Healing of Human Intestinal Organoids

Sunday, February 28, 2016 853-Pos Board B633 Quantifying Nanoscale Properties of Engineered Virus Capsids for Malaria Vaccines Albert J. Jin1, David Mertz1,2, Hsien-Shun Liao1, Aanchal Johri1, Luis Torres1, Yimin Wu3, David Narum3. 1 National Institute of Biomedical Imaging and Bioengineering/NIH, Bethesda, MD, USA, 2Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA, 3National Institute of Allergy and Infectious Diseases/NIH, Bethesda, MD, USA. An important challenge in targeted drug delivery and vaccine development is to create the most effective interface between the carrier and target cells. Parasitic diseases, for which no approved vaccines are available, remain a world health problem with over one million severe cases per year attributed to malaria alone. In parallel to developing potential malaria antigen constructs (1), we have also bioengineered and characterized nanoscale viruslike particles (VLPs) with tools such as atomic force microscopy (AFM) and quantitative nanomechanical mapping (QNM) in hopes that these might serve as possible carriers for drugs or vaccines because of how they uniquely interact with human cells. Through using AFM and QNM techniques we have revealed some morphological features of our own bioengineered VLP, a protein capsid of bacteriophage Qb (d = 25 nm), and analyzed deformation of the capsid under compressive force to quantify its elastic properties. We have found that these particles exhibit a linear relationship between compressive force and deformation as would a Hookean spring. Significant deformation (> 5 nm) was observed at forces of 0.1 - 0.5 nN. Our calculated spring constant for our Qb capsid, 0.059 N/m, indicates that our VLP is more pliable than others of similar protein structure, which is possibly a favorable characteristic for the capsid to bind with the cell membrane in an in vivo carrier scenario. We are also using this analytical technique for the characterization of chemically conjugated (malaria) vaccines. Ref (1) ‘‘Development of a Pfs25-EPA malaria transmission blocking vaccine as a chemically conjugated nanoparticle.’’ Shimp RL Jr, Rowe C, Reiter K, Chen B, Nguyen V, Aebig J, Rausch KM, Kumar K, Wu Y, Jin AJ, Jones DS, Narum DL. Vaccine. 2013. 31:2954-62. doi: 10.1016/ j.vaccine.2013.04.034. 854-Pos Board B634 Bio-AFM of Cancer Cells and Multifunctional Theranostics Xiao Fu1,2, Zhe Wang1, Ashwin Bhirde1,3, Jenny Zhu1, Hsien-Shun Liao1, Nicole Carvajal1, Gang Niu1, Henry Eden1, Xiaoyuan Chen1, Albert J. Jin1. 1 National Institute of Biomedical Imaging and Bioengineering/NIH, Bethesda, MD, USA, 2Beijing Institute of Technology, Beijing, China, 3Food and Drug Administration, Silver Spring, MD, USA. Biological atomic force microscopy (Bio-AFM) can quantify changes in cellular and macromolecular morphology and elasticity at the nanoscale and over a broad time scale, from fast dynamics to long-term cellular evolutions. Herein, we report quantitative Bio-AFM characterization of mechanobiological features of lung cancer and other cells in response to chemotherapeutics and multifunctional theranostic nanoparticles. Multidrug resistance (MDR) presents a daunting challenge for successful cancer management. Comprehensive investigation of mechanobiological characters of cancer cells with MDR provides insights in directing anti-cancer therapy. In our study, we have embarked on using multimodal and multiphasic Bio-AFM to identify cellular biomechanical markers and to investigate interactions among cells and theranostic nanoparticles as an aid to developing more efficient therapeutics. We have found a spectrum of distinct biomechanical signatures, such as stiffness moduli and adhesions that differ under physiological conditions between drug sensitive and resistant cancer cells. Multifunctional theranostic nanoparticles, such as polymeric nanovehicles (1), single-walled carbon nanotubes (SWCNTs), and carbon dots (Cdots) with various surface coatings are observed to have differential influences on cellular responses. As part of work to broaden the applicability of Bio-AFM for cellular imaging, we are investigating the development of docetaxel-sensitivity and resistance in a non-small-cell lung cancer cell line. Ref (1) ‘‘Polymeric Nanovehicle Regulated Spatiotemporal RealTime Imaging of the Differentiation Dynamics of Transplanted Neural Stem Cells after Traumatic Brain Injury.’’ Wang Z, Wang Y, Wang Z, Zhao J, Gutkind JS, Srivatsan A, Zhang G, Liao HS, Fu X, Jin A, Tong X, Niu G, Chen X. ACS Nano. 2015. 9:6683-95. doi:10.1021/acsnano.5b00690

