Open Access
Olfaction and neurodegeneration: olfactory proteotyping across proteinopathies
(2019) Lachén Montes, Mercedes; Santamaría Martínez, Enrique; Fernández Irigoyen, Joaquín; Ciencias de la Salud; Osasun Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa; Gobierno de Navarra / Nafarroako Gobernua
La disfunción olfatoria es un síntoma temprano y común en diversas enfermedades neurodegenerativas (ENs). Según el grado de disfunción olfatoria que presentan, existen ENs con disfunción olfatoria pronunciada, como la enfermedad de Alzheimer (EA) y la enfermedad de Parkinson (EP), y otras con déficits más leves, como ocurre en las demencias frontotemporales. Se ha sugerido que la existencia de un sustrato patológico común actuando mediante diferentes mecanismos en este amplio espectro de ENs podría ser el causante de estas diferencias en el déficit olfatorio. El bulbo olfatorio (BO) es la primera estructura del cerebro responsable de procesar la información olfatoria y el depósito de sustratos neuropatológicos como el péptido amiloide o la forma hiperfosforilada de la proteína tau en esta región se ha propuesto también como posible origen de este síntoma. Sin embargo, se desconoce si los agregados neuropatológicos son causa o consecuencia del proceso neurodegenerativo que ocurre en esta región. En esta tesis se ha realizado un análisis molecular de alto rendimiento en BOs procedentes de: a) dos modelos animales de la EA; y b) sujetos diagnosticados con cuatro ENs incluyendo la EA, la EP, la degeneración lobar fronto-temporal con depósitos de TAR DNA-binding protein (FTLD-TDP43) y la parálisis supranuclear progresiva (PSP), con el objetivo de caracterizar los mecanismos neuropatofisiológicos que ocurren en esta región durante el proceso neurodegenerativo. Este amplio análisis ha demostrado que existe una gran alteración en la proteostasis del BO durante la EA y la EP, donde los resultados mostraron un 20% del proteoma cuantificado diferencialmente expresado. Por otra parte, en el caso de los sujetos diagnosticados con FTLD-TDP43 y PSP, el número de alteraciones fue mucho menor, constituyendo alrededor del 1% del proteoma cuantificado. Es interesante resaltar que se han encontrado tanto similitudes como diferencias en los mediadores proteicos diferencialmente expresados entre las ENs analizadas y la población control. Por otra parte, el estudio en los dos modelos animales de EA ha demostrado que, a nivel de BO, existen alteraciones moleculares previas a la aparición de placas amiloides y deficits cognitivos. Finalmente, se ha demostrado la utilidad de la proteómica dirigida a estructuras olfatorias como fuente de biomarcadores en ENs. De hecho, se propone la proteína Glucosamine-6-phosphate isomerase 2 (GNPDA2) como potencial biomarcador de la EP.
Open Access
Neuropathological stage-dependent proteome mapping of the olfactory tract in Alzheimer's disease: from early olfactory-related omics signatures to computational repurposing of drug candidates
(Wiley, 2024) Cartas Cejudo, Paz; Cortés, Adriana; Lachén Montes, Mercedes; Anaya-Cubero, Elena; Puerta, Elena; Solas, Maite; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Ciencias de la Salud; Osasun Zientziak; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
Alzheimer's disease (AD) is the most common form of dementia, characterized by an early olfactory dysfunction, progressive memory loss, and behavioral deterioration. Albeit substantial progress has been made in characterizing AD-associated molecular and cellular events, there is an unmet clinical need for new therapies. In this study, olfactory tract proteotyping performed in controls and AD subjects (n = 17/group) showed a Braak stage-dependent proteostatic impairment accompanied by the progressive modulation of amyloid precursor protein and tau functional interactomes. To implement a computational repurposing of drug candidates with the capacity to reverse early AD-related olfactory omics signatures (OMSs), we generated a consensual OMSs database compiling differential omics datasets obtained by mass-spectrometry or RNA-sequencing derived from initial AD across the olfactory axis. Using the Connectivity Map-based drug repurposing approach, PKC, EGFR, Aurora kinase, Glycogen synthase kinase, and CDK inhibitors were the top pharmacologic classes capable to restore multiple OMSs, whereas compounds with targeted activity to inhibit PI3K, Insulin-like growth factor 1 (IGF-1), microtubules, and Polo-like kinase (PLK) represented a family of drugs with detrimental potential to induce olfactory AD-associated gene expression changes. To validate the potential therapeutic effects of the proposed drugs, in vitro assays were performed. These validation experiments revealed that pretreatment of human neuron-like SH-SY5Y cells with the EGFR inhibitor AG-1478 showed a neuroprotective effect against hydrogen peroxide-induced damage while the pretreatment with the Aurora kinase inhibitor Reversine reduced amyloid-beta (Aβ)-induced neurotoxicity. Taken together, our data pointed out that OMSs may be useful as substrates for drug repurposing to propose novel neuroprotective treatments against AD.
