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Open Access Publications from the University of California

Scholarly Works (21 results)

  • Thesis
  • Peer Reviewed
UCLA Electronic Theses and Dissertations (2019)

Hemodynamic shear force is an important determinant of cardiovascular function and development. While mammalian models combined with next regeneration sequencing are essential for diagnosis and drug discovery, zebrafish has emerged as an important developmental model that combines in vivo analyses of cardiovascular phenotypes and the advantages of forward and reverse genomic engineering. The following studies in this thesis utilize an advanced imaging-based technique to characterize hemodynamic regulation in cardiovascular development and regeneration in embryonic zebrafish. Cardiogenesis involves a series of complex signaling pathways, while imaging cellular dynamics including cardiac trabeculation requires high spatiotemporal resolution. By using four-dimensional light-sheet fluorescent microscopy, we constructed four-dimensional moving domain models of the contracting myocardial wall to investigate hemodynamic regulation of endocardial Notch signaling in facilitating cardiac trabeculation. In vivo modulations of hematopoiesis and atrial contraction or ectopic expressions of Notch Intracellular Cytoplasmic Domain (NICD), suggested distinct flow patterns in myocardial geometry differentially activate endocardial Notch activity for trabecular organization and contractile function. Vascular disorders characterized by ischemic reperfusion injury, such as stroke, myocardial infarction, and peripheral vascular disease, remain the most frequent causes of incapacitating disease and death. Despite numerous efforts, structural and functional vessel recovery remains challenging, while molecular events underlying vascular injury and regeneration are largely unknown. A plethora of epidemiological studies consistently supports a link between redox active ultrafine particles (UFP, diameter < 0.1 um) in primary pollutants and cardiopulmonary disease. Due to the small size and the light weight of the particle, aspiration of UFP allows for penetration of pulmonary systems as well as the endothelial barrier. Once particles enter the circulatory system, they systematically affect endothelial homeostasis by promoting vascular oxidative and inflammatory responses. However, epigenetic and pathological effects underlying vascular regeneration upon particle exposure remains elusive. For the second part of this thesis, we investigated the importance of the Forkhead Box Sub-family O1 (FOXO1)/Notch activation complex upon UFP exposure in vascular regeneration. Finally, our vascular protective metabolomic profiles suggested hemodynamic shear forces is a central player in modulating the expression levels of glycolytic metabolites. In this context, we investigated the Vascular Endothelial Growth Factor Receptor (VEGFR)-Protein Kinase C isoform epsilon (PKCɛ) pathway and its role in promoting pro-glycolytic metabolites to help facilitate vascular regeneration.

    Cover page: Hemodynamics: From Developmental Mechano-transduction to Vascular Injury and Regeneration
    • Article
    • Peer Reviewed
    UCLA Previously Published Works (2024)
    How do vessels find optimal radii? Capillaries are known to adapt their radii to maintain the shear stress of blood flow at the vessel wall at a set point, yet models of adaptation purely based on average shear stress have not been able to produce complex loopy networks that resemble real microvascular systems. For narrow vessels where red blood cells travel in a single file, the shear stress on vessel endothelium peaks sharply when a red blood cell passes through. We show that stable shear-stress-based adaptation is possible if vessel shear stress set points are cued to the stress peaks. Model networks that respond to peak stresses alone can quantitatively reproduce the observed zebrafish trunk microcirculation, including its adaptive trajectory when hematocrit changes or parts of the network are amputated. Our work reveals the potential for mechanotransduction alone to generate stable hydraulically tuned microvascular networks.
      Cover page: Hemodynamic regulation allows stable growth of microvascular networks.
      • Article
      • Peer Reviewed
      UCLA Previously Published Works (2018)

      Aims

      Hemodynamic shear stress participates in maintaining vascular redox status. Elucidating flow-mediated endothelial metabolites enables us to discover metabolic biomarkers and therapeutic targets. We posited that flow-responsive vascular endothelial growth factor receptor (VEGFR)-protein kinase C isoform epsilon (PKCɛ)-6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) signaling modulates glycolytic metabolites for vascular repair.

