Amyloid Precursor Protein Staining Research Articles

Diffuse axonal injury has a characteristic multidimensional MRI signature in the human brain.

Axonal injury is a major contributor to the clinical symptomatology in patients with traumatic brain injury. Conventional neuroradiological tools, such as CT and MRI, are insensitive to diffuse axonal injury (DAI) caused by trauma. Diffusion tensor MRI parameters may change in DAI lesions; however, the nature of these changes is inconsistent. Multidimensional MRI is an emerging approach that combines T1, T2, and diffusion, and replaces voxel-averaged values with distributions, which allows selective isolation of specific potential abnormal components. By performing a combined post-mortem multidimensional MRI and histopathology study, we aimed to investigate T1-T2-diffusion changes linked to DAI and to define their histopathological correlates. Corpora callosa derived from eight subjects who had sustained traumatic brain injury, and three control brain donors underwent post-mortem ex vivo MRI at 7 T. Multidimensional, diffusion tensor, and quantitative T1 and T2 MRI data were acquired and processed. Following MRI acquisition, slices from the same tissue were tested for amyloid precursor protein (APP) immunoreactivity to define DAI severity. A robust image co-registration method was applied to accurately match MRI-derived parameters and histopathology, after which 12 regions of interest per tissue block were selected based on APP density, but blind to MRI. We identified abnormal multidimensional T1-T2, diffusion-T2, and diffusion-T1 components that are strongly associated with DAI and used them to generate axonal injury images. We found that compared to control white matter, mild and severe DAI lesions contained significantly larger abnormal T1-T2 component (P = 0.005 and P < 0.001, respectively), and significantly larger abnormal diffusion-T2 component (P = 0.005 and P < 0.001, respectively). Furthermore, within patients with traumatic brain injury the multidimensional MRI biomarkers differentiated normal-appearing white matter from mild and severe DAI lesions, with significantly larger abnormal T1-T2 and diffusion-T2 components (P = 0.003 and P < 0.001, respectively, for T1-T2; P = 0.022 and P < 0.001, respectively, for diffusion-T2). Conversely, none of the conventional quantitative MRI parameters were able to differentiate lesions and normal-appearing white matter. Lastly, we found that the abnormal T1-T2, diffusion-T1, and diffusion-T2 components and their axonal damage images were strongly correlated with quantitative APP staining (r = 0.876, P < 0.001; r = 0.727, P < 0.001; and r = 0.743, P < 0.001, respectively), while producing negligible intensities in grey matter and in normal-appearing white matter. These results suggest that multidimensional MRI may provide non-invasive biomarkers for detection of DAI, which is the pathological substrate for neurological disorders ranging from concussion to severe traumatic brain injury.

Cerebral Microcirculation and Histological Mapping After Severe Head Injury: A Contusion and Acceleration Experimental Model.

Cerebral microcirculation after severe head injury is heterogeneous and temporally variable. Microcirculation is dependent upon the severity of injury, and it is unclear how histology relates to cerebral regional blood flow. This study assesses the changes of cerebral microcirculation blood flow over time after an experimental brain injury model in sheep and contrasts these findings with the histological analysis of the same regions with the aim of mapping cerebral flow and tissue changes after injury. Microcirculation was quantified using flow cytometry of color microspheres injected under intracardiac ultrasound to ensure systemic and homogeneous distribution. Histological analysis used amyloid precursor protein staining as a marker of axonal injury. A mapping of microcirculation and axonal staining was performed using adjacent layers of tissue from the same anatomical area, allowing flow and tissue data to be available from the same anatomical region. A mixed effect regression model assessed microcirculation during 4 h after injury, and those results were then contrasted to the amyloid staining qualitative score. Microcirculation values for each subject and tissue region over time, including baseline, ranged between 20 and 80 ml/100 g/min with means that did not differ statistically from baseline flows. However, microcirculation values for each subject and tissue region were reduced from baseline, although their confidence intervals crossing the horizontal ratio of 1 indicated that such reduction was not statistically significant. Histological analysis demonstrated the presence of moderate and severe score on the amyloid staining throughout both hemispheres. Microcirculation at the ipsilateral and contralateral site of a contusion and the ipsilateral thalamus and medulla showed a consistent decline over time. Our data suggest that after severe head injury, microcirculation in predefined areas of the brain is reduced from baseline with amyloid staining in those areas reflecting the early establishment of axonal injury.

