The purinergic receptor P2X7 (P2X7R) is implicated in all neurodegenerative diseases of the centr... more The purinergic receptor P2X7 (P2X7R) is implicated in all neurodegenerative diseases of the central nervous system. It is also involved in the retinal degeneration associated with glaucoma, age-related macular degeneration, and diabetic retinopathy, and its overexpression in the retina is evident in these disorders. Retinitis pigmentosa is a progressive degenerative disease that ultimately leads to blindness. Here, we investigated the expression of P2X7R during disease progression in the rd10 mouse model of RP. As the purinergic receptor P2X4 is widely co-expressed with P2X7R, we also studied its expression in the retina of rd10 mice. The expression of P2X7R and P2X4R was examined by immunohistochemistry, flow cytometry, and western blotting. In addition, we analyzed retinal functionality by electroretinographic recordings of visual responses and optomotor tests and retinal morphology. We found that the expression of P2X7R and P2X4R increased in rd10 mice concomitant with disease pr...
Multiple gene mutations have been associated with inherited retinal dystrophies (IRDs). Despite t... more Multiple gene mutations have been associated with inherited retinal dystrophies (IRDs). Despite the spectrum of phenotypes caused by the distinct mutations, IRDs display common physiopathology features. Cell death is accompanied by inflammation and oxidative stress. The vertebrate retina has several attributes that make this tissue vulnerable to oxidative and nitrosative imbalance. The high energy demands and active metabolism in retinal cells, as well as their continuous exposure to high oxygen levels and light-induced stress, reveal the importance of tightly regulated homeostatic processes to maintain retinal function, which are compromised in pathological conditions. In addition, the subsequent microglial activation and gliosis, which triggers the secretion of pro-inflammatory cytokines, chemokines, trophic factors, and other molecules, further worsen the degenerative process. As the disease evolves, retinal cells change their morphology and function. In disease stages where phot...
Inherited retinal dystrophies (IRDs) are a large group of genetically and clinically heterogeneou... more Inherited retinal dystrophies (IRDs) are a large group of genetically and clinically heterogeneous diseases characterized by the progressive degeneration of the retina, ultimately leading to loss of visual function. Oxidative stress and inflammation play fundamental roles in the physiopathology of these diseases. Photoreceptor cell death induces an inflammatory state in the retina. The activation of several molecular pathways triggers different cellular responses to injury, including the activation of microglia to eliminate debris and recruit inflammatory cells from circulation. Therapeutical options for IRDs are currently limited, although a small number of patients have been successfully treated by gene therapy. Many other therapeutic strategies are being pursued to mitigate the deleterious effects of IRDs associated with oxidative metabolism and/or inflammation, including inhibiting reactive oxygen species’ accumulation and inflammatory responses, and blocking autophagy. Several ...
Substance P (SP) is a neuropeptide that acts as a neurotransmitter or a neuromodulator in the ret... more Substance P (SP) is a neuropeptide that acts as a neurotransmitter or a neuromodulator in the retina. The aim of this study was to identify the type(s) and the distribution of the SP‐immunoreactive (SP‐IR) cells in the human retina. We have used an antiserum to SP to immunostain neurons in postmortem human retinae. Immunostained retinae were processedwith the avidin‐biotin complex (ABC) to visualize the cells either whole mounted in glycerol or embedded in plastic. Some retinae were also sectioned at 20 μm in order to obtain radial views of stained cells. SP‐IR amacrine cells stain intensely and appear to be of a single type in the human retina. They are large‐field cells with large cell bodies (16 μm diameter) lying in normal or displaced positions on either side of the inner plexiform layer (IPL). Their sturdy, spiny, and appendage‐bearing dendrites stratify in stratum 3 (S3) of the IPL, where many overlapping, fine dendrites intermingle to form a plexus of stained processes. Eith...
