Loss of DJ-1 does not affect mitochondrial respiration but increases ROS production and mitochondrial permeability transition pore opening

Background

Loss of function mutations in the DJ-1 gene have been linked to recessively inherited forms of Parkinsonism. Mitochondrial dysfunction and increased oxidative stress are thought to be key events in the pathogenesis of Parkinson’s disease. Although it has been reported that DJ-1 serves as scavenger for reactive oxidative species (ROS) by oxidation on its cysteine residues, how loss of DJ-1 affects mitochondrial function is less clear.

Methodology/Principal Findings

Using primary mouse embryonic fibroblasts (MEFs) or brains from DJ-1−/− mice, we found that loss of DJ-1 does not affect mitochondrial respiration. Specifically, endogenous respiratory activity as well as basal and maximal respiration are normal in intact DJ-1−/− MEFs, and substrate-specific state 3 and state 4 mitochondrial respiration are also unaffected in permeabilized DJ-1−/− MEFs and in isolated mitochondria from the cerebral cortex of DJ-1−/− mice at 3 months or 2 years of age. Expression levels and activities of all individual complexes composing the electron transport system are unchanged, but ATP production is reduced in DJ-1−/− MEFs. Mitochondrial transmembrane potential is decreased in the absence of DJ-1. Furthermore, mitochondrial permeability transition pore opening is increased, whereas mitochondrial calcium levels are unchanged in DJ-1−/− cells. Consistent with earlier reports, production of reactive oxygen species (ROS) is increased, though levels of antioxidative enzymes are unaltered. Interestingly, the decreased mitochondrial transmembrane potential and the increased mitochondrial permeability transition pore opening in DJ-1−/− MEFs can be restored by antioxidant treatment, whereas oxidative stress inducers have the opposite effects on mitochondrial transmembrane potential and mitochondrial permeability transition pore opening.

Conclusions/Significance

Our study shows that loss of DJ-1 does not affect mitochondrial respiration or mitochondrial calcium levels but increases ROS production, leading to elevated mitochondrial permeability transition pore opening and reduced mitochondrial transmembrane potential.     

Cigarette smoke-related hydroquinone dysregulates MCP-1, VEGF and PEDF expression in retinal pigment epithelium in vitro and in vivo

Background

Age-related macular degeneration (AMD) is the leading cause of legal blindness in the elderly population. Debris (termed drusen) below the retinal pigment epithelium (RPE) have been recognized as a risk factor for dry AMD and its progression to wet AMD, which is characterized by choroidal neovascularization (CNV). The underlying mechanism of how drusen might elicit CNV remains undefined. Cigarette smoking, oxidative damage to the RPE and inflammation are postulated to be involved in the pathophysiology of the disease. To better understand the cellular mechanism(s) linking oxidative stress and inflammation to AMD, we examined the expression of pro-inflammatory monocyte chemoattractant protein-1 (MCP-1), pro-angiogenic vascular endothelial growth factor (VEGF) and anti-angiogenic pigment epithelial derived factor (PEDF) in RPE from smoker patients with AMD. We also evaluated the effects of hydroquinone (HQ), a major pro-oxidant in cigarette smoke on MCP-1, VEGF and PEDF expression in cultured ARPE-19 cells and RPE/choroids from C57BL/6 mice.

Principal Findings

MCP-1, VEGF and PEDF expression was examined by real-time PCR, Western blot, and ELISA. Low levels of MCP-1 protein were detected in RPE from AMD smoker patients relative to controls. Both MCP-1 mRNA and protein were downregulated in ARPE-19 cells and RPE/choroids from C57BL/6 mice after 5 days and 3 weeks of exposure to HQ-induced oxidative injury. VEGF protein expression was increased and PEDF protein expression was decreased in RPE from smoker patients with AMD versus controls resulting in increased VEGF/PEDF ratio. Treatment with HQ for 5 days and 3 weeks increased the VEGF/PEDF ratio in vitro and in vivo.

