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Biomedical subjects

Michael A Moskowitz

Publications and source records attributed to Michael A Moskowitz.

53 records · Page 3Linked to original sources
PubMed2003

Poly(ADP-ribose) polymerase-1 activity promotes NF-kappaB-driven transcription and microglial activation: implication for neurodegenerative disorders.

Excessive release of proinflammatory products by activated glia causes neurotoxicity and participates in the pathogenesis of neurodegenerative disorders. Recently, poly(ADP-ribose) polymerase-1 (PARP-1) has been shown to play a key role in nuclear factor kappa B (NF-kappaB)-driven expression of inflammatory mediators by glia during the neuroimmune response. Here we report the novel finding that the enzymatic activity of PARP-1 promotes, in an beta-nicotinamide adenine dinucleotide-dependent fashion, the DNA binding of NF-kappaB in microglia exposed to lipopolysaccharides, interferon-gamma or beta-amyloid 1-40. Consistently, we found that targeting NF-kappaB-dependent glial activation with pharmacological inhibitors of PARP-1 enzymatic activity reduces expression of inflammatory mediators such as inducible nitric oxide synthase, interleukin 1beta, tumor necrosis factor alpha and amyloid precursor protein, and reduces the neurotoxic potential of activated glia in vitro. Importantly, pharmacological inhibition of lipopolysaccharide-induced poly(ADP-ribose) formation in vivo suppresses neuroinflammation and related neural cell death. Our findings build on prior published reports in PARP-1 null mice and highlight the importance of PARP-1 enzymatic activity in transcriptional control during glial activation, identifying PARP-1 activity-dependent regulation of NF-kappaB as a novel pharmacological target for therapeutic intervention in the treatment of acute and chronic neurodegenerative disorders.

Animals↗
PubMed2003

FGF-2 regulates neurogenesis and degeneration in the dentate gyrus after traumatic brain injury in mice.

We studied the role of FGF-2 on regulation of neurogenesis and cell loss in the granule cell layer (GCL) of the hippocampal dentate gyrus after experimental traumatic brain injury (TBI). In both FGF-2(-/-) and FGF-2(+/+) mice subjected to controlled cortical impact, the number of dividing cells labeled with BrdU, injected on posttrauma days 6 through 8, increased at 9 days after TBI, and the number of BrdU-positive cells colabeled with neuron-specific nuclear antigen significantly increased at 35 days. However, in injured FGF-2-/- mice, BrdU-positive cells and BrdU-positive neurons (days 9, 35) were fewer compared with FGF-2(+/+) mice. There was also a decrease in the volume of the GCL and the number of GCL neurons after TBI in both FGF-2(-/-) and FGF-2(+/+) mice, but the decrease in both was greater in FGF-2-/- mice at 35 days. Overexpression of FGF-2 by intracerebral injection of herpes simplex virus-1 amplicon vectors encoding this factor increased numbers of dividing cells (day 9) and BrdU-positive neurons (day 35) significantly in C57BL/6 mice. Furthermore, the decrease in GCL volume was also attenuated. These results suggest that FGF-2 upregulates neurogenesis and protects neurons against degeneration in the adult hippocampus after TBI, and that FGF-2 supplementation via gene transfer can reduce GCL degeneration after TBI.

Animals↗
PubMed2003

EGF amplifies the replacement of parvalbumin-expressing striatal interneurons after ischemia.

EGF promotes proliferation and migration of stem/progenitor cells in the normal adult brain. The effect of epidermal growth factor on neurogenesis in ischemic brain is unknown, however. Here we show that intraventricular administration of EGF and albumin augments 100-fold neuronal replacement in the injured adult mouse striatum after cerebral ischemia. Newly born immature neurons migrate into the ischemic lesion and differentiate into mature parvalbumin-expressing neurons, replacing more than 20% of the interneurons lost by 13 weeks after ischemia and representing 2% of the total BrdU-labeled cells. These data suggest that administration of EGF and albumin could be used to manipulate endogenous neurogenesis in the injured brain and to promote brain self-repair.