Investigative Opthalmology & Visual Science, 2007
PURPOSE. To study the involvement of stress-induced acetylcholinesterase (AChE) expression in lig... more PURPOSE. To study the involvement of stress-induced acetylcholinesterase (AChE) expression in light-induced retinal damage in albino rats. METHODS. Adult albino rats were exposed for 24 hours to bright, damaging light. AChE expression was monitored by in situ hybridization, by histochemistry for AChE activity, and by immunocytochemistry. An orphan antisense agent (Monarsen; Ester Neurosciences, Ltd., Herzlia Pituach, Israel) was administered intraperitoneally to minimize light-induced AChE expression. The electroretinogram (ERG) was recorded to assess retinal function. RESULTS. Twenty-four-hour exposure to bright light caused severe reduction in the ERG responses and augmented expression of mRNA for the "read-through" variant of AChE (AChE-R) in photoreceptor inner segments (IS), bipolar cells, and ganglion cells. AChE activity increased in IS. The expressed AChE protein was a novel variant, characterized by an extended N terminus (N-AChE). Systemic administration of the orphan antisense agent, Monarsen, reduced the photic induction of mRNA for AChE-R, and of the N-AChE protein. Rats exposed to bright, damaging light and treated daily with Monarsen exhibited larger ERG responses, relatively thicker outer nuclear layer (ONL), and more ONL nuclei than did rats exposed to the same damaging light but treated daily with saline. CONCLUSIONS. The findings indicate that the photic-induced novel variant of AChE (N-AChE-R) may be causally involved with retinal light damage and suggest the use of RNA targeting for limiting such damage.
Retinitis pigmentosa (RP) is a group of inherited retinal disorders that lead to photoreceptor lo... more Retinitis pigmentosa (RP) is a group of inherited retinal disorders that lead to photoreceptor loss. RP has been reported to be related to oxidative stress, autophagy, and inflammation. (-)-Epigallocatechin gallate (EGCG), the most abundant catechin-based flavonoid in green tea leaves, has significant antioxidant, anti-carcinogenic, antimicrobial, and neuroprotective properties. EGCG, given its low molecular weight and hydrophilic properties, can cross the blood-retinal barrier and is able to reach different ocular tissues such as the lens, cornea, and retina. EGCG has been shown to provide retinal protection against ischemia; sodium nitroprusside-, N-methyl-D-aspartate-, lipopolysaccharide-, light-, sodium iodate-, or H 2 O 2-induced damage and diabetic retinopathy. This suggests that systemic EGCG administration has the potential to protect against retinal degenerative or neurodegenerative diseases such as RP. The aim of this work was to investigate whether EGCG can protect against RP progression in the animal P23H line 1, the model of RP. Albino P23H rats were crossed with pigmented Long Evans rats to produce offspring exhibiting the clinical features of RP. Pigmented P23H rats were treated via intraperitoneal injection with saline or EGCG at a dose of 25 mg/kg every week from P100 to P160 and then compared to wild-type Long Evans rats. Rats treated with EGCG showed better visual and retinal electrical function with increased contrast sensitivity and b-wave values compared with those observed in P23H rats treated with vehicle. EGCG reduced lipid peroxidation and increased total antioxidant capacity and catalase and superoxide dismutase activities. No differences were observed in visual acuity, nitrate levels, nitrite levels or glutathione S-transferase activity. In conclusion, EGCG not only reduced the loss of visual function in P23H rats but also improved the levels of antioxidant enzymes and reduced oxidative damage. This study was approved by the Institutional Animal Care and Use Committee (CEICA) from the University of Zaragoza under project license PI12/14 on July 11, 2014.
Purpose To evaluate fundus autofluorescence (FAF) and OCT changes comparing with immunohistochemi... more Purpose To evaluate fundus autofluorescence (FAF) and OCT changes comparing with immunohistochemical (ICC) analysis in long term degeneration of P23H rat and to investigate retinal and choroidal vascularization using fluorescein and indocianin green angiography. Methods Twenty-albino homozygous P23H line 1 rats aging from 18 postnatal days (P18) to 27 months and wild-type albino Sprague-Dawley (SD rats) (2 and 15 months old) were used for this study. Normal pigmented Long Evans (LE) 2 months old rats were used to compare FAF findings. ICC was performed to correlate with the findings of OCT and AF changes. Results FAF pattern varied from not findings at P18 to a mosaic of hyperfluorescent dots in the rats of 6 months or older. Retinal thicknesses diminished during the time: 205.2–183.18 μm in SD rats and 189.88–58.15 μm in P23H rats. In long term degeneration, OCT showed severe changes at the retinal layers; ICC helped to identify the cell loss and remodeling changes. Conclusions Autofluorescent ophthalmoscopy is a non-invasive method that can detect changes in metabolic activity at the RPE in animal models of retinal degeneration in vivo. Retinal vascular plexus changed with aging. OCT showed a diminution of retinal thickness and retinal layer changes with the degeneration. ICC shows a good correlation.
