Small extracellular vesicles derived from human induced pluripotent stem cell-differentiated neural progenitor cells mitigate retinal ganglion cell degeneration in a mouse model of optic nerve injury.

JOURNAL/nrgr/04.03/01300535-202502000-00034/figure1/v/2024-05-28T214302Z/r/image-tiff Several studies have found that transplantation of neural progenitor cells (NPCs) promotes the survival of injured neurons. However, a poor integration rate and high risk of tumorigenicity after cell transplantation limits their clinical application. Small extracellular vesicles (sEVs) contain bioactive molecules for neuronal protection and regeneration. Previous studies have shown that stem/progenitor cell-derived sEVs can promote neuronal survival and recovery of neurological function in neurodegenerative eye diseases and other eye diseases. In this study, we intravitreally transplanted sEVs derived from human induced pluripotent stem cells (hiPSCs) and hiPSCs-differentiated NPCs (hiPSC-NPC) in a mouse model of optic nerve crush. Our results show that these intravitreally injected sEVs were ingested by retinal cells, especially those localized in the ganglion cell layer. Treatment with hiPSC-NPC-derived sEVs mitigated optic nerve crush-induced retinal ganglion cell degeneration, and regulated the retinal microenvironment by inhibiting excessive activation of microglia. Component analysis further revealed that hiPSC-NPC derived sEVs transported neuroprotective and anti-inflammatory miRNA cargos to target cells, which had protective effects on RGCs after optic nerve injury. These findings suggest that sEVs derived from hiPSC-NPC are a promising cell-free therapeutic strategy for optic neuropathy.

Dietary glutamine supplementation improves both Th1 and Th17 responses via CARD11-mTORC1 pathway in murine model of atopic dermatitis.

Glutamine (GLN) is considered an immunomodulatory nutrient, while caspase recruitment domain 11 (CARD11) is a susceptibility locus for atopic dermatitis (AD). T-cell antigen receptor (TCR)-stimulated GLN uptake requires CARD11. However, the specific pathogenesis of AD via GLN uptake remains unclear. This study aimed to elucidate the association between dietary GLN supplementation and the CARD11 pathway in the pathogenesis of AD, focusing on T helper type 1 (Th1) and Th17 cell expression in AD. Herein, wild-type (WT) mice with house dust mite epidermal-sensitized skin exhibited increased expression of interferon-gamma (IFN-gamma) and interleukin (IL)-17, whereas CARD11 deficiency impaired Th1 and Th17 responses at the same site. CARD11 is a key mediator of Th1 and Th17 expression in AD. Additionally, we suppressed mammalian target of rapamycin complex 1 (mTORC1) signaling, downstream of CARD11, to underscore the critical role of CARD11 in mediating Th1 and Th17 expression in AD. Further, dietary supplementation of GLN to CARD11-/- mice restored Th1 and Th17 responses, whereas inflammatory expression was reduced in WT mice, and p-CARD11 expression and mTORC1 signaling activity were increased in JPM50.6 cells and CARD11-/- mice. Upon inhibiting the GLN transporter, alanine-serine-cysteine transporter carrier 2 (ASCT2), we observed that the Th1 and Th17 response in AD was reduced. Conclusively, ASCT2-mediated GLN uptake improves the expression of Th1 and Th17 cells via CARD11-mTORC1 signaling pathway in AD, suggesting the potential of glutamine supplementation for AD treatment.

Circulating Small Extracellular Vesicles Involved in Systemic Regulation Respond to RGC Degeneration in Glaucoma.

Glaucoma is a leading cause of irreversible blindness worldwide and is characterized by progressive retinal ganglion cell (RGC) degeneration and vision loss. Since irreversible neurodegeneration occurs before diagnosable, early diagnosis and effective neuroprotection are critical for glaucoma management. Small extracellular vesicles (sEVs) are demonstrated to be potential novel biomarkers and therapeutics for a variety of diseases. In this study, it is found that intravitreal injection of circulating plasma-derived sEVs (PDEV) from glaucoma patients ameliorated retinal degeneration in chronic ocular hypertension (COH) mice. Moreover, it is found that PDEV-miR-29s are significantly upregulated in glaucoma patients and are associated with visual field defects in progressed glaucoma. Subsequently, in vivo and in vitro experiments are conducted to investigate the possible function of miR-29s in RGC pathophysiology. It is showed that the overexpression of miR-29b-3p effectively prevents RGC degeneration in COH mice and promotes the neuronal differentiation of human induced pluripotent stem cells (hiPSCs). Interestingly, engineered sEVs with sufficient miR-29b-3p delivery exhibit more effective RGC protection and neuronal differentiation efficiency. Thus, elevated PDEV-miR-29s may imply systemic regulation to prevent RGC degeneration in glaucoma patients. This study provides new insights into PDEV-based glaucoma diagnosis and therapeutic strategies for neurodegenerative diseases.

