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How many RBPMS+ retinal ganglion cells (RGCs) does a standard C57BL/6 laboratory mouse have on average and is this number substrain- or sex-dependent? Do RGCs of (European) C57BL/6J and -N mice show a different intrinsic vulnerability upon glaucomatous injury? Global RGC numbers and densities of common laboratory mice were previously determined via axon counts, retrograde tracing or BRN3A immunohistochemistry. Here, we report the global RGC number and density by exploiting the freely available tool RGCode to automatically count RGC numbers and densities on entire retinal wholemounts immunostained for the pan-RGC marker RBPMS. The intrinsic vulnerability of RGCs from different substrains to glaucomatous injury was evaluated upon introduction of the microbead occlusion model, followed by RBPMS counts, retrograde tracing and electroretinography five weeks post-injury. We demonstrate that the global RGC number and density varies between substrains, yet is not sex-dependent. C57BL/6J mice have on average 46K ± 2K RBPMS+ RGCs per retina, representing a global RGC density of 3268 ± 177 RGCs/mm2. C57BL/6N mice, on the other hand, have on average less RBPMS+ RGCs (41K ± 3K RGCs) and a lower density (3018 ± 189 RGCs/mm2). The vulnerability of the RGC population of the two C57BL/6 substrains to glaucomatous injury did, however, not differ in any of the interrogated parameters. [ABSTRACT FROM AUTHOR]

Retinal neurodegeneration and visual dysfunctions have been reported in a majority of Alzheimer's and Parkinson's patients, and, in light of the quest for novel biomarkers for these neurodegenerative proteinopathies, the retina has been receiving increasing attention as an organ for diagnosing, monitoring, and understanding disease. Thinning of retinal layers, abnormalities in vasculature, and protein deposition can be imaged at unprecedented resolution, which offers a unique systems biology view on the cellular and molecular changes underlying these pathologies. It makes the retina not only a promising target for biomarker development, but it also suggests that novel fundamental insights into the pathophysiology of Alzheimer's and Parkinson's disease can be obtained by studying the retina–brain axis. • Retinal changes and visual dysfunctions have been well documented in AD and PD. • Retinal biomarkers may allow early diagnosis and disease monitoring. • The retina offers a view on the cellular and molecular changes underlying AD and PD. • By understanding retina–brain differences, we can grasp AD/PD disease processes. [ABSTRACT FROM AUTHOR]

The rise in life expectancy in our modern societies concurs with an increasing prevalence of age‐associated neurodegenerative disorders and central nervous system (CNS) trauma. Despite many research attempts trying to unravel aging processes and their role in functional decline, the mechanisms underlying healthy CNS aging remain elusive. Moreover, the reparative approaches proposed to combat loss of neurons due to damage or (age‐associated) disease are failing. One of the reasons might be that the currently used animal models often lack the capability to properly recapitulate the aged environment in which these pathologies manifest and the repair processes need to occur. We introduce the African turquoise killifish as an opportune model to study CNS aging as well as neuroreparative strategies. Indeed, while sharing adult neuroregenerative potential with other teleost fish like zebrafish, the short‐lived killifish displays many of the human aging hallmarks, making it a promising biogerontology model. As vision decline manifests during human aging and in many neurodegenerative diseases, we performed a comprehensive characterization of the aged killifish visual system, comprising the retina, optic projections and optic tectum. Immunohistochemical, biochemical and molecular analyses revealed the presence of many aging traits in this system, including oxidative stress, genomic damage, cellular senescence, altered cellular communication, reduced neurogenic potential and even neurodegeneration. Specifically, microglia changed to a pro‐inflammatory secretion profile upon aging and macroglia became gliotic in the aging killifish visual system. Altogether, these age‐related changes led to a diminished visual performance in old killifish. Using an optic nerve injury model, we further disclosed an age‐related decline in functional circuit restoration, with different phases of the repair process being affected depending on the age. While young adult killifish were able to functionally recover after injury, old fish did not regain vision at all. Here we observed for the first time in a regenerative teleost fish model, a glial scar that formed a mechanical barrier physically preventing axonal regrowth. Moreover, in the optic tectum the inflammatory response shifted from acute to chronic upon aging. Macroglia, which showed a transient gliotic response after injury in young adult fish, displayed a permanent response in old animals. Overall, as in mammals, both a reduced intrinsic growth potential and a non‐supportive cellular environment, orchestrated by activated micro‐ and macroglia, were found to underlie the declined repair potential in aged killifish. To decode the cellular and molecular framework that defines visual system aging and the failed regenerative response in old fish, we are currently performing single‐cell RNA sequencing of the young adult and aged killifish retina. We envision our bioinformatic approaches to result in innovative druggable targets for future development of rejuvenative/curative therapies in the aged mammalian retina. In summary, our study highlights the importance of a neuron–glia crosstalk in health, disease and repair. It puts forward the strengths of the killifish as a vertebrate model with mammalian‐like characteristics upon aging, that offers unique opportunities to identify rejuvenating and neuroreparative strategies ensuring healthy aging. [ABSTRACT FROM AUTHOR]

Abstract: Mice with a targeted truncation in the gene encoding tissue factor of blood coagulation (TF) to eliminate the cytosolic domain and carrying a neo R cassette in intron 5 unexpectedly displayed severe spontaneous thrombosis in various vascular beds. Thrombosis was observed in heterozygous TF+/neo mice, causing death of over 50% of adults within 36 weeks of birth, and fulminantly exacerbating in pregnant females. Homozygous TFneo/neo mice were more severely affected and died within 7 weeks after birth. These TFneo mice primarily synthesized a mutant mRNA aberrantly spliced from exon 5 to neo R , encoding an apparently non-vesicle-binding soluble TF lacking both the transmembrane and cytosolic domain, but still capable of blood coagulation induction. This severe thrombotic phenotype associated with the presence of a non-anchored soluble TF variant underscores the recently recognized significance of circulating TF for thrombus formation and development. [Copyright &y& Elsevier]

Hypoxia stimulates angiogenesis through the binding of hypoxia-inducible factors to the hypoxia-response element in the vascular endothelial growth factor (Vegf) promotor. Here, we report that deletion of the hypoxia-response element in the Vegf promotor reduced hypoxic Vegf expression in the spinal cord and caused adult-onset progressive motor neuron degeneration, reminiscent of amyotrophic lateral sclerosis. The neurodegeneration seemed to be due to reduced neural vascular perfusion. In addition, Vegf165 promoted survival of motor neurons during hypoxia through binding to Vegf receptor 2 and neuropilin 1. Acute ischemia is known to cause nonselective neuronal death. Our results indicate that chronic vascular insufficiency and, possibly, insufficient Vegf-dependent neuroprotection lead to the select degeneration of motor neurons. [ABSTRACT FROM AUTHOR]

SY161-P5, a polyethylene glycol derivatized (PEGylated) mutant of the recombinant Staphylokinase (rSak) variant SakSTAR, exhibiting reduced antigenicity, is in clinical development for treatment of acute myocardial infarction as a single bolus injection. A series of safety studies were performed in vivo as a routine toxicology program with SY 161-P5 (PEG-rSakSTAR) and with the recombinant Staphylokinase variant Sak42D (rSak42D). For both compounds, intravenous single bolus injections of up to 100-fold therapeutic equivalent, as well as repeated injections during 7 to 28 days revealed no significant pathological findings in mice, rats or hamsters. However, New Zealand white rabbits developed clinically silent, multifocal myocarditis following single or repeat doses of SY 161-P5 or of Sak42D. These findings were dose-independent and reversible. A similar species-specific cardiotoxic effect has previously been described for other proteolytic proteins, including the approved drugs Streptokinase and Acetylated Plasminogen Streptokinase Complex (APSAC). The large experience with these drugs, as well as the clinical data accumulated both with PEGylated and non-PEGylated rSak variants to date, do not indicate cardiotoxic hazards associated with the use of these drugs in humans. [ABSTRACT FROM AUTHOR]

