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Low dose cranial irradiation-induced cerebrovascular damage is reversible in mice.

Sándor N, Walter FR, Bocsik A, Sántha P, Schilling-Tóth B, Léner V, Varga Z, Kahán Z, Deli MA, Sáfrány G, Hegyesi H - PLoS ONE (2014)

Bottom Line: Blood-brain barrier permeability for Evans blue, body weight and number of peripheral mononuclear and circulating endothelial progenitor cells were evaluated 1, 4 and 26 weeks postirradiation.Endothelial senescence was determined by senescence associated β-galactosidase staining.Extravasation of Evans blue increased in cerebrum and cerebellum in adult mice 1 week and in infant mice 4 weeks postirradiation at all treatment doses.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Radiobiology and Biodosimetry, "Frédéric Joliot-Curie" National Research Institute for Radiobiology and Radiohygiene, Budapest, Hungary; Doctoral Schools of Pathological Sciences, Semmelweis University, Budapest, Hungary.

ABSTRACT

Background: High-dose radiation-induced blood-brain barrier breakdown contributes to acute radiation toxicity syndrome and delayed brain injury, but there are few data on the effects of low dose cranial irradiation. Our goal was to measure blood-brain barrier changes after low (0.1 Gy), moderate (2 Gy) and high (10 Gy) dose irradiation under in vivo and in vitro conditions.

Methodology: Cranial irradiation was performed on 10-day-old and 10-week-old mice. Blood-brain barrier permeability for Evans blue, body weight and number of peripheral mononuclear and circulating endothelial progenitor cells were evaluated 1, 4 and 26 weeks postirradiation. Barrier properties of primary mouse brain endothelial cells co-cultured with glial cells were determined by measurement of resistance and permeability for marker molecules and staining for interendothelial junctions. Endothelial senescence was determined by senescence associated β-galactosidase staining.

Principle findings: Extravasation of Evans blue increased in cerebrum and cerebellum in adult mice 1 week and in infant mice 4 weeks postirradiation at all treatment doses. Head irradiation with 10 Gy decreased body weight. The number of circulating endothelial progenitor cells in blood was decreased 1 day after irradiation with 0.1 and 2 Gy. Increase in the permeability of cultured brain endothelial monolayers for fluorescein and albumin was time- and radiation dose dependent and accompanied by changes in junctional immunostaining for claudin-5, ZO-1 and β-catenin. The number of cultured brain endothelial and glial cells decreased from third day of postirradiation and senescence in endothelial cells increased at 2 and 10 Gy.

Conclusion: Not only high but low and moderate doses of cranial irradiation increase permeability of cerebral vessels in mice, but this effect is reversible by 6 months. In-vitro experiments suggest that irradiation changes junctional morphology, decreases cell number and causes senescence in brain endothelial cells.

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Effect of irradiation on GFAP immunostaining in mouse astroglia cells.Primary mouse astroglia cells 1, 2, 3 and 5 days after exposure to single dose of 0.1, 2 or 10 Gy irradiation were immunostained for glial fibrillary acidic protein (GFAP). Green color: immunostaining for GFAP. Blue color: H33343 staining of cell nuclei Bar  = 50 µm.
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pone-0112397-g006: Effect of irradiation on GFAP immunostaining in mouse astroglia cells.Primary mouse astroglia cells 1, 2, 3 and 5 days after exposure to single dose of 0.1, 2 or 10 Gy irradiation were immunostained for glial fibrillary acidic protein (GFAP). Green color: immunostaining for GFAP. Blue color: H33343 staining of cell nuclei Bar  = 50 µm.

Mentions: In MBECs the effect of irradiation on cell-cell adhesion and morphology was investigated by immunostaining for transmembrane tight junction protein claudin-5 (Figure 5), and cytoplasmic linker proteins β-catenin and ZO-1 (Figure S1–S2). In the control group all three junctional stainings were strong, and localized to the cell borders. The elongated shapes of endothelial cells were well delineated by the belt-like staining. Cell-cell attachment in MBECs was continuous, without gaps. No gross changes in junctional immunostainings were visible 24 h post-irradiation. Interendothelial junctional staining was changed in treated groups 2, 3 and 5 days post-irradiation, which effect was also reflected by permeability elevation at these time points. The pattern of the staining was especially altered at 3 and 5 days at 2 and 10 Gy: intercellular gaps, fragmented junctional staining and cytoplasmic redistribution of junctional proteins were observed (Figure 5). Morphological changes in astroglia cells were monitored by GFAP staining which revealed polygonal shape and several long processes in the sham irradiated group, typical for cultured type 1 astrocytes (Figure 6). This stellate shape was also observed in the 0.1 and 2 Gy groups, although more bipolar cells were present. The loss of polygonal shape and fine processes were more pronounced in cells exposed to 2 Gy irradiation. Striking morphological changes were visible in astrocytes irradiated with 10 Gy at all time points. A decreased intensity of the fluorescent immunostaining for junctional proteins was measured by quantitative image analysis in the moderate and high dose irradiation groups 3 and 5 days after treatment (Figure 7A, S3). In the high dose irradiation group at days 3 and 5 flattened morphology and increase of cell surface indicating cell stretching was prominent in MBECs (Figure 5, S1, S2). Changes in GFAP immunostaining in glial cells were quantified by image analysis, and significant decreases in intensity were observed at 2 and 10 Gy treatment doses on day 2, 3 and 5 postirradiation (Figure 7B).


