microvascular networks  (Nikon)

Journal: bioRxiv

Article Title: The Batch-Resourcing Angiogenesis Tool (BRAT) to enable high-content microscopy screening of microvascular networks

doi: 10.1101/2025.01.24.634836

Figure Lengend Snippet: GFP-hAECs co-cultured with hAD-MSCs for 7 days to form microvascular networks, with images captured every 10 minutes, as . BRAT and AngioTool were applied to measure microvascular network (A) end points, (B) branch points, and (C) vessel area as a percentage of the field of view area. (D) BRAT can also measure average or total vessel diameter and length (distances between end or branch points). Hashed vertical lines represent media change times. Individual timepoint analysis (dots) are overlaid with a regression and 95% confidence intervals. These results are compared with REAVER in .

Article Snippet: Finally, co-cultures were counterstained with DAPI (4′,6-diamidino-2-phenylindole, dihydrochloride, A-21137, ThermoFisher) for 10 minutes, washed thrice with PBS, and then imaged for CD31-positive microvascular networks on a Nikon ECLIPSE Ti2 Inverted Microscope (using a 555 nm excitation laser, 590-650 nm detector).

Techniques: Cell Culture

Journal: bioRxiv

Article Title: The Batch-Resourcing Angiogenesis Tool (BRAT) to enable high-content microscopy screening of microvascular networks

doi: 10.1101/2025.01.24.634836

Figure Lengend Snippet: (A) 3D microvascular networks formed by GFP-hAECs and hAD-MSCs co-cultured in a hydrogel microchip. (B) A 3D microvascular network of CD31-stained umbilical cord blood-outgrowth ECs (BOECs) and hAD-MSCs co-cultured in a fibrin hydrogel. (C) BRAT shows hAECs form thicker, longer, but fewer microvessels in 3D hAD-MSC hydrogel co-culture as opposed to BOECs. (A, B) 100 µm scale bars. (C)’s dimensions are reported in µm using 0.81 µm/pixel (Etaluma 720S) or 0.73 µm/pixel (Nikon Ti2) conversion factors. (C) datapoints represent independent cultures. Barplots represent mean ± standard deviation. *p<0.05, **p<0.01, ***p<0.001. Additional BRAT-generated metrics can be found in Fig. S9.

Article Snippet: Finally, co-cultures were counterstained with DAPI (4′,6-diamidino-2-phenylindole, dihydrochloride, A-21137, ThermoFisher) for 10 minutes, washed thrice with PBS, and then imaged for CD31-positive microvascular networks on a Nikon ECLIPSE Ti2 Inverted Microscope (using a 555 nm excitation laser, 590-650 nm detector).

Techniques: Cell Culture, MicroChIP Assay, Staining, Co-Culture Assay, Standard Deviation, Generated

Journal: bioRxiv

Article Title: The Batch-Resourcing Angiogenesis Tool (BRAT) to enable high-content microscopy screening of microvascular networks

doi: 10.1101/2025.01.24.634836

Figure Lengend Snippet: (A) A comparison of BRAT versus the leading four open-source software’s accuracy, sensitivity (true-positive), and specificity (true-negative) of microvascular network (MVN) pixel segmentation versus gold-standard manual segmentation. (B) Accuracy and precision across automatically detected vessel length, area, diameter, and branch points with reference to manual segmentations. (C) The most challenging MVN image to segment from the REAVER dataset; an image of CD31 + MVN in cardiac tissue (left), beside a manual annotation (ground truth), BRAT’s automated annotation, and BRAT’s false-positive (FP; green) and false-negative (FN; red) incorrectly-labelled MVN pixels. (A, B) Boxplots represent mean, quartile ranges, while whiskers extend to show the distribution, less outlier points. The annotations above or below each plot indicate significant pairwise comparisons between groups with Bonferroni adjusted p-values (letters).

Article Snippet: Finally, co-cultures were counterstained with DAPI (4′,6-diamidino-2-phenylindole, dihydrochloride, A-21137, ThermoFisher) for 10 minutes, washed thrice with PBS, and then imaged for CD31-positive microvascular networks on a Nikon ECLIPSE Ti2 Inverted Microscope (using a 555 nm excitation laser, 590-650 nm detector).

Techniques: Comparison

Journal: bioRxiv

Article Title: The Batch-Resourcing Angiogenesis Tool (BRAT) to enable high-content microscopy screening of microvascular networks

doi: 10.1101/2025.01.24.634836

Figure Lengend Snippet: Co-cultured GFP-hAECs and hAD-MSCs form microvascular networks over 7 days, as imaged every 10 minutes. BRAT analysed all 886 timesteps, while AngioTool only analysed 89 timesteps. In the bioRxiv version, the video is a download in the ‘Supplemental Material’ tab on the right of the abstract.

