Search

Diabetic retinopathy, a major cause of blindness, is characterized by a distinct phenotype. The molecular causes of the phenotype are not sufficiently clear. Here, we report that deletion of transforming growth factor β signaling in the retinal microenvironment of newborn mice induces changes that largely mimic the phenotype of nonproliferative and proliferative diabetic retinopathy in humans. Lack of transforming growth factor β signaling leads to the formation of abundant microaneurysms, leaky capillaries, and retinal hemorrhages. Retinal capillaries are not covered by differentiated pericytes, but by a coat of vascular smooth muscle-like cells and a thickened basal lamina. Reactive microglia is found in close association with retinal capillaries. In older animals, loss of endothelial cells and the formation of ghost vessels are observed, findings that correlate with the induction of angiogenic molecules and the accumulation of retinal hypoxia-inducible factor 1α, indicating hypoxia. Consequently, retinal and vitreal neovascularization occurs, a scenario that leads to retinal detachment, vitreal hemorrhages, neuronal apoptosis, and impairment of sensory function. We conclude that transforming growth factor β signaling is required for the differentiation of retinal pericytes during vascular development of the retina. Lack of differentiated pericytes initiates a scenario of structural and functional changes in the retina that mimics those of diabetic retinopathy strongly indicating a common mechanism., (Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Similar to other parts of the central nervous system, there are two types of programmed cell death during retinal development. In early development, the neuronal progenitor population is affected. In the mouse eye, this kind of programmed cell death begins at around embryonic day (E) 12.5 and peaks between E14.5 and E16.5. The second phase of programmed cell death occurs during synaptogenesis within the first 2 postnatal weeks. Important signaling mechanisms that induce programmed cell death of retinal progenitors appear to involve nerve growth factor acting on the proapoptotic receptor to p75 neurotrophin receptor (p75(NTR)) and transforming growth factor-β.

Glucocorticoids (GCs) have a wide range of functions on several mammalian cell types, most of which are aimed at boosting survival, which is the raison d'être of the acute stress response. The role GCs play in the survival and viability of neurons is incongruous, as studies have revealed neuroprotective as well as neurodegenerative effects. These effects seem to depend on multiple factors amongst which are; the cell type involved, the mode of injury or underlying cause of cell death, likewise the concentration and or duration of GC exposure.In this mini review, we discuss mechanisms of GC action and their effect on neurodegeneration in general, and specifically review the effect of GCs on retinal neurons, in animal models of retinal degeneration or acute neuronal damage. Finally, we summarize potential protective and harmful GC-mediated mechanisms, which might be involved in the determination of neuronal fate in the retina following injury or during degeneration.

We investigated the influence of transforming growth factor-β (TGF-β) signaling on developmental programmed cell death in the mouse retina by direct and specific molecular targeting of TGF-β type II receptor (TβRII) and Smad7 in retinal progenitor cells. Mice were generated carrying a conditional deletion of the TβRII in cells that originate from the inner layer of the optic cup. The animals showed a significant decrease of phosphorylated Smad3 in both the central and peripheral retina, which indicates the diminished activity of TGF-β signaling. TβRII deficiency significantly increased the apoptotic death of retinal neurons during embryonic and postnatal development without affecting their proliferation. In contrast, treatment with TGF-β2 inhibited cell death of retinal ganglion cells in dissociated retinal cell cultures, an effect that was blocked by inhibiting the phosphorylation of Smad3. The increase in apoptosis during development resulted in a significant reduction in the number of neurons in adult TβRII-deficient mice. The effect was most pronounced in the inner retina neurons and resulted in functional deficits as determined by electroretinography. In contrast, a conditional deletion of TGF-β-inhibiting Smad7 in retinal neurons significantly enhanced Smad3 phosphorylation and significantly decreased apoptosis of retinal neurons in embryos and pups. Moreover, the number of retinal ganglion cells was significantly higher in Smad7-deficient mice compared with control littermates. TβRII-deficient pups showed a lower level of nerve growth factor (NGF) in its mRNA; however, higher levels were observed in Smad7-deficient pups, which strongly suggests that the protective effects of TGF-β signaling on developmental cell death are mediated through NGF.

