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Microglia in mouse retina contralateral to experimental glaucoma exhibit multiple signs of activation in all retinal layers.

Rojas B, Gallego BI, Ramírez AI, Salazar JJ, de Hoz R, Valiente-Soriano FJ, Avilés-Trigueros M, Villegas-Perez MP, Vidal-Sanz M, Triviño A, Ramírez JM - J Neuroinflammation (2014)

Bottom Line: In comparison with the control group, a significant increase in the microglial number in the PL, OS, and in the area occupied by Iba-1+ cells in the NFL-GCL, and significant reduction of the arbor area in the PL.Such activation extended beyond the GCL, involving all retinal layers.Differences between the two eyes could help to elucidate glaucoma pathophysiology.

View Article: PubMed Central - HTML - PubMed

Affiliation: Instituto de Investigaciones Oftalmológicas Ramón Castroviejo, Facultad de Medicina, Pab VI, 4a, Avenida Complutense s/n, Universidad Complutense de Madrid, 28040 Madrid, Spain. ramirezs@med.ucm.es.

ABSTRACT

Background: Glaucomatous optic neuropathy, a leading cause of blindness, can progress despite control of intraocular pressure - currently the main risk factor and target for treatment. Glaucoma progression shares mechanisms with neurodegenerative disease, including microglia activation. In the present model of ocular hypertension (OHT), we have recently described morphological signs of retinal microglia activation and MHC-II upregulation in both the untreated contralateral eyes and OHT eyes. By using immunostaining, we sought to analyze and quantify additional signs of microglia activation and differences depending on the retinal layer.

Methods: Two groups of adult Swiss mice were used: age-matched control (naïve, n = 12), and lasered (n = 12). In the lasered animals, both OHT eyes and contralateral eyes were analyzed. Retinal whole-mounts were immunostained with antibodies against Iba-1, MHC-II, CD68, CD86, and Ym1. The Iba-1+ cell number in the plexiform layers (PL) and the photoreceptor outer segment (OS), Iba-1+ arbor area in the PL, and area of the retina occupied by Iba-1+ cells in the nerve fiber layer-ganglion cell layer (NFL-GCL) were quantified.

Results: The main findings in contralateral eyes and OHT eyes were: i) ameboid microglia in the NFL-GCL and OS; ii) the retraction of processes in all retinal layers; iii) a higher level of branching in PL and in the OS; iv) soma displacement to the nearest cell layers in the PL and OS; v) the reorientation of processes in the OS; vi) MHC-II upregulation in all retinal layers; vii) increased CD68 immunostaining; and viii) CD86 immunolabeling in ameboid cells. In comparison with the control group, a significant increase in the microglial number in the PL, OS, and in the area occupied by Iba-1+ cells in the NFL-GCL, and significant reduction of the arbor area in the PL. In addition, rounded Iba-1+ CD86+ cells in the NFL-GCL, OS and Ym1+ cells, and rod-like microglia in the NFL-GCL were restricted to OHT eyes.

Conclusions: Several quantitative and qualitative signs of microglia activation are detected both in the contralateral and OHT eyes. Such activation extended beyond the GCL, involving all retinal layers. Differences between the two eyes could help to elucidate glaucoma pathophysiology.

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Iba-1+ cell quantification in OS, OPL, and IPL. Each bar represents the mean ± SD of the sum of Iba-1+ cell number contained in the three retinal layers. ***P <0.001 versus naïve and contralateral retinas. *P <0.05 versus naïve retinas. (IPL: inner plexiform layer; OHT: ocular hypertension; OPL: outer plexiform layer; OS: photoreceptor outer segment).
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Figure 12: Iba-1+ cell quantification in OS, OPL, and IPL. Each bar represents the mean ± SD of the sum of Iba-1+ cell number contained in the three retinal layers. ***P <0.001 versus naïve and contralateral retinas. *P <0.05 versus naïve retinas. (IPL: inner plexiform layer; OHT: ocular hypertension; OPL: outer plexiform layer; OS: photoreceptor outer segment).

Mentions: OHT eyes had significantly more Iba-1+ cells in the IPL, OPL, and OS than did contralateral and naïve eyes, both when the comparison was made as the sum of Iba-1+ cells contained in the three layers (Iba-1+ total number, Figure 12) or when the layers were compared one by one between the study groups (Table 1) (P <0.001 in all instances; t-test). The Iba-1+ total number also significantly increased in contralateral eyes in comparison with naïve eyes (P <0.05; unpaired t-test; Figure 12). The analysis by layers revealed that the IPL of contralateral eyes had significantly more Iba-1+ cells than did naïve eyes (P <0.05; unpaired t-test) (Table 1). In addition, the comparison between OPL and IPL showed that the number of Iba-1+ cells was significantly greater in the OPL in naïve (P <0.01), contralateral (P <0.001) and OHT eyes (P <0.001; paired t-test in all instances; Table 1).