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855-Pos Board B635 Healing of Human Intestinal Organoids Emily A. Berglund1, Jonathan N.V. Martinson2, Jason R. Spence3, Seth T. Walk4, James N. Wilking1. 1 Chemical and Biological Engineering, Montana State University, Bozeman, MT, USA, 2Montana State University, Bozeman, MT, USA, 3University of Michigan, Ann Arbor, MI, USA, 4Microbiology and Immunology, Montana State University, Bozeman, MT, USA. Organoids are millimeter-scale tissues that replicate the structure and function of naturally formed organs. These tissues are grown in the lab through the directed differentiation of stem cells and have potential uses in biotechnology. Human intestinal organoids (HIO’s) serve as a model system for the small intestine and offer potential in regenerative medicine, drug formulation testing, and microbiome research. HIO’s are roughly spherical in shape, with a closed-shell structure containing an inner aqueous liquid. HIO’s exhibit healing when the tissue shell is compromised; in fact, a single HIO sliced in half can heal to form two closed, water-tight structures. Yet, the mechanism by which this occurs and the role that substrate mechanics plays in the healing process is not known. Here we describe the use of laser dissection, fluorescence and traction force microscopies to elucidate the mechanism by which threedimensional tissue shells recover from damage and form closed surfaces. Our results could offer insight into the healing of surfaces on three dimensional tissues. 856-Pos Board B636 Developing High-Speed AFM and Nanomechanical Characterizations for Biomedical Investigations Hsien-Shun Liao1,2, Nicole Carvajal1, Xiao Fu1,3, Maryam Raftari1, Albert J. Jin1. 1 National Institute of Biomedical Imaging and Bioengineering/NIH, Bethesda, MD, USA, 2National Taiwan University, Taipei, Taiwan, 3Beijing Institute of Technology, Beijing, China. Atomic force microscopy (AFM) is an advanced tool that enables the visualization of dynamic behaviors and the quantification of mechanical properties under physiological conditions of biological samples from macromolecules to cells and beyond. We have constructed a high-speed AFM toward a temporal resolution of 1s or shorter using a digital versatile disc (DVD) pickup head to detect deflections from a small cantilever. In addition, a flexure-guided scanner and a sinusoidal scan method were implemented to reach 100 line/s scan rate and successful images of clathrin cages and amyloid-b fibrils in buffers. To better define nanomechanical properties of biomedical samples ranging from nanomedicine constructs to live cells and EMC-like matrices (1), we have relied on optimizing commercial AFM systems and on using force-volume and quantitative nanomechanical mapping methods. This work showcases the value of high-speed and quantitative AFM in visualizing dynamic biological behavior at nanometer scale and in defining nanomechanical properties for understanding fundamental biomedical questions. Ref (1) ‘‘Local 3D matrix microenvironment regulates cell migration through spatiotemporal dynamics of contractility-dependent adhesions,’’ Doyle AD, Carvajal N, Jin A, Matsumoto K, and Yamada KM, Nature Commun., in press (2015). 857-Pos Board B637 Fluidic-Resistance Control in Arterial Pulsation Simulators Yuma Shiraishi1, Yun Jung Heo1, Atsushi Sakuma2. 1 Tokyo University of Agriculture and Technology, Tokyo, Japan, 2Kyoto Institute of Technology, Kyoto, Japan. Arterial pulsation is deeply related to the diseases and physical properties of arteries. Cultivation of an artificial artery under various pulsation will reveal the interaction between pulsation and arterial stiffness. The ideal system can culture an artificial artery with pulsation and measure arterial stiffness, simultaneously. To reach the ideal system, we first developed the arterial pulsation simulator that could generate a periodic solitary-wave. The system is composed of three parts: a pulsation input part, a silicone tube, and a resistance part correspond to the human heart, the artery, the peripheral resistance, respectively. Previously, we could create a periodic solitary-wave by installing solenoid valve in the input part of the system. Although we could obtain pulsationlike wave-profiles, there was pressure drop after a period of pulsation; such pressure drop is not shown in in vivo pulsation. Since the pressure drop is originated from a reflection wave, fluidic resistance has to be controlled to realize in-vivo-like pulsation. However, effects of fluidic resistance on pulsation