vascular research initiatives conference
TRANSCRIPT
Vascular Research Initiatives Conference
VRIC 2020 ONLINE: From Discovery to Translation
DAY 1
November 5th, 2020, 1:00-3:00 PM Central Time
ABSTRACT SESSION I: ARTERIAL REMODELING AND DISCOVERY SCIENCE FOR VENOUS DISEASE
ABSTRACT SESSION II: ATHEROSCLEROSIS AND ROLE OF THE IMMUNE SYSTEM
DAY 2
November 12th, 2020, 7:00-9:00 PM Central Time
ABSTRACT SESSION III: AORTOPATHIES AND NOVEL VASCULAR DEVICES
ABSTRACT SESSION IV: VASCULAR REGENERATION, STEM CELLS AND WOUND HEALING
Program Overview
The 34th annual Vascular Research Initiatives Conference (VRIC), presented by the Society for
Vascular Surgery®, is designed to encourage interaction and collaboration between vascular
surgeon investigators and scientists from other vascular biology‐related disciplines. An additional
objective is to stimulate interest in research among trainees who are aspiring academic vascular
surgeons. The VRIC 2020 ONLINE is taking place on November 5th and 12th as virtual online events
instead of the traditional in-person meeting in response to the 2020 COVID-19 pandemic.
2020-2021 SVS Basic and Translational Research Committee:
Luke Brewster – Chair, Emory University, Atlanta, GA
Jayer Chung, MD, University of Texas Southwestern Medical Center, Houston, TX
Scott Damrauer, MD, University of Pennsylvania School of Medicine, Philadelphia, PA
Sean English, MD, Washington University‐Barnes Hospital, St. Louis, MO
Katherine Gallagher, MD, University of Michigan, Ann Arbor, MI
Caitlin Hicks, MD, MS, Johns Hopkins University Program, Baltimore, MD
Karen Ho, MD, Northwestern University, Chicago, IL
Jason MacTaggart, MD, University of Nebraska Medical Center, Omaha, NE
Firas Mussa, MD, MS, University of South Carolina School of Medicine, New Hyde Park, NY
Ryan McEnaney, MD, University of Pittsburgh Medical Center, Pittsburgh, PA
Nicholas Osborne, MD, University of Michigan Hospitals, Ann Arbor, MI
Elsie Ross, MD, Stanford Hospital and Clinics, Stanford, CA
Ulka Sachdev, MD, University of Pittsburgh, Pittsburgh, PA
Kevin Southerland, MD, Duke University, Durham, NC
Bryan Tillman, MD, PhD, University of Pittsburg, Pittsburgh, PA
Shirling Tsai, MD, University of Texas Southwestern Medical Center, Houston, TX
Areck Ucuzian, MD, PhD, University of Maryland School of Medicine, Baltimore, MD
Dai Yamanouchi, MD, University of Wisconsin, Madison, WI
AGENDA
Thursday, November 5, 2020 (All times are Central Time US)
Luke Brewster, MD, Chair, Basic and Translational Research Committee
Moderator: Ulka Sachdev, MD
Moderator: Firas Mussa, MD
1:00 PM INTRODUCTORY REMARKS
VRIC 2020 ONLINE Day 1
ABSTRACT SESSION I: ARTERIAL REMODELING AND DISCOVERY SCIENCE FOR VENOUS DISEASE
1:05 PM
^* Elastic Fibers Of The Internal Elastic Lamina Are Unraveled But Not Created
With Expanding Arterial Diameter In Arteriogenesis
Derek Afflu, Univ of Pittsburgh Medical Ctr, PITTSBURGH, PA; Dylan D McCreary,
Nolan Skirtich, VA PHS, Pittsburgh, PA; Kathy Gonzalez, UPMC, Pittsburgh, PA;
Edith Tzeng, UNIVERSITY PITTSBURGH, Pittsburgh, PA; Ryan M McEnaney
VA PHS, Pittsburgh, PA
1:12 PM
A Synthetic Resolvin Analogue (Benzo-Rvd1) Attenuates Vascular Smooth
Muscle Cell (VSMC) Migration And Neointimal Hyperplasia
Alexander Kim, SMITH CARDIOVASCULAR RESEARCH BLDG, San Francisco, CA;
Hideo Kagaya, UCSF, San Francisco, CA; Mian Chen, SMITH CARDIOVASCULAR
RESEARCH BLDG, San Francisco, CA; Bian Wu, Giorgio Mottola, UCSF, San
Francisco, CA; Matthew R Spite, HARVARD MEDICAL SCHOOL, Boston, MA;
Brian Sansbury, Brigham and Women's Hosp, Boston, MA; Michael S Conte, UC
SAN FRANCISCO, San Francisco, CA
1:19 PM
Evaluation Of A Targeted Drug Eluting Intravascular Nanotherapy To Prevent
Neointimal Hyperplasia In A Novel Atherosclerotic Rat Model
Emily R Newton, Univ of North Carolina, Chapel Hill, NC; Brooke Dandurand,
UNC Sch of Med, Chapel Hill, NC; David C Gillis, Kui Sun, Robin Siletzky, Univ of
North Carolina, Chapel Hill, NC; Mark R Karver, Samuel I Stupp, Northwestern
Univ, Chicago, IL; NICK D TSIHLIS, UNC-CHAPEL HILL, Chapel Hill, NC; Melina R
KIBBE, Univ of North Carolina, Chapel Hill, NC
1:26 PM
Programmed Death Ligand-1 Regulates T-cells And M2 Macrophages To
Control Wall Thickening During Arteriovenous Fistula Maturation
Yutaka Matsubara, Luis Gonzalez, Jia Liu, Arash Fereydooni, John Langford,
Shin-rong Lee, Jolanta Gorecka, Mingjie Gao, Xixiang Gao, Ryosuke Taniguchi,
Bogdan Yatsula, Alan Dardik, Dept of Surgery, Yale Univ Sch of Med, New
Haven, CT
1:33 PM
1:40 PM
TRPC6 Depletion Results In Loss Of Myocardin And Phenotypic Modulation In
Vascular Smooth Muscle Cells
Andrew H. Smith, Priya Putta, Erin Driscoll, Pinaki Chaudhuri, Cleveland Clinic,
Cleveland, OH; Michael Rosenbaum, Louis Stokes Cleveland VAMC,
Cleveland, OH; Linda M. Graham, Cleveland Clinic, Cleveland, OH
Periadventitial Delivery Of Simvastatin From Microparticles Attenuates
Arteriovenous Fistula Outflow Vein Neointimal Hyperplasia
Chenglei Zhao, Mayo Clinic, Rochester, MN; Sean T Zuckerman, Affinity
Therapeutics, LLC, Cleveland, OH; Chuanqi Cai, Michael L Simeon, Avishek
Singh, Sreenivasulu Kilari, Mayo Clinic, Rochester, MN; Julius N Korley, Affinity
Therapeutics, LLC, Cleveland, OH; Sanjay Misra, MAYO CLINIC, Rochester, MN
1:50-2:00 Pause and Reset Presenters
Moderator: Ryan McEnaney, MD
Moderator: Caitlin Hicks, MD
2:00 PM ^ Absence Of Cpla2 In Lrp1 Smooth Muscle Cell Deficient Mice Promotes
Severe Aortic Atherosclerotic Disease
Roberto Ivan Mota Alvidrez, Shirling Tsai, Joachim Herz, Univ of Texas
Southwestern Medical Ctr, Dallas, TX
2:07 PM Serum Circulating Fatty Acid Synthase As A Diagnostic Biomarker For
Chronic Limb-Threatening Ischemia
Shirli Tay, Amanda Penrose, Gayan S De Silva, Yan Yan, Washington Univ Sch of
Med, St. Louis, MO; Clay F Semenkovich, WASHINGTON UNIV SCH OF MED, Saint
Louis, MO; Mohamed A Zayed, WASHINGTON UNIVERSITY SCHOOL M, Saint
Louis, MO
2:14 PM ZEB2 Regulates Activation And Exhaustion Programming Of CD8 T Cells In
Atherosclerosis
Dawn Fernandez, Nicolas F Fernandez, adeeb Rahman, Christopher Hill, Mount
Sinai, New York, NY; Roza Shamailova, Icahn Sch of Med, New York, NY;
Seunghee Kim-schulze, J Mocco, Peter Faries, Miriam Merad, Chiara
Giannarelli, Mount Sinai, New York, NY
2:21 PM The Role Of Mast Cells In Atherosclerotic Plaque Calcification
Nikolaos-Taxiarchis Skenteris, Malin Kronqvist, Mariette Lengquist, Karolinska
Inst, Solna, Sweden; Ulf Hedin, Karolinska Inst, Stockholm, Sweden; Erik L Biessen,
Maastricht Univ Medical Ctr, Maastricht, Netherlands; Ljubica Matic, Karolinska
Inst, Stockholm, Sweden
ABSTRACT SESSION II: ATHEROSCLEROSIS AND ROLE OF THE IMMUNE SYSTEM
2:28 PM
2:35 PM
2:42 PM
+ Inflammatory Activity of Human Perivascular Adipose Tissue in Abdominal
Aortic Aneurysms
Jorn Meekel, Marina Dias-Neto, Natalija Bogunovic, Gloria Conceigao, Claudia
Sousa-Mendes, Gawin R. Stoll, Adelino Leite-Moreira, Jennifer Huynh, Dimitra
Micha, Etto C. Eringa, Ron Balm, Jan D. Blankensteijn, Kak K. Yeung
Polygenic Risk Score Identifies Patients At Increased Risk For Abdominal Aortic
Aneurysm And May Benefit From Ultrasound Screening
Derek Klarin, Univ of Florida Coll of Med, Gainesville, FL; Ozan Dikilitas, Mayo
Clinic, Rochester, MN; Brooke Wolford, University of Michigan, Ann Arbor, MI;
Michael Levin, Univ of Pennsylvania Sch of Med, Philadelphia, PA; Ishan
Paranjpe, Mount Sinai Sch of Med, New York, NY; Renae Judy, Univ of
Pennsylvania Sch of Med, Philadelphia, PA; Julie Lynch, Dept of Veterans
Affairs, Salt Lake City Health Care System, Salt Lake City, UT; Themistocles L
Assimes, Stanford Univ Sch of Med, Palo Alto, CA; Yan Sun, Emory Univ, Atlanta,
GA; Daniel Rader, Univ of Pennsylvania Sch of Med, Philadelphia, PA; Peter W
Wilson, Emory Univ Sch of Med, Atlanta, GA; Salvatore Scali, Scott Berceli, Univ
of Florida Coll of Med, Gainesville, FL; Sekar Kathiresan, Verve Therapeutics,
Cambridge, MA; Pradeep Natarajan, Massachusetts General Hosp, Boston,
MA; Girish Nadkarni, Mount Sinai Sch of Med, New York, NY; Cristen Willer, Univ
of Michigan, Ann Arbor, MI; Iftikhar Kullo, Mayo Clinic, Rochester, MN; Scott M
Damrauer, University of Pennsylvania Sch of Med, Philadelphia, PA; Philip Tsao,
Stanford Univ Sch of Med, Palo Alto, CA
Vascular Surgery and the K Award: An Abundance of Successes: Presentation
will highlight the K award recipients, SVSF winners, and SVSF commitment to
helping young surgeons succeed in their academic pursuits.
