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Lithium increases mitochondrial function in cultured hippocampal neurons
S670 P3-483
Poster Presentations P3 INTRAVENOUS IMMUNOGLOBULIN BINDS TO TAU, pTAU AND PrPc IN ADDITION TO Ab
Alexander Schaub1, Alain Kropf1, Susann Cattepoel1, Miriam Ender1, Reinhard Bolli1, Louis Fabri2, Sylvia Miescher1, 1CSL Behring AG, Bern, Switzerland; 2CSL Limited, Parkville, Australia. Background: Intravenous Immunoglobulin (IVIG) is being used in phase III clinical studies to assess its efficacy in mild-to-moderate Alzheimer’s disease (AD) patients. Beside beneficial effects in smaller scale clinical settings, the rationale for using IVIG in AD is based on observations of reduced levels of anti-amyloid beta (Aß)antibodies in AD patients and the presence of these antibodies in normal IVIG preparations (R. Dodel, J Neurol 2004; N Relkin, Neurobiol Aging, 2009). We examined whether PrivigenÒ(10% IVIG product) contains specific antibodies that bind to proteins implicated in AD pathology that might explain mechanisms of action in addition to the direct binding of Aß. IVIG is hypothesized to mediate its beneficial effects in chronic inflammatory and autoimmune conditions by a variety of mechanisms. As this might also be the case for AD we measured PrivigenÒbinding to Tau, phosphorylated Tau (pTau) and the cellular prion protein (PrPc). Methods: The binding of PrivigenÒto antigens was measured by ELISA. Differences in IVIG binding were observed during method development as a result of both antibody and blocking solutions that were used. Results: We show herein that PrivigenÒ binds to Aß, as well as tau, ptau and PrPc, but not a-synuclein under the same experimental conditions. We also demonstrate that the influence of blocking solutions and antibody diluents on natural auto antibodies is greater than on monoclonal or high titre antibodies. Conclusions: The interaction of PrivigenÒ with additional antigens (tau, ptau, PrPc) points towards possible mechanisms of action of IVIG in AD that go beyond the direct binding of Aß. Tauhas been the target of immunotherapeutic strategies in the past, where both passive and active immunization improved pathology in neurofibrillary tangle mouse models. Additionally, we have measured binding to PrPc, which has been shown to function as a receptor for Aß and can induce neuronal apoptosis upon ligation. The binding of PrivigenÒ to these proteins might have beneficial effects by preventing tau-aggregation and PrPc-induced cell death. Further experiments will be necessary to address these putative mechanisms of action at a functional level.
P3-484
LITHIUM INCREASES MITOCHONDRIAL FUNCTION IN CULTURED HIPPOCAMPAL NEURONS
Leda Talib1, Wagner Gattaz2, Orestes Forlenza2, 1Laboratory of Neuroscience (LIM-27), Department and Institute of Psychiatry, Faculty of Medicine, Un, S~ ao Paulo - S.P., Brazil; 2Psychiatric institute Faculty of Medicine S~ ao Paulo, S~ ao Paulo - S.P., Brazil. Background: Studies have shown that lithium favors the activation of neurotrophic and neuroprotective cascades in neuronal cell cultures and animal
Figure 1.
models. Lithium is a potent inhibitor of GSK3b activity and also stimulates the production of neurotrophic (e.g., BDNF) and anti-apoptotic factors. Yet, the exact mechanisms that underlie the aforementioned properties of lithium are not fully understoods to evaluate the effect of chronic lithium treatment on mitochondrial function in cultured neurons Methods: Primary cultures of hippocampal neurons started from E18 embryonic rats were treated from the 7th to the 12th day in culture with different concentrations of LiCl (0.02, 0.2 and 2mM). Culture viability was quantitatively assessed by the MTT method. Because the conversion of MTT depends on the activity of mitochondrial dehydrogenases, it estimates the percentage of viablecellsin a given substrate as compared to controls, i.e., the functional state of the respiratory chain of living cells. Results: Preliminary results drawn from at least five independent experiments suggest a significant, dose dependent, increase in mitochondrial function in hippocampal neurons treated with LiCl (0.2mM: 7%, 2.mM: 5% of increase, Figure1). Conclusions: Our data suggest that lithium treatment is associated with an increment in mitochondrial function in cultured hippocampal neurons. These effects may have important implications for the understanding and management of neuropsychiatric diseases, namely bipolar disorder and Alzheimer’s disease (AD).
