- Email: [email protected]
Diet-induced hyperlipidemia accelerates amyloid deposition in APPv717F transgenic mouse model of Alzheimer disease
Symposium:
Apolipoprotein
E and Alzheimer’s
Disease
contrast, fibrillar A@ was never seen in the absence of apoE in the brain parenchyma of either APPsw or APPV7’7r mice through 15 months of age. CAA, which only occurs in APPsw but not APP”“” mice, was blocked by removing apoE. Importantly, despite AP deposition in the absence of apoE, neuritic degeneration (i.e. neuritic plaques) only developed when AP deposition occurred in the presence of apoE. Interestingly, the expression of human apoE &forms in the brain of both APPsw and APPv7’7” mice suppressed early AP deposition for several months. Despite this, AP deposition eventually developed in APPY7’7F, human apoE-positive rmce by 15 months of age. Further, the AP was fibrillar, and neuritic degeneration became evident. Of note, apoEA was associated with significantly greater A@ deposition and neuritic degeneration than apoE3. Our data suggest that apoE is involved in AP clearance and tibrillogenesis in viw and that it is a critical modulator of neuritic and cerebrovascular plaque formation. In viva and in vitro studies are underway to determine the bimhemical mechanisms underlying the effects of CNS apoE/lipoproteins on AP metabolism.
AS A ~THERAPEUTIC [6261 APOE OF ALZHEIMER’S DISEASE
TARGET
FOR THE TREATMENT
Judes Poiner, McGill Univrrsuy / Dou&s Hospital, Verdun. PQ Cunadu; Michrl Pmisset, McGi// Universitv. Verdun. PQ Canuda The discovery that the apolipoprotein ~4 (apoE4) allele is strongly linked to both sporadic and familial late onset Alzheimer’s disease (AD) raisea the possibility that a dysfunction of the lipid transport system could seriously affect lipid homeostasis in the brain. It has been shawn that the presence of the t4 allele is associated with reduced levels of apoE in both serum and brain tissues of AD subjects. Moreover, synaptic integrity and plasticity is markedly compromised in an ~4 allele dosedependent manner in several brain areas in AD. These observations led us to propose that the reduced concentrations of apoE observed in the brain of apoEl-AD subjects may compromise cholesterol and phospholipids transport in the CNS in a manner consistent with what has been described in apoE knockout and apoE knockin mice. This, in turn, would indirectly impair the synaptic integrity and plasticny in several key neurotransmitter systems in the CNS; particularly in the cholinergic system which relies heavily on lipids to synthesize acetylcholine. With the objective of reversing the reported apoE deficit in the brain of c4 allele carriers, we identified and characterised several apoE inducer druga from a low throughput astrocyte screening assay and characterised their effects in brain cells. Tacrine and Heptylphysostigmine were found to be potent inducers of apoE secretion in vitro. In contrast, donepesil (Aricept) and rlvastigmine (Exelon) both lack this particular activity. Another apoE inducer drug called probucol was examined in mice and rat brains as well as in humans suffenng from nuld to moderate sporadic AD. Cerebroapinal fluid (CSF) evels of apoE, beta amyloid I-40, lipid peroxides and Tau were determined before and during treatment and subsequently contrasted with clinical outcome measures (CIBIC, CDR and ADAS-Cog) at 3 and 6 months. Eleven Eubject with mild-to moderate AD were Invited to take pm in this studies. The proof-of-principle climcal trial revealed that apoE induction in the brain (CSF) indeed affects disease progressmn in an apoEconcentration dependent manner. Supported by grants from the Medical Research Council of Canada. Alzheimer Society of Canada, FRSQ, Alzheimer Group of Montreal and Nova Molecular Inc.
