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Influence of age on retinochoroidal healing processes after argon photocoagulation in C57bl/6j mice.

Dot C, Parier V, Behar-Cohen F, Benezra D, Jonet L, Goldenberg B, Picard E, Camelo S, de Kozak Y, May F, Soubrane G, Jeanny JC - Mol. Vis. (2009)

Bottom Line: In the cellular and morphologic study, the resultant retinal pigment epithelium interruption expanse was significantly different between the four groups.Our results were based on the study of four different age groups and included not only data from morphological observations but also from a molecular analysis of the various alterations of cytokine signaling and expression.The above results may provide some insight into possible therapeutic strategies in the future.

View Article: PubMed Central - PubMed

Affiliation: Centre de Recherche des Cordeliers, Université Pierre et Marie Curie, Paris, France.

ABSTRACT

Purpose: To analyze the influence of age on retinochoroidal wound healing processes and on glial growth factor and cytokine mRNA expression profiles observed after argon laser photocoagulation.

Methods: A cellular and morphometric study was performed that used 44 C57Bl/6J mice: 4-week-old mice (group I, n=8), 6-week-old mice (group II, n=8), 10-12-week-old mice (group III, n=14), and 1-year-old mice (group IV, n=14). All mice in these groups underwent a standard argon laser photocoagulation (50 microm, 400 mW, 0.05 s). Two separated lesions were created in each retina using a slit lamp delivery system. At 1, 3, 7, 14, 60 days, and 4 months after photocoagulation, mice from each of the four groups were sacrificed by carbon dioxide inhalation. Groups III and IV were also studied at 6, 7, and 8 months after photocoagulation. At each time point the enucleated eyes were either mounted in Tissue Tek (OCT), snap frozen and processed for immunohistochemistry or either flat mounted (left eyes of groups III and IV). To determine, by RT-PCR, the time course of glial fibrillary acidic protein (GFAP), vascular endothelial growth factor (VEGF), and monocyte chemotactic protein-1 (MCP-1) gene expression, we delivered ten laser burns (50 microm, 400 mW, 0.05 s) to each retina in 10-12-week-old mice (group III', n=10) and 1-year-old mice (group IV', n=10). Animals from Groups III' and IV' had the same age than those from Groups III and IV, but they received ten laser impacts in each eye and served for the molecular analysis. Mice from Groups III and IV received only two laser impacts per eye and served for the cellular and morphologic study. Retinal and choroidal tissues from these treated mice were collected at 16 h, and 1, 2, 3, and 7 days after photocoagulation. Two mice of each group did not receive photocoagulation and were used as controls.

Results: In the cellular and morphologic study, the resultant retinal pigment epithelium interruption expanse was significantly different between the four groups. It was more concise and smaller in the oldest group IV (112.1 microm+/-11.4 versus 219.1 microm+/-12.2 in group III) p<0.0001 between groups III and IV. By contrast, while choroidal neovascularization (CNV) was mild and not readily identifiable in group I, at all time points studied, CNV was more prominent in the (1-year-old mice) Group IV than in the other groups. For instance, up to 14 days after photocoagulation, CNV reaction was statistically larger in group IV than in group III ((p=0.0049 between groups III and IV on slide sections and p<0.0001 between the same groups on flat mounts). Moreover, four months after photocoagulation, the CNV area (on slide sections) was 1,282 microm(2)+/-90 for group III and 2,999 microm(2)+/-115 for group IV (p<0.0001 between groups III and IV). Accordingly, GFAP, VEGF, and MCP-1 mRNA expression profiles, determined by RT-PCR at 16 h, 1, 2, 3, and 7 days postphotocoagulation, were modified with aging. In 1-year-old mice (group IV), GFAP mRNA expression was already significantly higher than in the younger (10-12 week) group III before photocoagulation. After laser burns, GFAP mRNA expression peaked at 16-24 h and on day 7, decreasing thereafter. VEGF mRNA expression was markedly increased after photocoagulation in old mice eyes, reaching 2.7 times its basal level at day 3, while it was only slightly increased in young mice (1.3 times its level in untreated young mice 3 days postphotocoagulation). At all time points after photocoagulation, MCP-1 mRNA expression was elevated in old mice, reaching high levels of expression at 16 h and day 3 respectively.

