Enrique de la Rosa - Academia.edu (original) (raw)

Papers by Enrique de la Rosa

International Journal of Molecular Sciences

Genetic mosaicism is an intriguing physiological feature of the mammalian brain that generates al... more Genetic mosaicism is an intriguing physiological feature of the mammalian brain that generates altered genetic information and provides cellular, and prospectively functional, diversity in a manner similar to that of the immune system. However, both its origin and its physiological significance remain poorly characterized. Most, if not all, cases of somatic mosaicism require prior generation and repair of DNA double strand breaks (DSBs). The relationship between DSB generation, neurogenesis, and early neuronal cell death revealed by our studies in the developing retina provides new perspectives on the different mechanisms that contribute to DNA rearrangements in the developing brain. Here, we speculate on the physiological significance of these findings.

International Journal of Molecular Sciences, 2021

Although considered a rare retinal dystrophy, retinitis pigmentosa (RP) is the primary cause of h... more Although considered a rare retinal dystrophy, retinitis pigmentosa (RP) is the primary cause of hereditary blindness. Given its diverse genetic etiology (>3000 mutations in >60 genes), there is an urgent need for novel treatments that target common features of the disease. TLR2 is a key activator of innate immune response. To examine its role in RP progression we characterized the expression profile of Tlr2 and its adaptor molecules and the consequences of Tlr2 deletion in two genetically distinct models of RP: Pde6brd10/rd10 (rd10) and RhoP23H/+ (P23H/+) mice. In both models, expression levels of Tlr2 and its adaptor molecules increased in parallel with those of the proinflammatory cytokine Il1b. In rd10 mice, deletion of a single Tlr2 allele had no effect on visual function, as evaluated by electroretinography. However, in both RP models, complete elimination of Tlr2 attenuated the loss of visual function and mitigated the loss of photoreceptor cell numbers. In Tlr2 null rd1...

Scientific Reports, 2019

DNA double-strand breaks (DSBs), selectively visualized as γ-H2AX+ foci, occur during the develop... more DNA double-strand breaks (DSBs), selectively visualized as γ-H2AX+ foci, occur during the development of the central nervous system, including the retina, although their origin and biological significance are poorly understood. Mutant mice with DSB repair mechanism defects exhibit increased numbers of γ-H2AX+ foci, increased cell death during neural development, and alterations in axonogenesis in the embryonic retina. The aim of this study was to identify putative sources of DSBs. One of the identified DSBs sources is LINE-1 retrotransposition. While we did not detect changes in LINE-1 DNA content during the early period of cell death associated with retinal neurogenesis, retinal development was altered in mice lacking RAG-2, a component of the RAG-1,2-complex responsible for initiating somatic recombination in lymphocytes. Although γ-H2AX+ foci were less abundant in the rag2−/− mouse retina, retinal ganglion cell death was increased and axonal growth and navigation were impaired in...

Drug Discovery, 2018

As an extension of the central nervous system (CNS), the retina shares with the brain certain dev... more As an extension of the central nervous system (CNS), the retina shares with the brain certain developmental, physiological, and pathological characteristics. However, the underlying mechanisms and pathological signs common to neurodegenerative conditions of both the retina and brain have been relatively overlooked. In animal models and in human patients, marked retinal alterations have been demonstrated in Alzheimer´s disease, Parkinson´s disease, and multiple sclerosis, among other pathologies. Furthermore, neurodegeneration of the retina and brain appears to be mediated by similar mechanisms, which include protein aggregation, neuroinflammation, and cell death. Analysis of the retina, which is easily accessible to objective techniques, may therefore constitute an effective tool for the screening and follow-up of CNS neurodegeneration. Moreover, patients with retinal neurodegeneration could potentially benefit from the broad array of pharmacological compounds that have been designed and tested for the treatment of the aforementioned CNS pathologies. Supporting this view, we have shown that GSK-3 inhibitors, which have already been tested in clinical trials to treat several neurodegenerative conditions of the brain, attenuate retinal damage and vision loss in a mouse model of retinitis pigmentosa. Furthermore, systemic proinsulin treatment preserves visual and cognitive function in mouse models of retinitis pigmentosa and precocious aging, respectively.

