ApoE Mice Research Articles

APOE genotype and sex modulate Alzheimer's disease pathology in aged EFAD transgenic mice.

Increasing evidence supports that age, APOE and sex interact to modulate Alzheimer's disease (AD) risk, however the underlying pathways are unclear. One way that AD risk factors may modulate cognition is by impacting amyloid beta (Aβ) accumulation as plaques, and/or neuroinflammation Therefore, the goal of the present study was to evaluate the extent to which age, APOE and sex modulate Aβ pathology, neuroinflammation and behavior in vivo. To achieve this goal, we utilized the EFAD mice, which express human APOE3 or APOE4 and have five familial AD mutations (FAD) that result in Aβ42 overproduction. We assessed Aβ levels, reactive glia and Morris water maze performance in 6-, 10-, 14-, and 18-month-old EFAD mice. Female APOE4 mice had the highest Aβ deposition, fibrillar amyloid deposits and neuroinflammation as well as earlier behavior deficits. Interestingly, we found that female APOE3 mice and male APOE4 mice had similar levels of pathology. Collectively our data support that the combination of APOE4 and female sex is the most detrimental combination for AD, and that at older ages, female sex may be equivalent to APOE4 genotype.

Adropin inhibits the progression of atherosclerosis in ApoE-/-/Enho-/- mice by regulating endothelial-to-mesenchymal transition

Adropin, a secreted protein, coded by energy homeostasis-associated gene (Enho), is recently reported to modulate atherogenesis, with endothelial-to-mesenchymal transition (EndMT) involved in the early process. We explored whether adropin may alleviate atherosclerosis by regulating EndMT. We found that an intraperitoneal injection of adropin [105 μg/(kg·d) for 13 weeks] inhibited the progression of high-fat diet (HFD)-induced aortic atherosclerosis in apolipoprotein E-deficient mice (ApoE–/–) and those with double gene deletion (ApoE–/–/Enho–/–), as detected by Oil Red O and haematoxylin-eosin staining. In the aortas of ApoE–/– mouse, adropin treatment ameliorated the decrease in the mRNA expression of endothelial cell markers (leukocyte differentiation antigen 31, CD31, and vascular endothelial cadherin, VE-cadherin), but increased that of EndMT markers (alpha smooth muscle actin, α-SMA, and fibroblasts specific protein-1). In vitro, an adropin treatment (30 ng/ml) arrested the hydrogen peroxide (H2O2)-induced EndMT in human umbilical vein endothelial cells (HUVECs), attenuated the morphological changes of HUVECs, reduced the number of immunofluorescence-positive α-SMA, increased the mRNA and protein expressions of CD31 and VE-cadherin, and decreased those of α-SMA. Furthermore, the adropin treatment decreased the mRNA and protein expressions of transforming growth factor (TGF)-β1 and TGF-β2, and suppressed the phosphorylation of downstream signal protein Smad2/3 in HUVECs. These mitigative effects of adropin on H2O2-induced EndMT were reversed by the transfection of TGF-β plasmid. The findings signify that adropin treatment may alleviate the atherosclerosis in ApoE–/–/Enho–/– mice by inhibiting EndMT via the TGF-β/Smad2/3 signaling pathway.

Ferroptosis: A potential target of macrophages in plaque vulnerability.

Plaque vulnerability has been the subject of several recent studies aimed at reducing the risk of stroke and carotid artery stenosis. Atherosclerotic plaque development is a complex process involving inflammation mediated by macrophages. Plaques become more vulnerable when the equilibrium between macrophage recruitment and clearance is disturbed. Lipoperoxides, which are affected by iron levels in cells, are responsible for the cell death seen in ferroptosis. Ferroptosis results from lipoperoxide-induced mitochondrial membrane toxicity. Atherosclerosis in ApoE(-/-) mice is reduced when ferroptosis is inhibited and iron intake is limited. Single-cell sequencing revealed that a ferroptosis-related gene was substantially expressed in atherosclerosis-modeled macrophages. Since ferroptosis can be regulated, it offers hope as a non-invasive method of treating carotid plaque. In this study, we discuss the role of ferroptosis in atherosclerotic plaque vulnerability, including its mechanism, regulation, and potential future research directions.