DIET-INDUCED HYPERLIPIDEMIA ACCELERATES AMYLOID DEPOSITION IN THE APP”“= TRANSGENIC MOUSE MODEL OF ALZHEIMER DISEASE. Kelly R. Bales, Cindy l;ishmun, Cindy DeLon~, Eli Lilly and Company, Indianapolis, IN: Yanvhmg Du, lndiurru University School of Medicine. Indianapolis. IN; Willium Jordun. Strwn M. Paul, Eli Lilly and Cornpuny, Indiunapolis, IN Several genetic and nongenetic factors are associated with an increased risk for developing Alzheimer’s disease (AD) including the inheritance of an apolipoprotein t4 allele, increased age, and hyperlipidemia. Tu investigate the role of elevated serum lipids on amyloid deposition we fed male and female APPY7”’ transgenic mice (TG) mice a high fat (21.2% fat, 0.2% cholesterol) or normal (5.5% fat, 0% cholesterol) diet for various lengths of time. At 6,12,1X, and 24 weeks APPV717F TG mice were sacrificed and serum cholesterol, triglycerides as well as plaque burden in various brain regions were quantified. After 12 weeks on a high fat diet both male and female APPV7”r TG mice had a qigmticant elevation in serum triglycerides when compared to APP”“” TG mice fed a normal diet. Serum cholesterol, however, was signiticandy elevated after only 6 weeks on a high fat diet and remained elevated throughout the course of the study (24 weeks). Amyloid burden (thioflavine-S fluorescent deposits as well as AP immunorextivity) was significantly increased (pcO.05) in both the hippocampus as well as the cerebral cortex in APP”7’7F TG mice after 12 weeks on high fat diet. Although amyloid burden increased significantly in an age-dependent fashion in male APPV7”F TG rmce fed a control diet, no further accrual of amyloid occurred when mice were on a high fat diet for a longer (24 weeks)
s139
period of time, suggesting that a high fat diet-induced hyperlipidemia may accelerate amylold deposition in these mice. Interestingly APPV717F TG mice without apolipoprotein E have markedly elevated (greater than lo-fold) levels of serum cholesterol, and yet few if any amyloid deposits. Taken together, our results support the epidemiological association between the elevated risk to develop AD and hyperlipidemia perhaps through an apoE-dependent mechanism which leads to an increase in brain amyloid.
COGNITIVE DEFICITS 16281THEIR RELATIONSHIP
IN APOM TRANSGENIC MICE AND TO AGING, SEX, AND INCREASED
ARLEYELS lmnnart Muck<, Gladstone Institute of Neurological Disease, San Francisco, CA Deficits in spatial memory are a prominent feature of AD. Our analysis of transgenic mice expressing human apolipoprotein (ape) E &forms indicates that E4 plays a key role in the development of spatial memory impairments. In humans, E4 increases AD risk and accelerates disease manifestations compared with E3 and E2. We analyzed murine apoE-deficient (Apoe-‘-) mice in which the neuron-specific enolase (NSE) promoter directs expression of human E3 or E4 at comparable levels (PNAS 95: 10914; J. Neurosci. 19: 4867). These mice had normal basic visuomotor skills. At 3 months of age, E3 and E4 mice showed no deficits in spatial learning and memory in a water maze test compared with Apoe+‘+ or Apoe-‘-controls lacking human apoE. E3 mice performed well also at 6 and IX months. In contrast, E4 mice developed progressive impairments affecting spatial memory retention (probe trial) at 6 months and both spatial leaming and memory at I8 months. These E&associated deficits were detected in female, but not male, mice. Testosterone treatment significantly improved the performance of h-month-old female El mice (Raber et al., this meeting). Next, we tested whether human apoE &forms differentially affect spatial memory also in males if mice are challenged with AD-related copathogens. E3 and E4 mice were crossed with human amyloid precursor protein transgenic Apoe-‘~ mice expressing high levels of human Ahin neurons. In h-month-old male and female offspring, E3 prevented Ab-induced spatial memory deficits, but E4 did not. Molecular and morphological alterations that may relate to U-associated deficits will be discussed. Thus, E3 preserves cognitive function5 during aging and prevents Ab-induced neuronal deficits mere effectively than E4. In addition, E4 actively promotes age-related cognitive decline in a gender-dependent fashion and this effect can be counteracted with sex hormones. Ongoing studies focus on the development and assessment of apoE-targeted therapeutic strategies. Supported by the NIA, the Alzheimer’s Association, and The John Douglas French Alzheimer’s Foundation.
APOE AND 16291DISEASE.
