6.1.4. Cell-Derived Nanoparticles

Extracellular vesicles are membrane-containing vesicles from the endocytic pathway or plasma membrane, released into the extracellular space by virtually all cells. Three types of extracellular vesicles exist: (a) exosomes, the smallest vesicles (30–150 nm), derived from the inward budding of multivesicular bodies; (ii) microvesicles, or ectosomes (50 nm– 1 μm), result from outward budding of the plasma membrane, released under physiological conditions or in response to specific stimuli; and (c) apoptotic bodies (50 nm–5 μm), which are produced by cells undergoing apoptosis.

Exosomes contain cellular proteins, lipids, nucleic acids, mRNAs, and microRNAs (miRNAs) from host cells (Figure 3). Exosomes can be internalized by cells and functionally modify the cells that internalize them. Exosomes can be derived from activated or nonactivated cells or loaded with therapeutic molecules to be used as cargo.

The most commonly used cells for the use of exosomes as therapeutic cargo are mesenchymal stromal cells (MSCs). Exosomes derived from MSCs or MSCs preconditioned by hypoxia (increased miR-21 expression) were systemically administered to APP/PS1 transgenic mice mimicking AD [88]. In both cases, the use of MSC-derived exosomes or hypoxia-preconditioned MSCs had positive effects. Administration of exosomes from hypoxia-preconditioned MSCs improved memory and learning abilities; decreased plaque deposition and Aβ levels; increased expression of growth-associated protein 43, synapsin-1, and IL-10; and decreased levels of GFAP, Iba-1, TNF-α, IL-1β, and activation of STAT3 and NF-κB [85]. The use of these exosomes would correct synaptic dysfunction as well as inflammatory responses, which would lead to improvement of cognitive decline observed in AD via miR-21 signaling [88]. Human umbilical cord mesenchymal stem cells (hucMSC-exosomes) were injected into mouse models of AD and were found to improve cognitive decline and decrease Aβ deposition [89]. These hucMSC-exosomes also modulated microglial activation with a decrease in the number of activated microglia and a shift toward an M2 anti-inflammatory profile (increase in IL-10 and TGF-β cytokines) with an increase in Aβ-degrading enzymes [89]. Brain perfusion of neuroblastoma-derived exosomes may mediate Aβ clearance in an AD mouse model. Indeed, Aβ peptides can be taken up and transported by exosomes for presentation to microglia, resulting in their degradation [90,91].

Exosomes can also be loaded with a therapeutic molecule. The exosomes can be passively incubated with the therapeutic molecule, followed by purification. Curcumin was incubated at 22 ◦C for 5 min with exosomes, and the mixture was then effectively administered to the brain by intranasal route [92,93]. This decreased inflammation via microglia targeting [92,93]. Other exosome loading strategies exist, such as electroporation, incubation at room temperature, permeabilization with saponin, freeze/thaw cycles, sonication, and extrusion, and have been tested in neurodegenerative diseases but without microglia targeting or in other pathologies like cancer.

Exosomes are able to modulate the inflammatory response and phagocytosis activity of microglia and can be considered as a very interesting possibility in therapy.

#### 6.1.5. Antioxidant Nanoparticles

Nanoparticles could also be synthesized directly from an antioxidant like quercetin. Authors have obtained quercetin nanoparticles with a very heterogeneous size ranging from 520 to 750 nm. In a model of AD, these quercetin nanoparticles were able to reduce neuronal damage, decrease the formation of amyloid plaques and neurofibrillary tangles, and modulate the activity of microglia [94].

In the context of PD, a nanoparticle (NP) formulation containing two polyphenol antioxidants, tannic acid (TA) and a ferulic acid diacid molecule, was proposed. These antioxidant nanoparticles inhibited α-synuclein fibrillation and lowered intracellular αsynuclein oligomerization in the BV-2 microglial line subjected to a high concentration of extracellular α-synuclein, thereby ameliorating microglial oxidative stress [95]. Microglial

activation is also reduced with a modulation of the production of the pro-inflammatory cytokines TNF-α and IL-6 [95].