Tanshinone IIA Alleviates Atherosclerosis Through Inhibition of NF-κB and PPARα/ABCA1 Signaling Pathways

This study evaluated the role and underlying mechanisms of Tanshinone IIA (Tan IIA) in atherosclerosis. C57BL mice (control group) and ApoE mice (model group) were administered a conventional and high-fat diet for 20 weeks. The Tan IIA group was obtained by administering a high-fat diet plus 8 weeks of Tan IIA to other mice for 20 weeks, followed by oil red O staining and lipid examination. RAW264.7 cells were transfected with PPARα siRNA+Tan IIA to measure their expression. The results showed little change in body weight between the three groups (P < 0.05). Liver index was significantly increased in the model and Tan IIA groups (P <0.05). Atherosclerotic plaques, plaque cross-sectional area, human oxidized low-density lipoprotein (ox-LDL), low-density lipoprotein cholesterol (LDL-C) and triglyceride (TG) levels, p-NF-κB, p-IKKα, P-Ikkα/β, TNF-α and IL-1β levels were significantly increased in the model group and decreased in the Tan IIA group (P < 0.05). We also noted a decrease in PPARα, PGC-1α and ABCA1 in the model group and an increase in the Tan IIA group. NF-κB expression was increased in the nucleus and decreased in the cytoplasm in the model group, which was reversed by Tan IIA treatment. Tan IIA significantly reduced ox-LDL, LDL-C and TG levels, plaque size and plaque cross-sectional area in atherosclerosis. Tan IIA effectively inhibited NF-κB, activated the PPARα/ABCA1 signalling pathway, and reduce inflammatory pathways, thereby improving lipid deposition and acting as an anti-atherosclerotic agent.

Research on the Mechanism of Moxibustion Intervention in Atherosclerosis Based on Next Generation Sequencing

This study aimed to investigate the underlying mechanism through which moxibustion intervention therapy improves platelet activation in rats with atherosclerosis using Next Generation Sequencing (NGS). Atherosclerosis models were established in 15 Apolipoprotein E knockout mice (ApoE-/- mice) aged 8 weeks, while non-transgenic mice (C57BL/6) were used as the control group. The mice were divided into four groups: moxibustion group, clopidogrel group, model group, and blank control group. Following 12 weeks of intervention, platelet-rich plasma was prepared from venous blood collected from the mice’s eyeballs. RNA extraction and analysis were performed to assess gene expression patterns. In the moxibustion group, the expression of PPBP was downregulated, while PTPN7 expression was upregulated, indicating involvement in the MAPK classical pathway. The moxibustion group exhibited significantly higher PTPN7 mRNA expression compared to both the clopidogrel and model groups, while Robo4 mRNA expression was significantly reduced in both the moxibustion and clopidogrel groups. The PTPN7 gene locus, acting through the MAPK pathway, may serve as a potential target for enhancing atherosclerosis management by modulating platelet activation.

Prebiotic inulin enhances gut microbial metabolism and anti-inflammation in apolipoprotein E4 mice with sex-specific implications

Gut dysbiosis has been identified as a crucial factor of Alzheimer's disease (AD) development for apolipoprotein E4 (APOE4) carriers. Inulin has shown the potential to mitigate dysbiosis. However, it remains unclear whether the dietary response varies depending on sex. In the study, we fed 4-month-old APOE4 mice with inulin for 16 weeks and performed shotgun metagenomic sequencing to determine changes in microbiome diversity, taxonomy, and functional gene pathways. We also formed the same experiments with APOE3 mice to identify whether there are APOE-genotype dependent responses to inulin. We found that APOE4 female mice fed with inulin had restored alpha diversity, significantly reduced Escherichia coli and inflammation-associated pathway responses. However, compared with APOE4 male mice, they had less metabolic responses, including the levels of short-chain fatty acids-producing bacteria and the associated kinases, especially those related to acetate and Erysipelotrichaceae. These diet- and sex- effects were less pronounced in the APOE3 mice, indicating that different APOE variants also play a significant role. The findings provide insights into the higher susceptibility of APOE4 females to AD, potentially due to inefficient energy production, and imply the importance of considering precision nutrition for mitigating dysbiosis and AD risk in the future.

