Leader: Nicoletta Galeotti (UNIFI); Other collaborator(s):
We plan to better understand the mechanisms of microglia cellular senescence through i): in vitro models of senescent microglia cells to characterize aberrant molecular pathways involved in the modulation of senescence mechanisms; ii) animal models of neurodegenerative diseases to characterize microglia senescence during pathological conditions; iii) human microglia extracellular vesicles as biomarker for early diagnosis of disease and drug responsiveness monitoring; iv): exploitation of agents able to target aberrant pathways to unravel the clinical contribution of microglia senescence modulation such as herbal drug-derived phytoconstituents, drug reprofiling and drug repurposing, innovative multitarget agents, antisense oligonucleotide (ASO) strategies.
Brief description of the activities and of the intermediate results
To correlate microglia senescence and development of neurodegenerative disorders, we evaluated the presence of ß-galactosidase positive cells in the spinal cord of EAE mice, an animal model of chronic multiple sclerosis. While positive cells were detected in the whole spinal cord, a prominent expression in the dorsal rather than in the ventral horn was observed. Then we have evaluated CD11b protein expression, as a marker of microgliosis, that resulted drastically overexpressed. A prominent staining of ß-galactosidase in CD11b positive cells was also observed. The clinical relevance of targeting senescent microglia was, thus, investigated. First, we evaluated the pharmacological profile of some senotherapeutic agents in an in vitro model of microglial senescence obtained using BV2 murine microglial cells. Honokiol, its synthetic analogue Honokiol Hexafluoro, and zingiberene showed high efficacy in reducing the main senescence markers. Then, we evaluated whether a combination of two agents would produce a synergistic activity. The pharmacological effect of an association of Zingiber officinale Roscoe and Lavender essential oil was tested in vitro showing a promising effectiveness in reducing microglial senescence parameters. Then the association was tested in vivo in the EAE model. An attenuation of the EAE clinical symptoms, such as motor disability and pain hypersensitivity, was observed along with a modulation of microglial senescence markers at spinal level.
Brief description of the activities and of the intermediate results
Patients with multiple sclerosis (MS) experience, in addition to disability and pain, numerous comorbidities, including memory deficits, anxiety, and depression. Therefore, we assessed the potential role of microglial senescence in the main comorbidities associated with MS. First, we conducted a time-course study to evaluate the onset and progression of mood-related behavioral changes in EAE mice, an animal model of multiple sclerosis, in comparison with the development of motor and pain-related symptoms. The results revealed the onset of anxiety-like and depression-like phenotypes at later stages post-immunization, compared to the development of allodynia and motor disturbances that represent early symptoms.
Brief description of the activities and of the intermediate results
We investigated the presence of senescent cells at supraspinal sites in EAE mice and compared the findings to results from spinal cord samples obtained in previous months. After collecting tissue samples from the thalamus, prefrontal cortex, and hippocampus, we performed western blot analysis to detect the expression of key senescence markers. A significant increase in β-galactosidase, p16, and p21 protein expression was observed in all brain regions investigated of MOG-immunized animals, compared to sham animals, with intensities comparable to those detected at the spinal level. Ongoing experiments are assessing the cellular localization of β-galactosidase-positive cells through double-labeling immunostaining with classical markers of subpopulations in the hippocampal and cortical sections of EAE mice.
Brief description of the activities and of the intermediate results
To better correlate the induction of cellular senescence with the EAE phenotype, we investigated some peripheral parameters to be compared with neurological disturbances. We measured heart rate in EAE mice during model development, taking measurements before immunization and at 3,7,10,14,18,21,24, and 28 days post-immunization. EAE mice showed approximately 20-30 % reduction in heart rate, compared with sham, in about 50% of mice. No differences were recorded in blood pressure values. In collaboration with Prof. Marco Linari, (SPOKE 2, WP6), we analyzed some parameters on slow and fast muscles. For EDL (rapid) muscle: no changes in isometric force and maximum power developed were recorded while maximum shortening speed is reduced in EAE mice. For Soleus Muscle (slow): in EAE mice, isometric force is reduced by 30%, maximum shortening speed is increased by 26% and maximum power is reduced. Finally, circulating cytokine levels were assessed with ELISA kits. An increase in IL-6, IL-17 and TNF-alpha protein levels was observed in EAE mice compared with sham.
Scientific publications
Presentations