Biomedical subjects

Inez Vincent

Publications and source records attributed to Inez Vincent.

14 recordsLinked to original sources

PubMed2006

Neuronal cell cycle re-entry mediates Alzheimer disease-type changes.

Evidence showing the ectopic re-expression of cell cycle-related proteins in specific vulnerable neuronal populations in Alzheimer disease led us to formulate the hypothesis that neurodegeneration, like cancer, is a disease of inappropriate cell cycle control. To test this notion, we used adenoviral-mediated expression of c-myc and ras oncogenes to drive postmitotic primary cortical neurons into the cell cycle. Cell cycle re-entry in neurons was associated with increased DNA content, as determined using BrdU and DAPI, and the re-expression of cyclin B1, a marker for the G2/M phase of the cell cycle. Importantly, we also found that cell cycle re-entry in primary neurons leads to tau phosphorylation and conformational changes similar to that seen in Alzheimer disease. This study establishes that the cell cycle can be instigated in normally quiescent neuronal cells and results in a phenotype that shares features of degenerative neurons in Alzheimer disease. As such, our neuronal cell model may be extremely valuable for the development of novel therapeutic strategies.

Alzheimer Disease↗

PubMed2006

p35/p25 is not essential for tau and cytoskeletal pathology or neuronal loss in Niemann-Pick type C disease.

Hyperactivation of the cyclin-dependent kinase 5 (cdk5), triggered by proteolytic conversion of its neuronal activator, p35, to a more potent byproduct, p25, has been implicated in Alzheimer's disease (AD), amyotrophic lateral sclerosis, and Niemann-Pick type C disease (NPC). This mechanism is thought to lead to the development of neuropathological hallmarks, i.e., hyperphosphorylated cytoskeletal proteins, neuronal inclusions, and neurodegeneration, that are common to all three diseases. This pathological ensemble is recapitulated in a single model, the npc-1 (npc(-/-)) mutant mouse. Previously, we showed that pharmacological cdk inhibitors dramatically reduced hyperphosphorylation, lesion formation, and locomotor defects in npc(-/-) mice, suggesting that cdk activity is required for NPC pathogenesis. Here, we used genetic ablation of the p35 gene to examine the specific involvement of p35, p25, and hence cdk5 activation in NPC neuropathogenesis. We found that lack of p35/p25 does not slow the onset or progression or improve the neuropathology of NPC. Our results provide direct evidence that p35/p25-mediated cdk5 deregulation is not essential for NPC pathology and suggest that similar pathology in AD may also be cdk5 independent.

Age Factors↗

PubMed2006

Calpain-regulated p35/cdk5 plays a central role in dopaminergic neuron death through modulation of the transcription factor myocyte enhancer factor 2.

The mechanisms underlying dopamine neuron loss in Parkinson's disease (PD) are not clearly defined. Here, we delineate a pathway by which dopaminergic loss induced by 1-methyl-4-phenyl 1,2,3,6 tetrahydropyridine (MPTP) is controlled in vivo. We reported previously that calpains play a central required role in dopamine loss after MPTP treatment. Here, we provide evidence that the downstream effector pathway of calpains is through cyclin-dependent kinase 5 (cdk5)-mediated modulation of the transcription factor myocyte enhancer factor 2 (MEF2). We show that MPTP-induced conversion of the cdk5 activator p35 to a pathogenic p25 form is dependent on calpain activity in vivo. In addition, p35 deficiency attenuates MPTP-induced dopamine neuron loss and behavioral outcome. Moreover, MEF2 is phosphorylated on Ser444, an inactivating site, after MPTP treatment. This phosphorylation is dependent on both calpain and p35 activity, consistent with the model that calpain-mediated activation of cdk5 results in phosphorylation of MEF2 in vivo. Finally, we provide evidence that MEF2 is critical for dopaminergic loss because "cdk5 phosphorylation site mutant" of MEF2D provides neuroprotection in an MPTP mouse model of PD. Together, these data indicate that calpain-p35-p25/cdk5-mediated inactivation of MEF2 plays a critical role in dopaminergic loss in vivo.

Animals↗

PubMed2005

Differential roles of nuclear and cytoplasmic cyclin-dependent kinase 5 in apoptotic and excitotoxic neuronal death.