Environmental exposure to chlorpyrifos during gestation, APOE polymorphism and the risk on autistic-like behaviors

Autism spectrum disorder (ASD) encompasses several neurodevelopmental conditions characterized by communication and social impairment, as well as repetitive patterns of behavior. However, it can co-occur with other mental conditions such as anxiety. The massive use of chlorpyrifos (CPF) has been linked to the increased prevalence of developmental disorders. Likewise, ASD has also been closely linked to a wide variety of genetic factors. The aims of the present investigation are to study how gestational CPF exposure and APOE polymorphism affects communication skills, early development and mid-term anxiety-like behaviors, as well as, changes in gene expression related to the cholinergic system. C57BL/6J and humanized apoE3 and apoE4 homozygous mice were exposed to 0 or 1 mg/kg/day of CPF through the diet, from gestational day (GD) 12–18. In addition, a group of C57BL/6J females were injected subcutaneously with 300 mg/kg/day of valproic acid (VPA) on GD 12 and 13. This group was used as a positive control for studying some core and associated autism-like behaviors. Communication skills by means of ultrasonic vocalizations and physical/motor development were assessed during the preweaning period, whereas locomotor activity, anxiety-like behaviors and the gene expression of cholinergic elements were evaluated during adolescence. Our results showed that C57BL/6J mice prenatally exposed to CPF or VPA showed a decrease in body weight and a delay in eye opening. Communication and anxiety behavior were affected differently depending on treatment, while gene expression was altered by sex and treatment. In addition, none of the parameters evaluated in apoE transgenic mice exposed to CPF were affected, but there were differences between genotypes. Therefore, we suggest that prenatal CPF exposure and VPA produce divergent effects on communication and anxiety.

ApoE4 exacerbates the senescence of hippocampal neurons and spatial cognitive impairment by downregulating acetyl-CoA level.

Although aging and apolipoprotein E (APOE) ε4 allele have been documented as two major risk factors for late-onset Alzheimer's disease (LOAD), their interaction and potential underlying mechanisms remain unelucidated. Using humanized ApoE4- and ApoE3- target replacement mice, we found the accumulation of senescent neurons and the activation of mTOR and endosome-lysosome-autophagy (ELA) system in the hippocampus of aged ApoE4 mice. Further analyses revealed that ApoE4 aggravated the profile change of hippocampal transcription and metabolism in an age-dependent manner, accompanying with an disruption of metabolism, which is presented with the downregulating activity of citrate synthase, the level of ATP and, most importantly, the level of acetyl coenzyme A (Ac-CoA); GTA supplement, an Ac-CoA substrate, reversed the senescent characteristics, decreased the activation of mTOR and ELA system, and enhanced the synaptic structure and increasing level of pre-/post-synaptic plasticity-related protein, leading to cognitive improvement in aged ApoE4 mice. These data suggest that ApoE4 exacerbates neuronal senescence due to a deficiency of acetyl-CoA, which can be ameliorated by GTA supplement. The findings provide novel insights into the potential therapeutic value of GTA supplement for the cognitive improvement in aged APOE4 carriers.

Abstract P3022: Smyd2 Deficiency Protects Against Atherosclerotic Plaque Formation Through Modulation Of Oxidative Stress