Luke Brewster, MD, and Katherine Gallagher, MD
Adjourn (day 1)
Thursday, November 12, 2020 (All times are Central Time US)
Luke Brewster, MD, Chair, Basic and Translational Research Committee
Moderator: Jason MacTaggart, MD
Moderator: Elsie Ross, MD
7:05 PM
^* Epigenetic Modifications Influence Macrophage-mediated Inflammation
In abdominal Aortic Aneurysms
Frank Michael Davis, Aaron Dendekker, Amrita Joshi, Sonya Wolf, Christopher
Audu, William J Melvin, UNIVERSITY OF MICHIGAN, Ann Arbor, MI; Hong Lu, UNIV
KENTUCKY, Lexington, KY; Alan Daugherty, UNIVERSITY OF KENTUCKY, Lexington,
KY; Johann Gudjonsson, UNIVERSITY OF MICHIGAN, Ann Arbor, MI; Katherine A
Gallagher, UNIVERSITY MICHIGAN, Ann Arbor, MI
7:12 PM The Perfuse Dual Chamber Stent Improves Donor Organ Recovery In A Porcine
Model
Bryan W. Tillman, Catherine Go, Brian Frenz, UNIV OF PITTSBURGH MED CTR,
Pittsburgh, PA; Moataz Elsisy, UNIV OF PITTSBURGH, Pittsburgh, PA; Youngjae
Chun, UNIVERSITY OF PITTSBURGH, Pittsburgh, PA
7:19 PM Endovascular Repair Of Blunt Thoracic Aortic Trauma Is Associated With
Increased Left Ventricular Mass, Hypertension, And Off-target Aortic
Remodeling
Alexey Kamenskiy, Univ of Nebraska Omaha, Omaha, NE; Paul Aylward, Univ
of Nebraska Medical Cent, Omaha, NE; Anastasia Desyatova, Univ of
Nebraska Omaha, Omaha, NE; Matthew DeVries, Christopher Wichman, Univ
of Nebraska Medical Ctr, Omaha, NE; Jason N Mactaggart, Univ of Nebraska
Medical Cent, Omaha, NE
7:26 PM
Maresin 1 Attenuates Murine Abdominal Aortic Aneurysms Via Vascular
Smooth Muscle Cell Dependent TGF-β Signaling
Craig T Elder, Univ of Florida, Gainesville, FL; Gang Su, Gainesville, FL; Amanda
Filiberto, Univ of Florida Coll of Me, GAINESVILLE, FL; Guanyi Lu, Univ of Florida
Coll of Me, GAINESVILLE, FL, Gainesville, FL; Zhihua Jiang, UNIVERSITY OF
FLORIDA COM, Gainesville, FL; Ashish Sharma, UNIVERSITY OF FLORIDA,
Gainesville, FL; Gilbert R UPCHURCH Jr., Univ of FL Dept of Surgery, Gainesville,
FL
7:00 PM INTRODUCTORY REMARKS
VRIC 2020 ONLINE Day 2
ABSTRACT SESSION III: AORTOPATHIES AND NOVEL VASCULAR DEVICES
7:33 PM
7:40 PM
7:47 PM
7:54 PM
A Theranostic Role For Ccr2 In Rodent Abdominal Aortic Aneurysm
Development And Rupture
Sergio Sastriques Dunlop, Washington Univ Sch Med, Saint Louis, MO; Batool
Arif, Sean English, Washington Univ Sch Med, SAINT LOUIS, MO
Pharmacological Inhibition Of Bruton's Tyrosine Kinase Attenuates
Experimental Abdominal Aortic Aneurysms
Baohui Xu, STANFORD UNIVERSITY SCHOOL OF MEDIC, Stanford, CA; yankui li,
Stanford Univ Sch of Med, Stanford, CA; Gang Li, STANFORD UNIVERSITY
SCHOOL OF MEDIC, Stanford, CA; Yingbin Ge, Nanjing Medical Univ, Nanjing,
China; Jia Guo, Stanford Univ Sch of Med, Stanford, CA; Weirong Fang, China
Pharmaceutic Univ, Nanjing, China; Wei Wang, Xiangya Hosp, Central South
Univ, Chnagsha, China; Fanru Shen, STANFORD UNIVERSITY SCHOOL OF MEDIC,
Stanford, CA; Takahiro Shoji, Vascular Surgery, Stanford, Stanford, CA; Toru
Ikezoe, Xiaoya Zheng, STANFORD UNIVERSITY SCHOOL OF MEDIC, Stanford, CA;
Sihai Zhao, Stanford Univ Sch of Med, Stanford, CA; Xiaofeng Chen, TAIZHOU
HOSPITAL, Taizhou; Masaaki MIYATA, KAGOSHIMA CITY HOSPITAL, Kagoshima,
Japan; Alan Daugherty, UNIVERSITY OF KENTUCKY, Lexington, KY; Hong Lu, Univ
of Kentucky Sch of Med, Lexington, KY; Ronald L DALMAN, STANFORD
UNIVERSITY MEDICAL CE, Stanford, CA
Landscape Of Human Secretome In Abdominal Aortic Aneurysm Profiled
By Single-cell RNA Sequencing.
Bhama Ramkhelawon, NYU MEDICAL CENTER, New York, NY; Tarik Hadi, NYU
Langone Medical Ctr, New York, NY; Michele Silvestro, NYU Sch of Med, New
York, NY
Pause and Reset Presenters
Moderator: Jayer Chung, MD
Moderator: Kevin Southerland, MD
8:00 PM ^* Autophagy Remodels Mitochondria During Differentiation And Enhances
Longevity Through Ulk1 Kinase Signaling Of Induced Pluripotent Stem Cell-
derived Endothelial Cells
Katherine E Hekman, Kyle Koss, David Z Ivancic, Northwestern Univ, Chicago, IL;
Congcong He, Chicago, IL; Jason A Wertheim, Northwestern Univ, Chicago, IL
ABSTRACT SESSION IV: VASCULAR REGENERATION, STEM CELLS AND WOUND HEALING
8:07 PM
* Downregulation Of Inflammation And A Cascade Of Pro-angiogenic Signals
Mediate The Beneficial Effects Of Gene-modified Stem Cell Therapy In Hindlimb
Ischemia
Hallie J Quiroz, Univ of Miami, Miami, FL; Hongwei Shao, Yan Li, Samantha F
Valencia, Punam Parikh, Univ of Miami, Miami, FL; Yulexi Ortiz, Univ of Miami Miller
Sch of Med, Miami, FL; Roberta Lassance-Soares, Univ of Miami, Miami, FL; Zhao-
Jun Liu, Univ of Miami Miller, Miami, FL; Omaida Caridad Velazquez, Univ of Miami
Miller Sch of Med, Miami, FL
8:14 PM
Different Outcomes after Revascularization or Standard Supervised Exercise
Treadmill Training of Claudicating Patients with Peripheral Artery Disease
Shuai Li, Jonathan Robert Thompson, Univ of Nebraska Med Ctr, Omaha, NE;
Sara A Myers, Univ of Nebraska at Omaha, Omaha, NE; Julian K Kim, UNIV
NEBRASKA MEDICAL CENTER, Omaha, NE; Panagiotis Koutakis, Florida State Univ,
Tallahassee, FL; Mark Williams, Creighton Cardiac Ctr, Omaha, NE; Zhen Zhu, Univ
of Nebraska Med Ctr, Omaha, NE; Molly Schieber, Univ of Nebraska Medical
Cent, Omaha, NE; Timothy Lackner, Gregory Willcockson, Christina Shields,
Katyarina Brunette, Holly Despiegelaere, Univ of Nebraska Med Ctr, Omaha, NE;
Iraklis I Pipinos, UNIV NEBRASKA MEDICAL CTR, Omaha, NE; George Casale, Univ
of Nebraska Medical Cent, Omaha, NE
8:21 PM Differential Expression Of Canonical Mitosis And Dna-damage Repair Pathways
Characterize Muscle Satellite Cells Affected By Pad
Ricardo Ferrari, Guangzhi Cong, Bryan Thompson, Debbie Hollingshead,
Janette Lamb, Xiangdong Cui, Ulka Sachdev, UPMC, Pittsburgh, PA
8:28 PM Fluvastatin And Vegf Containing Resins Promote Angiogenesis And
Arteriogenesis
Furqan Muqri, SUNY Upstate Medical Ctr & Syracuse VA Medical Ctr, Syracuse,
NY; Dandan Guo, SUNY Upstate Medical Univ, Syracuse, NY; Mohammed
Kassem, Mary DaCosta, SUNY Upstate Medical Ctr & Syracuse VA Medical Ctr,
Syracuse, NY; David Bruch, SUNY Upstate Medical Univ, Syracuse, NY; Kristopher
Maier, SUNY Upstate Medical Ctr & Syracuse VA Medical Ctr, Syracuse, NY;
Juntao Luo, SUNY Upstate Medical Ctr, Syracuse, NY; Vivian Gahtan, SUNY
Upstate Medical Ctr & Syracuse VA Medical Ctr, Syracuse, NY
8:35 PM
8:42 PM
8:49 PM
8:56 PM
9:03 PM
9:10 PM
Increased Neutrophil Elastase Expression In Regions Of High Intraluminal
Thrombus Deposition In Human Abdominal Aortic Aneurysms.
Matthew Levesque, Lauren Bath, Max Rady Coll of Med, Winnipeg, MB, Canada;
Annie Ducas, Royal Victoria Regional Health Ctr, Barrie, ON, Canada; April Boyd,
Health Sciences Ctr, Winnipeg, MB, Canada
Adult and Pediatric Fibromuscular Dysplasia Are Genetically Distinct Dysplasia-
associated Arterial Diseases
Yu Wang, Dawn M. Coleman, Univ of Michigan Medical Sch, Ann Arbor, MI; Min-
Lee Yang, Issaquah, WA; Susan Blackburn, Issaquah, WA, Ann Arbor, MI; Kristina
L. Hunker, Univ of Michigan Medical Sch, Ann Arbor, MI; Heather L Gornik, Univ
Hosp Cleveland, Cleveland, OH; Jun Li, Univ of Michigan, Ann Arbor, MI; James
C Stanley, 2210 Taubman Ctr, Ann Arbor, MI; Santhi Ganesh, 2210 Taubman Ctr,
Ann Arbor, MI, Ann Arbor, MI
Impact Of Dietary Tungsten And Topical Nitrite In Diabetic Wound Healing And
The Composition Of The Wound Microbiome
Kathy Gonzalez, Karim M Salem, UPMC, Pittsburgh, PA; Barbara Methé, Kelvin
Li, Guiying Hong, Univ of Pittsburgh, Pittsburgh, PA; Edith Tzeng, UNIVERSITY
PITTSBURGH, Pittsburgh, PA
MicroRNA-181b Regulates Critical Limb Ischemia In Diabetic Mice
Henry S Cheng, Brigham and Women's Hosp, Boston, MA; Daniel Perez-
Cremades, BWH, Boston, MA; Marc P Bonaca, CPC Clinical Res, Aurora, CO;
Mark Feinberg, Brigham and Women's Hosp, Boston, MA
Nitric Oxide Bioavailability Is Reduced In Peripheral Artery Disease In Association
With Increased Oxidative Stress And An Altered Biopterin System
Ahmed Ismaeel, Evlampia Papoutsi, Florida State Univ, Tallahassee, FL; William T
Bohannon, Robert Smith, Baylor Scott and White Heath, Temple, TX; Robert
Brumberg, Vascular Surgery Associates, Tallahassee, FL; Carlos H Castro, Jeffrey S
Kirk, Capital Regional Medical Ctr, Tallahassee, FL; Iraklis I Pipinos, UNIV NEBRASKA
MEDICAL CTR, Omaha, NE; Panagiotis Koutakis, Florida State Univ, Tallahassee, FL
Adjourn (day 2)
* SVS Foundation VRIC Trainee Awardee
^ Best Abstract in Category
+ 2020 ESVS Young Researcher Prize winner
701
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
With a better understanding of the mechanisms governing arteriogenesis, we may find pharmacological
targets that could enhance collateral artery formation. This could lead to a pharmacological therapy that
effectively increases blood flow to an ischemic tissue which would revolutionize management of
patients with arterial occlusive diseases.
Professional Practice Gap:
Patients suffering from arterial occlusive diseases would benefit tremendously if there were an effective
medical therapy that could revascularize ischemic tissues. Collateral arteries develop (in a process
termed arteriogenesis) as a spontaneous adaptation when a large conductance artery becomes
occluded. The importance of collateral arteries for maintaining organ and limb perfusion has long been
recognized, but their development is incompletely understood. If enhanced collateral artery
development could be achieved thorough pharmacological means, it would offer a promising means to
noninvasively enhance perfusion for these patients.
Elastic Fibers of The Internal Elastic Lamina Are Unraveled But Not Created With Expanding Arterial
Diameter In Arteriogenesis
Derek Afflu, Univ of Pittsburgh Medical Ctr, PITTSBURGH, PA; Dylan D McCreary, Nolan Skirtich, VA PHS,
Pittsburgh, PA; Kathy Gonzalez, UPMC, Pittsburgh, PA; Edith Tzeng, UNIVERSITY PITTSBURGH,
Pittsburgh, PA; Ryan M McEnaney, VA PHS, Pittsburgh, PA
Objective: Outward remodeling of the arterial wall is essential during arteriogenesis to grow small
arterioles into conductance vessels. Prior reports suggest that internal elastic laminae (IEL) are degraded
and rebuilt in arteriogenesis. However, it is unclear whether new elastic fibers can be synthesized in
adult arteries. We sought to understand the IEL changes that occur in arteriogenesis. Methods: Rats age
8-12 wks had femoral artery ligation (FAL) with distal arteriovenous fistula (AVF) placement to enhance
collateral growth. Rats were fed β-aminopropionitrile (BAPN) in drinking water to inhibit lysyl
oxidoreductase (LOX). Rats were euthanized at 2 days to 12 wks and arteries were harvested (N=5/time
point) for multiphoton microscopy and quantitative measurement of desmosine content. Human
collateral arteries from amputation specimens were also analyzed for comparison. Results: At 2 days,
FAL+AVF treated profunda femoral arteries (PFA) showed maximal vasodilation, but IEL retained normal
structure. At 4 wks, PFA diameters increased over contralateral sham-operated PFAs (mean 255 +/-
18.5%) and IEL unraveled into a loose web of elastic fibers which persisted at 12 wks. This IEL pattern
was also seen in human collaterals. Despite PFA size increase, elastin content trended downward (Sham,
0.41 ± 0.07ng; AVF, 0.31 ± 0.06ng, p=0.058). LOX inhibition resulted in severe fragmentation of IEL in
PFAs and abnormally thickened elastic fibers in 3rd order collaterals. Conclusion: In arteriogenesis, the
IEL unravels and remains web like for up to 12 wks after FAL+AVF, a pattern also observed in human
collaterals. Despite the growth in arterial diameter, elastin content was not increased, suggesting new
elastic fibers are not created. Stabilization of IEL changes appeared to be mediated by LOX. Further
dissection of the mechanisms of arterial remodeling in arteriogenesis will allow us to harness this
adaptive process for therapeutic purposes.
