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Placental growth factor contributes to micro-vascular abnormalization and blood-retinal barrier breakdown in diabetic retinopathy.

Kowalczuk L, Touchard E, Omri S, Jonet L, Klein C, Valamanes F, Berdugo M, Bigey P, Massin P, Jeanny JC, Behar-Cohen F - PLoS ONE (2011)

Bottom Line: For a better understanding of its role on the retina, we have evaluated the effect of a sustained PGF over-expression in rat ocular media, using ciliary muscle electrotransfer (ET) of a plasmid encoding rat PGF-1 (pVAX2-rPGF-1). pVAX2-rPGF-1 ET in the ciliary muscle (200 V/cm) was achieved in non diabetic and diabetic rat eyes.After the control of rPGF-1 expression, PGF-induced effects on retinal vasculature and on the blood-external barrier were evaluated respectively by lectin and occludin staining on flat-mounts.PGF and its receptor Flt-1 may therefore be looked upon as a potential regulatory target at this stage of the disease.

View Article: PubMed Central - PubMed

Affiliation: Institut National pour la Santé Et la Recherche Médicale U872, Physiopathology of ocular diseases: Therapeutic innovations, Paris, France.

ABSTRACT

Objective: There are controversies regarding the pro-angiogenic activity of placental growth factor (PGF) in diabetic retinopathy (DR). For a better understanding of its role on the retina, we have evaluated the effect of a sustained PGF over-expression in rat ocular media, using ciliary muscle electrotransfer (ET) of a plasmid encoding rat PGF-1 (pVAX2-rPGF-1).

Materials and methods: pVAX2-rPGF-1 ET in the ciliary muscle (200 V/cm) was achieved in non diabetic and diabetic rat eyes. Control eyes received saline or naked plasmid ET. Clinical follow up was carried out over three months using slit lamp examination and fluorescein angiography. After the control of rPGF-1 expression, PGF-induced effects on retinal vasculature and on the blood-external barrier were evaluated respectively by lectin and occludin staining on flat-mounts. Ocular structures were visualized through histological analysis.

Results: After fifteen days of rPGF-1 over-expression in normal eyes, tortuous and dilated capillaries were observed. At one month, microaneurysms and moderate vascular sprouts were detected in mid retinal periphery in vivo and on retinal flat-mounts. At later stages, retinal pigmented epithelial cells demonstrated morphological abnormalities and junction ruptures. In diabetic retinas, PGF expression rose between 2 and 5 months, and, one month after ET, rPGF-1 over-expression induced glial activation and proliferation.

Conclusion: This is the first demonstration that sustained intraocular PGF production induces vascular and retinal changes similar to those observed in the early stages of diabetic retinopathy. PGF and its receptor Flt-1 may therefore be looked upon as a potential regulatory target at this stage of the disease.

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PGF expression in diabetic retina.(a–b) Comparison of PGF staining in non diabetic (a) and diabetic (b) retinas. (a) Sections of eyes from adult control non diabetic rat showed co-expression of PGF and GFAP in glial Muller cells from the gcl (arrowheads) to the inl, and PGF expression in glial Müller cells which are not immuno-reactive for GFAP (white star). Scale bar = 50 µm. (b) A similar pattern was observed on retinal sections of eyes from three-month-old diabetic rats, with a strong immuno-reactivity for PGF at the gcl level. (c) PGF detection by Western-blot in diabetic and non-diabetic retinas, from 1, 2, 5 and 12 month-old rats. For each lane in which 40 µg of proteins were deposited, the blood sugar level of the represented rats is indicated between parentheses. (d–e) Immunostaining for PGF and GFAP in sections from pVAX2-rPGF-1 ET- treated diabetic rat eyes, one month after ET. Sections show PGF-expressing infiltrating cells in the sub retinal space (d, arrows) and confirmed PGF expression by RMG cells (d, arrowheads). GFAP staining showed gliosis induced by RMG cells (d, e). ch, choroid; gcl, ganglion cell layer; inl, inner nuclear layer; ipl, inner plexiform layer; onl, outer nuclear layer; rpe, retinal pigmented epithelium; Scale bar = 100 µm.
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pone-0017462-g006: PGF expression in diabetic retina.(a–b) Comparison of PGF staining in non diabetic (a) and diabetic (b) retinas. (a) Sections of eyes from adult control non diabetic rat showed co-expression of PGF and GFAP in glial Muller cells from the gcl (arrowheads) to the inl, and PGF expression in glial Müller cells which are not immuno-reactive for GFAP (white star). Scale bar = 50 µm. (b) A similar pattern was observed on retinal sections of eyes from three-month-old diabetic rats, with a strong immuno-reactivity for PGF at the gcl level. (c) PGF detection by Western-blot in diabetic and non-diabetic retinas, from 1, 2, 5 and 12 month-old rats. For each lane in which 40 µg of proteins were deposited, the blood sugar level of the represented rats is indicated between parentheses. (d–e) Immunostaining for PGF and GFAP in sections from pVAX2-rPGF-1 ET- treated diabetic rat eyes, one month after ET. Sections show PGF-expressing infiltrating cells in the sub retinal space (d, arrows) and confirmed PGF expression by RMG cells (d, arrowheads). GFAP staining showed gliosis induced by RMG cells (d, e). ch, choroid; gcl, ganglion cell layer; inl, inner nuclear layer; ipl, inner plexiform layer; onl, outer nuclear layer; rpe, retinal pigmented epithelium; Scale bar = 100 µm.

