Olfactory sensitivity to odorant molecules is a complex biological function influenced by both endogenous factors, such as genetic background and physiological state, and exogenous factors, such as environmental conditions. In animals, this vital ability is mediated by olfactory sensory neurons (OSNs), which are distributed across several specialized olfactory subsystems depending on the species. Using the phosphorylation of the ribosomal protein S6 (rpS6) in OSNs following sensory stimulation, we developed an ex vivo assay allowing the simultaneous conditioning and odorant stimulation of different mouse olfactory subsystems, including the main olfactory epithelium, the vomeronasal organ, and the Grueneberg ganglion. This approach enabled us to observe odorant-induced neuronal activity within the different olfactory subsystems and to demonstrate the impact of environmental conditioning, such as temperature variations, on olfactory sensitivity, specifically in the Grueneberg ganglion. We further applied our rpS6-based assay to the human olfactory system and demonstrated its feasibility. Our findings show that analyzing rpS6 signal intensity is a robust and highly reproducible indicator of neuronal activity across various olfactory systems, while avoiding stress and some experimental limitations associated with in vivo exposure. The potential extension of this assay to other conditioning paradigms and olfactory systems, as well as its application to other animal species, including human olfactory diagnostics, is also discussed.
- Klíčová slova
- 3Rs, Grueneberg ganglion, environmental factors, human, mouse, neuronal activity, olfaction, olfactory subsystems, rpS6,
- MeSH
- čich fyziologie MeSH
- čichová sliznice metabolismus MeSH
- čichové buňky * metabolismus fyziologie MeSH
- fosforylace MeSH
- lidé MeSH
- myši inbrední C57BL MeSH
- myši MeSH
- odoranty analýza MeSH
- ribozomální protein S6 * metabolismus MeSH
- vomeronazální orgán metabolismus fyziologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- ribozomální protein S6 * MeSH
INTRODUCTION: Studies have correlated living close to major roads with Alzheimer's disease (AD) risk. However, the mechanisms responsible for this link remain unclear. METHODS: We exposed olfactory mucosa (OM) cells of healthy individuals and AD patients to diesel emissions (DE). Cytotoxicity of exposure was assessed, mRNA, miRNA expression, and DNA methylation analyses were performed. The discovered altered pathways were validated using data from the human population-based Rotterdam Study. RESULTS: DE exposure resulted in an almost four-fold higher response in AD OM cells, indicating increased susceptibility to DE effects. Methylation analysis detected different DNA methylation patterns, revealing new exposure targets. Findings were validated by analyzing data from the Rotterdam Study cohort and demonstrated a key role of nuclear factor erythroid 2-related factor 2 signaling in responses to air pollutants. DISCUSSION: This study identifies air pollution exposure biomarkers and pinpoints key pathways activated by exposure. The data suggest that AD individuals may face heightened risks due to impaired cellular defenses. HIGHLIGHTS: Healthy and AD olfactory cells respond differently to DE exposure. AD cells are highly susceptible to DE exposure. The NRF2 oxidative stress response is highly activated upon air pollution exposure. DE-exposed AD cells activate the unfolded protein response pathway. Key findings are also confirmed in a population-based study.
