Analyses of Endothelial Cells and Endothelial Progenitor Cells Released Microvesicles by Using Microbead and Q-dot Based Nanoparticle Tracking Analysis.
Accurate analysis of specific microvesicles (MVs) from biofluids is critical and challenging. Here we described novel methods to purify and detect MVs shed from endothelial cells (ECs) and endothelial progenitor cells (EPCs) by combining microbeads with fluorescence quantum dots (Q-dots) coupled nanoparticle tracking analysis (NTA). In the in vitro screening systems, we demonstrated that 1) anti-CD105 (EC marker) and anti-CD34 (EPC marker) conjugated-microbeads had the highest sensitivity and specificity for isolating respective MVs, which were confirmed with negative controls, CD41 and CD235a; 2) anti-CD144 (EC marker) and anti-KDR (EPC marker) conjugated-Q-dots exhibited the best sensitivity and specificity for their respective MV NTA detection, which were confirmed with positive control, anti-Annexin V (MV universal marker). The methods were further validated by their ability to efficiently recover the known amount of EC-MVs and EPC-MVs from particle-depleted plasma, and to detect the dynamical changes of plasma MVs in ischemic stroke patients, as compared with traditional flow cytometry. These novel methods provide ideal approaches for functional analysis and biomarker discovery of ECs- and EPCs- derived MVs.
Department of Pharmacology and Toxicology, Boonshoft School of Medicine, Wright State University, Dayton, Ohio, 45435, USA.
Jinju
J
Wang
Department of Pharmacology and Toxicology, Boonshoft School of Medicine, Wright State University, Dayton, Ohio, 45435, USA.|Department of Cardiology, Guangzhou Institute of Cardiovascular Disease, the Second Hospital of Guangzhou Medical University, Guangzhou, 510000, China.
Yun
Y
Zhong
Guangdong Key Laboratory of Age-Related Cardiac and Cerebral Diseases, Institute of Neurology, Affiliated Hospital of Guangdong Medical College, Zhanjiang, 524000, China.
Xiaotang
X
Ma
Department of Pharmacology and Toxicology, Boonshoft School of Medicine, Wright State University, Dayton, Ohio, 45435, USA.
Xiang
X
Xiao
Department of Cardiology, Guangzhou Institute of Cardiovascular Disease, the Second Hospital of Guangzhou Medical University, Guangzhou, 510000, China.
Chuanfang
C
Cheng
Guangdong Key Laboratory of Age-Related Cardiac and Cerebral Diseases, Institute of Neurology, Affiliated Hospital of Guangdong Medical College, Zhanjiang, 524000, China.
Yusen
Y
Chen
Department of Neurology, Ochsner Medical Center, Jefferson, LA, 70121, USA.
Ifeanyi
I
Iwuchukwu
Department of Neurology, Ochsner Medical Center, Jefferson, LA, 70121, USA.|Departments of Neurology and Internal Medicine, Boonshoft School of Medicine, Wright State University, Dayton, Ohio, 45435, USA.
Kenneth J
KJ
Gaines
Guangdong Key Laboratory of Age-Related Cardiac and Cerebral Diseases, Institute of Neurology, Affiliated Hospital of Guangdong Medical College, Zhanjiang, 524000, China.
Bin
B
Zhao
Department of Cardiology, Guangzhou Institute of Cardiovascular Disease, the Second Hospital of Guangzhou Medical University, Guangzhou, 510000, China.
Shiming
S
Liu
Department of Pharmacology and Toxicology, Boonshoft School of Medicine, Wright State University, Dayton, Ohio, 45435, USA.|Dayton VA Medical Center, Dayton, Ohio, 45428, USA.
Jeffrey B
JB
Travers
Department of Pharmacology and Toxicology, Boonshoft School of Medicine, Wright State University, Dayton, Ohio, 45435, USA.|Guangdong Key Laboratory of Age-Related Cardiac and Cerebral Diseases, Institute of Neurology, Affiliated Hospital of Guangdong Medical College, Zhanjiang, 524000, China.
Ji C
JC
Bihl
Department of Pharmacology and Toxicology, Boonshoft School of Medicine, Wright State University, Dayton, Ohio, 45435, USA.|Department of Cardiology, Guangzhou Institute of Cardiovascular Disease, the Second Hospital of Guangzhou Medical University, Guangzhou, 510000, China.|Guangdong Key Laboratory of Age-Related Cardiac and Cerebral Diseases, Institute of Neurology, Affiliated Hospital of Guangdong Medical College, Zhanjiang, 524000, China.|Departments of Neurology and Internal Medicine, Boonshoft School of Medicine, Wright State University, Dayton, Ohio, 45435, USA.
Yanfang
Y
Chen
D015415:Biomarkers; D055252:Cell-Derived Microparticles / Q000378:metabolism* / Q000648:ultrastructure; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism*; D066026:Endothelial Progenitor Cells / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D008863:Microspheres; D053758:Nanoparticles / Q000648:ultrastructure; D045663:Quantum Dots
D015415:Biomarkers