GSK-3β/mTORC1 Couples Synaptogenesis and Axonal Repair to Reduce Hypoxia Ischemia-Mediated Brain Injury in Neonatal Rats.

Glycogen synthase kinase 3 beta (GSK-3β) plays an important role in neurological outcomes after brain injury. However, its roles and mechanisms in hypoxia-ischemia (HI) are unclear. Activation of mTOR complex 1 (mTORC1) has been proven to induce the synthesis of proteins associated with regeneration. We hypothesized that GSK-3β inhibition could activate the mTORC1 signaling pathway, which may reduce axonal injury and induce synaptic protein synthesis and functional recovery of synapses after HI. By analyzing a P7 rat model of cerebral HI and an in vitro ischemic (oxygen glucose deprivation) model, we found that GSK-3β inhibitors (GSK-3β siRNA or lithium chloride) activated mTORC1 signaling, leading to increased expression of synaptic proteins, including synapsin 1, PSD95, and GluR1, and the microtubule-associated protein Tau and decreased expression of the axonal injury-associated protein amyloid precursor protein. These changes contributed to attenuated axonal injury (decreased amyloid precursor protein staining and axonal loss by silver staining), improved electrophysiological properties of synapses, and enhanced spatial memory performance in the Morris water maze. However, inhibition of mTORC1 by rapamycin blocked the benefits induced by GSK-3β inhibition, suggesting that GSK-3β inhibition induces synaptogenesis and axonal repair via mTORC1 signaling, which may benefit neonatal rats subjected to HI.

Cerebral microcirculation during mild head injury after a contusion and acceleration experimental model in sheep

Background: Cerebral microcirculation after head injury is heterogeneous and temporally variable. Regions at risk of infarction such as peri-contusional areas are vulnerable to anaemia. However, direct quantification of the cerebral microcirculation is clinically not feasible. This study describes a novel experimental head injury model correlating cerebral microcirculation with histopathology analysis.Objective: To test the hypothesis that cerebral microcirculation at the ischaemic penumbrae is reduced over time when compared with non-injured regions.Methods: Merino sheep were instrumented using a transeptal catheter to inject coded microspheres into the left cardiac atrium, ensuring systemic distribution. After a blunt impact over the left parietal region, cytometric analyses quantified cerebral microcirculation and amyloid precursor protein staining identified axonal injury in pre-defined anatomical regions. A mixed effect regression model assessed the hourly blood flow results during 4 hours after injury.Results: Cerebral microcirculation showed temporal reductions with minimal amyloid staining except for the ipsilateral thalamus and medulla.Conclusion: The spatial heterogeneity and temporal reduction of cerebral microcirculation in ovine models occur early, even after mild head injury, independent of the intracranial pressure and the level of haemoglobin. Alternate approaches to ensure recovery of regions with reversible injury require a targeted assessment of cerebral microcirculation.

Cerebral Microcirculation during Experimental Normovolaemic Anemia.

Anemia is accepted among critically ill patients as an alternative to elective blood transfusion. This practice has been extrapolated to head injury patients with only one study comparing the effects of mild anemia on neurological outcome. There are no studies quantifying microcirculation during anemia. Experimental studies suggest that anemia leads to cerebral hypoxia and increased rates of infarction, but the lack of clinical equipoise, when testing the cerebral effects of transfusion among critically injured patients, supports the need of experimental studies. The aim of this study was to quantify cerebral microcirculation and the potential presence of axonal damage in an experimental model exposed to normovolaemic anemia, with the intention of describing possible limitations within management practices in critically ill patients. Under non-recovered anesthesia, six Merino sheep were instrumented using an intracardiac transeptal catheter to inject coded microspheres into the left atrium to ensure systemic and non-chaotic distribution. Cytometric analyses quantified cerebral microcirculation at specific regions of the brain. Amyloid precursor protein staining was used as an indicator of axonal damage. Animals were exposed to normovolaemic anemia by blood extractions from the indwelling arterial catheter with simultaneous fluid replacement through a venous central catheter. Simultaneous data recording from cerebral tissue oxygenation, intracranial pressure, and cardiac output was monitored. A regression model was used to examine the effects of anemia on microcirculation with a mixed model to control for repeated measures. Homogeneous and normal cerebral microcirculation with no evidence of axonal damage was present in all cerebral regions, with no temporal variability, concluding that acute normovolaemic anemia does not result in short-term effects on cerebral microcirculation in the ovine brain.

