Thrombolysis remains the only effective therapy to reverse acute ischaemic stroke. However, delayed treatment may cause serious complications including hemorrhagic transformation and reperfusion ...injury. The level of lipocalin‐2 (LCN2) is elevated in the plasma of ischaemic stroke patients, but its role in stroke is unknown. Here, we show that LCN2 was acutely induced in mice after ischaemic stroke and is an important mediator of reperfusion injury. Increased levels of LCN2 were observed in mouse serum as early as 1 hr after transient middle cerebral artery occlusion (tMCAO), reaching peak levels at 23 hrs. LCN2 was also detected in neutrophils infiltrating into the ipsilateral hemisphere, as well as a subset of astrocytes after tMCAO, but not in neurons and microglia. Stroke injury, neurological deficits and infiltration of immune cells were markedly diminished in LCN2 null mice after tMCAO, but not after permanent MCAO (pMCAO). In vitro, recombinant LCN2 protein induced apoptosis in primary cultured neurons in a dose‐dependent manner. Our results demonstrate that LCN2 is a neurotoxic factor secreted rapidly in response to cerebral ischaemia, suggesting its potential usage as an early stroke biomarker and a novel therapeutic target to reduce stroke‐reperfusion injury.
Oxidative stress is a key contributor to the pathogenesis of stroke-reperfusion injury. Neuroinflammatory peptides released after ischemic stroke mediate reperfusion injury. Previous studies, ...including ours, have shown that lipocalin-2 (LCN2) is secreted in response to cerebral ischemia to promote reperfusion injury. Genetic deletion of LCN2 significantly reduces brain injury after stroke, suggesting that LCN2 is a mediator of reperfusion injury and a potential therapeutic target. Immunotherapy has the potential to harness neuroinflammatory responses and provides neuroprotection against stroke. Here we report that LCN2 was induced on the inner surface of cerebral endothelial cells, neutrophils, and astrocytes that gatekeep the blood-brain barrier (BBB) after stroke. LCN2 monoclonal antibody (mAb) specifically targeted LCN2 in vitro and in vivo, attenuating the induction of LCN2 and pro-inflammatory mediators (iNOS, IL-6, CCL2, and CCL9) after stroke. Administration of LCN2 mAb at 4 h after stroke significantly reduced neurological deficits, cerebral infarction, edema, BBB leakage, and infiltration of neutrophils. The binding epitope of LCN2 mAb was mapped to the β3 and β4 strands, which are responsible for maintaining the integrity of LCN2 cup-shaped structure. These data indicate that LCN2 can be pharmacologically targeted using a specific mAb to reduce reperfusion injury after stroke.
Lipocalin‐2 (LCN2) has been implicated in promoting apoptosis and neuroinflammation in neurological disorders; however, its role in neural transplantation remains unknown. In this study, we cultured ...and differentiated Lund human mesencephalic (LUHMES) cells into human dopaminergic‐like neurons and found that LCN2 mRNA was progressively induced in mouse brain after the intrastriatal transplantation of human dopaminergic‐like neurons. The induction of LCN2 protein was detected in a subset of astrocytes and neutrophils infiltrating the core of the engrafted sites, but not in neurons and microglia. LCN2‐immunoreactive astrocytes within the engrafted sites expressed lower levels of A1 and A2 astrocytic markers. Recruitment of microglia, neutrophils, and monocytes after transplantation was attenuated in LCN2 deficiency mice. The expression of M2 microglial markers was significantly elevated and survival of engrafted neurons was markedly improved after transplantation in LCN2 deficiency mice. Brain type organic cation transporter (BOCT), the cell surface receptor for LCN2, was induced in dopaminergic‐like neurons after differentiation, and treatment with recombinant LCN2 protein directly induced apoptosis in dopaminergic‐like neurons in a dose‐dependent manner. Our results, therefore, suggested that LCN2 is a neurotoxic factor for the engrafted neurons and a modulator of neuroinflammation. LCN2 inhibition may be useful in reducing rejection after neural transplantation.
Objective:
Mutations in the type IV collagen alpha 1 gene (COL4A1) cause dominantly inherited cerebrovascular disease. We seek to determine the extent to which COL4A1 mutations contribute to ...sporadic, nonfamilial, intracerebral hemorrhages (ICHs).
Methods:
We sequenced COL4A1 in 96 patients with sporadic ICH. The presence of putative mutations was tested in 145 ICH‐free controls. The effects of rare coding variants on COL4A1 biosynthesis were compared to previously validated mutations that cause porencephaly, small vessel disease, and hereditary angiopathy, nephropathy, aneurysms, and cramps (HANAC) syndrome.