Conclusions: Our results were based on the study of four different age groups and included not only data from morphological observations but also from a molecular analysis of the various alterations of cytokine signaling and expression. One-year-old mice demonstrated more extensive CNV formation and a slower pace of regression after laser photocoagulation than younger mice. These were accompanied by differences in growth factors and cytokine expression profiles indicate that aging is a factor that aggravates CNV. The above results may provide some insight into possible therapeutic strategies in the future.

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Evolution of the choroidal neovascularization reaction in young adult (group III) and old mice (group IV) as assessed on flat mounts. A: Choroidal neovascularization (CNV) reaction is still detected eight months after photocoagulation and is more important in 1-year-old mice during all tested time points. The difference between the two groups is significant on day 7 (p=0.028) and on month 8 (p=0.0094). B: The mean CNV area (during the first two weeks after photocoagulation) is larger in the 1-year-old mice; the difference is highly statistically significant according to t-test.
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f7: Evolution of the choroidal neovascularization reaction in young adult (group III) and old mice (group IV) as assessed on flat mounts. A: Choroidal neovascularization (CNV) reaction is still detected eight months after photocoagulation and is more important in 1-year-old mice during all tested time points. The difference between the two groups is significant on day 7 (p=0.028) and on month 8 (p=0.0094). B: The mean CNV area (during the first two weeks after photocoagulation) is larger in the 1-year-old mice; the difference is highly statistically significant according to t-test.

Mentions: CNV area was larger in one-year-old mice (group IV) at each time point from day 7 to month 8 in comparison with the young adult mice (group III). The difference was statistically significant: p=0.028 7 days and p=0.009 eight months following photocoagulation (Figure 7A). A compilation of the data collected during the first two weeks after photocoagulation showed that the mean CNV area was consistently and significantly larger in the group IV (1-year-old mice; 45,200 µm2±3,490 versus 24,770 µm2±2,160; p<0.001) than in the group III (10–12-week-old mice; Figure 7B).


Influence of age on retinochoroidal healing processes after argon photocoagulation in C57bl/6j mice.

Dot C, Parier V, Behar-Cohen F, Benezra D, Jonet L, Goldenberg B, Picard E, Camelo S, de Kozak Y, May F, Soubrane G, Jeanny JC - Mol. Vis. (2009)

Evolution of the choroidal neovascularization reaction in young adult (group III) and old mice (group IV) as assessed on flat mounts. A: Choroidal neovascularization (CNV) reaction is still detected eight months after photocoagulation and is more important in 1-year-old mice during all tested time points. The difference between the two groups is significant on day 7 (p=0.028) and on month 8 (p=0.0094). B: The mean CNV area (during the first two weeks after photocoagulation) is larger in the 1-year-old mice; the difference is highly statistically significant according to t-test.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2664845&req=5

f7: Evolution of the choroidal neovascularization reaction in young adult (group III) and old mice (group IV) as assessed on flat mounts. A: Choroidal neovascularization (CNV) reaction is still detected eight months after photocoagulation and is more important in 1-year-old mice during all tested time points. The difference between the two groups is significant on day 7 (p=0.028) and on month 8 (p=0.0094). B: The mean CNV area (during the first two weeks after photocoagulation) is larger in the 1-year-old mice; the difference is highly statistically significant according to t-test.
Mentions: CNV area was larger in one-year-old mice (group IV) at each time point from day 7 to month 8 in comparison with the young adult mice (group III). The difference was statistically significant: p=0.028 7 days and p=0.009 eight months following photocoagulation (Figure 7A). A compilation of the data collected during the first two weeks after photocoagulation showed that the mean CNV area was consistently and significantly larger in the group IV (1-year-old mice; 45,200 µm2±3,490 versus 24,770 µm2±2,160; p<0.001) than in the group III (10–12-week-old mice; Figure 7B).

Bottom Line: In the cellular and morphologic study, the resultant retinal pigment epithelium interruption expanse was significantly different between the four groups.Our results were based on the study of four different age groups and included not only data from morphological observations but also from a molecular analysis of the various alterations of cytokine signaling and expression.The above results may provide some insight into possible therapeutic strategies in the future.