Diffuse Axonal Injury: Pathological and Clinical Aspects

Diffuse axonal and vascular injuries are the initial pathological findings in victims of diffuse brain trauma Macroscopic findings include deep contusions and multiple petechial hemorrhages primarily involving cerebral white matter basal ganglia and upper brainstem Histologically amyloid precursor protein staining still remains the gold standard for microscopic detection of diffuse axonal damage A number of other histological and immunohistochemical techniques have emerged over time aiding the diagnosis of diffuse axonal injury There has been little success however in the microscopic differentiation of traumatic from non traumatic causes of diffuse axonal injury This review explains various pathological and clinical aspects of traumatic axonal and vascular injuries including current scenarios in microscopic detection of diffuse axonal injury as well as their limitations

Novel Rat Model of Weight Drop-Induced Closed Diffuse Traumatic Brain Injury Compatible with Electrophysiological Recordings of Vigilance States.

Traumatic brain injury (TBI) is a major cause of persistent disabilities such as sleep-wake disorders (SWD). Rodent studies of SWD after TBI are scarce, however, because of lack of appropriate TBI models reproducing acceleration-deceleration forces and compatible with electroencephalography/myography (EEG/EMG)-based recordings of vigilance states. We therefore adapted the Marmarou impact acceleration model to allow for compatibility with EEG-headset implantation. After implantation of EEG/EMG electrodes, we induced closed TBI by a frontal, angular hit with a weight-drop device (56 rats, weight 2500 g, fall height 25 cm). Subsequently, we tested our model's usefulness for long-term studies on a behavioral, electrophysiological, and histological level. Neurological, motor, and memory deficits were assessed with the neurological severity score, open field, and novel object recognition tests, respectively. EEG/EMG recordings were performed in both Sham (n = 7) and TBI (n = 7) rats before and 1, 7, and 28 days after trauma to evaluate sleep-wake proportions and post-traumatic implant stability. Histological assessments included hematoxylin and eosin staining for parenchymal damage and hemorrhage and amyloid precursor protein staining for diffuse axonal damage. All rats survived TBI without major neurological or motor deficits. Memory function was impaired after TBI at weeks 1, 2, and 3 and recovered at week 4. EEG implants were stable for at least 1 month and enabled qualitative and quantitative sleep analyses. Histological assessments revealed no major bleedings or necrosis but intense diffuse axonal damage after TBI. This approach fulfills major pre-conditions for experimental TBI models and offers a possibility to electrophysiologically study behavioral states before and after trauma.

Decompressive craniectomy reduces white matter injury after controlled cortical impact in mice.

Reduction and avoidance of increases in intracranial pressure (ICP) after severe traumatic brain injury (TBI) continue to be the mainstays of treatment. Traumatic axonal injury is a major contributor to morbidity after TBI, but it remains unclear whether elevations in ICP influence axonal injury. Here we tested the hypothesis that reduction in elevations in ICP after experimental TBI would result in decreased axonal injury and white matter atrophy in mice. Six-week-old male mice (C57BL/6J) underwent either moderate controlled cortical impact (CCI) (n=48) or Sham surgery (Sham, n=12). Immediately after CCI, injured animals were randomized to a loose fitting plastic cap (Open) or replacement of the previously removed bone flap (Closed). Elevated ICP was observed in Closed animals compared with Open and Sham at 15 min (21.4±4.2 vs. 12.3±2.9 and 8.8±1.8 mm Hg, p<0.0001) and 1 day (17.8±3.7 vs. 10.6±2.0 and 8.9±1.9 mm Hg, p<0.0001) after injury. Beta amyloid precursor protein staining in the corpus callosum and ipsilateral external capsule revealed reduced axonal swellings and bulbs in Open compared with Closed animals (32% decrease, p<0.01 and 40% decrease, p<0.001 at 1 and 7 days post-injury, respectively). Open animals were also found to have decreased neurofilament-200 stained axonal swellings at 7 days post-injury compared with Open animals (32% decrease, p<0.001). At 4 weeks post-injury, Open animals had an 18% reduction in white matter volume compared with 34% in Closed animals (p<0.01). Thus, our results indicate that CCI with decompressive craniectomy was associated with reductions in ICP and reduced pericontusional axonal injury and white matter atrophy. If similar in humans, therapeutic interventions that ameliorate intracranial hypertension may positively influence white matter injury severity.