Plasma exosomes from patients with active thyroid-associated orbitopathy induce inflammation and fibrosis in orbital fibroblasts.

BACKGROUND: The pathogenesis of thyroid-associated orbitopathy (TAO) remains incompletely understand. The interaction between immunocytes and orbital fibroblasts (OFs) play a critical role in orbital inflammatory and fibrosis. Accumulating reports indicate that a significant portion of plasma exosomes (Pla-Exos) are derived from immune cells; however, their impact upon OFs function is unclear. METHODS: OFs were primary cultured from inactive TAO patients. Exosomes isolated from plasma samples of patients with active TAO and healthy controls (HCs) were utilized for functional and RNA cargo analysis. Functional analysis in thymocyte differentiation antigen-1+ (Thy-1+) OFs measured expression of inflammatory and fibrotic markers (mRNAs and proteins) and cell activity in response to Pla-Exos. RNA cargo analysis was performed by RNA sequencing and RT-qPCR. Thy-1+ OFs were transfected with miR-144-3p mimics/inhibitors to evaluate its regulation of inflammation, fibrosis, and proliferation. RESULTS: Pla-Exos derived from active TAO patients (Pla-ExosTAO-A) induced stronger production of inflammatory cytokines and hyaluronic acid (HA) in Thy-1+ OFs while inhibiting their proliferation. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis and single sample gene set enrichment analysis (ssGSEA) suggested that the difference in mRNA expression levels between Pla-ExosTAO-A and Pla-ExosHC was closely related to immune cells. Differential expression analysis revealed that 62 upregulated and 45 downregulated miRNAs in Pla-ExosTAO-A, with the elevation of miR-144-3p in both Pla-Exos and PBMCs in active TAO group. KEGG analysis revealed that the target genes of differentially expressed miRNA and miR-144-3p enriched in immune-related signaling pathways. Overexpression of the miR-144-3p mimic significantly upregulated the secretion of inflammatory cytokines and HA in Thy-1+ OFs while inhibiting their proliferation. CONCLUSION: Pla-Exos derived from patients with active TAO were immune-active, which may be a long-term stimulus casual for inflammatory and fibrotic progression of TAO. Our finding suggests that Pla-Exos could be used as biomarkers or treatment targets in TAO patients.

Drp1-dependent mitochondrial fragmentation mediates photoreceptor abnormalities in type 1 diabetic retina.

Recent studies have highlighted that retinal neurodegeneration precedes microvascular changes in diabetic retinopathy (DR), but the specific mechanisms remain unclear. Given the pivotal role of dysfunctional mitochondria and oxidative stress in early DR, our objective was to observe mitochondria-related alterations in the neural retina of type one diabetic mellitus mice with no evidence of DR (T1DM-NDR). We aimed to identify the key mitochondrial-related proteins contributing to mitochondrial injury. Our study revealed that T1DM-NDR mice exhibited outer retina thinning, including the ellipsoid zone, inner segment, and outer segment. Additionally, there was an impaired amplitude of the b-wave in electroretinogram (ERG) and a disorganized arrangement of the photoreceptor layer. In both the retina of DM mice and high glucose (HG)-treated 661w cells, mitochondria appeared swollen and fragmented, with disrupted cristae, disorganized or shortened branches in the mitochondrial network, and decreased mitochondrial membrane potential. Among the mitochondrial-related proteins, dynamin-related protein 1 (Drp1) was upregulated, and the ratio of phosphorylated Drp1 protein at serine 616 (S616) and serine 637 (S637) sites significantly increased in the retina of DM mice. The administration of Mdivi-1 ameliorated high-glucose-induced dysfunctional mitochondria, thereby protecting T1DM-NDR mice retina from morphological and functional injuries. Our findings suggest that hyperglycemia promotes Drp1-mediated mitochondrial dysfunction, which may be a significant factor in the development of DR. The inhibition of high-glucose-induced mitochondrial fission emerges as a potential and innovative intervention strategy for preventing DR.

