Helena Passarelli Giroud Joaquim

Increased platelet GSK3B activity in patients with mild cognitive impairment and Alzheimer's disease

The disruption of glycogen synthase kinase 3-beta (GSK3B) homeostasis has implications in the pathophysiology of neuropsychiatric disorders, namely Alzheimers disease (AD). GSK3B activity is increased within the AD brain, favoring the hyperphosphorylation of microtubule-associated protein Tau and the formation of neurofibrillary tangles. Such abnormality has also been detected in leukocytes of patients with cognitive disorders. The aim of the present study was to determine the expression of total and phosphorylated GSK3B at protein level in platelets of older adults with varying degrees of cognitive impairment, and to compare GSK3B activity in patients with AD, mild cognitive impairment (MCI) and healthy controls. Sixty-nine older adults were included (24 patients with mild to moderate AD, 22 patients with amnestic MCI and 23 elderly controls). The expression of platelet GSK3B (total- and Ser-9 phosphorylated GSK3B) was determined by Western blot. GSK3B activity was indirectly assessed by means of the proportion between phospho-GSK3B to total GSK3B (GSK3B ratio), the former representing the inactive form of the enzyme. Ser-9 phosphorylated GSK3B was significantly reduced in patients with MCI and AD as compared to controls (p=0.04). Platelet GSK3B ratio was significantly decreased in patients with MCI and AD (p=0.04), and positively correlated with scores on memory tests (r=0.298, p=0.01). In conclusion, we corroborate previous evidence of increased GSK activity in peripheral tissues of patients with MCI and AD, and further propose that platelet GSK may be an alternative peripheral biomarker of this abnormality, provided samples are adequately handled in order to preclude platelet activation.

Platelet GSK3B activity in patients with late-life depression: marker of depressive episode severity and cognitive impairment?

Abstract Objective. Increased GSK3B activity has been reported as a state marker of major affective episodes in patients with depression and bipolar disorder. No study so far has addressed GSK3B activity in late-life depression. The aims of the present study were to determine GSK3B activity in platelets of elderly patients with major depression, and the association between GSK3B activity and the severity of depressive symptoms and cognitive impairment. Methods. Forty drug-free elderly patients with major depressive episode were compared to healthy older adults (n = 13). Severity of the depressive episode and current cognitive state were determined by the Hamilton Depression Scale (HAM-D) and the Cambridge Cognitive Test (CAMCOG), respectively. Total- and ser-9-phosphorylated GSK3B (tGSK3B and pGSK3B) were determined in platelets by enzyme immunometric assays (EIA). GSK3B activity was indirectly inferred by the GSK3B ratio (i.e. pGSK3B/tGSK3B). Results. Elderly depressed patients had significantly lower pGSK3B levels (P = 0.03) and GSK3B ratio (P = 0.03), indicating higher GSK3B activity. Higher GSK3B activity were observed in patients with severe depressive episode (HAM-D scores >22, P = 0.03) and with cognitive impairment (CAMCOG scores <86, P = 0.01). Conclusion. The present findings provide additional evidence of the involvement of GSK3B in the pathophysiology of late-life major depression. Higher GSK3B activity may be more relevant in those patients with more severe depressive symptoms and cognitive impairment.

Hippocampal serotonin depletion is related to the presence of generalized tonic–clonic seizures, but not to psychiatric disorders in patients with temporal lobe epilepsy

