Background Aging is a time-related deterioration of physiological functions, associated with chronic oxidative stress and inflammation. The brain, with proliferation limit of its cells, is particularly affected by it. Brain aging presents cognitive alterations and involves microglia (MG), responsible for the chronic neuroinflammation, that causes morpho-functional alteration of astrocytes (AS). AS lose the ability to maintain the physiological levels of glutamate (Glu). This leads to neuronal death. All these alterations cause memory loss and lack of motor coordination that occur with aging. MG and AS reactivities are linked to gender explaining the neuroprotective effects of sex steroids after brain injury. MG responds to estrogen by reducing its inflammatory response; AS distribution, reactivity, and interaction with other cells of the brain depend on gender. Extracellular vesicles (EVs) represent a mode of information exchange between donor and recipient cells.
Hypothesis The project proposes the exogenous administration of EVs deriving from "young" microglia or astrocytes to old mice to slow down neuroinflammation in aging. Objective To evaluate the effects of the administration of EVs deriving from ¿young¿ or ¿old¿ glial cells to mice of different sexes, to measure their efficacy in the aging processes linked to intercellular cross-talk. Experimental project EVs released from primary mouse MG and AS will be used to be administered to male and female mice. Aging will be evaluated in vivo as motor skills and memory, and ex vivo as level of neuroinflammation, neuronal death and synaptic plasticity. Expected results i) reduction of neuroinflammation slowdowns memory impairment and motor reflexes in old mice treated with EVs derived from ¿young¿ cell cultures; ii) acceleration of aging process in "young" mice treated with EVs derived from "old" cell cultures; iii) anti-aging effects of EVs from ¿young¿ cells in female mice more exhaustive compared to male mice.
The interest of the study is aimed at researching innovative strategies for slowing down aging. Many therapeutic approaches have been studied, some promising, others not so promising. Caloric restriction has the potential to delay aging, although the low-calorie diet induces many hormonal alterations (Kim et al 2015). In mice, administration of platelet-rich plasma reduces the senescence process of stem cells (Liu et al., 2014), but it is unclear whether stem cells affect longevity (Oh et al 2014). Exercise attenuates the effects of deterioration of cardiac function and vascular reactivity due to aging (Delbin et al. 2012), although the mechanisms are not yet known. Numerous drugs have been studied to combat pathological conditions linked to aging, such as heart failure (Little et al., 2005), diabetic nephropathy (Thallas-Bonke et al., 2004), type II diabetes (Freidja et al., 2012), ventricular and vascular stiffness (Steppan et al., 2012) but from preclinical experimentation no compound has so far emerged as a safe and effective agent. Another therapeutic attempt was the use of growth hormone (GH) because some studies had shown beneficial effects in the elderly (Taub et al., 2010), with an increase in muscle mass and strengthening of the immune system (de Magalhães, 2013). Side effects, such as metabolic alterations (Carroll et al., 1998), increase in arterial and intracranial pressure (Malozowski et al., 1993), onset of diabetes (Lewis et al., 2013) and the possibility that GH may stimulate the growth of neoplasms (Clayton et al., 2011) have decreed its failure. Antioxidant therapies were also thought to slow down the aging process, but large cohort studies have shown that these dietary supplements do not affect the aging process in patients who use them (Park et al., 2011); moreover, studies in mice have shown that antioxidant supplements accelerate tumorigenesis processes (Sayin et al., 2014). Even the overexpression of telomerase in mice, which preserves chromosomal stability, the use of telomerase did not show an anti-aging effect (de Magalhães and Toussaint, 2004), in addition to promoting tumor growth (Peterson et al., 2015). If the hypotheses of our study were verified, they would suggest the use EVs as a new approach to slow down the aging process, to attenuate chronic inflammation and to use them as possible therapy in neuroinflammation diseases. It could be hypothesized to take EVs released from microglial ¿young¿ cultures and administer them to patients in order to improve the quality of life in the aging phase, hypothesizing a reduction effect of neuroinflammation, responsible for the increased risk of diseases related to aging, and a slowing of cognitive deficits (such as memory and motor reflexes).
EVs include EXO and MVs, which carry proteins, lipids and RNA; they can bind to receptors on the target cell's membrane, merge with it, or be completely internalized. Among the biological effects of EVs identified so far are the fetus-maternal interaction (Pap et al 2008), blood coagulation (Leroyer et al. 2008), the transport of IL-1¿ (MacKenzie et al 2001). The potential therapeutic uses of engineered EVs have been demonstrated in Parkinson's disease (Kaney et al. 2015) and in Schwann cell cancer (Prabhakar et al. 2013). Studies conducted on mouse models of demyelinating diseases have shown that the administration of EVs improves the clinical picture (Zhang et al., 2014). The MV released by pro-inflammatory microglia (phenotype defined as anti-tumor) carry mRNAs capable of counteracting the growth and invasion of tumor cells and promoting the switch of anti-inflammatory, pro-tumor microglia towards the anti-tumor phenotype, pro-inflammatory (Grimaldi et al, 2019). Studies carried out on human genetic pathologies have revealed that immunotherapy is translatable to humans, because it is well tolerated by the body and with very encouraging clinical results (Cartier et al., 2009).
The potential economic impact of this study consists in a significant increase in the self-sufficiency of elderly subjects, reducing the economic efforts of the National Health Service and of families for assistance to the elderly. In addition, studies will be stimulated to search for new methods for the isolation of vesicles, for example, using human microglia lines in order to use them as an anti-aging therapy.