Conclusion

We propose that impaired RPE-derived MCP-1-mediated scavenging macrophages recruitment and phagocytosis might lead to incomplete clearance of proinflammatory debris and infiltration of proangiogenic macrophages which along with increased VEGF/PEDF ratio favoring angiogenesis might promote drusen accumulation and progression to CNV in smoker patients with dry AMD.     

Comparison of Mouse and Human Retinal Pigment Epithelium Gene Expression Profiles: Potential Implications for Age-Related Macular Degeneration

Background

The human retinal pigment epithelium (RPE) plays an important role in the pathogenesis of age related macular degeneration (AMD). AMD is the leading cause of blindness worldwide. There is currently no effective treatment available. Preclinical studies in AMD mouse models are essential to develop new therapeutics. This requires further in-depth knowledge of the similarities and differences between mouse and human RPE.

Methods

We performed a microarray study to identify and functionally annotate RPE specific gene expression in mouse and human RPE. We used a meticulous method to determine C57BL/6J mouse RPE signature genes, correcting for possible RNA contamination from its adjacent layers: the choroid and the photoreceptors. We compared the signature genes, gene expression profiles and functional annotations of the mouse and human RPE.

Results

We defined sets of mouse (64), human (171) and mouse–human interspecies (22) RPE signature genes. Not unexpectedly, our gene expression analysis and comparative functional annotation suggested that, in general, the mouse and human RPE are very similar. For example, we found similarities for general features, like “organ development” and “disorders related to neurological tissue”. However, detailed analysis of the molecular pathways and networks associated with RPE functions, suggested also multiple species-specific differences, some of which may be relevant for the development of AMD. For example, CFHR1, most likely the main complement regulator in AMD pathogenesis was highly expressed in human RPE, but almost absent in mouse RPE. Furthermore, functions assigned to mouse and human RPE expression profiles indicate (patho-) biological differences related to AMD, such as oxidative stress, Bruch’s membrane, immune-regulation and outer blood retina barrier.

Conclusion

These differences may be important for the development of new therapeutic strategies and translational studies in age-related macular degeneration.     

Microglia in the Mouse Retina Alter the Structure and Function of Retinal Pigmented Epithelial Cells: A Potential Cellular Interaction Relevant to AMD

Background

Age-related macular degeneration (AMD) is a leading cause of legal blindness in the elderly in the industrialized word. While the immune system in the retina is likely to be important in AMD pathogenesis, the cell biology underlying the disease is incompletely understood. Clinical and basic science studies have implicated alterations in the retinal pigment epithelium (RPE) layer as a locus of early change. Also, retinal microglia, the resident immune cells of the retina, have been observed to translocate from their normal position in the inner retina to accumulate in the subretinal space close to the RPE layer in AMD eyes and in animal models of AMD.

Methodology/Principal Findings

In this study, we examined the effects of retinal microglia on RPE cells using 1) an in vitro model where activated retinal microglia are co-cultured with primary RPE cells, and 2) an in vivo mouse model where retinal microglia are transplanted into the subretinal space. We found that retinal microglia induced in RPE cells 1) changes in RPE structure and distribution, 2) increased expression and secretion of pro-inflammatory, chemotactic, and pro-angiogenic molecules, and 3) increased extent of in vivo choroidal neovascularization in the subretinal space.

Conclusions/Significance

These findings share similarities with important pathological features found in AMD and suggest the relevance of microglia-RPE interactions in AMD pathogenesis. We speculate that the migration of retinal microglia into the subretinal space in early stages of the disease induces significant changes in RPE cells that perpetuate further microglial accumulation, increase inflammation in the outer retina, and fosters an environment conducive for the formation of neovascular changes responsible for much of vision loss in advanced AMD.     

Reduced Basal Autophagy and Impaired Mitochondrial Dynamics Due to Loss of Parkinson's Disease-Associated Protein DJ-1

Background

Mitochondrial dysfunction and degradation takes a central role in current paradigms of neurodegeneration in Parkinson''s disease (PD). Loss of DJ-1 function is a rare cause of familial PD. Although a critical role of DJ-1 in oxidative stress response and mitochondrial function has been recognized, the effects on mitochondrial dynamics and downstream consequences remain to be determined.