Total-body low-dose irradiation of mice induces neither learning disability and memory impairment in Morris water maze test nor Alzheimer's disease-like pathogensis in the brain

Purpose: Alzheimer's disease (AD) is the most common form of dementia, while its cause and progression are not well understood [ 1]. The possible cognitive and behavioral consequences induced by low-dose radiation are of great concern as humans are exposed to ionizing radiations from various sources including medical diagnosis [ 2]. A recent study in mice reported early transcriptional response in brain to low-dose X-rays (0.10 Gy) suggesting alterations of molecular networks and pathways associated with cognitive functions, advanced aging and AD. The present study is to investigate the late pathological, cognitive and behavioral consequences induced by low-dose radiation.Materials and methods: C57BL/6J mice were total-body irradiated with an acute dose from X-rays (0.10 Gy) or carbon ions (0.05 or 0.10 Gy). The hippocampus was collected and the expression of 84 AD-related genes was analyzed. Morris water maze test was applied to the measurement of the learning ability and memory of the animals. Amyloid imaging with positron emission tomography were performed to detect the accumulation of fibrillary amyloid β peptide (Aβ), and characteristic pathologies of AD were examined with immunohistochemical staining of amyloid precursor protein (APP), Aβ, tau, and phosphorylated tau.Results: For the transcriptional studies, results showed that a few genes out of 84 AD-related genes were significantly up-regulated at 4 h after irradiation and the other genes had no marked change; on the other hand, a few other genes showed a significant down-regulation, while the other genes had no marked change at 1 year after irradiation. For the behavioral studies, no significant difference on learning ability and memory was observed at 1 and 2 years after irradiation. Imaging and immunohistochemical staining showed no change in the accumulation of fibrillar amyloid and the expression of APP, Aβ, tau and phosphorylated tau were detectable in the animals 4 months and 2 years after irradiation.Conclusion: These findings suggest that total-body irradiation at a dose of 0.10 Gy could hardly induce significant early or late transcriptional alterations in most of the AD-related genes in the hippocampus, learning disability and memory impairment, and AD-like pathological change in the brain in mice [ 3].

Low-dose total-body carbon-ion irradiations induce early transcriptional alteration without late Alzheimer's disease-like pathogenesis and memory impairment in mice.

The cause and risk factors of Alzheimer's disease (AD) are largely unknown. Studies on possible radiation-induced AD-like pathogenesis and behavioral consequences are important because humans are exposed to ionizing radiation (IR) from various sources. It was reported that total-body irradiations (TBI) at 10 cGy of low linear energy transfer (LET) X-rays to mice triggered acute transcriptional alterations in genes associated with cognitive dysfunctions. However, it was unknown whether low doses of IR could induce AD-like changes late after exposure. We reported previously that 10 cGy X-rays induced early transcriptional response of several AD-related genes in hippocampi without late AD-like pathogenesis and memory impairment in mice. Here, further studies on two low doses (5 or 10 cGy) of high LET carbonion irradiations are reported. On expression of 84 AD-related genes in hippocampi, at 4 hr after TBI, 5 cGy induced a significant upregulation of three genes (Abca1, Casp3, and Chat) and 10 cGy led to a marked upregulation of one gene (Chat) and a downregulation of three genes (Apoe, Ctsd, and Il1α), and, at 1 year after TBI, one gene (Il1α) was significantly downregulated in 10 cGy-irradiated animals. Changes in spatial learning ability and memory and induction of AD-like pathogenesis were not detected by in vivo brain imaging for amyloid-β peptide accumulation and by immunohistochemical staining of amyloid precursor protein, amyloid-β protein, tau, and phosphorylated tau protein. These findings indicate that low doses of carbon-ion irradiations did not cause behavioral impairment or AD-like pathological change in mice.