Open Access
Network-driven proteogenomics unveils an aging-related imbalance in the olfactory IκBα-NFκB p65 complex functionality in Tg2576 Alzheimer’s disease mouse model
(MDPI, 2017) Palomino Alonso, Maialen; Lachén Montes, Mercedes; González Morales, Andrea; Ausín, Karina; Pérez Mediavilla, Alberto; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua, PC023-24; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
Olfaction is often deregulated in Alzheimer’s disease (AD) patients, and is also impaired in transgenic Tg2576 AD mice, which overexpress the Swedish mutated form of human amyloid precursor protein (APP). However, little is known about the molecular mechanisms that accompany the neurodegeneration of olfactory structures in aged Tg2576 mice. For that, we have applied proteome- and transcriptome-wide approaches to probe molecular disturbances in the olfactory bulb (OB) dissected from aged Tg2576 mice (18 months of age) as compared to those of age matched wild-type (WT) littermates. Some over-represented biological functions were directly relevant to neuronal homeostasis and processes of learning, cognition, and behavior. In addition to the modulation of CAMP responsive element binding protein 1 (CREB1) and APP interactomes, an imbalance in the functionality of the IκBα-NFκB p65 complex was observed during the aging process in the OB of Tg2576 mice. At two months of age, the phosphorylated isoforms of olfactory IκBα and NFκB p65 were inversely regulated in transgenic mice. However, both phosphorylated proteins were increased at 6 months of age, while a specific drop in IκBα levels was detected in 18-month-old Tg2576 mice, suggesting a transient activation of NFκB in the OB of Tg2576 mice. Taken together, our data provide a metabolic map of olfactory alterations in aged Tg2576 mice, reflecting the progressive effect of APP overproduction and β-amyloid (Aβ) accumulation on the OB homeostasis in aged stages.
Open Access
Progressive modulation of the human olfactory bulb transcriptome during Alzheimer´s disease evolution: novel insights into the olfactory signaling across proteinopathies
(Impact Journals, 2017) Lachén Montes, Mercedes; Zelaya Huerta, María Victoria; Segura, Víctor; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua, PC025; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
Alzheimer´s disease (AD) is characterized by progressive dementia, initially presenting olfactory dysfunction. Despite the olfactory bulb (OB) is the first central structure of the olfactory pathway, we lack a complete molecular characterization of the transcriptional events that occurs in this olfactory area during AD progression. To address this gap in knowledge, we have assessed the genome-wide expression in postmortem OBs from subjects with varying degree of AD pathology. A stagedependent deregulation of specific pathways was observed, revealing transmembrane transport, and neuroinflammation as part of the functional modules that are disrupted across AD grading. Potential drivers of neurodegeneration predicted by networkdriven transcriptomics were monitored across different types of dementia, including progressive supranuclear palsy (PSP), mixed dementia, and frontotemporal lobar degeneration (FTLD). Epidermal growth factor receptor (EGFR) expression was significantly increased in the OB of AD and mixed dementia subjects. Moreover, a significant increment in the activation of signal transducer and activator of transcription 3 (STAT3) was exclusively detected in advanced AD stages, whereas total STAT3 levels were specifically overexpressed in mixed dementia. Furthermore, transcription factors deregulated in the OB of mixed dementia subjects such as cAMP Responsive Element Binding Protein 1 (CREB1) and AP-1 Transcription Factor Subunit (c-Jun) were not differentially modulated at olfactory level across AD grading. On the other hand, olfactory expression of this signal transducer panel was unchanged in PSP and FTLD subjects. Taken together, this study unveils cross-disease similarities and differences for specific signal transducers, providing mechanistic clues to the intriguing divergence of AD pathology across proteinopathies.