Vascular endothelial growth factor (VEGF) stimulates angiogenesis by activating VEGF receptor-2 (VEGFR-2). The role of its homolog, placental growth factor (PlGF), remains unknown. Both VEGF and PlGF bind to VEGF receptor-1 (VEGFR-1), but it is unknown whether VEGFR-1, which exists as a soluble or a membrane-bound type, is an inert decoy or a signaling receptor for PlGF during angiogenesis. Here, we report that embryonic angiogenesis in mice was not affected by deficiency of PlGF (Pgf-/-). VEGF-B, another ligand of VEGFR-1, did not rescue development in Pgf-/- mice. However, loss of PlGF impaired angiogenesis, plasma extravasation and collateral growth during ischemia, inflammation, wound healing and cancer. Transplantation of wild-type bone marrow rescued the impaired angiogenesis and collateral growth in Pgf-/- mice, indicating that PlGF might have contributed to vessel growth in the adult by mobilizing bone-marrow?derived cells. The synergism between PlGF and VEGF was specific, as PlGF deficiency impaired the response to VEGF, but not to bFGF or histamine. VEGFR-1 was activated by PlGF, given that anti-VEGFR-1 antibodies and a Src-kinase inhibitor blocked the endothelial response to PlGF or VEGF/PlGF. By upregulating PlGF and the signaling subtype of VEGFR-1, endothelial cells amplify their responsiveness to VEGF during the 'angiogenic switch' in many pathological disorders. [ABSTRACT FROM AUTHOR]

Age‐related vision loss caused by retinal neurodegenerative pathologies is becoming more prevalent in our ageing society. To understand the physiological and molecular impact of ageing on retinal homeostasis, we used the short‐lived African turquoise killifish, a model known to naturally develop central nervous system (CNS) ageing hallmarks and vision loss. Bulk and single‐cell RNA‐sequencing (scRNAseq) of three age groups (6‐, 12‐, and 18‐week‐old) identified transcriptional ageing fingerprints in the killifish retina, unveiling pathways also identified in the aged brain, including oxidative stress, gliosis, and inflammageing. These findings were comparable to observations in the ageing mouse retina. Additionally, transcriptional changes in genes related to retinal diseases, such as glaucoma and age‐related macular degeneration, were observed. The cellular heterogeneity in the killifish retina was characterized, confirming the presence of all typical vertebrate retinal cell types. Data integration from age‐matched samples between the bulk and scRNAseq experiments revealed a loss of cellular specificity in gene expression upon ageing, suggesting potential disruption in transcriptional homeostasis. Differential expression analysis within the identified cell types highlighted the role of glial/immune cells as important stress regulators during ageing. Our work emphasizes the value of the fast‐ageing killifish in elucidating molecular signatures in age‐associated retinal disease and vision decline. This study contributes to the understanding of how age‐related changes in molecular pathways may impact CNS health, providing insights that may inform future therapeutic strategies for age‐related pathologies. [ABSTRACT FROM AUTHOR]

The human brain contains 86 billion neurons [[1]], which are connected to thousands of other neurons and thereby function in a complex spatiotemporal organization that forms the physiological and biological basis of cognition. Comparative neuroscience research may thus hold the key to future restorative therapies for CNS injury and disease in humans. Second, Yusifov and colleagues present novel research on the role of the primary cilium in peripheral nervous system formation and describe differences compared to the ciliation of neurons in the CNS [[12]]. [Extracted from the article]

Presents a study which discusses the role of urokinase-type plasminogen activator (u-PA). Revelation of the topographic analysis of arterial wound healing after electric injury; What formation of blood vessels during embryogenesis involves; Constituency of matrix metalloproteinases (MMPs).

Up to now neuroregenerative research mainly focused on improving axonal regrowth, leaving the dendrites, which form an essential component of the neuronal circuit, largely unstudied. Nevertheless, subtle changes in the dendritic arbors of neurons are one of the first stages of various neurodegenerative diseases, leading to dysfunctional neuronal networks and ultimately cellular death. Maintaining dendrites is therefore considered an essential neuroprotective strategy. Here, we postulate the intriguing hypothesis, that dendritic shrinkage is an important stimulant to boost axonal regeneration, and thus that preserving dendrites might not be the ideal therapeutic method to regain a full functional network upon CNS damage. Indeed, our study in zebrafish, a versatile animal model with robust regenerative capacity, recently unraveled that dendritic retraction is evoked prior to axonal regrowth after optic nerve crush (ONC) injury, and that the retinal ganglion cell (RGC) dendrites regenerate after axonal repair. This orderly sequence of neurite outgrowth is a recapitulation of development, where axogenesis also precedes dendritogenesis. Strikingly, inhibiting RGC dendritic pruning upon damage perturbed axonal regeneration in zebrafish. This constraining effect of dendrites on axonal regrowth has sporadically been proposed in literature. In line with this, we observed an increased loss of RGC dendrites and reduced dendritic arbor complexity in the mouse retina when axonal regeneration was induced by inflammatory stimulation. All together, these data point towards a conserved neuronal remodeling mechanism, where dendritic pruning upon neuronal injury supports axonal regeneration. We now hypothesize that a reprogramming and relocation of the neuronal energy production machinery towards the axon is needed for regeneration, a process in which the shrinking dendrites might play a key role. To test this idea, a combination of state‐of‐the‐art in vitro and in vivo research approaches are being pursued. We are characterizing, in the axonal versus dendritic cell compartment, the subcellular and molecular players modulating energy production and enabling axonal regrowth of injured CNS neurons. Overall, our findings could generate pivotal insights into how re‐directing intra‐neuronal energy channelling may promote neuronal repair in the mammalian CNS. [ABSTRACT FROM AUTHOR]

Neurodegenerative diseases and central nervous system (CNS) trauma are highly irreversible, in part because adult mammals lack a robust regenerative capacity. A multifactorial problem underlies the limited axonal regeneration potential. Strikingly, neuroinflammation seems able to induce axonal regrowth in the adult mammalian CNS. It is increasingly clear that both blood-borne and resident inflammatory cells as well as reactivated glial cells affect axonal regeneration. The scope of this review is to give a comprehensive overview of the knowledge that links inflammation (with a focus on the innate immune system) to axonal regeneration and to critically reflect on the controversy that still prevails about the cells, molecules and pathways that are dominating the scene. Also, a brief overview is given of what is already known about the crosstalk between and the heterogeneity of cell types that might play a role in axonal regeneration. Recent research indicates that inflammation-induced axonal regrowth is not solely driven by a single-cell population but probably relies on the crosstalk between multiple cell types and the strong regulation of these cell populations in time and space. Moreover, there is growing evidence that the different cell populations are highly heterogeneous and as such can react differently upon injury. This could explain the controversial results that have been obtained over the past years. The primary focus of this manuscript is the retinofugal system of adult mammals, however, when relevant, insights or examples of the spontaneous regenerating zebrafish model and spinal cord research are added. [ABSTRACT FROM AUTHOR]

Thanks to medical and technological improvements, our world population has become ever-greying. In consequence, the incidence and prevalence of age-related central nervous system neuropathies, such as Alzheimer's (AD) and Parkinson's disease (PD), are increasing tremendously. Despite many research efforts, the precise aetiology of these age-related neurodegenerative disorders remains elusive, highlighting the urgent need for more effective treatments. Current preclinical research mainly uses animal models that do not fully recapitulate the complex cellular context in which these diseases occur, thereby lacking good construct validity. Indeed, most investigations are performed using relatively young animals, thereby ignoring the ageing environment in which neurodegenerative diseases manifest. This points out a major hiatus in current research: a vertebrate model organism that combines the complex disease context (onset, spreading and further manifestation into functional impairment) with an ageing environment. In recent years, the African turquoise killifish has emerged as a promising novel animal model to study age-related neurodegenerative disorders that combines these essential features. In this review, we bundle all reported findings up till now and provide a detailed overview of the neurodegenerative events within the central nervous system of this teleost fish, with a focus on PD. • Killifish as novel gerontology model to study naturally occurring neurodegeneration. • Parkinson hallmarks: Lewy body formation/propagation and motor/cognitive symptoms. • Signs of Alzheimer's, amyotrophic lateral sclerosis, and frontotemporal dementia. [ABSTRACT FROM AUTHOR]