Total body 100-mGy X-irradiation does not induce Alzheimer's disease-like pathogenesis or memory impairment in mice

The cause and progression of Alzheimer's disease (AD) are poorly understood. Possible cognitive and behavioral consequences induced by low-dose radiation are important because humans are exposed to ionizing radiation from various sources. Early transcriptional response in murine brain to low-dose X-rays (100 mGy) has been reported, suggesting alterations of molecular networks and pathways associated with cognitive functions, advanced aging and AD. To investigate acute and late transcriptional, pathological and cognitive consequences of low-dose radiation, we applied an acute dose of 100-mGy total body irradiation (TBI) with X-rays to C57BL/6J Jms mice. We collected hippocampi and analyzed expression of 84 AD-related genes. Mouse learning ability and memory were assessed with the Morris water maze test. We performed in vivo PET scans with 11C-PIB, a radiolabeled ligand for amyloid imaging, to detect fibrillary amyloid beta peptide (Aβ) accumulation, and examined characteristic AD pathologies with immunohistochemical staining of amyloid precursor protein (APP), Aβ, tau and phosphorylated tau (p-tau). mRNA studies showed significant downregulation of only two of 84 AD-related genes, Apbb1 and Lrp1, at 4 h after irradiation, and of only one gene, Il1α, at 1 year after irradiation. Spatial learning ability and memory were not significantly affected at 1 or 2 years after irradiation. No induction of amyloid fibrillogenesis or changes in APP, Aβ, tau, or p-tau expression was detected at 4 months or 2 years after irradiation. TBI induced early or late transcriptional alteration in only a few AD-related genes but did not significantly affect spatial learning, memory or AD-like pathological change in mice.

Internal Jugular Vein Compression Mitigates Traumatic Axonal Injury in a Rat Model by Reducing the Intracranial Slosh Effect

Traumatic brain injury (TBI) remains a devastating condition for which extracranial protection traditionally has been in the form of helmets, which largely fail to protect against intracranial injury. To determine whether the pathological outcome after traumatic brain injury can be improved via slosh mitigation by internal jugular vein (IJV) compression. Two groups of 10 adult male Sprague-Dawley rats were subjected to impact-acceleration traumatic brain injury. One group underwent IJV compression via application of a collar before injury; the second group did not. Intracranial pressure and intraocular pressure were measured before and after IJV compression to assess collar performance. All rats were killed after a 7-day recovery period, and brainstem white matter tracts underwent fluorescent immunohistochemical processing and labeling of β-amyloid precursor protein, a marker of axonal injury. Digital imaging and statistical analyses were used to determine whether IJV compression resulted in a diminished number of injured axons. Compression of the IJV resulted in an immediate 30% increase in intraocular and intracranial pressures. Most notably, IJV compression resulted in > 80% reduction in the number of amyloid precursor protein-positive axons as indicated by immunohistochemical analysis. Using a standard acceleration-deceleration laboratory model of mild traumatic brain injury, we have shown successful prevention of axonal injury after IJV compression as indicated by immunohistochemical staining of amyloid precursor protein. We argue that IJV compression reduces slosh-mediated brain injury by increasing intracranial blood volume, which can be indirectly measured by intracranial and intraocular pressures.

Herpes Simplex Virus Dances with Amyloid Precursor Protein while Exiting the Cell