- Klíčová slova
- Alzheimer's disease (AD), air pollution, air–liquid interface (ALI), heat shock protein (HSP), next‐generation sequencing (NGS), nuclear factor erythroid 2–related factor 2 (NRF2), traffic emissions, traffic‐related air pollution (TRAP) olfactory mucosa (OM),
- MeSH
- Alzheimerova nemoc * genetika metabolismus MeSH
- čichová sliznice metabolismus MeSH
- epigenomika MeSH
- faktor 2 související s NF-E2 genetika metabolismus MeSH
- látky znečišťující vzduch škodlivé účinky MeSH
- lidé středního věku MeSH
- lidé MeSH
- metylace DNA * MeSH
- mikro RNA metabolismus genetika MeSH
- senioři MeSH
- stanovení celkové genové exprese MeSH
- transkriptom MeSH
- výfukové emise vozidel * toxicita MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- faktor 2 související s NF-E2 MeSH
- látky znečišťující vzduch MeSH
- mikro RNA MeSH
- NFE2L2 protein, human MeSH Prohlížeč
- výfukové emise vozidel * MeSH
Constituents of air pollution, the ultrafine particles (UFP) with a diameter of ≤0.1 μm, are considerably related to traffic emissions. Several studies link air pollution to Alzheimer's disease (AD), yet the exact relationship between the two remains poorly understood. Mitochondria are known targets of environmental toxicants, and their dysfunction is associated with neurodegenerative diseases. The olfactory mucosa (OM), located at the rooftop of the nasal cavity, is directly exposed to the environment and in contact with the brain. Mounting evidence suggests that the UFPs can impact the brain directly through the olfactory tract. By using primary human OM cultures established from nasal biopsies of cognitively healthy controls and individuals diagnosed with AD, we aimed to decipher the effects of traffic-related UFPs on mitochondria. The UFP samples were collected from the exhausts of a modern heavy-duty diesel engine (HDE) without aftertreatment systems, run with renewable diesel (A0) and petroleum diesel (A20), and from an engine of a 2019 model diesel passenger car (DI-E6d) equipped with state-of-the-art aftertreatment devices and run with renewable diesel (Euro6). OM cells were exposed to three different UFPs for 24-h and 72-h, after which cellular processes were assessed on the functional and transcriptomic levels. Our results show that UFPs impair mitochondrial functions in primary human OM cells by hampering oxidative phosphorylation (OXPHOS) and redox balance, and the responses of AD cells differ from cognitively healthy controls. RNA-Seq and IPA® revealed inhibition of OXPHOS and mitochondrial dysfunction in response to UFPs A0 and A20. Functional validation confirmed that A0 and A20 impair cellular respiration, decrease ATP levels, and disturb redox balance by altering NAD and glutathione metabolism, leading to increased ROS and oxidative stress. RNA-Seq and functional assessment revealed the presence of AD-related alterations in human OM cells and that different fuels and engine technologies elicit differential effects.
- Klíčová slova
- Mitochondrial dysfunction, Olfactory mucosa (OM), Oxidative phosphorylation (OXPHOS), RNA sequencing (RNA-Seq), Redox balance, Ultrafine particles (UFP),
- MeSH
- Alzheimerova nemoc * metabolismus etiologie patologie chemicky indukované MeSH
- čichová sliznice * metabolismus patologie účinky léků MeSH
- látky znečišťující vzduch toxicita škodlivé účinky MeSH
- lidé MeSH
- mitochondrie * metabolismus účinky léků MeSH
- oxidační stres účinky léků MeSH
- pevné částice * škodlivé účinky toxicita MeSH
- reaktivní formy kyslíku metabolismus MeSH
- senioři MeSH
- výfukové emise vozidel toxicita MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- látky znečišťující vzduch MeSH
- pevné částice * MeSH
- reaktivní formy kyslíku MeSH
- výfukové emise vozidel MeSH
BACKGROUND: The neurological effects of the coronavirus disease of 2019 (COVID-19) raise concerns about potential long-term consequences, such as an increased risk of Alzheimer's disease (AD). Neuroinflammation and other AD-associated pathologies are also suggested to increase the risk of serious SARS-CoV-2 infection. Anosmia is a common neurological symptom reported in COVID-19 and in early AD. The olfactory mucosa (OM) is important for the perception of smell and a proposed site of viral entry to the brain. However, little is known about SARS-CoV-2 infection at the OM of individuals with AD. METHODS: To address this gap, we established a 3D in vitro model of the OM from primary cells derived from cognitively healthy and AD individuals. We cultured the cells at the air-liquid interface (ALI) to study SARS-CoV-2 infection under controlled experimental conditions. Primary OM cells in ALI expressed angiotensin-converting enzyme 2 (ACE-2), neuropilin-1 (NRP-1), and several other known SARS-CoV-2 receptor and were highly vulnerable to infection. Infection was determined by secreted viral RNA content and confirmed with SARS-CoV-2 nucleocapsid protein (NP) in the infected cells by immunocytochemistry. Differential responses of healthy and AD individuals-derived OM cells to SARS-CoV-2 were determined by RNA sequencing. RESULTS: Results indicate that cells derived from cognitively healthy donors and individuals with AD do not differ in susceptibility to infection with the wild-type SARS-CoV-2 virus. However, transcriptomic signatures in cells from individuals with AD are highly distinct. Specifically, the cells from AD patients that were infected with the virus showed increased levels of oxidative stress, desensitized inflammation and immune responses, and alterations to genes associated with olfaction. These results imply that individuals with AD may be at a greater risk of experiencing severe outcomes from the infection, potentially driven by pre-existing neuroinflammation. CONCLUSIONS: The study sheds light on the interplay between AD pathology and SARS-CoV-2 infection. Altered transcriptomic signatures in AD cells may contribute to unique symptoms and a more severe disease course, with a notable involvement of neuroinflammation. Furthermore, the research emphasizes the need for targeted interventions to enhance outcomes for AD patients with viral infection. The study is crucial to better comprehend the relationship between AD, COVID-19, and anosmia. It highlights the importance of ongoing research to develop more effective treatments for those at high risk of severe SARS-CoV-2 infection.
