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Milos Pekny

Publications and source records attributed to Milos Pekny.

17 recordsLinked to original sources
PubMed2007

Cytoskeleton and vesicle mobility in astrocytes.

Exocytotic vesicles in astrocytes are increasingly viewed as essential in astrocyte-to-neuron communication in the brain. In neurons and excitable secretory cells, delivery of vesicles to the plasma membrane for exocytosis involves an interaction with the cytoskeleton, in particular microtubules and actin filaments. Whether cytoskeletal elements affect vesicle mobility in astrocytes is unknown. We labeled single vesicles with fluorescent atrial natriuretic peptide and monitored their mobility in rat astrocytes with depolymerized microtubules, actin, and intermediate filaments and in mouse astrocytes deficient in the intermediate filament proteins glial fibrillary acidic protein and vimentin. In astrocytes, as in neurons, microtubules participated in directional vesicle mobility, and actin filaments played an important role in this process. Depolymerization of intermediate filaments strongly affected vesicle trafficking and in their absence the fraction of vesicles with directional mobility was reduced.

Actins↗
PubMed2006

Redefining the concept of reactive astrocytes as cells that remain within their unique domains upon reaction to injury.

Reactive astrocytes in neurotrauma, stroke, or neurodegeneration are thought to undergo cellular hypertrophy, based on their morphological appearance revealed by immunohistochemical detection of glial fibrillary acidic protein, vimentin, or nestin, all of them forming intermediate filaments, a part of the cytoskeleton. Here, we used a recently established dye-filling method to reveal the full three-dimensional shape of astrocytes assessing the morphology of reactive astrocytes in two neurotrauma models. Both in the denervated hippocampal region and the lesioned cerebral cortex, reactive astrocytes increased the thickness of their main cellular processes but did not extend to occupy a greater volume of tissue than nonreactive astrocytes. Despite this hypertrophy of glial fibrillary acidic protein-containing cellular processes, interdigitation between adjacent hippocampal astrocytes remained minimal. This work helps to redefine the century-old concept of hypertrophy of reactive astrocytes.

Animals↗
PubMed2006

Complement: a novel factor in basal and ischemia-induced neurogenesis.

Through its involvement in inflammation, opsonization, and cytolysis, the complement protects against infectious agents. Although most of the complement proteins are synthesized in the central nervous system (CNS), the role of the complement system in the normal or ischemic CNS remains unclear. Here we demonstrate for the first time that neural progenitor cells and immature neurons express receptors for complement fragments C3a and C5a (C3a receptor (C3aR) and C5a receptor). Mice that are deficient in complement factor C3 (C3(-/-)) lack C3a and are unable to generate C5a through proteolytic cleavage of C5 by C5-convertase. Intriguingly, basal neurogenesis is decreased both in C3(-/-) mice and in mice lacking C3aR or mice treated with a C3aR antagonist. The C3(-/-) mice had impaired ischemia-induced neurogenesis both in the subventricular zone, the main source of neural progenitor cells in adult brain, and in the ischemic region, despite normal proliferative response and larger infarct volumes. Thus, in the adult mammalian CNS, complement activation products promote both basal and ischemia-induced neurogenesis.

Animals↗
PubMed2005

Re-establishing the regenerative potential of central nervous system axons in postnatal mice.

At a certain point in development, axons in the mammalian central nervous system lose their ability to regenerate after injury. Using the optic nerve model, we show that this growth failure coincides with two developmental events: the loss of Bcl-2 expression by neurons and the maturation of astrocytes. Before postnatal day 4, when astrocytes are immature, overexpression of Bcl-2 alone supported robust and rapid optic nerve regeneration over long distances, leading to innervation of brain targets by day 4 in mice. As astrocytes matured after postnatal day 4, axonal regeneration was inhibited in mice overexpressing Bcl-2. Concurrent induction of Bcl-2 and attenuation of reactive gliosis reversed the failure of CNS axonal re-elongation in postnatal mice and led to rapid axonal regeneration over long distances and reinnervation of the brain targets by a majority of severed optic nerve fibers up to 2 weeks of age. These results suggest that an early postnatal downregulation of Bcl-2 and post-traumatic reactive gliosis are two important elements of axon regenerative failure in the CNS.

Animals↗
PubMed2005

Astrocyte activation and reactive gliosis.

