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Solon Thanos

Publications and source records attributed to Solon Thanos.

35 records · Page 2Linked to original sources
PubMed2004

Microglia-targeted pharmacotherapy in retinal neurodegenerative diseases.

Microglial cells, members of the monocytic lineage, represent the resident immunocompetent cells of the central nervous system including the retina with its peculiarities like a double blood retinal barrier. Microglial cells invade the retina in response to naturally occurring neuronal death during embryonic development and remodelling. Resident microglial cells are extremely sensitive to changes in their microenvironment arising from either traumatic or chronic neurodegeneration, inproper wiring, hereditary diseases or infection and become rapidly activated. In their activated state, the cells undergo drastic morphological changes, upregulate a variety of receptors and secrete soluble factors, which contribute to recognition and phagocytotic cleareance of dying or malfunctioning neurons. In this review, we aim to summarise the current knowledge of microglial involvement in experimentally induced or naturally occurring retinal neurodegenerations with emphasising on mechanisms of microglia activation. Expanding on the mechanisms, we shall discuss on approaches to pharmacologically interfere with the microglial activation and neurophagy. The protagonistic role of these cells in the outcome of certain diseases may help designing microglial targeted treatments with potential benefit for neuronal survival and regeneration in clinically relevant conditions.

Animals↗
PubMed2003

Ink-jet printing for micropattern generation of laminin for neuronal adhesion.

In order to achieve defined adhesion and neurite outgrowth, the growth substrate must be patterned in an appropriate way. We utilised ink-jet printing by means of a piezo-based microdispenser to create defined line patterns of a polymer with typical dimensions of 100 microm width on glass, silicon, gold and carbon substrates. Vinnapas, a co-polymer of vinyl acetate and ethylene, was mixed with the extracellular matrix protein laminin to achieve neuronal adhesion on the surface of the patterns. It could be demonstrated that the laminin entrapped in the polymer lines can be recognised by a specific antibody. Adhesion of embryonic chicken forebrain neurones is following the prepared lines, and identity of adhering cells could be shown by neurofilament staining. These findings open the route for the generation of complex small neuronal arrays and for the electrochemical investigation of the obtained neuronal matrix.

Animals↗
PubMed2003

UV irradiation causes multiple cellular changes in cultured human retinal pigment epithelium cells.

BACKGROUND: The retinal pigment epithelium maybe causally involved in the development and progression of age-related macula degeneration; however, the mechanisms leading to the development of age-related macula degeneration remain largely unknown. The purpose of this study was to examine cellular changes in the retinal pigment epithelium induced by direct irradiation with UV light in culture. METHODS: Retinal pigment epithelium cells from post-mortem human retinas were used to obtain dissociated cultures with cells retaining the ability to differentiate in vitro. These cells were cultured over several days to weeks. The UV radiation (UV-A and UV-B) occurred under sterile conditions with a 100 HBO/mercury bulb attached to a dissecting microscope, delivering co-axial illumination. The time dependence of irradiation effects was analysed using morphometric, immunohistochemical, functional and apoptosis-detecting techniques. RESULTS: Vital and proliferating retinal pigment epithelium cell cultures could be prepared consistently. The cells showed tissue-specific morphologies in vitro for several days to weeks. Pigment epithelium-derived factor was detected in these cells using immunocytochemistry and Western blots. The UV irradiation but not white light resulted in measurable alterations of cell shape and size. The irradiated cells showed partial swelling and shrinkage reminiscent of progressing apoptotic degeneration. TUNEL staining revealed that apoptosis was induced by UV light, but not detectably by white light. The phagocytosis of fluorescent micro-particles diminished after irradiation. These effects were dependent on the duration of irradiation. CONCLUSIONS: Cultures of retinal pigment epithelium are suitable and sensitive models to study cell damage and may contribute to unravelling the pathogenetic mechanisms of retinal degeneration.

Apoptosis↗
PubMed2003

Measurement of nitric oxide production by the lesioned rat retina with a sensitive nitric oxide electrode.

Nitric oxide (NO) plays multiple roles in the nervous system. It is produced as a result of damage or injury of the retina as a part of the central nervous system. Detailed knowledge of the extent and the time course of NO production is of great importance for the understanding of pathological processes and their appropriate medical treatment. Sections of rat retina were stained with antibodies against the three isoforms of NO synthase (NOS) at several time points after a lesion of the optic nerve. No significant changes of NOS expression could be seen at any of the checked time points. For the electrochemical detection of NO production, we modified small platinum electrodes with a NO-sensitive nickel porphyrin by electrochemical polymerisation. Compared to other substances, electrochemically polymerised eugenol was found to be most suitable for protection against interferences. For the measurements, differential pulse amperometry was used. The response to nitric oxide was linear.NO production of adult rat retinas was measured post axotomy after different time points with electrochemical electrodes ex vivo. With non-treated retinas, an NO concentration of approximately 15 microM was measured. NO concentration is elevated after an axotomy reaching its highest value of up to 30 microM 5 days after the lesion. The NO concentration is decreased below the initial value after 9-14 days post axotomy.