Cyclin-dependent kinase 5 (cdk5) is a member of the cyclin-dependent kinase family whose activity is localized mainly to postmitotic neurons attributable to the selective expression of its activating partners p35 and p39. Deregulation of cdk5, as a result of calpain cleavage of p35 to a smaller p25 form, has been suggested to be a central component of neuronal death underlying numerous neurodegenerative diseases. However, the relevance of cdk5 in apoptotic death that relies on the mitochondrial pathway is unknown. Furthermore, evidence that cdk5 can also promote neuronal survival has necessitated a more complex understanding of cdk5 in the control of neuronal fate. Here we explore each of these issues using apoptotic and excitotoxic death models. We find that apoptotic death induced by the DNA-damaging agent camptothecin is associated with early transcription-mediated loss of p35 and with late production of p25 that is dependent on Bax, Apaf1, and caspases. In contrast, during excitotoxic death induced by glutamate, neurons rapidly produce p25 independent of the mitochondrial pathway. Analysis of the localization of p35 and p25 revealed that p35 is mainly cytoplasmic, whereas p25 accumulates selectively in the nucleus. By targeting a dominant-negative cdk5 to either the cytoplasm or nucleus, we show that cdk5 has a death-promoting activity within the nucleus and that this activity is required in excitotoxic death but not apoptotic death. Moreover, we also find that cdk5 contributes to pro-survival signaling selectively within the cytoplasm, and manipulation of this signal can modify death induced by both excitotoxicity and DNA damage.

Animals↗

PubMed2005

Multiple cyclin-dependent kinases signals are critical mediators of ischemia/hypoxic neuronal death in vitro and in vivo.

The mechanisms involving neuronal death after ischemic/hypoxic insult are complex, involving both rapid (excitotoxic) and delayed (apoptotic-like) processes. Recent evidence suggests that cell cycle regulators such as cyclin-dependent kinases are abnormally activated in neuropathological conditions, including stroke. However, the function of this activation is unclear. Here, we provide evidence that inhibition of the cell cycle regulator, Cdk4, and its activator, cyclinD1, plays critical roles in the delayed death component of ischemic/hypoxic stress by regulating the tumor suppressor retinoblastoma protein. In contrast, the excitotoxic component of ischemia/hypoxia is predominately regulated by Cdk5 and its activator p35, components of a cyclin-dependent kinase complex associated with neuronal development. Hence, our data both characterize the functional significance of the cell cycle Cdk4 and neuronal Cdk5 signals as well as define the pathways and circumstances by which they act to control ischemic/hypoxic damage.

Animals↗

PubMed2004

Intracellular accumulation of amyloidogenic fragments of amyloid-beta precursor protein in neurons with Niemann-Pick type C defects is associated with endosomal abnormalities.

Niemann-Pick type C disease (NPC) is characterized by neurodegeneration secondary to impaired cholesterol trafficking and excessive glycosphingolipid storage. Abnormal cholesterol and ganglioside metabolism may influence the generation and aggregation of amyloidogenic fragments (ie, C99 and Abeta) from amyloid-beta precursor protein (APP), crucial factors causing neurodegeneration in Alzheimer's disease. To reveal whether abnormal accumulation and aggregation of APP fragments also occurs in NPC, we studied their expression in cultured cortical neurons treated with U18666A, a compound widely used to induce NPC defects, and also in brain tissues from NPC patients. U18666A treatment resulted in increased intraneuronal levels of C99 and insoluble Abeta42, which were distributed among early and late endosomes, in compartments distinct from where endogenous cholesterol accumulates. Analyses of NPC brains revealed that C99 or other APP C-terminal fragments (APP-CTF), but not Abeta42, accumulated in Purkinje cells, mainly in early endosomes. In contrast, in hippocampal pyramidal neurons, the major accumulated species was Abeta42, in late endosomes. Similar to what has been shown in Alzheimer's disease, cathepsin D, a lysosomal hydrolase, was redistributed to early endosomes in NPC Purkinje cells, where it co-localized with C99/APP-CTF. Our results suggest that endosomal abnormalities related to abnormal lipid trafficking in NPC may contribute to abnormal APP processing and Abeta42/C99/APP-CTF deposition.

Adolescent↗

PubMed2004

Cyclin-dependent kinase inhibitors attenuate protein hyperphosphorylation, cytoskeletal lesion formation, and motor defects in Niemann-Pick Type C mice.

Dysregulation of cyclin-dependent kinases (cdks) and cytoskeletal protein hyperphosphorylation characterizes a subset of human neurodegenerative diseases, including Alzheimer's disease, amyotrophic lateral sclerosis, and Niemann-Pick Type C (NPC). It is thought that these cytoskeletal changes lead eventually to development of hallmark cytoskeletal lesions such as neurofibrillary tangles and axonal spheroids. Although many studies support an involvement of cdks in these neurodegenerative cascades, it is not known whether cdk activity is essential. The naturally occurring npc-1 mutant mouse mimics human NPC, in displaying activation of cdk5, mitotic cdc2, and cdk4, with concomitant cytoskeletal pathology and neurodegeneration. We availed of this model and specific pharmacological inhibitors of cdk activity, to determine whether cdks are necessary for NPC neuropathology. The inhibitors were infused intracerebroventricularly for a 2-week period, initiated at a pathologically incipient stage. While an inactive stereoisomer, iso-olomoucine, was ineffective, two potent inhibitors, roscovitine and olomoucine, attenuated significantly the hyperphosphorylation of neurofilament, tau, and mitotic proteins, reduced the number of spheroids, modulated Purkinje neuron death, and ameliorated motor defects in npc mice. These results suggest that cdk activity is required for neuropathology and subsequent motor impairment in NPC. Studies aimed at knocking down individual cdks in these mice will help identify the specific cdk(s) that are essential, and delineate their precise roles in the neurodegenerative process.

