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Photoreceptor degeneration, second order neuron remodeling and glia reactivity in an in vivo and in vitro model of retinal neurodegeneration
Örebro University, School of Medical Sciences.
2016 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

Photoreceptors have the ability to last during the entire lifespan of an individual. Being the first line of neurons in the visual transduction pathway, their health and maintenance is eminent for proper retinal function. However, photoreceptors are susceptible to neurodegenerative retinal dystrophies. A number of retinal pathologies such as retinitis pigmentosa, age-related amacular degeneration and diabetic retinopathy have been linked to photoreceptor death. Moreover, photoreceptor degeneration has been shown to affect downstream inner nuclear layer cells as well as induce reactive responses from Müller cells and microglia. Since current treatments are ineffective in preventing the degeneration of these neurons, intense research is still underway to discover novel treatment modalities. In this thesis, photoreceptor degeneration was assessed in an in vivo and in vitro model of neurodegeneration. Moreover, a possible mode of preserving these neurons by the use of human neural progenitor cells (hNPCs) was investigated. The in vivo pdgf-bret/ret (platelet derived growth factor-b retention motif knockout) mouse model, which shows severe vascular pathology as a result of detachment of pericytes from the vascular endothelium, was studied during the first postnatal month. In a short time span, i.e. between postnatal day (P)10 and P15, retinopathic features were observed. Photoreceptor degeneration related to cell death, cone outer segment (OS) shortening and synapse disassembly in the outer plexiform layer (OPL) was seen. The second order rod bipolar cells underwent remodeling and the Müller cells became gliotic with increased expression of GFAP (glial fibrillary acidic protein). Microglial cells were also observed to convert to their reactive amoeboid-like phenotype. These features seemed to become more severe in the older P28 mutants. In the in vitro porcine retinal explant model, photoreceptor death significantly increased by 3 days in vitro (div). This was associated with loss of cone OSs, opsin mislocalization and loss of synaptic integrity in the OPL. Horizontal cell death and remodeling was also observed together with a severe gliotic response from the Müller cells. Human neural progenitor cell cocultured explants for 3 div had the ability to preserve photoreceptor survival by means of OS conservation, better opsin trafficking and maintaining synaptic integrity. However, Müller cell gliosis was only mitigated by a decreased density of GFAP immunoreactive Müller cells. In conclusion, both the in vivo and in vitro model of neurodegeneration demonstrate the vulnerability of photoreceptors to various mechanisms of retinal injury. Interestingly, hNPC derived neurotrophic factors had neuroprotective qualities in 3 div porcine retinal explants.

Place, publisher, year, edition, pages
Örebro: Örebro University , 2016. , 40 p.
National Category
Other Basic Medicine
Research subject
Biomedicine
Identifiers
URN: urn:nbn:se:oru:diva-53596OAI: oai:DiVA.org:oru-53596DiVA: diva2:1048273
Presentation
2016-11-23, Örebro universitet, Campus USÖ, Hörsal C1, Södra Grev Rosengatan 30, Örebro, 13:15 (Swedish)
Available from: 2016-11-21 Created: 2016-11-21 Last updated: 2016-11-21Bibliographically approved
List of papers
1. Photoreceptor degeneration, structural remodeling and glial activation: a morphological study on a genetic mouse model for pericyte deficiency
Open this publication in new window or tab >>Photoreceptor degeneration, structural remodeling and glial activation: a morphological study on a genetic mouse model for pericyte deficiency
2014 (English)In: Neuroscience, ISSN 0306-4522, E-ISSN 1873-7544, Vol. 279, 269-284 p.Article in journal (Refereed) Published
Abstract [en]