Introduction: Oxidative stress induced vascular remodeling promotes atherosclerosis. By scavenging excessive ROS levels, NRF2 protects vascular cells against oxidative stress and exerts anti-inflammatory activity through modulation of HO-1. This study aimed to explore smooth muscle cell (SMC)-specific role of SMYD2 in modulating atherosclerosis associated oxidative stress in vivo and in vitro . Methods: SMC-specific SMYD2 knockout mice in ApoE -/- background (SMYD2 ΔSMC ApoE -/- ) and their littermate controls (SMYD2 fl/fl ApoE -/- ) were subjected to Western diet feeding for 18-26 weeks. Atherosclerotic plaque distribution was examined by en-face Oil Red O (ORO) staining of aortas from both groups. Hematoxylin & Eosin (H&E) staining on brachiocephalic artery tissues were performed to determine atherosclerotic plaque burden by measurements of total plaque size, lipid pool area, and necrotic core area. Lipid peroxidation was analyzed in aortic tissues and in blood serum. Primary VSMCs isolated from control and SMYD2 depleted mice and cultured C3H/ 10T1 / 2 cells were used for in vitro studies. Results: SMC-specific SMYD2 depletion attenuated atherosclerotic plaque establishment and progression. En -face ORO analysis revealed significant suppression of ORO positive areas in SMYD2 ΔSMC ApoE -/- mice compared to control mice. H&E analysis of lesions showed significant reduction of total plaque size, despite accumulation of lipids, in SMYD2 ΔSMC ApoE -/- mice as compared to control mice. Immunofluorescence staining of aortic tissues showed increased expression of NRF2, HO-1 and GPX4 in SMYD2 ΔSMC ApoE -/- mice compared to controls, as observed in serum by MDA assay. Mechanistic studies using C3H/ 10T1 / 2 cells treated with PAz-PC (oxLDL), GPX4 inhibitor, with or without SMYD2 knockout, demonstrated that SMYD2 modulates HO-1 and GPX4 expression through NRF2. Specifically, SMYD2 binds NRF2 and reduces its translocation into the nucleus, thereby repressing HO-1 expression and inducing oxidative stress. Conclusion: SMYD2 depletion induces atheroprotective effect via reducing plaque burden and promoting antioxidant activity of NRF2/HO-1 axis, suggesting SMYD2 could be a therapeutic target against atherosclerosis.

Viscoelastic Characteristics in Mouse Model of Hepatic Steatosis With Inflammation by Kelvin-Voigt Fractional Derivative Modeling.

The aim of the work described here was to measure the characteristics of viscoelasticity and fluidity in a mouse model of hepatic steatosis and inflammation using a nano-indentation test and the Kelvin-Voigt fractional derivative (KVFD) model and to explore the viscoelasticity and fluidity characteristics in mice with different degrees of hepatic steatosis with inflammation. Twenty-five ApoE mice were randomly divided into an experimental high-fat diet group (n=15) and an ordinary-food control group (n=10), then subdivided into four subgroups based on pathological degree of hepatic steatosis: S0 (normal), S1 (mild), S2 (moderate) and S3 (severe). The 25 liver specimens from these mice were evaluated by a slope-keeping relaxation nano-indentation test. Elasticity (E0) was significantly higher in the S3 group than in the S1 and S2 groups, while fluidity (α) and viscosity (τ) were significantly lower in S3 than in S1 and S2 (all p values < 0.05). The following cutoff values for the diagnosis of hepatic steatosis >33% with inflammation were also determined: E0 > 85.01 Pa (area under the curve [AUC]: 0.917, 95% confidence interval [CI]: 0.735-0.989), α ≤ 0.38 (AUC: 0.885, 95% CI: 0.695-0.977),\ and τ ≤ 3.92 (AUC: 0.813, 95% CI: 0.607-0.939). Increases in the degree of hepatic steatosis with inflammation in mice paralleled gradual increases in the stiffness of the liver and gradual decreases in the fluidity and viscosity of the liver.

Supplement of exogenous inorganic pyrophosphate inhibits atheromatous calcification in Apolipoprotein E knockout mice

