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Inorganic phosphate accelerates the migration of vascular smooth muscle cells: evidence for the involvement of miR-223.

Rangrez AY, M'Baya-Moutoula E, Metzinger-Le Meuth V, Hénaut L, Djelouat MS, Benchitrit J, Massy ZA, Metzinger L - PLoS ONE (2012)

Bottom Line: Interestingly, we also found that miR-223 (a marker of muscle damage and a key factor in osteoclast differentiation) is expressed in VSMCs and is significantly upregulated in Pi-treated cells.Additionally, we found that the expression of two of the known miR-223 targets, Mef2c and RhoB, was highly reduced in Pi treated as well as miR-223 over-expressing VSMCs.Our results suggest that (i) high levels of Pi increase VSMC migration and calcification, (ii) altered expression levels of miR-223 could play a part in this process and (iii) miR-223 is a potential new biomarker of VSMC damage.

View Article: PubMed Central - PubMed

Affiliation: INSERM U1088, Amiens, France.

ABSTRACT

Background: An elevated serum inorganic phosphate (Pi) level is a major risk factor for kidney disease and downstream vascular complications. We focused on the effect of Pi levels on human aortic vascular smooth muscle cells (VSMCs), with an emphasis on the role of microRNAs (miRNAs).

Methodology/principal findings: Exposure of human primary VSMCs in vitro to pathological levels of Pi increased calcification, migration rate and concomitantly reduced cell proliferation and the amount of the actin cytoskeleton. These changes were evidenced by significant downregulation of miRNA-143 (miR-143) and miR-145 and concomitant upregulation of their targets and key markers in synthetic VSMCs, such as Krüppel-like factors-4 and -5 and versican. Interestingly, we also found that miR-223 (a marker of muscle damage and a key factor in osteoclast differentiation) is expressed in VSMCs and is significantly upregulated in Pi-treated cells. Over-expressing miR-223 in VSMCs increased proliferation and markedly enhanced VSMC migration. Additionally, we found that the expression of two of the known miR-223 targets, Mef2c and RhoB, was highly reduced in Pi treated as well as miR-223 over-expressing VSMCs. To complement these in vitro findings, we also observed significant downregulation of miR-143 and miR-145 and upregulation of miR-223 in aorta samples collected from ApoE knock-out mice, which display vascular calcification.

Conclusions/significance: Our results suggest that (i) high levels of Pi increase VSMC migration and calcification, (ii) altered expression levels of miR-223 could play a part in this process and (iii) miR-223 is a potential new biomarker of VSMC damage.

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Related in: MedlinePlus

Expression of miR-143, miR-145 and miR-223 in wild-type and ApoE-KO mice.RNAs extracted from mouse aorta collected from 8- and 20-week-old ApoE-KO and wild-type mice were used for the qPCR expression analysis of miR-143, miR-145 and miR-223. There was significant downregulation of miR-143 and miR-145 and concomitant upregulation of miR-223 in 20-week-old ApoE-KO mice. Data are presented as the mean ± SD of 4 mice for each group.
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pone-0047807-g006: Expression of miR-143, miR-145 and miR-223 in wild-type and ApoE-KO mice.RNAs extracted from mouse aorta collected from 8- and 20-week-old ApoE-KO and wild-type mice were used for the qPCR expression analysis of miR-143, miR-145 and miR-223. There was significant downregulation of miR-143 and miR-145 and concomitant upregulation of miR-223 in 20-week-old ApoE-KO mice. Data are presented as the mean ± SD of 4 mice for each group.

Mentions: Lastly, we sought to study the fate of miR-143, miR-145 and miR-223 under in vivo vascular calcification conditions. We evaluated the levels of these miRNAs in aorta isolated from 8-week-old and 20-week-old ApoE-KO and WT mice from the same genetic background (C57BL6J, Figure 6). Our group has earlier published that 20-week-old ApoE-KO mice display more vascular calcification than WT mice[23]. The disease state in ApoE-KO mice was checked by determining blood cholesterol and triglycerides, which were constantly elevated (not shown). No significant differences in expression of either of the miRNAs were observed in young, 8-week-old animals (Figure 5A). In contrast, significantly greater expression of miR-223 (1.6-fold) was observed in older, 20-week-old ApoE-KO mice (which displayed vascular calcification), relative to WT mice (Figure 5B). As observed with VSMCs in culture, the expression levels of both miR-143 and miR-145 were significantly lower in ApoE-KO mice than in WT mice. This highly significant, approximately 2-fold difference was observed at the age of 20 weeks (Figure 5B). Our in vivo results in a well-established murine model thus reflected our in vitro findings, i.e. downregulation of miR-143 and miR-145 and upregulation of miR-223 in the presence of calcium-Pi deposits.