The Journal of Neuroscience, 2000

The signaling cascade Ras/Raf/mitogen-activated protein kinases modulates cell proliferation, dif... more The signaling cascade Ras/Raf/mitogen-activated protein kinases modulates cell proliferation, differentiation, and survival, all key cellular processes during neural development. To better define thein vivorole of Raf during chick retinal neurogenesis, we interfered with Raf-dependent signaling during days 4.5 to 7.5 of embryonic development by expressing a dominant negative mutant of c-Raf (ΔRaf), which blocks Ras-dependent Raf activation, and by overexpressing wild-type c-Raf. ΔRaf expression induced an increase in cell death by apoptosis, whereas it did not affect overall cell proliferation and differentiation. In parallel, the number of Islet-1/2-positive and TUJ1-positive retinal ganglion cells were diminished in their definitive layer, whereas there was an increase in the number of mislocated Islet-1/2-positive cells. This disturbed morphogenesis correlated with a disruption of the optic fiber layer. Conversely, c-Raf overexpression caused moderate opposite effects on apoptosi...

Frontiers in Molecular Neuroscience, 2018

Molecular neurodegeneration, Apr 16, 2018

Retinitis pigmentosa (RP) is a group of hereditary retinal neurodegenerative conditions character... more Retinitis pigmentosa (RP) is a group of hereditary retinal neurodegenerative conditions characterized by primary dysfunction and death of photoreceptor cells, resulting in visual loss and, eventually, blindness. To date, no effective therapies have been transferred to clinic. Given the diverse genetic etiology of RP, targeting common cellular and molecular retinal alterations has emerged as a potential therapeutic strategy. Using the Pde6b mouse model of RP, we investigated the effects of daily intraperitoneal administration of VP3.15, a small-molecule heterocyclic GSK-3 inhibitor. Gene expression was analyzed by quantitative PCR and protein expression and phosphorylation by Western blot. Photoreceptor preservation was evaluated by histological analysis and visual function was assessed by electroretinography. In rd10 retinas, increased expression of pro-inflammatory markers and reactive gliosis coincided with the early stages of retinal degeneration. Compared with wild-type controls...

Cell death & disease, Jul 13, 2017

ProNGF signaling through p75(NTR) has been associated with neurodegenerative disorders. Retinitis... more ProNGF signaling through p75(NTR) has been associated with neurodegenerative disorders. Retinitis pigmentosa (RP) comprises a group of inherited retinal dystrophies that causes progressive photoreceptor cell degeneration and death, at a rate dependent on the genetic mutation. There are more than 300 mutations causing RP, and this is a challenge to therapy. Our study was designed to explore a common mechanism for p75(NTR) in the progression of RP, and assess its potential value as a therapeutic target. The proNGF/p75(NTR) system is present in the dystrophic retina of the rd10 RP mouse model. Compared with wild-type (WT) retina, the levels of unprocessed proNGF were increased in the rd10 retina at early degenerative stages, before the peak of photoreceptor cell death. Conversely, processed NGF levels were similar in rd10 and WT retinas. ProNGF remained elevated throughout the period of photoreceptor cell loss, correlating with increased expression of α2-macroglobulin, an inhibitor of ...

Journal of enzyme inhibition and medicinal chemistry, 2017

Retinitis pigmentosa (RP) is an inherited retinal dystrophy that courses with progressive degener... more Retinitis pigmentosa (RP) is an inherited retinal dystrophy that courses with progressive degeneration of retinal tissue and loss of vision. Currently, RP is an unpreventable, incurable condition. We propose glycogen synthase kinase 3 (GSK-3) inhibitors as potential leads for retinal cell neuroprotection, since the retina is also a part of the central nervous system and GSK-3 inhibitors are potent neuroprotectant agents. Using a chemical genetic approach, diverse small molecules with different potency and binding mode to GSK-3 have been used to validate and confirm GSK-3 as a pharmacological target for RP. Moreover, this medicinal chemistry approach has provided new leads for the future disease-modifying treatment of RP.

Investigative ophthalmology & visual science, Jul 1, 2016

The induction of proinsulin expression by transgenesis or intramuscular gene therapy has been sho... more The induction of proinsulin expression by transgenesis or intramuscular gene therapy has been shown previously to retard retinal degeneration in mouse and rat models of retinitis pigmentosa (RP), a group of inherited conditions that result in visual impairment. We investigated whether intraocular treatment with biodegradable poly (lactic-co-glycolic) acid microspheres (PLGA-MS) loaded with proinsulin has cellular and functional neuroprotective effects in the retina. Experiments were performed using the Pde6brd10 mouse model of RP. Methionylated human recombinant proinsulin (hPI) was formulated in PLGA-MS, which were administered by intravitreal injection on postnatal days (P) 14 to 15. Retinal neuroprotection was assessed at P25 by electroretinography, and by evaluating outer nuclear layer (ONL) cellular preservation. The attenuation of photoreceptor cell death by hPI was determined by TUNEL assay in cultured P22 retinas, as well as Akt phosphorylation by immunoblotting. We successf...