      Results

      Bidirectional oscillatory flow (oscillatory shear stress [OSS]: 0.1 ± 3 dyne·cm-2 at 1 Hz) upregulated VEGFR-dependent PKCɛ expression to a greater degree than did unidirectional pulsatile flow (pulsatile shear stress [PSS]: 23 ± 8 dyne·cm-2 at 1 Hz) in human aortic endothelial cells (p < 0.05, n = 3). PSS and OSS further upregulated PKCɛ-dependent PFKFB3 expression for glycolysis (p < 0.05, n = 4). Constitutively active PKCɛ increased, whereas dominant-negative PKCɛ reduced both basal and maximal extracellular acidification rates for glycolytic flux (p < 0.01, n = 4). Metabolomic analysis demonstrated an increase in PKCɛ-dependent glycolytic metabolite, dihydroxyacetone (DHA), but a decrease in gluconeogenic metabolite, aspartic acid (p < 0.05 vs. control, n = 6). In a New Zealand White rabbit model, both PKCɛ and PFKFB3 immunostaining was prominent in the PSS- and OSS-exposed aortic arch and descending aorta. In a transgenic Tg(flk-1:EGFP) zebrafish model, GATA-1a morpholino oligonucleotide injection (to reduce viscosity-dependent shear stress) impaired vascular regeneration after tail amputation (p < 0.01, n = 20), which was restored with PKCɛ messenger RNA (mRNA) rescue (p < 0.05, n = 5). As a corollary, siPKCɛ inhibited tube formation and vascular repair, which were restored by DHA treatment in our Matrigel and zebrafish models. Innovation and Conclusion: Flow-sensitive VEGFR-PKCɛ-PFKFB3 signaling increases the glycolytic metabolite, dihydroxyacetone, to promote vascular repair. Antioxid. Redox Signal. 28, 31-43.
        Cover page: Flow-Responsive Vascular Endothelial Growth Factor Receptor-Protein Kinase C Isoform Epsilon Signaling Mediates Glycolytic Metabolites for Vascular Repair.
        • Article
        • Peer Reviewed
        UCLA Previously Published Works (2017)
        In animals, gas exchange between blood and tissues occurs in narrow vessels, whose diameter is comparable to that of a red blood cell. Red blood cells must deform to squeeze through these narrow vessels, transiently blocking or occluding the vessels they pass through. Although the dynamics of vessel occlusion have been studied extensively, it remains an open question why microvessels need to be so narrow. We study occlusive dynamics within a model microvascular network: the embryonic zebrafish trunk. We show that pressure feedbacks created when red blood cells enter the finest vessels of the trunk act together to uniformly partition red blood cells through the microvasculature. Using mathematical models as well as direct observation, we show that these occlusive feedbacks are tuned throughout the trunk network to prevent the vessels closest to the heart from short-circuiting the network. Thus occlusion is linked with another open question of microvascular function: how are red blood cells delivered at the same rate to each micro-vessel? Our analysis shows that tuning of occlusive feedbacks increase the total dissipation within the network by a factor of 11, showing that uniformity of flows rather than minimization of transport costs may be prioritized by the microvascular network.
          Cover page: Optimal occlusion uniformly partitions red blood cells fluxes within a microvascular network
          • Article
          • Peer Reviewed
          UCLA Previously Published Works (2018)

          Purpose of review

          Real-time 3-dimensional (3-D) imaging of cardiovascular injury and regeneration remains challenging. We introduced a multi-scale imaging strategy that uses light-sheet illumination to enable applications of cardiovascular injury and repair in models ranging from zebrafish to rodent hearts.