Animals↗

PubMed2003

Decreased cyclin-dependent kinase 5 (cdk5) activity is accompanied by redistribution of cdk5 and cytoskeletal proteins and increased cytoskeletal protein phosphorylation in p35 null mice.

Cdk5/p35 has been implicated in cytoskeletal protein phosphorylation in normal brain and in many human neurodegenerative disorders. Yet, mouse models of cdk5/p35 hyperactivity have not yielded corresponding changes in cytoskeletal protein phosphorylation. To elucidate the relationship between p35, cdk5, and the neuronal cytoskeleton, we deleted the p35 gene in mice having a pure C57BL/6 background. We found that p35 deficiency leads to a 38% reduction of cdk5 activity in adult brain. In addition, loss of p35 causes an anterograde redistribution of cdk5 toward peripheral neuronal processes. The unusual presence of nonphosphorylated neurofilament (NF) in aberrant axon fascicles and the relocation of tau and MAP2B from cell bodies and proximal neuronal processes to more distal sites of the neuropil in p35-/- mouse brain implicate p35 in neuronal trafficking, particularly in dynein-driven retrograde transport. In many axons of normal brain, cdk5 fails to colocalize with phosphorylated cytoskeletal protein epitopes. This observation, together with an unexpected increase of NF, tau, and MAP2B phosphoepitopes accompanying the decreased cdk5 activity in p35-/- mice, supports the idea that cdk5 does not phosphorylate cytoskeletal proteins directly. Rather, in structures where cdk5 does colocalize with phosphorylated cytoskeletal protein epitopes, it may function as a negative regulator of other proline-directed kinases that directly phosphorylate the proteins. Evidence for increased glycogen synthase kinase 3beta (GSK3beta) activity in p35-/- mice suggests that GSK3beta may be one such kinase regulated by cdk5. Our studies illustrate that p35 regulates the subcellular distribution of cdk5 and cytoskeletal proteins in neurons and that cdk5 has a hierarchical role in regulating the phosphorylation and function of cytoskeletal proteins.

Animals↗

PubMed2003

Proline-directed phosphorylation and isomerization in mitotic regulation and in Alzheimer's Disease.

The reversible phosphorylation of proteins on serine/threonine residues preceding proline (Ser/Thr-Pro) is a major regulatory mechanism for the control of a series of cell cycle events. Although phosphorylation is thought to regulate protein function by inducing conformational changes, little is known about most of these conformational changes and their significance. Recent studies indicate that the conformation and function of a subset of these phosphorylated proteins are controlled by the prolyl isomerase Pin1 through isomerization of specific phosphorylated Ser/Thr-Pro bonds. Furthermore, compelling evidence supports the idea that proline-directed phosphorylation and subsequent isomerization play a critical role not only in cell cycle control, but also in the development of Alzheimer's disease, where postmitotic neurons display various cell cycle markers, especially mitotic events, prior to degeneration.

Alzheimer Disease↗

PubMed2003

Understanding Niemann-Pick type C disease: a fat problem.

PURPOSE OF REVIEW: Progressive neurological deterioration is the ultimate cause of premature death in Niemann-Pick type C disease. Yet it remains unknown why a defect in basic cellular lipid homeostasis would lead to such profound neurological dysfunction and degeneration. The established belief that the central nervous system disorder of Niemann-Pick type C disease is secondary to lipid accumulation has led to a deficiency of research and information on the neurological manifestation of the disease. Lipid and vesicular trafficking have been studied extensively in non-neuronal cells such as fibroblasts and significant advances have been summarized in excellent reviews. The purpose of the present review is to consolidate the findings on brain lipid metabolism and Niemann-Pick type C disease neuropathology, so as to generate a current picture of the central nervous system disease, and emphasize potential aspects for future research. RECENT FINDINGS: Some key findings in the last couple of years have brought the neurological features of Niemann-Pick type C disease into focus. Most importantly, multiple strategies for reducing lipid overload have proven ineffective in preventing the onslaught of neurodegeneration, while efficiently reducing lipid disturbances in extracerebral tissues. Of much significance was the finding that the central nervous system disease is brain autonomous. Finally, investigation of lipid defects and neuropathological changes in Niemann-Pick type C disease gene-1-deficient mice has shed light on some novel putative therapeutic targets. SUMMARY: The independence of brain pathology from visceral complications in Niemann-Pick type C disease has implications for its treatment. Further studies of the neurological mechanisms underlying the disease will have a significant impact on future clinical diagnosis and management of patients.

