Publications (21) View all
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Article: Albumin induces upregulation of matrix metalloproteinase-9 in astrocytes via MAPK and reactive oxygen species-dependent pathways.
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ABSTRACT: Astrocytes are an integral component of the blood-brain barrier (BBB) which may be compromised by ischemic or traumatic brain injury. In response to trauma, astrocytes increase expression of the endopeptidase matrix metalloproteinase (MMP)-9. Compromise of the BBB leads to the infiltration of fluid and blood-derived proteins including albumin into the brain parenchyma. Albumin has been previously shown to activate astrocytes and induce the production of inflammatory mediators. The effect of albumin on MMP-9 activation in astrocytes is not known. We investigated the molecular mechanisms underlying the production of MMP-9 by albumin in astrocytes. Primary enriched astrocyte cultures were used to investigate the effects of exposure to albumin on the release of MMP-9. MMP-9 expression was analyzed by zymography. The involvement of mitogen-activated protein kinase (MAPK), reactive oxygen species (ROS) and the TGF-β receptor-dependent pathways were investigated using pharmacological inhibitors. The production of ROS was observed by dichlorodihydrofluorescein diacetate fluorescence. The level of the MMP-9 inhibitor tissue inhibitor of metalloproteinase (TIMP)-1 produced by astrocytes was measured by ELISA. We found that albumin induces a time-dependent release of MMP-9 via the activation of p38 MAPK and extracellular signal regulated kinase, but not Jun kinase. Albumin-induced MMP-9 production also involves ROS production upstream of the MAPK pathways. However, albumin-induced increase in MMP-9 is independent of the TGF-β receptor, previously described as a receptor for albumin. Albumin also induces an increase in TIMP-1 via an undetermined mechanism. These results link albumin (acting through ROS and the p38 MAPK) to the activation of MMP-9 in astrocytes. Numerous studies identify a role for MMP-9 in the mechanisms of compromise of the BBB, epileptogenesis, or synaptic remodeling after ischemia or traumatic brain injury. The increase in MMP-9 produced by albumin further implicates both astrocytes and albumin in the acute and long-term complications of acute CNS insults, including cerebral edema and epilepsy.Journal of Neuroinflammation 04/2012; 9:68. · 3.83 Impact Factor -
Article: Mild stretch-induced injury increases susceptibility to interleukin-1β-induced release of matrix metalloproteinase-9 from astrocytes.
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ABSTRACT: Traumatic brain injury (TBI) results in the activation of glia and the release of proinflammatory cytokines, including interleukin (IL)-1β. The response of astrocytes to mild TBI has not been well studied. We used an in vitro model of cell stretch to investigate the effects of mild mechanical insult on astrocyte injury (lactate dehydrogenase and propidium iodide), and on mediators of inflammation including IL-1β, the chemokine CX3CL1, and nitrite. Here, we tested the hypothesis that a mild mechanical insult would increase susceptibility of astrocytes to delayed exposure to IL-1β, including enhanced release of the matrix metalloproteinease-9 (MMP-9). We investigated the role of the mitogen protein-activated kinase (MAPK) pathway in these responses. Cells subjected to a mild stretch show an increase in activation of the ERK1/2 and JNK pathways, and an increase in lactate dehydrogenase (LDH), but no change in the levels of inflammatory mediators. An early increase in LDH was dependent on ERK activation. Exposure to IL-1β, or to stretch alone, did not increase MMP-9. In contrast, the combination of mild stretch followed by IL-1β resulted in greater activation of the ERK pathway compared to either stimulus alone, and also resulted in an increase in the production of MMP-9 by astrocytes. Inhibition of the ERK pathway suppressed the increase in MMP-9 induced by the combination of stretch and IL-1β treatment. These results suggest that a primary mild mechanical injury renders astrocytes more susceptible to a secondary exposure to a proinflammatory cytokine such as IL-1β via the activation of the ERK pathway, and suggest a mechanism by which a mild head injury may confer increased susceptibility to neurologic injury caused by a subsequent insult.Journal of neurotrauma 07/2011; 28(9):1757-66. · 4.25 Impact Factor -
Article: Albumin causes increased myosin light chain kinase expression in astrocytes via p38 mitogen-activated protein kinase.