Animals↗
PubMed2003

Benzodiazepine and kainate receptor binding sites in the RCS rat retina.

BACKGROUND: The effect of age and photoreceptor degeneration on the kainate subtype of glutamate receptors and on the benzodiazepine-sensitive gamma-aminobutyric acid-A receptors (GABA(A)) in normal and RCS (Royal College of Surgeons) rats were investigated. METHODS: [(3)H]Kainate and [(3)H]flunitrazepam were used as radioligands for kainate and GABA(A)/benzodiazepine()receptors, respectively, using the quantitative receptor autoradiography technique. RESULTS: In both normal and RCS rat retina we observed that [(3)Eta]flunitrazepam and [(3)Eta]kainate binding levels were several times higher in inner plexiform layer (IPL) than in outer plexiform layer (OPL) at all four ages studied (P17, P35, P60 and P180). Age-related changes in receptor binding were observed in normal rat retina: [(3)Eta]flunitrazepam binding showed a significant decrease of 25% between P17 and P60 in IPL,and [(3)Eta]kainate binding showed significant decreases between P17 and P35 in both synaptic layers (71% in IPL and 63% in OPL). Degeneration-related changes in benzodiazepine and kainate receptor binding were observed in RCS rat retina. In IPL, [(3)Eta]flunitrazepam and [(3)Eta]kainate binding levels were higher than in normal retina at P35 (by 24% and 86%, respectively). In OPL, [(3)Eta]flunitrazepam binding was higher in RCS than in normal retina on P35 (74%) and also on P60 (62%). CONCLUSIONS: The results indicate that postnatal changes occur in kainate and benzodiazepine receptor binding sites in OPL and IPL of the rat retina up to 6 months of age. The data also suggest that the receptor binding changes observed in the RCS retina could be a consequence of the primary photoreceptor degeneration.

Animals↗
PubMed2003

Sodium dodecyl sulfate versus acid-labile surfactant gel electrophoresis: comparative proteomic studies on rat retina and mouse brain.

A long-chain derivative of 1,3-dioxolane sodium propyloxy sulfate, with similar denaturing and electrophoretic properties as SDS, and facilitated protein identification following polyacrylamide gel electrophoresis (PAGE) for Coomassie-stained protein bands, has been tested. Comparative acid-labile surfactant/sodium dodecyl sulfate two-dimensional (ALS/SDS 2-D)-PAGE experiments of lower abundant proteins from the proteomes of regenerating rat retina and mouse brain show that peptide recovery for mass spectrometry (MS) mapping is significantly enhanced using ALS leading to more successful database searches. ALS may influence some procedures in proteomic analysis such as the determination of protein content and methods need to be adjusted to that effect. The promising results of the use of ALS in bioanalytics call for detailed physicochemical investigations of surfactant properties.

Acids↗
PubMed2003

Retinal ganglion cells resistant to advanced glaucoma: a postmortem study of human retinas with the carbocyanine dye DiI.

PURPOSE: The present study was conducted to examine whether the morphology of the retinal ganglion cells is altered in advanced glaucoma. Perikaryal, axonal, and dendritic alterations were monitored in glaucoma-resistant retinal ganglion cells by postvitam application of the fluorescent dye DiI. METHODS: The retinas of four amaurotic glaucomatous eyes and four normal eyes enucleated after death were used in this study. The retinas were freed from surrounding tissue, prepared as flatmounts on a nitrocellulose filter, and fixed overnight in 4% paraformaldehyde. The retinal ganglion cells were labeled by introducing crystals of the fluorescent carbocyanine dye DiI, into the optic fiber layer. This dye diffuses along membranes of ganglion cell axons, completely labeling them and their cell bodies and dendrites. Further characterization of the retinas and optic nerves included hematoxylin-eosin and van Gieson histochemical staining as well as immunohistochemistry against glial fibrillary acidic protein. RESULTS: Because of the advanced stage of the disease, the retinas were almost completely depleted of ganglion cells, which had degenerated and therefore could not be stained. The few remaining ganglion cells were considered to be resistant to glaucoma. They showed drastic morphologic alterations, such as abnormal axonal beading, the cell bodies were normal in size but had irregular silhouettes or swellings, and there were fewer dendritic bifurcations. The size of the dendritic trees was smaller, implicating pruning of smaller dendritic branches. Glial cells were also detected immunocytochemically indicating their involvement in the pathologic course of glaucoma. CONCLUSIONS: The data suggest that the few ganglion cells that survive the elevated intraocular pressure associated with loss of visual function display morphologic changes that are manifested both on the cell body and on their intraretinal processes, including axons and dendrites.

