Pengcheng Han

Pituitary adenylate cyclase-activating polypeptide protects against β-amyloid toxicity

Pituitary adenylate cyclase activating polypeptide (PACAP) is a neurotrophin. However, its role in human Alzheimers disease (AD) is largely unknown. We examined PACAP expression in postmortem human AD and triple transgenic mouse (3xTG, Psen1/APPSwe/TauP301L) brains. We established an in vitro model of primary neuronal cell culture to study the protective effects of PACAP against β-amyloid (Aβ) toxicity. We further studied the PACAP-Sirtuin 3 (Sirt3) pathway on mitochondrial function. PACAP expression was reduced in AD and 3xTG mouse brains. This reduction was inversely correlated with Aβ and tau protein levels. Treatment with PACAP effectively protected neurons against Aβ toxicity. PACAP stimulated mitochondrial Sirt3 production. Similar to PACAP, Sirt3 was reduced in AD and 3xTG brains. Knocking down Sirt3 compromised the neuroprotective effects of PACAP, and this was reversed by over-expressing Sirt3. PACAP is reduced in AD and may represent a novel therapeutic strategy.

Ketones block amyloid entry and improve cognition in an Alzheimer's model.

Sporadic Alzheimers disease (AD) is responsible for 60%-80% of dementia cases, and the most opportune time for preventive intervention is in the earliest stage of its preclinical phase. As traditional mitochondrial energy substrates, ketone bodies (ketones, for short), beta-hydroxybutyrate, and acetoacetate, have been reported to provide symptomatic improvement and disease-modifying activity in epilepsy and neurodegenerative disorders. Recently, ketones are thought as more than just metabolites and also as endogenous factors protecting against AD. In this study, we discovered a novel neuroprotective mechanism of ketones in which they blocked amyloid-β 42, a pathologic hallmark protein of AD, entry into neurons. The suppression of intracellular amyloid-β 42 accumulation rescued mitochondrial complex I activity, reduced oxidative stress, and improved synaptic plasticity. Most importantly, we show that peripheral administration of ketones significantly reduced amyloid burden and greatly improved learning and memory ability in a symptomatic mouse model of AD. These observations provide us insights to understand and to establish a novel therapeutic use of ketones in AD prevention.

Pituitary adenylate cyclase–activating polypeptide is reduced in Alzheimer disease

Objectives: There is growing evidence that pituitary adenylate cyclase–activating polypeptide (PACAP) is associated with Alzheimer disease (AD) pathology in animal models, but human studies are needed. Methods: We studied the brains of patients with pathologically confirmed late-onset AD and age-matched cognitively normal (CN) subjects to investigate the expression of PACAP messenger RNA (34 AD and 14 CN) and protein (12 AD and 11 CN) in a case-control study. Results: We report that PACAP levels are reduced in multiple brain regions, including the entorhinal cortex, the middle temporal gyrus, the superior frontal gyrus, and the primary visual cortex. This reduction is correlated with higher amyloid burden (CERAD plaque density) in the entorhinal cortex and superior frontal gyrus but not in the primary visual cortex, a region spared in most cases of AD. PACAP expression is lower in advanced Braak stages (V and VI) than in moderate stages (III and IV). Increased PACAP levels are associated with decreased scores on the Dementia Rating Scale, a global cognitive measure. Finally, CSF levels paralleled brain levels in AD but not in Parkinson dementia or frontotemporal dementia brains. Conclusions: The close relationship between PACAP reduction and the severity of AD pathology suggests that downregulation of PACAP may contribute to AD pathogenesis.

Association of pituitary adenylate cyclase-activating polypeptide with cognitive decline in mild cognitive impairment due to Alzheimer disease.