Article Snippet: Finally, co-cultures were counterstained with DAPI (4′,6-diamidino-2-phenylindole, dihydrochloride, A-21137, ThermoFisher) for 10 minutes, washed thrice with PBS, and then imaged for CD31-positive microvascular networks on a Nikon ECLIPSE Ti2 Inverted Microscope (using a 555 nm excitation laser, 590-650 nm detector).

Techniques: Cell Culture

Journal: Nano Convergence

Article Title: Technology for the formation of engineered microvascular network models and their biomedical applications

doi: 10.1186/s40580-024-00416-7

Figure Lengend Snippet: Technology for the formation of engineered microvascular network models (Created at smart.servier.com )

Article Snippet: Fig. 5 Fabrication of a microvascular network by microfluidic systems.

Techniques:

Journal: Nano Convergence

Article Title: Technology for the formation of engineered microvascular network models and their biomedical applications

doi: 10.1186/s40580-024-00416-7

Figure Lengend Snippet: Fabrication of a microvascular network by photolithography. A Photolithographic approach to generate cellular micropatterns. a Crosslinked chitosan pattern after 180 s of UV exposure. Bar = 100 μm. b , c Engineered tubular structures of BCAEC and HUVEC scanned by confocal laser-scanning microscopy (CLSM). 3D images showed a lumen within the tubular structures. Bar = 20 μm. (Figure reprinted with permission from Ref. ). B , a – c SEM image of the backside lithography technique at different magnifications. We can observe the gradation in height according to the width of the channels. (Figure reprinted with permission from Ref. ). C Culture of primary human lung microvascular endothelial cells (HLMECs) in the hourglass-shaped channels. Scale bar, 50 µm. (Figure reprinted with permission from Ref. )

Article Snippet: Fig. 5 Fabrication of a microvascular network by microfluidic systems.

Techniques: Confocal Laser Scanning Microscopy

Journal: Nano Convergence

Article Title: Technology for the formation of engineered microvascular network models and their biomedical applications

doi: 10.1186/s40580-024-00416-7

Figure Lengend Snippet: Fabrication of a microvascular network by laser degradation. A Schematic of ablation and perfusion process of a human alveolus. B Recreation of mouse brain microvasculature. (Figure reprinted with permission from Ref. ). C Schematic diagram of microvascular fabrication in a multifunctional hydrogel biomaterial. a – f 3D endothelialized channels generated within photodegradable fluorescent gels. Ten days following microvascular endothelialization with HUVECs, F-actin is shown in red, and nuclei are shown in blue. Endothelialization of g , h 60 μm × 60 μm and i , j 45 μm × 45 μm (width × height) channels were obtained. (Figure reprinted with permission from Ref. ). D Left column: time-lapse images of PEGDA during laser-induced degradation of a 500 × 100 × 100 μm (x, y, z) channel. right column: As microbubbles form, they migrate to the reservoir and coalesce to form a large bubble. (Figure reprinted with permission from Ref. ). E Laser illumination using 145 nJ pulse energy results in structures in which both photoablation and cavitation-molded sections are present, indicating that this pulse energy is a threshold value at which the transition between the modes of photoablation and cavitation molding occurs. Scale bar, 10 µm. (Figure reprinted with permission from Ref. )

Article Snippet: Fig. 5 Fabrication of a microvascular network by microfluidic systems.

Techniques: Generated

Journal: Nano Convergence

Article Title: Technology for the formation of engineered microvascular network models and their biomedical applications

doi: 10.1186/s40580-024-00416-7

Figure Lengend Snippet: Fabrication of a microvascular network by 3D printing. A , a Schematic of three different crosslinking strategies for bioprinting photo-crosslinkable inks (e.g., 5 wt% MeHA shown here), where crosslinking occurs before (pre-crosslink), after (post-crosslink), or during (in situ crosslink) extrusion. b Representative images of nozzles with extruded material and printed lattice structure. (Figure reprinted with permission from Ref. ). B , a Schematic diagram based on a coaxial bioprinting method. b confocal fluorescence image of a bioprinted HUVEC embedded construct under 30 s UV exposure. (Figure reprinted with permission from Ref. ). C A laser printing method generates capillaries. Green fluorescent endothelial cells. The printed cells formed a tubular structure with a lumen. The applied laser pulse energy was 6 µJ, and the patterns in panel a on the left were printed twice in the same place. Scale bars are 200 µm ( a left and b left), 50 µm ( a right and b center), and 10 µm ( c right). (Figure reprinted with permission from Ref. )

Article Snippet: Fig. 5 Fabrication of a microvascular network by microfluidic systems.