Adult↗
PubMed2003

Regeneration of ganglion cell axons into a peripheral nerve graft alters retinal expression of glial markers and decreases vulnerability to re-axotomy.

PURPOSE: To compare the effect of cutting the optic nerve versus replacing the cut optic nerve with a peripheral nerve (PN) graft on retinal glial markers, and to determine whether the PN graft can stabilize regenerating retinal ganglion cells (RGCs), thus preventing their death following re-axotomy. METHODS: Retinas harvested after ganglion cell regeneration into a sciatic nerve graft were compared to untreated control retinas and retinas obtained following optic nerve axotomy. Glial-specific proteins such as glial fibrillary acidic protein (GFAP), Bcl-2 and complement-3 receptor (Ox-42) were examined using immunohistochemistry. Ganglion cells that survived the second axotomy were quantified on retinal flat mounts by retrograde labeling from the graft. RESULTS: GFAP expression in astrocytes and Muller cells was elevated in axotomized retinas when compared to controls, and an additional up-regulation in Muller cells was found in retinas following ganglion cell regeneration. Increased GFAP expression in retinas containing regenerated neurons was accompanied by increased Bcl-2 expression with latter being confined to Muller cells. Moreover, re-axotomy of the regenerated axons within the graft did not result in significant retrograde degeneration of RGCs within 28 days. CONCLUSIONS: The data suggest that the graft stabilizes the regenerating RGCs to an extent reminiscent of peripheral neurons, a process that may involve the interaction between neuronal and glial elements.

Animals↗
PubMed2002

Reciprocal transmission of herpes simplex virus type 1 (HSV-1) between corneal epithelium and trigeminal neurites in an embryonic chick organ culture.

Reciprocal transmission between epithelia and sensory neurons of the peripheral nervous system is a crucial step in the life cycle of herpes simplex virus type 1 (HSV-1) and related alphaherpesviruses. In searching for an easy-to-perform and generally applicable experimental approach that enables the direct analysis of virus transfer between primary epithelial cells and sensory neurites, we investigated the spread of HSV-1 in a dual-chamber organ model comprising chick embryonic corneal epithelia and trigeminal sensory neurons. Embryonic chick corneal and trigeminal tissues were found to be permissive for productive infection with HSV-1. Our data show that HSV-1 efficiently enters neurites re-innervating the cornea and reaches the ganglion explant by retrograde axonal transport, with the first antigen-positive cells being detectable approximately 12 h postinfection. After direct infection of trigeminal tissues, the virus is transported by anterograde axonal transport to the corneal epithelium, causing a visible cytopathic effect approximately 48 h postinfection. These results suggest that the organ model presented in this study holds particular promise for the direct observation and molecular analysis of herpes simplex virus spread between primary epithelia and sensory neurons and that it may be an attractive alternative to current experimental approaches based on laboratory animals or human fetal tissues.

Animals↗
PubMed2002

In vivo FM: using conventional fluorescence microscopy to monitor retinal neuronal death in vivo.

Post-traumatic death of mature retinal neurons occurs in glaucoma and after optic nerve injury. The death is a dynamic process that can be fully analyzed with methods that monitor changes over time. We have coupled the development of retrogradely transportable fluorescent dyes with modification of conventional epifluorescence microscopy to manipulate and visualize rat retinal neurons in vivo. The method is a relatively new concept and has potential for the monitoring of retinal conditions, such as glaucoma or optic nerve transection, and for evaluation of neuroprotective strategies in the near future.

Animals↗
PubMed2002

Detection of early neuron degeneration and accompanying microglial responses in the retina of a rat model of glaucoma.