Open Access
Olfactory characterization and training in older adults: protocol study
(Frontiers Media, 2021) Zambom Ferraresi, Fabíola; Zambom Ferraresi, Fabrício; Fernández Irigoyen, Joaquín; Lachén Montes, Mercedes; Cartas Cejudo, Paz; Lasarte, Juan José; Casares, Noelia; Fernández, Secundino; Cedeño Veloz, Bernardo Abel; Maraví-Aznar, Enrique; Uzcanga-Lacabe, María Itziar; Galbete Jiménez, Arkaitz; Santamaría Martínez, Enrique; Martínez Velilla, Nicolás; Estadística, Informática y Matemáticas; Estatistika, Informatika eta Matematika; Gobierno de Navarra / Nafarroako Gobernua
The aim of this article is to present the research protocol for a prospective cohort study that will assess the olfactory function and the effect of an intervention based on olfactory training in healthy very old adults (≥75 years old). A convenience sample of 180 older people (50% female) will be recruited in three different environments: hospitalized control group (CH) with stable acute illness (n = 60); ambulatory control group (CA) of community-based living (n = 60); and an experimental odor training group (EOT) from nursing homes (n = 60). The odor training (OT) intervention will last 12 weeks. All the volunteers will be assessed at baseline; CA and EOT groups will also be assessed after 12 weeks. The primary end point will be change in olfactory capacity from baseline to 12 weeks period of intervention or control. The intervention effects will be assessed with the overall score achieved in Sniffin Sticks Test (SST) – Threshold, Discrimination, and Identification (TDI) extended version. Secondary end points will be changes in cognitive tasks, quality of life, mood, immune status, and functional capacity. All these measurements will be complemented with an immune fitness characterization and a deep proteome profiling of the olfactory epithelium (OE) cultured ex vivo. The current study will provide additional evidence to support the implementation of olfactory precision medicine and the development of immunomodulatory nasal therapies based on non-invasive procedures. The proposed intervention will also intend to increase the knowledge about the olfactory function in very elderly people, improve function and quality of life, and promote the recovery of the health.
Open Access
Tackling the biological meaning of the human olfactory bulb dyshomeostatic proteome across neurological disorders: an integrative bioinformatic approach
(MDPI, 2021) Cartas Cejudo, Paz; Lachén Montes, Mercedes; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua
Olfactory dysfunction is considered an early prodromal marker of many neurodegenerative diseases. Neuropathological changes and aberrant protein aggregates occur in the olfactory bulb (OB), triggering a tangled cascade of molecular events that is not completely understood across neurological disorders. This study aims to analyze commonalities and differences in the olfactory protein homeostasis across neurological backgrounds with different spectrums of smell dysfunction. For that, an integrative analysis was performed using OB proteomics datasets derived from subjects with Alzheimer’s disease (AD), Parkinson´s disease (PD), mixed dementia (mixD), dementia with Lewy bodies (DLB), frontotemporal lobar degeneration (FTLD-TDP43), progressive supranuclear palsy (PSP) and amyotrophic lateral sclerosis (ALS) with respect to OB proteome data from neurologically intact controls. A total of 80% of the differential expressed protein products were potentially disease-specific whereas the remaining 20% were commonly altered across two, three or four neurological phenotypes. A multi-level bioinformatic characterization revealed a subset of potential disease-specific transcription factors responsible for the downstream effects detected at the proteome level as well as specific densely connected protein complexes targeted by several neurological phenotypes. Interestingly, common or unique pathways and biofunctions were also identified, providing novel mechanistic clues about each neurological disease at olfactory level. The analysis of olfactory epithelium, olfactory tract and primary olfactory cortical proteotypes in a multi-disease format will functionally complement the OB dyshomeostasis, increasing our knowledge about the neurodegenerative process across the olfactory axis.
