Journal: Brain Sciences

Article Title: Histopathological Investigation of Dura-like Membrane in Vestibular Schwannomas

doi: 10.3390/brainsci11121649

Figure Lengend Snippet: Immunohistopathological analysis of CD34 staining. ( A ) Microvessel density (MVD) and diameter analyzed using CD34 staining are shown. Large CD34(+) vessels are observed in the tumors with DLM. Few CD34(+) vessels are identified in DLM (scale bar= 100 μm). ( B ) No significant difference is observed in the MVD in the tumors between DLM group and non-DLM groups ( p = 0.19). Vessel diameters are significantly larger in DLM group than that in non-DLM group ( p < 0.01). ( C ) The correlation between tumor volume and MVD are shown. The larger-sized VSs demonstrate higher MVD in the tumor with DLM ( p < 0.05, r = 0.89).

Article Snippet: To assess microvessel density (MVD) and vessel diameter, tissue sections were screened using CD34 immunohistochemistry in low-power fields and the three most vascularized regions (hot spots) were selected for automatic microvessel counting under high-power magnification microscopy (0.95 mm 2 ) (Biorevo BZ-9000, KEYENCE, Osaka, Japan), as previously described [ ].

Techniques: Staining

Journal: American Journal of Physiology - Heart and Circulatory Physiology

Article Title: Cell-generated traction forces and the resulting matrix deformation modulate microvascular alignment and growth during angiogenesis

doi: 10.1152/ajpheart.00995.2013

Figure Lengend Snippet: Z-projections from confocal images of microvessels after day 6. A: hexahedral boundary conditions U, LAC, and SAC. During imaging, the long axis of hexahedral constructs was aligned with the horizontal (x-) axis of the imaging plane. B: circular boundary conditions, circular unconstrained (CU) and circular constrained (CC).

Article Snippet: Microvessel fragments were isolated from epididymal fat pads harvested from male Sprague-Dawley rats as described previously ( 17 ).

Techniques: Imaging, Construct

Journal: American Journal of Physiology - Heart and Circulatory Physiology

Article Title: Cell-generated traction forces and the resulting matrix deformation modulate microvascular alignment and growth during angiogenesis

doi: 10.1152/ajpheart.00995.2013

Figure Lengend Snippet: Quantitative measurements of microvessel growth for different boundary conditions. A: microvessel vascularity tended to decrease as gels became more constrained. Vascularity for the long-short axis constrained (LSAC) group was significantly lower than that in the U (*P = 0.004) and LAC groups (*P = 0.044). Vascularity in the CC boundary condition was significantly less than that in the CU condition (*P = 0.017). B: average segment length tended to increase as the gel became more constrained. Segment length in the CC cultures was significantly greater than that in the CU cultures (*P = 0.043). C: microvessel branching tended to decrease as the gel became more constrained. Branching within the CC boundary condition culture was significantly reduced relative to the unconstrained control, CU (*P = 0.020). Bars indicate standard error.

Article Snippet: Microvessel fragments were isolated from epididymal fat pads harvested from male Sprague-Dawley rats as described previously ( 17 ).

Techniques: Control

Journal: American Journal of Physiology - Heart and Circulatory Physiology

Article Title: Cell-generated traction forces and the resulting matrix deformation modulate microvascular alignment and growth during angiogenesis

doi: 10.1152/ajpheart.00995.2013

Figure Lengend Snippet: Microvessel alignment and anisotropy within the XY-plane. A: length-normalized distributions of orientation for hexahedral constructs. The 10° bin represents angles from 0° to 10°, which are parallel to the long axis (x-axis). The 90° bin represents angles from 80° to 90° and is parallel to the short axis (y-axis). The LAC group was significantly different from at least 1 other boundary condition in all but the 30° and 40° angle bins (*P < 0.05). A completely random distribution should have ∼11% in each angle bin, represented by the dashed line. B: microvessel anisotropy as determined by fast Fourier transform (FFT) analysis. A value of 0.0 represents random organization, whereas a value of 1.0 represents complete alignment. The LAC boundary condition had a significantly higher anisotropy value than all the other boundary conditions, as expected based on the angle distributions in A. The cylindrical constructs had similar anisotropy values to the other randomly organized hexahedral boundary conditions. The LAC group had a high anisotropy value of 0.7, which was significantly greater than all other conditions when analyzed by FFT (*P < 0.001). Bars indicate standard error.

Article Snippet: Microvessel fragments were isolated from epididymal fat pads harvested from male Sprague-Dawley rats as described previously ( 17 ).

