Modified method for effective primary vascular smooth muscle progenitor cell culture from peripheral blood
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ORIGINAL ARTICLE
Modified method for effective primary vascular smooth muscle progenitor cell culture from peripheral blood Jin-Hee Seong . Yi-Sun Song . Hyun-Woo Joo . In-Hwa Park . Guang-Yin Shen . Na-Kyoung Shin . A-Hyeon Lee . Amy M. Kwon . Yonggu Lee . Hyuck Kim . Kyung-Soo Kim
Received: 10 February 2020 / Accepted: 2 September 2020 Ó The Author(s) 2020
Abstract In previous studies, vascular smooth muscle progenitor cells (vSMPCs) isolated from peripheral blood mononuclear cells (PBMCs) were cultured using medium containing platelet-derived growth factor-BB (PDGF-BB) for 4 weeks. However, this method requires long culture periods of up to 4 weeks and yields low cell counts. Therefore, we proposed the modified method to improve the cell yield and purity and to reduce the cell culture period. PBMCs were isolated from human peripheral blood and cultured by the conventional method using medium containing PDGF-BB alone or the modified method using medium containing PDGF-BB, basic fibroblast growth factor (bFGF), and insulin-transferrin-selenium ITS for 4 weeks. The purity of vSMPCs was analyzed for the expression of a- smooth muscle actin (SMA) by flow cytometry and significantly higher in the
modified method than conventional methods at the 1st and 2nd weeks. Also, mRNA expression of a-SMA by real-time PCR was significantly higher in the modified method than conventional method at the 2 weeks. The yield of vSMPCs by trypan blue exclusion assay was significantly higher in the modified method than conventional method at the 1st, 2nd and 3rd weeks. The primary culture using the modified method with PDGF-BB, bFGF, and ITS not only improved cell purity and yield, but also shortened the culture period, compared to the conventional culture method for vSMPCs. The modified method will be a time-saving and useful tool in various studies related to vascular pathology.
J.-H. Seong Y.-S. Song H.-W. Joo I.-H. Park N.-K. Shin A.-H. Lee K.-S. Kim Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul, South Korea
A. M. Kwon Biostatistical Consulting and Research Laboratory, Medical Research Collaborating Center, IndustryUniversity Cooperation Foundation, Hanyang University, Seoul, South Korea
G.-Y. Shen K.-S. Kim (&) Division of Cardiology, Department of Internal Medicine, Hanyang University College of Medicine, Seoul, South Korea e-mail: [email protected] G.-Y. Shen Division of Cardiology, Department of Internal Medicine, Jilin University Jilin Central Hospital, Jilin, China
Keywords VSMPCs PBMCs PDGF-BB Peripheral blood Cell culture
Y. Lee Department of Internal Medicine, Hanyang University Guri Hospital, Guri, South Korea H. Kim Department of Thoracic Surgery, Hanyang University Seoul Hospital, Seoul, South Korea
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Cytotechnology
Abbreviations a-SMA Alpha-smooth muscle actin bFGF Basic fibroblast growth factor BM Bone marrow CD31 Cluster of differentiation 31 HASMCs Human aortic smooth muscle cells ITS Insulin-transf
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