454-P: Conditioned Medium from Dental Pulp Stem Cells Promotes Angiogenesis
Authors: KATO MAKOTO , TSUNEKAWA SHIN , NAKAMURA NOBUHISA , MIURA-YURA EMIRI , YAMADA YUICHIRO , HAYASHI YUSUKE , INOUE RIEKO , MOHIUDDIN MOHAMMAD SARIF , MORISHITA YOSHIAKI , HIMENO TATSUHITO , KONDO MASAKI , KATO YOSHIRO , KAMIYA HIDEKI , NARUSE KEIKO , NAKAMURA JIRO 
Published: June 2020

Background/Aim: We previously reported that conditioned medium (CM) of stem cells from human exfoliated deciduous teeth (SHED) improved diabetic polyneuropathy in diabetic mice due to an increase of capillary blood flow in the vasa nervorum, and in in vitro study only secreted factors of less than 6 kDa fraction of SHED-CM promoted neurite outgrowth of mouse dorsal root ganglion neurons. In this study, we examined whether SHED-CM had beneficial effects on angiogenesis.
Methods: SHED-CM was collected 48 hours after culturing in serum-free DMEM, and was separated into two fractions according to molecular weight (less than 6 kDa and more than 6 kDa). Exosomes were isolated from SHED-CM by ultracentrifugation. Human umbilical-vein endothelial cells (HUVECs) were cultured with six different media (DMEM, DMEM with VEGF, SHED-CM, less than 6 kDa, more than 6 kDa or DMEM with exosomes) for 12-48 hours, then MTT assay, wound healing assay, Boyden chamber assay and tube formation assay were performed to evaluate the cell viability, migration ability and tube formation ability in HUVECs. Rat aortic ring assay and mouse Matrigel plug assay were performed to assess neovascularization and endothelial cell migration.
Results: SHED-CM, especially more than 6 kDa fraction, significantly promoted cell viability, migration and tube formation in HUVECs, compared with DMEM. Meanwhile, less than 6 kDa fraction promoted only tube formation, and the effects of exosomes were negligible. In aortic ring assay and Matrigel plug assay, SHED-CM accelerated neovascularization and increased endothelial cell migration in HUVECs, and the effects of more than 6 kDa fraction were stronger than that of less than 6 kDa fraction. These data suggested that the secreted factors in more than 6 kDa fraction played an essential role in promoting angiogenesis.
Conclusion: Soluble factors from SHED might have angiogenesis-promoting effects and hold promise for comprehensive clinical applications in vascular diseases and diabetic complications.

Disclosure
M. Kato: None. S. Tsunekawa: None. N. Nakamura: None. E. Miura-Yura: None. Y. Yamada: None. Y. Hayashi: None. R. Inoue: None. M. Mohiuddin: Other Relationship; Self; Abbott Japan Co. Ltd, ARKRAY, Astellas Pharma Inc., Astellas Pharma Inc., AstraZeneca K.K., Boehlinger Ingelheim Japan Co. Ltd., Boehringer Ingelheim Pharmaceuticals, Inc., Daiichi Sankyo, Daiichi Sankyo, Eli Lilly Japan K.K., Eli Lilly Japan K.K., Fukuda Denshi, Japan Tobacco Inc., Kaken Pharmaceutical Co., Ltd., Kissei Pharmaceutical Co., Ltd., Kowa pharmaceu. co. Ltd., Kowa Pharmaceu. Co., Ltd., Kyowa Kirin Co., Ltd., Mitsubishi Tanabe Pharma Corporation, Mitsubishi Tanabe Pharma Corporation, MSD K.K., MSD K.K., Mylan N. V., Novartis Pharma K.K., Novartis Pharma K.K., Novo Nordisk Pharma Ltd., Novo Nordisk Pharma Ltd., Ono Pharmaceu. Co., Ltd., Ono Pharmaceutical Co., Ltd., Otsuka Pharmaceutical Co., Ltd., Pfizer Japan Inc., Sanofi K.K., Sanofi K.K., Sanwa Kagaku Kenkyusho, Sanwa Kagaku Kenkyusho, Shionogi & Co., Ltd., Sumitomo Dainippon Pharma Co., Ltd., Sumitomo Dainippon Pharma Co., Ltd., Taisho Pharmaceutical Co., Ltd., Taisho Pharmaceutical Co., Ltd., Takeda Pharmaceutical Company Limited, Takeda Pharmaceutical Company Limited, Terumo Co. Ltd. Y. Morishita: None. T. Himeno: None. M. Kondo: None. Y. Kato: None. H. Kamiya: Speaker’s Bureau; Self; Astellas Pharma Inc., AstraZeneca K.K., Boehringer Ingelheim K.K., Daiichi Sankyo, Eli Lilly Japan K.K., Fukuda Denshi, Kissei Pharmaceutical Co., Ltd., Kowa Company, Ltd., Kyowa Hakko Kirin Co., Ltd., Mitsubishi Tanabe Pharma Corporation, MSD K.K., Novartis Pharma K.K., Novo Nordisk Pharma Ltd., Ono Pharmaceutical Co., Ltd., Sanofi K.K., Sumitomo Dainippon Pharma Co., Ltd., Taisho Pharmaceutical Co., Ltd., Takeda Pharmaceutical Company Limited. K. Naruse: None. J. Nakamura: Research Support; Self; Astellas Pharma Inc., Boehlinger Ingelheim Japan Co., Ltd., Daiichi Sankyo, Eli Lilly Japan K.K., Japan Tobacco Inc., Kaken Pharmaceutical Co., Ltd., Kowa Company, Ltd., Kyowa Hakko Kirin Co., Ltd., Mitsubishi Tanabe Pharma Corporation, MSD K.K., Novartis Pharma K.K., Novo Nordisk Pharma Ltd., Ono Pharmaceutical Co., Ltd., Otsuka Pharmaceutical Co., Ltd., Pfizer Japan Inc., Sanofi K.K., Sanwa Kagaku Kenkyusho, Shionogi & Co., Ltd., Sumitomo Dainippon Pharma Co., Ltd., Taisho Pharmaceutical Co., Ltd., Takeda Pharmaceutical Company Limited. Speaker’s Bureau; Self; Abbott Japan Co., Ltd., ARKRAY, Astellas Pharma Inc., AstraZeneca K.K., Boehlinger Ingelheim Japan Co., Ltd.,, Daiichi Sankyo, Eli Lilly Japan K.K., Fukuda Denshi, Kissei Pharmaceutical Co., Ltd., Kowa Company, Ltd., Mitsubishi Tanabe Pharma Corporation, MSD K.K., Mylan, Novartis Pharma K.K., Novo Nordisk Pharma Ltd, Ono Pharmaceutical Co., Ltd., Sanofi, Sanwa Kagaku Kenkyusho, Sumitomo Dainippon Pharma Co., Ltd., Taisho Pharmaceutical Co., Ltd., Takeda Pharmaceutical Company Limited, Terumo Medical Corporation.