Animals↗
PubMed2002

Caspase activation and neuroprotection in caspase-3- deficient mice after in vivo cerebral ischemia and in vitro oxygen glucose deprivation.

Caspase-3 is a major cell death effector protease in the adult and neonatal nervous system. We found a greater number and higher density of cells in the cortex of caspase-3(-/-) adult mice, consistent with a defect in developmental cell death. Caspase-3(-/-) mice were also more resistant to ischemic stress both in vivo and in vitro. After 2 h of ischemia and 48 h of reperfusion, cortical infarct volume was reduced by 55%, and the density of terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling-positive cells was decreased by 36% compared with wild type. When subjected to oxygen-glucose deprivation (2 h), cortical neurons cultured from mice deficient in caspase-3 expression were also more resistant to cell death by 59%. Mutant brains showed caspase-specific poly(ADP-ribose) polymerase cleavage product (85-kDa fragment) in vivo and in vitro, suggesting redundant mechanisms and persistence of caspase-mediated cell death. In the present study, we found that caspase-8 mediated poly(ADP-ribose) polymerase cleavage in caspase-3(-/-) neurons in vivo and in vitro. In addition, mutant neurons showed no evidence of compensatory activation by caspase-6 or caspase-7 after ischemia. Taken together, these data extend the pharmacological evidence supporting an important role for caspase-3 and caspase-8 as cell death mediators in mammalian cortex and indicate the potential advantages of targeting more than a single caspase family member to treat ischemic cell injury.

Animals↗
PubMed2002

Mechanisms of pain modulation in chronic syndromes.

Transmission of pain from the periphery to the cortex depends on integration and signal processing within the spinal cord, brainstem, and forebrain. Sensitization, a component of persistent or chronic pain, may develop either through peripheral mechanisms or as a consequence of altered physiology in the spinal cord or forebrain. Several molecular and biophysical mechanisms contribute to the phenomenon of sensitization and persistent pain, including upregulation of sensory neuron-specific sodium channels and vanilloid receptors, phenotypic switching of large myelinated axons, sprouting within the dorsal horn, and loss of inhibitory neurons due to apoptotic cell death. Recently, forebrain structures have been implicated in the pathophysiology of persistent pain. Although a number of treatment options are used, unfortunately pharmacotherapy for neuropathic pain is often ineffective. Unraveling the mysteries of chronic pain may lead to better treatment options, such as drugs that act specifically on sensory neuron-specific sodium channels or as NR2B-subunit-selective N-methyl-D-aspartate receptor antagonists.

Animals↗
PubMed2002

Lipid-sugar particles for intracranial drug delivery: safety and biocompatibility.

Controlled release of drugs to specific locales in the brain has engendered considerable interest. Here we evaluate the safety and biocompatibility of 6-microm diameter particles composed of dipalmitoylphosphatidylcholine and chondroitin sulfate A, when delivered into the cerebral parenchyma and ventricles, and in the case of intravascular injection. Some particles were loaded with fluorescein-labeled albumin to facilitate detection. Particles placed in medium with cultured murine primary cortical neurons did not increase cell death at concentrations as high as 4 mg/ml. When particles (100 microg in 2 microl) were placed stereotactically in the striatum and lateral ventricles, there was no histological evidence on hematoxylin-eosin stained sections of tissue injury outside of the needle track in any animal 3, 7, and 14 days after injection (n=6 each), and no inflammation. Ventricular size was not significantly different between animals given intraventricular injections of particles and albumin solution at those time points (n=4 each). Intracarotid injection of particles at concentrations of 0.2 and 1 mg/ml (n=4 each) did not affect relative cerebral blood flow, and there were no embolic events on histology. In one animal in the group injected with 5 mg/ml (n=3), there was a profound decrease in rCBF, with patchy emboli on histology. These novel biodegradable particles are biocompatible in and around the brain, and may be safe for intracranial sustained drug delivery either in the parenchyma or into the CSF.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed2002

Upregulation of the Fas receptor death-inducing signaling complex after traumatic brain injury in mice and humans.

