Franz Müller-Spahn

Reduced hippocampal MT2 melatonin receptor expression in Alzheimer's disease

Abstract:  The aim of the present study was to identify the distribution of the second melatonin receptor (MT2) in the human hippocampus of elderly controls and Alzheimers disease (AD) patients. This is the first report of immunohistochemical MT2 localization in the human hippocampus both in control and AD cases. The specificity of the MT2 antibody was ascertained by fluorescence microscopy using the anti‐MT2 antibody in HEK 293 cells expressing recombinant MT2, in immunoblot experiments on membranes from MT2 expressing cells, and, finally, by immunoprecipitation experiments of the native MT2. MT2 immunoreactivity was studied in the hippocampus of 16 elderly control and 16 AD cases. In controls, MT2 was localized in pyramidal neurons of the hippocampal subfields CA1‐4 and in some granular neurons of the stratum granulosum. The overall intensity of the MT2 staining was distinctly decreased in AD cases. The results indicate that MT2 may be involved in mediating the effects of melatonin in the human hippocampus, and this mechanism may be heavily impaired in AD.

Red wine ingredient resveratrol protects from β-amyloid neurotoxicity

Background: β-Amyloid peptide (Aβ), a neutrotoxic substance, has been implicated to a great degree in cell death during the course of AD. Resveratrol, a natural polyphenol mainly found in red wine, has been shown to be cardioprotective and chemoprotective. Since a moderate wine intake correlates with a lower risk for Alzheimer disease (AD), an additional neuroprotective effect has been postulated for resveratrol. Objective: The present study aimed at elucidating the possible neuroprotective effects of resveratrol against Aβ-induced neurotoxicity. Methods: The neuroprotective capacity against Aβ-related oxidative stress was studied in a cell culture model suitable for studying such potentially neuroprotective substances. Results: Resveratrol maintains cell viability and exerts an anti-oxidative action by enhancing the intracellular free-radical scavenger glutathione. Conclusion: Our findings suggest that red wine may be neuroprotective through the actions of resveratrol.

Cortical alterations of angiotensin converting enzyme, angiotensin II and AT1 receptor in Alzheimer’s dementia

We investigated the immunohistochemical alterations of angiotensin converting enzyme (ACE), angiotensin II and AT1 receptor in the parietal cortex in Alzheimers dementia (AD) to reveal the contributive role of the brain renin-angiotensin system in the disease process. In controls, ACE, angiotensin II and AT1 immunoreactivities were localized to pyramidal neurons of the cortex. The staining intensity was distinctly increased in AD for all three antigens, involving predominantly cortical layer V, which may reflect the enhanced brain renin-angiotensin system activity in the disease process. In addition, a prominent perivascular ACE and angiotensin II immunoreactivity surrounding some cortical vessels in aged controls and AD patients points to an underlying microvascular pathology in the process of neurodegeneration.

Age Dependency of Changes in Cerebral Hemoglobin Oxygenation during Brain Activation: A Near-Infrared Spectroscopy Study

We used near-infrared spectroscopy (NIRS) to study noninvasively the influence of aging on changes in the local concentration of oxygenated hemoglobin ([HbO2]), reduced hemoglobin ([HbR]), and total hemoglobin ([HbT] = [HbR] + [HbO2]) during activation of brain function. Young subjects (n = 12; age, 28 ± 4 years) performing calculation tasks showed an increase in [HbO2] [mean (arbitrary units) ± SD, 2.36 ± 1.07] and [HbT] (2.24 ± 1.13) in the frontal cortex, while [HbR] (−0.11 ± 0.48) decreased. Elderly subjects (n = 17; age, 52 ± 10 years) showed a significantly lower mean increase (p < 0.05) in [HbO2] and [HbT] levels (1.21 ± 1.38 and 0.72 ± 1.41, respectively). Regression analysis supports the hypothesis of an age-dependent decline in the activation-induced local increase in [HbO2] (y = −0.241x + 20.062; r = −0.431, p < 0.05) as well as [HbT] (y = −0.346x + 22.496; r = −0.568, p < 0.05). We conclude that NIRS is a promising approach for studying changes in Hb oxygenation during brain activation in physiological aging.

