The consideration of sex dimension is pivotal to improve our understanding of neurodegeneration and the biological mechanisms that contribute to the etiology, manifestation, and potential therapeutics of neurological syndromes [40]. In fact, multiple health incongruities in therapeutic and diagnostic fields have been associated with the lack of inclusion of women in basic research as well as women in clinical trials [41, 42]. Due to smell deficit has been proposed as an early indicator of AD and PD [43] and sex differences exist in terms of olfactory functionality [16–19], we consider that a better comprehension of the molecular mechanisms disrupted at olfactory level in a sex-dependent manner might offer unknown targets for earlier diagnosis and therapeutic intervention in neurological disorders. Up to now, the OT characterization has been mainly addressed in two different contexts. In humans, electrophysiological testing, together with morphological imaging approaches are used to evaluate the OT in the clinical assessment of the sense of smell [44]. In mice, the OT is broadly studied as a model to understand the mechanisms underlying the guidance of growing axons [7]. In this work, we report that: i) OT proteomic alteration is more severe in AD than in PD; ii) this OT proteostatic imbalance is higher in women than in men independent of the disease; iii) despite the dissimilar molecular profiles unveiled between women and men, common protein intermediates and biological pathways have been identified across both diseases; iv) experimentally-demonstrated protein interactors of canonical neuropathological substrates are differentially modulated across both sexes at OT level; v) specific olfactory signaling routes governed by NFkB are differentially modulated across sexes in AD and PD; vi) sex-dependent protein acetylation changes are evidenced across the olfactory axis in both diseases and vii) sex-specific sirtuin signaling imbalance occurs across the olfactory axis in AD and PD, in which protein expression changes of sirtuin family members at the level of OT are extensively more dynamic. All these sex-specific molecular profiles provide novel mechanistic clues on the divergent mechanisms involved in the olfactory neurodegeneration that occur in AD and PD [6].
The minimal overlap observed in OT proteome remodeling between women and men (in both diseases) support the hypothesis that distinctive pathophysiological processes are involved in the olfactory neurodegenerative process. This argument is clearly supported when the neuropathological stage is also considered (Supplementary Fig. 7). Although, common pathways are disrupted in AD advanced stages (Braak V-VI) and PD advanced stage (neocortical stage) independent of the sex variable, multiple biofunctions are specifically altered in a sex- and neuropathological stage-dependent manner (Supplementary Fig. 7). Due to the inherent limitations of our sample cohort, further research is needed to analyze the dual effect of sex and neuropathological grading in initial phases of AD and PD at olfactory level. SIRTs correspond to class III histone deacetylase enzymes (found throughout different cellular compartments) targeting both histone and non-histone substrates involved in metabolism, myelination and apoptosis [45–47]. From a neuropathological point of view, although SIRTs have been previously associated to multiple neurological syndromes [39, 48, 49], sex dimension has not been broadly considered. We have observed a tangled sex-dependent SIRT protein expression profile (SIRT1, 2, 3 and 5) that significantly differ in AD and PD across primary and secondary olfactory areas. Specifically, SIRT1 was increased in the OB and EC in AD women and men. It has been shown that SIRT1 protects neuronal axons, induces neurite outgrowth and regulates long-term potentiation and neurogenesis [50, 51]. SIRT1 levels are brain region-dependent in AD [49], being able to degrade the Aβ peptide in primary astrocytes and to reduce ROS and peroxidation levels in APP/PS1 AD model, decreasing senile plaques and improving learning and memory activities [52–54]. In addition, SIRT1 also interferes with Tau metabolism [55], reducing the acetylated tau levels and avoiding the propagation of pathological Tau [56, 57]. All these beneficial effects, together with the up-regulation of multiple neurotrophic factors (BDNF, GDNF and VEGF) by SIRT1 [58], could indicate a general neuroprotective mechanism through the olfactory axis in AD, specially at the level of the OB and EC. However, we observed a specific drop in SIRT1 levels in the OT derived from PD subjects. It has been demonstrated that SIRT1 activity slows down when extracellular α-synuclein is present as well as in PD-post mortem brain material [59, 60]. Moreover, pharmacological activation of SIRT1 triggers α-synuclein degradation through the deacetylation of LC3 and up-regulation of LC3-II [61]. Interestingly, it is known that SIRT1 presents known roles in recovery olfactory function. Specifically, SIRT1 is involved in olfactory function maintenance through the protection of subventricular zone-derived neural stem cells from DNA Double-Strand Breaks [62]. Moreover, the increment in olfactory bulb SIRT1 expression is associated with the recovery of olfactory function under bulbar excitotoxic insults [63]. Despite multiple evidence point out that SIRT1 is an olfactory-promoting factor and neuroprotective in different neurodegenerative contexts, it seems obvious that its positive role may depend on multiple factors such as oxidative stress levels [64] and the neuropathological damage across the olfactory axis.