790
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
This research is demonstrating the efficacy of a novel compound that may help to reduce neointimal
hyperplasia following vascular intervention
Professional Practice Gap:
The analogue discussed in this abstract may provide durable patency following vascular interventions
A Synthetic Resolvin Analogue (Benzo-Rvd1) Attenuates Vascular Smooth Muscle Cell (VSMC)
Migration And Neointimal Hyperplasia
Alexander Kim, Evan C Werlin, SMITH CARDIOVASCULAR RESEARCH BLDG, San Francisco, CA; Hideo
Kagaya, UCSF, San Francisco, CA; Mian Chen, SMITH CARDIOVASCULAR RESEARCH BLDG, San Francisco,
CA; Bian Wu, Giorgio Mottola, UCSF, San Francisco, CA; Matthew R Spite, HARVARD MEDICAL SCHOOL,
Boston, MA; Brian Sansbury, Brigham and Women's Hosp, Boston, MA; Michael S Conte, UC SAN
FRANCISCO, San Francisco, CA
Background: Persistent inflammation following vascular injury drives neointimal hyperplasia (NIH).
Specialized lipid mediators (SPM) play a critical role in the process of resolution. We investigated the
effects of a novel synthetic SPM on vascular cells and in a model of rat carotid angioplasty.
Methods: Human venous VSMC and endothelial cells (EC) were employed in toxicity, migration,
proliferation and NF-κB activation assays. A model of rat carotid angioplasty was used to evaluate the
effects of Benzo-RvD1 (BRvD) applied externally via 25% Pluronic gel. Drug concentration was measured
at 3 days after injury via LC-MS/MS, and effects on vessel morphometry were examined at 14 days after
injury.
Results: BRvD demonstrated no significant cytotoxicity, and modest anti-proliferative activity on VSMC
at 100 nM. BRvD significantly attenuated VSMC migration across a range of concentrations (0.1-100 nM)
that was equivalent or better than 17R-RvD1, a naturally-occurring SPM (Figure 1). BRvD (10-100nM)
inhibited NF-κB translocation in cytokine-stimulated EC by 12-21% (p<0.01), similar to 17R-RvD1.
Following external delivery, BRvD was detectable in rat carotid tissue at 3 days (mean 0.17 pg/mg; n=3)
after injury. Periadventitial treatment with BRvD reduced carotid neointimal thickness at 14 days
compared to controls, with similar efficacy to 17R-RvD1 (Figure 2).Conclusions: Benzo-RvD1 is a
synthetic analogue of the SPM 17R-RvD1 that demonstrates similar in vitro and in-vivo efficacy to inhibit
NIH as its naturally occurring cognate. The enhanced stability of BRvD may provide therapeutic
advantages for anti-restenosis strategies.
715
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
The results of this project, and subsequent projects it leads to, can lead to a paradigm shifting
technology that can completely alter how we treat vascular diseases, improving outcomes and the
overall cost of cardiovascular disease.
Professional Practice Gap:
Current vascular therapies often fail from restenosis due to neointimal hyperplasia development. Drug
eluting stents developed to combat this problem inhibit healing of the endothelial lining and lead to
catastrophic late in-stent thrombosis and now many of these products are being cautioned against by
the FDA for increasing mortality. This research is developing a new therapeutic to prevent restenosis
after vascular intervention, to make these procedures safer and more durable.
Evaluation of A Targeted Drug Eluting Intravascular Nanotherapy To Prevent Neointimal Hyperplasia
In A Novel Atherosclerotic Rat Model
Emily R Newton, Univ of North Carolina, Chapel Hill, NC; Brooke Dandurand, UNC Sch of Med, Chapel
Hill, NC; David C Gillis, Kui Sun, Robin Siletzky, Univ of North Carolina, Chapel Hill, NC; Mark R Karver,
Samuel I Stupp, Northwestern Univ, Chicago, IL; NICK D TSIHLIS, UNC-CHAPEL HILL, Chapel Hill, NC;
Melina R KIBBE, Univ of North Carolina, Chapel Hill, NC
Introduction: Vascular interventions often fail due to neointimal hyperplasia. We developed a
biocompatible nanotherapeutic that, when given systemically, targets sites of arterial injury and inhibits
neointimal hyperplasia in healthy arteries. Given the oxidative environment of atherosclerosis, the aim
of this study was to evaluate targeting and efficacy of our nanotherapy in a novel atherosclerotic rat
model. We hypothesize that our targeted drug-eluting nanofiber will bind sites of injury and inhibit
neointimal hyperplasia in an atherosclerotic environment. Methods: Knockout status, hyperlipidemia,
and plaque development were assessed in Sprague Dawley ApoE knockout (ApoE-/-) rats using Western
blot analysis, blood work, and Oil Red O (ORO) staining. Fluorescently labelled collagen-targeted, nitric
oxide-releasing peptide amphiphiles (CBP-NO PA) were synthesized by solid-phase peptide synthesis and
purified by HPLC. PA nanofiber formation was assessed by transmission electron microscopy (TEM).
Male 16-week-old rats underwent carotid balloon injury and tail vein injection of nanofiber (5 mg).
Targeting was quantified using fluorescent microscopy and efficacy was assessed via morphometric
analysis. Controls included non-targeted nanofibers and uninjured carotid arteries. Results: Western
blot analysis confirmed ApoE-/- status and blood work showed a 3-fold increase in cholesterol levels in
ApoE-/- vs. wild type rats (292±22 vs. 1050±4 mg/dL, p<0.05). ORO staining showed lipid-rich lesions in
ApoE-/- aortas. PAs were >95% pure and formed nanofibers on TEM. After carotid artery injury and
injection of the CBP-NO nanofiber, specific targeting to the site of arterial injury was seen after 20
minutes on fluorescent microscopy, while uninjured carotid and non-targeted NO nanofibers had
minimal fluorescence (3445±282, 11±2, and 4514±94 AU, p<0.05, n=4/group). Two weeks after injury,
the CBP-NO nanofiber resulted in 67% inhibition of neointimal area and 68% reduction in the
intima/media area ratio vs. injury alone (p<0.0001). Conclusion: We have shown our targeted, drug-
eluting nanofiber localizes to sites of vascular injury and significantly decreases neointimal hyperplasia
after 2 weeks in a novel atherosclerotic rat model.
786
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
PDL1 antibody increased T-helper cells and decreased Treg, consistent with increase of inflammation.
Moreover, PDL1 antibody reduced vascular wall thickening during AVF maturation. These results
indicate that PDL1 have a critical role to cause inflammation during AVF maturation and might be a
therapeutic target to improve AVF success.
Professional Practice Gap:
T-cells are essential for arteriovenous fistula (AVF) maturation, however, little is known about regulation
of T-cells (Treg) during AVF maturation. Programed death ligand-1 (PDL1) can induce regulatory T-cells
and reduce inflammation. We show that how PDL1 control T-cell population and AVF maturation.
Programmed Death Ligand-1 Regulates T-cells And M2 Macrophages to Control Wall Thickening
During Arteriovenous Fistula Maturation
Yutaka Matsubara, Luis Gonzalez, Jia Liu, Arash Fereydooni, John Langford, Shin-rong Lee, Jolanta
Gorecka, Mingjie Gao, Xixiang Gao, Ryosuke Taniguchi, Bogdan Yatsula, Alan Dardik, Dept of Surgery,
Yale Univ Sch of Med, New Haven, CT
Introduction: Vascular remodeling during arteriovenous fistula (AVF) maturation is characterized by
infiltration of T-cells and macrophages. We have previously shown that M2 macrophages play an
important role in AVF maturation to reduce inflammation and promote wall thickening. Although T-cells
can activate macrophages, little is known about T-cell regulation of macrophage function during AVF
maturation. Programed death ligand 1 (PD-L1) induces regulatory T-cells (Treg) to suppress other T-cells
and is expressed in endothelial cells. We hypothesized that PD-L1 induces Treg accumulation in the AVF
to promote AVF maturation. Methods: We used the mouse aortocaval AVF model. Intraperitoneal
injection of PD-L1 antibody (3x/wk) was used to inhibit PD-L1; control was matched IgG2 isotype
antibody. Helper T-cells (Th)-1, Th2, Treg, macrophage accumulation in the AVF was assessed by
immunofluorescence with their specific markers. Vascular wall thickening was assessed by elastin van
Gieson stain. Results: Inhibition of PD-L1 significantly increased accumulation of Th1 (15.9 vs 8.6
cells/hpf; p<0.05) and Th2 (18.0 vs 9.7 cells/hpf; p<0.05), but decreased Treg (3.8 vs 11.2 cells/hpf;
p<0.05) cells compared with control. PD-L1 significantly inhibited accumulation of TGM2+ (M2)
macrophages (5.8 vs 15.7 cells/hpf; p<0.05) and CD206+ (M2) macrophages (2.6 vs 13.2 cells/hpf;
p<0.05), but not iNOS+ (M1) macrophages (8.8 vs 10.0 cells/hpf; p=0.23) or TNF-α+ (M1) macrophages
(9.8 vs 12.3; p=0.29). There was less wall thickening in AVF treated with PD-L1 antibody compared with
control (8.2 µm vs. 17.55 µm; p<0.01) as well as reduced AVF maturation (p=0.03; n=16-17;
Figure). Conclusions: Inhibition of PD-L1 is associated with reduced vascular wall thickening as well as
less Treg and M2 macrophage accumulation in the maturing AVF. These results suggest that PD-L1
induces Treg cells to promote M2 macrophage accumulation during AVF maturation.
702
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
This research describes a novel, TRPC6-dependent pathway that regulates vascular smooth muscle cells
phenotype. The results of this study may lead to mechanism-based therapies to reduce phenotypic
switching, and thus mitigate neointimal hyperplasia, following vascular intervention.
Professional Practice Gap:
The durability of interventions for peripheral and coronary arterial occlusive disease is currently limited.
Neointimal hyperplasia is the major cause of early restenosis and failure of vascular bypass grafting,
angioplasty, and stenting. Strategies that combat neointimal hyperplasia therefore have the potential to
improve outcomes of vascular intervention.
TRPC6 Depletion Results in Loss of Myocardin And Phenotypic Modulation In Vascular Smooth Muscle
Cells
Andrew H. Smith, Priya Putta, Erin Driscoll, Pinaki Chaudhuri, Cleveland Clinic, Cleveland, OH; Michael
Rosenbaum, Louis Stokes Cleveland VAMC, Cleveland, OH; Linda M. Graham, Cleveland Clinic, Cleveland,
OH
Objective:The contractile phenotype of vascular smooth muscle cells (VSMC) is regulated by expression
of the master transcription factor, myocardin. Loss of myocardin is observed during VSMC phenotypic
switching and within the hyperplastic neointima after vascular intervention. We have shown that
depletion of the nonvoltage gated calcium-permeable channel, canonical transient receptor potential 6
(TRPC6) channel, promotes VSMC phenotypic modulation and more severe carotid stenosis following
wire injury. The goal of this study was to determine if TRPC6 regulates VSMC phenotype by altering
expression of myocardin.
Methods: Myocardin expression was assessed in primary aortic wild type (WT) and TRPC6-/-VSMC as well
as common carotid artery (CCA) explants from WT and TRPC6-/-mice. Acute TRPC6 knockdown was
performed in immortalized mouse VSMC (MOVAS cells) using siRNA. Functional alteration of VSMC was
assessed in vitro using a wound healing assay.
Results: Myocardin mRNA levels were significantly reduced in primary aortic TRPC6-/-VSMC compared to
primary aortic WT VSMC (mean 5.53x107fold reduction, n=4, p<0.001). Myocardin protein was also
reduced in TRPC6-/-primary aortic VSMC compared to WT, though not to the same degree (mean 2.03
fold reduction, n=3, p=0.001). Myocardin protein was significantly decreased in whole tissue explants of
TRPC6-/- CCA vs WT CCA (55.5% reduction, n=4 mice per genotype, p=0.007), demonstrating that
significant differences in myocardin expression was present in vivo. In MOVAS cells, acute siRNA-
mediated TRPC6 knockdown induced a 44.7% decrease in myocardin protein (n=3, p=0.044). Wound
healing assays showed a significant increase in the number of migrating TRPC6-/-VSMC compared WT
VSMC in response to growth factor stimulation, supporting our previous findings that TRPC6-/-VSMC are
modulated from a contractile to a proliferative phenotype.
Conclusions: TRPC6 depletion is associated with decreased myocardin and the emergence of pathogenic
VSMC behavior. TRPC6-dependent signaling may therefore be a therapeutic target to promote
myocardin expression and stabilize the VSMC contractile phenotype after arterial injury.
707
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
Periadventitial delivery of simvastatin from microparticles attenuates arteriovenous fistula outflow vein
neointimal hyperplasia. It provides a new option to prevent arteriovenous fistula failure.
Professional Practice Gap:
In the present study, there was no assembled marker on simvastatin or microparticles, which could
illustrate migration or residual dose in the outflow veins.
