[논문]miRNA-222 Modulates Differentiation of Mouse Embryonic Stem Cells

Ahn, Hee-Jin; Jung, Jee-Eun; Park, Kyung-Soon

[국내논문] miRNA-222 Modulates Differentiation of Mouse Embryonic Stem Cells 원문보기

Development & reproduction = 발생과 생식, v.15 no.4, 2011년, pp.331 - 338

Ahn, Hee-Jin (Department of Biomedical Science, College of Life Science, CHA University) , Jung, Jee-Eun (Department of Biomedical Science, College of Life Science, CHA University) , Park, Kyung-Soon (Department of Biomedical Science, College of Life Science, CHA University)

Abstract ▼ AI-Helper

MicroRNAs (miRNAs) function as a key regulator of diverse cellular functions. To find out novel miRNAs that promote the differentiation of mouse embryonic stem cells (mESCs), we compared the miRNAs expression profiles of mESCs under self-renewal vs. differentiation states. We noticed that miR-222 was highly expressed during the differentiation of mESCs. Quantitative RT-PCR analysis revealed that expression of miR-222 was up-regulated during the embryonic bodies formation and retinoic acid -dependent differentiation. When miR-222 was suppressed by antogomiR-222, the differentiation of mESCs was delayed compared to control. Self-renewal marker expression or cell proliferation was not affected but the expression of lineage specific marker was suppressed by the treatment of miR-222 inhibitor during the differentiation of mESCs. Taken together, these results suggest that miR-222 functions to promote the differentiation of mESCs by regulating expression of differentiation related genes.

Keyword

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제안 방법

In this study, we analyzed the expression profiles of miRNAs during the differentiation of mouse embryonic stem cells (mESCs). We found that miR-222 was prominently upregulated during the differentiation of mESCs.
com). To induced differentiation, we formed Embryonic bodies (EBs) in LIF-deficient ESCs medium (as described above) by suspension culture and media changed every two days. To induce spontaneous differentiation, mESCs were cultured in LIF-deficient ESC medium with 500 nM all-trans retinoic acid (RA) (Lee et al.
First-Strand Synthesis System (Invitrogen), according to the manufacturer’s instructions. Real-time PCR was carried out in triplicate with the Quantitect SYBR Green PCR kit (Qiagen, Valencia, CA, http://www.qiagen.com) and CFX96 Real-time System (Bio-Rad Laboratories, Richmond, CA http://www.bio-rad.com). For quantification, target genes were normalized with the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene.
Graphical data are presented as means ± SD. Each experiment was performed at least three times and subjected to statistical analysis. Statistical significance between two groups was determined using Student’s t-test.
Despite several studies reported the function of miR-222 in cancer, the role of miR-222 in ESCs has not been reported. To confirm the result of miRNAs array, we quantitatively analyzed the expression level of miR-222 in 2 days old EBs and spontaneously differentiated EBs. As shown in Figure 2A, the miR-222 expression was not activated in EBs but it was significantly increased during the differentiation of EBs.
To study the role of miR-222 for initiation of mESCs’ differentiation, we suppressed miR-222 using antagomiR-222 and then examined the differentiation pattern of mESCs.

대상 데이터

AntagomiR for miR-222 was purchased from Ambion (ABS-4427975, Gene ID: 000397) and transfection was performed according to the manufacturer’s instructions using Lipofectamin 2000 (Invitrogen).

데이터처리

Statistical significance between two groups was determined using Student’s t-test.

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