Abstract Intravenous and/or intrathecal administration of the anti-folate drug methotrexate is a common chemotherapeutic procedure in childhood leukemia. Therapeutic and prophylactic efficacy of these procedures notwithstanding, the occurrence of late adverse effects remains a cause of clinical concern in leukemia survivors. We propose an experimental mouse model to mimic the impact of methotrexate exposure on brain biochemistry and cell proliferation, as well as behavioral and neurocognitive functioning at adult age. Female C57Bl6/J mouse pups received saline or methotrexate injection (20 mg/kg, i.p.). CSF and serum concentrations of folate metabolites and toxicity makers were analyzed at 4 h, 24 h, and 1 week following injection. Behavioral test battery performance was assessed at adult age (3–4 months). We found acute changes in serum and CSF levels of folate in exposed pups that coincided with increases in CSF Tau, whereas homocysteine in serum and CSF, and CSF levels of pTau were unchanged or remained below detection. In addition, methotrexate injection coincided with diminished hippocampal cell proliferation 1 week after methotrexate injection. At adult age, exposed mice displayed hippocampus-dependent deficits in the Morris water maze, whereas exploration and anxiety-related behaviors were largely unaffected. Particularly during the reference memory (probe) trial after reversal learning, methotrexate-exposed animals were less precise than controls. These findings demonstrate adult neurocognitive sequelae in a mouse model that can be attributed to the biochemical and cellular impact of early-life methotrexate exposure. Highlights • Methotrexate-induced folate depletion associated with acute brain toxicity in mice. • Reduced hippocampal cell proliferation after methotrexate injection. • Early-life methotrexate induces adult cognitive deficits in mice. [ABSTRACT FROM AUTHOR]

The multifaceted nature of neuroinflammation is highlighted by its ability to both aggravate and promote neuronal health. While in mammals retinal ganglion cells (RGCs) are unable to regenerate following injury, acute inflammation can induce axonal regrowth. However, the nature of the cells, cellular states and signalling pathways that drive this inflammation-induced regeneration have remained elusive. Here, we investigated the functional significance of macrophages during RGC de- and regeneration, by characterizing the inflammatory cascade evoked by optic nerve crush (ONC) injury, with or without local inflammatory stimulation in the vitreous. By combining single-cell RNA sequencing and fate mapping approaches, we elucidated the response of retinal microglia and recruited monocyte-derived macrophages (MDMs) to RGC injury. Importantly, inflammatory stimulation recruited large numbers of MDMs to the retina, which exhibited long-term engraftment and promoted axonal regrowth. Ligand-receptor analysis highlighted a subset of recruited macrophages that exhibited expression of pro-regenerative secreted factors, which were able to promote axon regrowth via paracrine signalling. Our work reveals how inflammation may promote CNS regeneration by modulating innate immune responses, providing a rationale for macrophage-centred strategies for driving neuronal repair following injury and disease. [ABSTRACT FROM AUTHOR]

Purpose To determine the optimum administration route of bevacizumab after glaucoma filtering surgery ( GFS) and to investigate whether a reduced dose of mitomycin-C ( MMC) in combination with bevacizumab could improve surgical outcome with a reduced incidence of side-effects. Methods Plasma levels of bevacizumab were determined via ELISA after intracameral ( IC), subconjunctival ( SC) and intravitreal ( IV) injections in mice, subjected to a mouse model of GFS. Application of MMC was compared to bevacizumab ( SC, 25 μg) and to the combined use of both adjuvants. Surgical sponges soaked in MMC 0.02% or 0.01% were exposed to the sclera for 1 or 2 min. Treatment outcome was studied by bleb investigation. Results The three administration routes of bevacizumab equally improved surgical outcome. The VEGF antibody was detected at relatively high levels in plasma shortly after IV injection, whereas it was minimally absorbed after IC and SC injections. Both bevacizumab ( SC) and MMC 0.02% (2 min) similarly increased bleb area. As compared to MMC, the combined injection with bevacizumab induced an additional effect on surgical outcome. Exposure of MMC 0.02% for 1 or 2 min together with bevacizumab equally improved surgical outcome, but 2 min application induced corneal toxicity. The combined use of bevacizumab and 1-min MMC 0.01% also improved surgical outcome compared to monotherapy, although to a lesser extent than the combination with 1-min MMC 0.02%. Conclusions Adjunctive bevacizumab not only enhances the beneficial effect of MMC on surgical outcome, but also allows reducing the administration time of MMC 0.02%, thereby eliminating its toxic effects on the cornea. [ABSTRACT FROM AUTHOR]

Matrix metalloproteinases (MMPs) are Zn-dependent endopeptidases considered to be essential for normal brain development and neuroplasticity by modulating extracellular matrix proteins, receptors, adhesion molecules, growth factors and cytoskeletal proteins. Specifically, MMP-3 has recently been implicated in synaptic plasticity, hippocampus-dependent learning and neuronal development and migration in the cerebellum. However, the function(s) of this enzyme in the neocortex is understudied. Therefore, we explored the phenotypical characteristics of the neuronal architecture and the capacity for experience-dependent cortical plasticity in the visual cortex of adult MMP-3-deficient (MMP-3) mice. Golgi-Cox stainings revealed a significant reduction in apical dendritic length and an increased number of apical obliques for layer V pyramidal neurons in the visual cortex of adult MMP-3 mice compared to wild-type (WT) animals. In addition, a significant upregulation of both phosphorylated and non-phosphorylated neurofilament protein (NF)-high, phosphorylated NF-medium, NF-low and α-internexin was detected in the visual cortex of MMP-3 mice. To assess the effect of MMP-3 deficiency on cortical plasticity, we monocularly enucleated adult MMP-3 mice and analyzed the reactivation of the contralateral visual cortex 7 weeks post-enucleation. In contrast to previous results in C57Bl/6J adult mice, activity remained confined to the binocular zone and did not expand into the monocular regions indicative for an aberrant open-eye potentiation. Permanent hypoactivity in the monocular cortex lateral and medial to V1 also indicated a lack of cross-modal plasticity. These observations demonstrate that genetic inactivation of MMP-3 has profound effects on the structural integrity and plasticity response of the visual cortex of adult mice. [ABSTRACT FROM AUTHOR]

One important facet of glaucoma pathophysiology is axonal damage, which ultimately disrupts the connection between the retina and its postsynaptic brain targets. The concurrent loss of retrograde support interferes with the functionality and survival of the retinal ganglion cells (RGCs). Previous research has shown that stimulation of neuronal activity in a primary retinal target area—i.e., the superior colliculus—promotes RGC survival in an acute mouse model of glaucoma. To build further on this observation, we applied repeated chemogenetics in the superior colliculus of a more chronic murine glaucoma model—i.e., the microbead occlusion model—and performed bulk RNA sequencing on collicular lysates and isolated RGCs. Our study revealed that chronic target stimulation upon glaucomatous injury phenocopies the a priori expected molecular response: growth factors were pinpointed as essential transcriptional regulators both in the locally stimulated tissue and in distant, unstimulated RGCs. Strikingly, and although the RGC transcriptome revealed a partial reversal of the glaucomatous signature and an enrichment of pro-survival signaling pathways, functional rescue of injured RGCs was not achieved. By postulating various explanations for the lack of RGC neuroprotection, we aim to warrant researchers and drug developers for the complexity of chronic neuromodulation and growth factor signaling. [ABSTRACT FROM AUTHOR]