Although SIRT2 temporal cortical levels are increased in AD patients [65], we observed a specific significant decrease in the OT from AD men. In general, and in direct opposition to SIRT1, reduced expression levels of SIRT2 tend to be beneficial in the context of AD. In vitro and in vivo studies have revealed that low SIRT2 levels diminish Aβ toxicity [65–67] and Tau phosphorylation [68]. This conclusion has been also corroborated using SIRT2 inhibitors in several AD models [69, 70]. Respect to PD, despite SIRT2 levels are unchanged in the substantia nigra pars compacta of PD subjects [71], OT SIRT2 levels were down-regulated in PD women. Experimental data of SIRT2 expression in PD models are significantly divergent [72, 73]. Due to acetylation induces a reduction in α-synuclein oligomerization and aggregation, and SIRT2 interacts with alpha-synuclein leading to its deacetylation [74, 75], SIRT2 inhibition has been proposed as a potential therapeutics against synucleinopathies [76, 77].
SIRT3 expression diminishes in cortical regions from APP/PS1 AD model as well as in AD patients [78–80]. In our case, a significant decrease was observed in OT from AD women and EC from AD men. Although no correlation has been observed between SIRT3 and Aβ plaques across different human brain areas [49], SIRT3 activation protects neurons from Aβ toxicity [81–83] and reduces Tau and acetylated Tau [84], probably through a mitochondrial-related energetic route [85, 86]. In relation to PD, a decrease in SIRT3 levels was observed in the OT from PD men whereas SIRT3 increment was evidenced in the amygdala from the same group. SIRT3 reduction increases ROS production and α-synuclein aggregation, triggering the loss of dopaminergic neurons [87, 88]. Moreover, SIRT3 overexpression impacts on mitochondrial bioenergetics, mitigating the oxidative stress through the SIRT3-mediated deacetylation of Mn-SOD [89, 90] and increasing the autophagic capacity through the LKB1-AMPK-mTOR pathway, inducing ROS alleviation and inhibiting alpha-synuclein accumulation [91]. We have evidenced a significant drop in SIRT5 levels in the OT from AD women as well as an increment in protein levels at the level of OB and OT from AD and PD men. Unlike other SIRTs, SIRT5 is involved in deacetylation, demalonylation, deglutarylation and desuccinylation processes [64]. Specifically, the IDH2 desuccinylation and the G6PD deglutarylation by SIRT5 increase cellular antioxidant mechanisms [92]. It has been shown that a SIRT5 deficiency triggers ROS overproduction and mitochondrial imbalance inducing a motor dysfunction in a PD model [93], suggesting a neuroprotective effect.
Perspective And Significance
In general, the role of each SIRT protein form has been analyzed from a disease-centric perspective, leaving out the sexual factor. A hypothetical model has been proposed in which advantageous effects promoted by SIRTs could be presented in a sex-nonspecific manner. However, after reaching reproductive maturity, it has been hypothesized that SIRT dependency in modulating metabolic networks are evolutionarily decoupled from female longevity [94]. In this work, the neurodegenerative process differentially impacts on SIRT profile depending on disease, sex, and olfactory-related areas, affecting the global acetylation/deacetylation machinery of the human brain. Based on our sex-dependent data and because of the complex pattern of temporal and spatial SIRTs expression across the brain [95], the lack of knowledge regarding the substrate specificity of each SIRT isoform in healthy and diseased conditions [96] and the recent growing interest in the deployment of SIRT modulators to ameliorate cognitive deficits and treat neurodegenerative diseases [39, 97, 98], additional research is needed to address the importance of sex differences in animal models and human trials in the evaluation of emerging sirtuin-based therapies against neurological syndromes.