Techniques: Construct

Journal: American Journal of Physiology - Heart and Circulatory Physiology

Article Title: Cell-generated traction forces and the resulting matrix deformation modulate microvascular alignment and growth during angiogenesis

doi: 10.1152/ajpheart.00995.2013

Figure Lengend Snippet: Microvessel alignment relative to the z-axis. Length-normalized distributions of orientation for the hexahedral constructs (A) and cylindrical constructs (B) are shown. The 10° bin represents angles from 0° to 10°, which are parallel to the z-axis of the hexahedral constructs. The 90° bin represents angles from 80° to 90° and is parallel to the XY-plane of the hexahedral constructs. Most of the vessel segments for all of the boundary conditions were parallel to the XY-plane. Significant differences were detected between U and the other groups in the 70°, 80°, and 90° angle bins (*P < 0.01). The unconstrained cultures U and CU were significantly different than the other constrained cultures in all but the 10° and 20° angle bins (*P < 0.05). Bars indicate standard error.

Article Snippet: Microvessel fragments were isolated from epididymal fat pads harvested from male Sprague-Dawley rats as described previously ( 17 ).

Techniques: Construct

Journal: American Journal of Physiology - Heart and Circulatory Physiology

Article Title: Cell-generated traction forces and the resulting matrix deformation modulate microvascular alignment and growth during angiogenesis

doi: 10.1152/ajpheart.00995.2013

Figure Lengend Snippet: Finite element (FE) simulations of microvessel-induced contraction of the constrained vascularized constructs. A: LAC model is shown at left, and the SAC model is shown at right. The initial configuration is shown above, and the deformed configuration (i.e., day 6 of growth) is shown below. B–E: global engineering strain was calculated from both the experimental constructs (black) and FE models (gray) using edge displacement and the definition (L–Lo)/Lo. Local strain, or the principal components of the infinitesimal strain tensor, was measured for the geometric center of the FE models and reported in white. Engineering strain was calculated along the X-, Y-, and Z-dimension (B and C), and these results were used to calculate the volume change ratio that occurred over the deformation (E). Bars indicate standard error. No bars are present on the FE data (i.e., N = 1 simulation).

Article Snippet: Microvessel fragments were isolated from epididymal fat pads harvested from male Sprague-Dawley rats as described previously ( 17 ).

Techniques: Construct

Journal: American Journal of Physiology - Heart and Circulatory Physiology

Article Title: Cell-generated traction forces and the resulting matrix deformation modulate microvascular alignment and growth during angiogenesis

doi: 10.1152/ajpheart.00995.2013

Figure Lengend Snippet: Anisotropic deformation of the matrix due to cellular contractility is sufficient to cause microvascular alignment, independent of any other cellular behavior. Mapping the displacement field from the LAC FE model onto a distribution of randomly oriented microvessels results in a new alignment pattern that closely resembles alignment within the experimental LAC constructs. A: day 0, initial mesh geometry with the microvessel dataset positioned at geometric center. B: day 6, with the use of FE displacement field to map the microvessels, a new alignment pattern emerged due to contraction in the Y- and Z-directions. C: length-normalized distributions microvessel angle with the long axis (x-axis). The 10° angle bin represents angles from 0° to 10°, which were parallel to the long axis (x-axis). The 90° angle bin represents angles from 80° to 90° and was parallel to the short axis (y-axis). The angle orientation distribution before mapping (●) indicates random microvessel orientation. After mapping (○), microvessels were preferentially aligned along the long axis. Orientation data from LAC construct experiments are shown as well (▲). Error bars indicate standard error. Sim, simulation.

Article Snippet: Microvessel fragments were isolated from epididymal fat pads harvested from male Sprague-Dawley rats as described previously ( 17 ).

Techniques: Construct

Journal: Science Advances

Article Title: Biomimetic human small muscular pulmonary arteries

doi: 10.1126/sciadv.aaz2598

Figure Lengend Snippet: ( A ) Color photomicrograph of an adult hSMPA at the level of the respiratory bronchiole shown in cross section with hematoxylin and eosin staining. Black arrows indicate the EC of the intimal layer (E) and the VSMCs of the muscularis (M). Scale bar, 20 μm. Original magnification, ×200. This native artery exhibits several important structural characteristics including multicellular layering, curvature, and patterning. ( B ) Schematic illustration of the biomimetic hSMPA featuring patterning of cells and layering of VSMCs (M), laminin, and ECs (E). ( C ) Schematic illustration of the highly parallel, multistep patterning and assembly process for biomimetic hSMPA. Germanium (Ge) and bilayers of optically transparent silicon oxide and silicon dioxide (SiO/SiO 2 ) were deposited on silicon wafers using electron-beam evaporation, followed by adhesive protein patterning and cell layering, which were all achieved in 2D. Upon dissolution of the sacrificial germanium layer in cell culture medium, the 2D bilayer films were released and then self-folded into tubes. Additional fabrication details are shown in the schematic in fig. S1, and snapshots of the roll-up process are shown in fig. S2. Fn, fibronectin; Lm, laminin. ( D ) Confocal microscope images of tubular constructs with tunable 1 and 2 mm length and protein pattern. Luminal surfaces of the tubular constructs were patterned with fluorescently labeled fibronectin (red) or bovine serum albumin (green). The distribution of protein fluorescence intensity is shown in fig. S8A. For cell culture, fibronectin without fluorescence labeling was used. Scale bar, 500 μm. ( E ) Epifluorescence images of rhodamine-phalloidin–labeled ECs growing on the luminal surfaces of biomimetic microvessels. Scale bar, 500 μm.