Astrocytes become activated (reactive) in response to many CNS pathologies, such as stroke, trauma, growth of a tumor, or neurodegenerative disease. The process of astrocyte activation remains rather enigmatic and results in so-called "reactive gliosis," a reaction with specific structural and functional characteristics. In stroke or in CNS trauma, the lesion itself, the ischemic environment, disrupted blood-brain barrier, the inflammatory response, as well as in metabolic, excitotoxic, and in some cases oxidative crises--all affect the extent and quality of reactive gliosis. The fact that astrocytes function as a syncytium of interconnected cells both in health and in disease, rather than as individual cells, adds yet another dimension to this picture. This review focuses on several aspects of astrocyte activation and reactive gliosis and discusses its possible roles in the CNS trauma and ischemia. Particular emphasis is placed on the lessons learnt from mouse genetic models in which the absence of intermediate filament proteins in astrocytes leads to attenuation of reactive gliosis with distinct pathophysiological and clinical consequences.

Animals↗
PubMed2004

Under stress, the absence of intermediate filaments from Müller cells in the retina has structural and functional consequences.

In epithelial and muscle cells, intermediate filaments (IFs) are important for resistance to mechanical stress. The aim of this study was to elucidate whether IFs are also important for providing resistance to mechanical stress in the Müller cells of the retina and whether this has any pathophysiological consequences. We used mice deficient in IF proteins glial fibrillary acidic protein and/or vimentin (GFAP(-/-), Vim(-/-) and GFAP(-/-) Vim(-/-)), and stress on the retina was applied by excision of the eyes immediately post mortem (compared with in situ fixation) or by inducing a neovascular response to oxygen-induced retinopathy (OIR). The structure of unchallenged retinas was normal, but mechanical stress caused local separation of the inner limiting membrane (ILM) and adjacent tissue from the rest of the retina in GFAP(-/-) Vim(-/-) mice and, to a lesser extent, in Vim(-/-) mice. This detachment occurred within the endfeet of Müller cells, structures normally rich in IFs but IF-free in GFAP(-/-) Vim(-/-) mice. Hypoxia-induced neovascularization was comparable in all groups of mice with respect to the retinal surface area occupied by new vessels. However, the vessels traversed the ILM and penetrated the vitreous body less frequently than in wild-type retinas (31-55% in Vim(-/-), 66-79% in GFAP(-/-) Vim(-/-)). We conclude that IFs are important for maintaining the mechanical integrity of Müller-cell endfeet and the inner retinal layers under a mechanical challenge. Furthermore, the absence of IFs in Müller cells leads to an abnormal response of the vascular system to ischemia, specifically decreased ability of newly formed blood vessels to traverse the ILM.

Animals↗
PubMed2004

Absence of glial fibrillary acidic protein and vimentin prevents hypertrophy of astrocytic processes and improves post-traumatic regeneration.

The regenerative capacity of the CNS is extremely limited. The reason for this is unclear, but glial cell involvement has been suspected, and oligodendrocytes have been implicated as inhibitors of neuroregeneration (Chen et al., 2000, GrandPre et al., 2000; Fournier et al., 2001). The role of astrocytes in this process was proposed but remains incompletely understood (Silver and Miller, 2004). Astrocyte activation (reactive gliosis) accompanies neurotrauma, stroke, neurodegenerative diseases, or tumors. Two prominent hallmarks of reactive gliosis are hypertrophy of astrocytic processes and upregulation of intermediate filaments. Using the entorhinal cortex lesion model in mice, we found that reactive astrocytes devoid of the intermediate filament proteins glial fibrillary acidic protein and vimentin (GFAP-/-Vim-/-), and consequently lacking intermediate filaments (Colucci-Guyon et al., 1994; Pekny et al., 1995; Eliasson et al., 1999), showed only a limited hypertrophy of cell processes. Instead, many processes were shorter and not straight, albeit the volume of neuropil reached by a single astrocyte was the same as in wild-type mice. This was accompanied by remarkable synaptic regeneration in the hippocampus. On a molecular level, GFAP-/-Vim-/- reactive astrocytes could not upregulate endothelin B receptors, suggesting that the upregulation is intermediate filament dependent. These findings show a novel role for intermediate filaments in astrocytes and implicate reactive astrocytes as potent inhibitors of neuroregeneration.

Animals↗
PubMed2004

Astrocyte intermediate filaments in CNS pathologies and regeneration.

Astroglial cells are the most abundant cells in the mammalian central nervous system (CNS), yet our knowledge about their function in health and disease has been limited. This review focuses on the recent work addressing the function of intermediate filaments in astroglial cells under severe mechanical or osmotic stress, in hypoxia, and in brain and spinal cord injury. Recent data show that when astrocyte intermediate filaments are genetically ablated in mice, reactive gliosis is attenuated and the course of several CNS pathologies is altered, while the signs of CNS regeneration become more prominent. GFAP is the principal astrocyte intermediate filament protein and dominant mutations in the GFAP gene have been shown to lead to Alexander disease, a fatal neurodegenerative condition in humans.

