction between cells and nanofibrous scaffolds (NFS) is crucial in material selection, design and processing of the tissue engineering scaffolds. In this study, poly( l -lactide- co -ϵ-caprolactone) [P(LLA-CL)] (70 : 30) copolymer NFS were produced by electrospinning. Porcine coronary artery smooth muscle cells (PCASMCs) were seeded and cultured on the scaffold to evaluate cell–nanofibre interactions for up to 105 days. A favourable interaction between this scaffold and PCASMCs was demonstrated by cell viability assay, scanning electron microscopy, histological staining and extracellular matrix (ECM) secretion. Degradation behaviours of the scaffolds with or without PCASMC culture were determined by mechanical testing and gel permeation chromatography (GPC). The results showed that the PCASMCs attached and proliferated well on the P(LLA-CL) NFS. Large amount of ECM protein secretion was observed after 50 days of culture. Multilayers of aligned oriented PCASMCs were formed on the scaffold after two months of in vitro culture. In the degradation study, the PCASMCs were not shown to significantly increase the degradation rate of the scaffolds for up to 105 days of culture. The in vitro degradation time of the scaffold could be as long as eight months by extrapolating the results from GPC. These observations further supported the potential use of the P(LLA-CL) nanofibre in blood vessel tissue engineering.

“Open-top” microfluidic device for in vitro three-dimensional capillary beds

Lab on a Chip ◽  
2017 ◽  
Vol 17 (20) ◽  
pp. 3405-3414 ◽  
Author(s):  
Soojung Oh ◽  
Hyunryul Ryu ◽  
Dongha Tahk ◽  
Jihoon Ko ◽  
Yoojin Chung ◽  
...  
Keyword(s):  
Blood Vessel ◽  
Microfluidic Device ◽  
Three Dimensional ◽  
Vessel Network ◽  
Blood Vessel Network

We introduce a novel microfluidic device to co-culture a blood vessel network and cell tissues in an in vivo-like niche.

scholarly journals Vascular Tissue Engineering: Challenges and Requirements for an Ideal Large Scale Blood Vessel

2021 ◽  
Vol 9 ◽  
Author(s):  
Chloé D. Devillard ◽  
Christophe A. Marquette
Keyword(s):  
Tissue Engineering ◽  
Blood Vessel ◽  
Large Scale ◽  
Vascular Tissue ◽  
Autologous Blood ◽  
Biological Research ◽  
Cell Sources ◽  
Complex Engineering ◽  
Tissue Grafts

Since the emergence of regenerative medicine and tissue engineering more than half a century ago, one obstacle has persisted: the in vitro creation of large-scale vascular tissue (&gt;1 cm3) to meet the clinical needs of viable tissue grafts but also for biological research applications. Considerable advancements in biofabrication have been made since Weinberg and Bell, in 1986, created the first blood vessel from collagen, endothelial cells, smooth muscle cells and fibroblasts. The synergistic combination of advances in fabrication methods, availability of cell source, biomaterials formulation and vascular tissue development, promises new strategies for the creation of autologous blood vessels, recapitulating biological functions, structural functions, but also the mechanical functions of a native blood vessel. In this review, the main technological advancements in bio-fabrication are discussed with a particular highlights on 3D bioprinting technologies. The choice of the main biomaterials and cell sources, the use of dynamic maturation systems such as bioreactors and the associated clinical trials will be detailed. The remaining challenges in this complex engineering field will finally be discussed.

scholarly journals Production of Cell Enclosing Silk Derivative Microsphere in Uniform Size Distribution Through Coaxial Microfluidic Device and Horseradish Crosslinking Reaction

2021 ◽  
Author(s):  
Elham Badali ◽  
Mahshid Hosseini ◽  
Narges Mahmoodi ◽  
Sajad Hassanzadeh ◽  
Vajihe Taghdiri Nooshabadi ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Silk Fibroin ◽  
Microfluidic Device ◽  
Gelation Time ◽  
Biochemical Properties ◽  
Mitochondrial Activity ◽  
Crosslinking Reaction ◽  
Uniform Size ◽  
Cell Matrix ◽  
Cell Matrix Interactions