Herpes simplex type 1 (HSV1) replicates in epithelial cells and secondarily enters local sensory neuronal processes, traveling retrograde to the neuronal nucleus to enter latency. Upon reawakening newly synthesized viral particles travel anterograde back to the epithelial cells of the lip, causing the recurrent cold sore. HSV1 co-purifies with amyloid precursor protein (APP), a cellular transmembrane glycoprotein and receptor for anterograde transport machinery that when proteolyzed produces A-beta, the major component of senile plaques. Here we focus on transport inside epithelial cells of newly synthesized virus during its transit to the cell surface. We hypothesize that HSV1 recruits cellular APP during transport. We explore this with quantitative immuno-fluorescence, immuno-gold electron-microscopy and live cell confocal imaging. After synchronous infection most nascent VP26-GFP-labeled viral particles in the cytoplasm co-localize with APP (72.8+/−6.7%) and travel together with APP inside living cells (81.1+/−28.9%). This interaction has functional consequences: HSV1 infection decreases the average velocity of APP particles (from 1.1+/−0.2 to 0.3+/−0.1 µm/s) and results in APP mal-distribution in infected cells, while interplay with APP-particles increases the frequency (from 10% to 81% motile) and velocity (from 0.3+/−0.1 to 0.4+/−0.1 µm/s) of VP26-GFP transport. In cells infected with HSV1 lacking the viral Fc receptor, gE, an envelope glycoprotein also involved in viral axonal transport, APP-capsid interactions are preserved while the distribution and dynamics of dual-label particles differ from wild-type by both immuno-fluorescence and live imaging. Knock-down of APP with siRNA eliminates APP staining, confirming specificity. Our results indicate that most intracellular HSV1 particles undergo frequent dynamic interplay with APP in a manner that facilitates viral transport and interferes with normal APP transport and distribution. Such dynamic interactions between APP and HSV1 suggest a mechanistic basis for the observed clinical relationship between HSV1 seropositivity and risk of Alzheimer's disease.

Slowly Progressive Axonal Degeneration in a Rat Model of Chronic, Nonimmune-Mediated Demyelination

Axonal degeneration in the central nervous system (CNS) is associated with neurologic disability. In some diseases, it has been postulated that axonal degeneration may be caused by loss of trophic support normally provided by oligodendrocytes and myelin. To investigate this phenomenon, we studied axonal pathology in the taiep mutant rat, which develops nonimmune oligodendrocyte dysfunction and myelin loss. Using immunohistochemical analysis of several CNS regions, we show that accumulation of dephosphorylated neurofilaments occurs in taiep axons. These changes become more pronounced as myelin loss increases and characteristic spheroids, representing transected axons, become abundant. Amyloid precursor protein staining is increased in taiep white matter tracts, indicating abnormalities of axonal transport. These changes do not occur in wild type controls. Optic nerve counts demonstrate progressive axonal loss throughout the life of the rat; early axonal loss occurs in the context of dysmyelination; later axonal loss is likely to be related to chronic demyelination. The axonal pathology in the taiep rat provides evidence that CNS axonopathy may, in certain situations, be related to a loss of trophic support normally provided by cells of the oligodendrocyte lineage and/or myelin; this may occur in the absence of significant inflammation.

Temporal–spatial expression of presenilin 1 and the production of amyloid-β after acute spinal cord injury in adult rat

Regulated intramembrane proteolysis (RIP) is one of the signaling pathways mediating information transfer from the extracellular to the intracellular domain. γ-Secretase is an aspartyl protease of the RIP that cleaves the intramembrane region of type I integral membrane proteins, such as amyloid precursor protein (APP). Presenilin 1 (PS1) is the catalytic subunit of γ-secretase and PS1 mutations cause Alzheimer's disease, spastic paraplegia and spinal cord atrophy. The biological function of PS1 in the spinal cord has not been fully elucidated. Thus, to clarify the involvement of RIP in spinal cord injury, we examined the expression of PS1, APP and amyloid-β protein (Aβ) following rat spinal cord hemisection. Western blot analysis showed that PS1, APP and Aβ levels increased 1 day after spinal cord hemisection. Immunohistochemistry showed an increased number of PS1 immunopositive cells about 1 mm from the lesion site. PS1, APP and Aβ double staining with cell-type specific markers showed colocalization of PS1 with axons in the white matter of the lesioned side. These findings suggest that RIP signaling occurs following rat spinal cord injury. In the future, the control of RIP may offer a new strategy for the treatment of spinal cord injury.

