Ischemia leading to heart attacks and strokes is the major cause of deaths in the world. Whether an occlusion occurs or not, depends on the ability of a growing thrombus to resist forces exerted on its structure. This manuscript provides the first known in vivo measurement of the stresses that clots can withstand, before yielding to the surrounding blood flow. Namely, Lattice-Boltzmann Method flow simulations are performed based on 3D clot geometries. The latter are estimated from intravital microscopy images of laser-induced injuries in cremaster microvasculature of live mice. In addition to reporting the blood clot yield stresses, we also show that the thrombus 'core' does not experience significant deformation, while its 'shell' does. This indicates that the latter is more prone to embolization. Hence, drugs should be designed to target the shell selectively, while leaving the core intact (to minimize excessive bleeding). Finally, we laid down a foundation for a nondimensionalization procedure, which unraveled a relationship between clot mechanics and biology. Hence, the proposed framework could ultimately lead to a unified theory of thrombogenesis, capable of explaining all clotting events. Thus, the findings presented herein will be beneficial to the understanding and treatment of heart attacks, strokes and hemophilia.
Achieving hemostasis following penetrating injuries is essential for the survival of organisms that possess a closed high-pressure circulatory system. However, pathological manifestation of thrombosis and embolism can potentially lead to life-threatening complications when occurring in the heart (i.e., a heart attack), brain (i.e., a stroke), or lungs (i.e., DVT/PE). Among these, thromboembolic infarction is the leading cause of mortality and morbidity in the United States, while stroke is the 5 th . [1] Conversely, deficiencies in the clotting mechanisms (hemophilia or due to drug interactions) can result in bleeding risks that confront surgeons on a regular basis. Yet, despite tremendous efforts by the medical research community (e.g., ~$3 billion of annual expenditure on heart attack and brain stroke research alone [2]), the problem that essentially amounts to a clogged "pipe" remains largely unsolved to this day. Moreover, what makes one thrombus benign, while another one dangerous, is also not well understood. Therefore, gaining insight into the thrombi's tendency to occlude blood vessels would be beneficial for public health, since it could pave the way towards a better understanding of the risk factors involved; and subsequently to better disease treatments and thromboectomy devices. [3]
Whether an occlusion occurs or not, depends on the ability of the growing thrombus to resist the blood flow forces exerted on its structure. With development of advanced intravital microscopy experiments, the thrombi structures have been shown to be heterogeneous: consisting of a densely packed “core” nearest the injury site, and a loose “shell” overlaying the core (see Figure 1). [4,5]. It is also reported that the core is composed of highly “activated” platelets (as is measured by P-selectin expression), while the shell consists of loosely-packed P-selectin -negative cells. The biological purpose, as well as the cause of this heterogeneity, are unknown. One thing that is apparent, however, is that the core and the shell contribute differently to key parts of the thrombus formation and hemostasis: The shell is observed to shed the most mass (leading to the conclusion that embolism is mostly caused by this part of the clot); while the core can be seen to anchor the thrombus to the injury, and stop the escape of blood to the extravascular space by “sealing” the damage. This leads to an important conclusion that there are potentially significant material and functional differences between these two regions of blood clots.
Figure 1 LEFT -“Core-and-Shell” model schematic showing that the clot is comprised of two regions differing in degree of platelet activation and packing density; Image reprinted with permission, from Ref. [6] RIGHT -Confocal fluorescent microscopy image of a clot (Blue = Pselectin exposure marking the activated core; Red = anti-CD41 platelet marker). Scale bar is 10µm.
At the same time, the viscoplastic behavior exhibited by the thrombi resembles that of a Bingham plastic -a material, like toothpaste, that behaves as a rigid body at low stresses, but flows as a viscous fluid when its critical yield stress c is exceeded (see Equations 1 -2). Namely, like a Bingham plastic, the clot consists of discrete particles (in this case platelets) trapped in a liquid gel. The platelets interact with each other creating a weak solid structure known as a “false body”. A certain amount of stress corresponding to c is required to break this structure and allow the platelets to rearrange within the gel under viscous forces. After the stress is reduced however, the platelets associate again, solidifying the structure. Figure 2 illustrates the Bingham plastic-like behavior of a blood clot as observed from intravital microscopy. The figure shows three major progression stages of a typical thrombus formation: 1) initial platelet attachment to the injury site (see Figure 2-A); 2) clot growth radially outward from the injury site (see Figure 2-B); and 3) steady state stability (see Figure 2-C). The transition from the second to the third stage (Figure 2, panels BC) is the Bingham-like “yielding” of the thrombus. During this critical event, platelet mass from the upstream portion of the thrombus is forced to its downstream side. As a result, the thrombus changes shape from a “ball”-like structure, to the characteristic “comet-tail” shape typically seen in blood clots. This transition occurs because the obstructing thrombus experiences stronger forces from the surrounding fluid, which is trying to squeeze through the little remaining openings left in the lumen. Consequently, if there is no full occlusion of the blood vessel, the thrombus’ structure eventually yields to the flow forces, and rearranges its shape to minimize the fluid drag imposed on its surface.
Interestingly, despite the underlying complexity of the thrombo-genesis mechanism, the discrete regimes shown in Figure 2 appear to be common to all thrombi. The
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