Results:
We identified 2 rare nonsynonymous variants in ICH patients that were not detected in controls, 2 rare nonsynonymous variants in controls that were not detected in patients, and 2 common nonsynonymous variants that were detected in patients and controls. No variant found in controls affected COL4A1 biosynthesis. Both variants (COL4A1P352L and COL4A1R538G) found only in patients changed conserved amino acids and impaired COL4A1 secretion much like mutations that cause familial cerebrovascular disease.
Interpretation:
This is the first assessment of the broader role for COL4A1 mutations in the etiology of ICH beyond a contribution to rare and severe familial cases and the first functional evaluation of the biosynthetic consequences of an allelic series of COL4A1 mutations that cause cerebrovascular disease. We identified 2 putative mutations in 96 patients with sporadic ICH and showed that these and other previously validated mutations inhibit secretion of COL4A1. Our data support the hypothesis that increased intracellular accumulation of COL4A1, decreased extracellular COL4A1, or both, contribute to sporadic cerebrovascular disease and ICH. ANN NEUROL 2012;
Transient global cerebral ischemia (tGCI) resulting from cardiac arrest causes selective neurodegeneration in hippocampal CA1 neurons. Although the effect is clear, the underlying mechanisms ...directing this process remain unclear. Previous studies have shown that phosphorylation of Erk1/2 promotes cell survival in response to tGCI. DUSP6 (also named MKP3) serves as a cytosolic phosphatase that dephosphorylates Erk1/2, but the role of DUSP6 in tGCI has not been characterized. We found that DUSP6 was specifically induced in the cytoplasm of hippocampal CA1 neurons 4 to 24 h after tGCI. DUSP6-deficient mice showed normal spatial memory acquisition and retention in the Barnes maze. Impairment of spatial memory acquisition and retention after tGCI was attenuated in DUSP6-deficient mice. Neurodegeneration after tGCI, revealed by Fluoro-Jade C and H&E staining, was reduced in the hippocampus of DUSP6-deficient mice and DUSP6 deficiency enhanced the phosphorylation and nuclear translocation of Erk1/2 in the hippocampal CA1 region. These data support the role of DUSP6 as a negative regulator of Erk1/2 signaling and indicate the potential of DUSP6 inhibition as a novel therapeutic strategy to treat neurodegeneration after tGCI.
Global cerebral ischemia that accompanies cardiac arrest is a major cause of morbidity and mortality. Protein Kinase C epsilon (PKCε) is a member of the novel PKC subfamily and plays a vital role in ...ischemic preconditioning. Pharmacological activation of PKCε before cerebral ischemia confers neuroprotection. The role of endogenous PKCε after cerebral ischemia remains elusive. Here we used male PKCε‐null mice to assess the effects of PKCε deficiency on neurodegeneration after transient global cerebral ischemia (tGCI). We found that the cerebral vasculature, blood flow, and the expression of other PKC isozymes were not altered in the PKCε‐null mice. Spatial learning and memory was impaired after tGCI, but the impairment was attenuated in male PKCε‐null mice as compared to male wild‐type controls. A significant reduction in Fluoro‐Jade C labeling and mitochondrial release of cytochrome C in the hippocampus was found in male PKCε‐null mice after tGCI. Male PKCε‐null mice expressed increased levels of PKCδ in the mitochondria, which may prevent the translocation of PKCδ from the cytosol to the mitochondria after tGCI. Our results demonstrate the neuroprotective effects of PKCε deficiency on neurodegeneration after tGCI, and suggest that reduced mitochondrial translocation of PKCδ may contribute to the neuroprotective action in male PKCε‐null mice.
Neural cell death after transient global cerebral ischemia (tGCI) was attenuated in the hippocampus of mice lacking PKCε. Representative images of cresyl violet‐stained brain slices from Prkce+/+ and Prkce−/− mice 3 days after tGCI. Arrowheads indicate degenerative neurons with pyknotic and shrunken nuclei in the enlarged CA1 region of a Prkce+/+ brain slice (lower panels).
Hemorrhagic transformation (HT) is a serious complication after endovascular thrombectomy (EVT) for patients with acute ischemic stroke (AIS). We analyzed the plasma levels of MMP-9 before and after ...EVT and assessed the temporal changes of MMP-9 that may be associated with, and therefore predict, HT after EVT.
