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Experimental murine myopia induces collagen type Iα1 (COL1A1) DNA methylation and altered COL1A1 messenger RNA expression in sclera.

Zhou X, Ji F, An J, Zhao F, Shi F, Huang F, Li Y, Jiao S, Yan D, Chen X, Chen J, Qu J - Mol. Vis. (2012)

Bottom Line: MD was found to induce myopia in the treated eyes.Consistent with the CpG methylation, scleral COL1A1 mRNA was reduced by 57% in the MD-treated eyes compared to normal controls (p<0.05).After seven days of MD recovery, CpG methylation was significantly reduced (p=0.01).

View Article: PubMed Central - PubMed

Affiliation: School of Optometry & Ophthalmology and Eye Hospital, Wenzhou Medical College, Wenzhou, Zhejiang, China.

ABSTRACT

Purpose: To investigate whether myopia development is associated with changes of scleral DNA methylation in cytosine-phosphate-guanine (CpG) sites in the collagen 1A1 (COL1A1) promoter and messenger RNA (mRNA) levels following murine form deprivation myopia.

Methods: Fifty-seven C57BL/6 mice (postnatal day 23) were randomly assigned to four groups: (1) monocular form deprivation (MD) in which a diffuser lens was placed over one eye for 28 days; (2) normal controls without MD; (3) MD recovery in which the diffuser lens was removed for seven days; and (4) MD recovery normal controls. The DNA methylation pattern in COL1A1 promoter and exon 1 was determined by bisulfite DNA sequencing, and the COL1A1 mRNA level in sclera was determined by quantitative PCR.

Results: MD was found to induce myopia in the treated eyes. Six CpG sites in the promoter and exon 1 region of COL1A1 were methylated with significantly higher frequency in the treated eyes than normal control eyes (p<0.05), with CpG island methylation in MD-contralateral eyes being intermediate. Consistent with the CpG methylation, scleral COL1A1 mRNA was reduced by 57% in the MD-treated eyes compared to normal controls (p<0.05). After seven days of MD recovery, CpG methylation was significantly reduced (p=0.01). The methylation patterns returned to near normal level in five CpG sites, but the sixth was hypomethylated compared to normal controls.

Conclusions: In parallel with the development of myopia and the reduced COL1A1 mRNA, the frequency of methylation in CpG sites of the COL1A1 promoter/exon 1 increased during MD and returned to near normal during recovery. Thus, hypermethylation of CpG sites in the promoter/exon 1 of COL1A1 may underlie reduced collagen synthesis at the transcriptional level in myopic scleras.

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Proportion of sites that were methylated in the proximal promoter and a portion of exon 1. A: Form-deprived, contralateral control, and normal control eyes after four weeks of monocular form deprivation. B: Form-deprived, contralateral control, and normal control eyes after one week of recovery following four weeks of monocular deprivation (MD). Detailed maps of cytosine-phosphate-guanine (CpG) sites in the proximal promoter and first exon are shown. The beads in the horizontal lines illustrate the CpG sites, and the color of each indicates the corresponding degree of methylation: gray, 0–0.1; blue, 0.1–0.2; green, 0.2–0.3, red, >0.3. MD-T: monocular deprivation-treated eyes, MD-C: contralateral control eyes, NC: age-matched normal control eyes, Numbers: sample IDs.
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f2: Proportion of sites that were methylated in the proximal promoter and a portion of exon 1. A: Form-deprived, contralateral control, and normal control eyes after four weeks of monocular form deprivation. B: Form-deprived, contralateral control, and normal control eyes after one week of recovery following four weeks of monocular deprivation (MD). Detailed maps of cytosine-phosphate-guanine (CpG) sites in the proximal promoter and first exon are shown. The beads in the horizontal lines illustrate the CpG sites, and the color of each indicates the corresponding degree of methylation: gray, 0–0.1; blue, 0.1–0.2; green, 0.2–0.3, red, >0.3. MD-T: monocular deprivation-treated eyes, MD-C: contralateral control eyes, NC: age-matched normal control eyes, Numbers: sample IDs.

Mentions: DNA methylation profiles for MD-T, MD-C, and NC51 eyes were determined after four weeks of monocular form deprivation (Table 3). In MD-C and NC51 eyes, most of the CpG sites exhibited very low levels of DNA methylation, whereas in MD-T eyes, the levels were elevated at most of the sites (Figure 2A). The amount of methylation in MD-T eyes was higher than in MD-C eyes (Figure 3). The methylation percentages of six CpG sites (1, 3, 9, 14, 18, and 19) in MD-T eyes were significantly increased compared to the NC51 eyes (Figure 4). In MD-C eyes, the CpG sites were methylated at a level intermediate between the MD-T and NC51 eyes (Figure 3). The methylation percentages of four CpG sites (3, 8, 14, and 18) in MD-C eyes tended to increase compared to the NC51 eyes, although only site 14 was significantly increased (Figure 4).


