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Vps35 haploinsufficiency results in degenerative-like deficit in mouse retinal ganglion neurons and impairment of optic nerve injury-induced gliosis.

Liu W, Tang FL, Erion J, Xiao H, Ye J, Xiong WC - Mol Brain (2014)

Bottom Line: VPS35 (vacuolar protein sorting 35) is a major component of retromer that selectively promotes endosome-to-Golgi retrieval of transmembrane proteins.RGCs from young adult Vps35 heterozygotes (Vps35+/m) show degenerative-like features, such as dystrophic dendrites, reduced axon fibers, and increased TUNEL labeled RGCs.Additionally, gliosis in the optic nerve is transiently elevated in neonatal, but reduced in aged Vps35+/m mice.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Ophthalmology, Institute of Surgery Research, Daping Hospital, Third Military Medical University, Chongqing, China. yejian1979@163.com.

ABSTRACT
VPS35 (vacuolar protein sorting 35) is a major component of retromer that selectively promotes endosome-to-Golgi retrieval of transmembrane proteins. Dysfunction of retromer is a risk factor for the pathogenesis of Parkinson's disease (PD) and Alzheimer's disease (AD), both neuro-degeneration disorders. However, VPS35/retromer's function in retina or the contribution of Vps35-deficiency to retinal neuro-degenerative disorders has not been investigated. Here we provide evidence for a role of VPS35 in mouse retinal ganglion cell (RGC) survival and regeneration. VPS35 is selectively expressed in developing mouse RGCs. RGCs from young adult Vps35 heterozygotes (Vps35+/m) show degenerative-like features, such as dystrophic dendrites, reduced axon fibers, and increased TUNEL labeled RGCs. Additionally, gliosis in the optic nerve is transiently elevated in neonatal, but reduced in aged Vps35+/m mice. Optic nerve injury-induced gliosis is also attenuated in Vps35+/m mice. These results suggest that Vps35 is necessary for mouse RGC survival and regeneration, and Vps35-deficiency may contribute to the pathogenesis of retinal ganglion neuro-degeneration, a critical pathology leading to the blindness of many retinal degenerative disorders.

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Impairment of optic nerve injury-induced gliosis in vps35+/m mice. (A) Western blot analysis of homogenates from vps35+/+ and +/m optic nerves at indicated ages using indicated antibodies. (B) Illustration of mouse optic nerve injury. P30 vps35+/+ and +/m optic nerves were crushed as indicated. 3-days after injury, optic nerves were sectioned and subjected to immunostaining analysis. (C) Immunostaining analysis of optic nerves using indicated antibodies. Injury sites were marked with yellow arrows. Scale bar, 50 μm. (D) Quantification analysis of data from (C). Immunofluorescence intensity was normalized by the uninjured vps35+/+ controls. The values of mean +/- SEM (n = 3) were shown. *, P < 0.05, significance difference from the vps35+/+ control.
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Figure 6: Impairment of optic nerve injury-induced gliosis in vps35+/m mice. (A) Western blot analysis of homogenates from vps35+/+ and +/m optic nerves at indicated ages using indicated antibodies. (B) Illustration of mouse optic nerve injury. P30 vps35+/+ and +/m optic nerves were crushed as indicated. 3-days after injury, optic nerves were sectioned and subjected to immunostaining analysis. (C) Immunostaining analysis of optic nerves using indicated antibodies. Injury sites were marked with yellow arrows. Scale bar, 50 μm. (D) Quantification analysis of data from (C). Immunofluorescence intensity was normalized by the uninjured vps35+/+ controls. The values of mean +/- SEM (n = 3) were shown. *, P < 0.05, significance difference from the vps35+/+ control.

Mentions: Optic nerves are assembled by RGC axonal fibers and glial cells, such as oligodendricytes, astrocytes, and microglia [18]. To examine whether optic nerves are altered in vps35+/m mice, we first examined Vps35’s expression in optic nerve homogenates from various aged vps35+/+ and +/m mice. Vps35 protein levels were high in neonatal age (P15 and P30), but reduced in aged vps35+/+ mice (Figure 6A), in contrast from that in mouse retinas (Figure 1C). In agreement with the data from aged retinas, Vps35 protein levels were only decreased to ~50% in young age (P15 to P90), but not aged (e.g., P180), optic nerves (Figure 6A). Interestingly, increased GFAP (a marker for astrocytes) and MBP (myelin binding protein which marks oligodendricytes) levels were increased in neonatal, but decreased in aged (e.g., P90 and P180), vps35+/m optic nerves (Figure 6A). We thus further examined optic nerve phenotype in P30 vps35+/+ and +/m mice by immunostaining analysis using antibodies against Tuj1 (to label RGC axons), GFAP (to mark astrocytes), MBP (to view oligodendricytes), and IBA-1 (to stain microglia). RGC axons didn’t show obvious difference between P30 vps35+/+ and +/m optic nerves, based on immunostaining and Western blot analyses using anti-Tuj1 antibodies (data not shown). Oligodendricytes labeled by anti-MBP antibody were absent in the retina and optic nerve head, but present posterior to the lamina cribrosa-like region in both vps35+/+ and +/m optic nerves (Figure 6C). In contrast, astrocytes marked by GFAP were present in the retina and optic nerve head (Figure 6C). In agreement with data from Western blot analysis, Glial cells labeled by GFAP, MBP, and IBA-1 antibodies were all slightly increased in P30 vps35+/m optic nerves (Figures 6C-D). These results thus suggest an age-dependent alteration of gliosis in vps35+/m mutant optic nerves.