Methodology/Principal Findings

Using DJ-1 loss of function cellular models from knockout (KO) mice and human carriers of the E64D mutation in the DJ-1 gene we define a novel role of DJ-1 in the integrity of both cellular organelles, mitochondria and lysosomes. We show that loss of DJ-1 caused impaired mitochondrial respiration, increased intramitochondrial reactive oxygen species, reduced mitochondrial membrane potential and characteristic alterations of mitochondrial shape as shown by quantitative morphology. Importantly, ultrastructural imaging and subsequent detailed lysosomal activity analyses revealed reduced basal autophagic degradation and the accumulation of defective mitochondria in DJ-1 KO cells, that was linked with decreased levels of phospho-activated ERK2.

Conclusions/Significance

We show that loss of DJ-1 leads to impaired autophagy and accumulation of dysfunctional mitochondria that under physiological conditions would be compensated via lysosomal clearance. Our study provides evidence for a critical role of DJ-1 in mitochondrial homeostasis by connecting basal autophagy and mitochondrial integrity in Parkinson''s disease.     

A Pilot Study of Morphometric Analysis of Choroidal Vasculature In Vivo,Using En Face Optical Coherence Tomography

Purpose

To study the ability of volumetric spectral domain optical coherence tomography (SD-OCT) to perform quantitative measurement of the choroidal vasculature in vivo.

Methods

Choroidal vascular density and vessel size were quantified using en face choroidal scans from various depths below the retinal pigment epithelium (RPE) in 58 eyes of 58 patients with either epiretinal membranes (ERM), early age-related macular degeneration (AMD), or reticular pseudo-drusen (RPD). For each patient, we used the macular volume scan (6×6 mm cube) for vessel quantification, while high-definition (HD) cross-section raster scans were used to qualitatively assess vascularity of the choroidal sub-layers, and measure choroidal thickness.

Results

Of the 58 patients, more were female (66% versus 34% male), of whom 14 (24%) had ERM, 11 (19%) early AMD, and 33 (57%) RPD. Compared to intact choriocapillaris in all ERM (100%), none of the RPD and only 5/11 (45%) early AMD eyes had visible choriocapillaris on either cross section or C-scans (p-value<0.001). When comparing select regions from the most superficial C-scans, early AMD group had lowest vascular density and RPD had highest (p-value 0.04). Qualitative evaluation of C-scans from all three groups revealed a more granular appearance of the choriocapillaris in ERM versus increased stroma and larger vessels in the RPD eyes.

Conclusions

SD-OCT can be used to qualitatively and quantitatively assess choroidal vascularity in vivo. Our findings correlate to previously reported histopathologic studies. Lack of choriocapillaris on HD cross-sections or C-scans in all RPD and about half of early AMD eyes suggests earlier choroidal involvement in AMD and specifically, RPD.     

DJ-1 Modulates α-Synuclein Aggregation State in a Cellular Model of Oxidative Stress: Relevance for Parkinson's Disease and Involvement of HSP70

Background

Parkinson''s disease (PD) is a neurodegenerative pathology whose molecular etiopathogenesis is not known. Novel contributions have come from familial forms of PD caused by alterations in genes with apparently unrelated physiological functions. The gene coding for alpha-synuclein (α-syn) (PARK1) has been investigated as α-syn is located in Lewy bodies (LB), intraneuronal inclusions in the substantia nigra (SN) of PD patients. A-syn has neuroprotective chaperone-like and antioxidant functions and is involved in dopamine storage and release. DJ-1 (PARK7), another family-PD-linked gene causing an autosomal recessive form of the pathology, shows antioxidant and chaperone-like activities too.