Interaction between pericytes and endothelial cells via platelet-derived growth factor B (PDGF-B) signaling is critical for the development of the retinal microvasculature. The PDGF-B retention motif controls the spatial distribution range of the growth factor in the vicinity of its producing endothelial cells allowing its recognition by PDGF receptor beta-(PDGFR-beta)-carrying pericytes; this promotes recruitment of pericytes to the vascular basement membrane. Impairment of the PDGF-B signaling mechanism causes development of vascular abnormalities, and in the retina this consequently leads to defects in the neurological circuitry. The vascular pathology in the pdgf-b(ret/ret) (PDGF-B retention motif knockout) mouse retina has been previously reported; our study investigates the progressive neuronal defects and changes in the retinal morphology of this pericyte-deficient mouse model. Immunohistochemical analysis revealed retinal injuries to occur as early as postnatal day (P) 10 with substantial damage progressing from P15 and onward. Vascular abnormalities were apparent from P10, however, prominent neuronal defects were mostly observed from P15, beginning with the compromised integrity of the laminated retinal structure characterized by the presence of rosettes and focally distorted regions. Photoreceptor degeneration was observed by loss of both rod and cone cells, including the disassembly and altered structure of their synaptic terminals. Significant shortening of cone outer segments was observed from P10 and later stages; however, decrease in cone density was only observed at P28. Disorganization and dendrite remodeling of rod bipolar cells also added to the diminished neural and synaptic integrity. Moreover, in response to retinal injuries, Muller and microglial cells were observed to be in the reactive phenotype from P15 and onward. Such a sequence of events indicates that the pdgf-b(ret/ret) mouse model displays a short time frame between P10 and P15, during which the retina shifts to a retinopathic phase by the development of prominently altered morphological features.

Keyword
photoreceptor degeneration, cell death, rosette, synapse, microglia, Müller cells
National Category
Neurosciences
Research subject
Neurology
Identifiers
urn:nbn:se:oru:diva-38893 (URN)10.1016/j.neuroscience.2014.09.013 (DOI)000343633800022 ()25224828 (PubMedID)2-s2.0-84907487118 (ScopusID)
Note

Funding Agencies:

Faculty of Medicine at Örebro University

Signhild Engkvist Foundation

Ögonfonden

Crown Princess Margaretas Committee for the Blind

Gun and Bertil Stohnes Foundation

Karolinska Fonder och Stiftelser

Available from: 2014-11-24 Created: 2014-11-21 Last updated: 2016-11-21Bibliographically approved
2. Human neural progenitor cells decrease photoreceptor degeneration, normalize opsin distribution and support synapse structure in cultured porcine retina
Open this publication in new window or tab >>Human neural progenitor cells decrease photoreceptor degeneration, normalize opsin distribution and support synapse structure in cultured porcine retina
2016 (English)In: Brain Research, ISSN 0006-8993, E-ISSN 1872-6240, Vol. 1646, 522-534 p.Article in journal (Refereed) Published
Abstract [en]

Retinal neurodegenerative disorders like retinitis pigmentosa, age-related macular degeneration, diabetic retinopathy and retinal detachment decrease retinal functionality leading to visual impairment. The pathological events are characterized by photoreceptor degeneration, synaptic disassembly, remodeling of postsynaptic neurons and activation of glial cells. Despite intense research, no effective treatment has been found for these disorders. The current study explores the potential of human neural progenitor cell (hNPC) derived factors to slow the degenerative processes in adult porcine retinal explants. Retinas were cultured for 3 days with or without hNPCs as a feeder layer and investigated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), immunohistochemical, western blot and quantitative real time-polymerase chain reaction (qRT-PCR) techniques. TUNEL showed that hNPCs had the capacity to limit photoreceptor cell death. Among cone photoreceptors, hNPC coculture resulted in better maintenance of cone outer segments and reduced opsin mislocalization. Additionally, maintained synaptic structural integrity and preservation of second order calbindin positive horizontal cells was also observed. However, Müller cell gliosis only seemed to be alleviated in terms of reduced Müller cell density. Our observations indicate that at 3 days of coculture, hNPC derived factors had the capacity to protect photoreceptors, maintain synaptic integrity and support horizontal cell survival. Human neural progenitor cell applied treatment modalities may be an effective strategy to help maintain retinal functionality in neurodegenerative pathologies. Whether hNPCs can independently hinder Müller cell gliosis by utilizing higher concentrations or by combination with other pharmacological agents still needs to be determined.

Place, publisher, year, edition, pages
Amsterdam, Netherlands: Elsevier, 2016
Keyword
Photoreceptor degeneration, synapse, opsin, gliosis, neuroprotection
National Category
Neurosciences Ophthalmology
Research subject
Neurology
Identifiers
urn:nbn:se:oru:diva-51995 (URN)10.1016/j.brainres.2016.06.039 (DOI)000381844700059 ()27369448 (PubMedID)2-s2.0-84978831975 (ScopusID)
Note

Funding Agencies:

Faculty of Natural Sciences at Linnaeus University

Faculty of Medical Sciences at Örebro University 

Olle Engkvist Foundation

Ögonfonden

Crown Princess Margaretas Committee for the Blind

Edwin Jordan Foundation

Available from: 2016-09-07 Created: 2016-09-06 Last updated: 2016-11-21Bibliographically approved

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