INTRODUCTION

Alzheimer’s disease (AD) is the most common irreversible neurodegenerative disorder in the elderly population. The high variability in AD progression makes it difficult to predict when a patient will convert to dementia. We hypothesize that metabolic alterations in the brain may also be reflected at a systemic level in blood serum and that these alterations could serve as prognostic biomarkers.

METHODS

We investigated serum biomarkers using ¹H-NMR spectroscopy in a population that covers the entire spectrum of cognitive impairment: 57 patients with Alzheimer's disease at the dementia stage (AD-dem), 45 patients with AD at the mild cognitive impairment stage (MCI-AD), and 31 patients with MCI not associated with AD (MCI). 26 metabolites and 112 lipoprotein-related parameters were quantified. The logistic LASSO regression algorithm was employed to identify the optimal combination of metabolite-lipoprotein features and their ratios for group discrimination.

RESULTS

In the training set, our model distinguished AD-dem from MCI with 81.7% accuracy, which was reproduced in the validation set (82.1%). The progression of MCI-AD patients was evaluated over time, with those showing a ≥1.5-point MMSE (Mini–Mental State Examination test) decline per year classified as fast progressors. Our model differentiated fast- and slow-progressing MCI-AD patients with 73.9% accuracy.

CONCLUSIONS

The identification of potential novel peripheral biomarkers of AD paves the way for an innovative and minimally invasive method to identify AD in its early stages. Moreover, our model appears to be able to sub-stratify MCI-AD patients identifying those associated with a faster rate of clinical progression.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by cognitive decline, amyloid-β (Aβ) plaque accumulation and neurofibrillary tangles. Small Aβ oligomeric species exert pronounced neurotoxic and pro-inflammatory effects. Astrocytes, essentials for central nervous system homeostasis, show functional impairment in AD, contributing to disease progression. In this work, we investigated the effects of Aβ_1-42 oligomers on primary human astrocytes from healthy individuals and AD patients. We demonstrated that astrocytes from all individuals can internalize Aβ_1-42 oligomers, particularly females. This internalization enhanced proteasome activity in healthy astrocytes but suppressed it in AD astrocytes. Exposure to Aβ_1-42 oligomers alters calcium homeostasis and impairs mitochondrial membrane potential in astrocytes from all subjects. The treatment caused a reduction in cell viability by inducing apoptosis in astrocytes from healthy subjects; however, the surviving cells adopt a reactive phenotype characterized by increased proliferation and secretion of neuroinflammatory mediators. In contrast, the Aβ_1-42-treatment induced a senescent state in AD astrocytes, especially in those from female patients, as evidenced by the presence of relevant markers: elevated β-galactosidase activity, upregulation of p14^ARF, and the presence of senescence-associated heterochromatin foci. Notably, only conditioned media from Aβ-treated AD astrocytes induces cell death in differentiated neuroblastoma cells, suggesting that astrocyte senescence in AD could contribute to the neuronal damage observed in the disease. Our findings reveal distinct responses of healthy and AD astrocytes to Aβ_1-42-treatment, highlighting the close relationship between cellular senescence and Alzheimer's disease and suggesting its possible role in the pathogenesis and progression of the disease, especially in women.

Alzheimer’s disease (AD), one of the principal neurodegenerative disorders, is characterized by cognitive decline, plaque deposition, and neurofibrillary tangles. Neuroinflammation significantly contributes to the onset and progression of the disease, influenced by functionally impaired astrocytes, which are key regulators of CNS homeostasis. Cellular senescence, a hallmark of aging, contributes to tissue dysfunction and chronic inflammation and is particularly relevant in AD, where senescent astrocytes exacerbate neuronal damage and neuroinflammation. This study employed gemcitabine (GEM), a DNA-damaging chemotherapeutic capable of crossing the blood-brain barrier and used clinically in glioblastoma treatment, to induce senescence in primary human astrocytes derived from healthy individuals and AD patients, aiming to compare their responses.

Following the dose-response curve, 1µM GEM was selected as the optimal dose to induce senescence, preserving cell viability. In both healthy and AD astrocytes, treatment led to proliferation arrest in the G1 phase and increased expression of senescence-associated markers, although AD astrocytes exhibited a greater propensity toward senescence with higher SA-β-Gal activity and expression of p14^ARF, a gene involved in cell cycle arrest and γH2AX, a DNA damage marker. These preliminary results suggest that GEM can effectively induce senescence in human astrocytes, with a more pronounced response in AD cells, indicating a possible intrinsic susceptibility. This in vitro model offers a valuable tool for investigating astrocyte senescence in AD and may support the development of senescence-targeting therapeutic strategies for neurodegenerative diseases.