Methodology/Principal Findings

The present study addressed the question whether α-syn and DJ-1 interact functionally, with a view to finding some mechanism linking DJ-1 inactivation and α-syn aggregation and toxicity. We developed an in vitro model of α-syn toxicity in the human neuroblastoma cell line SK-N-BE, influencing DJ-1 and α-syn intracellular concentrations by exogenous addition of the fusion proteins TAT-α-syn and TAT-DJ-1; DJ-1 was inactivated by the siRNA method. On a micromolar scale TAT-α-syn aggregated and triggered neurotoxicity, while on the nanomolar scale it was neuroprotective against oxidative stress (induced by H₂O₂ or 6-hydroxydopamine). TAT-DJ-1 increased the expression of HSP70, while DJ-1 silencing made SK-N-BE cells more susceptible to oxidative challenge, rendering TAT-α-syn neurotoxic at nanomolar scale, with the appearance of TAT-α-syn aggregates.

Conclusion/Significance

DJ-1 inactivation may thus promote α-syn aggregation and the related toxicity, and in this model HSP70 is involved in the antioxidant response and in the regulation of α-syn fibril formation.     

Alternative Complement Pathway Deficiency Ameliorates Chronic Smoke-Induced Functional and Morphological Ocular Injury

Background

Age-related macular degeneration (AMD), a complex disease involving genetic variants and environmental insults, is among the leading causes of blindness in Western populations. Genetic and histologic evidence implicate the complement system in AMD pathogenesis; and smoking is the major environmental risk factor associated with increased disease risk. Although previous studies have demonstrated that cigarette smoke exposure (CE) causes retinal pigment epithelium (RPE) defects in mice, and smoking leads to complement activation in patients, it is unknown whether complement activation is causative in the development of CE pathology; and if so, which complement pathway is required.

Methods

Mice were exposed to cigarette smoke or clean, filtered air for 6 months. The effects of CE were analyzed in wildtype (WT) mice or mice without a functional complement alternative pathway (AP; CFB^−/−) using molecular, histological, electrophysiological, and behavioral outcomes.

Results

CE in WT mice exhibited a significant reduction in function of both rods and cones as determined by electroretinography and contrast sensitivity measurements, concomitant with a thinning of the nuclear layers as measured by SD-OCT imaging and histology. Gene expression analyses suggested that alterations in both photoreceptors and RPE/choroid might contribute to the observed loss of function, and visualization of complement C3d deposition implies the RPE/Bruch''s membrane (BrM) complex as the target of AP activity. RPE/BrM alterations include an increase in mitochondrial size concomitant with an apical shift in mitochondrial distribution within the RPE and a thickening of BrM. CFB^−/− mice were protected from developing these CE-mediated alterations.

Conclusions

Taken together, these findings provide clear evidence that ocular pathology generated in CE mice is dependent on complement activation and requires the AP. Identifying animal models with RPE/BrM damage and verifying which aspects of pathology are dependent upon complement activation is essential for developing novel complement-based treatment approaches for the treatment of AMD.     

Zinc deficiency leads to lipofuscin accumulation in the retinal pigment epithelium of pigmented rats

Background

Age-related macular degeneration (AMD) is associated with lipofuscin accumulation whereas the content of melanosomes decreases. Melanosomes are the main storage of zinc in the pigmented tissues. Since the elderly population, as the most affected group for AMD, is prone to zinc deficit, we investigated the chemical and ultrastructural effects of zinc deficiency in pigmented rat eyes after a six-month zinc penury diet.

Methodology/Principal Findings

Adult Long Evans (LE) rats were investigated. The control animals were fed with a normal alimentation whereas the zinc-deficiency rats (ZD-LE) were fed with a zinc deficient diet for six months. Quantitative Energy Dispersive X-ray (EDX) microanalysis yielded the zinc mole fractions of melanosomes in the retinal pigment epithelium (RPE). The lateral resolution of the analysis was 100 nm. The zinc mole fractions of melanosomes were significantly smaller in the RPE of ZD-LE rats as compared to the LE control rats. Light, fluorescence and electron microscopy, as well as immunohistochemistry were performed. The numbers of lipofuscin granules in the RPE and of infiltrated cells (Ø>3 µm) found in the choroid were quantified. The number of lipofuscin granules significantly increased in ZD-LE as compared to control rats. Infiltrated cells bigger than 3 µm were only detected in the choroid of ZD-LE animals. Moreover, the thickness of the Bruch''s membrane of ZD-LE rats varied between 0.4–3 µm and thin, rangy ED1 positive macrophages were found attached at these sites of Bruch''s membrane or even inside it.

