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miR-146a targets Fos expression in human cardiac cells.

Palomer X, Capdevila-Busquets E, Botteri G, Davidson MM, Rodríguez C, Martínez-González J, Vidal F, Barroso E, Chan TO, Feldman AM, Vázquez-Carrera M - Dis Model Mech (2015)

Bottom Line: These changes correlated with a diminution in the DNA-binding activity of AP-1, the Fos-containing transcription factor complex.The specific regulation of this MMP by miR-146a was further confirmed at the secretion and enzymatic activity levels, as well as after anti-miR-mediated miR-146a inhibition.The results reported here demonstrate that Fos is a direct target of miR-146a activity and that downregulation of the Fos-AP-1 pathway by miR-146a has the capacity to inhibit MMP-9 activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Therapeutic Chemistry, IBUB (Institut de Biomedicina de la Universitat de Barcelona) and CIBER de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Faculty of Pharmacy, University of Barcelona, Diagonal 643, Barcelona E-08028, Spain.

No MeSH data available.


Related in: MedlinePlus

miR-146a downregulates MMP-9 expression, secretion and activity in human cardiac cells. Relative quantification of MMP-2 and MMP-9 mRNA levels in non-differentiated human cardiac AC16 cells transfected with: (A) lacZ- or miR-146a-carrying plasmids, or (B) a human anti-miR-146a inhibitor or an anti-miR negative control (Anti-miR). The graphs represent the quantification of 18S-normalized mRNA levels, expressed as a percentage of control samples ±s.d. (C) Spearman rank correlation between TNF-α and miR-146a, miR-146a and Fos, and miR-146a and MMP-9 gene expression in left ventricular tissue obtained from patients undergoing heart transplantation. The relative transcript levels of the target genes, in arbitrary units, were used to calculate the Spearman correlation coefficients (n.s., non-significant). (D) Determination by ELISA of MMP-9 secretion into the culture media in AC16 cells transfected with lacZ- or miR-146a-carrying plasmids. (E) Representative gel zymography and corresponding densitometric analysis of MMP-9 gelatinolytic activity in culture media of cells transfected with lacZ- or miR-146a-carrying plasmids. MW, molecular weight; rMMP-9, human recombinant MMP-9. (A,D,E) *P<0.05, **P<0.01 and ***P<0.001 vs lacZ; (B) *P<0.05 and ***P<0.001 vs Anti-miR–TNF-α; ‡P<0.05 vs Anti-miR+TNF-α.
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DMM020768F5: miR-146a downregulates MMP-9 expression, secretion and activity in human cardiac cells. Relative quantification of MMP-2 and MMP-9 mRNA levels in non-differentiated human cardiac AC16 cells transfected with: (A) lacZ- or miR-146a-carrying plasmids, or (B) a human anti-miR-146a inhibitor or an anti-miR negative control (Anti-miR). The graphs represent the quantification of 18S-normalized mRNA levels, expressed as a percentage of control samples ±s.d. (C) Spearman rank correlation between TNF-α and miR-146a, miR-146a and Fos, and miR-146a and MMP-9 gene expression in left ventricular tissue obtained from patients undergoing heart transplantation. The relative transcript levels of the target genes, in arbitrary units, were used to calculate the Spearman correlation coefficients (n.s., non-significant). (D) Determination by ELISA of MMP-9 secretion into the culture media in AC16 cells transfected with lacZ- or miR-146a-carrying plasmids. (E) Representative gel zymography and corresponding densitometric analysis of MMP-9 gelatinolytic activity in culture media of cells transfected with lacZ- or miR-146a-carrying plasmids. MW, molecular weight; rMMP-9, human recombinant MMP-9. (A,D,E) *P<0.05, **P<0.01 and ***P<0.001 vs lacZ; (B) *P<0.05 and ***P<0.001 vs Anti-miR–TNF-α; ‡P<0.05 vs Anti-miR+TNF-α.

Mentions: Examination of MMP-9 expression in human cardiac cells transfected with miR-146a revealed that, unlike MMP-2, its transcript levels were downregulated (∼40% reduction, P<0.05, Fig. 5A). According to these results, anti-miR-146a stimulated MMP-9 expression regardless of the presence (1.5-fold, P<0.001) or absence (1.5-fold, P<0.001) of TNF-α (Fig. 5B). We next examined this issue in left ventricular tissue obtained from patients undergoing heart transplantation. The relative expression of miR-146a positively correlated with that of TNF-α (Fig. 5C, top panel, Spearman rank correlation r=0.5480), a finding that, despite being only marginally significant (P=0.08), matched the previous results obtained in AC16 cells fairly closely. Of note, miR-146a levels negatively correlated with the expression of Fos (Fig. 5C, middle panel, Spearman rank correlation r=−0.6208, P<0.05) and MMP-9 (Fig. 5C, bottom panel, r=−0.5659, P<0.05) in these patients. Finally, we aimed to examine the effects of MMP-9 downregulation by miR-146a on its enzymatic activity. As expected, miR-146a overexpression elicited a reduction in MMP-9 secretion to the media (∼25% reduction, P<0.001, Fig. 5D) and MMP-9 activity (∼35% reduction, P<0.05, Fig. 5E). The zymogram also yielded a stronger gelatinolytic activity band, which was concentrated in the area corresponding to the molecular mass of MMP-2 (62 kDa), which was not modified under our conditions.Fig. 5.


miR-146a targets Fos expression in human cardiac cells.

