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Abstract: Norrin is a secreted signaling molecule with structural and functional characteristics of an autocrine and/or paracrine acting growth factor. In the eye, Norrin is constitutively expressed in Müller cells. Norrin specifically binds to Frizzled-4 receptors and activates the canonical Wnt/β-catenin signaling pathway that is profoundly enhanced when Tspan12 is present at the Norrin/Frizzled-4 receptor complex. In the absence of Norrin or Frizzled-4, intraretinal capillaries are not formed during developmental angiogenesis. As a result there is considerable evidence that Norrin and Frizzled-4 are part of an essential signaling system that controls the formation of the retinal vasculature during eye development. Intriguingly, Norrin promotes vessel regrowth and induces the formation of intraretinal capillaries following oxygen-induced retinopathy in mice, an animal model of retinopathy of prematurity. Moreover, Norrin has pronounced neuroprotective properties on retinal ganglion cells (RGC) with the distinct potential to decrease the damaging effects of excitotoxic NMDA-induced RGC injury. The neuroprotective effects of Norrin similarly involve an activation of Wnt/β-catenin signaling and the subsequent induction of neuroprotective growth factor synthesis in Müller cells, such as that of fibroblast growth factor-2 (FGF2) or ciliary neurotrophic factor (CNTF). Overall, Norrin and the molecules involved in its signaling pathway appear to be promising targets to develop strategies that induce intraretinal vessel formation in patients suffering from ischemic retinopathies, or that increase RGC survival in glaucoma. [Copyright &y& Elsevier]

Mutations in the MYOC gene that encodes for myocilin are causative for some forms of juvenile and adult-onset primary open-angle glaucoma (POAG). Myocilin is a secreted 55–57 kDa glycoprotein that forms dimers and multimers. Characteristic structural motifs include a myosin-like domain, a leucine zipper region and an olfactomedin domain. Most of the mutations that have been identified in patients with POAG are localized in the olfactomedin domain, which is highly conserved among species. In the eye, myocilin is expressed in high amounts in the trabecular meshwork (TM), sclera, ciliary body and iris, and at considerable lower amounts in retina and optic nerve head. Secreted myocilin is present in the aqueous humor. In the TM, myocilin is found within the cytoplasm of TM cells and in the juxtacanalicular region in association with fibrillar extracellular matrix components. Since patients with mutations in myocilin may have high intraocular pressures, the role of myocilin for aqueous humor outflow has been investigated and conflicting results have been obtained. Recombinant myocilin increases outflow resistance in perfused anterior segment organ cultures, while overexpression of myocilin after viral gene transfer appears to reduce outflow resistance. In TM cells, the expression of myocilin is induced upon treatment with dexamethasone at a time course similar to that observed in steroid-induced glaucoma. Other factors that induce myocilin expression are transforming growth factor-β and mechanical stretch. Promoter elements that are important for the glucocorticoid induction have not been identified, but it has been shown that upstream stimulatory factor is critical for the basal promoter activity of MYOC. Mice with a targeted disruption of the myocilin gene do not express a phenotype, indicating that the glaucomatous phenotype in humans is not because of a loss-of-function effect. Experimental studies show that mutated myocilin is not secreted, but appears to accumulate in the cells. Such an accumulation might interfere with TM function and lead to impaired outflow resistance, but, so far, experimental evidence for such a scenario is lacking. In addition, the normal function(s) of myocilin is (are) still elusive. [Copyright &y& Elsevier]