Recent studies have implicated Fas in the pathogenesis of inflammatory, ischemic, and traumatic brain injury (TBI); however, a direct link between Fas activation and caspase-mediated cell death has not been established in injured brain. We detected Fas-Fas ligand binding and assembly of death-inducing signaling complexes (DISCs) [Fas, Fas-associated protein with death domain, and procaspase-8 or procaspase-10; receptor interacting protein (RIP)-RIP-associated interleukin-1beta converting enzyme and CED-3 homolog-1/Ced 3 homologous protein with a death domain-procaspase-2] by immunoprecipitation and immunoblotting within mouse parietal cortex after controlled cortical impact. At the time of DISC assembly, procaspase-8 was cleaved and the cleavage product appeared at 48 hr in terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling-positive neurons. Cleavage of caspase-8 was accompanied by caspase-3 processing detected at 48 hr by immunohistochemistry, and by caspase-specific cleavage of poly(ADP-ribose) polymerase at 12 hr. Fas pathways were also stimulated by TBI in human brain, because Fas expression plus Fas-procaspase-8 interaction were robust in contused cortical tissue samples surgically removed between 2 and 30 hr after injury. To address whether Fas functions as a death receptor in brain cells, cultured embryonic day 17 cortical neurons were transfected with an adenoviral vector containing the gene encoding Fas ligand. After 48 hr in culture, Fas ligand expression and Fas-procaspase-8 DISC assembly increased, and by 72 hr, cell death was pronounced. Cell death was decreased by approximately 50% after pan-caspase inhibition (Z-Val-ALa-Asp(Ome)-fluoromethylketone). These data suggest that Fas-associated DISCs assemble in neurons overexpressing Fas ligand as well as within mouse and human contused brain after TBI. Therefore, Fas may function as a death receptor after brain injury.

Adaptor Proteins, Signal Transducing↗
PubMed2002

Nuclear factor-kappaB as a molecular target for migraine therapy.

Nitric oxide (NO) generated from inducible NO synthase (iNOS) participates in immune and inflammatory responses in many tissues. The NO donor glyceryl trinitrate (GTN) provokes delayed migraine attacks when infused into migraineurs and also causes iNOS expression and delayed inflammation within rodent dura mater. Sodium nitroprusside, an NO donor as well, also increases iNOS expression. Because inflammation and iNOS are potential therapeutic targets, we examined transcriptional regulation of iNOS following GTN infusion and the consequences of its inhibition within dura mater. We show that intravenous GTN increases NO production within macrophages. L-N(6)-(1-iminoethyl)lysine, a selective iNOS inhibitor, attenuates the NO signal, emphasizing the importance of enzymatic activity to delayed NO production. iNOS expression is preceded by significant nuclear factor kappa B (NF-kappaB) activity, as reflected by a reduction in the inhibitory protein-kappa-Balpha (IkappaBalpha) and activation of NF-kappaB after GTN infusion. IkappaBalpha degradation, NF-kappaB activation, and iNOS expression were attenuated by parthenolide (3mg/kg), the active constituent of feverfew, an anti-inflammatory drug used for migraine treatment. These findings suggest that GTN promotes NF-kappaB activity and inflammation with a time course consistent with migraine attacks in susceptible individuals. We conclude, based on results with this animal model, that blockade of NF-kappaB activity provides a novel transcriptional target for the development of anti-migraine drugs.

Animals↗
PubMed2002

Neurotrophin-3 promotes cell death induced in cerebral ischemia, oxygen-glucose deprivation, and oxidative stress: possible involvement of oxygen free radicals.