PURPOSE: To characterize the early reaction of retinal ganglion cells (RGCs) in a rat model of glaucoma using in vivo imaging and to examine the involvement of retinal microglia in glaucomatous neuropathy. METHODS: Glaucoma was induced in adult female Sprague-Dawley rats by cauterizing two episcleral veins, which resulted in a 1.6-fold increase in intraocular pressure (IOP). Retinal ganglion cells were retrogradely labeled with the fluorescent dye, 4-[didecylaminostyryl]-N-methyl-pyridinium-iodide (4-Di-10ASP) and monitored in vivo after elevation of IOP using fluorescence microscopy imaging. The number of RGCs was quantified on retinal flatmounts. Dying RGCs were surrounded by activated microglia that became visible after taking up the fluorescent debris. Immunocytochemistry was conducted to characterize further the ganglion cells and microglia. RESULTS: Cauterizing two of the four episcleral veins resulted in a consistent increase of IOP to 25.3 +/- 2.0 mm Hg, as measured with a handheld tonometer. IOP remained high for at least 3 months in glaucomatous eyes. The earliest sign of RGC death was detected in anesthetized animals 20 hours after induction of glaucoma. RGCs continued to decrease in number over time, with 40% of RGCs having degenerated after 2.5 months. Fundoscopic examination of the optic nerve head revealed cupping 2 months after induction of glaucoma. In addition, microglia were detected on retinal flatmounts as early as 72 hours after induction. Activated microglia and RGCs were also identified immunocytochemically, with an antibody against ionized calcium-binding adaptor molecule (Iba)-1 and an antibody specific to the 200-kDa subunit of the neurofilament protein, respectively. CONCLUSIONS: Occlusion of episcleral veins is a reproducible method that mimics human glaucoma, with chronically elevated IOP-induced RGC loss. This study shows that in vivo imaging permits the detection of ganglion cells in the living animal in the early stages of the disease and highlights the importance of in vivo imaging in understanding ophthalmic disorders such as glaucoma. Secondly, activation of intraretinal microglia coincides with degeneration of RGCs in glaucoma.

Animals↗
PubMed2001

Microglia promote glioma migration.

Diffuse astrocytic gliomas extensively infiltrate brain tissue and contain numerous microglial cells, but it is unknown whether these two characteristic features are pathogenetically related. We therefore studied the effects of murine microglial cells on motility of GL261 mouse glioma cells using Boyden chamber assays. In the presence of microglia, glioma cell migration occurred earlier, and after 48 h it was threefold higher as compared to incubations without microglia. This effect was mediated by substances released from microglia, because similar effects were observed by microglia-conditioned medium, and it was specific to microglia, because oligodendroglia and endothelial cells only weakly stimulated glioma cell migration. Microglia activating substances (GM-CSF, LPS) led to a further increase of motility. These data support the notion that microglia accumulation in diffuse glial tumors does not merely represent a nonspecific reaction to tissue injury, but reflects participation of these cells in supporting and promoting the invasive phenotype of astrocytoma cells.

Animals↗
PubMed1992

Adult Retinofugal Axons Regenerating Through Peripheral Nerve Grafts Can Restore the Light-induced Pupilloconstriction Reflex.

To investigate the ability of mature regenerating retinal axons to form functional connections within central targets, severed axons were guided into the primary visual centres which subserve the pupillary constriction reflex in response to light. The ocular stump of the transected optic nerve of adult rats was connected by means of an autologous peripheral nerve graft with the pretectal region which contains the relay nucleus of pupillary constriction, the olivary pretectal nucleus. This nucleus is efferently connected with preganglionic neurons in the oculomotor nuclear complex which innervates parasympathetically the muscle constrictors of the iris. Six to sixteen weeks after optic nerve transection and peripheral nerve transplantation, brisk responses were observed in the pupils upon illumination of the transplanted eye. Recovery of the pupil responses indicated that retinal neurons used the peripheral nerve 'bridge' to access the pretectum, in which they established synaptic contacts in sufficient density and with appropriate specificity to reconstitute the function of the traumatically interrupted neuronal circuitry.

Journal Article↗
PubMed1991

The Relationship of Microglial Cells to Dying Neurons During Natural Neuronal Cell Death and Axotomy-induced Degeneration of the Rat Retina.