Inorganic pyrophosphate (PPi) is the endogenous inhibitor for vascular calcification (VC). The present study was to investigate the effects of adenosine disodium triphosphate (ADTP) and alendronate sodium (AL), two exogenous PPi sources, on the atheromatous calcification (AC) in Apolipoprotein E knockout (ApoE KO) mice. ApoE KO mice were randomly divided into five groups: ApoE KO group, ApoE KO + ADTP (Low) group, ApoE KO + ADTP (High) group, ApoE KO + AL (Low) group and ApoE KO + AL (High) group. The mice in ApoE KO + ADTP (Low) group and ApoE KO + ADTP (High) group were intraperitoneally injected with ADTP with dose of 0.5 and 1.0 mg/kg/day for 2 months respectively. The mice in ApoE KO + AL (Low) group and ApoE KO + AL (High) group were intraperitoneally injected with AL with dose of 0.6 and 1.2 mg/kg/day for 2 months respectively. The age matched C57 mice were used as control group. All ApoE KO and C57 mice were fed with normal chow throughout the experiment. The calcification was evaluated using von Kossa method. The contents of PPi, triglyceride (TG), total cholesterol (TC), high density lipoprotein (HDL) and low density lipoprotein (LDL), tumor necrosis factor α (TNF-α), interleukin-6 (IL-6), interferon-γ (IFN-γ) and interleukin-10 (IL-10) as well as the activity of alkaline phosphatase (ALP) in serum were measured. The results showed that compared with C57 mice, ApoE KO mice developed severe AC accompanied with high levels of TC, TG, LDL, IL-6, TNF-α and IFN-γ in serum and with low levels of PPi and IL-10 in serum. Both ADTP and AL dose-dependently reduced the AC in ApoE KO mice compared with that of ApoE mice, without affecting the contents of lipid profiles. In addition, ADTP and AL increased the contents of PPi and IL-10 while decreased the contents of TNF-α, IL-6 and IFN-γ in serum of ApoE KO mice, having no affection on ALP activity. The results suggested that ADTP and AL reduced AC in ApoE KO mice by increasing the PPi level and regulating the inflammation.

Intraperitoneal Injection of Human Ferritin Heavy Chain Attenuates the Atherosclerotic Process in APOE-Knockout Mice.

Background: Iron overload can accelerate the accumulation of lipid oxides and contribute to the progression of atherosclerosis. Ferritin heavy chain (FT-H) exhibits oxidase activity, which inhibits the toxicity of ferrous ions and reduces oxidative damage. We investigated the effect of the intraperitoneal injection of FT-H on the progression of atherosclerosis in APOE-knockout mice (Apo-E(-/-) mice). Methods: All mice were fed on a high-fat diet. After 10 weeks, the mice were divided into an injection group (n = 4) and a control group (n = 4). The injection group was injected intraperitoneally with FT-H (50 mg/kg, once a week), and the control group was treated with PBS buffer (at an equal volume to the injection group, once a week). After 10 weeks of intervention, MRI of the aortas was performed. Then, the animals were sacrificed, and tissues were taken. Hematoxylin-eosin (HE) staining was used for histomorphometry, Masson staining was used to quantify the collagen content in the arteries, Prussian blue staining was used to visualize iron deposition in the arteries, and MRI was used to analyze the structure of the aorta in vivo. Immunohistochemistry was performed to detect the expression of MCP-1, MMP-2, MMP-9, FT-H, FT-L, TfR1, NRF-2 and GPX-4. Results: The serological results showed that the injection group had lower levels of glucose (Glu), triacylglycerol (TG), cholesterol (CHO), low-density lipoprotein-C (LDL-C) and malondialdehyde (MDA) (p = 0.0058, p = 0.0098, p = 0.0019, p = 0.0368 and p = 0.0025, respectively), and their serum ferritin (SF) and superoxide dismutase (SOD) levels were higher (p = 0.0004 and p < 0.0001). The Masson staining and MRI results showed that the injection group had less collagen deposition (p = 0.0226), a larger arterial lumen area and arterial volume (p = 0.0006 and p = 0.0005), thinner arterial wall thickness (p = 0.0013) and a more stable arterial plaque structure (p < 0.0001). The immunohistochemical results showed reduced expression of FT-H, FT-L, TfR1, MMP-2, MMP-9, MCP-1 and NRF-2 in the injection group (p = 0.0054, p = 0.0242, p = 0.0221, p = 0.0477, p = 0.0131, p = 0.0435 and p = 0.0179). Prussian blue staining showed that the area of iron-positive areas in the aortic plaques of the control group was larger than that of injected group. The expression of GPX-4 was lower in the control group than in the injection group (p = 0.016). Conclusions: The intraperitoneal administration of FT-H to Apo-E(-/-) mice resulted in lower blood glucose and lipid levels; reduced iron and iron metabolism protein deposition in the aorta; reduced indices of their ferroptosis, oxidation and inflammatory aggregation; and reduced collagen deposition in the aorta, which delayed the process of aortic atherosclerosis in mice.