Periadventitial Delivery of Simvastatin From Microparticles Attenuates Arteriovenous Fistula Outflow
Vein Neointimal Hyperplasia
Chenglei Zhao, Mayo Clinic, Rochester, MN; Sean T Zuckerman, Affinity Therapeutics, LLC, Cleveland,
OH; Chuanqi Cai, Michael L Simeon, Avishek Singh, Sreenivasulu Kilari, Mayo Clinic, Rochester, MN;
Julius N Korley, Affinity Therapeutics, LLC, Cleveland, OH; Sanjay Misra, MAYO CLINIC, Rochester, MN
Background: Venous neointimal hyperplasia (VNH) is vexing problem to maintain arteriovenous fistula
(AVF) patency in end-stage renal disease patients. The available drug delivery systems to prevent VNH
formation are limited. VNH is characterized with increased expression of transforming growth factor- β1
(TGF- β1), vascular endothelial growth factor-A (VEGF-A) and monocyte chemoattractant protein-1
(MCP-1). Hypothesis: Periadventitial delivery of microparticles coated with simvastatin (MP-SIM) could
prevent VNH formation via inhibition of gene expression of TGF- β1, VEGF-A and MCP-1 in a murine AVF
model with chronic kidney disease (CKD). Methods: At day -28, eight-week C57BL/6J male mice were
randomly grouped into control group (MP alone) or MP-SIM group and nephrectomy was used to induce
CKD. At day 0 AVF was created. A volume of 20uL of PBS with 16.6mg/ml of either MP or MP-SIM was
applied to the periadventitia of the proximal AVF outflow vein at the time of AVF creation. Fistula
patency was assessed weekly using Doppler ultrasound. Mice were euthanized at day 3 and 28 for gene
expression and immunohistochemistry staining respectively. Results: At day 3, the gene expression of
TGF- β1, VEGF-A and MCP-1 was significantly decreased in MP-SIM group. At day 28, there was a
significant increase in peak systolic velocity and decrease in the average neointimal area and cell density
in MP-SIM group. At day 28, as assessed using Immunohistochemistry staining, there was a significant
increase in apoptosis and a decrease in the smooth muscle cell, fibroblasts, macrophages, fibrosis and
cellular proliferation in MP-SIM group.Conclusion: Our study indicates that periadventitial delivery of
MP-SIM attenuates VNH 4 weeks after AVF creation. Further studies using a porcine animal model to
confirm these findings are recommended. The potential clinical applicability of controlled-release
simvastatin to decrease expression of TGF-β1, VEGF-A and MCP-1 while increasing apoptosis and
decreasing cellular proliferation is encouraging.
832
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
Results from our studies will identify molecular mechanisms in the process of underpinning novel
therapeutic targets for the devastating consequences of treating atherosclerotic disease plaque
vulnerability and rupture by studying bioactive lipid dysregulation.
Professional Practice Gap:
Our studies aim to understand the molecular mechanisms of atherosclerotic plaque vulnerability and
rupture that continue to cause life-threatening complications affecting millions of people around the
world. Our studies aim to understand atherosclerosis disease development via the abrogation of the
inflammatory process by altered activation of the bioactive lipid machinery.
Absence Of Cpla2 In Lrp1 Smooth Muscle Cell Deficient Mice Promotes Severe Aortic Atherosclerotic
Disease
Roberto Ivan Mota Alvidrez, Shirling TSAI, Joachim Herz, Univ of Texas Southwestern Medical Ctr,
Dallas, TX
Smooth muscle cell targeted deficiency of LDL Receptor related protein 1 (LRP1) in a mouse model
(smLRP1-/-) results in accelerated aortic atherosclerosis through activation of cytoplasmic phospholipase
A2 (cPLA2), leading to reduced ABCA1 expression in vascular smooth muscle cells (VSMCs), and
increased intracellular cholesterol accumulation. We therefore hypothesized that deficiency of cPLA2
would impede atherogenesis in the smLRP1-/- mouse model. Methods: Adult male smLRP1-/-;cPLA2-/-
;LDLR-/- (Triple knockout) mice were placed on a high cholesterol diet (HCD) for 16 weeks and compared
to age- and diet- matched sibling control smLRP1+/+;cPLA2-/-;LDLR-/- mice. Histologic analysis was
performed using en face whole aorta Oil red O (ORO) staining, as well as cross-sectional analysis of the
aortic root with ORO, Picro Sirius Red, Alizarin Red and immunofluorescence. Immunoblot protein
analysis was performed using lysed whole aortas. Data is represented as mean±SEM. Statistical analysis
was performed using one and two way ANOVA with Tukey’s correction. Results: En face ORO analysis
revealed increased lipid accumulation in Triple Knockout mice as compared to controls (60+/-3% vs
13+/-2%, p<0.001) (Figure 1). Uniquely, Triple knockout mice develop extensive necrotic cores and thin
fibrous caps in atherosclerotic lesions in the aortic root (Figure 1). ABCA1 is paradoxically increased both
in whole aorta lysate as well as in immunofluorescence staining of the aortic
root. Conclusions: Deficiency of cPLA2 in the smLRP1-/- mouse model rescued ABCA1 expression, but
unexpectedly increased lipid accumulation within the plaque and generated more vulnerable plaque.
Future studies will underpin the mechanisms that guide severe disease development in our Triple
knockout mice via regulation of ABCA1. Funding: NHLBI R37 HL063762 and NINDS/NIA R01NS108115.
841
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
This study showed that circulating fatty acid synthase (cFAS) is independently associated with CLTI but
not DM. Future studies may explore using cFAS as biomarker for severity of PAD to guide clinical
management, or as a target for development of medical therapeutics.
Professional Practice Gap:
This study evaluates the potential association between serum biomarkers and clinical severity in
patients with diabetes and peripheral artery disease.
Serum Circulating Fatty Acid Synthase As A Diagnostic Biomarker For Chronic Limb-Threatening
Ischemia
Shirli Tay, Amanda Penrose, Gayan S De Silva, Yan Yan, Washington Univ Sch of Med, St. Louis, MO; Clay
F Semenkovich, WASHINGTON UNIV SCH OF MED, Saint Louis, MO; Mohamed A Zayed, WASHINGTON
UNIVERSITY SCHOOL M, Saint Louis, MO
Objectives: Circulating fatty acid synthase (cFAS), a de novo lipogenesis enzyme, is elevated in the serum
of patients with DM and carotid artery stenosis. However, it is unknown whether cFAS is similarly
elevated in patients with DM and advanced PAD. This study aims to evaluate whether cFAS content and
enzyme activity are biomarkers for clinical severity in patients with DM and CLTI. Methods: Serum
samples were prospectively collected from patients undergoing arterial revascularization procedures
and maintained in an IRB-approved institutional biobank. cFAS content and enzyme activity were
evaluated using colorimetric ELISA assays. Multivariable logistic regression was used to evaluate DM and
CLTI outcomes while adjusting for patient clinical characteristics. Hosmer-Lemeshow tests and C-index
assessed goodness-of-fit and classification accuracy. All tests were two-sided and P<0.05 was considered
significant. Results: A total of 86 patients underwent cFAS analysis (67 content; 63 activity). Mean age
was 65 (±8.5) with 67.4% male, 46.5% DM and 47.7% CLTI. Bivariable analyses demonstrated association
of CLTI with cFAS content (P<.01), DM (P=.01) and insulin use (P=.01); DM was associated with BMI
(P=.001), CKD (P=.001) and current smokers (P<.05). On multivariable analysis, CLTI was associated with
cFAS content (OR 1.16 [1.02-1.32] P=.02) and closely with DM (OR 2.72 [0.94-7.89] P=.066); while DM
was associated with BMI (OR 1.13 [1.05-1.23] P<.01) and CKD (OR 4.86 [1.72-13.73] P<.01). No
interactions were observed to be significant. Both models showed good fit (P>.15) and classification.
(Fig. 1) Conclusions: Serum cFAS content is associated with an increased risk of CLTI but not DM. Each
unit increase in cFAS content increases the odds of CLTI by 16%. Future analysis with a larger sample and
statistical bootstrapping will determine whether cFAS content is a clinically-relevant biomarker of CLTI
severity in patients with advanced atherosclerosis.
840
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy: N/A
Professional Practice Gap: N/A
ZEB2 Regulates Activation And Exhaustion Programming Of CD8+ T Cells In Atherosclerosis
Dawn Fernandez, Nicolas F Fernandez, adeeb Rahman, Christopher Hill, Mount Sinai, New York, NY;
Roza Shamailova, Icahn Sch of Med, New York, NY; Seunghee Kim-schulze, J Mocco, Peter Faries, Miriam
Merad, Chiara Giannarelli, Mount Sinai, New York, NY
T cells are among the most prevalent immune cells found in human atherosclerotic lesions, yet their role
remains obscure. In previous single-cell immuno-phenotyping studies we found that CD8+ T cells of
carotid atherosclerotic plaques display a spectrum of functionally heterogeneous states that vary based
on differentiation, activation, and exhaustion. Furthermore, CD8+ T cell profiles varied between patients
without (Asymptomatic, ASYM) and with (Symptomatic, SYM) recent cardiovascular (CV) events (i.e.
transient ischemic attack and stroke), suggesting that T cells might contribute to adverse outcomes. The
transcriptional regulator ZEB2 has dual roles in T cell differentiation and cardiovascular disease, as
genome wide association studies have reported ZEB2 polymorphisms as independent risk alleles for
coronary artery disease and myocardial infarction. Our independent analysis identified ZEB2 as a key
driver of CD8+ T cell alterations in atherosclerotic lesions. We found that ZEB2 was differentially
regulated between patient types, with ASYM patients expressing higher ZEB2 and GZMB levels
compared to SYM (Fig.1A). ZEB2high CD8+ T cells upregulated genes involved in cytotoxic functions, and
conversely ZEB2low CD8+ T cells upregulated PD-1 Signaling in T cell exhaustion (Fig.1B.). To probe the
mechanistic implications of ZEB2, we performed in vitro chronic stimulation assays of human primary
CD8+ T cells using depleted of ZEB2 using CRISPR/CAS9. ZEB2 knockout cells had reduced cytotoxic
function and had a dampened activation state upon acute stimulation. Persistent stimulation induce
exhaustion in these cells showed that ZEB2 knockout increased PD-1 expression levels, a protein marker
that is critical for T cell exhaustion (Fig.1C). Collectively these experiments suggest that ZEB2 may
contribute to the regulation of T cell activation and exhaustion states which is different between clinical
phenotypes.
693
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
Mast cell phenotype and secreting mediators could be detected in the plasma of patients with carotid
atherosclerotic plaque and predict the clinical outcome of the patient.
Professional Practice Gap:
The aim of this project is to associate mast cell activation with particular processes in human
atherosclerotic plaques and patient clinical data in a mean of associating the mast cell phenotype with
patient clinical outcome.
The Role Of Mast Cells In Atherosclerotic Plaque Calcification
Nikolaos-Taxiarchis Skenteris, Malin Kronqvist, Mariette Lengquist, Karolinska Inst, Solna, Sweden; Ulf
Hedin, Karolinska Inst, Stockholm, Sweden; Erik L Biessen, Maastricht Univ Medical Ctr, Maastricht,
Netherlands; Ljubica Matic, Karolinska Inst, Stockholm, Sweden
Background: Vascular calcification is a key feature of atherosclerosis and has been associated with
major adverse cardiovascular events. Unstable carotid atherosclerotic plaques cause stroke and lesions
from those patients are abundant with activated mast cells at the sites of rupture. Recent data from our
group showed a statistically significant upregulation of activated mast cells in low calcified whereas
resting mast cells were upregulated in high calcified plaques, indicating that mast cells fractions may
associate with various aspects of plaque pathology. Hypothesis: Our hypothesis is that mast cell
fractions associate with key features of plaque vulnerability such as calcification, intraplaque
hemorrhage and other immune cell fractions. Resources: Biobank of Karolinska Endarterectomies (BiKE)
prospectively enrolls patients (n=1300) treated for carotid atherosclerosis in Stockholm, comprising
BioBank with paraffin-embedded plaque tissues for histology, ImageBank with quantified diagnostic CT
images using VascuCap software and DataBank of 100 clinical variables as well as transcriptomics and
proteomics large-scale datasets. Results: Histological stainings of plaque tissue microarrays confirmed
the presence of mast cells in atheromatous lesions and revealed that mast cells were systematically
found in Perls+ regions. The average total number of mast cells per mm2 per patient correlated
negatively with the calcification content. In addition, immunohistochemical analysis demonstrated that
mast cells correlate positively with CD3+ cells while they did not correlate with markers of other
immune cells. By stratifying the results according to patient symptoms, we found that activated mast
cells were elevated in both symptomatic and asymptomatic patients and increased with severe
symptoms of plaque instability. However, patients’ medication does not impact mast cell
regulation. Conclusions: Systematic enumeration of mast cell fractions in human plaques indicates that
activated mast cells associate with increased vulnerability, both when it comes to clinical patient
symptoms and morphological plaque features.
792
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
Through the use of low cost genotyping, we are able to identify a subset of the population at higher risk
for AAA and who may benefit from screening beyond the current USPSTF recommendations.
Professional Practice Gap:
Currently, AAA screening guidelines as presented by the USPSTF recommend a single ultrasound
screening for AAA in male individuals over the age of 65. These recommendations miss a large portion of
the population at risk for AAA.