Adenine base editor-mediated splicing remodeling activates noncanonical splice sites.

Adenine base editors (ABEs) are genome-editing tools that have been harnessed to introduce precise A•T to G•C conversion. The discovery of split genes revealed that all introns contain two highly conserved dinucleotides, canonical "AG" (acceptor) and "GT" (donor) splice sites. ABE can directly edit splice acceptor sites of the adenine (A) base, leading to aberrant gene splicing, which may be further adopted to remodel splicing. However, spliced isoforms triggered with ABE have not been well explored. To address it, we initially generated a cell line harboring C-terminal enhanced GFP (eGFP)-tagged ß-actin (ACTB), in which the eGFP signal can track endogenous ß-actin expression. Expectedly, after the editing of splice acceptor sites, we observed a dramatical decrease in the percentage of eGFP-positive cells and generation of splicing products with the noncanonical splice site. Furthermore, we manipulated Peroxidasin in mouse embryos with ABE, in which a noncanonical acceptor was activated to remodel splicing, successfully generating a mouse disease model of anophthalmia and severely malformed microphthalmia. Collectively, we demonstrate that ABE-mediated splicing remodeling can activate a noncanonical acceptor to manipulate human and mouse genomes, which will facilitate the investigation of basic and translational medicine studies.

Schwann cell-derived extracellular vesicles as a potential therapy for retinal ganglion cell degeneration.

Optic neuropathy is the leading cause of irreversible blindness and is characterized by progressive degeneration of retinal ganglion cells (RGCs). Several studies have demonstrated that transplantation of Schwann cells (SCs) is a promising candidate therapy for optic neuropathy and that intravitreally transplanted cells exert their effect via paracrine actions. Extracellular vesicle (EV)-based therapies are increasingly recognized as a potential strategy for cell replacement therapy. In this study, we aimed to investigate the neuroprotective and regenerative effects of SC-EVs following optic nerve injury. We found that SC-EVs were internalized by RGCs in vitro and in vivo without any transfection reagents. Intriguingly, SC-EVs significantly enhanced the survival and axonal growth of primary RGCs in a coculture system. In a rat optic nerve crush model, SC-EVs mitigated RGC degeneration, prevented RGC loss, and preserved the thickness of the ganglion cell complex, as demonstrated by the statistically significant improvement in RGC counts and thickness measurements. Mechanistically, SC-EVs activated the cAMP-response element binding protein (CREB) signaling pathway and regulated reactive gliosis in ONC rats, which is crucial for RGC protection and axonal regeneration. These findings provide novel insights into the neuroprotective and regenerative properties of SC-EVs, suggesting their potential as a cell-free therapeutic strategy and natural biomaterials for neurodegenerative diseases of the central nervous system.

A Novel Heterozygous TGFBI c.1613C>A Pathogenic Variant is Associated With Lattice Corneal Dystrophy in a Chinese Family.

PURPOSE: To investigate the gene mutations and relationship of clinical manifestation in a Chinese family with familial lattice corneal dystrophy (LCD). DESIGN: Single-family case-control study. METHODS: A family with familial LCD was recruited for this study. A total of 10 affected and 13 healthy family members participated in this research. Clinical features were examined by slit-lamp examination and anterior segment optical coherence tomography (AS-OCT). Peripheral blood samples were collected from each participant, and genomic DNA was extracted. Whole-exome sequencing (WES) analysis was performed, and the pathogenic variants of LCD were identified using bioinformatics tools and confirmed by Sanger sequencing. RESULTS: Slit-lamp examination revealed diffuse grayish-white punctate, linear, and "lattice-like" opacities in the corneal epithelium and superficial corneal stroma. AS-OCT revealed an irregularly shaped cornea. The corneal epithelium and anterior corneal stroma showed high-reflective deposits and bulges. The clinical appearance of the patients fit the pattern and features of autosomal dominant inheritance of LCD type I (LCD I). A novel pathogenic variant of exon 12 in TGFBI was found by WES analysis, in which cytosine at position 1613 was substituted by adenine (c.1613C>A), and the amino acid was changed from threonine to lysine (p.T538K). Mutated genes and proteins were predicted to be deleterious. CONCLUSION: A novel heterozygous pathogenic variant (c.1613C>A) of TGFBI was identified in the Chinese family with LCD I.