Amyloid Precursor-like Protein 1 Influences Endocytosis and Proteolytic Processing of the Amyloid Precursor Protein

Ectodomain shedding of the amyloid precursor protein (APP) is a key regulatory step in the generation of the Alzheimer disease amyloid beta peptide (Abeta). The molecular mechanisms underlying the control of APP shedding remain little understood but are in part dependent on the low density lipoprotein receptor-related protein (LRP), which is involved in APP endocytosis. Here, we show that the APP homolog APLP1 (amyloid precursor-like protein 1) influences APP shedding. In human embryonic kidney 293 cells expression of APLP1 strongly activated APP shedding by alpha-secretase and slightly reduced beta-secretase cleavage. As revealed by domain deletion analysis, the increase in APP shedding required the NPTY amino acid motif within the cytoplasmic domain of APLP1. This motif is conserved in APP and is essential for the endocytosis of APP and APLP1. Unrelated membrane proteins containing similar endocytic motifs did not affect APP shedding, showing that the increase in APP shedding was specific to APLP1. In LRP-deficient cells APLP1 no longer induced APP shedding, suggesting that in wild-type cells APLP1 interferes with the LRP-dependent endocytosis of APP and there by increases APP alpha-cleavage. In fact, an antibody uptake assay revealed that expression of APLP1 reduced the rate of APP endocytosis. In summary, our study provides a novel mechanism for APP shedding, in which APLP1 affects the endocytosis of APP and makes more APP available for alpha-secretase cleavage.

Transformation of diffuse beta-amyloid precursor protein and beta-amyloid deposits to plaques in the thalamus following transient occlusion of the middle cerebral artery in rats

Cerebral ischemia leads to a transient upregulation and accumulation of -amyloid precursor protein (APP). For example, APP staining and expression are detected in the subcortical white matter and adjacent to the boundary of the ischemic lesion in the grey matter following transient occlusion of the middle cerebral artery (MCAO). Previous studies, however, have used only short survival periods ranging from days to weeks. The aim of the present study was to assess possible long-term accumulation of APP and A during a 9-month follow-up in rats subjected to transient MCAO. Male Wistar rats were subjected to transient middle cerebral artery occlusion (MCAO) for 2 hours. Sensorimotoric outcome was assessed using a tapered/ledged beam-walking task following operation. The distribution of APP and A was examined immunohistochemically at 1 week, 1 month, and 9 months after MCAO. Histologic analysis revealed severe corticostriatal damage in all MCAO rats. MCAO caused a long-lasting deficit in forelimb and hindlimb function assessed using the beam-walking test. In MCAO rats that survived 1 week, APP staining was present around the ischemic area, in the corpus callosum in crossing axons, in descending axons leaving the lesioned area, and in the terminal zone of these axons in the thalamus. There was staining for both N- and C-terminal APP. Similarly, there was positive A staining in all of these areas. After 1 month survival, some N- and C-terminal APP staining was present around the ischemic area, but most APP was present at the terminal zone of the deafferented axons in the thalamus. Most N-terminal APP was extracellular, in contrast to the C-terminal APP, which was predominantly intracellular. A was stained in a similar pattern in these areas; most A was extracellular, but some staining was present in axons. At 9 months following the MCAO, APP staining was not present around the lesioned area, in the corpus callosum, or in descending axons leaving the lesioned area. In the thalamus in the terminal zone of the deafferented corticothalamic axons, however, there were large, dense deposits, that resembled plaques and consisted of N-terminal APP. These deposits were also positively stained for A, but not for C-terminal APP. In conclusion, C- and N-terminal APP and A staining was present in areas adjacent to the infarct, the corpus callosum, and the thalamus for 1 week after MCAO. The staining of these proteins later disappeared from the cortical areas and white matter, but was still evident in the thalamus 9 months after MCAO. The N-terminal APP and A staining in the thalamus was diffuse acutely after the infarct, but accumulated, leading to dense plaque-like deposits in the ventroposterior lateral and ventroposterior medial nuclei, a finding that has not been reported previously. It is suggested that following focal cerebral ischemia APP and A are transported through corticothalamic axons and secreted at terminals where they form diffuse deposits, which over time develop into plaque-like structures.

Astroglial expression of the beta-amyloid in ischemia-reperfusion brain injury.

