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MicroRNA-145 regulates chondrogenic differentiation of mesenchymal stem cells by targeting Sox9.

Yang B, Guo H, Zhang Y, Chen L, Ying D, Dong S - PLoS ONE (2011)

Bottom Line: In addition, over-expression of miR-145 decreased expression of Sox9 only at protein levels and miR-145 inhibition significantly elevated Sox9 protein levels.Furthermore, over-expression of miR-145 decreased mRNA levels for three chondrogenic marker genes, type II collagen (Col2a1), aggrecan (Agc1), cartilage oligomeric matrix protein (COMP), type IX collagen (Col9a2) and type XI collagen (Col11a1) in C3H10T1/2 cells induced by TGF-β3, whereas anti-miR-145 inhibitor increased the expression of these chondrogenic marker genes.Thus, our studies demonstrated that miR-145 is a key negative regulator of chondrogenic differentiation by directly targeting Sox9 at early stage of chondrogenic differentiation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Biomechanics, Department of Anatomy, The Third Military Medical University, Chongqing, People's Republic of China.

ABSTRACT
Chondrogenic differentiation of mesenchymal stem cells (MSCs) is accurately regulated by essential transcription factors and signaling cascades. However, the precise mechanisms involved in this process still remain to be defined. MicroRNAs (miRNAs) regulate various biological processes by binding target mRNA to attenuate protein synthesis. To investigate the mechanisms for miRNAs-mediated regulation of chondrogenic differentiation, we identified that miR-145 was decreased during transforming growth factor beta 3 (TGF-β3)-induced chondrogenic differentiation of murine MSCs. Subsequently, dual-luciferase reporter gene assay data demonstrated that miR-145 targets a putative binding site in the 3'-UTR of SRY-related high mobility group-Box gene 9 (Sox9) gene, the key transcription factor for chondrogenesis. In addition, over-expression of miR-145 decreased expression of Sox9 only at protein levels and miR-145 inhibition significantly elevated Sox9 protein levels. Furthermore, over-expression of miR-145 decreased mRNA levels for three chondrogenic marker genes, type II collagen (Col2a1), aggrecan (Agc1), cartilage oligomeric matrix protein (COMP), type IX collagen (Col9a2) and type XI collagen (Col11a1) in C3H10T1/2 cells induced by TGF-β3, whereas anti-miR-145 inhibitor increased the expression of these chondrogenic marker genes. Thus, our studies demonstrated that miR-145 is a key negative regulator of chondrogenic differentiation by directly targeting Sox9 at early stage of chondrogenic differentiation.

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Differential expression of miR-145 during chondrogenic differentiation of murine MSCs.Down-regulated expression of miR-145 in murine MSCs induced by TGF-β3 was identified by qRT-PCR at different stage of chondrogenic differentiation (0 d, 7 d, 14 d). The relative expression level of miR-145 in untreated MSCs (0 d) was set to one, as control. Three independent cell culture experiments were done and data was represented as mean±sd. *, p<0.05, when compared with control.
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pone-0021679-g001: Differential expression of miR-145 during chondrogenic differentiation of murine MSCs.Down-regulated expression of miR-145 in murine MSCs induced by TGF-β3 was identified by qRT-PCR at different stage of chondrogenic differentiation (0 d, 7 d, 14 d). The relative expression level of miR-145 in untreated MSCs (0 d) was set to one, as control. Three independent cell culture experiments were done and data was represented as mean±sd. *, p<0.05, when compared with control.

Mentions: In our previous study, miRNA microarray technology was applied to detect miRNAs expression profiles of three different stages during chondrogenic differentiation, including murine MSCs, chondrogenic induction at 7 d, and 14 d after TGF-β3 treatment[23]. The expression of miR-145 significantly decreased during chondrogenic differentiation. Subsequently, we performed bioinformatic analyses to predict the target genes of miR-145 using Pictar [35] and Targetscan [36]. Noticeably, we found Sox9 was the potential target gene regulated by miR-145. According to the primary role of Sox9 in the process of MSCs differentiation into chondrocytes, we hypothesized that Sox9 may be inhibited by miR-145, which prevents MSCs from differentiating into chondrocytes. Furthermore, down-regulation of miR-145 may act as positive effect on chondrogenic differentiation. Thus qRT-PCR assay was performed to validate the expression pattern of miR-145 in this study. The results confirmed that miR-145 gradually decreased in MSCs, which were induced by TGF-β3 (Figure 1).