Conclusions/Significance

In pigmented rats, zinc deficiency yielded an accumulation of lipofuscin in the RPE and of large pigmented macrophages in the choroids as well as the appearance of thin, rangy macrophages at Bruch''s membrane. Moreover, we showed that a zinc diet reduced the zinc mole fraction of melanosomes in the RPE and modulated the thickness of the Bruch''s membrane.     

Transcript and Protein Analysis Reveals Better Survival Skills of Monocyte-Derived Dendritic Cells Compared to Monocytes during Oxidative Stress

Background

Dendritic cells (DCs), professional antigen-presenting cells with the unique ability to initiate primary T-cell responses, are present in atherosclerotic lesions where they are exposed to oxidative stress that generates cytotoxic reactive oxygen species (ROS). A large body of evidence indicates that cell death is a major modulating factor of atherogenesis. We examined antioxidant defence systems of human monocyte-derived (mo)DCs and monocytes in response to oxidative stress.

Methods

Oxidative stress was induced by addition of tertiary-butylhydroperoxide (tert-BHP, 30 min). Cellular responses were evaluated using flow cytometry and confocal live cell imaging (both using 5-(and-6)-chloromethyl-2,7-dichlorodihydrofluorescein diacetate, CM-H₂DCFDA). Viability was assessed by the neutral red assay. Total RNA was extracted for a PCR profiler array. Five genes were selected for confirmation by Taqman gene expression assays, and by immunoblotting or immunohistochemistry for protein levels.

Results

Tert-BHP increased CM-H₂DCFDA fluorescence and caused cell death. Interestingly, all processes occurred more slowly in moDCs than in monocytes. The mRNA profiler array showed more than 2-fold differential expression of 32 oxidative stress–related genes in unstimulated moDCs, including peroxiredoxin-2 (PRDX2), an enzyme reducing hydrogen peroxide and lipid peroxides. PRDX2 upregulation was confirmed by Taqman assays, immunoblotting and immunohistochemistry. Silencing PRDX2 in moDCs by means of siRNA significantly increased CM-DCF fluorescence and cell death upon tert-BHP-stimulation.

Conclusions

Our results indicate that moDCs exhibit higher intracellular antioxidant capacities, making them better equipped to resist oxidative stress than monocytes. Upregulation of PRDX2 is involved in the neutralization of ROS in moDCs. Taken together, this points to better survival skills of DCs in oxidative stress environments, such as atherosclerotic plaques.     

Identification of Active Retinaldehyde Dehydrogenase Isoforms in the Postnatal Human Eye

Background/Objectives

Retinaldehyde dehydrogenase 2 (RALDH2) has been implicated in regulating all-trans-retinoic acid (atRA) synthesis in response to visual signals in animal models of myopia. To explore the potential role of retinaldehyde dehydrogenase (RALDH) enzymes and atRA in human postnatal ocular growth, RALDH activity, along with the distribution of RALDH1, RALDH2, and RALDH3 in the postnatal eye was determined.

Methodology

Retina, retinal pigment epithelium (RPE), choroid, and sclera were isolated from donor human eyes. RALDH catalytic activity was measured in tissue homogenates using an in vitro atRA synthesis assay together with HPLC quantification of synthesized atRA. Homogenates were compared by western blotting for RALDH1, RALDH2, and RALDH3 protein. Immunohistochemistry was used to determine RALDH1 and RALDH2 localization in posterior fundal layers of the human eye.