The formation of fibrovascular membranes (FVMs) is a serious sight-threatening complication of proliferative diabetic retinopathy (PDR) that may result in retinal detachment and eventual blindness. During the formation of these membranes, neurite/process outgrowth occurs in retinal neurons and glial cells, which may both serve as a scaffold and have guiding or regulatory roles. To further understand this process, we investigated whether previously identified candidate proteins, from vitreous of PDR patients with FVMs, could induce neurite outgrowth in an experimental setting. Retinal explants of C57BL6/N mouse pups on postnatal day 3 (P3) were cultured in poly-L-lysine- and laminin-coated dishes. Outgrowth stimulation experiments were performed with the addition of potential inducers of neurite outgrowth. Automated analysis of neurite outgrowth was performed by measuring β–tubulin-immunopositive neurites using Image J. Expression of PDGF receptors was quantified by RT-PCR in FVMs of PDR patients. Platelet-derived growth factor (PDGF) induced neurite outgrowth in a concentration-dependent manner, whilst neuregulin 1 (NRG1) and connective tissue growth factor (CTGF) did not. When comparing three different PDGF dimers, treatment with PDGF-AB resulted in the highest neurite induction, followed by PDGF-AA and -BB. In addition, incubation of retinal explants with vitreous from PDR patients resulted in a significant induction of neurite outgrowth as compared to non-diabetic control vitreous from patients with macular holes, which could be prevented by addition of CP673451, a potent PDGF receptor (PDGFR) inhibitor. Abundant expression of PDGF receptors was detected in FVMs. Our findings suggest that PDGF may be involved in the retinal neurite outgrowth, which is associated with the formation of FVMs in PDR. [ABSTRACT FROM AUTHOR]

As the mammalian central nervous system matures, its regenerative ability decreases, leading to incomplete or non‐recovery from the neurodegenerative diseases and central nervous system insults that we are increasingly facing in our aging world population. Current neuroregenerative research is largely directed toward identifying the molecular and cellular players that underlie central nervous system repair, yet it repeatedly ignores the aging context in which many of these diseases appear. Using an optic nerve crush model in a novel biogerontology model, that is, the short‐living African turquoise killifish, the impact of aging on injury‐induced optic nerve repair was investigated. This work reveals an age‐related decline in axonal regeneration in female killifish, with different phases of the repair process being affected depending on the age. Interestingly, as in mammals, both a reduced intrinsic growth potential and a non‐supportive cellular environment seem to lie at the basis of this impairment. Overall, we introduce the killifish visual system and its age‐dependent regenerative ability as a model to identify new targets for neurorepair in non‐regenerating individuals, thereby also considering the effects of aging on neurorepair. [ABSTRACT FROM AUTHOR]

The angiogenic mechanism and therapeutic potential of PDGF-CC, a recently discovered member of the VEGF/ PDGF superfamily, remain incompletely characterized. Here we report that PDGF-CC mobilized endothelial progenitor cells in ischemic conditions; induced differentiation of bone marrow cells into ECs; and stimulated migration of ECs. Furthermore, PDGF-CC induced the differentiation of bone marrow cells into smooth muscle cells and stimulated their growth during vessel sprouting. Moreover, delivery of PDGF-CC enhanced postischemic revascularization of the heart and limb. Modulating the activity of PDGF-CC may provide novel opportunities for treating ischemic diseases. [ABSTRACT FROM AUTHOR]

The therapeutic potential of placental growth factor (PlGF) and its receptor Flt1 in angiogenesis is poorly understood. Here, we report that PlGF stimulated angiogenesis and collateral growth in ischemic heart and limb with at least a comparable efficiency to vascular endothelial growth factor (VEGF). An antibody against Flt1 suppressed neovascularization in tumors and ischemic retina, and angiogenesis and inflammatory joint destruction in autoimmune arthritis. Anti-Flt1 also reduced atherosclerotic plaque growth and vulnerability, but the atheroprotective effect was not attributable to reduced plaque neovascularization. Inhibition of VEGF receptor Flk1 did not affect arthritis or atherosclerosis, indicating that inhibition of Flk1-driven angiogenesis alone was not sufficient to halt disease progression. The anti-inflammatory effects of anti-Flt1 were attributable to reduced mobilization of bone marrow?derived myeloid progenitors into the peripheral blood; impaired infiltration of Flt1-expressing leukocytes in inflamed tissues; and defective activation of myeloid cells. Thus, PlGF and Flt1 constitute potential candidates for therapeutic modulation of angiogenesis and inflammation. [ABSTRACT FROM AUTHOR]

The aging central nervous system (CNS) of mammals displays progressive limited regenerative abilities. Recovery after loss of neurons is extremely restricted in the aged brain. Many research models fall short in recapitulating mammalian aging hallmarks or have an impractically long lifespan. We established a traumatic brain injury model in the African turquoise killifish (Nothobranchius furzeri), a regeneration‐competent vertebrate that evolved to naturally age extremely fast. Stab‐wound injury of the aged killifish dorsal telencephalon unveils an impaired and incomplete regeneration response when compared to young individuals. In the young adult killifish, brain regeneration is mainly supported by atypical non‐glial progenitors, yet their proliferation capacity clearly declines with age. We identified a high inflammatory response and glial scarring to also underlie the hampered generation of new neurons in aged fish. These primary results will pave the way to unravel the factor age in relation to neurorepair, and to improve therapeutic strategies to restore the injured and/or diseased aged mammalian CNS. [ABSTRACT FROM AUTHOR]

• ONC triggers autophagy in the retinotectal system of adult zebrafish. • Injured RGCs induce autophagy in their soma and regrowing axons. • Macroglial cells in the optic nerve/optic tectum display autophagy after ONC. • Systemic autophagy inhibition accelerates axonal regeneration. • Increased mTOR activity might underlie the positive effect of autophagy inhibition. Optic neuropathies comprise a group of disorders in which the axons of retinal ganglion cells (RGCs), the retinal projection neurons conveying visual information to the brain, are damaged. This results in visual impairment or even blindness, which is irreversible as adult mammals lack the capacity to repair or replace injured or lost neurons. Despite intensive research, no efficient treatment to induce axonal regeneration in the central nervous system (CNS) is available yet. Autophagy, the cellular recycling response, was shown repeatedly to be elevated in animal models of optic nerve injury, and both beneficial and detrimental effects have been reported. In this study, we subjected spontaneously regenerating adult zebrafish to optic nerve damage (ONC) and revealed that autophagy is enhanced after optic nerve damage in zebrafish, both in RGC axons and somas, as well as in macroglial cells of the retina, the optic nerve and the visual target areas in the brain. Interestingly, the pattern of the autophagic response in the axons followed the spatiotemporal window of axonal regrowth, which suggests that autophagy is ongoing at the growth cones. Pharmacological inhibition of the recycling pathway resulted in accelerated RGC target reinnervation, possibly linked to increased mechanistic target of rapamycin (mTOR) activity, known to stimulate axonal regrowth. Taken together, these intriguing findings underline that further research is warranted to decipher if modulation of autophagy could be an effective therapeutic method to induce CNS regeneration. [ABSTRACT FROM AUTHOR]

Depression is characterized by the disruption of both neural circuitry and neurogenesis. Defects in hippocampal activity and volume, indicative of reduced neurogenesis, are associated with depression-related behaviors in both humans and animals. Neurogenesis in adulthood is considered an activity-dependent process; therefore, hippocampal neurogenesis defects in depression can be a result of defective neural circuitry activity. However, the mechanistic understanding of how defective neural circuitry can induce neurogenesis defects in depression remains unclear. This review highlights the current findings supporting the neural circuitry-regulated neurogenesis, especially focusing on hippocampal neurogenesis regulated by the entorhinal cortex, with regard to memory, pattern separation, and mood. Taken together, these findings may pave the way for future progress in neural circuitry–neurogenesis coupling studies of depression. [ABSTRACT FROM AUTHOR]