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ABSTRACT: Myosin light chain kinase (MLCK) plays an important role in the reorganization of the cytoskeleton, leading to disruption of vascular barrier integrity in multiple organs, including the blood-brain barrier (BBB), after traumatic brain injury (TBI). MLCK has been linked to transforming growth factor (TGF) and rho kinase signaling pathways, but the mechanisms regulating MLCK expression following TBI are not well understood. Albumin leaks into the brain parenchyma following TBI, activates glia, and has been linked to TGF-β receptor signaling. We investigated the role of albumin in the increase of MLCK in astrocytes and the signaling pathways involved in this increase. After midline closed-skull TBI in mice, there was a significant increase in MLCK-immunoreactive (IR) cells and albumin extravasation, which was prevented by treatment with the MLCK inhibitor ML-7. Using immunohistochemical methods, we identified the MLCK-IR cells as astrocytes. In primary astrocytes, exposure to albumin increased both isoforms of MLCK, 130 and 210. Inhibition of the TGF-β receptor partially prevented the albumin-induced increase in both isoforms, which was not prevented by inhibition of smad3. Inhibition of p38 MAPK, but not ERK, JNK, or rho kinase, also prevented this increase. These results are further evidence of a role of MLCK in the mechanisms of BBB compromise following TBI and identify astrocytes as a cell type, in addition to endothelium in the BBB, that expresses MLCK. These findings implicate albumin, acting through p38 MAPK, in a novel mechanism by which activation of MLCK following TBI may lead to compromise of the BBB.Journal of Neuroscience Research 02/2011; 89(6):852-61. · 2.74 Impact Factor -
Article: Arylsulfanyl pyrazolones block mutant SOD1-G93A aggregation. Potential application for the treatment of amyotrophic lateral sclerosis.
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ABSTRACT: Amyotrophic lateral sclerosis (ALS) is an orphan neurodegenerative disease currently without a cure. Mutations in copper/zinc superoxide dismutase 1 (SOD1) have been implicated in the pathophysiology of this disease. Using a high-throughput screening assay expressing mutant G93A SOD1, two bioactive chemical hit compounds (1 and 2), identified as arylsulfanyl pyrazolones, were identified. The structural optimization of this scaffold led to the generation of a more potent analogue (19) with an EC(50) of 170nM. To determine the suitability of this class of compounds for further optimization, 1 was subjected to a battery of pharmacokinetic assays; most of the properties of 1 were good for a screening hit, except it had a relatively rapid clearance and short microsomal half-life stability. Compound 2 was found to be blood-brain barrier penetrating with a brain/plasma ratio=0.19. The optimization of this class of compounds could produce novel therapeutic candidates for ALS patients.Bioorganic & medicinal chemistry 01/2011; 19(1):613-22. · 2.82 Impact Factor -
Article: Albumin activates the canonical TGF receptor-smad signaling pathway but this is not required for activation of astrocytes.
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ABSTRACT: The use of albumin as a resuscitation fluid is considered safe for most critically ill patients. However, clinical data suggest albumin may increase mortality in neurotrauma, but improve outcome after stroke. Albumin has been shown to activate glia, and to play a role in the mechanisms of epileptogenesis via the TGFβ-receptor (TGFβR). We have previously shown that albumin induces the production of inflammatory mediators including IL-1β via activation of MAPK pathways in primary astrocytes and microglia. The extracellular signaling mechanisms leading to the activation of glial cells in response to albumin are not well understood. Here, we investigated the role of the TGFβR and the canonical TGFβ receptor-smad signaling pathway in astrocyte activation by albumin. In primary astrocyte cultures, albumin activated the smad pathway downstream of the TGFβR by increasing the phosphorylation of smad2, and in the level of smad3 and smad4 translocated to the nucleus. Albumin produced an increase in IL-1β which was not dependent on smad activation, but was prevented by blockade of the TGFβR. Increase in the chemokine CX3CL1, and the decrease in S100B produced by albumin were independent of the TGFβR and smad activation. Albumin induced an increase in LDH release that was inhibited by blockade of the TGFβR and by inhibition of smad activation. These findings show that albumin activates the canonical TGF receptor-smad signaling pathway. The albumin-induced increase in the pro-epileptogenic cytokine IL-1β involves the TGFβR, but is independent of smad activation. Taken together, the effects of albumin on both IL-1β and activation of the TGFβR pathway are further evidence for a role for albumin in neurotrauma-related epileptogenesis.Experimental Neurology 12/2010; 226(2):310-9. · 4.70 Impact Factor
