Sandy Chan, Morgan Brophy, Nozomi Nishimura, Chris B. Schaffer
PLoS ONE (2019)
Microhemorrhages are common in the aging brain and are thought to contribute to cognitive decline and the development of neurodegenerative diseases, such as Alzheimer’s disease. Chronic aspirin therapy is widespread in older individuals and decreases the risk of coronary artery occlusions and stroke. There remains a concern that such aspirin usage may prolong bleeding after a vessel rupture in the brain, leading to larger bleeds that cause more damage to the surrounding tissue. Here, we aimed to understand the influence of aspirin usage on the size of cortical microhemorrhages and explored the impact of age. We used femtosecond laser ablation to rupture arterioles in the cortex of both young (2–5 months old) and aged (18–29 months old) mice dosed on aspirin in their drinking water and measured the extent of penetration of both red blood cells and blood plasma into the surrounding tissue. We found no difference in microhemorrhage size for both young and aged mice dosed on aspirin, as compared to controls (hematoma diameter = 104 +/- 39 (97 +/- 38) μm in controls and 109 +/- 25 (101 +/- 28) μm in aspirin-treated young (aged) mice; mean +/- SD). In contrast, young mice treated with intravenous heparin had an increased hematoma diameter of 136 +/- 44 μm. These data suggest that aspirin does not increase the size of microhemorrhages, supporting the safety of aspirin usage.
Amy F. Smith, Vincent Doyeux, Maxime Berg, Myriam Peyrounette, Mohammad Haft-Javaherian, Anne-Edith Larue, John H. Slater, Frédéric Lauwer, Pablo Blinder, Philbert Tsai, David Kleinfeld, Chris B. Schaffer, Nozomi Nishimura, Yohan Davit and Sylvie Lorthois
Frontiers in physiology (2019)
Despite the key role of the capillaries in neurovascular function, a thorough characterization of cerebral capillary network properties is currently lacking. Here, we define a range of metrics (geometrical, topological, flow, mass transfer, and robustness) for quantification of structural differences between brain areas, organs, species, or patient populations and, in parallel, digitally generate synthetic networks that replicate the key organizational features of anatomical networks (isotropy, connectedness, space-filling nature, convexity of tissue domains, characteristic size). To reach these objectives, we first construct a database of the defined metrics for healthy capillary networks obtained from imaging of mouse and human brains. Results show that anatomical networks are topologically equivalent between the two species and that geometrical metrics only differ in scaling. Based on these results, we then devise a method which employs constrained Voronoi diagrams to generate 3D model synthetic cerebral capillary networks that are locally randomized but homogeneous at the network-scale. With appropriate choice of scaling, these networks have equivalent properties to the anatomical data, demonstrated by comparison of the defined metrics. The ability to synthetically replicate cerebral capillary networks opens a broad range of applications, ranging from systematic computational studies of structure-function relationships in healthy capillary networks to detailed analysis of pathological structural degeneration, or even to the development of templates for fabrication of 3D biomimetic vascular networks embedded in tissue-engineered constructs.
Mohammad Haft-Javaherian, Linjing Fang, Victorine Muse, Chris B. Schaffer, Nozomi Nishimura, Mert R. Sabuncu
PLoS ONE (2019)
The health and function of tissue rely on its vasculature network to provide reliable blood perfusion. Volumetric imaging approaches, such as multiphoton microscopy, are able to generate detailed 3D images of blood vessels that could contribute to our understanding of the role of vascular structure in normal physiology and in disease mechanisms. The segmentation of vessels, a core image analysis problem, is a bottleneck that has prevented the systematic comparison of 3D vascular architecture across experimental populations. We explored the use of convolutional neural networks to segment 3D vessels within volumetric in vivo images acquired by multiphoton microscopy. We evaluated different network architectures and machine learning techniques in the context of this segmentation problem. We show that our optimized convolutional neural network architecture with a customized loss function, which we call DeepVess, yielded a segmentation accuracy that was better than state-of-the-art methods, while also being orders of magnitude faster than the manual annotation. To explore the effects of aging and Alzheimer’s disease on capillaries, we applied DeepVess to 3D images of cortical blood vessels in young and old mouse models of Alzheimer’s disease and wild type littermates. We found little difference in the distribution of capillary diameter or tortuosity between these groups, but did note a decrease in the number of longer capillary segments (>75μm) in aged animals as compared to young, in both wild type and Alzheimer’s disease mouse models.