Srgap2 suppression ameliorates retinal ganglion cell degeneration in mice.

Slit-Robo GTPase-activating protein 2 (SRGAP2) plays important roles in axon guidance, neuronal migration, synapse formation, and nerve regeneration. However, the role of SRGAP2 in neuroretinal degenerative disease remains unclear. In this study, we found that SRGAP2 protein was first expressed in the retina of normal mice at the embryonic stage and was mainly located in the mature retinal ganglion cell layer and the inner nuclear layer. SRGAP2 protein in the retina and optic nerve increased after optic nerve crush. Then, we established a heterozygous knockout (Srgap2+/-) mouse model of optic nerve crush and found that Srgap2 suppression increased retinal ganglion cell survival, lowered intraocular pressure, inhibited glial cell activation, and partially restored retinal function. In vitro experiments showed that Srgap2 suppression activated the mammalian target of rapamycin signaling pathway. RNA sequencing results showed that the expression of small heat shock protein genes (Cryaa, Cryba4, and Crygs) related to optic nerve injury were upregulated in the retina of Srgap2+/- mice. These results suggest that Srgap2 suppression reduced the robust activation of glial cells, activated the mammalian target of rapamycin signaling pathway related to nerve protein, increased the expression of small heat shock protein genes, inhibited the degeneration of retinal ganglion cells, and partially restored optic nerve function.

Vitreous metabolomic signatures of pathological myopia with complications.

BACKGROUND: Pathological myopia (PM) is closely associated with blinding ocular morbidities. Identifying biomarkers can provide clues on pathogeneses. This study aimed to identify metabolic biomarkers and underlying mechanisms in the vitreous humour (VH) of PM patients with complications. METHODS: VH samples were collected from 39 PM patients with rhegmatogenous retinal detachment (RRD) (n = 23) or macular hole (MH)/myopic retinoschisis (MRS) (n = 16) and 23 controls (MH with axial length < 26 mm) who underwent surgical treatment. VH metabolomic profiles were investigated using ultra-performance liquid chromatography‒mass spectrometry. The area under the receiver operating characteristic curve (AUC) was computed to identify potential biomarkers for PM diagnosis. RESULTS: Bioinformatics analysis identified nineteen and four metabolites altered in positive and negative modes, respectively, and these metabolites were involved in tryptophan metabolism. Receiver operating characteristic analysis showed that seventeen metabolites (AUC > 0.6) in the positive mode and uric acid in the negative mode represent potential biomarkers for PM with complications (AUC = 0.894). Pairwise and pathway analyses among the RRD-PM, MH/MRS-PM and control groups showed that tryptophan metabolism and uric acid were closely correlated with PM. Altered metabolites and pathways in our study were characterized by increased oxidative stress and altered energy metabolism. These results contribute to a better understanding of myopia progression with or without related complications. CONCLUSIONS: Our study provides metabolomic signatures and related immunopathological features in the VH of PM patients, revealing new insight into the prevention and treatment of PM and related complications.

Anti-angiogenic properties of microRNA-29a in preclinical ocular models.

Abnormal neovascularization is an important cause of blindness in many ocular diseases, for which the etiology and pathogenic mechanisms remain incompletely understood. Recent studies have revealed the diverse roles of noncoding RNAs in various biological processes and facilitated the research and development of the clinical application of numerous RNA drugs, including microRNAs. Here, we report the antiangiogenic activity of microRNA-29a (miR-29a) in three animal models of ocular neovascularization. The miR-29a knockout (KO) mice displayed enhanced vessel pruning, resulting in a decreased vascularized area during retinal development. In contrast, miR-29a deletion in adult mice accelerated angiogenesis in preclinical disease models, including corneal neovascularization, oxygen-induced retinopathy, and choroidal neovascularization, while the administration of agomir-29a ameliorated pathological neovascularization. Furthermore, miR-29a exerted inhibitory effects on endothelial cell proliferation, migration, and tube formation capacities. RNA sequencing analysis of retinas from miR-29a KO mice and RNA interference experiments identified platelet-derived growth factor C and several extracellular matrix genes as downstream targets of miR-29a involved in regulating ocular angiogenesis. Our data suggest that miR-29a may be a promising clinical candidate for the treatment of neovascular diseases.