Palomer X, Capdevila-Busquets E, Botteri G, Davidson MM, Rodríguez C, Martínez-González J, Vidal F, Barroso E, Chan TO, Feldman AM, Vázquez-Carrera M - Dis Model Mech (2015)

miR-146a downregulates MMP-9 expression, secretion and activity in human cardiac cells. Relative quantification of MMP-2 and MMP-9 mRNA levels in non-differentiated human cardiac AC16 cells transfected with: (A) lacZ- or miR-146a-carrying plasmids, or (B) a human anti-miR-146a inhibitor or an anti-miR negative control (Anti-miR). The graphs represent the quantification of 18S-normalized mRNA levels, expressed as a percentage of control samples ±s.d. (C) Spearman rank correlation between TNF-α and miR-146a, miR-146a and Fos, and miR-146a and MMP-9 gene expression in left ventricular tissue obtained from patients undergoing heart transplantation. The relative transcript levels of the target genes, in arbitrary units, were used to calculate the Spearman correlation coefficients (n.s., non-significant). (D) Determination by ELISA of MMP-9 secretion into the culture media in AC16 cells transfected with lacZ- or miR-146a-carrying plasmids. (E) Representative gel zymography and corresponding densitometric analysis of MMP-9 gelatinolytic activity in culture media of cells transfected with lacZ- or miR-146a-carrying plasmids. MW, molecular weight; rMMP-9, human recombinant MMP-9. (A,D,E) *P<0.05, **P<0.01 and ***P<0.001 vs lacZ; (B) *P<0.05 and ***P<0.001 vs Anti-miR–TNF-α; ‡P<0.05 vs Anti-miR+TNF-α.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4582106&req=5

DMM020768F5: miR-146a downregulates MMP-9 expression, secretion and activity in human cardiac cells. Relative quantification of MMP-2 and MMP-9 mRNA levels in non-differentiated human cardiac AC16 cells transfected with: (A) lacZ- or miR-146a-carrying plasmids, or (B) a human anti-miR-146a inhibitor or an anti-miR negative control (Anti-miR). The graphs represent the quantification of 18S-normalized mRNA levels, expressed as a percentage of control samples ±s.d. (C) Spearman rank correlation between TNF-α and miR-146a, miR-146a and Fos, and miR-146a and MMP-9 gene expression in left ventricular tissue obtained from patients undergoing heart transplantation. The relative transcript levels of the target genes, in arbitrary units, were used to calculate the Spearman correlation coefficients (n.s., non-significant). (D) Determination by ELISA of MMP-9 secretion into the culture media in AC16 cells transfected with lacZ- or miR-146a-carrying plasmids. (E) Representative gel zymography and corresponding densitometric analysis of MMP-9 gelatinolytic activity in culture media of cells transfected with lacZ- or miR-146a-carrying plasmids. MW, molecular weight; rMMP-9, human recombinant MMP-9. (A,D,E) *P<0.05, **P<0.01 and ***P<0.001 vs lacZ; (B) *P<0.05 and ***P<0.001 vs Anti-miR–TNF-α; ‡P<0.05 vs Anti-miR+TNF-α.
Mentions: Examination of MMP-9 expression in human cardiac cells transfected with miR-146a revealed that, unlike MMP-2, its transcript levels were downregulated (∼40% reduction, P<0.05, Fig. 5A). According to these results, anti-miR-146a stimulated MMP-9 expression regardless of the presence (1.5-fold, P<0.001) or absence (1.5-fold, P<0.001) of TNF-α (Fig. 5B). We next examined this issue in left ventricular tissue obtained from patients undergoing heart transplantation. The relative expression of miR-146a positively correlated with that of TNF-α (Fig. 5C, top panel, Spearman rank correlation r=0.5480), a finding that, despite being only marginally significant (P=0.08), matched the previous results obtained in AC16 cells fairly closely. Of note, miR-146a levels negatively correlated with the expression of Fos (Fig. 5C, middle panel, Spearman rank correlation r=−0.6208, P<0.05) and MMP-9 (Fig. 5C, bottom panel, r=−0.5659, P<0.05) in these patients. Finally, we aimed to examine the effects of MMP-9 downregulation by miR-146a on its enzymatic activity. As expected, miR-146a overexpression elicited a reduction in MMP-9 secretion to the media (∼25% reduction, P<0.001, Fig. 5D) and MMP-9 activity (∼35% reduction, P<0.05, Fig. 5E). The zymogram also yielded a stronger gelatinolytic activity band, which was concentrated in the area corresponding to the molecular mass of MMP-2 (62 kDa), which was not modified under our conditions.Fig. 5.

Bottom Line: These changes correlated with a diminution in the DNA-binding activity of AP-1, the Fos-containing transcription factor complex.The specific regulation of this MMP by miR-146a was further confirmed at the secretion and enzymatic activity levels, as well as after anti-miR-mediated miR-146a inhibition.The results reported here demonstrate that Fos is a direct target of miR-146a activity and that downregulation of the Fos-AP-1 pathway by miR-146a has the capacity to inhibit MMP-9 activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Therapeutic Chemistry, IBUB (Institut de Biomedicina de la Universitat de Barcelona) and CIBER de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Faculty of Pharmacy, University of Barcelona, Diagonal 643, Barcelona E-08028, Spain.

No MeSH data available.


Related in: MedlinePlus