#### *6.2. Dendrosomal Nanoparticles*

An Iranian team used the dendrosome, a neutral, amphipathic, and biodegradable nanomaterial, to transport a molecule of interest such as curcumin. In a cuprizone-induced model of MS, these curcumin-loaded dendrosomes suppressed the accumulation of microglia and astrocytes, highlighting their possible use in therapy [96].

## *6.3. Dendrimers and Other Dendritic Polymers*

Dendritic polymers belong to the synthetic polymers with linear, cross-linked, and branched polymers. Among these are the dendrimers which are obtained after generational synthesis (Figure 3), resulting in the formation of theoretically monodisperse structures with a narrow molecular weight distribution. In contrast to dendrimers, hyperbranched polymers are polydisperse, with a broad molecular weight distribution. The third members are dendrigrafts and have a configuration that shares commonalities between dendrimers and hyperbranched polymers (Figure 3).

Dendrimers, which are synthetic molecules with a tree-like structure, can be constructed by different methods: the hyperbranched structure can be built from the core, layer by layer, or by attaching dendrons to a central core [97,98]. Among the best-known dendrimers are poly(amidoamine) (PAMAM), poly(propylene imine) (PPI), phosphorus, and dimethylolpropionic acid-based dendrimers. One team has developed a dendrimer that targets the mitochondria (using triphenyl-phosphonium (TPP)) and delivers the antioxidant N-acetylcysteine (NAC):mitochondrial targeting hydroxyl PAMAM dendrimer-drug construct (TPP-D-NAC) [99]. The authors had already shown in previous studies that this PAMAM polymer was able to cross the BBB by selectively targeting activated microglia/macrophages and was able to deliver NAC to these cells (dendrimer D-NAC) [100]. These TPP-D-NAC dendrimers show preferential targeting to mitochondria (but not only) of activated microglial/macrophage cells. They showed superior efficacy in terms of oxidative stress inhibition to dendrimers developed from NAC but without organelle targeting (D-NAC) and to NAC used alone [99]. Concerning the dendrimers, the functionalization of the surface is an important factor to take into account, as it can play on the toxicity of the molecule [101]. Peroxisome proliferator-activated receptor (PPAR) α and PPARγ agonists can switch microglia from an M1-like to an M2-like phenotype. Because hydroxyl-terminated polyamidoamine dendrimers cross the altered BBB at the site of neuroinflammation and accumulate in activated microglia, they are conjugated with a PPARα/γ dual agonist [102,103]. The dual agonist dendrimer-PPARα/γ conjugate (Dtesaglitazar) induced the following: (a) an "M1 to M2" phenotype change, (b) a decrease in reactive oxygen species secretion, (c) an increase in the expression of phagocytosis and enzymatic degradation genes of pathogenic proteins, and (d) an increase in phagocytosis of β-amyloid [104].

Dendrigrafts have also been used to deliver treatment to the brain. Caspase-3 is involved in cell death and inhibiting it could help prevent the progression of neurodegenerative diseases and, in this study, PD. For this purpose, RNA interference was used as well as a vector constituted by dendrigraft poly-L-lysines, on which a peptide glycoprotein of the rabies virus with 29 amino acids was bound, allowing it to cross the BBB by transcytosis mediated by a specific receptor [105]. Plasmid DNA encoding the short hairpin RNA of caspase-3 was complexed with this vector to give nanoparticles. Injection by weekly intravenous administration of the nanoparticles reduced the levels of activated caspase-3, which decreased dopaminergic neuronal loss in the brains of rats with PD [105]. In addition, the rat model was obtained by treating them with rotenone, which increases TNF-α and NO levels in the brain. The use of these nanoparticles reduced the levels of TNF-α and NO in the brain [105]. These nanoparticles would thus have an anti-inflammatory effect on the microglia according to this study [105].
