The term 'extracellular vesicles' refers to a heterogeneous population of vesicular bodies of cellular origin that derive either from the endosomal compartment (exosomes) or as a result of shedding from the plasma membrane (microvesicles, oncosomes and apoptotic bodies). Extracellular vesicles carry a variety of cargo, including RNAs, proteins, lipids and DNA, which can be taken up by other cells, both in the direct vicinity of the source cell and at distant sites in the body via biofluids, and elicit a variety of phenotypic responses. Owing to their unique biology and roles in cell-cell communication, extracellular vesicles have attracted strong interest, which is further enhanced by their potential clinical utility. Because extracellular vesicles derive their cargo from the contents of the cells that produce them, they are attractive sources of biomarkers for a variety of diseases. Furthermore, studies demonstrating phenotypic effects of specific extracellular vesicle-associated cargo on target cells have stoked interest in extracellular vesicles as therapeutic vehicles. There is particularly strong evidence that the RNA cargo of extracellular vesicles can alter recipient cell gene expression and function. During the past decade, extracellular vesicles and their RNA cargo have become better defined, but many aspects of extracellular vesicle biology remain to be elucidated. These include selective cargo loading resulting in substantial differences between the composition of extracellular vesicles and source cells; heterogeneity in extracellular vesicle size and composition; and undefined mechanisms for the uptake of extracellular vesicles into recipient cells and the fates of their cargo. Further progress in unravelling the basic mechanisms of extracellular vesicle biogenesis, transport, and cargo delivery and function is needed for successful clinical implementation. This Review focuses on the current state of knowledge pertaining to packaging, transport and function of RNAs in extracellular vesicles and outlines the progress made thus far towards their clinical applications.

/pdf/rna-delivery-by-extracellular-vesicles-in-mammalian-cells-pub81smyc0.pdf

RNA delivery by extracellular vesicles in mammalian cells and its applications.

Iron oxide nanoparticles (IONPs) have been extensively used during the last two decades, either as effective bio-imaging contrast agents or as carriers of biomolecules such as drugs, nucleic acids and peptides for controlled delivery to specific organs and tissues. Most of these novel applications require elaborate tuning of the physiochemical and surface properties of the IONPs. As new IONPs designs are envisioned, synergistic consideration of the body's innate biological barriers against the administered nanoparticles and the short and long-term side effects of the IONPs become even more essential. There are several important criteria (e.g. size and size-distribution, charge, coating molecules, and plasma protein adsorption) that can be effectively tuned to control the in vivo pharmacokinetics and biodistribution of the IONPs. This paper reviews these crucial parameters, in light of biological barriers in the body, and the latest IONPs design strategies used to overcome them. A careful review of the long-term biodistribution and side effects of the IONPs in relation to nanoparticle design is also given. While the discussions presented in this review are specific to IONPs, some of the information can be readily applied to other nanoparticle systems, such as gold, silver, silica, calcium phosphates and various polymers.

/pdf/in-vivo-delivery-pharmacokinetics-biodistribution-and-4qn10hldat.pdf

In vivo delivery, pharmacokinetics, biodistribution and toxicity of iron oxide nanoparticles

Extracellular vesicles (EVs) are nanometer-sized, lipid membrane-enclosed vesicles secreted by most, if not all, cells and contain lipids, proteins, and various nucleic acid species of the source cell. EVs act as important mediators of intercellular communication that influence both physiological and pathological conditions. Given their ability to transfer bioactive components and surmount biological barriers, EVs are increasingly being explored as potential therapeutic agents. EVs can potentiate tissue regeneration, participate in immune modulation, and function as potential alternatives to stem cell therapy, and bioengineered EVs can act as delivery vehicles for therapeutic agents. Here, we cover recent approaches and advances of EV-based therapies.