ESTROGEN
IN MOUSE MODELS OF ALZHEIMER
Jonurkun D. Smrrk, Justine A. Lrvin, Ckru Lmninska, Yuri Bunimuvich, Rockefellrr Umwsiry. New York, NY ApoE is the most prevalent genetic susceptibility factor for Alzheimer disease (AD). Females are more likely to develop AD than males, and this differece can be accounted for by the increased likelihood of AD among EA/3 females compared to males. Also, women who take estrogen replacement therapy are less likely to develop AD. ! fs20Head injury is an environmental risk factor for AD, but only among apoE carriers. Thus we have studied the effects of various genetic and environmental risk factor5 in various inbred mouse models for AD. In C57BL/6 mice, estrogen induces apoE in the cortex, hippocampus, and diencephalon. We determined that the estrogen induction of apoE in the diencephalon, but not the hippocampus and cortex, is mediated by estrogen receptor a (for details, see abstract by Justine Levi”). Amyloid precursor protein (APP) transgenic and wildtype mice were ovariectomized (OVX) at 4 weeks of age and survival through 8 months of age was followed versus sham OVX and intact mice. Mortality was 60% in the OVX APP mice, compared to 0% in the sham OVX and intact APP plainmice, and 10% in the OVX wildtype mice. This increased death was not associated with increased brain levels of total AP, measured by ELISA. The effects of 6 week treatments with pharmacological doses of 17@- and l7a-estradiol on AP levels were determined in young OVX APP transgenic mice. 17p- and 17a-estradiol led to 10% and 25% decreases in total AP levels, respectively, compared to placebo. Fluid percusive brain injury was performed unilaterally to determine the effects of trauma on apoE and A@ levels in APP transgenic mice. 3 days! after injury, the apoE levels were about 4.fold higher in the injured versus control brain hemisphere, but apoE levels decreased close to baseline 6 days after injury. In contrast, AP levels in the injured hemisphere increased about 2.fold, but only 15 days after the injury, the latest time point examined thus far. The effect of apoE-deficiency on learning was examined using an a-arm radial maze, cued by odorants. We observed that the genetic background strain had an impact on the effect of apoE-deficiency, such that apoE-deficient mice on the FVB/N background learned as well as wildype FVBlN mice, while apoE-deficient C57BW6 mice had decreased learning Compared to their wildtype controls.
Apolipoprotein
E and Alzheimer’s
Disease
contrast, fibrillar A@ was never seen in the absence of apoE in the brain parenchyma of either APPsw or APPV7’7r mice through 15 months of age. CAA, which only occurs in APPsw but not APP”“” mice, was blocked by removing apoE. Importantly, despite AP deposition in the absence of apoE, neuritic degeneration (i.e. neuritic plaques) only developed when AP deposition occurred in the presence of apoE. Interestingly, the expression of human apoE &forms in the brain of both APPsw and APPv7’7” mice suppressed early AP deposition for several months. Despite this, AP deposition eventually developed in APPY7’7F, human apoE-positive rmce by 15 months of age. Further, the AP was fibrillar, and neuritic degeneration became evident. Of note, apoEA was associated with significantly greater A@ deposition and neuritic degeneration than apoE3. Our data suggest that apoE is involved in AP clearance and tibrillogenesis in viw and that it is a critical modulator of neuritic and cerebrovascular plaque formation. In viva and in vitro studies are underway to determine the bimhemical mechanisms underlying the effects of CNS apoE/lipoproteins on AP metabolism.