Developmental brain lipidomics is influenced by postnatal chlorpyrifos exposure and APOE genetic background in mice

Lipids are a major component of the brain, and are involved in structural and neurodevelopmental processes such as neurogenesis, synaptogenesis and signaling. Apolipoprotein E (apoE) is the main lipoprotein involved in lipid transport in the brain. The apoE isoforms can determine vulnerability to the toxic effects of the pesticide chlorpyrifos (CPF), which can interfere with normal neurodevelopment. We aimed to study the effects of postnatal exposure to CPF and of the APOE genotype on the lipid composition of the brain at early ages. For it, we used apoE3 and apoE4 targeted-replacement (TR) male mice, as well as wild-type C57BL/6. The mice were orally exposed to 1 mg/kg/day of CPF on postnatal days 10–15 and, four hours after the treatment, we obtained samples to assess the cerebral lipid composition. Differences between APOE genotypes were found in the cerebral lipid profile in the postnatal period. ApoE4-TR mice exhibited higher lipid concentrations compared to the other groups in most of the cases. CPF exposure led to a decrease in cholesteryl ester and triglyceride concentrations, while modulating the levels of phosphatidylcholine species based on the apoE isoform. Specifically, CPF treatment decreased the concentration of some species of this lipid (PC30:0, PC31:0, PC32:2, PC36:5, PC40:4 and PC40:5) in C57BL/6 mice exposed to CPF, increased (PC31:0 and PC37:6) in apoE3-TR exposed mice while exposed apoE4-TR mice remained unaltered. These results provide further insights into the lipid composition of the brain at early ages, and how it can be modulated by environmental and genetic factors.

Prenatal stress aggravates age-dependent cognitive decline, insulin signaling dysfunction, and the pro-inflammatory response in the APPNL-F/NL-F mouse model of Alzheimer's disease

Accumulating evidence indicates that early adverse life experiences may be involved in the pathogenesis of Alzheimer's disease (AD). Prenatal stress (PS) can affect brain maturation and neuroimmune and metabolic interactions, leading to age-dependent cognitive deficits in offspring. However, a multi-faceted cause-and-effect impact of PS on the development of cognitive deficits in the process of physiological ageing and in the APPNL-F/NL-F mouse model of Alzheimer's disease has not yet been evaluated.We have identified age-dependent cognitive learning and memory deficits using male C57BL/6 J (wild type, WT) and the knock-in APPNL-F/NL-F (KI) aged 12, 15, and 18 months. An increase in the Aβ42/Aβ40 ratio and mouse ApoE levels in the hippocampus and frontal cortex preceded the onset of cognitive deficits in the KI mice. Moreover, dysfunction in insulin signaling, including increased IRS-1 serine phosphorylation in both brain areas and the tyrosine phosphorylation deficit in the frontal cortex, suggested age-dependent insulin/IGF-1 resistance. Resistance was reflected by disturbances in mTOR or ERK1/2 kinase phosphorylation and excessive pro-inflammatory (TNF-α, IL-6, and IL-23) status in the KI mice. Importantly, our study has provided insights into the higher vulnerability to PS-induced exacerbation of age-dependent cognitive deficits and biochemical dysfunction in KI mice than in WT animals.We anticipate our study will lead to future investigation of a multi-faceted cause-and-effect relationship between stress during neurodevelopment and the onset of AD pathology, distinguishing it from changes in the course of dementia during normal ageing.

Synaptojanin 1 Modulates Functional Recovery After Incomplete Spinal Cord Injury in Male Apolipoprotein E Epsilon 4 Mice.