Mentions: In non-diabetic untreated eyes, PGF immuno-localization showed that PGF-1 is expressed in retinal Müller glial (RMG) cells and in the inner segments of photoreceptors (Fig. 6a). In three-month-old GK diabetic rats, PGF-1 was strongly expressed in the inner part the retina, mostly in astrocytes and in swollen RMG cells, as shown by the co labeling with the glial marker, GFAP (Fig. 6b). PGF expression over time in diabetic retinas was compared to PGF expression in control retinas using western-blot (Fig. 6c). This kinetic showed that PGF expression increased in the neuroretinas from diabetic rats aged between two and five months, and remained elevated as the diabetes develops at later stages (12 months), whereas it remained constant between one month and one year in retinas from non diabetic control rats. PGF-1 is therefore upregulated in the retina of GK diabetic rats, increasing at the early stages of diabetes and remaining elevated thereafter.


Placental growth factor contributes to micro-vascular abnormalization and blood-retinal barrier breakdown in diabetic retinopathy.

Kowalczuk L, Touchard E, Omri S, Jonet L, Klein C, Valamanes F, Berdugo M, Bigey P, Massin P, Jeanny JC, Behar-Cohen F - PLoS ONE (2011)

PGF expression in diabetic retina.(a–b) Comparison of PGF staining in non diabetic (a) and diabetic (b) retinas. (a) Sections of eyes from adult control non diabetic rat showed co-expression of PGF and GFAP in glial Muller cells from the gcl (arrowheads) to the inl, and PGF expression in glial Müller cells which are not immuno-reactive for GFAP (white star). Scale bar = 50 µm. (b) A similar pattern was observed on retinal sections of eyes from three-month-old diabetic rats, with a strong immuno-reactivity for PGF at the gcl level. (c) PGF detection by Western-blot in diabetic and non-diabetic retinas, from 1, 2, 5 and 12 month-old rats. For each lane in which 40 µg of proteins were deposited, the blood sugar level of the represented rats is indicated between parentheses. (d–e) Immunostaining for PGF and GFAP in sections from pVAX2-rPGF-1 ET- treated diabetic rat eyes, one month after ET. Sections show PGF-expressing infiltrating cells in the sub retinal space (d, arrows) and confirmed PGF expression by RMG cells (d, arrowheads). GFAP staining showed gliosis induced by RMG cells (d, e). ch, choroid; gcl, ganglion cell layer; inl, inner nuclear layer; ipl, inner plexiform layer; onl, outer nuclear layer; rpe, retinal pigmented epithelium; Scale bar = 100 µm.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3049767&req=5