Learning and memory are known to depend on synaptic plasticity. Whereas the involvement of plastic changes at excitatory synapses is well established, plasticity mechanisms at inhibitory synapses only start to be discovered. Extracellular proteolysis is known to be a key factor in glutamatergic plasticity but nothing is known about its role at GABAergic synapses. We reveal that pharmacological inhibition of MMP3 activity or genetic knockout of the Mmp3 gene abolishes induction of postsynaptic iLTP. Moreover, the application of exogenous active MMP3 mimics major iLTP manifestations: increased mIPSCs amplitude, enlargement of synaptic gephyrin clusters, and a decrease in the diffusion coefficient of synaptic GABAA receptors that favors their entrapment within the synapse. Finally, we found that MMP3 deficient mice show faster spatial learning in Morris water maze and enhanced contextual fear conditioning. We conclude that MMP3 plays a key role in iLTP mechanisms and in the behaviors that presumably in part depend on GABAergic plasticity. [ABSTRACT FROM AUTHOR]

Glaucoma is a disease associated with the loss of retinal ganglion cells (RGCs), and remains one of the primary causes of blindness worldwide. Major research efforts are presently directed towards the understanding of disease pathogenesis and the development of new therapies, with the help of rodent models as an important preclinical research tool. The ultimate goal is reaching neuroprotection of the RGCs, which requires a tool to reliably quantify RGC survival. Hence, we demonstrate a novel deep learning pipeline that enables fully automated RGC quantification in the entire murine retina. This software, called RGCode (Retinal Ganglion Cell quantification based On DEep learning), provides a user-friendly interface that requires the input of RBPMS-immunostained flatmounts and returns the total RGC count, retinal area and density, together with output images showing the computed counts and isodensity maps. The counting model was trained on RBPMS-stained healthy and glaucomatous retinas, obtained from mice subjected to microbead-induced ocular hypertension and optic nerve crush injury paradigms. RGCode demonstrates excellent performance in RGC quantification as compared to manual counts. Furthermore, we convincingly show that RGCode has potential for wider application, by retraining the model with a minimal set of training data to count FluoroGold-traced RGCs. [ABSTRACT FROM AUTHOR]

In this study, we report the results of a comprehensive phenotyping of the retina of the AppNL-G-F mouse. We demonstrate that soluble Aβ accumulation is present in the retina of these mice early in life and progresses to Aβ plaque formation by midlife. This rising Aβ burden coincides with local microglia reactivity, astrogliosis, and abnormalities in retinal vein morphology. Electrophysiological recordings revealed signs of neuronal dysfunction yet no overt neurodegeneration was observed and visual performance outcomes were unaffected in the AppNL-G-F mouse. Furthermore, we show that hyperspectral imaging can be used to quantify retinal Aβ, underscoring its potential as a biomarker for AD diagnosis and monitoring. These findings suggest that the AppNL-G-F retina mimics the early, preclinical stages of AD, and, together with retinal imaging techniques, offers unique opportunities for drug discovery and fundamental research into preclinical AD. [ABSTRACT FROM AUTHOR]

• Aging undeniably impacts central nervous system functioning and repair. • Ignoring the aging context is detrimental for neuroregenerative research. • Aged regenerating animal models are vital in the search for successful therapies. • Spontaneously regenerating animal models inform about cellular and molecular players. In our ever-aging world population, the risk of age-related neuropathies has been increasing, representing both a social and economic burden to society. Since the ability to regenerate in the adult mammalian central nervous system is very limited, brain trauma and neurodegeneration are often permanent. As a consequence, novel scientific challenges have emerged and many research efforts currently focus on triggering repair in the damaged or diseased brain. Nevertheless, stimulating neuroregeneration remains ambitious. Even though important discoveries have been made over the past decades, they did not translate into a therapy yet. Actually, this is not surprising; while these disorders mainly manifest in aged individuals, most of the research is being performed in young animal models. Aging of neurons and their environment, however, greatly affects the central nervous system and its capacity to repair. This review provides a detailed overview of the impact of aging on central nervous system functioning and regeneration potential, both in non-regenerating and spontaneously regenerating animal models. Additionally, we highlight the need for aging animal models with regenerative capacities in the search for neuroreparative strategies. [ABSTRACT FROM AUTHOR]

• X-ray exposure at E7.5 decreases gene and protein expression of Pax6 and Lhx2 at E9. • Irradiation at E7.5 decreases melanogenic gene expression at E11. • Reduced embryonic eye pigmentation at E11 may precede radiation-induced eye defects. • Radiation exposure during neurulation decreases retinal mitosis at E18. Recent studies highlighted a link between ionizing radiation exposure during neurulation and birth defects such as microphthalmos and anophthalmos. Because the mechanisms underlying these defects remain largely unexplored, we irradiated pregnant C57BL/6J mice (1.0 Gy, X-rays) at embryonic day (E)7.5, followed by histological and gene/protein expression analyses at defined days. Irradiation impaired embryonic development at E9 and we observed a delayed pigmentation of the retinal pigment epithelium (RPE) at E11. In addition, a reduced RNA expression and protein abundance of critical eye-development genes (e.g. Pax6 and Lhx2) was observed. Furthermore, a decreased expression of Mitf, Tyr and Tyrp1 supported the radiation-induced perturbation in RPE pigmentation. Finally, via immunostainings for proliferation (Ki67) and mitosis (phosphorylated histone 3), a decreased mitotic index was observed in the E18 retina after exposure at E7.5. Overall, we propose a plausible etiological model for radiation-induced eye-size defects, with RPE melanogenesis as a major determining factor. [ABSTRACT FROM AUTHOR]

Glaucoma and other optic neuropathies are characterized by axonal transport deficits. Axonal cargo travels back and forth between the soma and the axon terminus, a mechanism ensuring homeostasis and the viability of a neuron. An example of vital molecules in the axonal cargo are neurotrophic factors (NTFs). Hindered retrograde transport can cause a scarcity of those factors in the retina, which in turn can tilt the fate of retinal ganglion cells (RGCs) towards apoptosis. This postulation is one of the most widely recognized theories to explain RGC death in the disease progression of glaucoma and is known as the NTF deprivation theory. For several decades, research has been focused on the use of NTFs as a novel neuroprotective glaucoma treatment. Until now, results in animal models have been promising, but translation to the clinic has been highly disappointing. Are we lacking important knowledge to lever NTF therapies towards the therapeutic armamentarium? Or did we get the wrong end of the stick regarding the NTF deprivation theory? In this review, we will tackle the existing evidence and caveats advocating for and against the target-derived NTF deprivation theory in glaucoma, whilst digging into associated therapy efforts. [ABSTRACT FROM AUTHOR]

mRNA therapeutics have recently experienced a new wave of interest, mainly due to the discovery that chemical modifications to mRNA's molecular structure could drastically reduce its inherent immunogenicity and perceived instability. On this basis, we aimed to explore the potential of chemically stabilized mRNA for ocular applications. More specifically, we investigated the behavior of mRNA-loaded lipid-based carriers in human retinal cells (in vitro) , in bovine retinal explants (ex vivo) and in mouse retinas (in vivo). We demonstrate a clear superiority of mRNA over pDNA to induce protein expression in different retinal cell types, which was further enhanced by chemical modification of the mRNA, providing up to ~1800-fold higher reporter gene expression compared to pDNA. Moreover, transgene expression could be detected for at least 20 days after a single administration of chemically modified mRNA in vitro. We furthermore determined the localization and extent of mRNA expression depending on the administration route. After subretinal (SR) administration, mRNA expression was observed in vivo and ex vivo. By contrast, intravitreal (IVT) administration resulted in limited expression in vivo. Using ex vivo bovine explants with an intact vitreoretinal (VR) interface we could attribute this to the inner limiting membrane (ILM), which presents a large barrier for non-viral delivery of mRNA, trapping mRNA complexes at the vitreal side. When the vitreous was removed, which compromises the ILM, mRNA expression was apparent and seemed to colocalize with Müller cells or photoreceptors after respectively IVT or SR administration. Taken together, this study represents a first step towards mRNA-mediated therapy for retinal diseases. Unlabelled Image [ABSTRACT FROM AUTHOR]