Abstract BackgroundSilk fibroin (SF) as a natural polymer holds great potential in biomedical research because of its biocompatibility, easy processing, high toughness, and strength. However, slow gelation time has narrowed its applications, specifically in cell-laden microparticles that are versatile structures for tissue engineering due to their unique features. In addition, most crosslinking methods used to decrease gelation time did not occur in a mid-condition. Methods This study aimed to use modified SF with phenol conjugation to accelerate crosslinking mediated via horseradish peroxidase (HRP)/ hydrogen peroxide (H2O2) in a co-flow high-throughput microfluidic device for the ultimate goal of cell-laden silk fibroin-phenol (SF-Ph) microparticles formation. The physical and biochemical properties of fabricated cell-laden SF-Ph were evaluated to reveal its potential for tissue engineering.ResultsThe monodisperse microparticles in shape and size were formed in various diameters changing from 300 to 80 µm by altering oil phase velocity from SF-Ph substrate. More than 90% cell viability and three times cells upregulation of mitochondrial activity of enclosed-cells in microparticles with 150 ± 32 µm diameters revealed that these structures were suitable subcultures produced through a mild process based on morphological and MTT assays. It was noticed that cells approximately cover the microparticles until the 15th day. ConclusionSpherical micro-tissue formation in microparticles, resulting from cell growth promoted by cell-cell and cell-matrix interactions, adds significant weight to this method's applications.

Modulation of Fibrin Gel Extracellular Matrix Properties by Fibrinogen and Thrombin Concentrations for Angiogenesis Assay

2014 ◽  
Vol 911 ◽  
pp. 342-346
Author(s):  
Siti Amirah Ishak ◽  
Irza Sukmana
Keyword(s):  
Tissue Engineering ◽  
Extracellular Matrix ◽  
Blood Vessel ◽  
Clotting Time ◽  
Fibrin Gel ◽  
Angiogenesis Assay ◽  
Matrix Properties ◽  
In Vitro Angiogenesis

Angiogenesis is the formation of new microvascular network from the pre-existing blood vessel. In tissue engineering approaches, angiogenesis is essential for the promotion of micro-vascular network inside an engineered scaffold construct, mimicking a functional blood vessel in vivo. In the in vivo system, the formation of new blood vessels depends on the properties fibrin gel extracellular matrix. In this study, we have investigated the effect of different fibrinogen and thrombin composition on the biophysical properties of fibrin gel. Higher concentration of thrombin (4.0 Units/milliliter) yields a shorter clotting time of the fibrin gel and result in better water uptake property while at lower concentration of thrombin (0.5 Units/milliliter), the clotting time takes much longer. Also, at lowest concentration ratio of fibrinogen to thrombin (0.5 milligram/milliliter to 4.0 Units/milliliter), the turbidity study shows the lowest absorbance compared to other samples. Different concentration of fibrinogen and thrombin also affect the microstructure of the fibrin gel. The variation of these properties will be then manipulated to be used for in vitro angiogenesis. This study opens broader application of fibrin extracellular matrix in regenerative medicine and tissue engineering researches.

Vacuoles Induced in Mammalian Cells by Helicobacter Cytotoxin are Acidic Organelles

1990 ◽  
Vol 48 (3) ◽  
pp. 168-169
Author(s):  
M. H. Chestnut ◽  
C. E. Catrenich
Keyword(s):  
Acridine Orange ◽  
Mammalian Cells ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Ulcer Disease ◽  
Gastroduodenal Disease ◽  
H Pylori

Helicobacter pylori is a non-invasive, Gram-negative spiral bacterium first identified in 1983, and subsequently implicated in the pathogenesis of gastroduodenal disease including gastritis and peptic ulcer disease. Cytotoxic activity, manifested by intracytoplasmic vacuolation of mammalian cells in vitro, was identified in 55% of H. pylori strains examined. The vacuoles increase in number and size during extended incubation, resulting in vacuolar and cellular degeneration after 24 h to 48 h. Vacuolation of gastric epithelial cells is also observed in vivo during infection by H. pylori. A high molecular weight, heat labile protein is believed to be responsible for vacuolation and to significantly contribute to the development of gastroduodenal disease in humans. The mechanism by which the cytotoxin exerts its effect is unknown, as is the intracellular origin of the vacuolar membrane and contents. Acridine orange is a membrane-permeant weak base that initially accumulates in low-pH compartments. We have used acridine orange accumulation in conjunction with confocal laser scanning microscopy of toxin-treated cells to begin probing the nature and origin of these vacuoles.

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