AS A ~THERAPEUTIC [6261 APOE OF ALZHEIMER’S DISEASE
TARGET
FOR THE TREATMENT
Judes Poiner, McGill Univrrsuy / Dou&s Hospital, Verdun. PQ Cunadu; Michrl Pmisset, McGi// Universitv. Verdun. PQ Canuda The discovery that the apolipoprotein ~4 (apoE4) allele is strongly linked to both sporadic and familial late onset Alzheimer’s disease (AD) raisea the possibility that a dysfunction of the lipid transport system could seriously affect lipid homeostasis in the brain. It has been shawn that the presence of the t4 allele is associated with reduced levels of apoE in both serum and brain tissues of AD subjects. Moreover, synaptic integrity and plasticity is markedly compromised in an ~4 allele dosedependent manner in several brain areas in AD. These observations led us to propose that the reduced concentrations of apoE observed in the brain of apoEl-AD subjects may compromise cholesterol and phospholipids transport in the CNS in a manner consistent with what has been described in apoE knockout and apoE knockin mice. This, in turn, would indirectly impair the synaptic integrity and plasticny in several key neurotransmitter systems in the CNS; particularly in the cholinergic system which relies heavily on lipids to synthesize acetylcholine. With the objective of reversing the reported apoE deficit in the brain of c4 allele carriers, we identified and characterised several apoE inducer druga from a low throughput astrocyte screening assay and characterised their effects in brain cells. Tacrine and Heptylphysostigmine were found to be potent inducers of apoE secretion in vitro. In contrast, donepesil (Aricept) and rlvastigmine (Exelon) both lack this particular activity. Another apoE inducer drug called probucol was examined in mice and rat brains as well as in humans suffenng from nuld to moderate sporadic AD. Cerebroapinal fluid (CSF) evels of apoE, beta amyloid I-40, lipid peroxides and Tau were determined before and during treatment and subsequently contrasted with clinical outcome measures (CIBIC, CDR and ADAS-Cog) at 3 and 6 months. Eleven Eubject with mild-to moderate AD were Invited to take pm in this studies. The proof-of-principle climcal trial revealed that apoE induction in the brain (CSF) indeed affects disease progressmn in an apoEconcentration dependent manner. Supported by grants from the Medical Research Council of Canada. Alzheimer Society of Canada, FRSQ, Alzheimer Group of Montreal and Nova Molecular Inc.
DIET-INDUCED HYPERLIPIDEMIA ACCELERATES AMYLOID DEPOSITION IN THE APP”“= TRANSGENIC MOUSE MODEL OF ALZHEIMER DISEASE. Kelly R. Bales, Cindy l;ishmun, Cindy DeLon~, Eli Lilly and Company, Indianapolis, IN: Yanvhmg Du, lndiurru University School of Medicine. Indianapolis. IN; Willium Jordun. Strwn M. Paul, Eli Lilly and Cornpuny, Indiunapolis, IN Several genetic and nongenetic factors are associated with an increased risk for developing Alzheimer’s disease (AD) including the inheritance of an apolipoprotein t4 allele, increased age, and hyperlipidemia. Tu investigate the role of elevated serum lipids on amyloid deposition we fed male and female APPY7”’ transgenic mice (TG) mice a high fat (21.2% fat, 0.2% cholesterol) or normal (5.5% fat, 0% cholesterol) diet for various lengths of time. At 6,12,1X, and 24 weeks APPV717F TG mice were sacrificed and serum cholesterol, triglycerides as well as plaque burden in various brain regions were quantified. After 12 weeks on a high fat diet both male and female APPV7”r TG mice had a qigmticant elevation in serum triglycerides when compared to APP”“” TG mice fed a normal diet. Serum cholesterol, however, was signiticandy elevated after only 6 weeks on a high fat diet and remained elevated throughout the course of the study (24 weeks). Amyloid burden (thioflavine-S fluorescent deposits as well as AP immunorextivity) was significantly increased (pcO.05) in both the hippocampus as well as the cerebral cortex in APP”7’7F TG mice after 12 weeks on high fat diet. Although amyloid burden increased significantly in an age-dependent fashion in male APPV7”F TG rmce fed a control diet, no further accrual of amyloid occurred when mice were on a high fat diet for a longer (24 weeks)
s139
period of time, suggesting that a high fat diet-induced hyperlipidemia may accelerate amylold deposition in these mice. Interestingly APPV717F TG mice without apolipoprotein E have markedly elevated (greater than lo-fold) levels of serum cholesterol, and yet few if any amyloid deposits. Taken together, our results support the epidemiological association between the elevated risk to develop AD and hyperlipidemia perhaps through an apoE-dependent mechanism which leads to an increase in brain amyloid.