Vascular damage and reactive gliosis are found colocalized with amyloid deposits in Alzheimer's disease brain, suggesting that the cerebrovasculature may be a clinically relevant site of Alzheimer's disease pathology and may contribute to the neurodegeneration process in Alzheimer's disease. In ischemic conditions, amyloid precursor protein and amyloid peptide are reported to be upregulated in neurons and in extracellular space. Expression and distribution of amyloid precursor protein and amyloid peptide in astroglial cells were examined immunohistochemically after 10-min cardiac arrest. After reperfusion for 2, 7, and 14 days and 1, 6, 9, and 12 months, brains were immunostained. The indirect reactive astrocytes with fragments of the full-length amyloid precursor protein were observed in brains until 7 days postischemia. Direct immunoreactivity only of amyloid peptide and the C-terminal of amyloid precursor protein was also localized in the reactive astrocytes in ischemic brains at 6 months after ischemia. Ischemia temporarily induced amyloid peptide overexpression in reactive astrocytes, but this expression peaked at 7 days and 6 months. A glial appearance of amyloid peptide and C-terminal of amyloid precursor protein staining occurred when extensive neuronal loss and the onset of brain atrophy were evident. The localization of the C-terminal of amyloid precursor protein within ischemic astroglial cells underscores the likely importance of the C-terminal of amyloid precursor protein in the pathogenesis of ischemia as in Alzheimer's disease.

Uptake of HIV-1 tat protein mediated by low-density lipoprotein receptor-related protein disrupts the neuronal metabolic balance of the receptor ligands.

Neurological disorders develop in most people infected with human immunodeficiency virus type 1 (HIV-1). However, the underlying mechanisms remain largely unknown. Here we report that binding of HIV-1 transactivator (Tat) protein to low-density lipoprotein receptor-related protein (LRP) promoted efficient uptake of Tat into neurons. LRP-mediated uptake of Tat was followed by translocation to the neuronal nucleus. Furthermore, the binding of Tat to LRP resulted in substantial inhibition of neuronal binding, uptake and degradation of physiological ligands for LRP, including alpha2-macroglobulin, apolipoprotein E4, amyloid precursor protein and amyloid beta-protein. In a model of macaques infected with a chimeric strain of simian-human immunodeficiency virus, increased staining of amyloid precursor protein was associated with Tat expression in the brains of simian-human immunodeficiency virus-infected macaques with encephalitis. These results indicate that HIV-1 Tat may mediate HIV-1-induced neuropathology through a pathway involving disruption of the metabolic balance of LRP ligands and direct activation of neuronal genes.

Damage and repair of DNA in HIV encephalitis.

Neuronal damage and dementia are common sequelae of HIV encephalitis. The mechanism by which HIV infection of CNS macrophages results in neuronal damage is not known. We examined the brains from 15 AIDS autopsies (8 with HIV encephalitis and 7 without) and 4 non-infected control autopsies for the presence of DNA strand breaks, for associated changes in the expression of the DNA repair enzymes KU80 and Poly (ADP-ribose) polymerase (PARP), and for accumulation of amyloid precursor protein (APP). Abundant DNA damage was observed with terminal transferase-mediated dUTP nick end-labeling (TUNEL), however, there was no morphologic evidence of significant neuroglial apoptosis. The DNA repair enzyme KU80 was immunocytochemically detectable in neuronal and glial cells in autopsy brains from patients with and without HIV encephalitis; however, in cases with HIV encephalitis the staining was more prominent than in the infected or non-infected controls without encephalitis. There was no difference in KU80 immunostaining in oligodendroglia from autopsies with and without encephalitis. Immunostaining for PARP was more intense in gray and white matter of cases with HIV encephalitis. No clear spatial relationship existed between expression of DNA repair enzymes and the spatial proximity of microglial nodules or HIV-infected macrophages. The cytoplasm of cortical and subcortical neurons immunostained for APP Stronger cortical neuronal APP staining was observed in cases without HIV encephalitis. Staining of deep gray matter neurons was similar, irrespective of the presence or absence of encephalitis. While foci of intense APP staining were noted in white matter not related to HIV infection, they were associated with foci of opportunistic infections (e.g. due to CMV or PML). We conclude that damaged DNA and altered patterns of expression of DNA repair proteins and APP are common findings in the brains of AIDS patients at autopsy, but do not have a spatial relationship to HIV-infected macrophages.