Polygenic Risk Score Identifies Patients At Increased Risk For Abdominal Aortic Aneurysm And May
Benefit From Ultrasound Screening
Derek Klarin, Univ of Florida Coll of Med, Gainesville, FL; Ozan Dikilitas, Mayo Clinic, Rochester, MN;
Brooke Wolford, Univeristy of Michigan, Ann Arbor, MI; Michael Levin, Univ of Pennsylvania Sch of Med,
Philadelphia, PA; Ishan Paranjpe, Mount Sinai Sch of Med, New York, NY; Renae Judy, Univ of
Pennsylvania Sch of Med, Philadelphia, PA; Julie Lynch, Dept of Veterans Affairs, Salt Lake City Health
Care System, Salt Lake City, UT; Themistocles L Assimes, Stanford Univ Sch of Med, Palo Alto, CA; Yan
Sun, Emory Univ, Atlanta, GA; Daniel Rader, Univ of Pennsylvania Sch of Med, Philadelphia, PA; Peter W
Wilson, Emory Univ Sch of Med, Atlanta, GA; Salvatore Scali, Scott Berceli, Univ of Florida Coll of Med,
Gainesville, FL; Sekar Kathiresan, Verve Therapeutics, Cambridge, MA; Pradeep Natarajan,
Massachusetts General Hosp, Boston, MA; Girish Nadkarni, Mount Sinai Sch of Med, New York, NY;
Cristen Willer, Univ of Michigan, Ann Arbor, MI; Iftikhar Kullo, Mayo Clinic, Rochester, MN; Scott M
Damrauer, Universtiy of Pennsylvania Sch of Med, Philadelphia, PA; Philip Tsao, Stanford Univ Sch of
Med, Palo Alto, CA
Introduction: Abdominal aortic aneurysm (AAA) is a significant heritable cause of cardiovascular-related
mortality, yet published GWAS have only identified 10 genome-wide significant (P<5x10-8) risk loci to
date. In addition, current AAA screening recommendations remain limited to men age 65-75 with a
history of smoking. Hypothesis: Genetic variants affecting multiple biological pathways are associated
with AAA risk and may help identify asymptomatic individuals at higher risk for disease. Methods: Using
EHR data, we identified individuals with and without clinical AAA in Million Veteran Program (MVP)
participants. Individuals were genotyped on a customized Affymetrix array, and we tested 18 million
genotyped and imputed DNA variants for association with AAA using logistic regression models adjusting
for age, sex and population structure. We then performed replication in external datasets and set a
P<5x10-8 for statistical significance. In downstream analyses, we tested and validated a series of AAA
polygenic risk scores (PRS) and assessed the associated AAA risk per standard deviation (SD) increase in
PRS using prevalent data from an independent set of MVP participants (1,656 AAA cases; 44,908
controls). We set P < 0.05 for statistical significance. Results: We identified 7,642 AAA cases and 172,172
controls. Following replication, we identified 14 novel AAA loci implicating known risk factors including
lipids (LPA, PCSK9) and smoking (CHRNA3). We generated a 29 variant PRS and observed that a 1 SD
increase in the AAA PRS was associated with a 32% increased risk of AAA (OR = 1.32, PPRS =1.7x10-34).
Men in MVP with the 5% highest PRS and over 50 years of age had a disease prevalence of 7.8%
(142/1680), higher than that observed in AAA screening trials informing current guidelines. Conclusions:
Here, we identify novel AAA genetic associations with therapeutic implications and identify a subset of
the population at significantly increased risk of AAA. Our data suggest that extending current screening
guidelines to include testing for those with high polygenic AAA risk would significantly increase the yield
of current screening.
755
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
The results of this study allow us to better understand vascular inflammation driving AAA formation and
potential intervene medically.
Professional Practice Gap:
There is a lack of proven medical therapy to prevent AAA growth. This abstract demonstrates that
macrophage epigenetic modifications regulate inflammation in AAAs and highlights a new mechanism to
target
Epigenetic Modifications Influence Macrophage-mediated Inflammation Inabdominal Aortic
Aneurysms
Frank Michael Davis, Aaron Dendekker, Amrita Joshi, Sonya Wolf, Christopher Audu, William J Melvin,
UNIVERSITY OF MICHIGAN, Ann Arbor, MI; Hong Lu, UNIV KENTUCKY, Lexington, KY; Alan Daugherty,
UNIVERSITY OF KENTUCKY, Lexington, KY; Johann Gudjonsson, UNIVERSITY OF MICHIGAN, Ann Arbor,
MI; Katherine A Gallagher, UNIVERSITY MICHIGAN, Ann Arbor, MI
Introduction: Abdominal aortic aneurysms (AAA) are characterized by inflammatory macrophage (Mφ)
infiltration and pathological vascular remodeling. The mechanisms regulating Mφ polarization during
AAA development remain unknown. There is increasing evidence that epigenetic enzymes, specifically
the histone demethylase JMJD3, direct Mφ polarization. The purpose of this study was to investigate if
JMJD3-mediated epigenetic modifications regulate Mφ phenotype and hence inflammation during AAA
formation.
Methods: Single-cell sequencing was conducted on human AAA and control tissue samples. For our
murine model, male C57BL/6 mice (n=60) were intraperitoneally injected with an AAV encoding a mouse
PCSK9 gain-of-function mutation (D377Y). Mice were then fed a saturated fat-enriched diet and infused
with AngII (1 µg/kg/min) or saline for 4 weeks. AAA maximum diameters were quantified and Mφs
(CD11b+,CD3-,CD19-,Ly6G-) were sorted. Jmjd3 and NFκB mediated inflammatory gene expression was
examined. Chromatin immunoprecipitation (ChIP) was used to evaluate histone 3 lysine 27
trimethylation (H3K27me3). Statistical significance was determined using Student’s t-test or ANOVA.
Results: Single-cell transcriptome analysis of human AAAs demonstrated a significant upregulation
of JMJD3 and inflammatory cytokines in myeloid subsets compared to controls. Congruently, Mφs from
AngII-induced AAAs displayed increased Jmjd3 and NFκB-mediated cytokine expression (i.e. IL-1β, IL-12)
(p<0.05). Histone methylation was evaluated by ChIP in AngII-induced AAA Mφs and demonstrated
decreased levels of the Jmjd3-mediated repressive histone methylation mark, H3K27me3, on
inflammatory gene promoters. In vitro inhibition of JMJD3 using siRNA significantly reduced Mφ-
mediated inflammatory cytokine expression (p<0.05). Finally, pharmacological inhibition of JMJD3 (GSK-
J4) in the AngII-induced murine model resulted in a significant reduction in AngII-induced AAA formation
and inflammatory cytokines.
Conclusions: These results suggest an important role for JMJD3 in regulating Mφ-mediated
inflammation in human and murine AAAs as well as identify a potential target for the treatment of
chronic inflammation.
759
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
Use of a dual chamber stent to improve organ recovery and reduce ischemic injury.
Professional Practice Gap:
Perfusion of abdominal organs during organ recovery.
The Perfuse Dual Chamber Stent Improves Donor Organ Recovery In A Porcine Model
Bryan W. Tillman, Catherine Go, Brian Frenz, UNIV OF PITTSBURGH MED CTR, Pittsburgh, PA; Moataz
Elsisy, UNIV OF PITTSBURGH, Pittsburgh, PA; Youngjae Chun, UNIVERSITY OF PITTSBURGH, Pittsburgh,
PA
Background: Amidst a critical shortage of organs for transplantation, ischemic injury from malperfusion
during the agonal period remains a prohibitive barrier. We hypothesized that a dual chamber stent graft
could isolate visceral perfusion from the agonal systemic circulation, while respecting the ethics of organ
donation. Methods: A retrievable dual chamber stent graft was welded from nitinol and covered with
polymer. A central lumen maintained aortic flow, with an outer visceral chamber perfused by an
oxygenator. Anesthetized pigs were assigned to either control (n=7) or the dual chamber stent (n=6). A
one hour agonal phase of hypoxia (saturations < 60-70%) and hypotension (MAP < 25) was simulated
both medically and with partial balloon occlusion. The Perfuse stent visceral flow totaled 500 ml/min
during the agonal phase followed by stent recapture and resuscitation to an endpoint of 2
days. Results: Study groups had comparable agonal O2 saturations, HR and MAP. Cardiac output and
right ventricular end diastolic volume did not change during stent graft deployment. Compared to the
low pO2 of controls (48 mm Hg) and systemic stent animals (49 mm Hg), the visceral pO2 averaged 413
mm Hg and visceral flow was significantly higher in stent animals. 5/7 controls were euthanized from
acute renal failure and volume overload while all stent animals survived without renal impairment.
Transaminases were between 1.8 to 3 fold elevated in control as compared to stented
animals. Conclusions: During a simulation of the agonal period, a dual chamber stent provided
endovascular separation and marked improvement in perfusion and organ outcome. This was
accomplished without significant impact on cardiac function, respecting current ethical considerations
of the DCD donor. The ability to separate the perfusion of the abdominal organs from the agonal
systemic circulation without the need for open surgery might significantly improve the availability of
donor organs for transplant.
795
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
Trauma TEVAR is associated with LV mass increase, development of hypertension, and off-target
accelerated expansile ascending aortic remodeling. Our findings call for better follow-up of young
patients receiving stent-grafts and for improved endovascular devices capable of preserving optimal
ventricular-arterial coupling.
Professional Practice Gap:
Thoracic endovascular aortic repair (TEVAR) is first-line therapy for blunt thoracic aortic trauma, but its
effects on the left ventricle (LV) and off-target aortic zones are poorly understood. We have measured
changes in LV mass, LV mass index, and diameters and lengths of the ascending thoracic aorta in trauma
TEVAR patients, and compared these patients with similarly-aged control subjects without aortic repair
evaluated at similar follow-ups.
Endovascular Repair Of Blunt Thoracic Aortic Trauma Is Associated With Increased Left Ventricular
Mass, Hypertension, And Off-target Aortic Remodeling
Alexey Kamenskiy, Univ of Nebraska Omaha, Omaha, NE; Paul Aylward, Univ of Nebraska Medical Cent,
Omaha, NE; Anastasia Desyatova, Univ of Nebraska Omaha, Omaha, NE; Matthew DeVries, Christopher
Wichman, Univ of Nebraska Medical Ctr, Omaha, NE; Jason N Mactaggart, Univ of Nebraska Medical
Cent, Omaha, NE
Introduction: Aortic elasticity creates a cushion that protects the heart from pressure injury, and a recoil
that helps perfuse the coronary arteries. Thoracic endovascular aortic repair (TEVAR) has become first-
line therapy for many aortic pathologies including trauma, but stent-grafts stiffen the aorta and likely
increase left ventricular (LV) afterload. We hypothesized that trauma TEVAR is associated with LV mass
increase and adverse off-target aortic remodeling.
Methods: CTA scans of 20 trauma TEVAR patients (17M/3F) at baseline (age 34.9±18.5[11.4-71.5] years)
and 5.1±3.1[1.1-12.3] years after repair were used to measure changes in LV mass, LV mass index
(LVMi), and diameters and lengths of the ascending thoracic aorta (ATA). Measurements were
compared with similarly-aged control patients without aortic repair (21M/21F) evaluated at similar
follow-ups.
Results: LV mass and LVMi of TEVAR patients increased from 138.5±39.6g and 72.35±15.17g/m2 to
173.5±50.1g and 85.48±18.34g/m2 at the rate of 10.03±12.79g/year and 6.25±10.28g/m2/year, while in
control patients LV characteristics did not change (Figure). ATA diameters of TEVAR patients increased at
a rate of 0.60±0.80mm/year, which was 2.4-fold faster than in controls. ATA length in both TEVAR and
control patients increased at 0.58mm/year. Half of TEVAR patients had hypertension at follow-up
compared to only 5% at baseline.
Conclusions: TEVAR is associated with LV mass increase, development of hypertension, and accelerated
expansile remodeling of the ascending aorta. While younger trauma patients may adapt to these effects,
these changes may be even more important in older patients with other aortic pathologies and
diminished baseline cardiac function.
809
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
There currently exists no successful medical management of abdominal aortic aneurysms. This research
helps to identify potential therapeutic targets and mechanisms in the treatment of this disease.
Professional Practice Gap:
This research addresses potential medical therapies in the treatment of AAA as currently operative
intervention remains the only definitive treatment.
Maresin 1 Attenuates Murine Abdominal Aortic Aneurysms Via Vascular Smooth Muscle Cell
Dependent TGF-β Signaling
Craig T Elder, Univ of Florida, Gainesville, FL; Gang Su, Gainesville, FL; Amanda Filiberto, Univ of Florida
Coll of Me, GAINESVILLE, FL; Guanyi Lu, Univ of Florida Coll of Me, GAINESVILLE, FL, Gainesville, FL;
Zhihua Jiang, UNIVERSITY OF FLORIDA COM, Gainesville, FL; Ashish Sharma, UNIVERSITY OF FLORIDA,
Gainesville, FL; Gilbert R UPCHURCH Jr., Univ of FL Dept of Surgery, Gainesville, FL
Introduction: The endogenous pro-resolving lipid mediator Maresin 1 (MaR1) is derived from the ω-3
polyunsaturated fatty acid docosahexanoic acid and is involved in the resolution phase of inflammation.
Specifically, MaR1 has been shown to attenuate inflammatory signaling in smooth muscle cells. It was
hypothesized that exogenous administration of MaR1 would attenuate AAA growth via smooth muscle
cell dependent TGF-β signaling.