Michael G. McCoy, Dennis Nyanyo, Carol K. Hung, Julian Palacios Goerger, Warren R. Zipfel, Rebecca M. Williams, Nozomi Nishimura & Claudia Fischbach
Scientific Reports (2019)
Rapid growth and perivascular invasion are hallmarks of glioblastoma (GBM) that have been attributed to the presence of cancer stem-like cells (CSCs) and their association with the perivascular niche. However, the mechanisms by which the perivascular niche regulates GBM invasion and CSCs remain poorly understood due in part to a lack of relevant model systems. To simulate perivascular niche conditions and analyze consequential changes of GBM growth and invasion, patient-derived GBM spheroids were co-cultured with brain endothelial cells (ECs) in microfabricated collagen gels. Integrating these systems with 3D imaging and biochemical assays revealed that ECs increase GBM invasiveness and growth through interleukin-8 (IL-8)-mediated enrichment of CSCs. Blockade of IL-8 inhibited these effects in GBM-EC co-cultures, while IL-8 supplementation increased CSC-mediated growth and invasion in GBM-monocultures. Experiments in mice confirmed that ECs and IL-8 stimulate intracranial tumor growth and invasion in vivo. Collectively, perivascular niche conditions promote GBM growth and invasion by increasing CSC frequency, and IL-8 may be explored clinically to inhibit these interactions.
Jean C. Cruz Hernández, Oliver Bracko, Calvin J. Kersbergen , Victorine Muse , Mohammad Haft-Javaherian, Maxime Berg, Laibaik Park , Lindsay K. Vinarcsik, Iryna Ivasyk, Daniel A. Rivera, Yiming Kang, Marta Cortes-Canteli, Myriam Peyrounette, Vincent Doyeux, Amy Smith , Joan Zhou, Gabriel Otte, Jeffrey D. Beverly, Elizabeth Davenport, Yohan Davit, Charles P. Lin, Sidney Strickland, Costantino Iadecola , Sylvie Lorthois , Nozomi Nishimura , Chris B. Schaffer
Nature Neuroscience (2019)
Cerebral blood flow (CBF) reductions in Alzheimer’s disease patients and related mouse models have been recognized for decades, but the underlying mechanisms and resulting consequences for Alzheimer’s disease pathogenesis remain poorly understood. In APP/PS1 and 5xFAD mice we found that an increased number of cortical capillaries had stalled blood flow as compared to in wild-type animals, largely due to neutrophils that had adhered in capillary segments and blocked blood flow. Administration of antibodies against the neutrophil marker Ly6G reduced the number of stalled capillaries, leading to both an immediate increase in CBF and rapidly improved performance in spatial and working memory tasks. This study identified a previously uncharacterized cellular mechanism that explains the majority of the CBF reduction seen in two mouse models of Alzheimer’s disease and demonstrated that improving CBF rapidly enhanced short-term memory function. Restoring cerebral perfusion by preventing neutrophil adhesion may provide a strategy for improving cognition in Alzheimer’s disease patients
Tejapratap Bollu, Nathan R Cornelius, John Sunwoo, Nozomi Nishimura, Chris B Schaffer and Peter C Doerschuk
Journal of Cerebral Blood Flow & Metabolism (2017)
Computations are described which estimate flows in all branches of the cortical surface arteriole network from two-photon excited fluorescence (2PEF) microscopy images which provide the network topology and, in selected branches red blood cell (RBC) speeds and lumen diameters. Validation is done by comparing the flow predicted by the model with experimentally measured flows and by comparing the predicted flow redistribution in the network due to single-vessel strokes with experimental observations. The model predicts that tissue is protected from RBC flow decreases caused by multiple occlusions of surface arterioles but not penetrating arterioles. The model can also be used to study flow rerouting due to vessel dilations and constrictions.
M. Koyama, F. Minale, J. Shum, N. Nishimura, C. B. Schaffer, and J. R. Fetcho
Animals collect sensory information from the world and make adaptive choices about how to respond to it. Here, we reveal a network motif in the brain for one of the most fundamental behavioral choices made by bilaterally symmetric animals: whether to respond to a sensory stimulus by moving to the left or to the right. We define network connectivity in the hindbrain important for the lateralized escape behavior of zebrafish and then test the role of neurons by using laser ablations and behavioral studies. Key inhibitory neurons in the circuit lie in a column of morphologically similar cells that is one of a series of such columns that form a developmental and functional ground plan for building hindbrain networks. Repetition within the columns of the network motif we defined may therefore lie at the foundation of other lateralized behavioral choices.