The interactions between dying neurons and phagocytotic cells within the developing and injured retina remain controversial. The present work explored the role of microglia and investigated whether so-called resident microglial cells are permanently responsible for removing cell debris whenever it is produced. As a first goal, I characterized some quantitative and morphometric features of the small ipsilateral retinocollicular projections and analysed the permanent function of phagocytosing microglia with these projections as a paradigm. To achieve this, I combined the fluorescent dyes Dil and 4Di-10ASP, both of which persist in the labelled ganglion cells after injection into the superior colliculus (SC), and retrograde labelling. After neuronal degradation, the dyes accompany the degradation products, become interiorized and then persist within the phagocytosing microglia. Consequently, early labelling of microglial cells can be assessed by injecting one dye into the SC during the first postnatal day of life, that is, prior to advanced natural neuronal cell death. Labelling of the remaining ipsilaterally projecting neurons with the second dye following intraorbital axotomy in adulthood and during subsequent neuronal death would therefore result in double labelling of some microglial cells, if these were involved in phagocytosis during both the natural and the induced phases of neuronal degradation. The ganglion cells which survived natural neuronal cell death remained fluorescent for 3 months after labelling with either dye on the day of the animal's birth, indicating that both fluorescent probes persisted within neurons. Quantitatively, 1770+/-220 ganglion cells/mm2 were labelled within the contralateral retina and a total population of 1442+/-120 cells/retina were observed within the periphery of the inferior/temporal quadrant of the ipsilateral retina. A smaller, ipsilateral projection of 150+/-24 cells/retina was uniformly scattered throughout the rest of the retinal surface. Transient projections of ganglion cells to either the contralateral or the ipsilateral colliculi and death of labelled ganglion cells during the first postnatal days resulted in labelling of 210+/-36 microglial cells/mm2 within the contralateral retina and a total number of 800+/-120 cells/retina within the inferior/temporal and 200+/-22 cells/retina within the rest of the retina. These labelled microglial cells were observed in adulthood and indicated that after taking away the neuronal cell debris they persisted within the retinal tissue. The small number of prelabelled ganglion cells which formed persistent ipsilateral projections until adulthood were axotomized by transecting the optic nerve, and resulted in additional labelling of microglial cells with the second fluorescent dye as well. Double-labelled microglia were observed within the inferior/temporal quadrant (3500+/-240 cells/retina) and to a lesser extent (340+/-40 cells/retina) scattered over the entire retinal surface. The chronotopological sequence of microglial labelling paralleled that of ganglion cell degeneration. Injection of protease inhibitors into the vitreous body during optic nerve transection retarded retrograde glial cell degeneration, probably by blocking microglial proteases. The results directly proved that the same microglial cells which remove neuronal cell debris in the postnatal retina were reactivated later in life to proteolytically degrade and then phagocytose neurons which had altered because of the axotomy.

Journal Article↗
PubMed1989

Survival and Axonal Elongation of Adult Rat Retinal Ganglion Cells.

A peripheral nerve exudate, collected in situ from the proximal end of a severed rat sciatic nerve, can induce substantial axonal elongation from ganglion cells when tested on explanted adult rat retinae. The responsive cells are identified on the basis of their Thy 1.1 immunostaining properties. Similar outgrowth can be obtained from explants when the culture medium is supplemented with brain-derived neurotrophic factor (BDNF). In addition, both BDNF and the sciatic nerve exudate can prevent ganglion cell degeneration as shown by the retrograde transport of a fluorescent dye that had been applied to the superior colliculus prior to explantation. The results demonstrate that soluble components, released by lesioned peripheral nerves, can effect adult retinal ganglion cells in a way that is reminiscent of that which has been described in vivo using sciatic nerve grafts after sectioning of the optic nerve. The molecular nature of these components is discussed.

Journal Article↗
PubMedDate not supplied

Multifocal electroretinography changes in the macula at high altitude: a report of three cases.

BACKGROUND: To evaluate the short- and long-term effects of high-altitude hypobaric hypoxia on macula morphology and function during ascents, acclimatizations, and descents between 500 m and 5,650 m, macula function was evaluated in three healthy climbers of a trekking expedition. METHODS: Macula physiology was tested with multifocal electroretinography (MF ERG), near and farvisual acuity, and Amsler grid tests. Macula morphology was tested with optical coherence tomography (OCT) and with stereoscopic fundoscopy obtained 1 week before ascent, as well as 1 week and 2 weeks after high-altitude exposure. The following physiological parameters indicative of acclimatization were compared daily during the expedition at altitudes between 500 m and 5,050 m: hemoglobin, oxygen saturation, resting heart rate, retinal findings, and the Lake Louise score of acclimatization. RESULTS: The central macula MF ERG responses were significantly reduced 1 week after high-altitude exposure, and had recovered by the follow-up examination performed during the following week. Near visual acuity and Amsler grid tests remained unaffected at both follow-up examinations. No significant changes were found in the follow-up OCT and daily fundoscopy examinations in all three well-acclimatized climbers. CONCLUSIONS: High-altitude hypobaric hypoxia affects the function of the highly sensitive macula region. This suggests that the exposure of persons with macula diseases such as age-related macula degeneration, tapetoretinal degeneration, or diabetic retinopathy to high altitudes may influence the disease progression. For this reason, this population should avoid prolonged and unnecessary high-altitude exposure without proper acclimatization.

Acclimatization↗