Apolipoprotein E epsilon 4 (ApoE4) is the second most common variant of ApoE, being present in ∼14% of the population. Clinical reports identify ApoE4 as a genetic risk factor for poor outcomes after traumatic spinal cord injury (SCI) and spinal cord diseases such as cervical myelopathy. To date, there is no intervention to promote recovery of function after SCI/spinal cord diseases that is specifically targeted at ApoE4-associated impairment. Studies in the human and mouse brain link ApoE4 to elevated levels of synaptojanin 1 (synj1), a lipid phosphatase that degrades phosphoinositol 4,5-bisphosphate (PIP2) into inositol 4-monophosphate. Synj1 regulates rearrangements of the cytoskeleton as well as endocytosis and trafficking of synaptic vesicles. We report here that, as compared to ApoE3 mice, levels of synj1 messenger RNA and protein were elevated in spinal cords of healthy ApoE4 mice associated with lower PIP2 levels. Using a moderate-severity model of contusion SCI in mice, we found that genetic reduction of synj1 improved locomotor function recovery at 14 days after SCI in ApoE4 mice without altering spared white matter. Genetic reduction of synj1 did not alter locomotor recovery of ApoE3 mice after SCI. Bulk RNA sequencing revealed that at 14 days after SCI in ApoE4 mice, genetic reduction of synj1 upregulated genes involved in glutaminergic synaptic transmission just above and below the lesion. Overall, our findings provide evidence for a link between synj1 to poor outcomes after SCI in ApoE4 mice, up to 14 days post-injury, through mechanisms that may involve the function of excitatory glutaminergic neurons.

Downregulation of SIRT1, SIRT3, and IGF-1 in ApoE-deficient mice exacerbates neuronal damage induced by chronic cerebral hypoperfusion

Chronic cerebral hypoperfusion is a widespread pathological condition caused by chronically reduced cerebral blood flow leading to brain damage, but the specific molecular mechanisms that regulate these phenomena remain poorly understood. In this study, we investigated brain damage and neuronal DNA injury in a vulnerable region of the brain, the hippocampus, as well as the involvement of apolipoprotein E (ApoE), sirtuins of 1 (SIRT1) and 3 (SIRT3) types and insulin-like growth factor 1 (IGF-1) in pathogenetic mechanisms in mice with chronic cerebral hypoperfusion caused by the permanent occlusion of the left unilateral common carotid artery. Male C57/6j (C57, wild type) and ApoE(-/-) mice were divided into four experimental groups (10 mice per group): sham-operated С57, С57 with chronic cerebral hypoperfusion, sham-operated ApoE(-/-) mice, ApoE(-/-) mice with chronic cerebral hypoperfusion. Our results showed that the number of damaged neurons in the hippocampus at 8 weeks after surgical manipulation increased in both groups of mice with chronic cerebral hypoperfusion, with more pronounced rates in ApoE(-/-) mice than in C57 mice. However, ApoE deficiency in moderate chronic cerebral hypoperfusion was accompanied by a higher level of undamaged DNA (class 0) and a low level of maximally damaged DNA (class 4) in brain cell nuclei in contrast to group C57. In ApoE-deficient mice, reduced expression of SIRT1, SIRT3, and IGF-1 was found. In chronic cerebral hypoperfusion, expression of sirtuins was preserved, but IGF-1 expression was significantly reduced in ApoE(-/-) mice in comparison to C57. The obtained results indicate that ApoE deficiency leads to downregulation of SIRT1, SIRT3 and IGF-1 in the brain; this lack of cytoprotection is enhanced in chronic cerebral hypoperfusion and may participate in the mechanisms of neuronal damage.

APOE4 expression confers a mild, persistent reduction in neurovascular function in the visual cortex and hippocampus of awake mice

Vascular factors are known to be early and important players in Alzheimer’s disease (AD) development, however the role of the ε4 allele of the Apolipoprotein (APOE) gene (a risk factor for developing AD) remains unclear. APOE4 genotype is associated with early and severe neocortical vascular deficits in anaesthetised mice, but in humans, vascular and cognitive dysfunction are focused on the hippocampal formation and appear later. How APOE4 might interact with the vasculature to confer AD risk during the preclinical phase represents a gap in existing knowledge. To avoid potential confounds of anaesthesia and to explore regions most relevant for human disease, we studied the visual cortex and hippocampus of awake APOE3 and APOE4-TR mice using 2-photon microscopy of neurons and blood vessels. We found mild vascular deficits: vascular density and functional hyperaemia were unaffected in APOE4 mice, and neuronal or vascular function did not decrease up to late middle-age. Instead, vascular responsiveness was lower, arteriole vasomotion was reduced and neuronal calcium signals during visual stimulation were increased. This suggests that, alone, APOE4 expression is not catastrophic but stably alters neurovascular physiology. We suggest this state makes APOE4 carriers more sensitive to subsequent insults such as injury or beta amyloid accumulation.