M. J. Farrar, J. D. Rubin, D. M. Diago, and C. B. Schaffer
Journal of Cerebral Blood flow and Metabolism (2015)
The availability of transgenic strains has made the laboratory mouse a popular model for the study of healthy and diseased state spinal cord (SC). Essential to identifying physiologic and pathologic events is an understanding of the microvascular network and flow patterns of the SC. Using 2-photon excited fluorescence (2PEF) microscopy we performed in vivo measurements of blood flow in the lower thoracic portion of the mouse dorsal spinal vein (dSV) and in the first upstream branches supplying it, denoted as dorsal ascending venules (dAVs). We found that the dSV had large radiculomedullary veins (RMVs) exiting the SC, and that flow in the dSV between pairs of RMVs was bidirectional. Volumetric flow increased in each direction away from the point of bifurcation. Flow in the upstream dAVs varied with diameter in a manner consistent with a constant distal pressure source. By performing ex vivo 2PEF microscopy of fluorescent-gel perfused tissue, we created a 3-D map of the dorsal spinal vasculature. From these data, we constructed a simple model that predicted changes in the flow of upstream branches after occlusion of the dSV in different locations. Using an atraumatic model of dSV occlusion, we confirmed the predictions of this model in vivo.
Lindsey SE, Menon PG, Kowalski WJ, Shekhar A, Yalcin HC, Nishimura N, Schaffer CB, Butcher JT, Pekkan K.
Biomech Model Mechanobiol (2015)
The majority of severe clinically significant forms of congenital heart disease (CHD) are associated with great artery lesions, including hypoplastic, double, right or interrupted aortic arch morphologies. While fetal and neonatal interventions are advancing, their potential ability to restore cardiac function, optimal timing, location, and intensity required for intervention remain largely unknown. Here, we combine computational fluid dynamics (CFD) simulations with in vivo experiments to test how individual pharyngeal arch artery hemodynamics alter as a result of local interventions obstructing individual arch artery flow. Simulated isolated occlusions within each pharyngeal arch artery were created with image-derived three-dimensional (3D) reconstructions of normal chick pharyngeal arch anatomy at Hamburger-Hamilton (HH) developmental stages HH18 and HH24. Acute flow redistributions were then computed using in vivo measured subject-specific aortic sinus inflow velocity profiles. A kinematic vascular growth-rendering algorithm was then developed and implemented to test the role of changing local wall shear stress patterns in downstream 3D morphogenesis of arch arteries. CFD simulations predicted that altered pressure gradients and flow redistributions were most sensitive to occlusion of the IVth arches. To evaluate these simulations experimentally, a novel in vivo experimental model of pharyngeal arch occlusion was developed and implemented using two-photon microscopy-guided femtosecond laser-based photodisruption surgery. The right IVth arch was occluded at HH18, and resulting diameter changes were followed for up to 24 h. Pharyngeal arch diameter responses to acute hemodynamic changes were predicted qualitatively but poorly quantitatively. Chronic growth and adaptation to hemodynamic changes, however, were predicted in a subset of arches. Our findings suggest that this complex biodynamic process is governed through more complex forms of mechanobiological vascular growth rules. Other factors in addition to wall shear stress or more complex WSS rules are likely important in the long-term arterial growth and patterning. Combination in silico/experimental platforms are essential for accelerating our understanding and prediction of consequences from embryonic/fetal cardiovascular occlusions and lay the foundation for noninvasive methods to guide CHD diagnosis and fetal intervention.
A.Y. Shih, C. Ruhlmann, P. Blinder, A. Devor, P. J. Drew, B. Friedman, P. M. Knutsen, P. D. Lyden, C. Mateo, L. Mellander, N. Nishimura, C. B. Schaffer, P. S. Tsai, and D. Kleinfeld
We review the organizational principles of the cortical vasculature and the underlying patterns of blood flow under normal conditions and in response to occlusion of single vessels. The cortex is sourced by a two-dimensional network of pial arterioles that feeds a three-dimensional network of subsurface microvessels in close proximity to neurons and glia. Blood flow within the surface and subsurface networks is largely insensitive to occlusion of a single vessel within either network. However, the penetrating arterioles that connect the pial network to the subsurface network are bottlenecks to flow; occlusion of even a single penetrating arteriole results in the death of a 500 μm diameter cylinder of cortical tissue despite the potential for collateral flow through microvessels. This pattern of flow is consistent with that calculated from a full reconstruction of the angioarchitecture. Conceptually, collateral flow is insufficient to compensate for the occlusion of a penetrating arteriole because penetrating venules act as shunts of blood that flows through collaterals. Future directions that stem from the analysis of the angioarchitecture concern cellular-level issues, in particular the regulation of blood flow within the subsurface microvascular network, and system-level issues, in particular the role of penetrating arteriole occlusions in human cognitive impairment.