IMPORTANCE There is a deficit of pituitary adenylate cyclase-activating polypeptide (PACAP) in patients with neuropathologically confirmed Alzheimer dementia. However, whether this deficit is associated with the earlier stages of Alzheimer disease (AD) is unknown. This study was conducted to clarify the association between PACAP biomarkers and preclinical, mild cognitive impairment (MCI), and dementia stages of AD in postmortem brain tissue. OBJECTIVES To examine PACAP and PACAP receptor levels in postmortem brain tissues and cerebrospinal fluid from cognitively and neuropathologically normal control individuals, patients with MCI due to AD (MCI-AD), and individuals with AD; analyze the relationship between PACAP, cognitive, and pathologic features; and propose a model to assess these relationships. DESIGN, SETTING, AND PARTICIPANTS We measured PACAP and its receptor (PAC1) levels using enzyme-linked immunoassay. A total of 35 cases were included. All the brain tissue and cerebrospinal fluid samples were selected from Banner Sun Health Research Institute Brain and Body Donation Program. All cognitive test results were in record with the Arizona Alzheimers Consortium. MAIN OUTCOMES AND MEASURES A comparison of PACAP and PAC1 levels among the healthy controls, MCI-AD, and AD dementia groups, as well as a systematic correlation analysis between PACAP level, cognitive performance, and pathologic severity. RESULTS The PACAP levels in cerebrospinal fluid, the superior frontal gyrus, and the middle temporal gyrus were inversely related to dementia severity. The PACAP levels in cerebrospinal fluid correlated with the Mattis Dementia Rating Scale score (Pearson r = 0.50; P = .03) and inversely correlated with total amyloid plaques (Pearson r = -0.48; P < .01) and tangles (Pearson r = -0.55; P = .01) in the brain. The PACAP in the superior frontal gyrus and middle temporal gyrus correlated with the Stroop Color-Word Interference Test (Pearson r = 0.58; P < .01) and the Auditory Verbal Learning Test-Total Learning (Pearson r = 0.33; P = .02), respectively. The PACAP in the primary visual cortex did not correlate with the Judgment of Line orientation test (P = .14). Furthermore, the PAC1 level in the superior frontal gyrus showed an upregulation in MCI-AD but not in AD. The pharmacodynamic model of the PACAP-PAC1 interaction best predicted cognitive function in the superior frontal gyrus, but it was less predictive in the middle temporal gyrus and failed to be predictive in the primary visual cortex. CONCLUSIONS AND RELEVANCE Deficits in PACAP are associated with clinical severity in the MCI and dementia stages of AD. Additional studies are needed to clarify the role of PACAP deficits in the predisposition to, pathogenesis of, and treatment of AD.

Pertussis toxin reduces calcium influx to protect ischemic stroke in a middle cerebral artery occlusion model.

Increased calcium influx secondary to glutamate induced excitotoxicity initiates and potentiates devastating pathological changes following ischemic stroke. Pertussis toxin (PTx), a G‐protein blocker, is known to suppress intracellular calcium accumulation. We hypothesize that PTx can protect against stroke by blocking calcium influx. In a permanent middle cerebral artery occlusion model, PTx (1000 ng) was given intraperitoneally 30 min after inducing stroke. Magnetic Resonance Imaging of perfusion and T2‐weighted brain scans were obtained to evaluate cerebral blood flow (CBF) and infarct volume. Primary neuronal culture was used to test glutamate induced excitotoxicity and calcium influx. We established a non‐linear exponential curve model to minimize variations in animal cerebrovasculature. A reduction of 40–60% in relative CBF was a critical window where infarct volume started to increase as rCBF reduced. PTx showed maximal effects in reducing infarct volume at this window. In vitro studies further demonstrated PTx increased neuronal cell survival by decreasing glutamate‐induced calcium influx into neurons and preventing neurons from apoptosis. PTx salvages the ischemic penumbra by blocking calcium influx. This provides us a new mechanism upon which experimental therapies can be explored to treat ischemic stroke.

Ischemia-induced Neuronal Cell Death Is Mediated by Chemokine Receptor CX3CR1

The chemokine fractalkine (CX3CL1) and its receptor CX3CR1 play a fundamental role in the pathophysiology of stroke. Previous studies have focused on a paracrine interaction between neurons that produce fractalkine and microglia that express CX3CR1 receptors in the central nervous system. Recent findings have demonstrated the functional expression of CX3CR1 receptors by hippocampal neurons, suggesting their involvement in neuroprotective and neurodegenerative actions. To elucidate the roles of neuronal CX3CR1 in neurodegeneration induced by ischemic stroke, a mouse model of permanent middle cerebral artery occlusion (pMCAO) was employed. In the pMCAO mice, increased CX3CR1 levels, apoptosis-associated morphological changes, and Caspase 3-positive neuronal cells were observed in the striatum and in the hippocampus 24 hours after occlusion. Upregulation of CX3CR1 in ischemic neurons is associated with neuronal apoptotic cell death. In contrast, ischemia-induced apoptotic neuronal cell death was decreased in CX3CR1 deficient mice. Cultured primary hippocampal neurons obtained from CX3CR1 deficient mice were more resistant to glutamate-induced excitotoxicity by blocking calcium influx than those from wild-type mice. For the first time, we reported that neuronal CXCR1 mediates neuronal apoptotic cell death in ischemia. Our results suggest that modulating CXCR1 activity offers a novel therapeutic strategy for stroke.