Methods: AAAs were induced in C57BL/6 wild-type mice (n=9 elastase + MaR1 [4ng/g] or n=10 elastase
+ vehicle) using an established topical elastase AAA model. Mice were treated with MaR1 or vehicle via
IP injection on days 7, 9, 11, and 13 post AAA induction. Abdominal aortas were harvested on day 14 for
phenotypic evaluation of aortic diameter. Histological analysis of smooth muscle actin (n=3-4/group)
and western blot of TGF-β1 expression (n=6-7/group) were performed. Additionally, AAAs were induced
in smooth muscle cell specific TGF-β2 receptor knockout mice and treated with MaR1 (n=7) vs vehicle
(n=10) and harvested on day 14 for phenotypic evaluation of aortic diameter. Groups were analyzed
using one-way ANOVA with post hoc Tukey’s test and data presented as mean ± SEM.
Results: MaR1 treatment significantly attenuated AAA growth compared to vehicle (121.4% ± 9.4 vs.
165.3% ± 9.4; p<0.01). A significant increase in aortic wall smooth muscle cell actin was identified in
MaR1 treated mice compared to vehicle (27.1% ± 3.4 vs 14.2% ± 2.4; p=0.03). TGF-β1 expression was
also significantly higher in MaR1 treated mice compared to vehicle (2.95 x106 ± 11.1 x105 vs 1.63 x106 ±
2.6 x105 densitometry units; p=0.02). Finally, smooth muscle cell specific TGF-β2 receptor knockout mice
showed no difference in AAA diameter in MaR1 treated mice compared to vehicle (133.9% ± 8.0 vs
123.8 % ± 7.9; p=0.4).
Conclusion: These results demonstrate that MaR1, important in the resolution of inflammation,
attenuates murine AAA growth, preserves aortic wall smooth muscle cell actin, and increases TGF-β1
expression. This attenuation in AAA growth is lost in smooth muscle cell specific TGF-β2 receptor
knockout mice further suggesting that MaR1 modulates TGF-β signaling in aortic smooth muscle cells.
787
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
Develop new diagnostic and treatment modalities for AAA
Professional Practice Gap:
Experimental AAA models are subacute.
A Theranostic Role For Ccr2 In Rodent Abdominal Aortic Aneurysm Development And Rupture
Sergio Sastriques Dunlop, Washington Univ Sch Med, Saint Louis, MO; Batool Arif, Sean English,
Washington Univ Sch Med, SAINT LOUIS, MO
Introduction: Abdominal aortic aneurysm (AAA) is common in the aging population, and its rupture
carries a high mortality risk. AAA is characterized by mononuclear phagocyte destructive aortic extra
cellular matrix remodeling. Monocyte chemoattractant protein (MCP-1)/C-C chemokine receptor type 2
(CCR2) axis plays an important role in AAA development. We sought to evaluate if increased murine
AAA 64Cu-DOTA-ECL1i uptake, a radiolabeled CCR2 binding peptide, by positron emission tomography
(PET), is predictive of rupture, as well as if CCR2 inhibition prevents rupture.
Methods: Sprague-Dawley rats and C57BL/6 (wild type, WT) mice underwent intraluminal aortic
exposure to porcine pancreatic elastase (PPE). Four groups were considered: 1. Active PPE (AAA), 2.
Heat-inactivated PPE (Sham), 3. APPE + β-aminopropionitrile (BAPN, a lysyl oxidase inhibitor to stimulate
rupture) (RAAA), and 4. APPE + BAPN + RS-504393 (CCR2 inhibitor, CCR2i). WT and CCR2-/- mice were
also exposed to Angiotensin II. 64Cu-DOTA-ECL1i PET imaging was performed at 7 and 14 days post AAA
induction.
Results: RATS: AAA demonstrated significantly greater 64Cu-DOTA-ECL1i uptake by PET and
autoradiaography compared to Sham (P<0.05). RAAA that subsequently ruptured demonstrated
significantly greater uptake compared to those animals that did not rupture (P<0.001). RAAA and CCR2i
rupture rates were 56% and 0%, with mean aortic diameter % increases of 353 ± 43% and 171 ± 26 %,
respectively (P<0.0001). CCR2i PET uptake decreased by 60%. MICE: WT, CCR2i and CCR2 -/- rupture
rates were 53, 33 and 6%, with mean aortic diameter % increases of 330 ± 62, 191 ± 67 and 101 ± 16%,
respectively (P<0.01). Quantification of tracer uptake by PET was significantly greater in AAA compared
to Sham and CCR2-/- (p<0.0001).
Conclusion: CCR2 targeted PET imaging demonstrated inflammation associated with rodent AAA
development. Increased radiotracer uptake by AAA that subsequently ruptured may aid in assessing
AAA rupture potential. CCR2 inhibition with our model prevents AAA rupture and inhibits AAA
associated inflammation, suggesting a theranostic role for CCR2 in the management of AAA patients.
762
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
This study demonstrated that BTK inhibitor ibrutinib, FDA-approved drug, attenuated the formation of,
and further progression of established, experimental AAAs in two mechanistically distinct but
complementary AAA model systems. Our results suggest that pharmacological inhibition of BTK may
represent a novel translational strategy for AAA disease suppression.
Professional Practice Gap:
Macrophages are critical for abdominal aortic aneurysm (AAA) pathogenesis. Bruton’s tyrosine kinase
(BTK) contributes to macrophage-driven diseases s. It is not known whether BTK plays a role in AAA
disease. Additionally, currently there is no approved drugs for treating aortic aneurysm disease.
Pharmacological Inhibition Of Bruton's Tyrosine Kinase Attenuates Experimental Abdominal Aortic
Aneurysms
Baohui Xu, STANFORD UNIVERSITY SCHOOL OF MEDIC, Stanford, CA; yankui li, Stanford Univ Sch of
Med, Stanford, CA; Gang Li, STANFORD UNIVERSITY SCHOOL OF MEDIC, Stanford, CA; Yingbin Ge,
Nanjing Medical Univ, Nanjing, China; Jia Guo, Stanford Univ Sch of Med, Stanford, CA; Weirong Fang,
China Pharmaceutic Univ, Nanjing, China; Wei Wang, Xiangya Hosp, Central South Univ, Chnagsha,
China; Fanru Shen, STANFORD UNIVERSITY SCHOOL OF MEDIC, Stanford, CA; Takahiro Shoji, Vascular
Surgery, Stanford, Stanford, CA; Toru Ikezoe, Xiaoya Zheng, STANFORD UNIVERSITY SCHOOL OF MEDIC,
Stanford, CA; Sihai Zhao, Stanford Univ Sch of Med, Stanford, CA; Xiaofeng Chen, TAIZHOU HOSPITAL,
Taizhou; Masaaki MIYATA, KAGOSHIMA CITY HOSPITAL, Kagoshima, Japan; Alan Daugherty, UNIVERSITY
OF KENTUCKY, Lexington, KY; Hong Lu, Univ of Kentuky Sch of Med, Lexington, KY; Ronald L DALMAN,
STANFORD UNIVERSITY MEDICAL CE, Stanford, CA
Objective: Macrophages are critical for abdominal aortic aneurysm (AAA) pathogenesis. Bruton’s
tyrosine kinase (BTK) contributes to macrophage driven diseases such as atherosclerosis and ischemic
stroke by modulating proinflammatory macrophage activation. It is not known whether BTK plays a role
in AAA disease. This study was to investigate the influence of the FDA-approved BTK inhibitor ibrutinib
on the formation and progression of experimental AAAs. Methods: AAAs were generated via intra-aortic
elastase infusion in male normolipidemic C57BL/6J mice or subcutaneous angiotensin II infusion in male
hyperlipidemic C57BL/6J mice (inoculated an adeno-associated virus expressing a gain-of-functional
mutation of PCSK9 and fed high fat diet). Ibrutinib administration (30 mg/kg) was initiated one day prior
to, or indicated days after, AAA creation. Influence on AAAs was evaluated via ultrasonography and
histology. The effect of ibrutinib on macrophage mediator mRNAs was assessed via quantitative RT-PCR
assay. Results: By ultrasonography, ibrutinib pretreatment remarkably attenuated elastase infusion-
induced aortic enlargement, in association with delayed onset and reduced incidence of AAAs, as
compared to vehicle treatment. In established AAA mice, ibrutinib treatment limited further aneurysmal
enlargement. On histology, ibrutinib treatment preserved medial elastin and smooth muscle cell
cellularity and reduced mural macrophage, lymphocyte and neocapillary density. Ibrutinib
administration also reduced the incidence and severity of angiotensin II-induced AAAs in hyperlipidemic
mice. Additionally, BTK treatment attenuated CCL2 and TNF-alpha mRNA expression, but accentuated IL-
10 mRNA, in classically or alternatively activated macrophages. Conclusion: BTK mediates AAA
progression in two complementary experimental AAA models. Pharmacological inhibition of BTK may
represent a novel translational strategy for AAA disease suppression.
694
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy: N/A
Professional Practice Gap: N/A
Landscape Of Human Secretome In Abdominal Aortic Aneurysm Profiled By Single-cell RNA
Sequencing.
Bhama Ramkhelawon, NYU MEDICAL CENTER, New York, NY; Tarik Hadi, NYU Langone Medical Ctr,
New York, NY; Michele Silvestro, NYU Sch of Med, New York, NY
Introduction: Abdominal aortic aneurysms (AAA) is a lethal vascular disease upon aortic rupture.
Evidence suggests pathological cellular crosstalks that fuel extracellular matrix (ECM) degradation in
AAA. However, the comprehensive ligand-receptor intra-aortic signaling networks that persists in
human AAA is undefined. Hypothesis: Heterogeneous cell-cell communication via specific ligand-
receptor signaling contributes to human AAA.
Methods: Human AAA, control organ donor, mouse control and AAA modeled by angiotensin II infusion,
samples were processed to for single cell RNA Sequencing (scRNA seq) libraries. Reads were aligned to
reference genome and analyzed for significant differential expression profiles. Pathway analysis were
used to outline biologically relevant networks. Human secretome of ligand-receptor interactions were
applied to human and mouse scRNA seq libraries. Results: Analysis of human scRNA seq showed 16 cell
clusters (9 immune and 7 vascular) expressing 923 significantly different genes in AAA. Secretome
analysis revealed a highly connected network of 262 transcripts enriched in ECM remodeling,
metabolites, leukocyte migration, Wnt signaling and cytokines. Comparison between mouse and human
showed that only macrophages displayed similar gene profile to that of human AAA. Amongst the 2005
increased genes in murine macrophages, about 10% was common in human, 20 of which were in the
secretome analysis. We observed an emergence of a novel population of IL7 expressing B cells only in
human AAA. CD4 and CD8, but not naïve B cells and dendritic cells expressed
elevated IL7 receptor suggesting active recruitment and differentiation of lymphoid progenitors of T cell
lineage in AAA sac. Further studies are needed to delineate the role of IL7+ B cells and its crosstalk with
T cells in AAA. Circulating IL7 in serum might be of reliable predictive value to predict AAA. Conclusion:
We uncovered novel insights in the landscape of intercellular crosstalk highlighting the similarity
between murine and human transmural macrophages and the distinct role of lymphoid signaling in
human AAA. These candidates that directly stem from AAA could be used as valuable biomarkers much
needed to predict clinical AAA.
788
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
This research represents a significant step in overcoming prior major limitations for patient-specific
induced pluripotent stem cell-derived endothelial cells (iPSC-ECs) regarding longevity and function in
culture that have been a barrier for clinical applications. These findings open the door for future
investigations into clinical applications for iPSC-ECs such as therapeutic injections for critical limb
ischemia, or for engineering patient-specific bypass conduits for peripheral arterial disease.
Professional Practice Gap:
To date, clinical applications for patient-specific stem cell-derived endothelial cells have been limited by
poor longevity in culture and loss of mature cellular phenotype and function due to premature
senescence.
Autophagy Remodels Mitochondria During Differentiation And Enhances Longevity Through Ulk1
Kinase Signaling Of Induced Pluripotent Stem Cell-derived Endothelial Cells
Katherine E Hekman, Kyle Koss, David Z Ivancic, Northwestern Univ, Chicago, IL; Congcong He, Chicago,
IL; Jason A Wertheim, Northwestern Univ, Chicago, IL
Background Stem cells hold great promise for the future of peripheral arterial disease therapy, and
patient-specific stem cell-derived endothelial cells (ECs) present major advantages as a therapeutic
modality. However, ECs derived from patient-specific induced pluripotent stem cells (iPSCs) suffer from
premature replicative senescence with rapid loss of mature cellular function in culture currently limiting
clinical applications. In the present study we characterize autophagy during differentiation and
maintenance of iPSC-ECs as a target to modulate longevity.
Methods and Results Autophagy, measured by autophagosome number in TEM samples and by
quantitation of the expression of microtubule-associated proteins 1A/1B light chain 3B (LC3),
demonstrates multiple significant increases during differentiation suggesting an active role in
remodeling. Mitotracker staining shows increasing mitochondrial membrane potential and TEM
demonstrates that maturation continues through serial passages; however, these stagnate, coinciding
with decreased cellular proliferation as measured with Edu labeling. Plant-based small molecule Rg2
significantly improved proliferative capacity of iPSC-ECs over multiple passages (Figure 1). This benefit
was not observed in cells treated with either rapamycin or the autophagy inducer ML246, or with
mitophagy inducers Antimycin A or oligomycin. The benefit of Rg2 was blocked by treatment with ULK1
kinase inhibitor SBI-0206965 in a dose-dependent manner.