Alexander Disease↗
PubMed2004

Increased cell proliferation and neurogenesis in the hippocampal dentate gyrus of old GFAP(-/-)Vim(-/-) mice.

In response to central nervous system (CNS) injury, and more discretely so also during aging, astrocytes become reactive and increase their expression of the intermediate filament proteins glial fibrillary acidic protein (GFAP) and vimentin. Studies of mice deficient in astrocytic intermediate filaments have provided insights into the function of reactive gliosis. Recently we demonstrated robust integration of retinal transplants (1) and increased posttraumatic synaptic regeneration (2) in GFAP(-/-)Vim(-/-) mice, suggesting that modulation of astrocyte activity affects the permissiveness of the CNS environment for regeneration. Neurogenesis in the adult mammalian CNS is restricted to essentially two regions, the hippocampus and the subventricular zone. Here, we assessed neurogenesis in the hippocampus of 18-month-old GFAP(-/-)Vim(-/-) mice. In the granular layer of the dentate gyrus, cell proliferation/survival was 34% higher and neurogenesis 36% higher in GFAP(-/-)Vim(-/-) mice than in wildtype controls. These findings suggest that the adult hippocampal neurogenesis in healthy old mice can be increased by modulating astrocyte reactivity.

Aging↗
PubMed2004

Bfsp2 mutation found in mouse 129 strains causes the loss of CP49 and induces vimentin-dependent changes in the lens fibre cell cytoskeleton.

Here we report the first natural mutation in the mouse Bfsp2 gene. Characterisation of mouse Bfsp2 in the 129X1/SvJ revealed a mutation that deleted the acceptor site of exon 2. This results in exon 1 being erroneously spliced to exon 3 causing a frameshift in the reading frame and the introduction of a stop codon at position 2 of exon 3 in the Bfsp2 transcript. RT-PCR studies of lens RNA isolated from 129S1/SvImJ, 129S2/SvPas and 129S4/SvJae strains confirmed the presence of this mutation in these diverse 129 strains and similar mutations were found in both CBA and 101 strains, but not in C3H or C57BL/6J mouse strains. This mutation is predicted to result in a severely truncated protein product called CP49, comprising essentially only exon 1, but polyclonal antibodies to CP49 failed to detect either full length or fragments of CP49 in extracts made from either 129S1/SvImJ or 129S4/SvJae suggesting that these 129 strains lack CP49 protein. Like the knockout of Bfsp2 reported recently, filensin protein levels and its proteolytic processing were altered also in the 129S1/SvImJ and 129S4/SvJae strains compared to C57BL/6J. Electron microscopy of the lens cytoskeleton from 129S2/SvPas revealed similar morphological changes in the cytoskeleton as compared to the CP49 knockout, with beaded and intermediate filaments being apparently replaced by poorly defined filament-like material. Vimentin was a key component of this residual material as shown by immunoelectron microscopy and by the generation of a CP49/vimentin double knockout mouse. This report of a natural mutation in Bfsp2 in the 129 and other mouse strains also has important implications for lens studies that have used the 129X1/SvJ strain in knockout strategies.

Animals↗
PubMed2004

Bfsp2 mutation found in mouse 129 strains causes the loss of CP49' and induces vimentin-dependent changes in the lens fibre cell cytoskeleton.

Here we report the first natural mutation in the mouse Bfsp2 gene. Characterisation of mouse Bfsp2 in the 129X1/SvJ revealed a mutation that deleted the acceptor site of exon 2. This results in exon 1 being erroneously spliced to exon 3 causing a frameshift in the reading frame and the introduction of a stop codon at position 2 of exon 3 in the Bfsp2 transcript. RT-PCR studies of lens RNA isolated from 129S1/SvImJ, 129S2/SvPas and 129S4/SvJae strains confirmed the presence of this mutation in these diverse 129 strains and similar mutations were found in both CBA and 101 strains, but not in C3H or C57BL/6J mouse strains. This mutation is predicted to result in a severely truncated protein product called CP49, comprising essentially only exon 1, but polyclonal antibodies to CP49 failed to detect either full length or fragments of CP49 in extracts made from either 129S1/SvImJ or 129S4/SvJae suggesting that these 129 strains lack CP49 protein. Like the knockout of Bfsp2 reported recently, filensin protein levels and its proteolytic processing were altered also in the 129S1/SvImJ and 129S4/SvJae strains compared to C57BL/6J. Electron microscopy of the lens cytoskeleton from 129S2/SvPas revealed similar morphological changes in the cytoskeleton as compared to the CP49 knockout, with beaded and intermediate filaments being apparently replaced by poorly defined filament-like material. Vimentin was a key component of this residual material as shown by immunoelectron microscopy and by the generation of a CP49/vimentin double knockout mouse. This report of a natural mutation in Bfsp2 in the 129 and other mouse strains also has important implications for lens studies that have used the 129X1/SvJ strain in knockout strategies.