Humanized APOE genotypes influence lifespan independently of tau aggregation in the P301S mouse model of tauopathy

Apolipoprotein (APOE) E4 isoform is a major risk factor of Alzheimer’s disease and contributes to metabolic and neuropathological abnormalities during brain aging. To provide insights into whether APOE4 genotype is related to tau-associated neurodegeneration, we have generated human P301S mutant tau transgenic mice (PS19) that carry humanized APOE alleles (APOE2, APOE3 or APOE4). In aging mice that succumbed to paralysis, PS19 mice homozygous for APOE3 had the longest lifespan when compared to APOE4 and APOE2 homozygous mice (APOE3 > APOE4 ~ APOE2). Heterozygous mice with one human APOE and one mouse Apoe allele did not show any variations in lifespan. At end-stage, PS19 mice homozygous for APOE3 and APOE4 showed equivalent levels of phosphorylated tau burden, inflammation levels and ventricular volumes. Compared to these cohorts, PS19 mice homozygous for APOE2 showed lower induction of phosphorylation on selective epitopes, though the effect sizes were small and variable. In spite of this, the APOE2 cohort showed shorter lifespan relative to APOE3 homozygous mice. None of the cohorts accumulated appreciable levels of phosphorylated tau compartmentalized in the insoluble cell fraction. RNAseq analysis showed that the induction of immune gene expression was comparable across all the APOE genotypes in PS19 mice. Notably, the APOE4 homozygous mice showed additional induction of transcripts corresponding to the Alzheimer’s disease-related plaque-induced gene signature. In human Alzheimer’s disease brain tissues, we found no direct correlation between higher burden of phosphorylated tau and APOE4 genotype. As expected, there was a strong correlation between phosphorylated tau burden with amyloid deposition in APOE4-positive Alzheimer’s disease cases. Overall, our results indicate that APOE3 genotype may confer some resilience to tauopathy, while APOE4 and APOE2 may act through multiple pathways to increase the pathogenicity in the context of tauopathy.

Amelioration of Cognitive and Olfactory System Deficits in APOE4 Transgenic Mice with DHA Treatment.

Olfactory dysfunction and atrophy of olfactory brain regions are observed early in mild cognitive impairment and Alzheimer disease. Despite substantial evidence showing neuroprotective effects in MCI/AD with treatment of docosahexaenoic acid (DHA), an omega-3 fatty acid, few studies have assessed DHA and its effects on the olfactory system deficits. We therefore performed structural (MRI), functional (olfactory behavior, novel object recognition), and molecular (markers of apoptosis and inflammation) assessments of APOE4 and wild-type mice ± DHA treatment at 3, 6, and 12 months of age. Our results demonstrate that APOE4 mice treated with the control diet show recognition memory deficits, abnormal olfactory habituation, and discrimination abilities and an increase in IBA-1 immunoreactivity in the olfactory bulb. These phenotypes were not present in APOE4 mice treated with a DHA diet. Alterations in some brain regions' weights and/or volumes were observed in the APOPE4 mice and may be due to caspase activation and/or neuroinflammatory events. These results suggest that the consumption of a diet rich in DHA may provide some benefit to E4 carriers but may not alleviate all symptoms.

Allelic variation within the major APOE CpG island affects its methylation in the brain of targeted replacement mice expressing human APOE

•The number of cytosine-phosphate-guanine (CpG) sites differs due to sequence variation in the human apolipoprotein E (APOE) gene.•APOE DNA methylation is allele-dependently altered corresponding to the total number of CpG pairs in the brain of APOE targeted replacement mice (APOE εpsilon 4 > εpsilon 3 > εpsilon 2).•Binding of the methyl-CpG binding protein 2 to genomic APOE was in trend less pronounced in the brain of APOE4 mice.