Techniques: In Situ, Fluorescence, Construct

Journal: Nano Convergence

Article Title: Technology for the formation of engineered microvascular network models and their biomedical applications

doi: 10.1186/s40580-024-00416-7

Figure Lengend Snippet: Fabrication of a microvascular network by microfluidic systems. A Cross-sectional imaging thickness and Z-position = 10 μm for representative channels (optical channels) after 9 days of microfluidic perfusion culture of endothelial cells in a sacrificial lattice. (Figure reprinted with permission from Ref. ). B , a Schematic diagram of the printed vascular channel construct. b Fluorescence image of the printed vascular channel construct by wide-field microscope. HUVECs are visualized in red, beads flow in green. (Figure reprinted with permission from Ref. ). C Endothelialized channels are readily fabricated in the presence of encapsulated stromal cells. A single-layer channel was generated by photodegradation. Channels were then endothelialized with HUVECs, cultured for 4 days, fixed, and stained for F-actin (red). The sample is viewed ( a – c ) as Z-, X-, and Y-direction maximum intensity projections. (Figure reprinted with permission from Ref. ). D In vitro microvascular network model of the peritoneum. a PDMS mold with patterned channels were fabricated using soft lithography and bonded to a glass coverslip. The central gel region (green) contained cells and a fibrin hydrogel. The side channels and reservoirs (purple) as well as the top channel and reservoir (orange) were filled with cell culture medium. Scale bar, 3 mm. b A confocal microscopy image of the microvascular networks within the device, in which ECs express GFP, cell nuclei are stained with DAPI (blue), and lipid droplets in Acs are stained with LipidTox (white). Scale bar, 30 μm. (Figure reprinted with permission from Ref. ). E Create a tricompartmental model of the arteriole-to-capillary-to-venule microvasculature. Capillaries (middle) modeled by perfusable MVNs made from endothelial cells (EC, green) and fibroblasts (FB) in fibrin gel, venule (left) modeled by collagen channel with EC monolayer, arteriole (right) modeled by collagen channel with smooth muscle cells (SMC, magenta); the scale bar is 250 μm. (Figure reprinted with permission from Ref. ) F The modular microfluidic system combines two PDMS layers. The different morphological properties of the capillaries generated using diamond-, half-rectangle and rectangle-shaped chambers were analyzed. The rectangle-shaped tissue chambers generated the largest capillaries. (Figure reprinted with permission from Ref. )

Article Snippet: Fig. 5 Fabrication of a microvascular network by microfluidic systems.

Techniques: Imaging, Construct, Fluorescence, Microscopy, Generated, Cell Culture, Staining, In Vitro, Confocal Microscopy

Journal: Nano Convergence

Article Title: Technology for the formation of engineered microvascular network models and their biomedical applications

doi: 10.1186/s40580-024-00416-7

Figure Lengend Snippet: Techniques for microvascular network formation

Article Snippet: Fig. 5 Fabrication of a microvascular network by microfluidic systems.

Techniques: Control

Journal: Nano Convergence

Article Title: Technology for the formation of engineered microvascular network models and their biomedical applications

doi: 10.1186/s40580-024-00416-7

Figure Lengend Snippet: Properties of various biomaterials for microvascular network formation

Article Snippet: Fig. 5 Fabrication of a microvascular network by microfluidic systems.

Techniques: Activation Assay, Viscosity, Cell Culture, Binding Assay

Journal: Nano Convergence

Article Title: Technology for the formation of engineered microvascular network models and their biomedical applications

doi: 10.1186/s40580-024-00416-7

Figure Lengend Snippet: Advantages and disadvantages of cell types used for microvascular network formation

Article Snippet: Fig. 5 Fabrication of a microvascular network by microfluidic systems.

Techniques: Modification

Journal: Biomicrofluidics

Article Title: Vascularized liver-on-a-chip model to investigate nicotine-induced dysfunction

doi: 10.1063/5.0172677

Figure Lengend Snippet: Formation of microvascular networks. (a) General schematic of microvascular networks and timeline. (b) Day-by-day microvascular network formation of 10 × 10 6 HUVECs + 100 000 HDFs/mL. (c) Day 6 immunofluorescent max intensity images of (i) 2.5, (ii) 5, and (iii) 10 × 10 6 HUVECs/mL with 100:1 ratio of HDFs. (d) Perfusion of 1 μ m FluoSpheres into the microvascular networks showing perfusability. (e) Quantification of key microvascular parameters. Each region of interest covers 0.11 mm 2 .

Article Snippet: While these findings agree with the literature, they have not previously been shown in biomimetic microvascular networks.