View Article: PubMed Central - PubMed

Affiliation: Centre de Recherche des Cordeliers, Université Pierre et Marie Curie, Paris, France.

ABSTRACT

Purpose: To analyze the influence of age on retinochoroidal wound healing processes and on glial growth factor and cytokine mRNA expression profiles observed after argon laser photocoagulation.

Methods: A cellular and morphometric study was performed that used 44 C57Bl/6J mice: 4-week-old mice (group I, n=8), 6-week-old mice (group II, n=8), 10-12-week-old mice (group III, n=14), and 1-year-old mice (group IV, n=14). All mice in these groups underwent a standard argon laser photocoagulation (50 microm, 400 mW, 0.05 s). Two separated lesions were created in each retina using a slit lamp delivery system. At 1, 3, 7, 14, 60 days, and 4 months after photocoagulation, mice from each of the four groups were sacrificed by carbon dioxide inhalation. Groups III and IV were also studied at 6, 7, and 8 months after photocoagulation. At each time point the enucleated eyes were either mounted in Tissue Tek (OCT), snap frozen and processed for immunohistochemistry or either flat mounted (left eyes of groups III and IV). To determine, by RT-PCR, the time course of glial fibrillary acidic protein (GFAP), vascular endothelial growth factor (VEGF), and monocyte chemotactic protein-1 (MCP-1) gene expression, we delivered ten laser burns (50 microm, 400 mW, 0.05 s) to each retina in 10-12-week-old mice (group III', n=10) and 1-year-old mice (group IV', n=10). Animals from Groups III' and IV' had the same age than those from Groups III and IV, but they received ten laser impacts in each eye and served for the molecular analysis. Mice from Groups III and IV received only two laser impacts per eye and served for the cellular and morphologic study. Retinal and choroidal tissues from these treated mice were collected at 16 h, and 1, 2, 3, and 7 days after photocoagulation. Two mice of each group did not receive photocoagulation and were used as controls.

Results: In the cellular and morphologic study, the resultant retinal pigment epithelium interruption expanse was significantly different between the four groups. It was more concise and smaller in the oldest group IV (112.1 microm+/-11.4 versus 219.1 microm+/-12.2 in group III) p<0.0001 between groups III and IV. By contrast, while choroidal neovascularization (CNV) was mild and not readily identifiable in group I, at all time points studied, CNV was more prominent in the (1-year-old mice) Group IV than in the other groups. For instance, up to 14 days after photocoagulation, CNV reaction was statistically larger in group IV than in group III ((p=0.0049 between groups III and IV on slide sections and p<0.0001 between the same groups on flat mounts). Moreover, four months after photocoagulation, the CNV area (on slide sections) was 1,282 microm(2)+/-90 for group III and 2,999 microm(2)+/-115 for group IV (p<0.0001 between groups III and IV). Accordingly, GFAP, VEGF, and MCP-1 mRNA expression profiles, determined by RT-PCR at 16 h, 1, 2, 3, and 7 days postphotocoagulation, were modified with aging. In 1-year-old mice (group IV), GFAP mRNA expression was already significantly higher than in the younger (10-12 week) group III before photocoagulation. After laser burns, GFAP mRNA expression peaked at 16-24 h and on day 7, decreasing thereafter. VEGF mRNA expression was markedly increased after photocoagulation in old mice eyes, reaching 2.7 times its basal level at day 3, while it was only slightly increased in young mice (1.3 times its level in untreated young mice 3 days postphotocoagulation). At all time points after photocoagulation, MCP-1 mRNA expression was elevated in old mice, reaching high levels of expression at 16 h and day 3 respectively.

Conclusions: Our results were based on the study of four different age groups and included not only data from morphological observations but also from a molecular analysis of the various alterations of cytokine signaling and expression. One-year-old mice demonstrated more extensive CNV formation and a slower pace of regression after laser photocoagulation than younger mice. These were accompanied by differences in growth factors and cytokine expression profiles indicate that aging is a factor that aggravates CNV. The above results may provide some insight into possible therapeutic strategies in the future.

Show MeSH
Related in: MedlinePlus