The Expression of Rab8, Ezrin, Radixin and Moesin in the Ciliary Body of Cynomolgus Monkeys.

Purpose: The purpose of this study was to determine what proteins are present in the ciliary body (CB). To accomplish this, we conducted a proteomic analysis of the CB of cynomolgus monkeys. We also determined the location of the proteins in CB by immunohistology. Methods: The eyes of euthanized cynomolgus monkeys were enucleated, and the CB, were isolated from the eyes. Proteins were extracted from the CB and determined by liquid chromatography-mass spectrometry. Separated CB epithelial cells were cultured, and the proteins expressed in the CB were determined by Western blotting. The location of these proteins in the CB was determined by immunohistochemical staining. We also investigated whether adding dexamethasone to the culture medium changed protein expression by the epithelial cells. Results: Proteomic analysis of the CBs showed that 813 proteins were expressed in the epithelium and stroma. These proteins included the small guanosine triphosphate-binding protein Rab8 and the ezrin/radixin/moesin (ERM) family. Tissue and immunohistological staining confirmed the colocalization of these proteins in non-pigmented CB epithelium. Western blotting of cultured CB epithelial cell lysates showed a tendency that adding dexamethasone changed Rab8 protein expression levels. Conclusions: Proteomic analysis of CBs identified several proteins involved in the transport and secretion of proteins. These proteins may be involved in the production of aqueous humor and protein secretion by the CB.

Suppressor of Cytokine Signaling 2 Regulates Retinal Pigment Epithelium Metabolism by Enhancing Autophagy.

Retinal pigment epithelium (RPE) serves critical functions in maintaining retinal homeostasis. An important function of RPE is to degrade the photoreceptor outer segment fragments daily to maintain photoreceptor function and longevity throughout life. An impairment of RPE functions such as metabolic regulation leads to the development of age-related macular degeneration (AMD) and inherited retinal degenerative diseases. As substrate recognition subunit of a ubiquitin ligase complex, suppressor of cytokine signaling 2 (SOCS2) specifically binds to the substrates for ubiquitination and negatively regulates growth hormone signaling. Herein, we explore the role of SOCS2 in the metabolic regulation of autophagy in the RPE cells. SOCS2 knockout mice exhibited the irregular morphological deposits between the RPE and Bruch's membrane. Both in vivo and in vitro experiments showed that RPE cells lacking SOCS2 displayed impaired autophagy, which could be recovered by re-expressing SOCS2. SOCS2 recognizes the ubiquitylated proteins and participates in the formation of autolysosome by binding with autophagy receptors and lysosome-associated membrane protein2 (LAMP-2), thereby regulating the phosphorylation of glycogen synthase kinase 3ß (GSK3ß) and mammalian target of rapamycin (mTOR) during the autophagy process. Our results imply that SOCS2 participates in ubiquitin-autophagy-lysosomal pathway and enhances autophagy by regulating GSK3ß and mTOR. This study provides a potential therapeutic target for AMD.

Whole transcriptome sequencing identifies key circRNAs, lncRNAs, and miRNAs regulating neurogenesis in developing mouse retina.

BACKGROUND: The molecular complexity of neural retina development remains poorly studied. Knowledge of retinal neurogenesis regulation sheds light on retinal degeneration therapy exploration. Therefore, we integrated the time-series circRNA, lncRNA, miRNA, and mRNA expression profiles of the developing retina through whole-transcriptome sequencing. The key functional ncRNAs and the ceRNA network regulating retinal neurogenesis were identified. RESULTS: Transcriptomic analysis identified circRNA as the most variable ncRNA subtype. We screened a series of neurogenesis-related circRNAs, lncRNAs, and miRNAs using different strategies based on their diversified molecular functions. The expression of circCDYL, circATXN1, circDYM, circPRGRIP, lncRNA Meg3, and lncRNA Vax2os was validated by quantitative real-time PCR. These circRNAs and lncRNAs participate in neurotransmitter transport and multicellular organism growth through the intricate circRNA/lncRNA-miRNA-mRNA network. CONCLUSION: Whole-transcriptome sequencing and bioinformatics analysis systematically screened key ncRNAs in retinal neurogenesis. The validated ncRNAs and their circRNA/lncRNA-miRNA-mRNA network involve neurotransmitter transport and multicellular organism growth during retinal development.