• Most retinal diseases underly multifactorial pathomechanisms. • The retinal pigment epithelium (RPE) plays a key part in these pathomechanisms. • Targeted RPE therapy can be used to efficiently treat retinal diseases. • Intravitreal and systemic nanoparticle systems can reach the RPE. • Guidance for the development of RPE-specific nanotherapeutics is provided. The retinal pigment epithelium (RPE) plays a crucial part in sight-threatening diseases. In this review, we shed light on the pivotal implication of the RPE in age-related macular degeneration, diabetic retinopathy and retinopathy of prematurity; and explain why a paradigm shift toward targeted RPE therapy is needed to efficiently fight these retinal diseases. We provide guidance for the development of RPE-specific nanotherapeutics by giving a comprehensive overview of the possibilities and challenges of drug delivery to the RPE and highlight successful nanotherapeutic approaches targeting the RPE. [ABSTRACT FROM AUTHOR]

Abstract: Intraocular pressure, the most critical risk factor for primary open-angle glaucoma is generated in the trabecular meshwork outflow pathways, which provide resistance to aqueous humor outflow. The resistance is increased in primary open-angle glaucoma, and changes in the quality and amount of the extracellular matrix in the juxtacanalicular region of the trabecular meshwork appear to be causatively involved. The extracellular matrix changes are very likely under control of transforming growth factor-β2 (TGF-β2), which is found at high concentrations in the aqueous humor of patients with primary open-angle glaucoma. Additional factors are thrombospondin-1, which activates TGF-β2 in vivo, and connective tissue growth factor, which is an important downstream mediator of the effects of TGF-β2 on trabecular meshwork extracellular matrix turnover. In contrast, bone morphogenetic protein-7 (BMP-7) strongly antagonizes fibrogenic actions of TGF-β2 on human trabecular meshwork cells, indicating that a pharmacological modulation of BMP-7 signalling might be a promising strategy to treat primary open-angle glaucoma. [Copyright &y& Elsevier]

Intraocular pressure (IOP) is the critical risk factor for glaucoma, a neurodegenerative disease and frequent cause of blindness worldwide. As of today, all effective strategies to treat glaucoma aim at lowering IOP. IOP is generated and maintained via the aqueous humor circulation system in the anterior eye. Aqueous humor is secreted by the ciliary processes and exits the eye through the trabecular meshwork (TM) or the uveoscleral outflow pathways. The TM outflow pathways provide resistance to aqueous humor outflow and IOP builds up in response to it. In the normal eye, the resistance is localized in the inner wall region, which comprises the juxtacanalicular connective tissue (JCT) and the inner wall endothelium of Schlemm’s canal (SC). Outflow resistance in the inner wall region is lowered through the contraction of the ciliary muscle or the relaxation of contractile myofibroblasts in the posterior part of the TM and the adjacent scleral spur. Patients with primary open-angle glaucoma (POAG), the most frequent form of glaucoma, typically suffer from an abnormally high outflow resistance of the inner wall region. There is increasing evidence that the increase in TM outflow resistance in POAG is the result of a characteristic change in the biological properties of the resident cells in the JCT, which increasingly acquire the phenotype of contractile myofibroblasts. This scenario strengthens simultaneously both their actin cytoskeleton and their directly associated extracellular matrix fibrils, leads to overall stiffening of the tissue, and is modulated by transforming growth factor-β (TGF-β)/connective tissue growth factor (CTGF) signaling. Essentially comparable changes appear to occur in SC endothelial cells in glaucoma. Causative therapy concepts targeting the aqueous outflow pathways in glaucoma should aim at interfering with this process either by attenuating TM or SC stiffness, and/or by modulating TGF-β/CTGF signaling. [ABSTRACT FROM AUTHOR]