Amyloid-beta Leads to Impaired Cellular Respiration, Energy Production and Mitochondrial Electron Chain Complex Activities in Human Neuroblastoma Cells

Evidence suggests that amyloid-beta (Aβ) protein is a key factor in the pathogenesis of Alzheimer’s disease (AD) and it has been recently proposed that mitochondria are involved in the biochemical pathway by which Aβ can lead to neuronal dysfunction. Here we investigated the specific effects of Aβ on mitochondrial function under physiological conditions. Mitochondrial respiratory functions and energy metabolism were analyzed in control and in human wild-type amyloid precursor protein (APP) stably transfected human neuroblastoma cells (SH-SY5Y). Mitochondrial respiratory capacity of mitochondrial electron transport chain (ETC) in vital cells was measured with a high-resolution respirometry system (Oxygraph-2k). In addition, we determined the individual activities of mitochondrial complexes I–IV that compose ETC and ATP cellular levels. While the activities of complexes I and II did not change between cell types, complex IV activity was significantly reduced in APP cells. In contrast, activity of complex III was significantly enhanced in APP cells, as compensatory response in order to balance the defect of complex IV. However, this compensatory mechanism could not prevent the strong impairment of total respiration in vital APP cells. As a result, the respiratory control ratio (state3/state4) together with ATP production decreased in the APP cells in comparison with the control cells. Chronic exposure to soluble Aβ protein may result in an impairment of energy homeostasis due to a decreased respiratory capacity of mitochondrial electron transport chain which, in turn, may accelerate neurons demise.

Impaired Cu/Zn-SOD activity contributes to increased oxidative damage in APP transgenic mice.

Oxidative stress plays an important role in the pathogenesis of Alzheimers disease. To determine which mechanisms cause the origin of oxidative damage, we analyzed enzymatic antioxidant defense (Cu/Zn-superoxide dismutase Cu/Zn-SOD, glutathione peroxidase GPx and glutathione reductase GR) and lipid peroxidation products malondialdehyde MDA and 4-hydroxynonenal HNE in two different APP transgenic mouse models at 3-4 and 12-15 months of age. No changes in any parameter were observed in brains from PDGF-APP695(SDL) mice, which have low levels of Abeta and no plaque load. In contrast, Thy1-APP751(SL) mice show high Abeta accumulation with aging and plaques from an age of 6 months. In brains of these mice, HNE levels were increased at 3 months (female transgenic mice) and at 12 months (both gender), that is, before and after plaque deposition, and the activity of Cu/Zn-SOD was reduced. Interestingly, beta-amyloidogenic cleavage of APP was increased in female Thy1-APP751(SL) mice, which also showed increased HNE levels with simultaneously reduced Cu/Zn-SOD activity earlier than male Thy1-APP751(SL) mice. Our results demonstrate that impaired Cu/Zn-SOD activity contributes to oxidative damage in Thy1-APP751(SL) transgenic mice, and these findings are closely linked to increased beta-amyloidogenic cleavage of APP.

Soluble beta-amyloid leads to mitochondrial defects in amyloid precursor protein and tau transgenic mice

Background: Mitochondrial dysfunction has been identified in neurodegenerative disorders including Alzheimer’s disease, where accumulation of β-amyloid (Aβ) and oxidative stress seem to play central roles in the pathogenesis, by probably directly leading to mitochondrial dysfunction. Objective: In order to study the in vivo effect of Aβ load during aging, we evaluated the mitochondrial function of brain cells from transgenic mice bearing either mutant amyloid precursor protein (tgAPP) or mutant amyloid precursor protein and mutant PS1 (tgAPP/PS1) as well as from nontransgenic wild-type littermates. tgAPP mice exhibit onset of Aβ plaques at an age of 6 months, but the intracellular soluble Aβ load is already increased at 3 months of age. In contrast, onset of Aβ plaques starts at an age of 3 months in tgAPP/PS1 mice. In addition, we investigated the effects of different Aβ preparations on mitochondrial function of brain cells from tau transgenic mice. Results: Of note, mitochondrial damage such as reduced mitochondrial membrane potential and ATP levels can already be detected in the brains from these mice before the onset of plaques. In agreement with our findings in tgAPP mice, soluble Aβ induced mitochondrial dysfunction in brain cells from tau transgenic mice. Conclusion: Our results indicate that mitochondrial dysfunction is exacerbated by the presence of soluble Aβ species as a very early event during pathogenesis.