Optineurin is a Golgi complex-associated ubiquitous protein with high expression levels in retinal ganglion cells (RGCs). Mutations in optineurin have been observed in rare hereditary cases of primary open-angle glaucoma and in amyotrophic lateral sclerosis. We explored the possibility that optineurin deficiency will compromise neuronal exocytosis leading to a diminished secretion of neurotrophic factors that are critically required for neuronal survival. To this end, we used RNA interference to induce depletion of optineurin in RGC-5 cells derived from retinal neurons. SiRNA specific for optineurin was transiently transfected. Moreover, a stable cell line with constitutive optineurin deficiency (RGC-5 pSilencer OPTN) was generated. In addition, we investigated the subcellular localization of optineurin in primary RGCs in retinal cell cultures isolated from eyes of mature mice. In RGC-5 cells, optineurin localized to the periphery of the Golgi complex and was observed in vesicular structures throughout the cytoplasm and close to the plasma membrane. A comparable Golgi-associated localization of optineurin was observed in cultured primary RGCs that were identified by TUJ1 labeling. Optineurin deficiency caused a marked increase in the number of RGC-5 cells with fragmented Golgi complex. RGC-5 pSilencer OPTN with stable optineurin deficiency showed a pronounced increase in the number of cells undergoing apoptotic cell death. Furthermore, the amounts of secreted neurotrophin-3 (NT-3) and ciliary neurotrophic factor were significantly lower in culture medium of RGC-5 pSilencer OPTN cells when compared to controls. Adding exogenous NT-3 to the culture medium to achieve amounts seen in control cultures completely prevented the increase in apoptotic cell death. We propose that lack of neurotrophic support due to impaired secretion of neurotrophic proteins is a critical factor that causes or contributes to RGC or motor neuron death in patients with mutated optineurin., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Purpose: To investigate whether and how leukemia inhibitory factor (Lif) is involved in mediating the neuroprotective effects of Norrin on retinal ganglion cells (RGC) following excitotoxic damage. Norrin is a secreted protein that protects RGC from N-methyl-d-aspartate (NMDA)-mediated excitotoxic damage, which is accompanied by increased expression of protective factors such as Lif, Edn2 and Fgf2. Methods: Lif-deficient mice were injected with NMDA in one eye and NMDA plus Norrin into the other eye. RGC damage was investigated and quantified by TUNEL labeling 24 h after injection. Retinal mRNA expression was analyzed by quantitative real-time polymerase chain reaction following retinal treatment. Results: After intravitreal injection of NMDA and Norrin in wild-type mice approximately 50% less TUNEL positive cells were observed in the RGC layer when compared to NMDA-treated littermates, an effect which was lost in Lif-deficient mice. The mRNA expression for Gfap, a marker for Müller cell gliosis, as well as Edn2 and Fgf2 was induced in wild-type mice following NMDA/Norrin treatment but substantially blocked in Lif-deficient mice. Conclusions: Norrin mediates its protective properties on RGC via Lif, which is required to enhance Müller cell gliosis and to induce protective factors such as Edn2 or Fgf2. [ABSTRACT FROM AUTHOR]

Abstract: Apoptotic death of photoreceptors in hereditary retinal degenerations can be prevented by neuroprotective molecules. Here, we report that adrenal glucocorticoids (GC) released during psychosocial stress protect photoreceptors from apoptosis after light damage. Psychosocial stress is known to be the main type of stressor humans are exposed to and was induced here in mice by 10h of chronic subordinate colony housing (CSC). Photoreceptor damage was generated by subsequent exposure to white light. Short-term psychosocial stress prior to illumination significantly reduced the number of apoptotic photoreceptors, an effect that was absent in adrenalectomized (ADX) mice. The neuroprotective effect was completely restored in ADX mice substituted with GC. Moreover, phosphorylation of retinal AKT increased following CSC or exogenous GC treatment, an effect that was again absent in ADX mice exposed to CSC. Finally, inhibition of AKT signaling with triciribine blocked the stress- and GC-mediated neuroprotective effects on photoreceptors. In summary, we provide evidence that 1) short-term psychosocial stress protects photoreceptors from light-induced damage and 2) the protective effect is most likely mediated by GC-induced activation of the AKT signaling pathway. [Copyright &y& Elsevier]