Glaucoma is characterized by a progressive loss of retinal ganglion cells (RGCs) in the eye, which ultimately results in visual impairment or even blindness. Because current therapies often fail to halt disease progression, there is an unmet need for novel neuroprotective therapies to support RGC survival. Various research lines suggest that visual target centers in the brain support RGC functioning and survival. Here, we explored whether increasing neuronal activity in one of these projection areas could improve survival of RGCs in a mouse glaucoma model. Prolonged activation of an important murine RGC target area, the superior colliculus (SC), was established via a novel optogenetic stimulation paradigm. By leveraging the unique channel kinetics of the stabilized step function opsin (SSFO), protracted stimulation of the SC was achieved with only a brief light pulse. SSFO-mediated collicular stimulation was confirmed by immunohistochemistry for the immediate-early gene c-Fos and behavioral tracking, which both demonstrated consistent neuronal activity upon repeated stimulation. Finally, the neuroprotective potential of optogenetic collicular stimulation was investigated in mice of either sex subjected to a glaucoma model and a 63% reduction in RGC loss was found. This work describes a new paradigm for optogenetic collicular stimulation and a first demonstration that increasing target neuron activity can increase survival of the projecting neurons. [ABSTRACT FROM AUTHOR]

Despite decades of research, accurate diagnosis of Parkinson's disease remains a challenge, and disease-modifying treatments are still lacking. Research into the early (presymptomatic) stages of Parkinson's disease and the discovery of novel biomarkers is of utmost importance to reduce this burden and to come to a more accurate diagnosis at the very onset of the disease. Many have speculated that non-motor symptoms could provide a breakthrough in the quest for early biomarkers of Parkinson's disease, including the visual disturbances and retinal abnormalities that are seen in the majority of Parkinson's disease patients. An expanding number of clinical studies have investigated the use of in vivo assessments of retinal structure, electrophysiological function, and vision-driven tasks as novel means for identifying patients at risk that need further neurological examination and for longitudinal follow-up of disease progression in Parkinson's disease patients. Often, the results of these studies have been interpreted in relation to α-synuclein deposits and dopamine deficiency in the retina, mirroring the defining pathological features of Parkinson's disease in the brain. To better understand the visual defects seen in Parkinson's disease patients and to propel the use of retinal changes as biomarkers for Parkinson's disease, however, more conclusive neuropathological evidence for the presence of retinal α-synuclein aggregates, and its relation to the cerebral α-synuclein burden, is urgently needed. This review provides a comprehensive and critical overview of the research conducted to unveil α-synuclein aggregates in the retina of Parkinson's disease patients and animal models, and thereby aims to aid the ongoing discussion about the potential use of the retinal changes and/or visual symptoms as biomarkers for Parkinson's disease. [ABSTRACT FROM AUTHOR]

Placental growth factor (PlGF) is a member of the vascular endothelial growth factor (VEGF) family. Upon binding to VEGF- and neuropilin-receptor sub-types, PlGF modulates a range of neural, glial and vascular cell responses that are distinct from VEGF-A. As PlGF expression is selectively associated with pathological angiogenesis and inflammation, its blockade does not affect the healthy vasculature. PlGF actions have been extensively described in tumor biology but more recently there has been accumulating preclinical evidence that indicates that this growth factor could have an important role in retinal diseases. High levels of PlGF have been found in aqueous humor, vitreous and/or retina of patients exhibiting retinopathies, especially those with diabetic retinopathy (DR) and neovascular age-related macular degeneration (nvAMD). Expression of this growth factor seems to correlate closely with many of the key pathogenic features of early and late retinopathy in preclinical models. For example, studies using genetic modification and/or pharmacological treatment to block PlGF in the laser-induced choroidal neovascularization (CNV) model, oxygen-induced retinopathy model, as well as various murine diabetic models, have shown that PlGF deletion or inhibition can reduce neovascularization, retinal leakage, inflammation and gliosis, without affecting vascular development or inducing neuronal degeneration. Moreover, an inhibitory effect of PlGF blockade on retinal scarring in the mouse CNV model has also been recently demonstrated and was found to be unique for PlGF inhibition, as compared to various VEGF inhibition strategies. Together, these preclinical results suggest that anti-PlGF therapy might have advantages over anti-VEGF treatment, and that it may have clinical applications as a standalone treatment or in combination with anti-VEGF. Additional clinical studies are clearly needed to further elucidate the role of PlGF and its potential as a therapeutic target in ocular diseases. [ABSTRACT FROM AUTHOR]

As adult mammals lack the capacity to replace or repair damaged neurons, degeneration and trauma (and subsequent dysfunction) of the central nervous system (CNS) seriously constrains the patient's life quality. Recent work has shown that appropriate modulation of acute neuroinflammation upon CNS injury can trigger a regenerative response; yet, the underlying cellular and molecular mechanisms remain largely elusive. In contrast to mammals, zebrafish retain high regenerative capacities into adulthood and thus form a powerful model to study the contribution of neuroinflammation to successful regeneration. Here, we used pharmacological immunosuppression methods to study the role of microglia/macrophages during optic nerve regeneration in adult zebrafish. We first demonstrated that systemic immunosuppression with dexamethasone (dex) impedes regeneration after optic nerve injury. Secondly, and strikingly, local intravitreal application of dex or clodronate liposomes prior to injury was found to sensitize retinal microglia. Consequently, we observed an exaggerated inflammatory response to subsequent optic nerve damage, along with enhanced tectal reinnervation. In conclusion, we found a strong positive correlation between the acute inflammatory response in the retina and the regenerative capacity of the optic nerve in adult zebrafish subjected to nerve injury. [ABSTRACT FROM AUTHOR]

Due to the lack of axonal regeneration, age-related deterioration in the central nervous system (CNS) poses a significant burden on the wellbeing of a growing number of elderly. To overcome this regenerative failure and to improve the patient’s life quality, the search for novel regenerative treatment strategies requires valuable (animal) models and techniques. As an extension of the CNS, the retinofugal system, consisting of retinal ganglion cells that send their axons along the optic nerve to the visual brain areas, has importantly contributed to the current knowledge on mechanisms underlying the restricted regenerative capacities and to the development of novel strategies to enhance axonal regeneration. It provides an extensively used research tool, not only in amniote vertebrates including rodents, but also in anamniote vertebrates, such as zebrafish. Indeed, the latter show robust regeneration capacities, thereby providing insights into the factors that contribute to axonal regrowth and proper guidance, complementing studies in mammals. This review provides an integrative and critical overview of the classical and state-of-the-art models and methods that have been employed in the retinofugal system to advance our knowledge on the signaling pathways underlying the restricted versus robust axonal regeneration in rodents and zebrafish, respectively. In vitro, ex vivo and in vivo models and techniques to improve the visualization and analysis of regenerating axons are summarized. As such, the retinofugal system is presented as a valuable model to further facilitate research on axonal regeneration and to open novel therapeutic avenues for CNS pathologies. [ABSTRACT FROM AUTHOR]

