miR-335 orchestrates cell proliferation, migration and differentiation in human mesenchymal stem cells

Abstract

In spite of the extensive potential of human mesenchymal stem cells (hMSCs) in cell therapy, little is known about the molecular mechanisms that regulate their therapeutic properties. We aimed to identify microRNAs (miRNAs) involved in controlling the transition between the resting and reparative phenotypes of hMSCs, hypothesizing that these miRNAs must be present in the undifferentiated cells and downregulated to allow initiation of distinct activation/differentiation programs. Differential miRNA expression analyses revealed that miR-335 is significantly downregulated upon hMSC differentiation. In addition, hMSCs derived from a variety of tissues express miR-335 at a higher level than human skin fibroblasts, and overexpression of miR-335 in hMSCs inhibited their proliferation and migration, as well as their osteogenic and adipogenic potential. Expression of miR-335 in hMSCs was upregulated by the canonical Wnt signaling pathway, a positive regulator of MSC self-renewal, and downregulated by interferon-c (IFN-c), a pro-inflammatory cytokine that has an important role in activating the immunomodulatory properties of hMSCs. Differential gene expression analyses, in combination with computational searches, defined a cluster of 62 putative target genes for miR-335 in hMSCs. Western blot and 3 0 UTR reporter assays confirmed RUNX2 as a direct target of miR-335 in hMSCs. These results strongly suggest that miR-335 downregulation is critical for the acquisition of reparative MSC phenotypes. Mesenchymal stem cells (MSCs) are multipotent mesodermderived somatic stem cell (SSC) precursors of non-hematopoietic connective tissues that are present in the stroma of virtually all mammalian organs, especially bone marrow and subcutaneous fat (reviewed in Bernardo et al. 1 ). Upon activation by tissue damage, MSCs contribute to tissue-repair processes through a multitude of activities, including cell proliferation, differentiation and migration, and the regulation of angiogenesis and immune responses. There is growing evidence, in both animal and clinical models, that administration of ex vivo-expanded human MSCs (hMSCs) has potential to ameliorate many degenerative disorders; however, the specific molecular mechanisms underlying this therapeutic potential remain mostly unknown.MicroRNAs (miRNAs) are an extensive family of small (18-24 nucleotide), single-stranded non-coding RNAs, which regulate gene expression in eukaryotic cells by controlling the translation (usually by repression), stability and localization of specific mRNA targets. Computational predictions of miRNA targets indicate that each miRNA regulates hundreds of mRNAs, and that approximately one third of all mammalian protein-coding genes are regulated by miRNAs. 2 Functional studies show that miRNAs participate in virtually each cellular process investigated, and that alterations in their expression levels might underlie human diseases, including cardiovascular disease and cancer. There are also data indicating that mammalian miRNAs can be imported into the nucleu...

MicroRNAs in liver cancer: a model for investigating pathogenesis and novel therapeutic approaches

“…RB1 is a target of miR-335. 15 miR-335 is downregulated during mesenchymal stem cell differentiation 16 and upregulated by the Wnt signaling pathway, 16 which is often activated in HCC. RB1 is also a target of miR-221, a miRNA that is frequently overexpressed in HCC.…”

Epigenetic Aspects of Osteoporosis

“…Наиболее изучено влияние мкРНК на транскрипционный фактор, участвующий в регулировании остеобластогенеза, RUNX2 (фактор транскрипции 2, содержащий домен Runt). К груп-пе мкРНК, блокирующих дифференцировку остеобластов посредством влияния на RUNX2, относятся мкРНК23а [37], мкРНК30а-d [38], мкРНК135а [39], мкРНК204 [40], мкРНК335 [41], мкРНК433 [42], мкРНК34с, мкРНК133а, мкРНК137, мкРНК205, мкРНК211, мкРНК217 [43,44]. Некоторые мкРНК со схожими функциями образуют кла-стеры: например, кластер мкРНК23а-27а-24-2, который является одним из основных в регуляции остеобластоге-неза.…”

“…Таргетная молекула Источник мкРНК, стимулирующие дифференцировку остеобластов мкРНК27, микроРНК142-3p APC [51,52] мкРНК29a Дкк1, KREMEN2, сФРЗ2, SPARC [48,54] мкРНК29b COL1A1, COL5A3, COL4A2, HDAC4, ТФРβ3, ACVR2A, CTNNBIP1, DUSP2 [55] мкРНК218 SOST, Дкк2, сФРЗ2 [49] мкРНК335-5p Дкк1 [50] мкРНК, блокирующие дифференцировку остеобластов мкРНК23a RUNX2, SATB2 [37,43,44] мкРНК30a-d RUNX2, SMAD1 [38,43,44] мкРНК34c, мкРНК133a, мкРНК137, мкРНК204, мкРНК205, мкРНК211, мкРНК217, мкРНК335, мкРНК338, мкРНК433, мкРНК3077-5p RUNX2 [39][40][41][42][43][44]53] мкРНК93 OSX [47] мкРНК135a RUNX2, SMAD5 [39,43,44] мкРНК155 SOCS1 [56] Примечание. APC -аденоматозная полипозная палочка; Дкк1 и 2 -Диккопф 1 и 2; KREMEN2 -кринглсодержащий трансмембран-ный белок; сФРЗ2 -секреторный белок, связанный с фризельдом 2; SPARC -секретируемый белок, кислый, богатый цистеином (остеонектин); COL1A1 -коллаген, тип I, α 1; COL5A3 -коллаген, тип V, α 3; COL4A2 -коллаген, тип IV, α 2; HDAC4 -гистонде-ацетилаза 4; ТФРβ 3 -трансформирующий фактор роста β 3 ; ACVR2A -рецептор типа 2А к активину А; CTNNBIP1 -белок, взаимо-действующий с β-катенином, 1; DUSP2 -фосфатаза двойной специфичности, тип 2; SOST -склеростин; RUNX2 -фактор транс-крипции 2, содержащий домен Runt; SATB2 -ДНК-связывающий белок; SMAD1 и 5 -семейство SMAD тип 1 и 5; OSX -остерикс; SOCS1 -супрессор цитокиновых сигналов 1.…”

MicroRNAs: Synthesis, Gene Regulation and Osteoblast Differentiation

“…There are several miRNAs namely miR-23a, miR-30c, miR-34c, miR-133a, miR-135a, miR-137, miR-204, miR-205, miR-217, and miR-338 that can target and decrease expression of Runx2 protein resulting inhibition of osteoblast differentiation (Hassan et al, 2010;. miR-355 was found to be directly targeting Runx2 mRNA, and overexpression of miR-335 in hMSCs inhibits their proliferation and migration, as well as their osteogenic and adipogenic potential (Tome et al, 2011).…”