Techniques:

Journal: Biomicrofluidics

Article Title: Vascularized liver-on-a-chip model to investigate nicotine-induced dysfunction

doi: 10.1063/5.0172677

Figure Lengend Snippet: Microvascular network response to nicotine. (a) Reactive oxygen species (ROS) generation within microvascular networks detected by H 2 DCFDA after 48 h of nicotine exposure. (b) Quantification of ROS through H 2 DCFDA MFI. (c) Immunofluorescent images of angiogenesis assay on day 7. (d) Quantification of the sprout length and number of sprouts from angiogenesis assay. (e) Microvascular network perfusion of 70 kDa FITC-dextran after 120 s, showing leakiness. (f) Normalized mean fluorescence intensity (MFI) of extravasated FITC-dextran over 120 s and fold change after 120 s.

Article Snippet: While these findings agree with the literature, they have not previously been shown in biomimetic microvascular networks.

Techniques: Angiogenesis Assay, Fluorescence

Journal: Biomicrofluidics

Article Title: Vascularized liver-on-a-chip model to investigate nicotine-induced dysfunction

doi: 10.1063/5.0172677

Figure Lengend Snippet: Vascularized liver-on-a-chip model provides functional outputs. (a) Secreted albumin from microvascular networks, non-vascularized, and vascularized liver-on-a-chip. (i) The normalized (to day 2) time course values are shown along with (ii) day 8 values. (b) Change in albumin values after 48 h exposure to 5 mM APAP when compared to the corresponding control. (c) Secreted urea from non-vascularized and vascularized liver-on-a-chip. (d) Fold change in transcriptional CYP2A6 and CYP3A4 of the liver-on-a-chip. (e) Functional CYP3A4 metabolism with non-vascularized and vascularized liver-on-a-chip when treated with 5 mM APAP and/or 10 μ M nicotine.

Article Snippet: While these findings agree with the literature, they have not previously been shown in biomimetic microvascular networks.

Techniques: Functional Assay, Control

Journal: Biomicrofluidics

Article Title: Vascularized liver-on-a-chip model to investigate nicotine-induced dysfunction

doi: 10.1063/5.0172677

Figure Lengend Snippet: Formation of microvascular networks. (a) General schematic of microvascular networks and timeline. (b) Day-by-day microvascular network formation of 10 × 10 6 HUVECs + 100 000 HDFs/mL. (c) Day 6 immunofluorescent max intensity images of (i) 2.5, (ii) 5, and (iii) 10 × 10 6 HUVECs/mL with 100:1 ratio of HDFs. (d) Perfusion of 1 μ m FluoSpheres into the microvascular networks showing perfusability. (e) Quantification of key microvascular parameters. Each region of interest covers 0.11 mm 2 .

Article Snippet: These findings indicate a dose-dependent ROS production with increasing concentrations of nicotine in biomimetic microvascular networks.

Techniques:

Journal: Biomicrofluidics

Article Title: Vascularized liver-on-a-chip model to investigate nicotine-induced dysfunction

doi: 10.1063/5.0172677

Figure Lengend Snippet: Microvascular network response to nicotine. (a) Reactive oxygen species (ROS) generation within microvascular networks detected by H 2 DCFDA after 48 h of nicotine exposure. (b) Quantification of ROS through H 2 DCFDA MFI. (c) Immunofluorescent images of angiogenesis assay on day 7. (d) Quantification of the sprout length and number of sprouts from angiogenesis assay. (e) Microvascular network perfusion of 70 kDa FITC-dextran after 120 s, showing leakiness. (f) Normalized mean fluorescence intensity (MFI) of extravasated FITC-dextran over 120 s and fold change after 120 s.

Article Snippet: These findings indicate a dose-dependent ROS production with increasing concentrations of nicotine in biomimetic microvascular networks.

Techniques: Angiogenesis Assay, Fluorescence

Journal: Biomicrofluidics

Article Title: Vascularized liver-on-a-chip model to investigate nicotine-induced dysfunction

doi: 10.1063/5.0172677

Figure Lengend Snippet: Vascularized liver-on-a-chip model provides functional outputs. (a) Secreted albumin from microvascular networks, non-vascularized, and vascularized liver-on-a-chip. (i) The normalized (to day 2) time course values are shown along with (ii) day 8 values. (b) Change in albumin values after 48 h exposure to 5 mM APAP when compared to the corresponding control. (c) Secreted urea from non-vascularized and vascularized liver-on-a-chip. (d) Fold change in transcriptional CYP2A6 and CYP3A4 of the liver-on-a-chip. (e) Functional CYP3A4 metabolism with non-vascularized and vascularized liver-on-a-chip when treated with 5 mM APAP and/or 10 μ M nicotine.

Article Snippet: These findings indicate a dose-dependent ROS production with increasing concentrations of nicotine in biomimetic microvascular networks.

Techniques: Functional Assay, Control