Open Access
Involvement of glucosamine 6 phosphate isomerase 2 (GNPDA2) overproduction in beta-amyloid- and Tau P301L-driven pathomechanisms
(MDPI, 2024) Lachén Montes, Mercedes; Cartas Cejudo, Paz; Cortés, Adriana; Anaya-Cubero, Elena; Peral, Erika; Ausín, Karina; Díaz-Peña, Ramón; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Ciencias de la Salud; Osasun Zientziak
Alzheimer’s disease (AD) is a neurodegenerative olfactory disorder affecting millions of people worldwide. Alterations in the hexosamine- or glucose-related pathways have been described through AD progression. Specifically, an alteration in glucosamine 6 phosphate isomerase 2 (GNPDA2) protein levels has been observed in olfactory areas of AD subjects. However, the biological role of GNPDA2 in neurodegeneration remains unknown. Using mass spectrometry, multiple GNPDA2 interactors were identified in human nasal epithelial cells (NECs) mainly involved in intraciliary transport. Moreover, GNPDA2 overexpression induced an increment in NEC proliferation rates, accompanied by transcriptomic alterations in Type II interferon signaling or cellular stress responses. In contrast, the presence of beta-amyloid or mutated Tau-P301L in GNPDA2-overexpressing NECs induced a slowdown in the proliferative capacity in parallel with a disruption in protein processing. The proteomic characterization of Tau-P301L transgenic zebrafish embryos demonstrated that GNPDA2 overexpression interfered with collagen biosynthesis and RNA/protein processing, without inducing additional changes in axonal outgrowth defects or neuronal cell death. In humans, a significant increase in serum GNPDA2 levels was observed across multiple neurological proteinopathies (AD, Lewy body dementia, progressive supranuclear palsy, mixed dementia and amyotrophic lateral sclerosis) (n = 215). These data shed new light on GNPDA2-dependent mechanisms associated with the neurodegenerative process beyond the hexosamine route.
Open Access
Amyotrophic lateral sclerosis is accompanied by protein derangements in the olfactory bulb-tract axis
(MDPI, 2020) Lachén Montes, Mercedes; Mendizuri, Naroa; Ausín, Karina; Andrés Benito, Pol; Ferrer, Isidro; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua, Ref. 0011-1411-2020-000028
Amyotrophic lateral sclerosis (ALS) is a fatal disease characterized by progressive muscle paralysis due to the degeneration of upper and lower motor neurons. Recent studies point out an involvement of the non-motor axis during disease progression. Despite smell impairment being considered a potential non-motor finding in ALS, the pathobiochemistry at the olfactory level remains unknown. Here, we applied an olfactory quantitative proteotyping approach to analyze the magnitude of the olfactory bulb (OB) proteostatic imbalance in ALS subjects (n = 12) with respect to controls (n = 8). Around 3% of the quantified OB proteome was differentially expressed, pinpointing aberrant protein expression involved in vesicle-mediated transport, macroautophagy, axon development and gliogenesis in ALS subjects. The overproduction of olfactory marker protein (OMP) points out an imbalance in the olfactory signal transduction in ALS. Accompanying the specific overexpression of glial fibrillary acidic protein (GFAP) and Bcl-xL in the olfactory tract (OT), a tangled disruption of signaling routes was evidenced across the OB–OT axis in ALS. In particular, the OB survival signaling dynamics clearly differ between ALS and frontotemporal lobar degeneration (FTLD), two faces of TDP-43 proteinopathy. To the best of our knowledge, this is the first report on high-throughput molecular characterization of the olfactory proteostasis in ALS.