          Recent findings

          Light-sheet imaging enables rapid data acquisition with high spatiotemporal resolution and with minimal photo-bleaching or photo-toxicity. We demonstrated the capacity of this novel light-sheet approach for scanning a region of interest with specific fluorescence contrast, thereby providing axial and temporal resolution at the cellular level without stitching image columns or pivoting illumination beams during one-time imaging. This cutting-edge imaging technique allows for elucidating the differentiation of stem cells in cardiac regeneration, providing an entry point to discover novel micro-circulation phenomenon with clinical significance for injury and repair. These findings demonstrate the multi-scale applications of this novel light-sheet imaging strategy to advance research in cardiovascular development and regeneration.
            Cover page: Light-Sheet Imaging to Elucidate Cardiovascular Injury and Repair
            • Article
            • Peer Reviewed
            UCLA Previously Published Works (2022)
            Wall shear stress (WSS) contributes to the mechanotransduction underlying microvascular development and regeneration. Using computational fluid dynamics, we elucidated the interplay between WSS and vascular remodelling in a zebrafish model of tail amputation and regeneration. The transgenic Tg (fli1:eGFP; Gata1:ds-red) zebrafish line was used to track the three-dimensional fluorescently labelled vascular endothelium for post-image segmentation and reconstruction of the fluid domain. Particle image velocimetry was used to validate the blood flow. Following amputation to the dorsal aorta and posterior cardinal vein (PCV), vasoconstriction developed in the dorsal longitudinal anastomotic vessel (DLAV) along with increased WSS in the proximal segmental vessels (SVs) from amputation. Angiogenesis ensued at the tips of the amputated DLAV and PCV where WSS was minimal. At 2 days post amputation (dpa), vasodilation occurred in a pair of SVs proximal to amputation, followed by increased blood flow and WSS; however, in the SVs distal to amputation, WSS normalized to the baseline. At 3 dpa, the blood flow increased in the arterial SV proximal to amputation and through anastomosis with DLAV formed a loop with PCV. Thus, our in silico modelling revealed the interplay between WSS and microvascular adaptation to changes in WSS and blood flow to restore microcirculation following tail amputation.
              Cover page: Computational simulations of the 4D micro-circulatory network in zebrafish tail amputation and regeneration
              • Article
              • Peer Reviewed
              UCLA Previously Published Works (2017)
              Blood viscosity provides the rheological basis to elucidate shear stress underlying developmental cardiac mechanics and physiology. Zebrafish is a high throughput model for developmental biology, forward-genetics, and drug discovery. The micro-scale posed an experimental challenge to measure blood viscosity. To address this challenge, a microfluidic viscometer driven by surface tension was developed to reduce the sample volume required (3μL) for rapid (<2 min) and continuous viscosity measurement. By fitting the power-law fluid model to the travel distance of blood through the micro-channel as a function of time and channel configuration, the experimentally acquired blood viscosity was compared with a vacuum-driven capillary viscometer at high shear rates (>500 s-1), at which the power law exponent (n) of zebrafish blood was nearly 1 behaving as a Newtonian fluid. The measured values of whole blood from the micro-channel (4.17cP) and the vacuum method (4.22cP) at 500 s-1 were closely correlated at 27 °C. A calibration curve was established for viscosity as a function of hematocrits to predict a rise and fall in viscosity during embryonic development. Thus, our rapid capillary pressure-driven micro-channel revealed the Newtonian fluid behavior of zebrafish blood at high shear rates and the dynamic viscosity during development.
                Cover page: A Rapid Capillary-Pressure Driven Micro-Channel to Demonstrate Newtonian Fluid Behavior of Zebrafish Blood at High Shear Rates.
                • Article
                • Peer Reviewed
                UCLA Previously Published Works (2018)
