Granzyme B degrades extracellular matrix and promotes inflammation and choroidal neovascularization.

Age-related macular degeneration (AMD) is a common retinal neurodegenerative disease among the elderly. Neovascular AMD (nAMD), a leading cause of AMD-related blindness, involves choroidal neovascularization (CNV), which can be suppressed by anti-angiogenic treatments. However, current CNV treatments do not work in all nAMD patients. Here we investigate a novel target for AMD. Granzyme B (GzmB) is a serine protease that promotes aging, chronic inflammation and vascular permeability through the degradation of the extracellular matrix (ECM) and tight junctions. Extracellular GzmB is increased in retina pigment epithelium (RPE) and mast cells in the choroid of the healthy aging outer retina. It is further increased in donor eyes exhibiting features of nAMD and CNV. Here, we show in RPE-choroidal explant cultures that exogenous GzmB degrades the RPE-choroid ECM, promotes retinal/choroidal inflammation and angiogenesis while diminishing anti-angiogenic factor, thrombospondin-1 (TSP-1). The pharmacological inhibition of either GzmB or mast-cell degranulation significantly reduces choroidal angiogenesis. In line with our in vitro data, GzmB-deficiency reduces the extent of laser-induced CNV lesions and the age-related deterioration of electroretinogram (ERG) responses in mice. These findings suggest that targeting GzmB, a serine protease with no known endogenous inhibitors, may be a potential novel therapeutic approach to suppress CNV in nAMD.

Granzyme B Contributes to Choroidal Neovascularization and Age-Related Macular Degeneration Through Proteolysis of Thrombospondin-1.

Age-related macular degeneration (AMD) is a leading cause of irreversible central vision loss in the elderly. The pathology of neovascular age-related macular degeneration (nAMD), also known as wet AMD, is associated with an abnormal blood vessel growth in the eye and involves an imbalance of proangiogenic and antiangiogenic factors. Thrombospondin (TSP)-1 and TSP-2 are endogenous matricellular proteins that inhibit angiogenesis. TSP-1 is significantly diminished in eyes with AMD, although the mechanisms involved in its reduction are unknown. Granzyme B (GzmB) is a serine protease with an increased extracellular activity in the outer retina and choroid of human eyes with nAMD-related choroidal neovascularization (CNV). This study investigated whether TSP-1 and TSP-2 are GzmB substrates using in silico and cell-free cleavage assays and explored the relationship between GzmB and TSP-1 in human eyes with nAMD-related CNV and the effect of GzmB on TSP-1 in retinal pigment epithelial culture and an explant choroid sprouting assay (CSA). In this study, TSP-1 and TSP-2 were identified as GzmB substrates. Cell-free cleavage assays substantiated the GzmB proteolysis of TSP-1 and TSP-2 by showing dose-dependent and time-dependent cleavage products. TSP-1 and TSP-2 proteolysis were hindered by the inhibition of GzmB. In the retinal pigment epithelium and choroid of human eyes with CNV, we observed a significant inverse correlation between TSP-1 and GzmB, as indicated by lower TSP-1 and higher GzmB immunoreactivity. In CSA, the vascular sprouting area increased significantly with GzmB treatment and reduced significantly with TSP-1 treatment. Western blot showed significantly reduced expression of TSP-1 in GzmB-treated retinal pigment epithelial cell culture and CSA supernatant compared with that in controls. Together, our findings suggest that the proteolysis of antiangiogenic factors such as TSP-1 by extracellular GzmB might represent a mechanism through which GzmB may contribute to nAMD-related CNV. Future studies are needed to investigate whether pharmacologic inhibition of extracellular GzmB can mitigate nAMD-related CNV by preserving intact TSP-1.

Potential roles of branched-chain amino acids in neurodegeneration.

Despite having disease-specific pathologic features and symptoms, neurodegenerative diseases share common mechanisms, such as excitotoxicity, neuroinflammation, and neurotransmitter dysregulation. Although the common underlying cause of these neurodegenerative processes has yet to be identified, accumulating evidence suggests that branched-chain amino acids (BCAAs) could be involved. BCAAs have been shown to not only influence the central levels of neurotransmitters but also to induce excitotoxicity, hyperexcitability, inflammation, and oxidative stress. Furthermore, emerging evidence indicates that BCAA metabolism may be dysregulated in major neurodegenerative diseases, namely Alzheimer's and Parkinson's diseases and amyotrophic lateral sclerosis. In this review, we identified the neurodegenerative mechanisms of BCAAs and outlined their potential roles in neurodegenerative diseases, suggesting that targeting BCAA metabolism may represent a new approach to identifying new therapeutic targets for multifaceted neurodegenerative diseases.

Harnessing Astrocytes and Müller Glial Cells in the Retina for Survival and Regeneration of Retinal Ganglion Cells.

Astrocytes have been associated with the failure of axon regeneration in the central nervous system (CNS), as it undergoes reactive gliosis in response to damages to the CNS and functions as a chemical and physical barrier to axon regeneration. However, beneficial roles of astrocytes have been extensively studied in the spinal cord over the years, and a growing body of evidence now suggests that inducing astrocytes to become more growth-supportive can promote axon regeneration after spinal cord injury (SCI). In retina, astrocytes and Müller cells are known to undergo reactive gliosis after damage to retina and/or optic nerve and are hypothesized to be either detrimental or beneficial to survival and axon regeneration of retinal ganglion cells (RGCs). Whether they can be induced to become more growth-supportive after retinal and optic nerve injury has yet to be determined. In this review, we pinpoint the potential molecular pathways involved in the induction of growth-supportive astrocytes in the spinal cord and suggest that stimulating the activation of these pathways in the retina could represent a new therapeutic approach to promoting survival and axon regeneration of RGCs in retinal degenerative diseases.

Recombinant growth differentiation factor 11 influences short-term memory and enhances Sox2 expression in middle-aged mice.

Previous evidence suggests that a significant decline in cognitive ability begins during middle-age and continues to deteriorate with increase in age. Recent work has demonstrated the potential rejuvenation impact of growth differentiation factor-11 (GDF-11) in aged mice. We carried out experiments to evaluate the impact of a single dose of recombinant (rGDF-11) on short-term visual and spatial memory in middle-aged male mice. On the novel object recognition task, we observed middle-aged mice treated rGDF-11 showed improved performance on the novel object recognition task. However, middle-aged mice did not show increased expression of phosphorylated-Smad2/3, a downstream effector of GDF-11. We noted however that the expression of the transcription factor, Sox2 was increased within the dentate gyrus. Our data suggest that a single injection of rGDF-11 contributes to improvements in cognitive function of middle-aged animals, which may be critical in the preservation of short-term memory capacity in old age.