OBJECTIVE Previous studies suggest that concentration of serotonin ([5-HT]) plays a pathogenic role in various types of epilepsy inhibiting seizures. However, most have not considered the clinical variables of epilepsy, and all of these studies included small and heterogeneous samples with refractory epilepsy, regardless of etiology. In this work, we measured [5-HT]s in hippocampal tissues from a large series of patients with refractory temporal lobe epilepsy caused by hippocampal sclerosis who underwent epilepsy surgery and evaluated the relationship between [5HT] and epilepsy-related clinical variables and psychiatric disorders. METHODS We included 44 patients with refractory unilateral TLE-HS who underwent surgical treatment for epilepsy. Hippocampal samples were collected, and serotonin concentrations were measured with high-pressure liquid chromatography (HPLC). RESULTS Lower [5-HT]s were correlated with a history of GTC seizures (Students t-test: p 0.041). There were no differences in [5-HT]s according to the other clinical variables and the presence of psychiatric disorders. SIGNIFICANCE Our findings demonstrated that serotonin depletion in the hippocampus play an important role in some aspects of the severity of epilepsy (i.e., the presence of GTC seizures) in a homogeneous sample of patients with refractory temporal lobe epilepsy determined by hippocampal sclerosis, but not with the presence of psychiatric disorders.

DONEPEZIL MODULATES ADAM10 AND BACE1 EXPRESSION IN PLATELETS OF ALZHEIMER'S DISEASE PATIENTS

brain. To investigate this, we are characterising the effects of CD2AP knockdown in an in vitro model of the BBB. Methods: Endogenous CD2AP expression was knocked down in H4 and hMEC/D3 cells using siRNA. Knockdown was quantified by western blot and Image J analysis. Levels of APP, Ab, and APP metabolites were quantified using ELISAs. The rate of receptor-mediated endocytosis was measured by a transferrin uptake assay. Co-localisation of CD2AP with APP and endosomal markers was assessed using immunocytochemistry. Transcytosis in hMEC/D3 cells will be assessed using Transwell assays. Results:We observed a significant 30.2760.07 % increase in Ab40 expression (p1⁄40.0046). Expression levels of APP and APP processing metabolites was not significantly altered. Results from our experiments in progress will be made available at AAIC 2016. Conclusions:Knockdown of CD2AP expression resulted in a significant increase in extracellular amyloid-B (Ab) levels, which did not appear to be directly related to changes in APP processing. This study is ongoing and novel data will be presented at AAIC 2016. Overall, our study provides valuable insights into how CD2AP contributes to LOAD susceptibility.

Higher proportion of inactive Gsk3β in platelets of elderly patients with bipolar disorder: an effect of treatment?

OBJECTIVE It has been postulated that mood stabilizers inhibit glycogen synthase kinase 3-beta (Gsk3β) activity, mainly through its phosphorylation on serine-9 (Ser9). However, in vivo studies addressing Gsk3β activity in patients with bipolar disorder are scarce. Here, we compare Gsk3β inactivation (as indicated by Ser9-phosphorylation) in platelets of elderly patients with bipolar disorder undergoing clinical treatment and healthy elderly adults not taking medication. METHODS Platelet samples were obtained from 37 elderly adults (bipolar disorder = 19, controls = 18). Relative changes in Gsk3β inactivation was estimated by comparing the ratios of phosphorylated Gsk3β to total Gsk3β (p-Gsk3β Ser9/Gsk3β) between the disease and control groups. RESULTS Phosphorylated-Gsk3β (p < 0.001) and the p-Gsk3β Ser9/Gsk3β ratio (p = 0.006) were elevated in bipolar patients. In the bipolar disorder group, p-Gsk3β Ser9/Gsk3β was positively correlated with serum lithium levels (r = 0.478, p = 0.039). CONCLUSIONS Gsk3β inactivation is higher in this group of elderly adults undergoing treatment for bipolar disorder. However, whether the treatment or the disease causes Gsk3β inactivation was confounded by the lack of an unmedicated, bipolar control group and the non-uniform treatment regimens of the bipolar disorder group. Thus, further studies should help distinguish whether Gsk3β inactivation is an effect of drug treatment or an intrinsic characteristic of bipolar disorder.