Background Carotid endarterectomy (CEA) to prevent upcoming cerebrovascular events is necessary in patients with high-grade stenosis (≥ 70%). A blood molecular signature i.e., % of monocytes, albumin, CRP, miR-1271-5p, miR-126-5p CXCL9, GDF8 and, IL-6, has been proposed being able to distinguish between elderly asymptomatic/symptomatic patients as previously published (Capri et al., 2024). The challenge is the validation of the signature in a different cohort of volunteers (≥ 65 years) suffering of comorbidities and with different level of carotid stenosis in a prospective enrolment. The possibility to monitor patients with blood predictive circulating biomarker could have great translational interest and impact on National Health System, if it is possible to avoid or reduce dramatic cerebrovascular injuries.

Methods Forty-three volunteers have been enrolled, including 20 males and 23 females (mean age 80 ± 4). The recruitment phase is still ongoing, since it foresees a short longitudinal study of 50 patients in time series (T1, T2 after ˜6-8 months). Specifically, the patients suffering of different morbidities/co-morbidities (hypertension, type2 diabetes, ischemic cardiopathy, chronic kidney disease, etc.) undergo practitioner’s visit with the carotid stenosis evaluation by means of color doppler ultrasound instrument. Some volunteers with stenosis ≥70% will be evaluated for CEA and listed for hospital admission, in collaboration with the Vascular Surgery Unit, IRCCS-AOU- S.Orsola (Bologna). Haemato-biochemical and urine analyses have been performed. Plasma cytokine measurement such as CXCL9, GDF8 and, IL-6 are ongoing measurements through standard ELISA technique and other molecules such as follistatin, activin, myostatin and fibronectin as well. Plasma miR-1271-5p and miR-126-5p are currently under determination by RT-qPCR. Plaque biopsies from the previous cohort (Capri et al., 2024) will be investigate to deepen the molecular difference between asymptomatic and symptomatic patients starting from miR- profiling.

Results and conclusion T1 and T2 recruitment of the cohort of volunteers is not yet completed. First analysis identifies a positive correlation (r = 0.3401; p = 0.0023) between % of stenosis and plasma albumin concentration in T1 volunteers. T1-T2 patients, enrolled until now, show significant differences for almost all the parameters tested when compared with asymptomatic/symptomatic patients (first cohort of inpatients previously recruited). Plasma albumin, miR-126-5p, follistatin and fibronectin show an increase trend at T2 period. When the recruitment and analyses will be completed a predictive power of carotid stenosis progress through different biomarkers will be applied (training a Random Forest Classifier with 100 decision tree). MiRs emerging from plaque biopsies (asymptomatic vs symptomatic inpatients) with highest fold changes have been bioinformatically investigated and mRNA targets/protein analysis will be investigated in lab wet.

Reference: Capri M, Fronterrè S, Collura S, Giampieri E, Carrino S, Feroldi FM, Ciurca E, Conte M, Olivieri F, Ullo I, Pini R, Vacirca A, Astolfi A, Vasuri F, La Manna G, Pasquinelli G, Gargiulo M. Circulating CXCL9, monocyte percentage, albumin, and C-reactive protein as a potential, non-invasive, molecular signature of carotid artery disease in 65+ patients with multimorbidity: a pilot study in Age.It. Front Endocrinol (Lausanne). 2024 Jul 23;15:1407396. doi: 10.3389/fendo.2024.1407396.

Cellular senescence is a key mechanism underlying aging and frailty, causing chronic inflammation that predisposes to many age-associated diseases. A major contributor to cellular senescence is telomere dysfunction. In post-mitotic cells, telomere dysfunction triggers the permanent activation of a DNA damage response (DDR), which in turns leads to a senescent phenotype. The discovery of telomeric non-coding RNAs responsible for the DDR persistence has led to the development of specific telomeric antisense oligonucleotides (tASO), able to inhibit the telomeric DDR activation.

In this study, we inhibited the telomeric DDR using the tASO strategy on naturally aged C57BL/6 mice. In a first longitudinal experiment, old mice (26 months) were treated with tASO and monitored for several parameters including frailty, cognitive and functional performances, senescent cells accumulation, inflammatory markers, epigenetic age, and lifespan. The first study revealed the absence of acute and long-term toxicity following treatment, and highlighted, only in females, a slight improvement in survival and in the frailty condition. An overall reduction of senescent cells was observed, with a specific effect in the intestine. However, no effects on senescence were observed in other organs, nor on the inflammatory status or on the epigenetic age.

To better understand the tASO efficacy, a second longitudinal experiment (still ongoing) was setup, anticipating the tASO treatment at 18 months of age (at the beginning of aging). The results from this second study will reveal if an earlier intervention could highlight the best potential of the tASO strategy, with important implications for healthspan and longevity.