We enrolled 30 AIS patients who received EVT, and 16 (53.3%) developed HT. The levels of MMP-9 in plasma collected from the arteries of AIS patients before and immediately after EVT were measured using ELISA. The percent change in MMP-9 after EVT (after/before) was calculated and compared between patients with and without HT.
The median age of the AIS patients was 70 years, and 13 patients (43.3%) were men. The median National Institutes of Health Stroke Scale (NIHSS) scores of patients with HT were 18 on admission and 18 after EVT. The median NIHSS scores of patients without HT were 17 on admission and 11 after EVT. Patients with HT demonstrated significantly greater percentage increases in arterial MMP-9 levels after EVT.
Patients with AIS who developed HT had significantly increased arterial MMP-9 levels after EVT, suggesting that the upregulation of MMP-9 following EVT could serve as a predictive biomarker for HT.
Global cerebral ischemia triggers neurodegeneration in the hippocampal CA1 region, but the mechanism of neuronal death remains elusive. The epsilon isoform of protein kinase C (PKCε) has recently ...been identified as a master switch that controls the nucleocytoplasmic trafficking of ATF2 and the survival of melanoma cells. It is of interest to assess the role of PKCε-ATF2 signaling in neurodegeneration.
Phosphorylation of ATF2 at Thr-52 was reduced in the hippocampus of PKCε null mice, suggesting that ATF2 is a phosphorylation substrate of PKCε. PKCε protein concentrations were significantly reduced 4, 24, 48 and 72 h after transient global cerebral ischemia, resulting in translocation of nuclear ATF2 to the mitochondria. Degenerating neurons staining positively with Fluoro-Jade C exhibited cytoplasmic ATF2.
Our results support the hypothesis that PKCε regulates phosphorylation and nuclear sequestration of ATF2 in hippocampal neurons during ischemia-induced neurodegeneration.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Stroke is an important risk factor for dementia. Epidemiological studies have indicated a high incidence of dementia in stroke patients. There is currently no effective biomarker for the diagnosis of ...post-stroke dementia (PSD). D-amino acid oxidase (DAO) is a flavin-dependent enzyme widely distributed in the central nervous system. DAO oxidizes D-amino acids, a process which generates neurotoxic hydrogen peroxide and leads to neurodegeneration. This study aimed to examine post-stroke plasma DAO levels as a biomarker for PSD. In total, 53 patients with PSD, 20 post-stroke patients without dementia (PSNoD), and 71 age- and gender-matched normal controls were recruited. Cognitive function was evaluated at more than 30 days post-stroke. Plasma DAO was measured using the enzyme-linked immunosorbent assay. White matter hyperintensity (WMH), a neuroimaging biomarker of cerebral small vessel diseases, was determined by magnetic resonance imaging. We found that plasma DAO levels were independently higher in PSD subjects than in PSNoD subjects or the controls and were correlated with the WMH load in stroke patients. Using an area under the curve (AUC)/receiver operating characteristic analysis, plasma DAO levels were significantly reliable for the diagnosis of PSD. The sensitivity and specificity of the optimal cut-off value of 321 ng/ml of plasma DAO for the diagnosis of PSD were 75 and 88.7%, respectively. In conclusion, our data support that plasma DAO levels were increased in PSD patients and correlated with brain WMH, independent of age, gender, hypertension, and renal function. Plasma DAO levels may therefore aid in PSD diagnosis.
Ataxia-telangiectasia (A-T) and Nijmegen breakage syndrome (NBS) are recessive
genetic disorders with susceptibility to cancer and similar cellular phenotypes. The protein product of the gene ...responsible for A-T, designated
ATM, is a member of a family of kinases characterized by a carboxy-terminal
phosphatidylinositol 3-kinase-like domain. The NBS1 protein
is specifically mutated in patients with Nijmegen breakage syndrome and forms
a complex with the DNA repair proteins Rad50 and Mre11.
Here we show that phosphorylation of NBS1, induced by ionizing radiation,
requires catalytically active ATM. Complexes containing ATM and NBS1 exist
in vivo in both untreated cells and cells treated with ionizing radiation.
We have identified two residues of NBS1, Ser 278 and Ser 343
that are phosphorylated in vitro by ATM and whose modification
in vivo is essential for the cellular response to DNA damage. This response
includes S-phase checkpoint activation, formation of the NBS1/Mre11/Rad50
nuclear foci and rescue of hypersensitivity to ionizing radiation. Together,
these results demonstrate a biochemical link between cell-cycle checkpoints
activated by DNA damage and DNA repair in two genetic diseases with overlapping
phenotypes.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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