Increased melatonin 1a-receptor immunoreactivity in the hippocampus of Alzheimer's disease patients.

The pineal secretory product melatonin has, in addition to regulating retinal, circadian and vascular functions, neuroprotective effects. Blood melatonin levels are often decreased in Alzheimers disease (AD), a progressively disabling neurodegenerative disorder. In this study we provide the first immunohistochemical evidence for the localization of melatonin 1a‐receptor (MT1) in aged human hippocampus and a comparison of AD cases. MT1 was localized to pyramidal neurons in the hippocampal cornu ammonis (CA)1‐4 subfields. There was a distinct increase in staining intensity in all AD cases indicating an up‐regulation of the receptor, possibly as a compensatory response to impaired melatonin levels in order to augment melatonins neuroprotective effects.

Decreased trkA neurotrophin receptor expression in the parietal cortex of patients with Alzheimer's disease

The cholinergic neurons of the basal forebrain system are sensitive to nerve growth factor (NGF), a member of the neurotrophin gene family. Since the cholinergic system is affected early in the course of Alzheimers disease (AD), it was hypothesized that a deficit in NGF, e.g. reduced neurotrophin uptake by specific receptors, may play a role in neuronal cell death in AD. We quantitated mRNA levels of neurotrophins (NGF, BDNF, NT-3, NT-4/5) and their receptors (trkA, trkB, trkC, p75) in AD postmortem parietal cortex (n = 16) and cerebellum (n = 11). We applied highly sensitive reverse transcription-polymerase chain reaction (RT-PCR) in rapid autopsy derived brain tissue (mean postmortem delay 147+/-96 min., n = 53) to minimize postmortem mRNA variations. In the AD parietal cortex trkA mRNA levels were more than two times lower as compared to controls (n = 16, mean+/-SEM 0.26+/-0.07 units/S12, range, 0-1.78, and n = 11, 0.59+/-0.10 units/S12, range, 0.17-1.10, respectively, P = 0.015). TrkA mRNA levels did not appear to be altered in the AD cerebellum as compared to normal human cerebellum. NGF, BDNF, NT-3, NT-4/5, as well as trkB, trkC and p75 mRNA levels were unchanged in AD parietal cortex and cerebellum as compared to controls. This finding suggests that a reduced expression of the trkA receptor may contribute to impaired NGF-trkA signalling and a reduced transport of NGF in cholinergic neurons. These results reveal a central specific role of the high affinity NGF receptor during neurodegeneration in AD.

Treatment with the selective muscarinic ml agonist talsaclidine decreases cerebrospinal fluid levels of Aβ42 in patients with Alzheimer's disease

The amyloid p-peptides Aβ40 and Aβ42 are highly amyloidogenic constituents of brain Pamyloid plaques in Alzheimers disease (AD). Lowering their formation may be achieved by modulating the activities of proteases that cleave the amyloid precursor protein (AβPP), including α-β-, and γ-secretases. Talsaclidine is a functionally selective muscarinic ml agonist that stimulates non-amyloidogenic a-secretase processing in vitro. We compared cerebrospinal fluid (CSF) levels of Aβ40 and Aβ42 measured by ELISA before and at the end of 4 weeks of treatment with talsaclidine. The medication was administered in a double- blind, placebo-controlled, and randomized clinical study to 40 patients with AD. Talsaclidine (n=34) decreased CSF levels of Aβ42 by a median of 19% (p < 0.001) as compared to baseline. The mean diflerence between CSF levels of Aβ42 before and after treatment with talsaclidine (n=34) was -46.73 (SD) pg/ml as compared to 0.8 (SD)pg/ml with placebo (n=6) (p < 0.05). CSF levels of Aβ40 increased during treatement with placebo (n=6) while they remained stable during treatment with talsaclidine (n=31) (1.118±1.710 ng/ ml, and-0.170A0.967 ng/ml, respectively; p < 0.05). These data show that treatment with the ml agonist talsaclidine reduced Aβ peptides, and particularly Aβ42, in AD patients, suggesting it as a potential amyloid lowering therapy of AD.