• Decorin deficiency leads to elevated intraocular pressure in mice. • TGF-β target genes are enriched in the conventional outflow pathways of decorin-deficient mice. • Decorin attenuates the expression of TGF-β2 in cultured HTM cells. • Decorin levels are reduced in the chamber angle of glaucomatous eyes. Primary open-angle glaucoma, a neurodegenerative disorder characterized by degeneration of optic nerve axons, is a frequent cause of vision loss and blindness worldwide. Several randomized multicenter studies have identified intraocular pressure as the major risk factor for its development, caused by an increased outflow resistance to the aqueous humor within the trabecular meshwork. However, the molecular mechanism for increased outflow resistance in POAG has not been fully established. One of the proposed players is the pro-fibrotic transforming growth factor (TGF)-β2, which is found in higher amounts in the aqueous humor of patients with POAG. In this study we elucidated the role of decorin, a small leucine-rich proteoglycan and known antagonist of TGF-β, in the region of aqueous humor outflow tissue. Utilizing decorin deficient mice, we discovered that decorin modulated TGF-β signaling in the canonical outflow pathways and the lack of decorin in vivo caused an increase in intraocular pressure. Additionally, the Dcn −/− mice showed significant loss of optic nerve axons and morphological changes in the glial lamina, typical features of glaucoma. Moreover, using human trabecular meshwork cells we discovered that soluble decorin attenuated TGF-β2 mediated synthesis and expression of typical downstream target genes including CCN2/CTGF, FN and COL IV. Finally, we found a negative reciprocal regulation of decorin and TGF-β, with a dramatic downregulation of decorin in the canonical outflow pathways of patients with primary open-angle glaucoma. Collectively, our results indicate that decorin plays an important role in the pathogenesis of primary open-angle glaucoma and offers novel perspectives in the treatment of this serious disease. [ABSTRACT FROM AUTHOR]

The angiotensin II receptor type 1 (AT 1 R), which is expressed in blood vessels of the posterior eye, is of paramount significance in the pathogenesis of severe ocular diseases such as diabetic retinopathy and age-related macular degeneration. However, small molecule angiotensin receptor blockers (ARBs) have not proven to be a significant therapeutic success. We report here on a nanoparticle system consisting of ARB molecules presented in a multivalent fashion on the surface of quantum dots (Qdots). As a result of the multivalent receptor binding, nanoparticles targeted cells with high AT 1 R expression and inhibited their angiotensin receptor signaling with an IC 50 of 3.8 nM while showing only minor association to cells with low AT 1 R expression. After intravenous injection into the tail vein of mice, multivalent nanoparticles accumulated in retinal and choroidal blood vessels of the posterior eye. At the same time, multivalent ligand display doubled the Qdot concentration in the blood vessels compared to non-targeted Qdots. Remarkably, ARB-targeted Qdots showed no pronounced accumulation in AT 1 R-expressing off-target tissues such as the kidney. Following systemic application, this multivalent targeting approach has the potential to amplify AT 1 R blockade in the eye and concomitantly deliver a therapeutic payload into ocular lesions. [ABSTRACT FROM AUTHOR]

Summary: Schwann cells produce myelin sheaths and thereby permit rapid saltatory conductance in the vertebrate peripheral nervous system. Their stepwise differentiation from neural crest cells is driven by a defined set of transcription factors. How this is linked to chromatin changes is not well understood. Here we show that the glial transcription factor Sox10 functions in Schwann cells by recruiting Brg1-containing chromatin-remodeling complexes via Baf60a to regulatory regions of Oct6 and Krox20 genes. It thereby stimulates expression of these transcriptional regulators that then cooperate with Sox10 to convert immature into myelinating Schwann cells. The functional interaction between Sox10 and Brg1 is evident from gain- and loss-of-function studies, similar neuropathies in the corresponding mouse mutants, and an aggravated neuropathy in compound mutants. Our results demonstrate that the transcription factor-mediated recruitment of the chromatin-remodeling machinery to specific genomic loci is an essential driving force for Schwann cell differentiation and myelination. [ABSTRACT FROM AUTHOR]