Animals↗
PubMed2003

Loss of GFAP expression in high-grade astrocytomas does not contribute to tumor development or progression.

In astrocytic neoplasms, the number of cells expressing glial fibrillary acidic protein (GFAP) is inversely proportional to the extent of anaplasia. The loss of GFAP expression, the principal marker of astroglial cells, in these tumors has been proposed to constitute a step in their development and progression. To test this hypothesis, we crossed p53-negative (p53(-/-)) mice, which frequently develop astrocytomas after intrauterine exposure to ethylnitrosourea, with GFAP-negative (GFAP(-/-)) mice or GFAP(+/+) controls. Brain tumors of glial origin were found in 12 of 35 GFAP(+/+) p53(-/-) mice (34%) and in 11 of 27 GFAP(-/-) p53(-/-) mice (41%). The two groups did not differ in the age at which tumors were detected or in tumor histology or progression. Thus, the loss of GFAP expression frequently seen in high-grade astrocytomas does not constitute a step in tumor development. Rather, it may represent the undifferentiated state of these cells.

Animals↗
PubMed2003

Neuron-specific ablation of PDGF-B is compatible with normal central nervous system development and astroglial response to injury.

Members of the PDGF family have multiple roles during embryogenesis and in a variety of pathological situations in the adult. One of the major sites of PDGF-B expression in adult mammals are postmitotic CNS neurons. Combined with reported neurotrophic and neuroprotective effects of exogenously administered PDGFs, this has led to the speculation that PDGF-B may have a role in CNS development, in maintenance, or in response to CNS injury. To test these hypotheses, we developed mice in which PDGF-B was ablated genetically in postmitotic neurons at sites where PDGF-B is normally expressed. We found that these mice develop to adulthood without apparent defects. We demonstrate PDGF-B expression in the postnatal mouse hippocampus and forebrain cortex. We show that neuron-specific knockout of PDGF-B does not influence the astroglial and angiogenic responses to injury in the hippocampus or forebrain cortex. We conclude that the role of neuron-derived PDGF-B remains obscure. A role for neuron-derived PDGF-B, if existing, might be redundant with other CNS growth factors. Alternatively, other and more specific analyses of CNS functions in the normal and injured states will be required to demonstrate such a role.

Animals↗
PubMed2003

Robust neural integration from retinal transplants in mice deficient in GFAP and vimentin.

With recent progress in neuroscience and stem-cell research, neural transplantation has emerged as a promising therapy for treating CNS diseases. The success of transplantation has been limited, however, by the restricted ability of neural implants to survive and establish neuronal connections with the host. Little is known about the mechanisms responsible for this failure. Neural implantation triggers reactive gliosis, a process accompanied by upregulation of intermediate filaments in astrocytes and formation of astroglial scar tissue. Here we show that the retinas of adult mice deficient in glial fibrillary acidic protein and vimentin, and consequently lacking intermediate filaments in reactive astrocytes and Müller cells, provide a permissive environment for grafted neurons to migrate and extend neurites. The transplanted cells integrated robustly into the host retina with distinct neuronal identity and appropriate neuronal projections. Our results indicate an essential role for reactive astroglial cells in preventing neural graft integration after transplantation.

Animals↗
PubMed2002

Mice lacking the adenosine A1 receptor are anxious and aggressive, but are normal learners with reduced muscle strength and survival rate.

Behavioural assessment of mice lacking adenosine A1 receptors (A1Rs) showed reduced activity in some phases of the light-dark cycle, reduced exploratory behaviour in the open-field and in the hole-board, increased anxiety in the plus maze and dark-light box and increased aggressiveness in the resident-intruder test. No differences were found in spatial reference and working memory in several Morris water maze tasks. Both mutant mice had reduced muscle strength and survival rate. These results confirm the involvement of adenosine in motor activity, exploratory behaviour, anxiety and aggressiveness. A1Rs also appear to play a critical role in ageing-related deterioration.

Adenosine↗