Advances in therapeutic applications of extracellular vesicles

Functional neural competence and integrity require interactive exchanges among sensory and motor neurons, interneurons and glial cells. Recent studies have attributed some of the tasks needed for these exchanges to extracellular vesicles (such as exosomes and microvesicles), which are most prominently involved in shuttling reciprocal signals between myelinating glia and neurons, thus promoting neuronal survival, the immune response mediated by microglia, and synapse assembly and plasticity. Such vesicles have also been identified as important factors in the spread of neurodegenerative disorders and brain cancer. These extracellular vesicle functions add a previously unrecognized level of complexity to transcellular interactions within the nervous system.

Extracellular vesicles round off communication in the nervous system

Patients with chronic inflammation are often associated with the emergence of depression symptoms, while diagnosed depressed patients show increased levels of circulating cytokines. Further studies revealed the activation of the brain immune cell microglia in depressed patients with a greater magnitude in individuals that committed suicide, indicating a crucial role for neuroinflammation in depression brain pathogenesis. Rapid advances in the understanding of microglial and astrocytic neurobiology were obtained in the past fifteen to twenty years. Indeed, recent data reveal that microglia play an important role in managing neuronal cell death, neurogenesis, and synaptic interactions, besides their involvement in immune-response generating cytokines. The communication between microglia and neurons is essential to synchronize these diverse functions with brain activity. Evidence is accumulating that secreted extracellular vesicles (EVs), comprising ectosomes and exosomes with a size ranging from 0.1 to 1 μm, are key players in intercellular signaling. These EVs may carry specific proteins, mRNAs and microRNAs (miRNAs). Transfer of exosomes to neurons was shown to be mediated by oligodendrocytes, microglia and astrocytes that may either be supportive to neurons, or instead disseminate the disease. Interestingly, several recent reports have identified changes in miRNAs in depressed patients, which target not only crucial pathways associated with synaptic plasticity, learning and memory but also the production of neurotrophic factors and immune cell modulation. In this article, we discuss the role of neuroinflammation in the emergence of depression, namely dynamic alterations in the status of microglia response to stimulation, and how their activation phenotypes may have an etiological role in neurodegeneneration, in particular in depressive-like behavior. We will overview the involvement of miRNAs, exosomes, ectosomes and microglia in regulating critical pathways associated with depression and how they may contribute to other brain disorders including amyotrophic lateral sclerosis, Alzheimer's disease and Parkinson disease, which share several neuroinflammatory-associated processes. Specific reference will be made to EVs as potential biomarkers and disease monitoring approaches, focusing on their potentialities as drug delivery vehicles and on putative therapeutic strategies using autologous exosome-based delivery systems to treat neurodegenerative and psychiatric disorders.

/pdf/neuroinflammation-and-depression-microglia-activation-3pndmc8ud1.pdf

Neuroinflammation and Depression: Microglia Activation, Extracellular Microvesicles and microRNA Dysregulation

IFNγ-stimulated dendritic cell exosomes as a potential therapeutic for remyelination.

Environmental enrichment (EE) consists of increased physical, intellectual, and social activity, and has wide-ranging effects, including enhancing cognition, learning and memory, and motor coordination. Animal studies have demonstrated that EE improves outcome of brain trauma and neurodegenerative disorders, including demyelinating diseases like multiple sclerosis, making it a promising therapeutic option. However, the complexity of applying a robust EE paradigm makes clinical use difficult. A better understanding of the signaling involved in EE-based neuroprotection may allow for development of effective mimetics as an alternative. In prior work, we found that exosomes isolated from the serum of rats exposed to EE impact CNS myelination. Exosomes are naturally occurring nanovesicles containing mRNA, miRNA, and protein, which play important roles in cell function, disease, and immunomodulation. When applied to hippocampal slice cultures or nasally administered to naive rats, EE-serum exosomes significantly increase myelin content, oligodendrocyte precursor (OPC) and neural stem cell levels, and reduce oxidative stress (OS). We found that rat EE exosomes were enriched in miR-219, which is necessary and sufficient for OPC differentiation into myelinating cells. Thus, peripherally produced exosomes may be a useful therapy for remyelination. Here, we aim to better characterize the impact of EE on CNS health and to determine the cellular source of nutritive exosomes found in serum. We found that exosomes isolated from various circulating immune cell types all increased slice culture myelin content, contained miR-219, and reduced OS, suggesting that EE globally alters immune function in a way that supports brain health.