- Klíčová slova
- Air–liquid interface, Alzheimer’s disease, Anosmia, COVID-19, Immune responses, Inflammation, Neurological manifestations, Olfactory, SARS-CoV-2,
- MeSH
- Alzheimerova nemoc * metabolismus MeSH
- anosmie metabolismus MeSH
- čichová sliznice metabolismus MeSH
- COVID-19 * MeSH
- lidé MeSH
- neurozánětlivé nemoci MeSH
- SARS-CoV-2 MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Olfactory function, orchestrated by the cells of the olfactory mucosa at the rooftop of the nasal cavity, is disturbed early in the pathogenesis of Alzheimer's disease (AD). Biometals including zinc and calcium are known to be important for sense of smell and to be altered in the brains of AD patients. Little is known about elemental homeostasis in the AD patient olfactory mucosa. Here we aimed to assess whether the disease-related alterations to biometal homeostasis observed in the brain are also reflected in the olfactory mucosa. We applied RNA sequencing to discover gene expression changes related to metals in olfactory mucosal cells of cognitively healthy controls, individuals with mild cognitive impairment and AD patients, and performed analysis of the elemental content to determine metal levels. Results demonstrate that the levels of zinc, calcium and sodium are increased in the AD olfactory mucosa concomitantly with alterations to 17 genes related to metal-ion binding or metal-related function of the protein product. A significant elevation in alpha-2-macroglobulin, a known metal-binding biomarker correlated with brain disease burden, was observed on the gene and protein levels in the olfactory mucosa cells of AD patients. These data demonstrate that the olfactory mucosa cells derived from AD patients recapitulate certain impairments of biometal homeostasis observed in the brains of patients.
- Klíčová slova
- Alzheimer’s disease, alpha-2-macroglobulin, biometals, calcium, olfactory dysfunction, olfactory mucosa cells, sodium, zinc,
- MeSH
- Alzheimerova nemoc * metabolismus MeSH
- chelátory metabolismus MeSH
- čichová sliznice metabolismus MeSH
- lidé MeSH
- stopové prvky * metabolismus MeSH
- vápník metabolismus MeSH
- zinek metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- chelátory MeSH
- stopové prvky * MeSH
- vápník MeSH
- zinek MeSH
Facial shape is the basis for facial recognition and categorization. Facial features reflect the underlying geometry of the skeletal structures. Here, we reveal that cartilaginous nasal capsule (corresponding to upper jaw and face) is shaped by signals generated by neural structures: brain and olfactory epithelium. Brain-derived Sonic Hedgehog (SHH) enables the induction of nasal septum and posterior nasal capsule, whereas the formation of a capsule roof is controlled by signals from the olfactory epithelium. Unexpectedly, the cartilage of the nasal capsule turned out to be important for shaping membranous facial bones during development. This suggests that conserved neurosensory structures could benefit from protection and have evolved signals inducing cranial cartilages encasing them. Experiments with mutant mice revealed that the genomic regulatory regions controlling production of SHH in the nervous system contribute to facial cartilage morphogenesis, which might be a mechanism responsible for the adaptive evolution of animal faces and snouts.
