A new study shows that APOE4 reshapes brain-linked immune and lymphatic pathways in strikingly different ways in females and males, with opposite cognitive effects when innate immunity is suppressed.
Study: Sex-specific APOE4-dependent innate immunity regulates meningeal lymphatics, brain lipids, neuroinflammation, and cognition. Image Credit: ahmetmapush / Shutterstock
In a recent study published in the journal Neuron, researchers showed that apolipoprotein E ε4 (APOE4) expression leads to sexually dimorphic inflammatory, lymphatic, lipid, neuroinflammatory, and cognitive responses.
APOE is the primary lipid carrier in the brain. In humans, the APOE gene has three alleles - APOE2, APOE3 (E3), and APOE4 (E4), with E3/E3 being the most common genotype in the population. In Caucasians, the expression of one or two E4 alleles increases AD risk by 3- to 4-fold or 9- to 15-fold, respectively.
APOE4 and sex can independently modulate glial, immune, and vascular responses in the brain. Recent rodent studies have shown that peripheral and neural cells expressing E4 exhibit sexual dimorphism. However, it is unclear why females have a greater risk of AD and how E4 shifts the balance toward dementia.
Meningeal Immune Cell Changes in APOE4 Mice
In the present study, researchers showed that E4 expression leads to sexually dimorphic inflammatory and lymphatic responses. First, meningeal dura samples were collected from middle-aged mice (12-13 months old) expressing two alleles of E4 or E3 instead of murine Apoe. The frequencies of innate immune cells were assessed by flow cytometry.
Despite similar frequencies and numbers of innate immune cells in E4/E4 and E3/E3 mice within each sex, there was an increase in macrophages expressing major histocompatibility complex (MHC) class II and cluster of differentiation 206 (CD206) in females, irrespective of their genotype. Next, the researchers investigated the effects of E4 or E3 expression on the lymphatic vasculature.
Further, four-week exposure to PLX5622 (PLX), a CSF1R inhibitor, reduced macrophage frequencies. In particular, there was a consistent and selective depletion of CD206+MHC class II+ macrophages in both male and female mice, irrespective of their APOE genotype. PLX exposure also decreased APOE expression in macrophages, blood endothelial cells (BECs), mast cells, and fibroblasts, regardless of sex and genotype.
Macrophages showed the highest differentially expressed genes (DEGs) across comparisons between groups. E4/E4 PLX female mice had particularly elevated DEGs in macrophages, B cells, and BECs compared to female E4/E4 controls. In E4/E4 PLX male mice, BECs and B cells showed the highest DEGs compared to male E4/E4 controls; they also had substantially higher DEGs in natural killer cells, T cells, dendritic cells, fibroblasts, and innate lymphoid cells than male E4/E4 controls.
APOE4 Effects on Lymphatic Vessels and CSF Drainage
Analyses of meningeal dura from younger mice (2-4 months old) or middle-aged mice showed a sex- and age-dependent effect of E4 expression on the total length of vessels expressing the lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1). Specifically, longer LYVE-1+ vessels were noted in male E4/E4 mice, but not in females, at 12-13 months compared to age-matched E3/E3 males. There were no group differences in lymphatic morphology at the younger age.
In addition, middle-aged male E4/E4 mice showed reduced outflow of cerebrospinal fluid (CSF) into the deep cervical lymph nodes (LNs) compared to age-matched E4/E4 females. This finding suggests that greater meningeal lymphatic vessel coverage in males was not necessarily beneficial, because it was accompanied by impaired drainage. Single-cell RNA sequencing analysis revealed that two E4 alleles result in distinct, significant changes in gene expression in meningeal dural immune cells of middle-aged mice.
Neuroinflammation, Brain Lipids, and Cognitive Performance
Next, the team investigated the influence of E4 expression on inflammatory chemokines and cytokines in the forebrain of middle-aged mice. E4/E4 females had higher levels of inflammatory cytokines and chemokines than E3/E3 females. In contrast, E3/E3 and E4/E4 males had comparable inflammatory profiles, consistent with the authors’ interpretation that males may show relative resilience to APOE4-linked neuroinflammatory changes at this stage. PLX exposure was associated with higher interleukin (IL)-22 and lower IL-1α, IL-16, and C-C motif chemokine ligand 12 (CCL12) levels, irrespective of APOE genotype and sex.
In addition, E4/E4 PLX females showed lower levels of programmed cell death ligand 2, while E4/E4 PLX males had higher levels of CCL4, CSF1, fibroblast growth factor 21 (FGF21), CCL2, and hepatocyte growth factor (HGF) than sex-matched E4/E4 controls. The study also identified sex-specific effects of APOE4 on brain lipid profiles. Next, the researchers examined the cognitive function of middle-aged E4/E4 and E3/E3 mice in fear-conditioning and open field tests. In the open field test, the total distance traveled was similar across the female groups.
However, the distance traveled in the center of the arena was significantly lower between female E4/E4 control and PLX groups. Male groups had no differences in the open field test. In the fear-conditioning test, E4/E4 females showed a shorter freezing time during the cued trial, indicating worse cognitive function than E3/E3 females. Notably, PLX exposure led to poor performance in the context trial for E3/E3 females but improved performance in the cued trial for E4/E4 females, suggesting that suppressing innate immune responses may lessen cognitive vulnerability in female E4/E4 mice.
In contrast, E4/E4 males, which had similar fear-freezing times to E3/E3 males, showed shorter freezing times in both trials after PLX exposure, indicating worsened cognitive function. Thus, suppressing innate immunity had divergent effects, appearing beneficial in E4/E4 females but detrimental in E4/E4 males. Finally, the researchers integrated six publicly available single-nucleus RNA sequencing datasets of human brain cells from patients with and without AD. They noted that each brain leukocyte population exhibited distinct responses to E4 expression in males and females.
Sex-Specific APOE4 Implications for Alzheimer’s Therapy
In sum, the results emphasize that the meningeal dura could be a relevant neuroimmune niche in the context of aging-related neurodegenerative disorders, whose risk varies by sex, age, and E4 status. Determining the roles and mechanisms underlying immune responses in males and females will be crucial to developing tailored immunotherapeutic approaches for cognitive decline due to APOE4. Overall, the findings suggest earlier APOE4-linked neuroinflammatory and cognitive vulnerability in females, while males may show relative resilience at this stage despite altered lymphatic structure and drainage.
Scientists discover similarities in brain aging between mice and humans