MicroRNA-145 regulates chondrogenic differentiation of mesenchymal stem cells by targeting Sox9.

Yang B, Guo H, Zhang Y, Chen L, Ying D, Dong S - PLoS ONE (2011)

Differential expression of miR-145 during chondrogenic differentiation of murine MSCs.Down-regulated expression of miR-145 in murine MSCs induced by TGF-β3 was identified by qRT-PCR at different stage of chondrogenic differentiation (0 d, 7 d, 14 d). The relative expression level of miR-145 in untreated MSCs (0 d) was set to one, as control. Three independent cell culture experiments were done and data was represented as mean±sd. *, p<0.05, when compared with control.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3140487&req=5

pone-0021679-g001: Differential expression of miR-145 during chondrogenic differentiation of murine MSCs.Down-regulated expression of miR-145 in murine MSCs induced by TGF-β3 was identified by qRT-PCR at different stage of chondrogenic differentiation (0 d, 7 d, 14 d). The relative expression level of miR-145 in untreated MSCs (0 d) was set to one, as control. Three independent cell culture experiments were done and data was represented as mean±sd. *, p<0.05, when compared with control.
Mentions: In our previous study, miRNA microarray technology was applied to detect miRNAs expression profiles of three different stages during chondrogenic differentiation, including murine MSCs, chondrogenic induction at 7 d, and 14 d after TGF-β3 treatment[23]. The expression of miR-145 significantly decreased during chondrogenic differentiation. Subsequently, we performed bioinformatic analyses to predict the target genes of miR-145 using Pictar [35] and Targetscan [36]. Noticeably, we found Sox9 was the potential target gene regulated by miR-145. According to the primary role of Sox9 in the process of MSCs differentiation into chondrocytes, we hypothesized that Sox9 may be inhibited by miR-145, which prevents MSCs from differentiating into chondrocytes. Furthermore, down-regulation of miR-145 may act as positive effect on chondrogenic differentiation. Thus qRT-PCR assay was performed to validate the expression pattern of miR-145 in this study. The results confirmed that miR-145 gradually decreased in MSCs, which were induced by TGF-β3 (Figure 1).

Bottom Line: In addition, over-expression of miR-145 decreased expression of Sox9 only at protein levels and miR-145 inhibition significantly elevated Sox9 protein levels.Furthermore, over-expression of miR-145 decreased mRNA levels for three chondrogenic marker genes, type II collagen (Col2a1), aggrecan (Agc1), cartilage oligomeric matrix protein (COMP), type IX collagen (Col9a2) and type XI collagen (Col11a1) in C3H10T1/2 cells induced by TGF-β3, whereas anti-miR-145 inhibitor increased the expression of these chondrogenic marker genes.Thus, our studies demonstrated that miR-145 is a key negative regulator of chondrogenic differentiation by directly targeting Sox9 at early stage of chondrogenic differentiation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Biomechanics, Department of Anatomy, The Third Military Medical University, Chongqing, People's Republic of China.

ABSTRACT
Chondrogenic differentiation of mesenchymal stem cells (MSCs) is accurately regulated by essential transcription factors and signaling cascades. However, the precise mechanisms involved in this process still remain to be defined. MicroRNAs (miRNAs) regulate various biological processes by binding target mRNA to attenuate protein synthesis. To investigate the mechanisms for miRNAs-mediated regulation of chondrogenic differentiation, we identified that miR-145 was decreased during transforming growth factor beta 3 (TGF-β3)-induced chondrogenic differentiation of murine MSCs. Subsequently, dual-luciferase reporter gene assay data demonstrated that miR-145 targets a putative binding site in the 3'-UTR of SRY-related high mobility group-Box gene 9 (Sox9) gene, the key transcription factor for chondrogenesis. In addition, over-expression of miR-145 decreased expression of Sox9 only at protein levels and miR-145 inhibition significantly elevated Sox9 protein levels. Furthermore, over-expression of miR-145 decreased mRNA levels for three chondrogenic marker genes, type II collagen (Col2a1), aggrecan (Agc1), cartilage oligomeric matrix protein (COMP), type IX collagen (Col9a2) and type XI collagen (Col11a1) in C3H10T1/2 cells induced by TGF-β3, whereas anti-miR-145 inhibitor increased the expression of these chondrogenic marker genes. Thus, our studies demonstrated that miR-145 is a key negative regulator of chondrogenic differentiation by directly targeting Sox9 at early stage of chondrogenic differentiation.

Show MeSH
Related in: MedlinePlus