Vps35 haploinsufficiency results in degenerative-like deficit in mouse retinal ganglion neurons and impairment of optic nerve injury-induced gliosis.

Liu W, Tang FL, Erion J, Xiao H, Ye J, Xiong WC - Mol Brain (2014)

Impairment of optic nerve injury-induced gliosis in vps35+/m mice. (A) Western blot analysis of homogenates from vps35+/+ and +/m optic nerves at indicated ages using indicated antibodies. (B) Illustration of mouse optic nerve injury. P30 vps35+/+ and +/m optic nerves were crushed as indicated. 3-days after injury, optic nerves were sectioned and subjected to immunostaining analysis. (C) Immunostaining analysis of optic nerves using indicated antibodies. Injury sites were marked with yellow arrows. Scale bar, 50 μm. (D) Quantification analysis of data from (C). Immunofluorescence intensity was normalized by the uninjured vps35+/+ controls. The values of mean +/- SEM (n = 3) were shown. *, P < 0.05, significance difference from the vps35+/+ control.
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Figure 6: Impairment of optic nerve injury-induced gliosis in vps35+/m mice. (A) Western blot analysis of homogenates from vps35+/+ and +/m optic nerves at indicated ages using indicated antibodies. (B) Illustration of mouse optic nerve injury. P30 vps35+/+ and +/m optic nerves were crushed as indicated. 3-days after injury, optic nerves were sectioned and subjected to immunostaining analysis. (C) Immunostaining analysis of optic nerves using indicated antibodies. Injury sites were marked with yellow arrows. Scale bar, 50 μm. (D) Quantification analysis of data from (C). Immunofluorescence intensity was normalized by the uninjured vps35+/+ controls. The values of mean +/- SEM (n = 3) were shown. *, P < 0.05, significance difference from the vps35+/+ control.
Mentions: Optic nerves are assembled by RGC axonal fibers and glial cells, such as oligodendricytes, astrocytes, and microglia [18]. To examine whether optic nerves are altered in vps35+/m mice, we first examined Vps35’s expression in optic nerve homogenates from various aged vps35+/+ and +/m mice. Vps35 protein levels were high in neonatal age (P15 and P30), but reduced in aged vps35+/+ mice (Figure 6A), in contrast from that in mouse retinas (Figure 1C). In agreement with the data from aged retinas, Vps35 protein levels were only decreased to ~50% in young age (P15 to P90), but not aged (e.g., P180), optic nerves (Figure 6A). Interestingly, increased GFAP (a marker for astrocytes) and MBP (myelin binding protein which marks oligodendricytes) levels were increased in neonatal, but decreased in aged (e.g., P90 and P180), vps35+/m optic nerves (Figure 6A). We thus further examined optic nerve phenotype in P30 vps35+/+ and +/m mice by immunostaining analysis using antibodies against Tuj1 (to label RGC axons), GFAP (to mark astrocytes), MBP (to view oligodendricytes), and IBA-1 (to stain microglia). RGC axons didn’t show obvious difference between P30 vps35+/+ and +/m optic nerves, based on immunostaining and Western blot analyses using anti-Tuj1 antibodies (data not shown). Oligodendricytes labeled by anti-MBP antibody were absent in the retina and optic nerve head, but present posterior to the lamina cribrosa-like region in both vps35+/+ and +/m optic nerves (Figure 6C). In contrast, astrocytes marked by GFAP were present in the retina and optic nerve head (Figure 6C). In agreement with data from Western blot analysis, Glial cells labeled by GFAP, MBP, and IBA-1 antibodies were all slightly increased in P30 vps35+/m optic nerves (Figures 6C-D). These results thus suggest an age-dependent alteration of gliosis in vps35+/m mutant optic nerves.

Bottom Line: VPS35 (vacuolar protein sorting 35) is a major component of retromer that selectively promotes endosome-to-Golgi retrieval of transmembrane proteins.RGCs from young adult Vps35 heterozygotes (Vps35+/m) show degenerative-like features, such as dystrophic dendrites, reduced axon fibers, and increased TUNEL labeled RGCs.Additionally, gliosis in the optic nerve is transiently elevated in neonatal, but reduced in aged Vps35+/m mice.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Ophthalmology, Institute of Surgery Research, Daping Hospital, Third Military Medical University, Chongqing, China. yejian1979@163.com.

ABSTRACT
VPS35 (vacuolar protein sorting 35) is a major component of retromer that selectively promotes endosome-to-Golgi retrieval of transmembrane proteins. Dysfunction of retromer is a risk factor for the pathogenesis of Parkinson's disease (PD) and Alzheimer's disease (AD), both neuro-degeneration disorders. However, VPS35/retromer's function in retina or the contribution of Vps35-deficiency to retinal neuro-degenerative disorders has not been investigated. Here we provide evidence for a role of VPS35 in mouse retinal ganglion cell (RGC) survival and regeneration. VPS35 is selectively expressed in developing mouse RGCs. RGCs from young adult Vps35 heterozygotes (Vps35+/m) show degenerative-like features, such as dystrophic dendrites, reduced axon fibers, and increased TUNEL labeled RGCs. Additionally, gliosis in the optic nerve is transiently elevated in neonatal, but reduced in aged Vps35+/m mice. Optic nerve injury-induced gliosis is also attenuated in Vps35+/m mice. These results suggest that Vps35 is necessary for mouse RGC survival and regeneration, and Vps35-deficiency may contribute to the pathogenesis of retinal ganglion neuro-degeneration, a critical pathology leading to the blindness of many retinal degenerative disorders.

Show MeSH
Related in: MedlinePlus