To explore the role of neurotrophin-3 (NT-3) during cerebral ischemia, NT-3-deficient brains were subjected to transient focal ischemia. Conditional mutant brains produced undetectable amounts of NT-3 mRNA, whereas the expression of the neurotrophin, BDNF, the NT-3 receptor, TrkC, and the nonselective, low-affinity neurotrophin receptor p75NTR, were comparable to wild-type. Baseline absolute blood flow, vascular and neuroanatomical features, as well as physiological measurements were also indistinguishable from wild-type. Interestingly, the absence of NT-3 led to a significantly decreased infarct volume 23 h after middle cerebral artery occlusion. Consistent with this, the addition of NT-3 to primary cortical cell cultures exacerbated neuronal death caused by oxygen-glucose deprivation. Coincubation with the oxygen free radical chelator, trolox, diminished potentiation of neuronal death. NT-3 also enhanced neuronal cell death and the production of reactive oxygen species caused by oxidative damage inducing agents. We conclude that endogenous NT-3 enhanced neuronal injury during acute stroke, possible by increasing oxygen-radical mediated cell death.

Animals↗
PubMed2002

The HD mutation does not alter neuronal death in the striatum of Hdh(Q92) knock-in mice after mild focal ischemia.

Huntington's disease, with its dominant loss of striatal neurons, is triggered by an expanded glutamine tract in huntingtin. To investigate a proposed role for increased activation of the apoptotic cascade in mutant huntingtin's trigger mechanism, we examined huntingtin cleavage and lesion severity after mild ischemic injury in Hdh(Q92) mice. We found activation of calpain and caspase proteases and proteolysis of huntingtin in lesioned striatum. However, huntingtin fragments resembled products of calpain I, not caspase-3, cleavage and turnover was accompanied by augmented levels of full-length normal and mutant protein. By contrast, the number of apoptotic cells, total and striatal infarct size, and degree of neurologic deficit were similar in Hdh(Q92) and wild-type mice, indicating that the disease process neither strongly protected nor sensitized striatal neurons to apoptotic death. Thus, our findings do not support a role for increased apoptosis or caspase-3 cleavage in the mechanism by which mutant huntingtin triggers disease. However, they suggest that calpain activation and huntingtin regulation merit investigation as modifiers of disease progression in neurons injured by the harmful consequences of full-length mutant huntingtin.

Animals↗
PubMed2002

Intrinsic brain activity triggers trigeminal meningeal afferents in a migraine model.

Although the trigeminal nerve innervates the meninges and participates in the genesis of migraine headaches, triggering mechanisms remain controversial and poorly understood. Here we establish a link between migraine aura and headache by demonstrating that cortical spreading depression, implicated in migraine visual aura, activates trigeminovascular afferents and evokes a series of cortical meningeal and brainstem events consistent with the development of headache. Cortical spreading depression caused long-lasting blood-flow enhancement selectively within the middle meningeal artery dependent upon trigeminal and parasympathetic activation, and plasma protein leakage within the dura mater in part by a neurokinin-1-receptor mechanism. Our findings provide a neural mechanism by which extracerebral cephalic blood flow couples to brain events; this mechanism explains vasodilation during headache and links intense neurometabolic brain activity with the transmission of headache pain by the trigeminal nerve.

Brain↗
PubMed2002

Acute cardiovascular protective effects of corticosteroids are mediated by non-transcriptional activation of endothelial nitric oxide synthase.

Corticosteroids have been shown to exert beneficial effects in the treatment of acute myocardial infarction, but the precise mechanisms underlying their protective effects are unknown. Here we show that high-dose corticosteroids exert cardiovascular protection through a novel mechanism involving the rapid, non-transcriptional activation of endothelial nitric oxide synthase (eNOS). Binding of corticosteroids to the glucocorticoid receptor (GR) stimulated phosphatidylinositol 3-kinase and protein kinase Akt, leading to eNOS activation and nitric oxide dependent vasorelaxation. Acute administration of pharmacological concentrations of corticosteroids in mice led to decreased vascular inflammation and reduced myocardial infarct size following ischemia and reperfusion injury. These beneficial effects of corticosteroids were abolished by GR antagonists or eNOS inhibitors in wild-type mice and were completely absent in eNOS-deficient (Nos3(-/-)) mice. The rapid activation of eNOS by the non-nuclear actions of GR, therefore, represents an important cardiovascular protective effect of acute high-dose corticosteroid therapy.