Myocilin is expressed in the glomerulus of the kidney and induced in mesangioproliferative glomerulonephritis.Background.Myocilin is a 55 to 57 kD secreted glycoprotein and member of the olfactomedin protein family. It is expressed in high amounts in the outflow tissues of the aqueous humor in the eye where it is supposed to contribute to outflow resistance. Myocilin is mutated in some forms of primary open angle glaucoma and affected patients show very high intraocular pressures because of an increase in resistance to aqueous humor outflow. To obtain information, if myocilin may play a comparable role in other tissues with transendothelial fluid flow, we investigated its expression in the rat kidney.Methods.The expression of myocilin in the normal rat kidney and its changes during mesangioproliferative glomerulonephritis were investigated by immunohistochemistry, one- and two-dimensional gel electrophoresis with Western blotting, and reverse transcription-polymerase chain reaction (RT-PCR).Results.Myocilin and its mRNA were detected in isolated glomeruli. Immunohistochemistry showed specific labeling of glomerular cells, while tubular and interstitial regions were essentially negative. Double staining with the podocyte-specific markers synaptopodin and ezrin indicated that myocilin-positive cells were predominately podocytes. During mesangioproliferative glomerulonephritis, an induction of myocilin immunoreactivity was observed. Labeling for myocilin was now observed in activated mesangial cells and areas of glomerular sclerosis. In parallel cell culture experiments, mRNA for myocilin was detected in cultured murine podocytes and rat mesangial cells.Conclusion.Myocilin is expressed in podocytes of the kidney and induced in mesangial cells during experimental mesangioproliferative glomerulonephritis. The specific function of myocilin in the kidney is not clear, but in a parallel to functions of other olfactomedin proteins, it might have a role in cell-cell adhesion and/or signaling processes. [ABSTRACT FROM AUTHOR]

The development of the chamber angle was studied in the eyes of heterozygous Pax6lacZ/+ mutant mice (Nature 387 (1997) 406). Mutations in PAX6 cause aniridia, a condition that is frequently associated with glaucoma, a blinding disease that may be associated with chamber angle defects. Mesenchymal cells were seen in the chamber angle at P1–P5. In wild-type mice, these cells differentiated into typical trabecular meshwork (TM) cells next to Schlemm''s canal. In Pax6lacZ/+ mice, TM cells remained undifferentiated and Schlemm''s canal was absent. From P1 to P4, staining for β-galactosidase and immunoreactivity for Pax6 were observed in chamber angle mesenchyme, but were absent later. Cultured murine TM cells expressed Pax6. The defects in chamber angle and TM differentiation were associated with a wide spectrum of other anterior eye defects, which included various degrees of iris hypoplasia and corneal haze, isolated iridocorneal adhesions and atypical coloboma, and a vascularized cornea in all adult animals. A third of the animals showed Peters'' anomaly including corneal opacity and iridocorneal adhesions. The separation of the lens from the cornea was incomplete, and epithelial layers of lens and cornea were continuous. Pax6 activity is directly required for differentiation of the chamber angle. Variations in phenotype of Pax6lacZ/+ mice appear not to involve direct dominant-negative or dose-dependent effects. [Copyright &y& Elsevier]

Lipid nanocapsules are treasured nanoparticulate systems, although they lack detectability in biological environments. To overcome this, we designed LNCs loaded simultaneously with fluorescent dye and superparamagnetic iron oxide nanoparticles (Dual LNCs). The introduction of both labels did not alter nanoparticle characteristics such as size (50 nm), size distribution (polydispersity index < 0.1) or surface modifications, including the effectiveness of targeting ligands. Furthermore, the colloidal stability, particle integrity and biocompatibility of the nanoparticles were not negatively affected by label incorporation. These Dual LNCs are concomitantly visualizable via fluorescence and transmitted light imaging after either the internalization by cells or systemic administration to mice. Importantly, they are detectable in liver sections of mice using transmission electron microscopy without additional enhancement. The iron content of 0.24% (m/m) is sufficiently high for precise quantification of nanoparticle concentrations via inductively coupled plasma optical emission spectroscopy. Dual LNCs are precious tools for the investigation of in vitro and in vivo performances of lipid nanocapsule formulations, since they allow for the use of complementary imaging methods for broad range detectability. [ABSTRACT FROM AUTHOR]