Open Access
Early-onset molecular derangements in the olfactory bulb of Tg2576 mice: novel insights into the stress-responsive olfactory kinase dynamics in Alzheimer’s disease
(Frontiers Media, 2019) Lachén Montes, Mercedes; González Morales, Andrea; Palomino Alonso, Maialen; Ausín, Karina; Gómez-Ochoa, Marta; Zelaya Huerta, María Victoria; Ferrer, Isidro; Pérez Mediavilla, Alberto; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
The olfactory bulb (OB) is the first processing station in the olfactory pathway. Despite smell impairment, which is considered an early event in Alzheimer’s disease (AD), little is known about the initial molecular disturbances that accompany the AD development at olfactory level. We have interrogated the time-dependent OB molecular landscape in Tg2576 AD mice prior to the appearance of neuropathological amyloid plaques (2-, and 6-month-old), using combinatorial omics analysis. The metabolic modulation induced by overproduction of human mutated amyloid precursor protein (APP) clearly differs between both time points. Besides the progressive perturbation of the APP interactome, functional network analysis unveiled an inverse regulation of downstream extracellular signal-regulated kinase (ERK1/2), and p38 mitogen-activated protein kinase (MAPK) routes in 2-month-old Tg2576 mice with respect to wild-type (WT) mice. In contrast, Akt and MAPK kinase 4 (SEK1)/ stress-activated protein kinase (SAPK) axis were parallel activated in the OB of 6-months-old-Tg2576 mice. Furthermore, a survival kinome profiling performed during the aging process (2-, 6-, and 18-month-old) revealed that olfactory APP overexpression leads to changes in the activation dynamics of protein kinase A (PKA), and SEK1/MKK4-SAPK/JNK between 6 and 18 months of age, when memory deficits appear and AD pathology is well established in transgenic mice. Interestingly, both olfactory pathways were differentially activated in a stage-dependent manner in human sporadic AD subjects with different neuropathological grading. Taken together, our data reflect the early impact of mutated APP on the OB molecular homeostasis, highlighting the progressive modulation of specific signaling pathways during the olfactory amyloidogenic pathology.
Open Access
Olfactory bulb proteomics reveals widespread proteostatic disturbances in mixed dementia and guides for potential serum biomarkers to discriminate alzheimer disease and mixed dementia phenotypes
(MDPI, 2021) Lachén Montes, Mercedes; Íñigo-Marco, Ignacio; Cartas Cejudo, Paz; Fernández Irigoyen, Joaquín; Santamaría Martínez, Enrique; Ciencias de la Salud; Osasun Zientziak; Gobierno de Navarra / Nafarroako Gobernua
The most common form of mixed dementia (MixD) is constituted by abnormal protein deposits associated with Alzheimer's disease (AD) that coexist with vascular disease. Although olfactory dysfunction is considered a clinical sign of AD-related dementias, little is known about the impact of this sensorial impairment in MixD at the molecular level. To address this gap in knowledge, we assessed olfactory bulb (OB) proteome-wide expression in MixD subjects (n = 6) respect to neurologically intact controls (n = 7). Around 9% of the quantified proteins were differentially expressed, pinpointing aberrant proteostasis involved in synaptic transmission, nucleoside monophosphate and carbohydrate metabolism, and neuron projection regeneration. In addition, network-driven proteomics revealed a modulation in cell-survival related pathways such as ERK, AKT, and the PDK1-PKC axis. Part of the differential OB protein set was not specific of MixD, also being deregulated across different tauopathies, synucleinopathies, and tardopathies. However, the comparative functional analysis of OB proteome data between MixD and pure AD pathologies deciphered commonalities and differences between both related phenotypes. Finally, olfactory pro-teomics allowed to propose serum Prolow-density lipoprotein receptor-related protein 1 (LRP1) as a candidate marker to differentiate AD from MixD phenotypes.