Conclusion In summary, stimulating mTOR-independent ULK1-mediated autophagy with Rg2 improved
the longevity of iPSC-ECs in culture. This represents a novel approach to overcome a major barrier to the
use of iPSC-ECs for clinical applications.
828
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
We attempt to increase stem cell's inherent therapeutic potential with a novel vector to increase cell's
homing capabilities and overall potency.
Professional Practice Gap:
Patients with critical limb ischemia often have progressed on current medical therapies and have either
failure surgical interventions or cannot undergo a surgical procedure. These no-option patients currently
have no other option other than amputation, our pre-clinical study attempts to discovery new
therapeutics for CLI patients in hopes of future limb salvage
Downregulation Of Inflammation And A Cascade Of Pro-angiogenic Signals Mediate The Beneficial
Effects Of Gene-modified Stem Cell Therapy In Hindlimb Ischemia
Hallie J Quiroz, Univ of Miami, Miami, FL; Hongwei Shao, Yan Li, Samantha F Valencia, Punam Parikh,
Univ of Miami, Miami, FL; Yulexi Ortiz, Univ of Miami Miller Sch of Med, Miami, FL; Roberta Lassance-
Soares, Univ of Miami, Miami, FL; Zhao-Jun Liu, Univ of Miami Miller, Miami, FL; Omaida Caridad
Velazquez, Univ of Miami Miller Sch of Med, Miami, FL
Introduction: Novel therapies to afford limb salvage in patients with imminent amputation due to
critical limb ischemia are needed. We have previously shown E-selectin, a cell adhesion molecule, as an
essential participant in neovascularization. Thus, we hypothesized E-selectin supercharged
mesenchymal stem cells (E-selectin+/MSC) would augment limb reperfusion, tissue regeneration, and
functionality.
Methods: C57Bl6 mice underwent femoral artery ligation and received either vehicle (PBS, n=9) or
syngeneic donor MSCs, transduced ex vivo to express either GFP+ (control, n=20) or E-selectin-
GFP+ (treatment, n=18). Laser doppler imaging (LDI), confocal laser microscopy, and treadmill
exhaustion test were utilized to determine neovascularization and limb function. Extent of ischemic
skeletal muscle atrophy (mean myocyte size,µm²) was assessed via leg muscle histology. RT2 Profiler PCR
Array analysis of 84 genes involved in angiogenesis assessed therapeutic mechanism of action. Student’s
t-test or ANOVA was utilized to compare means and significance set at p<0.05.
Results: Compared with GFP+/MSC and PBS, treatment with E-selectin-GFP+/MSC increased ischemic leg
LDI reperfusion (55% vs. 39% vs. 24%, p<0.001), ischemic mouse footpad vessel density (23% vs. 14% vs.
14%, p<0.01) and treadmill distance traversed (162m vs. 111m vs. 110m, p<0.01). The ischemic limb in
mice treated with E-selectin-GFP+/MSC were less atrophic than controls (793µm² vs. 556µm² vs.
546µm², p<0.001). Seven pro-angiogenic genes were upregulated in E-selectin-GFP+/MSC treated
ischemic leg tissue while tumor necrosis factor (TNF) was downregulated, when compared with
GFP+/MSC treated tissues.
Conclusion: This innovative E-selectin supercharged stem cell therapy confers increased limb
reperfusion, function, and decreased atrophy, likely via upregulation of pro-angiogenic cytokines and
downregulated TNF.
717
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy: N/A
Professional Practice Gap: N/A
Prediction Of Walking Performance After Revascularization Of Arteries Supplying The Lower
Extremities Of Claudicating Patients With Peripheral Artery Disease
Shuai Li, Jonathan Robert Thompson, Univ of Nebraska Med Ctr, Omaha, NE; Sara A Myers, Univ of
Nebraska at Omaha, Omaha, NE; Julian K Kim, UNIV NEBRASKA MEDICAL CENTER, Omaha, NE;
Panagiotis Koutakis, Florida State Univ, Tallahassee, FL; Mark Williams, Creighton Cardiac Ctr, Omaha,
NE; Zhen Zhu, Univ of Nebraska Med Ctr, Omaha, NE; Molly Schieber, Univ of Nebraska Medical Cent,
Omaha, NE; Timothy Lackner, Gregory Willcockson, Christina Shields, Katyarina Brunette, Holly
Despiegelaere, Univ of Nebraska Med Ctr, Omaha, NE; Iraklis I Pipinos, UNIV NEBRASKA MEDICAL CTR,
Omaha, NE; George Casale, Univ of Nebraska Medical Cent, Omaha, NE
Rationale and Objectives: Peripheral Artery Disease (PAD) affects over 200 million patients globally.
Revascularization is a mainline treatment for PAD, however, change in walking performance and
Quality-of-Life (QoL) varies substantially among patients, ranging from no to marked improvement. We
evaluated biochemical and histopathological measurements of calf muscle and leg hemodynamics,
before revascularization, as potential predictors of change in walking performance and QoL, in response
to revascularization.
Methods and Results: We measured, at baseline and 6-months post-revascularization, Ankle-Brachial
Index (ABI), Systolic Pedal Pressure at 15 seconds after Post-occlusive Reactive Hyperemia (SPP15sec),
Six-Minute Walking Distance (SMWD), Initial Claudication Time (ICT), Peak Walking Time (PWT) and
Quality of Life (QoL) survey scores of forty claudicating PAD patients. In addition, we determined a)
activities of mitochondrial Citrate Synthase (CS), Complexes 1-4 and MnSOD, b) myofiber morphology
and oxidative damage (carbonyl adducts), and c) fibrosis, in calf muscle. Prediction of walking
performance and QoL was evaluated by Spearman Rho and ROC curve. Baseline SPP15sec and change in
SMWD were most strongly correlated (Rs = -0.72 at P < 0.0001). A ROC curve identified patients whose
SMWD improved > 20 m at a baseline SPP15sec cutoff of 50 mmHg, with sensitivity of 89% and
specificity of 100%. In addition, we detected significant improvements (P < 0.05) of hemodynamics (ABI,
SPP15sec), walking performance (SMWD, ICT and PWT), QoL survey scores, and calf muscle pathology
(CS, Complex I and II and MnSOD activities, myofiber shape and carbonyl adducts).
Conclusions: Our study shows that revascularization improves hemodynamics, walking performance,
QoL and calf muscle pathology in claudicating PAD patients. Baseline SPP15sec (cutoff = 50 mmHg)
strongly predicts which patients will experience > 20 m improvement of SMWD.
695
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
We may opt to treat patients with claudication more aggressively
Professional Practice Gap:
Resting perfusion to muscle in PAD may cause long term effects beyond schema and may alter how
disease should be managed
Differential Expression Of Canonical Mitosis And Dna-damage Repair Pathways Characterize Muscle
Satellite Cells Affected By Pad
Ricardo Ferrari, Guangzhi Cong, Bryan Thompson, Debbie Hollingshead, Janette Lamb, Xiangdong Cui,
Ulka Sachdev, UPMC, Pittsburgh, PA
Introduction: Peripheral arterial disease (PAD) is associated with diminished muscle strength, even in
claudicants with adequate resting perfusion. Muscle Satellite Cells (MuSC) regeneration may be
deterred by PAD. We propose that MuSC in PAD have transcriptomic differences causing ischemia-
induced myopathy. Methods: Anterior tibialissegments were harvested during amputations for critical
limb ischemia from viable (proximal; P) and ischemic (distal; D) muscle. Pressure indices documented
perfusion. MuSC were isolated by antibody-mediated magnetic cell sorting. Control myocytes were
purchased. Both were measured for myocyte fusion index (MFI; 1&5d). TruSeq stranded mRNA kits
facilitated library creation for next-generation sequencing. Ingenuity Pathway Analysis (IPA) was
performed on differentially expressed genes with a false discovery rate p-value of <0.05 for pathway
enrichment. Results: MFI in PAD (N=8, P and D) MuSC was half that of controls (p<0.03; N=4, ANOVA).
Pressure indices confirmed a perfusion gradient along the limb. The # of differentially expressed genes
among PAD and controls are shown (Fig.1). MuSC in PAD vs. controls had significant differences in
canonical pathway expression including mitotic roles of polo-like kinase (PLK; z score = 7.5 distal, 13.5
proximal), and G2/M DNA damage checkpoint regulation (z score = 6.5 distal, 9.0 proximal). Genetic
expression did not differ within PAD muscle as a function of perfusion. Conclusion: Compared to
controls, MuSC in PAD have significant differences in gene expression even in better perfused muscle.
Pathways affected included those essential to mitosis and DNA-damage repair, both critical to cell
survival and differentiation. These findings suggest perfusion-independent PAD effects on muscle that
persist despite optimization. Adjunctive treatments beyond reperfusion may be required to mitigate
muscle damage from arterial insufficiency.
781
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
Through nanotechnology created sustained release of growth factors or statin drugs in the stimulation
of angiogenesis.
Professional Practice Gap:
Potential intervention in the treatment of critical limb ischemia.
Fluvastatin And Vegf Containing Resins Promote Angiogenesis And Arteriogenesis
Furqan Muqri, SUNY Upstate Medical Ctr & Syracuse VA Medical Ctr, Syracuse, NY; Dandan Guo, SUNY
Upstate Medical Univ, Syracuse, NY; Mohammed Kassem, Mary DaCosta, SUNY Upstate Medical Ctr &
Syracuse VA Medical Ctr, Syracuse, NY; David Bruch, SUNY Upstate Medical Univ, Syracuse, NY;
Kristopher Maier, SUNY Upstate Medical Ctr & Syracuse VA Medical Ctr, Syracuse, NY; Juntao Luo, SUNY
Upstate Medical Ctr, Syracuse, NY; Vivian Gahtan, SUNY Upstate Medical Ctr & Syracuse VA Medical Ctr,
Syracuse, NY
Select patients with critical limb ischemia who are not candidates for surgical revascularization
rely on angiogenesis to ameliorate symptoms. Statin drugs and vascular endothelial growth factor
(VEGF) stimulate angiogenesis. Hypothesis: resins used for local statin delivery to the ischemic
bed would increase angiogenesis and be augmented by dual delivery with VEGF.
Functionalized hydrogel resins were loaded with fluvastatin or VEGF-165A. Matrigel or
proliferation assays were performed on endothelial cells (ECs) treated with free drug, resin, or
resin bound with drugs. Free or bound resins, or a combination of both resins were injected into
male rat hindlimbs after femoral artery ligation. Doppler flow was measured pre and postoperative
day 1, 3, 7 and 14 and compared to the contralateral uninjured leg. On day 14, the ischemic
vastus and gastrocnemius muscles were stained with alpha smooth muscle actin (arteriogenesis)
and CD34 (angiogenesis). In vitro findings: statin release rate from resin was 10% per 24 hrs;
unloaded resin no EC toxicity and tubule formation was unaffected; both free and resin statin or
VEGF promoted tubule formation (p<0.05); tubule formation was the same for free statin and resin
statin; resin VEGF or VEGF plus resin statin had less tubule formation compared to its respective
free form (p<0.05); free and resin statin and VEGF equally increased EC proliferation. Doppler
blood flow studies (day 14) showed: untreated control and empty resin had an 80% blood flow
recovery; resin statin returned flow to preoperative levels (p<0.001); the combination group
increased blood flow more than statin alone (p<0.001); and resin VEGF had the greatest effect
increasing flow above preoperative levels (p<0.05). Resin statin, resin VEGF or both increased
angiogenesis and arteriogenesis (p<0.05); however, the empty resin had no effect. In conclusion,
in vitro, free fluvastatin and free VEGF were more effective than the respective resin, likely due
to slow release of the drugs from the resin. Both angiogenesis and arteriogenesis improved with
VEGF or fluvastatin bound resins. Resin VEGF alone was strongest in vivo. Single dosing of
resin VEGF effectively promoted angiogenesis, which may make this revascularization method
more feasible clinically.
726
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
Understanding of the potential role of intraluminal thrombus in aortic aneurysm development, growth,
and rupture.
Professional Practice Gap:
Understanding mechanisms that lead to aortic aneursym expansion and rupture.
Increased Neutrophil Elastase Expression in Regions of High Intraluminal Thrombus Deposition In
Human Abdominal Aortic Aneurysms.
Matthew Levesque, Lauren Bath, Max Rady Coll of Med, Winnipeg, MB, Canada; Annie Ducas, Royal
Victoria Regional Health Ctr, Barrie, ON, Canada; April Boyd, Health Sciences Ctr, Winnipeg, MB, Canada
INTRODUCTION
Abdominal aortic aneurysms (AAA) have previously been shown to rupture at sites of high intraluminal
thrombus (ILT) deposition. Neutrophil and macrophage activity have also been associated with AAA
development, but their precise roles are not known. The purpose of this study was to evaluate the
possible differential expression of macrophage chemoattractant protein 1 (MCP1), macrophage-derived
protease (CD68), neutrophil chemoattractant factor (IL8), and neutrophil elastase (MMP8) in regions of
human AAA with high and low ILT deposition.