Microglia in mouse retina contralateral to experimental glaucoma exhibit multiple signs of activation in all retinal layers.

Rojas B, Gallego BI, Ramírez AI, Salazar JJ, de Hoz R, Valiente-Soriano FJ, Avilés-Trigueros M, Villegas-Perez MP, Vidal-Sanz M, Triviño A, Ramírez JM - J Neuroinflammation (2014)

Iba-1+ cell quantification in OS, OPL, and IPL. Each bar represents the mean ± SD of the sum of Iba-1+ cell number contained in the three retinal layers. ***P <0.001 versus naïve and contralateral retinas. *P <0.05 versus naïve retinas. (IPL: inner plexiform layer; OHT: ocular hypertension; OPL: outer plexiform layer; OS: photoreceptor outer segment).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4128533&req=5

Figure 12: Iba-1+ cell quantification in OS, OPL, and IPL. Each bar represents the mean ± SD of the sum of Iba-1+ cell number contained in the three retinal layers. ***P <0.001 versus naïve and contralateral retinas. *P <0.05 versus naïve retinas. (IPL: inner plexiform layer; OHT: ocular hypertension; OPL: outer plexiform layer; OS: photoreceptor outer segment).
Mentions: OHT eyes had significantly more Iba-1+ cells in the IPL, OPL, and OS than did contralateral and naïve eyes, both when the comparison was made as the sum of Iba-1+ cells contained in the three layers (Iba-1+ total number, Figure 12) or when the layers were compared one by one between the study groups (Table 1) (P <0.001 in all instances; t-test). The Iba-1+ total number also significantly increased in contralateral eyes in comparison with naïve eyes (P <0.05; unpaired t-test; Figure 12). The analysis by layers revealed that the IPL of contralateral eyes had significantly more Iba-1+ cells than did naïve eyes (P <0.05; unpaired t-test) (Table 1). In addition, the comparison between OPL and IPL showed that the number of Iba-1+ cells was significantly greater in the OPL in naïve (P <0.01), contralateral (P <0.001) and OHT eyes (P <0.001; paired t-test in all instances; Table 1).

Bottom Line: In comparison with the control group, a significant increase in the microglial number in the PL, OS, and in the area occupied by Iba-1+ cells in the NFL-GCL, and significant reduction of the arbor area in the PL.Such activation extended beyond the GCL, involving all retinal layers.Differences between the two eyes could help to elucidate glaucoma pathophysiology.

View Article: PubMed Central - HTML - PubMed

Affiliation: Instituto de Investigaciones Oftalmológicas Ramón Castroviejo, Facultad de Medicina, Pab VI, 4a, Avenida Complutense s/n, Universidad Complutense de Madrid, 28040 Madrid, Spain. ramirezs@med.ucm.es.

ABSTRACT

Background: Glaucomatous optic neuropathy, a leading cause of blindness, can progress despite control of intraocular pressure - currently the main risk factor and target for treatment. Glaucoma progression shares mechanisms with neurodegenerative disease, including microglia activation. In the present model of ocular hypertension (OHT), we have recently described morphological signs of retinal microglia activation and MHC-II upregulation in both the untreated contralateral eyes and OHT eyes. By using immunostaining, we sought to analyze and quantify additional signs of microglia activation and differences depending on the retinal layer.

Methods: Two groups of adult Swiss mice were used: age-matched control (naïve, n = 12), and lasered (n = 12). In the lasered animals, both OHT eyes and contralateral eyes were analyzed. Retinal whole-mounts were immunostained with antibodies against Iba-1, MHC-II, CD68, CD86, and Ym1. The Iba-1+ cell number in the plexiform layers (PL) and the photoreceptor outer segment (OS), Iba-1+ arbor area in the PL, and area of the retina occupied by Iba-1+ cells in the nerve fiber layer-ganglion cell layer (NFL-GCL) were quantified.

Results: The main findings in contralateral eyes and OHT eyes were: i) ameboid microglia in the NFL-GCL and OS; ii) the retraction of processes in all retinal layers; iii) a higher level of branching in PL and in the OS; iv) soma displacement to the nearest cell layers in the PL and OS; v) the reorientation of processes in the OS; vi) MHC-II upregulation in all retinal layers; vii) increased CD68 immunostaining; and viii) CD86 immunolabeling in ameboid cells. In comparison with the control group, a significant increase in the microglial number in the PL, OS, and in the area occupied by Iba-1+ cells in the NFL-GCL, and significant reduction of the arbor area in the PL. In addition, rounded Iba-1+ CD86+ cells in the NFL-GCL, OS and Ym1+ cells, and rod-like microglia in the NFL-GCL were restricted to OHT eyes.

Conclusions: Several quantitative and qualitative signs of microglia activation are detected both in the contralateral and OHT eyes. Such activation extended beyond the GCL, involving all retinal layers. Differences between the two eyes could help to elucidate glaucoma pathophysiology.

Show MeSH
Related in: MedlinePlus