Low dose cranial irradiation-induced cerebrovascular damage is reversible in mice.

Sándor N, Walter FR, Bocsik A, Sántha P, Schilling-Tóth B, Léner V, Varga Z, Kahán Z, Deli MA, Sáfrány G, Hegyesi H - PLoS ONE (2014)

Effect of irradiation on GFAP immunostaining in mouse astroglia cells.Primary mouse astroglia cells 1, 2, 3 and 5 days after exposure to single dose of 0.1, 2 or 10 Gy irradiation were immunostained for glial fibrillary acidic protein (GFAP). Green color: immunostaining for GFAP. Blue color: H33343 staining of cell nuclei Bar  = 50 µm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4231057&req=5

pone-0112397-g006: Effect of irradiation on GFAP immunostaining in mouse astroglia cells.Primary mouse astroglia cells 1, 2, 3 and 5 days after exposure to single dose of 0.1, 2 or 10 Gy irradiation were immunostained for glial fibrillary acidic protein (GFAP). Green color: immunostaining for GFAP. Blue color: H33343 staining of cell nuclei Bar  = 50 µm.
Mentions: In MBECs the effect of irradiation on cell-cell adhesion and morphology was investigated by immunostaining for transmembrane tight junction protein claudin-5 (Figure 5), and cytoplasmic linker proteins β-catenin and ZO-1 (Figure S1–S2). In the control group all three junctional stainings were strong, and localized to the cell borders. The elongated shapes of endothelial cells were well delineated by the belt-like staining. Cell-cell attachment in MBECs was continuous, without gaps. No gross changes in junctional immunostainings were visible 24 h post-irradiation. Interendothelial junctional staining was changed in treated groups 2, 3 and 5 days post-irradiation, which effect was also reflected by permeability elevation at these time points. The pattern of the staining was especially altered at 3 and 5 days at 2 and 10 Gy: intercellular gaps, fragmented junctional staining and cytoplasmic redistribution of junctional proteins were observed (Figure 5). Morphological changes in astroglia cells were monitored by GFAP staining which revealed polygonal shape and several long processes in the sham irradiated group, typical for cultured type 1 astrocytes (Figure 6). This stellate shape was also observed in the 0.1 and 2 Gy groups, although more bipolar cells were present. The loss of polygonal shape and fine processes were more pronounced in cells exposed to 2 Gy irradiation. Striking morphological changes were visible in astrocytes irradiated with 10 Gy at all time points. A decreased intensity of the fluorescent immunostaining for junctional proteins was measured by quantitative image analysis in the moderate and high dose irradiation groups 3 and 5 days after treatment (Figure 7A, S3). In the high dose irradiation group at days 3 and 5 flattened morphology and increase of cell surface indicating cell stretching was prominent in MBECs (Figure 5, S1, S2). Changes in GFAP immunostaining in glial cells were quantified by image analysis, and significant decreases in intensity were observed at 2 and 10 Gy treatment doses on day 2, 3 and 5 postirradiation (Figure 7B).

Bottom Line: Blood-brain barrier permeability for Evans blue, body weight and number of peripheral mononuclear and circulating endothelial progenitor cells were evaluated 1, 4 and 26 weeks postirradiation.Endothelial senescence was determined by senescence associated β-galactosidase staining.Extravasation of Evans blue increased in cerebrum and cerebellum in adult mice 1 week and in infant mice 4 weeks postirradiation at all treatment doses.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Radiobiology and Biodosimetry, "Frédéric Joliot-Curie" National Research Institute for Radiobiology and Radiohygiene, Budapest, Hungary; Doctoral Schools of Pathological Sciences, Semmelweis University, Budapest, Hungary.

ABSTRACT

Background: High-dose radiation-induced blood-brain barrier breakdown contributes to acute radiation toxicity syndrome and delayed brain injury, but there are few data on the effects of low dose cranial irradiation. Our goal was to measure blood-brain barrier changes after low (0.1 Gy), moderate (2 Gy) and high (10 Gy) dose irradiation under in vivo and in vitro conditions.

Methodology: Cranial irradiation was performed on 10-day-old and 10-week-old mice. Blood-brain barrier permeability for Evans blue, body weight and number of peripheral mononuclear and circulating endothelial progenitor cells were evaluated 1, 4 and 26 weeks postirradiation. Barrier properties of primary mouse brain endothelial cells co-cultured with glial cells were determined by measurement of resistance and permeability for marker molecules and staining for interendothelial junctions. Endothelial senescence was determined by senescence associated β-galactosidase staining.

Principle findings: Extravasation of Evans blue increased in cerebrum and cerebellum in adult mice 1 week and in infant mice 4 weeks postirradiation at all treatment doses. Head irradiation with 10 Gy decreased body weight. The number of circulating endothelial progenitor cells in blood was decreased 1 day after irradiation with 0.1 and 2 Gy. Increase in the permeability of cultured brain endothelial monolayers for fluorescein and albumin was time- and radiation dose dependent and accompanied by changes in junctional immunostaining for claudin-5, ZO-1 and β-catenin. The number of cultured brain endothelial and glial cells decreased from third day of postirradiation and senescence in endothelial cells increased at 2 and 10 Gy.

Conclusion: Not only high but low and moderate doses of cranial irradiation increase permeability of cerebral vessels in mice, but this effect is reversible by 6 months. In-vitro experiments suggest that irradiation changes junctional morphology, decreases cell number and causes senescence in brain endothelial cells.

Show MeSH
Related in: MedlinePlus