The purinergic receptor P2X7 (P2X7R) is implicated in all neurodegenerative diseases of the centr... more The purinergic receptor P2X7 (P2X7R) is implicated in all neurodegenerative diseases of the central nervous system. It is also involved in the retinal degeneration associated with glaucoma, age-related macular degeneration, and diabetic retinopathy, and its overexpression in the retina is evident in these disorders. Retinitis pigmentosa is a progressive degenerative disease that ultimately leads to blindness. Here, we investigated the expression of P2X7R during disease progression in the rd10 mouse model of RP. As the purinergic receptor P2X4 is widely co-expressed with P2X7R, we also studied its expression in the retina of rd10 mice. The expression of P2X7R and P2X4R was examined by immunohistochemistry, flow cytometry, and western blotting. In addition, we analyzed retinal functionality by electroretinographic recordings of visual responses and optomotor tests and retinal morphology. We found that the expression of P2X7R and P2X4R increased in rd10 mice concomitant with disease pr...
Multiple gene mutations have been associated with inherited retinal dystrophies (IRDs). Despite t... more Multiple gene mutations have been associated with inherited retinal dystrophies (IRDs). Despite the spectrum of phenotypes caused by the distinct mutations, IRDs display common physiopathology features. Cell death is accompanied by inflammation and oxidative stress. The vertebrate retina has several attributes that make this tissue vulnerable to oxidative and nitrosative imbalance. The high energy demands and active metabolism in retinal cells, as well as their continuous exposure to high oxygen levels and light-induced stress, reveal the importance of tightly regulated homeostatic processes to maintain retinal function, which are compromised in pathological conditions. In addition, the subsequent microglial activation and gliosis, which triggers the secretion of pro-inflammatory cytokines, chemokines, trophic factors, and other molecules, further worsen the degenerative process. As the disease evolves, retinal cells change their morphology and function. In disease stages where phot...
Inherited retinal dystrophies (IRDs) are a large group of genetically and clinically heterogeneou... more Inherited retinal dystrophies (IRDs) are a large group of genetically and clinically heterogeneous diseases characterized by the progressive degeneration of the retina, ultimately leading to loss of visual function. Oxidative stress and inflammation play fundamental roles in the physiopathology of these diseases. Photoreceptor cell death induces an inflammatory state in the retina. The activation of several molecular pathways triggers different cellular responses to injury, including the activation of microglia to eliminate debris and recruit inflammatory cells from circulation. Therapeutical options for IRDs are currently limited, although a small number of patients have been successfully treated by gene therapy. Many other therapeutic strategies are being pursued to mitigate the deleterious effects of IRDs associated with oxidative metabolism and/or inflammation, including inhibiting reactive oxygen species’ accumulation and inflammatory responses, and blocking autophagy. Several ...
Substance P (SP) is a neuropeptide that acts as a neurotransmitter or a neuromodulator in the ret... more Substance P (SP) is a neuropeptide that acts as a neurotransmitter or a neuromodulator in the retina. The aim of this study was to identify the type(s) and the distribution of the SP‐immunoreactive (SP‐IR) cells in the human retina. We have used an antiserum to SP to immunostain neurons in postmortem human retinae. Immunostained retinae were processedwith the avidin‐biotin complex (ABC) to visualize the cells either whole mounted in glycerol or embedded in plastic. Some retinae were also sectioned at 20 μm in order to obtain radial views of stained cells. SP‐IR amacrine cells stain intensely and appear to be of a single type in the human retina. They are large‐field cells with large cell bodies (16 μm diameter) lying in normal or displaced positions on either side of the inner plexiform layer (IPL). Their sturdy, spiny, and appendage‐bearing dendrites stratify in stratum 3 (S3) of the IPL, where many overlapping, fine dendrites intermingle to form a plexus of stained processes. Eith...