MicroRNA-18a-5p Administration Suppresses Retinal Neovascularization by Targeting FGF1 and HIF1A.

Pathologic ocular neovascularization commonly results in visual impairment or even blindness in numerous fundus diseases, including proliferative diabetic retinopathy (PDR), retinopathy of prematurity (ROP), and age-related macular degeneration (AMD). MicroRNAs regulate angiogenesis through modulating target genes and disease progression, making them a new class of targets for drug discovery. In this study, we investigated the potential role of miR-18a-5p in retinal neovascularization using a mouse model of oxygen-induced proliferative retinopathy (OIR). We found that miR-18a-5p was highly expressed in the retina of pups as well as retinal endothelial cells, and was consistently down-regulated during retinal development. On the other hand, miR-18a-5p was increased significantly during pathologic neovascularization in the retinas of OIR mice. Moreover, intravitreal administration of miRNA mimic, agomiR-18a-5p, significantly suppressed retinal neovascularization in OIR models. Accordingly, agomir-18a-5p markedly suppressed human retinal microvascular endothelial cell (HRMEC) function including proliferation, migration, and tube formation ability. Additionally, we demonstrated that miR-18a-5p directly down-regulated known vascular growth factors, fibroblast growth factor 1 (FGF1) and hypoxia-inducible factor 1-alpha (HIF1A), as the target genes. In conclusion, miR-18a-5p may be a useful drug target for pathologic ocular neovascularization.

Circulating S100A8/A9 Levels Reflect Intraocular Inflammation in Uveitis Patients.

Purpose: To investigate whether there is an association between circulating S100A8/A9 levels and uveitis activity.Methods: A total of 549 plasma samples were collected from uveitis patients and non-uveitic controls.Results: S100A8/A9 plasma levels were elevated in uveitis patients compared to non-uveitic controls (P < 0.001). S100A8/A9 plasma levels in patients with active acute anterior uveitis (AAU) were significantly elevated and remarkably decreased in parallel with the severity of intraocular inflammation after corticosteroid treatment (P < 0.001). S100A8/A9 plasma levels were also higher in AAU patients with ankylosing spondylitis (AS) than in patients without AS (P = 0.02). S100A8/A9 plasma levels were significantly increased in uveitis patients with elevated C-reactive protein (CRP, P = 0.004) or erythrocyte sedimentation rates (ESR, P = 0.049) levels compared to uveitis patients with normal CRP or ESR values.Conclusion: Circulating S100A8/A9 might be a useful biomarker for the measurement of intraocular inflammation.

Elevated Plasma Levels of Drebrin in Glaucoma Patients With Neurodegeneration.

Glaucoma is an optic neuropathy characterized by progressive degeneration of retinal ganglion cells (RGCs). Aberrations in several cytoskeletal proteins, such as tau have been implicated in the pathogenesis of neurodegenerative diseases, could be initiating factors in glaucoma progression and occurring prior to axon degeneration. Developmentally regulated brain protein (Drebrin or DBN1) is an evolutionarily conserved actin-binding protein playing a prominent role in neurons and is implicated in neurodegenerative diseases. However, the relationship between circulating DBN1 levels and RGC degeneration in glaucoma patients remains unclear. In our preliminary study, we detected drebrin protein in the plasma of glaucoma patients using proteomic analysis. Subsequently, we recruited a total of 232 patients including primary angle-closure glaucoma (PACG), primary open-angle glaucoma (POAG) and Posner-Schlossman syndrome (PS) and measured its DBN1 plasma levels. We observed elevated DBN1 plasma levels in patients with primary glaucoma but not in patients with PS compared to nonaxonopathic controls. Interestingly, in contrast to tau plasma levels increased in all groups of patients, elevated drebrin plasma levels correlated with retinal nerve fiber layer defect (RNFLD) in glaucoma patients. To further explore the expression of DBN1 in neurodegeneration, we conducted experiment of optic nerve crush (ONC) models, and observed increased expression of DBN1 in the serum as well as in the retina and then decreased after ONC. This result reinforces the potentiality of circulating DBN1 levels are increased in glaucoma patients with neurodegeneration. Taken together, our findings suggest that circulating DBN1 levels correlated with RNFLD and may reflect the severity of RGCs injury in glaucoma patients. Combining measurement of circulating drebrin and tau levels may be a useful indicator for monitoring progression of neurodegenerative diseases.

