EFFECTS OF MICROVESICLES DERIVED FROM BONE MARROW MESENCHYMAL STEM CELLS IN EXPERIMENTAL MODELS OF ALZHEIMER¿S DISEASE

Alzheimer's disease (AD) is the most common form of neurodegenerative illness leading to dementia characterized by the accumulation of abnormally folded β-Amyloid (Aβ) and tau proteins, forming amyloid plaques and neurofibrillary tangles, respectively. Recent evidences have highlighted that inflammation plays a critical role in AD, even though it remains unclear whether it represents a cause or a consequence of the pathology. Deposition of Aβ peptides and tangles are able to stimulate a chronic inflammatory reaction, involving microglial activation and production of inflammatory cytokines likely contributing to neuronal dysfunction and cell death per se. Regarding immunomodulatory strategy development, during the last years, it has been shown that Mesenchymal Stem Cells (MSCs) play a strongly immunomodulatory role, protecting the injured tissue and guiding anti-inflammatory processes by the secretion of cytokines and microvesicles (MVs), involved in their paracrine effects. Furthermore many clinical studies are now performing therapies with MSCs and some phase I and phase II clinical trials in the oncology field are also studying MSC derived MVs (Dai et al, 2008 and Chaput and Théry, 2011). Since the possibility that inflammation is not a mere consequence but a primary contributing factor in AD is becoming concrete, and given upregulation of inflammatory molecules (pro-inflammatory cytokines and chemokines) and activated glial cells surrounding the senile plaques in AD patients brains and AD transgenic animal models are now recognized as typical features of AD, the aim of this project is to assess the anti-inflammatory effects of MVs released by Bone Marrow Mesenchymal Stem Cells (BM-MSCs) in an AD context. Both in vitro and in vivo experimental approaches are used to investigate the MV immunomodulatory effect and their ability to affect Aβ deposition and degradation. Pro- inflammatory (TNFα and IL6) and anti-inflammatory cytokines (IL10) release was investigated by in vitro experiments performed on both microglial N9 cell line and on primary cortical cells exposed to human-Aβ1-42 (h- Aβ1-42 ) and MSC derived MVs. For a more complete analysis of the cell inflammatory state, the microglia phenotype was assessed in order to determine whether a change from M1 to M2 cell phenotype was detectable. The in vivo approach consisted of MV intracranial injections in a well-established transgenic AD chimeric murine model (APPswe/PS1dE9) that recapitulates many of the aspects of the human disease; to investigate whether MVs effects could be effective on the clearance and production of Aβ, possibly ameliorating the neurodegenerative context, we analyzed Aβ load, plaque area and density in three different brain areas: cerebral cortex, hippocampus and cerebellum. The results obtained in vitro indicate that i) the administration of MVs promotes in vitro the secretion of anti-inflammatory cytokines, such as IL10; ii) MVs promote the switch of microglial cell to M2 phenotype, characterized by the typical amoeboid morphology, and increase the expression of CD206, a marker associated to the anti-inflammatory abilities; iii) MV administration significantly inhibits the release of the pro- inflammatory cytokines TNFα and IL 6 in vitro and accordingly, MHC II expression, which is associated with a pro-inflammatory phenotype, is down regulated in the presence of MSC-MVs. On the other hand, in vivo results show that a single MV administration causes a significant reduction of Aβ load into amyloid plaques and a decreased plaque area, into all three brain regions analyzed in 6 month old mice. In 3 month old treated mice, MV administration only affected Aβ plaque density, that resulted smaller respect to untreated mice. In conclusion, MVs released from BM-derived MSCs can exert, in vivo, a protective effect reducing the accumulation in plaques of Aβ, possibly promoting the degradation and elimination of Aβ42 and participating in the regulation of neuroinflammation. Future experiments will be directed to assess the safety of MV administration and to define whether a significant improvement of cognitive impairment is detectable in AD mice treated with BM-derived MSCs .