Scientific reports, May 13, 2016

Programmed cell death occurs naturally at different stages of neural development, including neuro... more Programmed cell death occurs naturally at different stages of neural development, including neurogenesis. The functional role of this early phase of neural cell death, which affects recently differentiated neurons among other cell types, remains undefined. Some mouse models defective in DNA double-strand break (DSB) repair present massive cell death during neural development, occasionally provoking embryonic lethality, while other organs and tissues remain unaffected. This suggests that DSBs occur frequently and selectively in the developing nervous system. We analyzed the embryonic retina of a mouse model deficient in the error-prone DNA polymerase μ (Polμ), a key component of the non-homologous end-joining (NHEJ) repair system. DNA DSBs were increased in the mutant mouse at embryonic day 13.5 (E13.5), as well as the incidence of cell death that affected young neurons, including retinal ganglion cells (RGCs). Polμ(-/-) mice also showed disturbed RGC axonal growth and navigation, an...

Advanced materials (Deerfield Beach, Fla.), Jan 9, 2015

A novel suspended planar-array chips technology is described, which effectively allows molecular ... more A novel suspended planar-array chips technology is described, which effectively allows molecular multiplexing using a single suspended chip to analyze extraordinarily small volumes. The suspended chips are fabricated by combining silicon-based technology and polymer-pen lithography, obtaining increased molecular pattern flexibility, and improving miniaturization and parallel production. The chip miniaturization is so dramatic that it permits the intracellular analysis of living cells.

During the development of the vertebrate nervous system, multiple physiological processes are inv... more During the development of the vertebrate nervous system, multiple physiological processes are involved in the generation of its complex cytoarchitecture and functionality. Among them, programmed cell death has been recognized as a key process that affects connecting neurons. By contrast, there is limited information available regarding the cell death that affects neuroepithelial cells, and recently born neurons and glia, hindering the comprehensive understanding of neural development. We have demonstrated that exquisitely regulated PCD occurs during early stages of neural development such as neurulation and neurogenesis. We have characterized how survival signals from proteins like proinsulin/insulin, c-Raf, and HSC70 counteract caspase-dependent apoptosis, which affects neuroepithelial cells proliferation and the generation of retinal ganglion cells. Furthermore, the characterization of these physiological signals during retinal neurogenesis has the potential to provide new therape...

The Scientific World Journal, 2013

Orchestrated proliferation, differentiation, and cell death contribute to the generation of the c... more Orchestrated proliferation, differentiation, and cell death contribute to the generation of the complex cytoarchitecture of the central nervous system, including that of the neuroretina. However, few studies have comprehensively compared the spatiotemporal patterns of these 3 processes, or their relative magnitudes. We performed a parallel study in embryonic chick and mouse retinas, focusing on the period during which the first neurons, the retinal ganglion cells (RGCs), are generated. The combination ofin vivoBrdU incorporation, immunolabeling of retinal whole mounts for BrdU and for the neuronal markers Islet1/2 andβIII-tubulin, and TUNEL allowed for precise cell scoring and determination the spatiotemporal patterns of cell proliferation, differentiation, and death. As predicted, proliferation preceded differentiation. Cell death and differentiation overlapped to a considerable extent, although the magnitude of cell death exceeded that of neuronal differentiation. Precise quantifi...

Proceedings of the National Academy of Sciences, 1995

Early neurogenesis progresses by an initial massive proliferation of neuroepithelial cells follow... more Early neurogenesis progresses by an initial massive proliferation of neuroepithelial cells followed by a sequential differentiation of the various mature neural cell types. The regulation of these processes by growth factors is poorly understood. We intend to understand, in a well-defined biological system, the embryonic chicken retina, the role of the insulin-related growth factors in neurogenesis. We demonstrate the local presence of signaling elements together with a biological response to the factors. Neuroretina at days 6-8 of embryonic development (E6-E8) expressed proinsulin/insulin and insulin-like growth factor I (IGF-I) mRNAs as well as insulin receptor and IGF type I receptor mRNAs. In parallel with this in vivo gene expression, E5 cultured neuroretinas synthesized and released to the medium a metabolically radiolabeled immunoprecipitable insulin-related peptide. Furthermore, insulin-related immunoreactive material with a HPLC mobility close to that of proinsulin was foun...