COGNITIVE DEFICITS 16281THEIR RELATIONSHIP
IN APOM TRANSGENIC MICE AND TO AGING, SEX, AND INCREASED
ARLEYELS lmnnart Muck<, Gladstone Institute of Neurological Disease, San Francisco, CA Deficits in spatial memory are a prominent feature of AD. Our analysis of transgenic mice expressing human apolipoprotein (ape) E &forms indicates that E4 plays a key role in the development of spatial memory impairments. In humans, E4 increases AD risk and accelerates disease manifestations compared with E3 and E2. We analyzed murine apoE-deficient (Apoe-‘-) mice in which the neuron-specific enolase (NSE) promoter directs expression of human E3 or E4 at comparable levels (PNAS 95: 10914; J. Neurosci. 19: 4867). These mice had normal basic visuomotor skills. At 3 months of age, E3 and E4 mice showed no deficits in spatial learning and memory in a water maze test compared with Apoe+‘+ or Apoe-‘-controls lacking human apoE. E3 mice performed well also at 6 and IX months. In contrast, E4 mice developed progressive impairments affecting spatial memory retention (probe trial) at 6 months and both spatial leaming and memory at I8 months. These E&associated deficits were detected in female, but not male, mice. Testosterone treatment significantly improved the performance of h-month-old female El mice (Raber et al., this meeting). Next, we tested whether human apoE &forms differentially affect spatial memory also in males if mice are challenged with AD-related copathogens. E3 and E4 mice were crossed with human amyloid precursor protein transgenic Apoe-‘~ mice expressing high levels of human Ahin neurons. In h-month-old male and female offspring, E3 prevented Ab-induced spatial memory deficits, but E4 did not. Molecular and morphological alterations that may relate to U-associated deficits will be discussed. Thus, E3 preserves cognitive function5 during aging and prevents Ab-induced neuronal deficits mere effectively than E4. In addition, E4 actively promotes age-related cognitive decline in a gender-dependent fashion and this effect can be counteracted with sex hormones. Ongoing studies focus on the development and assessment of apoE-targeted therapeutic strategies. Supported by the NIA, the Alzheimer’s Association, and The John Douglas French Alzheimer’s Foundation.
APOE AND 16291DISEASE.
ESTROGEN
IN MOUSE MODELS OF ALZHEIMER
Jonurkun D. Smrrk, Justine A. Lrvin, Ckru Lmninska, Yuri Bunimuvich, Rockefellrr Umwsiry. New York, NY ApoE is the most prevalent genetic susceptibility factor for Alzheimer disease (AD). Females are more likely to develop AD than males, and this differece can be accounted for by the increased likelihood of AD among EA/3 females compared to males. Also, women who take estrogen replacement therapy are less likely to develop AD. ! fs20Head injury is an environmental risk factor for AD, but only among apoE carriers. Thus we have studied the effects of various genetic and environmental risk factor5 in various inbred mouse models for AD. In C57BL/6 mice, estrogen induces apoE in the cortex, hippocampus, and diencephalon. We determined that the estrogen induction of apoE in the diencephalon, but not the hippocampus and cortex, is mediated by estrogen receptor a (for details, see abstract by Justine Levi”). Amyloid precursor protein (APP) transgenic and wildtype mice were ovariectomized (OVX) at 4 weeks of age and survival through 8 months of age was followed versus sham OVX and intact mice. Mortality was 60% in the OVX APP mice, compared to 0% in the sham OVX and intact APP plainmice, and 10% in the OVX wildtype mice. This increased death was not associated with increased brain levels of total AP, measured by ELISA. The effects of 6 week treatments with pharmacological doses of 17@- and l7a-estradiol on AP levels were determined in young OVX APP transgenic mice. 17p- and 17a-estradiol led to 10% and 25% decreases in total AP levels, respectively, compared to placebo. Fluid percusive brain injury was performed unilaterally to determine the effects of trauma on apoE and A@ levels in APP transgenic mice. 3 days! after injury, the apoE levels were about 4.fold higher in the injured versus control brain hemisphere, but apoE levels decreased close to baseline 6 days after injury. In contrast, AP levels in the injured hemisphere increased about 2.fold, but only 15 days after the injury, the latest time point examined thus far. The effect of apoE-deficiency on learning was examined using an a-arm radial maze, cued by odorants. We observed that the genetic background strain had an impact on the effect of apoE-deficiency, such that apoE-deficient mice on the FVB/N background learned as well as wildype FVBlN mice, while apoE-deficient C57BW6 mice had decreased learning Compared to their wildtype controls.