P3-485
COMPLEMENT COMPONENTS C5-DERIVED C5a ANAPHYLATOXIN-MEDIATED ATTENUATION OF COGNITIVE DETERIORATION IS A POTENTIAL NOVEL PHARMACOLOGICAL TARGET OF IVIG REGULATED HORMONAL IMMUNOTHERAPY IN ALZHEIMER’S DISEASE
Giulio Pasinetti, Mount Sinai School of Medicine, New York, New York, United States. Background: Human intravenous immunoglobulin (IVIG), a safe immunotherapeutic approach, recently, it appears to be a promising target in Alzheimer’s disease (AD) immunotherapeutic studies. However, the underlying mechanisms through which IVIG improves cognitive functions are not clear. Evidence showed that genetic inhibition of molecular mechanisms resulting in a reduction of levels of the complement C3, and therefore C3-derived C3a anaphylatoxin, in the brain of an AD model exacerbates the AD-type phenotype and cognitive deterioration. Moreover, previous we reported that C5a may influence upstream signal transduction pathways associated with CREB/CBP activation and the key roles of CREB are associated with cognitive deterioration in AD. Based on the evidence that IVIG selectively activates certain complement anaphylatoxin, we propose that the mechanism of IVIG’s beneficial effects in the improvement of cognitive function might be through the hormonal immune regulation of these complement components such as C3 and C5a. Methods: A total of 214 postmortem brain hippocampal samples were grouped by of Clinical Dementia Rating (CDR) scores. Their C5aR levels, neuropath logical changes such as amyloid plaques, and neurofibril tangles were assessed and the correlation with CDR scores was analyzed. TgC5aR-/-, TgC5aR+/+, Tg2576 mouse and wild-type littermates were used for the evaluation of the behavioral performance, synaptic function, and signaling transduction pathways. Results: 1) Cognitive function was severely impaired in C5aR deficits mice and correlated with the down-regulation of the levels of the pCaMKII, pCREB/ CEBP. 2) Activation of C5aR via either application of hrC5a or in the C5aR over expression mice, improved the CaMKII-pCREB pathway. 3) The C5a and GluR1 levels were significantly higher in normal oldest elder lies than in those with severe cognitive deficits, and their cognitive impairments are not directly correlated with the number and size of amyloid-plaques and neurofibril tangles; 4) exogenous application of hrC5a peptide in Tg2576 mice significantly improved the deficits of LTP in hippocampal CA1 region. Moreover, our studies related to the IVIG mediated induction of C3 and C5a content in the brain will be discussed. Conclusions: One of the therapeutic mechanisms of IVIG in AD may be through the local regulation of the C5a complement mediated CaMKII-pCREB signaling pathway in the brain.
Poster Presentations P3 INTRAVENOUS IMMUNOGLOBULIN BINDS TO TAU, pTAU AND PrPc IN ADDITION TO Ab
Alexander Schaub1, Alain Kropf1, Susann Cattepoel1, Miriam Ender1, Reinhard Bolli1, Louis Fabri2, Sylvia Miescher1, 1CSL Behring AG, Bern, Switzerland; 2CSL Limited, Parkville, Australia. Background: Intravenous Immunoglobulin (IVIG) is being used in phase III clinical studies to assess its efficacy in mild-to-moderate Alzheimer’s disease (AD) patients. Beside beneficial effects in smaller scale clinical settings, the rationale for using IVIG in AD is based on observations of reduced levels of anti-amyloid beta (Aß)antibodies in AD patients and the presence of these antibodies in normal IVIG preparations (R. Dodel, J Neurol 2004; N Relkin, Neurobiol Aging, 2009). We examined whether PrivigenÒ(10% IVIG product) contains specific antibodies that bind to proteins implicated in AD pathology that might explain mechanisms of action in addition to the direct binding of Aß. IVIG is hypothesized to mediate its beneficial effects in chronic inflammatory and autoimmune conditions by a variety of mechanisms. As this might also be the case for AD we measured PrivigenÒbinding to Tau, phosphorylated Tau (pTau) and the cellular prion protein (PrPc). Methods: The binding of PrivigenÒto antigens was measured by ELISA. Differences in IVIG binding were observed during method development as a result of both antibody and blocking solutions that were used. Results: We show herein that PrivigenÒ binds to Aß, as well as tau, ptau and PrPc, but not a-synuclein under the same experimental conditions. We also demonstrate that the influence of blocking solutions and antibody diluents on natural auto antibodies is greater than on monoclonal or high titre antibodies. Conclusions: The interaction of PrivigenÒ with additional antigens (tau, ptau, PrPc) points towards possible mechanisms of action of IVIG in AD that go beyond the direct binding of Aß. Tauhas been the target of immunotherapeutic strategies in the past, where both passive and active immunization improved pathology in neurofibrillary tangle mouse models. Additionally, we have measured binding to PrPc, which has been shown to function as a receptor for Aß and can induce neuronal apoptosis upon ligation. The binding of PrivigenÒ to these proteins might have beneficial effects by preventing tau-aggregation and PrPc-induced cell death. Further experiments will be necessary to address these putative mechanisms of action at a functional level.