Investigative Opthalmology & Visual Science, 2007
PURPOSE. To study the involvement of stress-induced acetylcholinesterase (AChE) expression in lig... more PURPOSE. To study the involvement of stress-induced acetylcholinesterase (AChE) expression in light-induced retinal damage in albino rats. METHODS. Adult albino rats were exposed for 24 hours to bright, damaging light. AChE expression was monitored by in situ hybridization, by histochemistry for AChE activity, and by immunocytochemistry. An orphan antisense agent (Monarsen; Ester Neurosciences, Ltd., Herzlia Pituach, Israel) was administered intraperitoneally to minimize light-induced AChE expression. The electroretinogram (ERG) was recorded to assess retinal function. RESULTS. Twenty-four-hour exposure to bright light caused severe reduction in the ERG responses and augmented expression of mRNA for the "read-through" variant of AChE (AChE-R) in photoreceptor inner segments (IS), bipolar cells, and ganglion cells. AChE activity increased in IS. The expressed AChE protein was a novel variant, characterized by an extended N terminus (N-AChE). Systemic administration of the orphan antisense agent, Monarsen, reduced the photic induction of mRNA for AChE-R, and of the N-AChE protein. Rats exposed to bright, damaging light and treated daily with Monarsen exhibited larger ERG responses, relatively thicker outer nuclear layer (ONL), and more ONL nuclei than did rats exposed to the same damaging light but treated daily with saline. CONCLUSIONS. The findings indicate that the photic-induced novel variant of AChE (N-AChE-R) may be causally involved with retinal light damage and suggest the use of RNA targeting for limiting such damage.
Retinitis pigmentosa (RP) is a group of inherited retinal disorders that lead to photoreceptor lo... more Retinitis pigmentosa (RP) is a group of inherited retinal disorders that lead to photoreceptor loss. RP has been reported to be related to oxidative stress, autophagy, and inflammation. (-)-Epigallocatechin gallate (EGCG), the most abundant catechin-based flavonoid in green tea leaves, has significant antioxidant, anti-carcinogenic, antimicrobial, and neuroprotective properties. EGCG, given its low molecular weight and hydrophilic properties, can cross the blood-retinal barrier and is able to reach different ocular tissues such as the lens, cornea, and retina. EGCG has been shown to provide retinal protection against ischemia; sodium nitroprusside-, N-methyl-D-aspartate-, lipopolysaccharide-, light-, sodium iodate-, or H 2 O 2-induced damage and diabetic retinopathy. This suggests that systemic EGCG administration has the potential to protect against retinal degenerative or neurodegenerative diseases such as RP. The aim of this work was to investigate whether EGCG can protect against RP progression in the animal P23H line 1, the model of RP. Albino P23H rats were crossed with pigmented Long Evans rats to produce offspring exhibiting the clinical features of RP. Pigmented P23H rats were treated via intraperitoneal injection with saline or EGCG at a dose of 25 mg/kg every week from P100 to P160 and then compared to wild-type Long Evans rats. Rats treated with EGCG showed better visual and retinal electrical function with increased contrast sensitivity and b-wave values compared with those observed in P23H rats treated with vehicle. EGCG reduced lipid peroxidation and increased total antioxidant capacity and catalase and superoxide dismutase activities. No differences were observed in visual acuity, nitrate levels, nitrite levels or glutathione S-transferase activity. In conclusion, EGCG not only reduced the loss of visual function in P23H rats but also improved the levels of antioxidant enzymes and reduced oxidative damage. This study was approved by the Institutional Animal Care and Use Committee (CEICA) from the University of Zaragoza under project license PI12/14 on July 11, 2014.
Purpose To evaluate fundus autofluorescence (FAF) and OCT changes comparing with immunohistochemi... more Purpose To evaluate fundus autofluorescence (FAF) and OCT changes comparing with immunohistochemical (ICC) analysis in long term degeneration of P23H rat and to investigate retinal and choroidal vascularization using fluorescein and indocianin green angiography. Methods Twenty-albino homozygous P23H line 1 rats aging from 18 postnatal days (P18) to 27 months and wild-type albino Sprague-Dawley (SD rats) (2 and 15 months old) were used for this study. Normal pigmented Long Evans (LE) 2 months old rats were used to compare FAF findings. ICC was performed to correlate with the findings of OCT and AF changes. Results FAF pattern varied from not findings at P18 to a mosaic of hyperfluorescent dots in the rats of 6 months or older. Retinal thicknesses diminished during the time: 205.2–183.18 μm in SD rats and 189.88–58.15 μm in P23H rats. In long term degeneration, OCT showed severe changes at the retinal layers; ICC helped to identify the cell loss and remodeling changes. Conclusions Autofluorescent ophthalmoscopy is a non-invasive method that can detect changes in metabolic activity at the RPE in animal models of retinal degeneration in vivo. Retinal vascular plexus changed with aging. OCT showed a diminution of retinal thickness and retinal layer changes with the degeneration. ICC shows a good correlation.
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Papers by Nicolás Cuenca