Chronic nSMase2 Inhibition Suppresses Neuronal Exosome Spreading In Vivo & Induces Sex‐Specific Attenuation of AD Pathology in APPNL‐F/NL‐F Knock‐In Mice

Abstract

AbstractBackgroundAltered ceramide metabolism is implicated in Alzheimer’s disease (AD) progression. Although prior work shows that inhibition of neutral sphingomyelinase (nSMase) 2, a key enzyme responsible for hydrolysis of sphingomyelin to ceramide, reduces exosome secretion, attenuates Aβ pathology, and improves cognitive function, these studies have been limited to in vitro application or short‐term treatments in fast‐progressing models.MethodWe employed the slow‐progressing APPNL‐F/NL‐F knock‐in (APP KI) AD mouse model and cell type‐specific exosome reporter CD63‐GFPf/f mice to investigate the impact of chronic long‐term nSmase2 inhibition on in vivo neuron‐derived exosome dynamics, region‐specific Aβ plaque load, and cognitive function. Male and female APPNL‐F/NL‐F‐CD63‐GFPf/+ (APP KI‐CD63) mice were treated daily with the nSmase2 inhibitor, GW4869 (GW; 1.25mg/kg BW; i.p.), or vehicle (VEH, DMSO in saline), from 8 months old (mo.) to 13mo. and from 10 mo. to 18mo. We also analyzed ceramide levels in both APP KI and WT mice. Brain‐derived exosomes of 14mo. APP KI females versus males and age‐ and sex‐matched WT mice were also isolated and quantified.ResultWe found significant elevations in multiple ceramide species from somatosensory cortex (SC) samples of 14mo. APP KI mice compared to age‐ and sex‐matched WT controls. We also found a sex‐specific increase in total exosomes in 14mo. APP KI females versus males that was absent in WT mice. Chronic inhibition of nSMase2 by GW4869 leads to significantly suppression of neuronal exosome spreading. Non‐linear regression analyses of size distribution curves showed consistent shifts to smaller plaque sizes in GW‐treated APP KI females, which were accompanied by decreased plaque numbers in each region examined. Conversely, plaque load was largely unaffected by GW treatment in APP KI males. T maze trials to examine spatial working memory revealed a female‐specific reduction in spontaneous alternation rate in APP KI vs. WT mice, which was significantly reversed with GW treatment.ConclusionOur results demonstrate that chronic nSMase2 inhibition suppresses neuronal exosome spreading in APPNL‐F/NL‐F knock‐in mice and preferentially attenuates Aβ pathology and cognitive dysfunction in females, pointing to a strong contribution of sex‐specific alterations in ceramide metabolism and exosome dynamics to the progression of AD‐like pathology in this model.