title: CC chemokine ligand 2 and leukemia inhibitory factor cooperatively promote pluripotency in mouse induced pluripotent cells.
abstract: The pluripotency of mouse embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) can be maintained by feeder cells, which secrete leukemia inhibitory factor (LIF). We found that feeder cells provide a relatively low concentration (25 unit/ml) of LIF, which is insufficient to maintain the ESCs/iPSCs pluripotency in feeder free conditions. To identify additional factors involved in the maintenance of pluripotency, we carried out a global transcript expression profiling of mouse iPSCs cultured on feeder cells and in feeder-free (LIF-treated) conditions. This identified 17 significantly differentially expressed genes (adjusted p value <0.05) including seven chemokines overexpressed in iPSCs grown on feeder cells. Ectopic expression of these chemokines in iPSCs revealed that CC chemokine ligand 2 (Ccl2) induced the key transcription factor genes for pluripotency, Klf4, Nanog, Sox2, and Tbx3. Furthermore, addition of recombinant Ccl2 protein drastically increased the number of Nanog-green fluorescent protein-positive iPSCs grown in low-LIF feeder free conditions. We further revealed that pluripotency promotion by Ccl2 is mediated by activating the Stat3-pathway followed by Klf4 upregulation. We demonstrated that Ccl2-mediated increased pluripotency is independent of phosphoinositide 3-kinase and mitogen-activated protein kinase pathways and that Tbx3 may be upregulated by Klf4. Overall, Ccl2 cooperatively activates the Stat3-pathway with LIF in feeder-free conditions to maintain pluripotency for ESCs/iPSCs.
authors.affiliation: RIKEN Omics Science Center, RIKEN Yokohama Institute, Tsurumi-ku, Yokohama, Kanagawa, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Hasegawa
authors.forename: Naoko
authors.initials: N
authors.lastname: Takahashi
authors.forename: Alistair R R
authors.initials: AR
authors.lastname: Forrest
authors.forename: Jay W
authors.initials: JW
authors.lastname: Shin
authors.forename: Yohei
authors.initials: Y
authors.lastname: Kinoshita
authors.forename: Harukazu
authors.initials: H
authors.lastname: Suzuki
authors.forename: Yoshihide
authors.initials: Y
authors.lastname: Hayashizaki
mesh_terms: D000818:Animals; D002460:Cell Line; D018932:Chemokine CCL2 / Q000235:genetics / Q000378:metabolism*; D018920:Coculture Techniques; D015870:Gene Expression; D020869:Gene Expression Profiling; D005819:Genetic Markers; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D057026:Induced Pluripotent Stem Cells / Q000187:drug effects / Q000502:physiology*; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000235:genetics / Q000378:metabolism; D053605:Leukemia Inhibitory Factor / Q000378:metabolism* / Q000494:pharmacology; D051379:Mice; D000071317:Nanog Homeobox Protein; D020411:Oligonucleotide Array Sequence Analysis; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D055748:SOXB1 Transcription Factors / Q000235:genetics / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000378:metabolism; D020825:T-Box Domain Proteins / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation
chemical_list: C491768:Ccl2 protein, mouse; D018932:Chemokine CCL2; D005819:Genetic Markers; D018398:Homeodomain Proteins; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D053605:Leukemia Inhibitory Factor; D000071317:Nanog Homeobox Protein; C475540:Nanog protein, mouse; D011994:Recombinant Proteins; D055748:SOXB1 Transcription Factors; D050796:STAT3 Transcription Factor; C527783:Sox2 protein, mouse; C494087:Stat3 protein, mouse; D020825:T-Box Domain Proteins; C491460:Tbx3 protein, mouse