pone-0017462-g006: PGF expression in diabetic retina.(a–b) Comparison of PGF staining in non diabetic (a) and diabetic (b) retinas. (a) Sections of eyes from adult control non diabetic rat showed co-expression of PGF and GFAP in glial Muller cells from the gcl (arrowheads) to the inl, and PGF expression in glial Müller cells which are not immuno-reactive for GFAP (white star). Scale bar = 50 µm. (b) A similar pattern was observed on retinal sections of eyes from three-month-old diabetic rats, with a strong immuno-reactivity for PGF at the gcl level. (c) PGF detection by Western-blot in diabetic and non-diabetic retinas, from 1, 2, 5 and 12 month-old rats. For each lane in which 40 µg of proteins were deposited, the blood sugar level of the represented rats is indicated between parentheses. (d–e) Immunostaining for PGF and GFAP in sections from pVAX2-rPGF-1 ET- treated diabetic rat eyes, one month after ET. Sections show PGF-expressing infiltrating cells in the sub retinal space (d, arrows) and confirmed PGF expression by RMG cells (d, arrowheads). GFAP staining showed gliosis induced by RMG cells (d, e). ch, choroid; gcl, ganglion cell layer; inl, inner nuclear layer; ipl, inner plexiform layer; onl, outer nuclear layer; rpe, retinal pigmented epithelium; Scale bar = 100 µm.
Mentions: In non-diabetic untreated eyes, PGF immuno-localization showed that PGF-1 is expressed in retinal Müller glial (RMG) cells and in the inner segments of photoreceptors (Fig. 6a). In three-month-old GK diabetic rats, PGF-1 was strongly expressed in the inner part the retina, mostly in astrocytes and in swollen RMG cells, as shown by the co labeling with the glial marker, GFAP (Fig. 6b). PGF expression over time in diabetic retinas was compared to PGF expression in control retinas using western-blot (Fig. 6c). This kinetic showed that PGF expression increased in the neuroretinas from diabetic rats aged between two and five months, and remained elevated as the diabetes develops at later stages (12 months), whereas it remained constant between one month and one year in retinas from non diabetic control rats. PGF-1 is therefore upregulated in the retina of GK diabetic rats, increasing at the early stages of diabetes and remaining elevated thereafter.

Bottom Line: For a better understanding of its role on the retina, we have evaluated the effect of a sustained PGF over-expression in rat ocular media, using ciliary muscle electrotransfer (ET) of a plasmid encoding rat PGF-1 (pVAX2-rPGF-1). pVAX2-rPGF-1 ET in the ciliary muscle (200 V/cm) was achieved in non diabetic and diabetic rat eyes.After the control of rPGF-1 expression, PGF-induced effects on retinal vasculature and on the blood-external barrier were evaluated respectively by lectin and occludin staining on flat-mounts.PGF and its receptor Flt-1 may therefore be looked upon as a potential regulatory target at this stage of the disease.

View Article: PubMed Central - PubMed

Affiliation: Institut National pour la Santé Et la Recherche Médicale U872, Physiopathology of ocular diseases: Therapeutic innovations, Paris, France.

ABSTRACT

Objective: There are controversies regarding the pro-angiogenic activity of placental growth factor (PGF) in diabetic retinopathy (DR). For a better understanding of its role on the retina, we have evaluated the effect of a sustained PGF over-expression in rat ocular media, using ciliary muscle electrotransfer (ET) of a plasmid encoding rat PGF-1 (pVAX2-rPGF-1).

Materials and methods: pVAX2-rPGF-1 ET in the ciliary muscle (200 V/cm) was achieved in non diabetic and diabetic rat eyes. Control eyes received saline or naked plasmid ET. Clinical follow up was carried out over three months using slit lamp examination and fluorescein angiography. After the control of rPGF-1 expression, PGF-induced effects on retinal vasculature and on the blood-external barrier were evaluated respectively by lectin and occludin staining on flat-mounts. Ocular structures were visualized through histological analysis.

Results: After fifteen days of rPGF-1 over-expression in normal eyes, tortuous and dilated capillaries were observed. At one month, microaneurysms and moderate vascular sprouts were detected in mid retinal periphery in vivo and on retinal flat-mounts. At later stages, retinal pigmented epithelial cells demonstrated morphological abnormalities and junction ruptures. In diabetic retinas, PGF expression rose between 2 and 5 months, and, one month after ET, rPGF-1 over-expression induced glial activation and proliferation.

Conclusion: This is the first demonstration that sustained intraocular PGF production induces vascular and retinal changes similar to those observed in the early stages of diabetic retinopathy. PGF and its receptor Flt-1 may therefore be looked upon as a potential regulatory target at this stage of the disease.

Show MeSH
Related in: MedlinePlus