Mutations in the myocilin gene (MYOC) are causative for 10% of cases with juvenile open-angle glaucoma and 3-4% of those with primary open-angle glaucoma. Myocilin is a secreted protein with relatively ill-defined matricellular properties. Despite its high expression in the eye, myocilin-deficient mice have originally been reported to have no obvious ocular phenotype. Here we revisited the ocular phenotype of myocilin-deficient mice and detected a higher number of neurons in their inner (INL) and outer (ONL) nuclear layers, as well as a higher number of retinal ganglion cells (RGC) and their axons. The increase in retinal neurons appears to be caused by a decrease in programmed developmental cell death, as apoptosis of retinal neurons between postnatal days 4 and 10 was found to be attenuated when compared to that of wildtype littermates. In contrast, when Myoc-/- mice were crossed with βB1-crystallin-MYOC mice with ectopic overexpression of myocilin in the eye, no differences in developmental apoptosis, RGC number and INL thickness were observed when compared to wildtype littermates. The amounts of the anti-apoptotic Bcl-2-like protein 1 (BCL2L1, Bcl-xL) and its mRNA were increased in retinae of Myoc-/- mice, while lower amounts of BCL2L1 and its mRNA were detected in mixed Myoc-/-/βB1-crystallin-MYOC mice. The structural differences between Myoc-/- mice and wildtype littermates did not result in functional differences as measured by electroretinography. Noteworthy though mixed Myoc-/-/βB1-crystallin-MYOC mice with ocular overexpression of myocilin had significant cone function deficits. Myocilin appears to modulate apoptotic death of retinal neurons likely by interacting with the intrinsic apoptotic pathway. [ABSTRACT FROM AUTHOR]

Abstract: Mutations in the OPTN gene encoding for optineurin have been associated with primary open-angle glaucoma. The functional role(s) of optineurin in the normal and glaucomatous eye are unclear. As optineurin interferes with TNF-α mediated cell death in vitro, an involvement of optineurin in the regulatory pathways leading to apoptosis of retinal ganglion cells has been suggested. The goal of the present study was to study the molecular properties of optineurin and its capabilities to prevent apoptosis in vivo in the eyes of transgenic mice. The chicken βB1-crystallin promoter was used to overexpress ectopic optineurin in the lenses of transgenic mice. The expression of transgenic mRNA was monitored by northern blot analysis. The localization of transgenic optineurin was investigated by one- and two-dimensional western blot analysis and by immunohistochemistry, and compared with that of endogenous optineurin. To assess effects of transgenic optineurin on apoptosis, βB1-crystallin-OPTN mice were crossbred with βB1-crystallin-TGFβ1 mice that undergo substantial TGF-β1-induced apoptotic cell death in the lens. Two independent βB1-crystallin-OPTN transgenic lines were established, in which transgenic optineurin was expressed strictly lens-specific as assessed by Northern and Western blotting, and by immunohistochemistry. In contrast, endogenous optineurin was preferentially expressed in the retina, where retinal ganglion cells showed strong labeling. Immunostaining for endogenous optineurin in the anterior eye was considerably weaker than in the posterior eye and was seen in iris, ciliary epithelium, cells of corneal stroma and endothelium, and in the trabecular meshwork. Neither transgenic nor endogenous optineurin was found in the aqueous humor. Transgenic overexpression of optineurin did not have measurable effects on TGFβ1-induced apoptosis in mixed βB1-crystallin-OPTN/βB1-crystallin-TGFβ1 transgenic mice. Our results show that optineurin is a cytoplasmatic rather than a secretory protein that is preferentially expressed in retinal ganglion cells, and argue against a major role of optineurin for the modulation of apoptosis in vivo. [Copyright &y& Elsevier]