研究目的
To measure in vivo the mechanical strength of blood clots, specifically the yield stress, using a hybrid approach combining intravital microscopy and computational fluid dynamics to understand thrombogenesis and its implications for heart attacks, strokes, and hemophilia.
研究成果
The study provides the first in vivo measurement of blood clot yield stress, revealing that the thrombus shell deforms under higher fluid-induced stresses while the core remains intact, indicating the shell is more prone to embolization. This suggests targeted drug therapies could dissolve the shell selectively to reduce bleeding risks. The nondimensionalization approach offers a foundation for a unified theory of thrombogenesis, with potential benefits for understanding and treating cardiovascular diseases.
研究不足
1. 3D clot shapes are extrapolated from 2D cross-sections assuming parabolic profiles, which may not be accurate for early stages. 2. Pseudo steady-state assumption in simulations may not fully capture transient effects. 3. Thrombi are assumed impermeable, though reality may involve porosity. 4. Blood escape from injury sites and vessel deformations are not modeled. 5. Blood is treated as Newtonian, though it has non-Newtonian properties at low shear rates.
1：Experimental Design and Method Selection:
A semi-empirical approach is used, combining in vivo imaging with Lattice-Boltzmann Method (LBM) simulations. The design involves estimating 3D clot geometries from 2D intravital microscopy images and simulating fluid flow to calculate shear stresses.
2：Sample Selection and Data Sources:
Laser-induced injuries in the cremaster microvasculature of live male C57Bl/6J mice (8-12 weeks old) are used. Data includes 2D confocal microscopy images with a time interval of 0.619 seconds and spatial resolution of 0.22 μm/pixel, from 10 different experiments.
3：619 seconds and spatial resolution of 22 μm/pixel, from 10 different experiments. List of Experimental Equipment and Materials:
3. List of Experimental Equipment and Materials: Confocal microscope, SlideBook 5 software, Fiji plug-ins, Matlab code, in-house LBM code, Tecplot 360 EX 2017 for visualization. Materials include Alexa-Fluor? monoclonal antibody labeling kits, anti-CD41 F(ab)2 fragments, anti-P-selectin antibodies, caged fluorescein conjugated albumin.
4：Experimental Procedures and Operational Workflow:
Induce thrombus formation via laser injury, acquire 2D images, deblur and stabilize images, segment 2D shapes, estimate 3D geometries using parabolic assumptions, perform LBM simulations with D3Q15 lattice and bounce-back boundary conditions, calculate shear stresses using finite difference approximations.
5：Data Analysis Methods:
Nondimensionalization of time and stress data, moving averages and variances for trend analysis, validation against COMSOL? simulations, statistical analysis of stress and morphology changes.
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confocal microscope
Used for acquiring 2D intravital microscopy images of thrombus formation in live mice.
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SlideBook 5 software
5
Intelligent Imaging Innovations
Software for image acquisition in confocal microscopy.
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Fiji plug-ins
Fiji
Used for image stabilization and processing, including StackReg and Image Stabilizer.
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Matlab code
MathWorks Inc.
Custom code for image enhancement, segmentation, and 3D geometry estimation.
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LBM code
D3Q15 lattice
in-house
In-house code for Lattice-Boltzmann Method simulations of fluid flow around thrombi.
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Tecplot 360 EX
2017
Tecplot Inc.
Software for 3D visualization of simulation results.
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COMSOL
COMSOL
Commercial computational fluid dynamics package used for validation against idealized models.
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Alexa-Fluor monoclonal antibody labeling kits
Invitrogen
Used to label platelet-specific antibodies for visualization in microscopy.
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anti-CD41 F(ab)2 fragments
clone MWReg30
BD Biosciences
Antibody for visualizing platelet surfaces in microscopy.
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anti-P-selectin
clone RB40.34
BD Bioscience
Antibody for specific labeling of activated platelets.
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caged fluorescein conjugated albumin
Used for visualizing the lumen and laser injury site in microscopy.
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