P3-484
LITHIUM INCREASES MITOCHONDRIAL FUNCTION IN CULTURED HIPPOCAMPAL NEURONS
Leda Talib1, Wagner Gattaz2, Orestes Forlenza2, 1Laboratory of Neuroscience (LIM-27), Department and Institute of Psychiatry, Faculty of Medicine, Un, S~ ao Paulo - S.P., Brazil; 2Psychiatric institute Faculty of Medicine S~ ao Paulo, S~ ao Paulo - S.P., Brazil. Background: Studies have shown that lithium favors the activation of neurotrophic and neuroprotective cascades in neuronal cell cultures and animal
Figure 1.
models. Lithium is a potent inhibitor of GSK3b activity and also stimulates the production of neurotrophic (e.g., BDNF) and anti-apoptotic factors. Yet, the exact mechanisms that underlie the aforementioned properties of lithium are not fully understoods to evaluate the effect of chronic lithium treatment on mitochondrial function in cultured neurons Methods: Primary cultures of hippocampal neurons started from E18 embryonic rats were treated from the 7th to the 12th day in culture with different concentrations of LiCl (0.02, 0.2 and 2mM). Culture viability was quantitatively assessed by the MTT method. Because the conversion of MTT depends on the activity of mitochondrial dehydrogenases, it estimates the percentage of viablecellsin a given substrate as compared to controls, i.e., the functional state of the respiratory chain of living cells. Results: Preliminary results drawn from at least five independent experiments suggest a significant, dose dependent, increase in mitochondrial function in hippocampal neurons treated with LiCl (0.2mM: 7%, 2.mM: 5% of increase, Figure1). Conclusions: Our data suggest that lithium treatment is associated with an increment in mitochondrial function in cultured hippocampal neurons. These effects may have important implications for the understanding and management of neuropsychiatric diseases, namely bipolar disorder and Alzheimer’s disease (AD).
P3-485
COMPLEMENT COMPONENTS C5-DERIVED C5a ANAPHYLATOXIN-MEDIATED ATTENUATION OF COGNITIVE DETERIORATION IS A POTENTIAL NOVEL PHARMACOLOGICAL TARGET OF IVIG REGULATED HORMONAL IMMUNOTHERAPY IN ALZHEIMER’S DISEASE
Giulio Pasinetti, Mount Sinai School of Medicine, New York, New York, United States. Background: Human intravenous immunoglobulin (IVIG), a safe immunotherapeutic approach, recently, it appears to be a promising target in Alzheimer’s disease (AD) immunotherapeutic studies. However, the underlying mechanisms through which IVIG improves cognitive functions are not clear. Evidence showed that genetic inhibition of molecular mechanisms resulting in a reduction of levels of the complement C3, and therefore C3-derived C3a anaphylatoxin, in the brain of an AD model exacerbates the AD-type phenotype and cognitive deterioration. Moreover, previous we reported that C5a may influence upstream signal transduction pathways associated with CREB/CBP activation and the key roles of CREB are associated with cognitive deterioration in AD. Based on the evidence that IVIG selectively activates certain complement anaphylatoxin, we propose that the mechanism of IVIG’s beneficial effects in the improvement of cognitive function might be through the hormonal immune regulation of these complement components such as C3 and C5a. Methods: A total of 214 postmortem brain hippocampal samples were grouped by of Clinical Dementia Rating (CDR) scores. Their C5aR levels, neuropath logical changes such as amyloid plaques, and neurofibril tangles were assessed and the correlation with CDR scores was analyzed. TgC5aR-/-, TgC5aR+/+, Tg2576 mouse and wild-type littermates were used for the evaluation of the behavioral performance, synaptic function, and signaling transduction pathways. Results: 1) Cognitive function was severely impaired in C5aR deficits mice and correlated with the down-regulation of the levels of the pCaMKII, pCREB/ CEBP. 2) Activation of C5aR via either application of hrC5a or in the C5aR over expression mice, improved the CaMKII-pCREB pathway. 3) The C5a and GluR1 levels were significantly higher in normal oldest elder lies than in those with severe cognitive deficits, and their cognitive impairments are not directly correlated with the number and size of amyloid-plaques and neurofibril tangles; 4) exogenous application of hrC5a peptide in Tg2576 mice significantly improved the deficits of LTP in hippocampal CA1 region. Moreover, our studies related to the IVIG mediated induction of C3 and C5a content in the brain will be discussed. Conclusions: One of the therapeutic mechanisms of IVIG in AD may be through the local regulation of the C5a complement mediated CaMKII-pCREB signaling pathway in the brain.