Inorganic phosphate accelerates the migration of vascular smooth muscle cells: evidence for the involvement of miR-223.

Rangrez AY, M'Baya-Moutoula E, Metzinger-Le Meuth V, Hénaut L, Djelouat MS, Benchitrit J, Massy ZA, Metzinger L - PLoS ONE (2012)

Expression of miR-143, miR-145 and miR-223 in wild-type and ApoE-KO mice.RNAs extracted from mouse aorta collected from 8- and 20-week-old ApoE-KO and wild-type mice were used for the qPCR expression analysis of miR-143, miR-145 and miR-223. There was significant downregulation of miR-143 and miR-145 and concomitant upregulation of miR-223 in 20-week-old ApoE-KO mice. Data are presented as the mean ± SD of 4 mice for each group.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3475714&req=5

pone-0047807-g006: Expression of miR-143, miR-145 and miR-223 in wild-type and ApoE-KO mice.RNAs extracted from mouse aorta collected from 8- and 20-week-old ApoE-KO and wild-type mice were used for the qPCR expression analysis of miR-143, miR-145 and miR-223. There was significant downregulation of miR-143 and miR-145 and concomitant upregulation of miR-223 in 20-week-old ApoE-KO mice. Data are presented as the mean ± SD of 4 mice for each group.
Mentions: Lastly, we sought to study the fate of miR-143, miR-145 and miR-223 under in vivo vascular calcification conditions. We evaluated the levels of these miRNAs in aorta isolated from 8-week-old and 20-week-old ApoE-KO and WT mice from the same genetic background (C57BL6J, Figure 6). Our group has earlier published that 20-week-old ApoE-KO mice display more vascular calcification than WT mice[23]. The disease state in ApoE-KO mice was checked by determining blood cholesterol and triglycerides, which were constantly elevated (not shown). No significant differences in expression of either of the miRNAs were observed in young, 8-week-old animals (Figure 5A). In contrast, significantly greater expression of miR-223 (1.6-fold) was observed in older, 20-week-old ApoE-KO mice (which displayed vascular calcification), relative to WT mice (Figure 5B). As observed with VSMCs in culture, the expression levels of both miR-143 and miR-145 were significantly lower in ApoE-KO mice than in WT mice. This highly significant, approximately 2-fold difference was observed at the age of 20 weeks (Figure 5B). Our in vivo results in a well-established murine model thus reflected our in vitro findings, i.e. downregulation of miR-143 and miR-145 and upregulation of miR-223 in the presence of calcium-Pi deposits.

Bottom Line: Interestingly, we also found that miR-223 (a marker of muscle damage and a key factor in osteoclast differentiation) is expressed in VSMCs and is significantly upregulated in Pi-treated cells.Additionally, we found that the expression of two of the known miR-223 targets, Mef2c and RhoB, was highly reduced in Pi treated as well as miR-223 over-expressing VSMCs.Our results suggest that (i) high levels of Pi increase VSMC migration and calcification, (ii) altered expression levels of miR-223 could play a part in this process and (iii) miR-223 is a potential new biomarker of VSMC damage.

View Article: PubMed Central - PubMed

Affiliation: INSERM U1088, Amiens, France.

ABSTRACT

Background: An elevated serum inorganic phosphate (Pi) level is a major risk factor for kidney disease and downstream vascular complications. We focused on the effect of Pi levels on human aortic vascular smooth muscle cells (VSMCs), with an emphasis on the role of microRNAs (miRNAs).

Methodology/principal findings: Exposure of human primary VSMCs in vitro to pathological levels of Pi increased calcification, migration rate and concomitantly reduced cell proliferation and the amount of the actin cytoskeleton. These changes were evidenced by significant downregulation of miRNA-143 (miR-143) and miR-145 and concomitant upregulation of their targets and key markers in synthetic VSMCs, such as Krüppel-like factors-4 and -5 and versican. Interestingly, we also found that miR-223 (a marker of muscle damage and a key factor in osteoclast differentiation) is expressed in VSMCs and is significantly upregulated in Pi-treated cells. Over-expressing miR-223 in VSMCs increased proliferation and markedly enhanced VSMC migration. Additionally, we found that the expression of two of the known miR-223 targets, Mef2c and RhoB, was highly reduced in Pi treated as well as miR-223 over-expressing VSMCs. To complement these in vitro findings, we also observed significant downregulation of miR-143 and miR-145 and upregulation of miR-223 in aorta samples collected from ApoE knock-out mice, which display vascular calcification.

Conclusions/significance: Our results suggest that (i) high levels of Pi increase VSMC migration and calcification, (ii) altered expression levels of miR-223 could play a part in this process and (iii) miR-223 is a potential new biomarker of VSMC damage.

Show MeSH
Related in: MedlinePlus