Environmental Enrichment Stimulates Immune Cell Secretion of Exosomes that Promote CNS Myelination and May Regulate Inflammation.

Microglia play an important role in fine-tuning neuronal activity. In part, this involves their production of tumor necrosis factor-alpha (TNFa), which increases neuronal excitability. Excessive synaptic activity is necessary to initiate spreading depression (SD). Increased microglial production of proinflammatory cytokines promotes initiation of SD, which, when recurrent, may play a role in conversion of episodic to high frequency and chronic migraine. Previous work shows that this potentiation of SD occurs through increased microglial production of TNFa and reactive oxygen species, both of which are associated with an M1-skewed microglial population. Hence, we explored the role of microglia and their M1 polarization in SD initiation. Selective ablation of microglia from rat hippocampal slice cultures confirmed that microglia are essential for initiation of SD. Application of minocycline to dampen M1 signaling led to increased SD threshold. In addition, we found that SD threshold was increased in rats exposed to environmental enrichment. These rats had increased neocortical levels of interleukin-11 (IL-11), which decreases TNFa signaling and polarized microglia to an M2a-dominant phenotype. M2a microglia reduce proinflammatory signaling and increase production of anti-inflammatory cytokines, and therefore may protect against SD. Nasal administration of IL-11 to mimic effects of environmental enrichment likewise increased M2a polarization and increased SD threshold, an effect also seen in vitro. Similarly, application of conditioned medium from M2a polarized primary microglia to slice cultures also increased SD threshold. Thus, microglia and their polarization state play an essential role in SD initiation, and perhaps by extension migraine with aura and migraine.

/pdf/spreading-depression-requires-microglia-and-is-decreased-by-1yx34sk3po.pdf

Spreading depression requires microglia and is decreased by their M2a polarization from environmental enrichment.

This study explored the novel use of iron oxide (IO) nanoparticles ( 90%) internalized IO nanoparticles, but only the latter were significantly activated, with elevated expression/secretion of CD86, cytokines (IL-6, TNF-α, IL1-b, IFN-γ, and IL-12), and chemokines (CXCL1, CXCL2, CCL2, CCL3, CCL4, and CXCL10). Thus, the IO nanoparticles is a novel, safe, and effective vaccine platform, with built-in adjuvancy, that is highly stable and field deployable for cost-effective vaccine delivery.

Iron oxide nanoparticles as a clinically acceptable delivery platform for a recombinant blood-stage human malaria vaccine

A nanoparticle-based delivery system and methods for its use are disclosed. In one aspect, a nanoparticle-based delivery system comprising at least one molecule such as proteins, DNA/RNA or fragments thereof, carbohydrates, enzymes, chemicals, virus cells, bacteria, parts of a virus, parts of a bacteria, parts of a cell, part of a tissue, or a combination of one or more of these, which shall be referred to as immunogens, are chemically or physically combined with water soluble nanoparticles which, when administered to a living system, is capable of eliciting a desired immunological response. More particularly, the invention relates to nanoparticle-based delivery systems that are specifically engineered to enhance humoral or cellular immune response without the use of adjuvants.

Kae M. Pusic

Papers

IFNγ-stimulated dendritic cell exosomes as a potential therapeutic for remyelination.

Environmental Enrichment Stimulates Immune Cell Secretion of Exosomes that Promote CNS Myelination and May Regulate Inflammation.

Spreading depression requires microglia and is decreased by their M2a polarization from environmental enrichment.

Iron oxide nanoparticles as a clinically acceptable delivery platform for a recombinant blood-stage human malaria vaccine

Nanoparticle based immunological stimulation