Adrenal Cortex Hormones↗
PubMed2002

STAT1 is activated in neurons after ischemia and contributes to ischemic brain injury.

Signal transducers and activators of transcription (STAT) proteins are a family of transcription factors that play a crucial role in growth and differentiation in a variety of cell types. Among them, STAT1, which is expressed in the brain and directly activated by reactive oxygen species, participates in the regulation of cytokine-signaling and cellular responses, particularly to interferon-gamma. Very little, however, is known about the importance of STAT1 during brain injury. The authors found that STAT1 was phosphorylated at tyrosine and serine727 and translocated into neuronal nuclei within hours after middle cerebral artery occlusion. At later time points, STAT1 immunoreactivity colocalized with TUNEL-positive neurons, thereby suggesting a role in cell death. In mice genetically deficient in STAT1 expression, the volume of ischemic brain injury was reduced, neurologic deficits were less severe, and TUNEL-positive neurons were also less numerous compared with wild-type mice. STAT1-knockout mice showed increased phosphorylated Akt and decreased procaspase-3 cleavage. Major strain differences in phosphorylated STAT3 or cyclooxygenase-2 protein expression were not found after ischemia. These results indicate that STAT1 is activated and translocated within ischemic neurons and may contribute to brain injury by regulating transcription and phosphorylation of proteins related to apoptosis and cell death.

Animals↗
PubMed2002

Rapid nontranscriptional activation of endothelial nitric oxide synthase mediates increased cerebral blood flow and stroke protection by corticosteroids.

Many cellular responses to corticosteroids involve the transcriptional modulation of target genes by the glucocorticoid receptor (GR). A rapid, non-nuclear effect of GR was found to mediate neuroprotection. High-dose corticosteroids (20 mg/kg intraperitoneally), given within 2 hours of transient cerebral ischemia, acutely increased endothelial nitric oxide synthase (eNOS) activity, augmented regional cerebral blood flow (CBF) by 40% to 50%, and reduced cerebral infarct size by 32%. These neuroprotective effects of corticosteroids were abolished by the GR antagonist RU486 and by inhibition of phosphatidylinositol 3-kinase (PI3K), and were absent in eNOS(-/-) mice. To determine the mechanism by which GR activated eNOS, we measured the effect of corticosteroids on PI3K and the protein kinase Akt. In a ligand-dependent manner, GR activated PI3K and Akt in vitro and in vivo caused NO-dependent vasodilation, which was blocked by cotreatment with RU486 or the PI3K inhibitor LY294002 but not by transcriptional inhibitors. Indeed, a mutant GR, which cannot dimerize and bind to DNA, still activated PI3K and Akt in response to corticosteroids. These findings indicate that non-nuclear GR rapidly activates eNOS through the PI3K/Akt pathway and suggest that this mechanism mediates the acute neuroprotective effects of corticosteroids through augmentation of CBF.

Adrenal Cortex Hormones↗
PubMedDate not supplied

Function of BID -- a molecule of the bcl-2 family -- in ischemic cell death in the brain.

Mitochondrial mechanisms, particularly the release of cytochrome c, play a role in the death of nerve and glial cells in cerebral ischemia. We have currently investigated whether BID, a proapoptotic molecule of the bcl-2 family and promoter of the release of cytochrome c is expressed in the brain, activated by cerebral ischemia in vivo, and contributes to ischemic cell death. We found BID in the cytosol of mouse brain and of primary cultured mouse neurons and showed that neuronal BID is a substrate for caspase 8. BID was cleaved in vivo 4 h after transitory occlusion of the middle cerebral artery. Further, BID(-/-) mice had a significant attenuation of infarction (-67%) and significantly lower release of cytochrome c (-41%). The findings indicate that the proapoptotic molecule BID may contribute to the demise of nerve cells from cerebral ischemia by release of cytochrome c and activation of caspase.

Animals↗