- Klíčová slova
- cartilage induction, cleft palate, developmental biology, embryonic development, facial shaping, mammalian face, mouse, regenerative medicine, sonic hedgehog, stem cells,
- MeSH
- chondrocyty cytologie účinky léků metabolismus MeSH
- čichová sliznice cytologie účinky léků růst a vývoj metabolismus MeSH
- embryo savčí MeSH
- homeoboxový protein Nkx-2.2 MeSH
- homeodoménové proteiny genetika metabolismus MeSH
- integrasy genetika metabolismus MeSH
- kolagen typ II genetika metabolismus MeSH
- lidé MeSH
- maxilofaciální vývoj genetika MeSH
- morfogeneze účinky léků genetika MeSH
- mozek účinky léků růst a vývoj metabolismus MeSH
- mutageny aplikace a dávkování MeSH
- myši transgenní MeSH
- myši MeSH
- nosní chrupavky cytologie účinky léků růst a vývoj metabolismus MeSH
- obličej anatomie a histologie embryologie MeSH
- obličejové kosti cytologie účinky léků růst a vývoj metabolismus MeSH
- proteiny dánia pruhovaného MeSH
- proteiny hedgehog genetika metabolismus MeSH
- rekombinantní fúzní proteiny genetika metabolismus MeSH
- signální transdukce * MeSH
- tamoxifen aplikace a dávkování MeSH
- transkripční faktory genetika metabolismus MeSH
- vývojová regulace genové exprese MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- Col2a1 protein, mouse MeSH Prohlížeč
- Cre recombinase MeSH Prohlížeč
- homeoboxový protein Nkx-2.2 MeSH
- homeodoménové proteiny MeSH
- integrasy MeSH
- kolagen typ II MeSH
- mutageny MeSH
- proteiny dánia pruhovaného MeSH
- proteiny hedgehog MeSH
- rekombinantní fúzní proteiny MeSH
- SHH protein, human MeSH Prohlížeč
- tamoxifen MeSH
- transkripční faktory MeSH
Drug access to the CNS is hindered by the presence of the blood-brain barrier (BBB), and the intranasal route has risen as a non-invasive route to transport drugs directly from nose-to-brain avoiding the BBB. In addition, nanoparticles (NPs) have been described as efficient shuttles for direct nose-to-brain delivery of drugs. Nevertheless, there are few studies describing NP nose-to-brain transport. Thus, the aim of this work was (i) to develop, characterize and validate in vitro olfactory cell monolayers and (ii) to study the transport of polymeric- and lipid-based NPs across these monolayers in order to estimate NP access into the brain using cell penetrating peptide (CPPs) moieties: Tat and Penetratin (Pen). All tested poly(d,l-lactide-co-glycolide) (PLGA) and nanostructured lipid carrier (NLC) formulations were stable in transport buffer and biocompatible with the olfactory mucosa cells. Nevertheless, 0.7% of PLGA NPs was able to cross the olfactory cell monolayers, whereas 8% and 22% of NLC and chitosan-coated NLC (CS-NLC) were transported across them, respectively. Moreover, the incorporation of CPPs to NLC surface significantly increased their transport, reaching 46% of transported NPs. We conclude that CPP-CS-NLC represent a promising brain shuttle via nose-to-brain for drug delivery.
- Klíčová slova
- CPP, Lipid nanoparticles, NLC, Nanoparticles, Nose-to-brain delivery, Olfactory mucosa,
- MeSH
- aplikace intranazální MeSH
- biologický transport MeSH
- chemie farmaceutická metody MeSH
- chitosan chemie MeSH
- čichová sliznice cytologie metabolismus MeSH
- hematoencefalická bariéra metabolismus MeSH
- kopolymer kyseliny glykolové a mléčné MeSH
- krysa rodu Rattus MeSH
- kyselina mléčná chemie MeSH
- kyselina polyglykolová chemie MeSH
- lipidy chemie MeSH
- mozek metabolismus MeSH
- nanočástice * MeSH
- nosní sliznice metabolismus MeSH
- penetrační peptidy chemie MeSH
- polymery chemie MeSH
- potkani Wistar MeSH
- systémy cílené aplikace léků * MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- validační studie MeSH
- Názvy látek
- chitosan MeSH
- kopolymer kyseliny glykolové a mléčné MeSH
- kyselina mléčná MeSH
- kyselina polyglykolová MeSH
- lipidy MeSH
- penetrační peptidy MeSH
- polymery MeSH
