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MicroRNA-132/212 family enhances arteriogenesis after hindlimb ischaemia through modulation of the Ras-MAPK pathway.

Lei Z, van Mil A, Brandt MM, Grundmann S, Hoefer I, Smits M, El Azzouzi H, Fukao T, Cheng C, Doevendans PA, Sluijter JP - J. Cell. Mol. Med. (2015)

Bottom Line: Moreover, in in vitro pericyte-endothelial co-culture cell assays, overexpression of miR-132 and mir-212 in endothelial cells results in enhanced vascularization, as shown by an increase in tubular structures and junctions.Our results suggested that miR-132/212 may exert their effects by enhancing the Ras-Mitogen-activated protein kinases MAPK signalling pathway through direct inhibition of Rasa1, and Spred1.The miR-132/212 cluster promotes arteriogenesis by modulating Ras-MAPK signalling via direct targeting of its inhibitors Rasa1 and Spred1.

View Article: PubMed Central - PubMed

Affiliation: Division Heart and Lungs, Department of Cardiology, University Medical Center Utrecht, Utrecht, The Netherlands.

No MeSH data available.


Related in: MedlinePlus

Expression of miR-132/212 targets and phosophorylated ERK1/2 in ischaemia limb. (A) Spred1, Rasa1 and Spry1 expression as determined by Western blot, normalized by β-Actin. (B) Quantification Spred1, Rasa1 and Spry1 expression as determined by Western blot in B. (C) Phosophorylated ERK1/2 expression in the thigh WT and KO mice on 14 days after hindlimb ischaemia as determined by Western blot on day 14. (D) Quantification of phosophorylated ERK1/2 expression in the thigh WT and KO mice on 14 days after hindlimb ischaemia as determined by Western blot, normalized by β-Actin.
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fig07: Expression of miR-132/212 targets and phosophorylated ERK1/2 in ischaemia limb. (A) Spred1, Rasa1 and Spry1 expression as determined by Western blot, normalized by β-Actin. (B) Quantification Spred1, Rasa1 and Spry1 expression as determined by Western blot in B. (C) Phosophorylated ERK1/2 expression in the thigh WT and KO mice on 14 days after hindlimb ischaemia as determined by Western blot on day 14. (D) Quantification of phosophorylated ERK1/2 expression in the thigh WT and KO mice on 14 days after hindlimb ischaemia as determined by Western blot, normalized by β-Actin.

Mentions: We reasoned that if Spred1, Spry1 and Rasa1 are in vivo targets of miR-132 or 212, they should be expressed in the arteries of the thigh muscle. By immunofluorescent staining, we observed that Spred1, Spry1 and Rasa1 could all be detected in the vascular wall (Fig. S2). By comparing the expression of Spred1, Spry1 and Rasa1 in the adductor muscle from WT and KO after femoral artery ligation using Western blot, we observed that Spred1 and Rasa1 are significantly more present in the KO mice (Fig.7A and B). Surprisingly we found that there is no difference in the Spry1 protein between WT and KO mice (Fig.7A and B). Next we asked if higher level of Spred1 and Rasa1 expression could have an effect on the Ras-MAPK pathway. By Western blotting we detected lower phosphorylated ERK1/2 in the area of blood vessel growth in KO mice 14 days after hindlimb ischaemia (Fig.7C and D).


MicroRNA-132/212 family enhances arteriogenesis after hindlimb ischaemia through modulation of the Ras-MAPK pathway.

Lei Z, van Mil A, Brandt MM, Grundmann S, Hoefer I, Smits M, El Azzouzi H, Fukao T, Cheng C, Doevendans PA, Sluijter JP - J. Cell. Mol. Med. (2015)

Expression of miR-132/212 targets and phosophorylated ERK1/2 in ischaemia limb. (A) Spred1, Rasa1 and Spry1 expression as determined by Western blot, normalized by β-Actin. (B) Quantification Spred1, Rasa1 and Spry1 expression as determined by Western blot in B. (C) Phosophorylated ERK1/2 expression in the thigh WT and KO mice on 14 days after hindlimb ischaemia as determined by Western blot on day 14. (D) Quantification of phosophorylated ERK1/2 expression in the thigh WT and KO mice on 14 days after hindlimb ischaemia as determined by Western blot, normalized by β-Actin.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4549050&req=5

fig07: Expression of miR-132/212 targets and phosophorylated ERK1/2 in ischaemia limb. (A) Spred1, Rasa1 and Spry1 expression as determined by Western blot, normalized by β-Actin. (B) Quantification Spred1, Rasa1 and Spry1 expression as determined by Western blot in B. (C) Phosophorylated ERK1/2 expression in the thigh WT and KO mice on 14 days after hindlimb ischaemia as determined by Western blot on day 14. (D) Quantification of phosophorylated ERK1/2 expression in the thigh WT and KO mice on 14 days after hindlimb ischaemia as determined by Western blot, normalized by β-Actin.
Mentions: We reasoned that if Spred1, Spry1 and Rasa1 are in vivo targets of miR-132 or 212, they should be expressed in the arteries of the thigh muscle. By immunofluorescent staining, we observed that Spred1, Spry1 and Rasa1 could all be detected in the vascular wall (Fig. S2). By comparing the expression of Spred1, Spry1 and Rasa1 in the adductor muscle from WT and KO after femoral artery ligation using Western blot, we observed that Spred1 and Rasa1 are significantly more present in the KO mice (Fig.7A and B). Surprisingly we found that there is no difference in the Spry1 protein between WT and KO mice (Fig.7A and B). Next we asked if higher level of Spred1 and Rasa1 expression could have an effect on the Ras-MAPK pathway. By Western blotting we detected lower phosphorylated ERK1/2 in the area of blood vessel growth in KO mice 14 days after hindlimb ischaemia (Fig.7C and D).

Bottom Line: Moreover, in in vitro pericyte-endothelial co-culture cell assays, overexpression of miR-132 and mir-212 in endothelial cells results in enhanced vascularization, as shown by an increase in tubular structures and junctions.Our results suggested that miR-132/212 may exert their effects by enhancing the Ras-Mitogen-activated protein kinases MAPK signalling pathway through direct inhibition of Rasa1, and Spred1.The miR-132/212 cluster promotes arteriogenesis by modulating Ras-MAPK signalling via direct targeting of its inhibitors Rasa1 and Spred1.

View Article: PubMed Central - PubMed

Affiliation: Division Heart and Lungs, Department of Cardiology, University Medical Center Utrecht, Utrecht, The Netherlands.

No MeSH data available.


Related in: MedlinePlus