Animals↗

PubMed2003

The cell cycle and human neurodegenerative disease.

The mechanism by which neurons die in human neurodegenerative diseases remains an enigma till today. Terminally differentiated neurons of normal brain are incapable of cell division. However, accumulating evidence has suggested that aberrant activation of the cell cycle in certain degenerative diseases leads to their demise. In Alzheimer's disease, regulators spanning every phase of the cell cycle are upregulated in affected neurons, leading to successful DNA replication, but unsuccessful mitosis. The end point of this nonproductive cycle of division is death. Elucidating the details of this cell cycle-mediated degenerative cascade may lead to novel strategies for curbing the onset and progression of degenerative diseases.

Animals↗

PubMed2002

Deregulation of cdk5, hyperphosphorylation, and cytoskeletal pathology in the Niemann-Pick type C murine model.

NPC-1 gene mutations cause Niemann-Pick type C (NPC), a neurodegenerative storage disease resulting in premature death in humans. Spontaneous mutation of the NPC-1 gene in mice generates a similar phenotype, usually with death ensuing by 12 weeks of age. Both human and murine NPC are characterized neuropathologically by ballooned neurons distended with lipid storage, axonal spheroid formation, demyelination, and widespread neuronal loss. To elucidate the biochemical mechanism underlying this neuropathology, we have investigated the phosphorylation of neuronal cytoskeletal proteins in the brains of npc-1 mice. A spectrum of antibodies against phosphorylated epitopes in neurofilaments (NFs) and MAP2 and tau were used in immunohistochemical and immunoblotting analyses of 4- to 12-week-old mice. Multiple sites in NFs, MAP2, and tau were hyperphosphorylated as early as 4 weeks of age and correlated with a significant increase in activity of the cyclin-dependent kinase 5 (cdk5) and accumulation of its more potent activator, p25, a proteolytic fragment of p35. At 5 weeks of age, the development of axonal spheroids was noted in the pons. p25 and cdk5 coaccumulated with hyperphosphorylated cytoskeletal proteins in axon spheroids. These various abnormalities escalated with each additional week of age, spreading to other regions of the brainstem, basal ganglia, cerebellum, and eventually, the cortex. Our data suggest that focal deregulation of cdk5/p25 in axons leads to cytoskeletal abnormalities and eventual neurodegeneration in NPC. The npc-1 mouse is a valuable in vivo model for determining how and when cdk5 becomes deregulated and whether cdk5 inhibitors would be useful in blocking NPC neurodegeneration.

Animals↗

PubMed2002

Niemann-Pick disease type C yields possible clue for why cerebellar neurons do not form neurofibrillary tangles.

It is unknown why cerebellar neurons resist neurofibrillary tangle (NFT) formation. In Niemann-Pick disease Type C (NPC), NFT-mediated neurodegeneration occurs throughout brain, but the cerebellum degenerates conspicuously without NFT. To understand why, we have studied markers of NFT pathogenesis in cerebellum from 17 NPC cases, all having abundant NFT in forebrain. Remarkably, we found that NPC cerebella display several early markers of NFT formation, i.e., hyperphosphorylated tau and an array of cell cycle regulators, suggesting that cerebellar neurons in NPC undergo similar modifications as other neurons that develop NFT. However, cerebellar neurons are deficient in tau, the building block of NFT, and this may be one reason for their inability to form NFT. Even without NFT, cerebellar neurodegeneration may be triggered by the inappropriate activation of the cell cycle cdc2 kinase, and the npc-1 murine model provides an opportunity to test this hypothesis.

Adolescent↗

PubMed2001

Constitutive Wee1 activity in adult brain neurons with M phase-type alterations in Alzheimer neurodegeneration.

The cdc2/cyclin B1 kinase is absent from neurons that are terminally differentiated. However, unscheduled activation of Cdc2/cyclin B and accumulation of mitotic phosphoepitopes have been described in degenerating neurons of Alzheimer's disease (AD), and their appearance precedes hallmark lesion formation. In cycling cells the timing of cdc2 activation and onset of mitosis are determined by the Wee1 tyrosine kinase. We therefore investigated the Wee1 kinase in human brain. Surprisingly, we have found that the enzyme is constitutively active in neurons of normal brain. Consistent with its behavior in M phase, Wee1 in AD has decreased activity, becomes MPM-2 immunoreactive, and is redistributed from its normally nuclear domain to the cytoplasm of affected neurons. These data suggest that Wee1 functions in normal postmitotic neurons, but is altered in AD so as to promote activation of Cdc2/cyclin B1. Thus, Wee1 is yet another mitotic regulator that participates in the AD neurodegenerative process.

Journal Article↗