HYPOTHESIS
We hypothesized that higher ILT deposition would be associated with increased MCP1, CD68, IL8, and
MMP8 levels compared with low ILT deposition and control aorta.
METHODS
Full thickness AAA samples were collected from 15 participants using a systematic map. For each case,
samples were taken from regions with low or high ILT. Infrarenal aortic control tissue was harvested
from 6 participants undergoing aortobifemoral bypass. A cytokine array assay was performed to
measure MMP-8, IL8, and MCP-1. CD68 immunohistochemistry was also performed.
RESULTS
AAA tissue had significantly higher levels of IL8 (145 vs 64 pg/mg protein, p=0.044), MMP8 (42566 vs
3740 pg/mg protein, p=0.015), MCP1 (473 vs 64 pg/mg protein, p=0.003), and CD68 (1.77% vs 0.246%,
p<0.001) compared to control tissue. In AAA patients, the presence of ILT was associated with
significantly higher levels of IL8 (185 vs 99 pg/mg protein, p=0.035), but not MMP8, MCP1, or CD68.
CONCLUSIONS
Human AAA tissue is associated with higher macrophage and neutrophil activity compared to control
tissue as measured by MMP8, IL8, MCP1, and CD68. Within AAA tissue, areas of high ILT deposition were
associated with higher IL8 levels, suggesting that there is increased neutrophil recruitment in high ILT
regions.
773
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
Our result indicated adult FMD is genetically different with pediatric FMD, suggesting a separate clinical
practice for both adult and pediatric FMD.
Professional Practice Gap:
We are working on the genetic difference between adult and pediatric fibromuscular dysplasia (FMD),
filling the gap of understanding the mechanism of adult FMD.
Adult and Pediatric Fibromuscular Dysplasia Are Genetically Distinct Dysplasia-associated Arterial
Diseases
Yu Wang, Dawn M. Coleman, Univ of Michigan Medical Sch, Ann Arbor, MI; Min-Lee Yang, Issaquah,
WA; Susan Blackburn, Issaquah, WA, Ann Arbor, MI; Kristina L. Hunker, Univ of Michigan Medical Sch,
Ann Arbor, MI; Heather L Gornik, Univ Hosp Cleveland, Cleveland, OH; Jun Li, Univ of Michigan, Ann
Arbor, MI; James C Stanley, 2210 Taubman Ctr, Ann Arbor, MI; Santhi Ganesh, 2210 Taubman Ctr, Ann
Arbor, MI, Ann Arbor, MI
Introduction: Fibromuscular dysplasia (FMD) is an obstructive, non-atherosclerotic arterial disease with
an estimated population prevalence of 3-5% in adults. FMD occurs primarily in women (> 90%), and its
genetic basis is poorly defined. Pediatric FMD is rare and may represent a more severe FMD phenotype,
with earlier onset hypothesized to be due to high impact deleterious genetic variants. Differences
between adult and pediatric forms of FMD have been defined for angiographic pattern and sex
distribution (with pediatric FMD afflicting males and females equally). Monogenic genes underlying
adult FMD have not yet been defined. Syndromic associations with pediatric FMD are known; for
example genetic variants predicted to cause loss-of-function of the NF1 gene have been identified in up
to 33% of pediatric FMD series. The impact of these known genes have not been systematically
evaluated in adult FMD.
Methods: We performed a whole exome sequencing of 264 unrelated adults with multifocal FMD and
annotated genetic variants in genes described in pediatric renovascular hypertension and FMD: NF1,
JAG1, RNF213, ELN, and GATA6. Pathogenicity criteria included: 1) low population frequency (gnomAD
MAF <0.01%); 2) CADD score >20. Statistical tests of enrichment of variants in FMD cases as compared
to gnomAD were performed using a Fisher's exact test.
Results and Conclusion: For all genes tested, we did not observe any significant enrichment (P>0.05).
Specifically, for the NF1 gene, we observed no loss-of-function variants, and only 3 carriers of
nonsynonymous, predicted deleterious variants (1.1%) in 264 samples. These specific variants were
observed in gnomAD in 2001/251,486 (0.8%) samples. None of these individuals carried any diagnosis of
neurofibromatosis type 1 or associated clinical features. We identified no loss-of-function variants
in JAG1, RNF213, ELN, and GATA6, nor did we find any statistical enrichment of variants predicted to be
deleterious in these genes or NF1, as compared to gnomAD and our lab controls (N=284). In summary,
adult FMD patients lacked pathogenic variations in genes underlying ~30-45% pediatric arterial dysplasia
cases. These genetic findings support that adult and pediatric FMD are likely due to disparate arterial
pathologies.
746
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
The results of this project may lead to the development of new therapies for the treatment of diabetic
wounds, which cause significant morbidity and mortality in a large proportion of the diabetic population.
These therapies will likely include topical nitrite and inhibitors of XOR. Additionally, our research is
providing mechanistic data on the causes of impaired diabetic wound healing and methods to target
these mechanisms.
Professional Practice Gap:
Our lab has been researching the role of xanthine oxidoreductase (XOR) in diabetic wound healing.
While XOR is necessary for normal wound healing, it has been suggested that it causes excessive
oxidative stress in diabetic wounds and impairs diabetic wound healing. Our prior experiments with oral
and topical febuxostat, an XOR inhibitor, have failed to consistently inhibit XOR and demonstrate a clear
benefit for diabetic wound healing. With this research, we now demonstrate that inhibition of XOR with
dietary tungsten does accelerate diabetic wound healing. Additionally, despite inhibiting XOR, which
also functions as a nitrite reductase, topical nitrite has still been demonstrated to be effective in
accelerating wound healing. Our current results indicate that this may be secondary to changes effected
by nitrite on the wound microbiome.
Impact Of Dietary Tungsten And Topical Nitrite In Diabetic Wound Healing And The Composition Of
The Wound Microbiome
Kathy Gonzalez, Karim M Salem, UPMC, Pittsburgh, PA; Barbara Methé, Kelvin Li, Guiying Hong, Univ of
Pittsburgh, Pittsburgh, PA; Edith Tzeng, UNIVERSITY PITTSBURGH, Pittsburgh, PA
Background:
Impaired diabetic (DM) wound healing is attributed to increased reactive oxygen species (ROS) and
decreased nitric oxide (NO) production. Xanthine oxidoreductase (XOR) produces ROS and can convert
nitrite to NO. XOR is expressed in wounds and necessary for normal healing. In DM wounds, XOR may
contribute to excess ROS and poor healing. We have shown that topical nitrite accelerates DM wound
healing. We hypothesize that XOR inhibition will improve DM wound repair and that concurrent topical
nitrite would not be effective.
Methods:
Diabetic db/db mice were fed a tungsten (W)-enriched diet, to inhibit XOR, or standard chow for 2 wks
before excisional wounding. Wounds were treated with either topical Aquaphor or Aquaphor/2% nitrite
(nitrite). Wounds areas were quantified every other day for 28 days. Healing rates were compared with
ANOVA. Wound swabs were collected on Days 2 and 14 for microbiota analysis using 16S rRNA
sequencing and paired-difference analysis.
Results:
While topical nitrite or W diet improved DM wound healing compared to chow + Aquaphor, W + nitrite
mice healed significantly faster than these groups at all wound sizes (Figure). Microbiota composition
over time was compared between the same taxon in the same mouse. Greater differences between taxa
were found in the nitrite and W + nitrite-treated groups while differences decreased with W diet.
While Staphylococcus (Staph) was the predominant taxon in all groups, nitrite significantly
increased Staph over time in chow fed mice.
Conclusion:
XOR inhibition with W improved DM wound healing. Surprisingly, the combination of W + nitrite further
accelerated healing. This supports the benefit of XOR inhibition to reduce ROS and that nitrite
conversion to NO is supported by other nitrite reductases. W and nitrite both mediate wound
microbiome changes favoring Staph species which may contribute to improved wound repair. The
contribution of the microbiome to nitrite conversion to NO needs to be assessed.
727
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
The results from this project may highlight new biomarkers involved with PAD/CLI. Furthermore, RNA
based therapies can be developed from the findings of this project.
Professional Practice Gap:
The mechanisms involved with the pathogenesis of peripheral artery diseases and critical limb ischemia.
MicroRNA-181b Regulates Critical Limb Ischemia In Diabetic Mice
Henry S Cheng, Brigham and Women's Hosp, Boston, MA; Daniel Perez-Cremades, BWH, Boston, MA;
Marc P Bonaca, CPC Clinical Res, Aurora, CO; Mark Feinberg, Brigham and Women's Hosp, Boston, MA
Despite current therapeutic strategies, human subjects with peripheral artery disease (PAD) suffer from
an increased risk of critical limb ischemia (CLI) and limb amputation. PAD and CLI are associated with
several risk factors, such as aging and diabetes. MicroRNAs are crucial regulators of gene expression
with important therapeutic potential. However, the role of microRNAs in diabetic CLI remains poorly
understood. Due to the high conservation of microRNAs across species, plasma microRNA sequencing
from both human patients with PAD and mouse models were used to identify new microRNAs that
regulate diabetic limb ischemia. Two methods were used to induce limb ischemia in obese diabetic
(db/db) mice: acute limb ischemia model - ligation of the femoral artery; and subacute limb ischemia
model - insertion of ameroid constrictors around the femoral artery to induce slow occlusion. Through
stratification of the microRNA sequencing screens in mouse and human diabetic CLI cohorts, we
identified miR-181b as a top differentially expressed miRNA capable of regulating angiogenesis in vitro.
Furthermore, miR-181b inhibitors can promote proliferation of cultured mouse vascular smooth muscle
cells (VSMC). Systemic miR-181a2b2 knockout (KO) mice demonstrate markedly improved blood flow
recovery (by 3.7-fold at 28 days) after limb ischemia compared to controls (P<0.002). Analysis of skeletal
muscle RNA expression revealed a 2-fold increase of VSMC markers Myosin heavy chain 11 (P<0.011)
and 1.65-fold increase of Myosin light polypeptide 6 (P<0.022) in systemic miR-181a2b2 KO mice. In
contrast, endothelial-specific miR-181a2b2 KO mice showed no difference in blood flow recovery.
Collectively, the stark increase of blood flow recovery and VSMC markers in systemic miR-181a2b2 KO
mice suggests enhancement of arteriogenesis in lieu of angiogenesis. Further investigation of miR-181b
in diabetic CLI may prove to be a key therapeutic target for promoting tissue perfusion in PAD.
817
There are no unlabeled/unapproved uses of drugs or products
Knowledge Strategy:
To inform the vascular surgeons that oxidative stress can disrupt the biopterin system impairing vascular
function.
Professional Practice Gap:
The development of a blood biomarker that can be used for early detection of PAD.
Nitric Oxide Bioavailability Is Reduced In Peripheral Artery Disease In Association With Increased
Oxidative Stress And An Altered Biopterin System
Ahmed Ismaeel, Evlampia Papoutsi, Florida State Univ, Tallahassee, FL; William T Bohannon, Robert
Smith, Baylor Scott and White Heath, Temple, TX; Robert Brumberg, Vascular Surgery Associates,
Tallahassee, FL; Carlos H Castro, Jeffrey S Kirk, Capital Regional Medical Ctr, Tallahassee, FL; Iraklis I
Pipinos, UNIV NEBRASKA MEDICAL CTR, Omaha, NE; Panagiotis Koutakis, Florida State Univ, Tallahassee,
FL
Introduction: Peripheral artery disease (PAD) manifestations are the result of several pathophysiologic
mechanisms including endothelial dysfunction. This process is believed to be mediated by decreased
bioavailability of nitric oxide (NO), and oxidative stress may play a role in inducing endothelial
dysfunction by increasing oxidation of the NO synthase (NOS) co-factor tetrahydrobiopterin (BH4) to
dihydrobiopterin (BH2) as well as by inducing the expression of the endogenous NOS inhibitor
asymmetric dimethylarginine (ADMA). Hypothesis: We hypothesized that the regulators of NO synthesis
and bioavailability are compromised in PAD as a result of elevated oxidative stress. Methods: Blood was
collected from 35 PAD patients with intermittent claudication (IC), 26 patients with critical limb ischemia
(CLI), and 35 non-PAD controls, and serum was analyzed using specific assay kits to determine levels of
BH4, BH2, arginine, ADMA, nitrate/nitrite (NOx, used to quantify NO), and the oxidative stress markers
peroxynitrite, protein carbonyls, 8-OHdG, and 4-HNE. Results: IC and CLI patients demonstrated reduced
serum NOx levels compared to control (control: 84.89 ± 40.81, IC: 50.12 ± 17.48, CLI: 43.96 ± 19.47
μmol/L). Furthermore, BH4 was significantly reduced and BH2 significantly elevated in IC and CLI
compared to control, resulting in a lower BH4/BH2 ratio (control: 5.28 ± 6.71, IC: 1.08 ± 0.64, CLI: 1.09 ±
0.78). The arginine/ADMA ratio was significantly lower in CLI compared to both IC and control.
Furthermore, all the oxidative stress markers were significantly elevated in IC compared to control, with
further elevations in CLI. There was a significant inverse correlation between all the oxidative stress
markers and the BH4/BH2 ratio as well as NOx levels. Conclusions: The NO system is dysregulated in
PAD, which appears to be driven by elevated oxidative stress, worsening as the disease progresses from
IC to CLI.