Principal Findings

In the postnatal human eye, RALDH catalytic activity was detected in the choroid (6.84 ± 1.20 pmol/hr/ug), RPE (5.46 ± 1.18 pmol/hr/ug), and retina (4.21 ± 1.55 pmol/hr/ug), indicating the presence of active RALDH enzymes in these tissues. RALDH2 was most abundant in the choroid and RPE, in moderate abundance in the retina, and in relatively low abundance in sclera. RALDH1 was most abundant in the choroid, in moderate abundance in the sclera, and substantially reduced in the retina and RPE. RALDH3 was undetectable in human ocular fundal tissues. In the choroid, RALDH1 and RALDH2 localized to slender cells in the stroma, some of which were closely associated with blood vessels.

Conclusions/Significance

Results of this study demonstrated that: 1) Catalytically active RALDH is present in postnatal human retina, RPE, and choroid, 2) RALDH1 and RALDH2 isoforms are present in these ocular tissues, and 3) RALDH1 and RALDH2 are relatively abundant in the choroid and/or RPE. Taken together, these results suggest that RALDH1 and 2 may play a role in the regulation of postnatal ocular growth in humans through the synthesis of atRA.     

The role of DJ-1 in the pathogenesis of endometriosis

Background

Endometriosis is an estrogen-dependent disease causing pelvic pain and infertility in 10% of reproductive-aged women. Despite a long history of the disease the pathogenesis of endometriosis is poorly understood. It is known that the expression of several proteins is either up or down regulated during endometriosis, but their precise role remains to be determined. DJ-1 is one such protein that is upregulated in eutopic endometrium of women having endometriosis suggesting that DJ-1 may be involved in the pathogenesis of endometriosis.

Methodology and Principal Findings

The role of DJ-1 in the pathogenesis of endometriosis was investigated. For this purpose the influence of DJ-1 on endometrial cell survival, attachment, proliferation, migration, and invasion either by overexpressing DJ-1 in normal endometrial cells or by knocking down DJ-1 expression in endometriotic cells using siRNA was investigated. The results indicated that DJ-1 protects endometrial cells from oxidative stress mediated apoptosis. Overexpression of DJ-1 in normal endometrial epithelial cells increases the adhesion on collagen type IV. However, no significant difference was observed incase of stromal cells. It was further demonstrated that DJ-1 regulates cell proliferation, migration, and invasion in normal endometrial and endometriotic epithelial cells whereas in the case of normal endometrial and endometriotic stromal cells, it regulates cell proliferation and invasion but not migration. Furthermore, the present study also indicated that DJ-1 regulates these cellular processes by modulating PI3K/Akt pathway by interacting and negatively regulating PTEN.

Conclusions

Abnormally high levels of DJ-1 expression may be involved in endometriosis, possibly by stimulating endometrial cell survival, proliferation, migration, and invasion.     

Hyperglycemia induces oxidative stress and impairs axonal transport rates in mice

Background

While hyperglycemia-induced oxidative stress damages peripheral neurons, technical limitations have, in part, prevented in vivo studies to determine the effect of hyperglycemia on the neurons in the central nervous system (CNS). While olfactory dysfunction is indicated in diabetes, the effect of hyperglycemia on olfactory receptor neurons (ORNs) remains unknown. In this study, we utilized manganese enhanced MRI (MEMRI) to assess the impact of hyperglycemia on axonal transport rates in ORNs. We hypothesize that (i) hyperglycemia induces oxidative stress and is associated with reduced axonal transport rates in the ORNs and (ii) hyperglycemia-induced oxidative stress activates the p38 MAPK pathway in association with phosphorylation of tau protein leading to the axonal transport deficits.

Research Design and Methods

T₁-weighted MEMRI imaging was used to determine axonal transport rates post-streptozotocin injection in wildtype (WT) and superoxide dismutase 2 (SOD2) overexpressing C57Bl/6 mice. SOD2 overexpression reduces mitochondrial superoxide load. Dihydroethidium staining was used to quantify the reactive oxygen species (ROS), specifically, superoxide (SO). Protein and gene expression levels were determined using western blotting and Q-PCR analysis, respectively.