Pertussis toxin attenuates experimental autoimmune encephalomyelitis by upregulating neuronal vascular endothelial growth factor.

We have reported earlier that pertussis toxin (PTx) attenuates the motor deficits in experimental autoimmune encephalomyelitis (EAE), an animal model for human multiple sclerosis. PTx protects neurons from inflammatory insults. Vascular endothelial growth factor (VEGF) is also neuroprotective. However, the effect of PTx on VEGF has never been studied. We investigated whether PTx modulates neuronal VEGF expression and how it affects the pathogenesis of EAE. EAE was induced by injecting myelin oligodendrocyte glycoprotein 35–55 peptides with adjuvants into C57BL/6 mice. Clinical scores of EAE were evaluated daily for 19 days. Brain and spinal cord samples were collected and assessed for inflammation and demyelination. VEGF, NeuN for neurons, and Caspase-3 for apoptosis were stained for localization using immunohistochemistry techniques, followed by western blot analysis for quantification. Primary neurons were cultured to assess the direct effect of PTx on neuronal VEGF expression. PTx treatment increases neuronal VEGF expression by up to ∼75% in vitro and ∼60% in vivo, preventing neurons from apoptosis. This leads to resolution in inflammation and remyelination and amendment in motor deficits. Our findings suggest that upregulation of endogenous neuronal VEGF by PTx protects motor deficits in EAE and it is a potential therapeutic option for multiple sclerosis.

THE EFFECT OF SIRTUIN 3 ON TAU ACETYLATION IN ALZHEIMER'S DISEASE

Background:Proteomic studies have identified many potential proteins associated with early diagnosis, differential diagnosis, prognosis and therapeutic response in diverse diseases. Recent proteomic analysis of Alzheimer’s disease (AD) plaques has identified a novel amyloid associated protein, secernin-1, which is a 50 kDa cytosolic protein (Acta Neuropathol 133: 933-954, 2017). Secernin-1 is known to regulate exocytosis in mast cells and increases both the extent of secretion and the sensitivity of mast cells to stimulation with calcium. It has been identified as a novel immunotherapy target for gastric cancer and a novel marker for prognosis in colorectal cancer. It is highly expressed in brain tissue and has dipeptidase activity. However, secernin-1 had not been previously known to be associated with neurodegenerative disorders.Methods: The presence of neuritic amyloid plaques and intracellular neurofibrillary tangles are hallmarksofADHowever the soluble oligomersof amyloid beta (Ab) and hyperphosphorylated tau are thought to be the major toxic species in AD. To determine the potential role of secernin-1 in AD neurodegeneration we initially performed in vitro binding experiments of human secernin-1 to theAbpeptide and to recombinant human Tau-His tag. These two AD proteins were polymerized by glutaraldehyde treatment to form stable oligomers. Binding was assessed by western blot analysis with mouse monoclonal antibodies to Ab (6E10, epitope residues 1-16; 4G8, epitope residues 17-24), Tau and rabbit polyclonal antiHuman secernin-1.Results:Surprisingly, the anti-secernin-1 antibody immunolabeled oligomers of Ab and tau at approximately their original molecular weight. There were no discrete higher molecular weight positive bands detected by any antibody that would correspond to a complex of Ab or tau oligomer and intact secernin-1. Therefore only a small fragment of secernin-1 may be complexedwith Ab and Tau oligomers. These complexes are being processed and analyzed by label-free quantitative LC–MS/MS analysis. Experiments with AD brain tissue extracts of Ab and tau oligomers are in progress. Conclusions: This secernin-1 peptide may be a novel marker for detection, prognosis and therapeutic evaluation in Alzheimer’s disease.