Protein Expression of BACE1 is Downregulated by Donepezil in Alzheimer’s Disease Platelets

BACKGROUND Abnormal amyloid-β protein precursor (AβPP) metabolism is a key feature of Alzheimers disease (AD). Platelets contain most of the enzymatic machinery required for AβPP processing, and correlates of intracerebral abnormalities have been demonstrated in platelets of patients with AD. Thus, AβPP-related molecules in platelets may be regarded as peripheral markers of AD. OBJECTIVE We sought to determine the protein expression of the AβPP secretases (ADAM10, BACE1, and PSEN1) and AβPP ratio in platelets of patients with mild or moderate AD compared to healthy controls. We further determined whether the protein expression of these markers might be modified by chronic treatment with donepezil. METHODS Platelet samples were obtained from patients and controls at baseline and after 3 and 6 months of continuous treatment with therapeutic doses of donepezil. The protein expression of platelet markers was determined by western blotting. RESULTS AD patients had a significant decrease in AβPP ratio, ADAM10, and PSEN1 compared to controls at baseline, but these differences were not modified by the treatment. Nonetheless, a significant reduction in the protein expression of BACE1 was observed in patients treated with donepezil for 6 months. CONCLUSION Our results corroborate previous findings from our group and others of decreased AβPP ratio and protein expression of ADAM10 in AD. We further show that PSEN1 is decreased in AD platelets, and that the protein expression of BACE1 is downregulated by chronic treatment with donepezil. This effect may be interpreted as evidence of disease modification.

Increased platelet glycogen sysnthase kinase 3beta in first-episode psychosis

Past studies have linked intracellular pathways related to psychotic disorders to the GSK3B enzyme. This study aimed to investigate GSK3B protein expression and phosphorylation in drug-naïve first-episode psychosis patients (n=43) at baseline and following symptom remission, and in healthy controls (n=77). At baseline GSK3B total level was higher in patients (p<0.001). In schizophrenia spectrum patients (n=25) GSK3B total and phosphorylated levels were higher than in controls and patients with other non-affective psychotic disorders (n=18) (p<0.001; p=0.027; p=0.05 respectively). No enzyme changes were found after clinical remission. The implication of this finding for the biology of psychoses warrants further studies to clarify whether increased GSK3B may be useful as a biomarker for psychosis in general, and schizophrenia in particular.

5-hydroxytryptamine1A receptor density in the hippocampus of patients with temporal lobe epilepsy is associated with disease duration

To date, no study has evaluated the association between serotonin receptor density and clinical variables in patients with temporal lobe epilepsy caused by hippocampal sclerosis (TLE‐HS) using hippocampal tissue. We evaluated 5‐hydroxytryptamine1A receptor (5‐HT1AR) density in hippocampal tissue from patients with TLE‐HS.

LIPID METABOLITE PLASMA LEVELS AS BIOMARKERS OF EARLY-ONSET ALZHEIMER DISEASE

Alana Caroline Costa, Helena Passarelli Giroud Joaquim, Wagner Farid Gattaz, Leda Leme Talib, Laboratory of Neuroscience (LIM-27), Institut of Psychiatry, Faculty of Medicine, S~ao Paulo, Brazil; Laboratory of Neuroscience (LIM-27), Institut of Psychiatry, Faculty of Medicine, Sao Paulo, Brazil; University of S~ao Paulo, S~ao Paulo, Brazil; Laboratory of Neuroscience (LIM-27), Department and Institute of Psychiatry, Faculty of Medicine, University of S~ao Paulo, Sao Paulo, Brazil. Contact e-mail: alanacosta@usp.br

PLATELET APP RATIO IN AD PATIENTS TREATED WITH DONEPEZIL

amyloid-b 40 with GAPDH aggregates revealed the significant pyramidal cell death and inflammation in the field of CA3, concomitant with the mitochondrial dysfunction. Conclusions: These findings suggest that GAPDH aggregates might act as a seed for amyloid-b 40 amyloidogenesis and enhances the cell death both in vitro and in vivo, providing a novel insight to investigate mechanisms underlying formation of amyloidal deposits such as senile and dendritic plaques in Alzheimer disease.