Results

STZ-treated WT mice exhibited significantly reduced axonal transport rates and significantly higher levels of ROS, phosphorylated p38 MAPK and tau protein as compared to the WT vehicle treated controls and STZ-treated SOD2 mice. The gene expression levels of p38 MAPK and tau remained unchanged.

Conclusion

Increased oxidative stress in STZ-treated WT hyperglycemic mice activates the p38 MAPK pathway in association with phosphorylation of tau and attenuates axonal transport rates in the olfactory system. In STZ-treated SOD-overexpressing hyperglycemic mice in which superoxide levels are reduced, these deficits are reversed.     

Inhibition of renin-angiotensin system reverses endothelial dysfunction and oxidative stress in estrogen deficient rats

Background

Estrogen deficiency increases the cardiovascular risks in postmenopausal women. Inhibition of the renin-angiotensin system (RAS) and associated oxidative stress confers a cardiovascular protection, but the role of RAS in estrogen deficiency-related vascular dysfunction is unclear. The present study investigates whether the up-regulation of RAS and associated oxidative stress contributes to the development of endothelial dysfunction during estrogen deficiency in ovariectomized (OVX) rats.

Methodology/Principal Findings

Adult female rats were ovariectomized with and without chronic treatment with valsartan and enalapril. Isometric force measurement was performed in isolated aortae. The expression of RAS components was determined by immunohistochemistry and Western blotting method while ROS accumulation in the vascular wall was evaluated by dihydroethidium fluorescence. Ovariectomy increased the expression of angiotensin-converting enzyme (ACE), angiotensin II type 1 receptor (AT₁R), NAD(P)H oxidase, and nitrotyrosine in the rat aorta. An over-production of angiotensin II and ROS was accompanied by decreased phosphorylation of eNOS at Ser¹¹⁷⁷ in OVX rat aortae. These pathophysiological changes were closely coupled with increased oxidative stress and decreased nitric oxide bioavailability, culminating in markedly impaired endothelium-dependent relaxations. Furthermore, endothelial dysfunction and increased oxidative stress in aortae of OVX rats were inhibited or reversed by chronic RAS inhibition with enalapril or valsartan.

Conclusions/Significance

The novel findings highlight a significant therapeutic benefit of RAS blockade in the treatment of endothelial dysfunction-related vascular complications in postmenopausal states.     

Orally active multi-functional antioxidants are neuroprotective in a rat model of light-induced retinal damage

Background

Progression of age-related macular degeneration has been linked to iron dysregulation and oxidative stress that induce apoptosis of neural retinal cells. Since both antioxidants and chelating agents have been reported to reduce the progression of retinal lesions associated with AMD in experimental animals, the present study evaluates the ability of multi-functional antioxidants containing functional groups that can independently chelate redox metals and quench free radicals to protect the retina against light-induced retinal degeneration, a rat model of dry atrophic AMD.

Methods/Results

Proof of concept studies were conducted to evaluate the ability of 4-(5-hydroxypyrimidin-2-yl)-N,N-dimethyl-3,5-dioxopiperazine-1-sulfonamide (compound 4) and 4-(5-hydroxy-4,6-dimethoxypyrimidin-2-yl)-N,N-dimethyl-3,5-dioxopiperazine-1-sulfonamide (compound 8) to reduce retinal damage in 2-week dark adapted Wistar rats exposed to 1000 lx of light for 3 hours. Assessment of the oxidative stress markers 4- hydroxynonenal and nitrotyrosine modified proteins and Thioredoxin by ELISA and Western blots indicated that these compounds reduced the oxidative insult caused by light exposure. The beneficial antioxidant effects of these compounds in providing significant functional and structural protection were confirmed by electroretinography and quantitative histology of the retina.

Conclusions/Significance

The present study suggests that multi-functional compounds may be effective candidates for preventive therapy of AMD.