Microglia play an important role in the pathology of CNS disorders, however, there remains significant uncertainty about the neuroprotective/degenerative role of these cells due to a lack of techniques to adequately assess their complex behaviour in response to injury. Advancing microscopy techniques, transgenic lines and well-characterized molecular markers, have made histological assessment of microglia populations more accessible. However, there is a distinct lack of tools to adequately extract information from these images to fully characterise microglia behaviour. This, combined with growing economic pressures and the ethical need to minimise the use of laboratory animals, led us to develop tools to maximise the amount of information obtained. This study describes a novel approach, combining image analysis with spatial statistical techniques. In addition to monitoring morphological parameters and global changes in microglia density, nearest neighbour distance, and regularity index, we used cluster analyses based on changes in soma size and roundness to yield novel insights into the behaviour of different microglia phenotypes in a murine optic nerve injury model. These methods should be considered a generic tool to quantitatively assess microglia activation, to profile phenotypic changes into microglia subpopulations, and to map spatial distributions in virtually every CNS region and disease state. [ABSTRACT FROM AUTHOR]

Dysfunction of the central nervous system (CNS) in neurodegenerative diseases or after brain lesions seriously affects life quality of a growing number of elderly, since the adult CNS lacks the capacity to replace or repair damaged neurons. Despite intensive research efforts, full functional recovery after CNS disease and/or injury remains challenging, especially in an aging environment. As such, there is a rising need for an aging model in which the impact of aging on successful regeneration can be studied. Here, we introduce the senescent zebrafish retinotectal system as a valuable model to elucidate the cellular and molecular processes underlying age-related decline in axonal regeneration capacities. We found both intrinsic and extrinsic response processes to be altered in aged fish. Indeed, expression levels of growth-associated genes are reduced in naive and crushed retinas, and the injury-associated increase in innate immune cell density appears delayed, suggesting retinal inflammaging in old fish. Strikingly, however, despite a clear deceleration in regeneration onset and early axon outgrowth leading to an overall slowing of optic nerve regeneration, reinnervation of the optic tectum and recovery of visual function occurs successfully in the aged zebrafish retinotectal system. [ABSTRACT FROM AUTHOR]

The current standard of care in clinical practice for diabetic retinopathy (DR), anti-vascular endothelial growth factor (VEGF) therapy, has shown a significant improvement in visual acuity. However, treatment response can be variable and might be associated with potential side effects. This study was designed to investigate inhibition of placental growth factor (PlGF) as a possible alternative therapy for DR. The effect of the anti-PlGF antibody (PL5D11D4) was preclinically evaluated in various animal models by investigating different DR hallmarks, including inflammation, neurodegeneration, vascular leakage and fibrosis. The in vivo efficacy was tested in diabetic streptozotocin (STZ) and Akimba models and in the laser induced choroidal neovascularization (CNV) mouse model. Intravitreal (IVT) administration of the anti-PlGF antibody was compared to anti-VEGFR-2 antibody (DC101), anti-VEGF antibody (B20), VEGF-Trap (aflibercept) and triamcinolone acetonide (TAAC). Vascular leakage was investigated in the mouse STZ model by fluorescein isothiocyanate labeled bovine serum albumin (FITC-BSA) perfusion and in the Akimba model by fluorescein angiography (FA). Repeated IVT administration of the anti-PlGF antibody reduced vascular leakage, which was comparable to a single administration of VEGFR-2 inhibition in the mouse STZ model. PL5D11D4 treatment did not alter retinal ganglion cell (RGC) density, as demonstrated by Brn3a staining, whereas DC101 significantly reduced RGC number with 20%. Immunohistological stainings were performed to investigate inflammation (CD45, F4/80) and fibrosis (collagen type 1a). In the CNV model, IVT injection(s) of PL5D11D4 dose-dependently reduced inflammation and fibrosis, as compared to PBS treatment. Equimolar single administration of the anti-PlGF antibody and aflibercept (21 nM) and TAAC decreased leukocyte and macrophage infiltration with 50%, whereas DC101 and B20 (21 nM) had no effect on the inflammatory response. Similar results were observed in the mouse STZ model on the number of microglia and macrophages in the retina. Repeated administration of PL5D11D4 (21 nM) and TAAC similarly reduced fibrosis, while no effect was observed after equimolar DC101, B20 nor aflibercept administration (21 nM). In summary, the anti-PlGF antibody showed comparable efficacy as well-characterized VEGF-inhibitor on the process of vascular leakage, but differentiates itself by also reducing inflammation and fibrosis, without triggering a neurodegenerative response. [ABSTRACT FROM AUTHOR]

Growing evidence suggests that mitochondrial dysfunction might play a key role in the pathogenesis of age-related neurodegenerative inner retinal diseases such as diabetic retinopathy and glaucoma. Therefore, the present review provides a perspective on the impact of functional mitochondria in the most predominant glial cells of the retina, the Müller cells. Müller cells span the entire thickness of the neuroretina and are in close proximity to retinal cells including the retinal neurons that provides visual signaling to the brain. Among multiple functions, Müller cells are responsible for the removal of neurotransmitters, buffering potassium, and providing neurons with essential metabolites. Thus, Müller cells are responsible for a stable metabolic dialogue in the inner retina and their crucial role in supporting retinal neurons is indisputable. Müller cell functions require considerable energy production and previous literature has primarily emphasized glycolysis as the main energy provider. However, recent studies highlight the need of mitochondrial ATP production to upheld Müller cell functions. Therefore, the present review aims to provide an overview of the current evidence on the impact of mitochondrial functions in Müller cells. [ABSTRACT FROM AUTHOR]

Dysfunction of photoreceptors, retinal pigment epithelium (RPE) or both contribute to the initiation and progression of several outer retinal disorders. Disrupted Müller glia function might additionally subsidize to these diseases. Mitochondrial malfunctioning is importantly associated with outer retina pathologies, which can be classified as primary and secondary mitochondrial disorders. This review highlights the importance of oxidative stress and mitochondrial DNA damage, underlying outer retinal disorders. Indeed, the metabolically active photoreceptors/RPE are highly prone to these hallmarks of mitochondrial dysfunction, indicating that mitochondria represent a weak link in the antioxidant defenses of outer retinal cells. [ABSTRACT FROM AUTHOR]

Purpose: Diabetic retinopathy (DR) is characterized by an early stage of inflammation and vessel leakage, and an advanced vasoproliferative stage. Also, neurodegeneration might play an important role in disease pathogenesis. The aim of this study was to investigate the effect of the Rho kinase (ROCK) inhibitor, AMA0428, on these processes. Methods: The response to ROCK inhibition by AMA0428 (1 µg) was studiedin vivousing the murine model for streptozotocin (STZ)-induced diabetes, focusing on early non-proliferative DR features and the oxygen-induced retinopathy (OIR) model to investigate proliferative DR. Intravitreal (IVT) administration of AMA0428 was compared with murine anti-VEGF-R2 antibody (DC101, 6.2 µg) and placebo (H2O/PEG; 1C8). Outcome was assessed by analyzing leukostasis using fluorescein isothiocyanate coupled concanavalin A (FITC-ConA) and vessel leakage (bovine serum albumin conjugated with fluorescein isothiocyanate; FITC-BSA)/neovascularization and neurodegeneration by immunohistological approaches (hematoxylin and eosin (H&E), terminal deoxynucleotidyl transferase–mediated biotinylated UTP nick end labeling (TUNEL), Brn3a). ELISA and Western blotting were employed to unravel the consequences of ROCK inhibition (1 µM AMA0428) on myosin phosphatase target protein (MYPT)-1 phosphorylation, endothelial nitric oxide synthase (eNOS) phosphorylation, and vascular endothelial growth factor (VEGF) levels in retinas of diabetic mice, on NF-κβ activity and ICAM-1 expression in endothelial cells (ECs). Results: In vivo, AMA0428 significantly reduced vessel leakage and neovascularization, respectively, in the STZ and OIR model, comparable to DC101 therapy. Additionally, the ROCK inhibitor decreased neurodegeneration in both models and inhibited leukostasis by 30% (p< 0.05) in the STZ model (p< 0.05), while DC101 had no positive effect on the outcome of these latter processes. ROCK activity was upregulated in the diabetic retina and AMA0428 administration resulted in decreased phospho-MYPT-1, enhanced phospho-eNOS, and reduced VEGF levels.In vitro, AMA0428 interfered with NF-κβ activity, thereby inhibiting ICAM-1 expression in ECs. Conclusions: Targeting ROCK with AMA0428 effectively attenuated outcome in an early DR model (STZ) and a late vasoproliferative retinopathy model (OIR). These findings make AMA0428 a promising candidate with an additional anti-inflammatory and neuroprotective benefit for DR patients, as compared with anti-VEGF treatment. [ABSTRACT FROM PUBLISHER]