                The advent of 4-dimensional (4D) light-sheet fluorescence microscopy (LSFM) has provided an entry point for rapid image acquisition to uncover real-time cardiovascular structure and function with high axial resolution and minimal photo-bleaching/-toxicity. We hereby review the fundamental principles of our LSFM system to investigate cardiovascular morphogenesis and regeneration after injury. LSFM enables us to reveal the micro-circulation of blood cells in the zebrafish embryo and assess cardiac ventricular remodeling in response to chemotherapy-induced injury using an automated segmentation approach. Next, we review two distinct mechanisms underlying zebrafish vascular regeneration following tail amputation. We elucidate the role of endothelial Notch signaling to restore vascular regeneration after exposure to the redox active ultrafine particles (UFP) in air pollutants. By manipulating the blood viscosity and subsequently, endothelial wall shear stress, we demonstrate the mechanism whereby hemodynamic shear forces impart both mechanical and metabolic effects to modulate vascular regeneration. Overall, the implementation of 4D LSFM allows for the elucidation of mechanisms governing cardiovascular injury and regeneration with high spatiotemporal resolution.
                  Cover page: Advanced microscopy to elucidate cardiovascular injury and regeneration: 4D light-sheet imaging.
                  • Article
                  • Peer Reviewed
                  UC San Diego Previously Published Works (2014)
                  Prognosis of patients with lung metastases of soft-tissue sarcoma is still poor. Therefore, novel systemic therapy is needed to improve the survival of soft-tissue sarcoma. In the present study, tumor-targeting therapy with a genetically-modified auxotrophic strain of Salmonella typhimurium, termed A1-R, was evaluated. Mouse models of primary soft tissue sarcoma and spontaneous lung metastasis were obtained by orthotopic intra-muscular injection of HT1080-RFP human fibrosarcoma cells. S. typhimurium A1-R was administered from day 14, once a week for two weeks. On day 28, lung samples were excised and observed with a fluorescence imaging system. The number of lung metastasis was 8.8 ± 3.4 in the untreated group and 0.8 ± 0.8 in the treated group (P = 0.024). A mouse model of experimental lung metastasis was obtained by tail vein injection of HT1080-RFP cells. The mice were treated with S. typhimurium A1-R (i.v.) on day 7, once a week for three weeks. S. typhimurium A1-R significantly reduced lung metastases and improved overall survival (P = 0.004). S. typhimurium A1-R bacterial therapy has future potential for treating advanced soft tissue sarcoma and improving prognosis of patients with lung metastasis.
                    Cover page: Inhibition of spontaneous and experimental lung metastasis of soft-tissue sarcoma by tumor-targeting Salmonella typhimurium A1-R
                    • Article
                    • Peer Reviewed
                    UCLA Previously Published Works (2017)
                    Currently, there is a limited ability to interactively study developmental cardiac mechanics and physiology. We therefore combined light-sheet fluorescence microscopy (LSFM) with virtual reality (VR) to provide a hybrid platform for 3D architecture and time-dependent cardiac contractile function characterization. By taking advantage of the rapid acquisition, high axial resolution, low phototoxicity, and high fidelity in 3D and 4D (3D spatial + 1D time or spectra), this VR-LSFM hybrid methodology enables interactive visualization and quantification otherwise not available by conventional methods, such as routine optical microscopes. We hereby demonstrate multiscale applicability of VR-LSFM to (a) interrogate skin fibroblasts interacting with a hyaluronic acid-based hydrogel, (b) navigate through the endocardial trabecular network during zebrafish development, and (c) localize gene therapy-mediated potassium channel expression in adult murine hearts. We further combined our batch intensity normalized segmentation algorithm with deformable image registration to interface a VR environment with imaging computation for the analysis of cardiac contraction. Thus, the VR-LSFM hybrid platform demonstrates an efficient and robust framework for creating a user-directed microenvironment in which we uncovered developmental cardiac mechanics and physiology with high spatiotemporal resolution.
                      Cover page: Integrating light-sheet imaging with virtual reality to recapitulate developmental cardiac mechanics.
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