Journal of Cell Science, 2002

Nerve growth factor induces cell death in organotypic cultures of otic vesicle explants. This cel... more Nerve growth factor induces cell death in organotypic cultures of otic vesicle explants. This cell death has a restricted pattern that reproduces the in vivo pattern of apoptosis occurring during inner ear development. In this study, we show that binding of nerve growth factor to its low affinity p75 neurotrophin receptor is essential to achieve the apoptotic response. Blockage of binding to p75 receptor neutralized nerve-growth-factor-induced cell death,as measured by immunoassays detecting the presence of cytosolic oligonucleosomes and by TUNEL assay to visualize DNA fragmentation. Nerve growth factor also induced a number of cell-death-related intracellular events including ceramide generation, caspase activation and poly-(ADP ribose)polymerase cleavage. Again, p75 receptor blockade completely abolished all of these effects. Concerning the intracellular pathway, ceramide increase depended on initiator caspases, whereas its actions depended on both initiator and effector caspases,...

Investigative Opthalmology & Visual Science, 2011

PURPOSE. To characterize the effect of IGF-I in the rd10 mouse model of retinitis pigmentosa at t... more PURPOSE. To characterize the effect of IGF-I in the rd10 mouse model of retinitis pigmentosa at the cellular level, focusing on the role of microglia in the neurodegenerative process. METHODS. Both organotypic retinal explants and intravitreal injections were used to assess the effect of IGF-I on photoreceptor cell death in the Pde6b rd10 mice. Cell death was determined by TUNEL in retinal sections and by ELISA of free nucleosomes in retinal extracts. The number and distribution of microglial cells was visualized by immunolabeling with Cd11b and Iba1 antibodies. Depletion of microglia in culture was achieved by treatment with liposomes containing clodronate. RESULTS. Both ex vivo and in vivo IGF-I treatment reduced the number of TUNEL-positive nuclei in rd10 mouse retinas. In addition, IGF-I treatment in explants increased the number of microglial cells in the ONL. Depletion of microglia in explants with liposomes containing clodronate diminished the neuroprotective effect of IGF-I but also moderately reduced photoreceptor cell death in rd10 retinas cultured in the absence of IGF-I. CONCLUSIONS. IGF-I is able to attenuate photoreceptor cell death both ex vivo and in vivo in the rd10 mouse retina. Microglia is required for the neuroprotective effect of IGF-I in the dystrophic retina. In addition, microglial cells play a detrimental role, seemingly led to neuroprotection by IGF-I. (Invest Ophthalmol Vis Sci. 2011;52:9124-9130

Investigative Opthalmology & Visual Science, 2008

EMBO reports, 2005

Proinsulin gene expression regulation and function during early embryonic development differ rema... more Proinsulin gene expression regulation and function during early embryonic development differ remarkably from those found in postnatal organisms. The embryonic proinsulin protein content decreased from gastrulation to neurulation in contrast with the overall proinsulin messenger RNA increase. This is due to increasing levels of a proinsulin mRNA variant generated by intron 1 retention in the 5 0 untranslated region. Inclusion of intron 1 inhibited proinsulin translation almost completely without affecting nuclear export or cytoplasmic decay. The novel proinsulin mRNA isoform expression was developmentally regulated and tissue specific. The proportion of intron retention increased from gastrulation to organogenesis, was highest in the heart tube and presomitic region, and could not be detected in the pancreas. Notably, proinsulin addition induced cardiac marker gene expression in the early embryonic stages when the translationally active transcript was expressed. We propose that regulated unproductive splicing and translation is a mechanism that regulates proinsulin expression in accordance with specific requirements in developing vertebrates.

Diabetologia, 2006

In postnatal organisms, insulin is well known as an essential anabolic hormone responsible for ma... more In postnatal organisms, insulin is well known as an essential anabolic hormone responsible for maintaining glucose homeostasis. Its biosynthesis by the pancreatic beta cell has been considered a model of tissue-specific gene expression. However, proinsulin mRNA and protein have been found in embryonic stages before the formation of the pancreatic primordium, and later, in extrapancreatic tissues including the nervous system. Phylogenetic studies have also confirmed that production of insulin-like peptides antecedes the morphogenesis of a pancreas, and that these peptides contribute to normal development. In recent years, other roles for insulin distinct from its metabolic function have emerged also in vertebrates. During embryonic development, insulin acts as a survival factor and is involved in early morphogenesis. These findings are consistent with the observation that, at these stages, the proinsulin gene product remains as the precursor form, proinsulin. Independent of its low metabolic activity, proinsulin stimulates proliferation in developing neuroretina, as well as cell survival and cardiogenesis in early embryos. Insulin/proinsulin levels are finely regulated during development, since an excess of the protein interferes with correct morphogenesis and is deleterious for the embryo. This fine-tuned regulation is achieved by the expression of alternative embryonic proinsulin transcripts that have diminished translational activity.