Experimental murine myopia induces collagen type Iα1 (COL1A1) DNA methylation and altered COL1A1 messenger RNA expression in sclera.

Zhou X, Ji F, An J, Zhao F, Shi F, Huang F, Li Y, Jiao S, Yan D, Chen X, Chen J, Qu J - Mol. Vis. (2012)

Proportion of sites that were methylated in the proximal promoter and a portion of exon 1. A: Form-deprived, contralateral control, and normal control eyes after four weeks of monocular form deprivation. B: Form-deprived, contralateral control, and normal control eyes after one week of recovery following four weeks of monocular deprivation (MD). Detailed maps of cytosine-phosphate-guanine (CpG) sites in the proximal promoter and first exon are shown. The beads in the horizontal lines illustrate the CpG sites, and the color of each indicates the corresponding degree of methylation: gray, 0–0.1; blue, 0.1–0.2; green, 0.2–0.3, red, >0.3. MD-T: monocular deprivation-treated eyes, MD-C: contralateral control eyes, NC: age-matched normal control eyes, Numbers: sample IDs.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3369898&req=5

f2: Proportion of sites that were methylated in the proximal promoter and a portion of exon 1. A: Form-deprived, contralateral control, and normal control eyes after four weeks of monocular form deprivation. B: Form-deprived, contralateral control, and normal control eyes after one week of recovery following four weeks of monocular deprivation (MD). Detailed maps of cytosine-phosphate-guanine (CpG) sites in the proximal promoter and first exon are shown. The beads in the horizontal lines illustrate the CpG sites, and the color of each indicates the corresponding degree of methylation: gray, 0–0.1; blue, 0.1–0.2; green, 0.2–0.3, red, >0.3. MD-T: monocular deprivation-treated eyes, MD-C: contralateral control eyes, NC: age-matched normal control eyes, Numbers: sample IDs.
Mentions: DNA methylation profiles for MD-T, MD-C, and NC51 eyes were determined after four weeks of monocular form deprivation (Table 3). In MD-C and NC51 eyes, most of the CpG sites exhibited very low levels of DNA methylation, whereas in MD-T eyes, the levels were elevated at most of the sites (Figure 2A). The amount of methylation in MD-T eyes was higher than in MD-C eyes (Figure 3). The methylation percentages of six CpG sites (1, 3, 9, 14, 18, and 19) in MD-T eyes were significantly increased compared to the NC51 eyes (Figure 4). In MD-C eyes, the CpG sites were methylated at a level intermediate between the MD-T and NC51 eyes (Figure 3). The methylation percentages of four CpG sites (3, 8, 14, and 18) in MD-C eyes tended to increase compared to the NC51 eyes, although only site 14 was significantly increased (Figure 4).

Bottom Line: MD was found to induce myopia in the treated eyes.Consistent with the CpG methylation, scleral COL1A1 mRNA was reduced by 57% in the MD-treated eyes compared to normal controls (p<0.05).After seven days of MD recovery, CpG methylation was significantly reduced (p=0.01).

View Article: PubMed Central - PubMed

Affiliation: School of Optometry & Ophthalmology and Eye Hospital, Wenzhou Medical College, Wenzhou, Zhejiang, China.

ABSTRACT

Purpose: To investigate whether myopia development is associated with changes of scleral DNA methylation in cytosine-phosphate-guanine (CpG) sites in the collagen 1A1 (COL1A1) promoter and messenger RNA (mRNA) levels following murine form deprivation myopia.

Methods: Fifty-seven C57BL/6 mice (postnatal day 23) were randomly assigned to four groups: (1) monocular form deprivation (MD) in which a diffuser lens was placed over one eye for 28 days; (2) normal controls without MD; (3) MD recovery in which the diffuser lens was removed for seven days; and (4) MD recovery normal controls. The DNA methylation pattern in COL1A1 promoter and exon 1 was determined by bisulfite DNA sequencing, and the COL1A1 mRNA level in sclera was determined by quantitative PCR.

Results: MD was found to induce myopia in the treated eyes. Six CpG sites in the promoter and exon 1 region of COL1A1 were methylated with significantly higher frequency in the treated eyes than normal control eyes (p<0.05), with CpG island methylation in MD-contralateral eyes being intermediate. Consistent with the CpG methylation, scleral COL1A1 mRNA was reduced by 57% in the MD-treated eyes compared to normal controls (p<0.05). After seven days of MD recovery, CpG methylation was significantly reduced (p=0.01). The methylation patterns returned to near normal level in five CpG sites, but the sixth was hypomethylated compared to normal controls.

Conclusions: In parallel with the development of myopia and the reduced COL1A1 mRNA, the frequency of methylation in CpG sites of the COL1A1 promoter/exon 1 increased during MD and returned to near normal during recovery. Thus, hypermethylation of CpG sites in the promoter/exon 1 of COL1A1 may underlie reduced collagen synthesis at the transcriptional level in myopic scleras.

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Related in: MedlinePlus