Article Snippet: A confluent monolayer of HPMEC, with tightly contiguous vascular endothelial (VE)–cadherin–mediated junctions, was visualized on the luminal surface of the biomimetic microvessel 48 hours after cell seeding ( ; movie S1; and figs. S3A, S4A, and S6).

Techniques: Staining, Evaporation, Adhesive, Dissolution, Cell Culture, Microscopy, Construct, Labeling, Fluorescence

Journal: Science Advances

Article Title: Biomimetic human small muscular pulmonary arteries

doi: 10.1126/sciadv.aaz2598

Figure Lengend Snippet: Reconstructions of confocal Z -stacks of biomimetic microvessels populated by HPMECs, featuring the following: ( A ) VE-cadherin at endothelial adherens junctions (antibody labeling, green), ( B ) F-actin (phalloidin, red), and ( C ) merged images that include nuclei [4′,6-diamidino-2-phenylindole(DAPI), gray scale]. Insets in (A) and (B) show the cross-sectional views in the corresponding color channel and demonstrate the amount of overlap due to the roll-up process. Scale bars, 100 μm. A green-magenta rendering of this figure is shown in fig. S6.

Article Snippet: A confluent monolayer of HPMEC, with tightly contiguous vascular endothelial (VE)–cadherin–mediated junctions, was visualized on the luminal surface of the biomimetic microvessel 48 hours after cell seeding ( ; movie S1; and figs. S3A, S4A, and S6).

Techniques: Antibody Labeling

Journal: Science Advances

Article Title: Biomimetic human small muscular pulmonary arteries

doi: 10.1126/sciadv.aaz2598

Figure Lengend Snippet: ( A ) Confocal image (side view) of HPASMCs on an unpatterned tubular construct, stained for F-actin (phalloidin, red), smooth muscle α-actin (antibody labeling, green), and nuclei (DAPI, gray scale). The polar plot is based on image analysis of the gray scale F-actin image and shows that without patterning, HPASMCs attached and spread with random orientation. ( B ) Confocal image (side view) of HPASMCs grown on a patterned tubular construct, stained for F-actin (phalloidin, red), smooth muscle α-actin (antibody labeling, green), and nuclei (DAPI, gray scale). On the basis of image analysis of the gray scale F-actin image, the polar plot shows that F-actin filaments demonstrated alignment in parallel helical structures on the fibronectin-patterned scaffold. Binning in the polar plots is 10°. ( C ) 3D views of biomimetic microvessels demonstrating tunable variations in orientation angles and patterning periodicity, with labeled F-actin (phalloidin, red), smooth muscle α-actin (antibody labeling, green), and nuclei (DAPI, gray scale). Scale bars in (A to C), 100 μm.

Article Snippet: A confluent monolayer of HPMEC, with tightly contiguous vascular endothelial (VE)–cadherin–mediated junctions, was visualized on the luminal surface of the biomimetic microvessel 48 hours after cell seeding ( ; movie S1; and figs. S3A, S4A, and S6).

Techniques: Construct, Staining, Antibody Labeling, Labeling

Journal: Science Advances

Article Title: Biomimetic human small muscular pulmonary arteries

doi: 10.1126/sciadv.aaz2598

Figure Lengend Snippet: ( A ) Cell viability was calculated as the percentage of live cells compared with baseline (day 3) using the CyQUANT assay. Coculture of HPMEC and HPASMC populations were assayed. Data from flat (red) and tubular constructs (green) were compared. Data are displayed as means ± SEM. ( B ) HPMECs in the biomimetic microvessels exhibited a fourfold rise in nitric oxide production over the levels observed in HPMEC monolayers in flat culture. Nitrite levels in cell culture medium were determined by using a Sievers bioluminescence nitric oxide analyzer. Nitrite levels were normalized to total protein content for each sample. The numbers in the y axis indicate picomoles of nitrite per micrograms of protein. Medium was collected 48 hours after confluency and medium replacement. Data are displayed as means ± SEM (** P < 0.01). ( C ) Phosphorylation of eNOS (at Ser 1177 ) was found to be greater in HPMECs that were seeded and cultured on biomimetic microvessels than that in cells on flat SiO/SiO 2 films. HPMECs were grown at each condition for 48 hours. Western blots of cell lysates from each population were analyzed with antibodies against phosphorylated eNOS (p-eNOS), total eNOS, and β-tubulin (protein loading control). Data are displayed as means ± SEM (** P < 0.01).

Article Snippet: A confluent monolayer of HPMEC, with tightly contiguous vascular endothelial (VE)–cadherin–mediated junctions, was visualized on the luminal surface of the biomimetic microvessel 48 hours after cell seeding ( ; movie S1; and figs. S3A, S4A, and S6).

Techniques: CyQUANT Assay, Construct, Cell Culture, Phospho-proteomics, Western Blot, Control