Long-term potentiation (LTP) is widely perceived as a memory substrate and in the hippocampal CA3-CA1 pathway, distinct forms of LTP depend on NMDA receptors (nmdaLTP) or L-type voltage-gated calcium channels (vdccLTP). LTP is also known to be effectively regulated by extracellular proteolysis that is mediated by various enzymes. Herein, we investigated whether in mice hippocampal slices these distinct forms of LTP are specifically regulated by different metalloproteinases (MMPs). We found that MMP-3 inhibition or knock-out impaired late-phase LTP in the CA3-CA1 pathway. Interestingly, late-phase LTP was also decreased by MMP-9 blockade. When both MMP-3 and MMP-9 were inhibited, both early- and late-phase LTP was impaired. Using immunoblotting, in situ zymography, and immunofluorescence, we found that LTP induction was associated with an increase in MMP-3 expression and activity in CA1 stratum radiatum. MMP-3 inhibition and knock-out prevented the induction of vdccLTP, with no effect on nmdaLTP. L-type channel-dependent LTP is known to be impaired by hyaluronic acid digestion. We found that slice treatment with hyaluronidase occluded the effect of MMP-3 blockade on LTP, further confirming a critical role for MMP-3in this form of LTP. Incontrast to the CA3-CA1 pathway, LTP in the mossy fiber-CA3 projection did not depend on MMP-3, indicating the pathway specificity of the actions of MMPs. Overall, our study indicates that the activation of perisynaptic MMP-3 supports L-type channel-dependent LTP in the CA1 region, whereas nmdaLTP depends solely on MMP-9. [ABSTRACT FROM AUTHOR]

Damage to the central nervous system (CNS) is one of the leading causes of morbidity and mortality in elderly, as repair after lesions or neurodegenerative disease usually fails because of the limited capacity of CNS regeneration. The causes underlying this limited regenerative potential are multifactorial, but one critical aspect is neuroinflammation. Although classically considered as harmful, it is now becoming increasingly clear that inflammation can also promote regeneration, if the appropriate context is provided. Here, we review the current knowledge on how acute inflammation is intertwined with axonal regeneration, an important component of CNS repair. After optic nerve or spinal cord injury, inflammatory stimulation and/or modification greatly improve the regenerative outcome in rodents. Moreover, the hypothesis of a beneficial role of inflammation is further supported by evidence from adult zebrafish, which possess the remarkable capability to repair CNS lesions and even restore functionality. Lastly, we shed light on the impact of aging processes on the regenerative capacity in the CNS of mammals and zebrafish. As aging not only affects the CNS, but also the immune system, the regeneration potential is expected to further decline in aged individuals, an element that should definitely be considered in the search for novel therapeutic strategies. [ABSTRACT FROM AUTHOR]

Matrix metalloproteinase-3 (MMP-3) is known to mediate neuroinflammatory processes by activating microglia, disrupting blood-central nervous system barriers and supporting neutrophil influx into the brain. In addition, the posterior part of the eye, more specifically the retina, the retinal pigment epithelium (RPE) and the blood-retinal barrier, is affected upon neuroinflammation, but a role for MMP-3 during ocular inflammation remains elusive. We investigated whether MMP-3 contributes to acute inflammation in the eye using the endotoxin-induced uveitis (EIU) model. Systemic administration of lipopolysaccharide induced an increase in MMP-3 mRNA and protein expression level in the posterior part of the eye. MMP-3 deficiency or knockdown suppressed retinal leukocyte adhesion and leukocyte infiltration into the vitreous cavity in mice subjected to EIU. Moreover, retinal and RPE mRNA levels of intercellular adhesion molecule 1 (Icam1), interleukin 6 (Il6), cytokine-inducible nitrogen oxide synthase (Nos2) and tumor necrosis factorα (Tnfα), which are key molecules involved in EIU, were clearly reduced in MMP-3 deficient mice. In addition, loss of MMP-3 repressed the upregulation of the chemokines monocyte chemoattractant protein (MCP)-1 and (C-X-C motif) ligand 1 (CXCL1). These findings suggest a contribution of MMP-3 during EIU, and its potential use as a therapeutic drug target in reducing ocular inflammation. [ABSTRACT FROM AUTHOR]

ABSTRACT Overcoming the failure of axon regeneration in the mammalian central nervous system (CNS) after injury remains a major challenge, which makes the search for proregenerative molecules essential. Matrix metalloproteinases (MMPs) have been implicated in axonal outgrowth during CNS development and show increased expression levels during vertebrate CNS repair. In mammals, MMPs are believed to alter the suppressive extracellular matrix to become more permissive for axon regrowth. We investigated the role of MMPs in axonal regeneration following optic nerve crush (ONC) in adult zebrafish, which fully recover from such injuries due to a high intrinsic axon growth capacity and a less inhibitory environment. Lowering general retinal MMP activity through intravitreal injections of GM6001 after ONC strongly reduced retinal ganglion cell (RGC) axonal regrowth, without influencing RGC survival. Based on a recently performed transcriptome profiling study, the expression pattern of four MMPs after ONC was determined via combined use of western blotting and immunostainings. Mmp-2 and -13a were increasingly present in RGC somata during axonal regrowth. Moreover, Mmp-2 and -9 became upregulated in regrowing RGC axons and inner plexiform layer (IPL) synapses, respectively. In contrast, after an initial rise in IPL neurites and RGC axons during the injury response, Mmp-14 expression decreased during regeneration. Altogether, a phase-dependent expression pattern for each specific MMP was observed, implicating them in axonal regrowth and inner retina remodeling after injury. In conclusion, these data suggest a novel, neuron-intrinsic function for multiple MMPs in axon regrowth that is distinct from breaking down environmental barriers. J. Comp. Neurol. 524:1472-1493, 2016. © 2015 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Mouse disease models have proven indispensable in glaucoma research, yet the complexity of the vast number of models and mouse strains has also led to confusing findings. In this study, we evaluated baseline intraocular pressure, retinal histology, and retinofugal projections in three mouse strains commonly used in glaucoma research, i.e. C57Bl/6, C57Bl/6-Tyr c , and CD-1 mice. We found that the mouse strains under study do not only display moderate variations in their intraocular pressure, retinal architecture, and retinal ganglion cell density, also the retinofugal projections to the dorsal lateral geniculate nucleus and the superior colliculus revealed striking differences, potentially underlying diverging optokinetic tracking responses and visual acuity. Next, we reviewed the success rate of three models of (glaucomatous) optic neuropathies (intravitreal N-methyl- d -aspartic acid injection, optic nerve crush, and laser photocoagulation-induced ocular hypertension), looking for differences in disease susceptibility between these mouse strains. Different genetic backgrounds and albinism led to differential susceptibility to experimentally induced retinal ganglion cell death among these three mouse strains. Overall, CD-1 mice appeared to have the highest sensitivity to retinal ganglion cell damage, while the C57Bl/6 background was more resistant in the three models used. [ABSTRACT FROM AUTHOR]