The role of microRNA-145 in human embryonic stem cell differentiation into vascular cells

Shintaro Yamaguchi, Kenichi Yamahara, Koichiro Honma, Sayuri Suzuki, Shizuka Fujii, Ryuji Morizane, Toshiaki Monkawa, Yumi Matsuzaki, Kenji Kangawa, Hiroshi Itoh

Research output: Contribution to journalArticle

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Abstract

Background: Recent studies have reported that microRNA-145 (miR-145) is a critical mediator in the regulation of proliferation, differentiation, and phenotype expression of smooth muscle cells (SMCs). Previously, we established a system for differentiating human ESCs into vascular cells including endothelial cells (ECs) and vascular smooth muscle cells (SMCs). In the present study, we investigated the role of miR-145 in the differentiation process from human ESCs into ECs and SMCs. Methods and results: Undifferentiated human ESCs were induced to differentiate into vascular lineage according to our established method. Quantitative RT-PCR analysis revealed that human ESC-derived precursor of SMCs (ES-pre-SMCs), similar to human aortic SMCs, expressed a significant amount of miR-145 as well as smooth muscle-specific proteins, compared to undifferentiated human ESCs, adult ECs, or ESC-derived ECs (ES-ECs). However, morphological analysis revealed that human ES-pre-SMCs appeared round and flattened in shape, though human aortic SMCs exhibited the typical spindle-like morphology of SMCs. In addition, Krüppel-like factor 4 and 5 (KLF4 and 5), direct targets of miR-145 and suppressors of smooth muscle differentiation, were upregulated in ES-pre-SMCs compared to aortic SMCs, indicating ES-pre-SMCs were not fully differentiated SMCs. Overexpression of miR-145 in ES-pre-SMCs upregulated the expression of smooth muscle markers, repressed KLF4 and 5 expressions, and changed their morphology into a differentiated spindle-like shape. Furthermore, by introduction of miR-145, ES-pre-SMC proliferation was significantly inhibited and carbachol-stimulated contraction of ES-pre-SMCs was significantly increased. In contrast, downregulation of miR-145 in ES-pre-SMCs upregulated KLF4 and 5 expressions, suppressed the expression of smooth muscle markers, and left unchanged their proliferation and contractility. In ES-ECs, miR-145 overexpression did not induce the synthesis of smooth muscle-related proteins nor suppress the expression of endothelial nitric oxide synthase. Conclusion: We showed that miR-145 can regulate the fate and phenotype of human ES-pre-SMCs as they become fully differentiated SMCs. Overexpression of miR-145 on human ES-pre-SMCs is a promising method to obtain functional mature SMCs from human ESCs, which are required for reliable experimental research in the fields of atherosclerosis, hypertension and other vascular diseases.

Original languageEnglish
Pages (from-to)468-474
Number of pages7
JournalAtherosclerosis
Volume219
Issue number2
DOIs
Publication statusPublished - 2011 Dec

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MicroRNAs
Smooth Muscle Myocytes
Blood Vessels
Cell Differentiation
Smooth Muscle
Endothelial Cells
Muscle Proteins
Human Embryonic Stem Cells
Phenotype
Nitric Oxide Synthase Type III
Myoblasts
Carbachol
Vascular Diseases
Vascular Smooth Muscle
Muscle Cells
Atherosclerosis
Down-Regulation
Hypertension
Polymerase Chain Reaction

Keywords

  • Embryonic stem cells
  • MicroRNA-145
  • Smooth muscle cells
  • Vascular differentiation

ASJC Scopus subject areas

  • Cardiology and Cardiovascular Medicine

Cite this

The role of microRNA-145 in human embryonic stem cell differentiation into vascular cells. / Yamaguchi, Shintaro; Yamahara, Kenichi; Honma, Koichiro; Suzuki, Sayuri; Fujii, Shizuka; Morizane, Ryuji; Monkawa, Toshiaki; Matsuzaki, Yumi; Kangawa, Kenji; Itoh, Hiroshi.

In: Atherosclerosis, Vol. 219, No. 2, 12.2011, p. 468-474.

Research output: Contribution to journalArticle

Yamaguchi, S, Yamahara, K, Honma, K, Suzuki, S, Fujii, S, Morizane, R, Monkawa, T, Matsuzaki, Y, Kangawa, K & Itoh, H 2011, 'The role of microRNA-145 in human embryonic stem cell differentiation into vascular cells', Atherosclerosis, vol. 219, no. 2, pp. 468-474. https://doi.org/10.1016/j.atherosclerosis.2011.09.004
Yamaguchi, Shintaro ; Yamahara, Kenichi ; Honma, Koichiro ; Suzuki, Sayuri ; Fujii, Shizuka ; Morizane, Ryuji ; Monkawa, Toshiaki ; Matsuzaki, Yumi ; Kangawa, Kenji ; Itoh, Hiroshi. / The role of microRNA-145 in human embryonic stem cell differentiation into vascular cells. In: Atherosclerosis. 2011 ; Vol. 219, No. 2. pp. 468-474.
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abstract = "Background: Recent studies have reported that microRNA-145 (miR-145) is a critical mediator in the regulation of proliferation, differentiation, and phenotype expression of smooth muscle cells (SMCs). Previously, we established a system for differentiating human ESCs into vascular cells including endothelial cells (ECs) and vascular smooth muscle cells (SMCs). In the present study, we investigated the role of miR-145 in the differentiation process from human ESCs into ECs and SMCs. Methods and results: Undifferentiated human ESCs were induced to differentiate into vascular lineage according to our established method. Quantitative RT-PCR analysis revealed that human ESC-derived precursor of SMCs (ES-pre-SMCs), similar to human aortic SMCs, expressed a significant amount of miR-145 as well as smooth muscle-specific proteins, compared to undifferentiated human ESCs, adult ECs, or ESC-derived ECs (ES-ECs). However, morphological analysis revealed that human ES-pre-SMCs appeared round and flattened in shape, though human aortic SMCs exhibited the typical spindle-like morphology of SMCs. In addition, Kr{\"u}ppel-like factor 4 and 5 (KLF4 and 5), direct targets of miR-145 and suppressors of smooth muscle differentiation, were upregulated in ES-pre-SMCs compared to aortic SMCs, indicating ES-pre-SMCs were not fully differentiated SMCs. Overexpression of miR-145 in ES-pre-SMCs upregulated the expression of smooth muscle markers, repressed KLF4 and 5 expressions, and changed their morphology into a differentiated spindle-like shape. Furthermore, by introduction of miR-145, ES-pre-SMC proliferation was significantly inhibited and carbachol-stimulated contraction of ES-pre-SMCs was significantly increased. In contrast, downregulation of miR-145 in ES-pre-SMCs upregulated KLF4 and 5 expressions, suppressed the expression of smooth muscle markers, and left unchanged their proliferation and contractility. In ES-ECs, miR-145 overexpression did not induce the synthesis of smooth muscle-related proteins nor suppress the expression of endothelial nitric oxide synthase. Conclusion: We showed that miR-145 can regulate the fate and phenotype of human ES-pre-SMCs as they become fully differentiated SMCs. Overexpression of miR-145 on human ES-pre-SMCs is a promising method to obtain functional mature SMCs from human ESCs, which are required for reliable experimental research in the fields of atherosclerosis, hypertension and other vascular diseases.",
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AU - Fujii, Shizuka

AU - Morizane, Ryuji

AU - Monkawa, Toshiaki

AU - Matsuzaki, Yumi

AU - Kangawa, Kenji

AU - Itoh, Hiroshi

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KW - MicroRNA-145

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