PR1P ameliorates neurodegeneration through activation of VEGF signaling pathway and remodeling of the extracellular environment.

Neurodegenerative diseases affect millions of people worldwide. Optic neuropathies are the most commonly occurring neurodegenerative diseases, characterized by progressive retinal ganglion cell (RGC) degeneration. We recently reported that Prominin-1, a protein found on the surface of stem cells, interacts with VEGF and enhances its activity. VEGF is known to have various protective roles in the nervous system. Subsequently, we have developed a 12-mer peptide derived from Prominin-1, named PR1P, and investigated its effects on neuronal survival of damaged RGCs in a rat model of optic nerve crush (ONC). PR1P prevented RGC apoptosis resulting in improvement of retinal function in the rat ONC model. PR1P treatment significantly increased phosphorylation of ERK and AKT and expression its downstream proteins c-fos and Egr-1 in the retina. Additionally, PR1P beneficially increased the MMP-9/TIMP-1 ratio and promoted glial activation in the retina of ONC rats. Thus, PR1P displayed neuroprotective effects through enhanced VEGF-driven neuronal survival and reconstruction of the extracellular environment in ONC model. Our data indicate that PR1P may be a promising new clinical candidate for the treatment of neurodegenerative diseases.

A novel strategy to enhance angiogenesis in vivo using the small VEGF-binding peptide PR1P.

Therapeutic angiogenesis is an experimental frontier in vascular biology that seeks to deliver angiogenic growth factors to ischemic or injured tissues to promote targeted formation of new blood vessels as an alternative approach to surgical revascularization procedures. Vascular endothelial growth factor (VEGF) is a potent angiogenic signal protein that is locally upregulated at sites of tissue injury. However, therapies aimed at increasing VEGF levels experimentally by injecting VEGF gene or protein failed to improve outcomes in human trials in part due to its short half-life and systemic toxicity. We recently designed a novel 12-amino acid peptide (PR1P) whose sequence was derived from an extracellular VEGF-binding domain of the pro-angiogenic glycoprotein prominin-1. In this study, we characterized the molecular binding properties of this novel potential therapeutic for targeted angiogenesis and provided the foundation for its use as an angiogenic molecule that can potentiate endogenous VEGF. We showed that PR1P bound VEGF directly and enhanced VEGF binding to endothelial cells and to VEGF receptors VEGFR2 and neuropilin-1. PR1P increased angiogenesis in the murine corneal micropocket assay when combined with VEGF, but had no activity without added VEGF. In addition, PR1P also enhanced angiogenesis in murine choroidal neovascularization and wound-healing models and augmented reperfusion in a murine hind-limb ischemia model. Together our data suggest that PR1P enhanced angiogenesis by potentiating the activity of endogenous VEGF. In so doing, this novel therapy takes advantage of endogenous VEGF gradients generated in injured tissues and may improve the efficacy of and avoid systemic toxicity seen with previous VEGF therapies.

S100A8 promotes migration and infiltration of inflammatory cells in acute anterior uveitis.

Uveitis, the pathologic condition of inflammation of the uvea, frequently leads to severe vision loss and blindness. S100A8 is a calcium-binding protein which mainly expresses in granulocytes and monocytes and plays a prominent role in the regulation of inflammatory processes and immune response. Here, we determined the role of S100A8-positive cells in acute anterior uveitis (AAU) and keratitis. In rat models of endotoxin (lipopolisaccharide, LPS) -induced uveitis (EIU) and keratitis, S100A8-positive granulocytes and monocytes increased significantly in the iris-ciliary body and cornea as well as in the blood. Interestingly, Glucocorticoids slightly increased S100A8 levels in leukocytes, but reduced its presence significantly in the iris-ciliary body after LPS injection. Moreover, inhibition of NF-kB activation remarkably suppressed both progression of AAU and total S100A8 levels in leukocytes and the iris-ciliary body after LPS administration. Additionally, S100A8 protein level was also found to be elevated in the serum of AAU patients parallel with the progression of AAU through the designated clinical stages. Thus, S100A8 plays a pivotal role in the processes of AAU through involvement in migration and infiltration of S100A8-positive cells. Our findings suggest that serum levels of S100A8 protein can be used to monitor inflammatory activity in AAU.