The use of exosomes in clinical settings is progressively becoming a reality, as clinical trials testing exosomes for diagnostic and therapeutic applications are generating remarkable interest from the scientific community and investors. Exosomes are small extracellular vesicles secreted by all cell types playing intercellular communication roles in health and disease by transferring cellular cargoes such as functional proteins, metabolites and nucleic acids to recipient cells. An in-depth understanding of exosome biology is therefore essential to ensure clinical development of exosome based investigational therapeutic products. Here we summarise the most up-to-date knowkedge about the complex biological journey of exosomes from biogenesis and secretion, transport and uptake to their intracellular signalling. We delineate the major pathways and molecular players that influence each step of exosome physiology, highlighting the routes of interest, which will be of benefit to exosome manipulation and engineering. We highlight the main controversies in the field of exosome research: their adequate definition, characterisation and biogenesis at plasma membrane. We also delineate the most common identified pitfalls affecting exosome research and development. Unravelling exosome physiology is key to their ultimate progression towards clinical applications.
Blazek et al. demonstrate that treatment with IL-28A reduces inflammation in collagen-induced arthritis by restricting the recruitment of IL-1β+ neutrophils.
Whereas the importance of macrophages in chronic inflammatory diseases is well recognized, there is an increasing awareness that neutrophils may also play an important role. In addition to the well-documented heterogeneity of macrophage phenotypes and functions, neutrophils also show remarkable phenotypic diversity among tissues. Understanding the molecular pathways that control this heterogeneity should provide abundant scope for the generation of more specific and effective therapeutics. We have shown that the transcription factor IFN regulatory factor 5 (IRF5) polarizes macrophages toward an inflammatory phenotype. IRF5 is also expressed in other myeloid cells, including neutrophils, where it was linked to neutrophil function. In this study we explored the role of IRF5 in models of acute inflammation, including antigen-induced inflammatory arthritis and lung injury, both involving an extensive influx of neutrophils. Mice lacking IRF5 accumulate far fewer neutrophils at the site of inflammation due to the reduced levels of chemokines important for neutrophil recruitment, such as the chemokine (C-X-C motif) ligand 1. Furthermore we found that neutrophils express little IRF5 in the joints and that their migratory properties are not affected by the IRF5 deficiency. These studies extend prior ones suggesting that inhibiting IRF5 might be useful for chronic macrophage-induced inflammation and suggest that IRF5 blockade would ameliorate more acute forms of inflammation, including lung injury.macrophages | neutrophils | IRF5 | acute inflammation | arthritis
For inherited genetic diseases, fetal gene therapy offers the potential of prophylaxis against early, irreversible and lethal pathological change. To explore this, we studied neuronopathic Gaucher disease (nGD), caused by mutations in GBA. In adult patients, the milder form presents with hepatomegaly, splenomegaly and occasional lung and bone disease; this is managed, symptomatically, by enzyme replacement therapy. The acute childhood lethal form of nGD is untreatable since enzyme cannot cross the blood-brain barrier. Patients with nGD exhibit signs consistent with hindbrain neurodegeneration, including neck hyperextension, strabismus and, often, fatal apnea. We selected a mouse model of nGD carrying a loxP-flanked neomycin disruption of Gba plus Cre recombinase regulated by the keratinocyte-specific K14 promoter. Exclusive skin expression of Gba prevents fatal neonatal dehydration. Instead, mice develop fatal neurodegeneration within 15 days. Using this model, fetal intracranial injection of adeno-associated virus (AAV) vector reconstituted neuronal glucocerebrosidase expression. Mice lived for up to at least 18 weeks, were fertile and fully mobile. Neurodegeneration was abolished and neuroinflammation ameliorated. Neonatal intervention also rescued mice but less effectively. As the next step to clinical translation, we also demonstrated the feasibility of ultrasound-guided global AAV gene transfer to fetal macaque brains.
Macrophages are an integral part of the innate immune system and key players in pathogen clearance and tissue remodelling. Both functions are accomplished by a pivotal network of different macrophage subtypes, including proinflammatory M1 and anti-inflammatory M2 macrophages. Previously, our laboratory identified the transcription factor interferon regulatory factor 5 (IRF5) as the master regulator of the M1 macrophage polarisation. IRF5 was found to be highly expressed in human M1 compared to M2 macrophages. Furthermore, IRF5 dictates the expression of proinflammatory genes such as IL12b and IL23a whilst repressing anti-inflammatory genes like IL10. Here we show that murine bone marrow derived macrophages differentiated in vitro with GM-CSF are also characterised by high levels of IRF5 mRNA and protein and express proinflammatory cytokines upon LPS stimulation. These macrophages display characteristic expression of M1-marker MHC II but lack the M2-marker CD206. Significantly, we develop intracellular staining of IRF5- expressing macrophages and utilise it to recapitulate the in vitro results in an in vivo model of antigen-induced arthritis, emphasising their physiological relevance. Thus, we establish the species-invariant role of IRF5 in controlling the inflammatory macrophage phenotype both in vitro and in in vivo.
IntroductionType 4 phosphodiesterases (PDE4) play an important role in immune cells through the hydrolysis of the second messenger, cAMP. Inhibition of PDE4 has previously been shown to suppress immune and inflammatory responses, demonstrating PDE4 to be a valid therapeutic target for immune-mediated pathologies. We assessed the anti-inflammatory effects of a novel PDE4 inhibitor, apremilast, in human synovial cells from rheumatoid arthritis (RA) patients, as well as two murine models of arthritis.MethodsCells liberated from tissue excised from arthritic joints of RA patients were cultured in the presence of increasing concentrations of apremilast for 48 hours and spontaneous tumour necrosis factor-alpha (TNFα) production was analysed in culture supernatants by ELISA. In addition, arthritis was induced in BALB/c and DBA/1 mice by passive transfer of anti-type II collagen mAb and immunisation with type II collagen, respectively. Mice with established arthritis received 5 or 25 mg/kg apremilast and disease severity was monitored relative to mice receiving vehicle alone. At the end of the study, paws were removed and processed for histopathological assessment. Behavioural effects of apremilast, relative to rolipram, were assessed in naïve DBA/1 mice using an automated activity monitor (LABORAS).ResultsApremilast dose dependently inhibited spontaneous release of TNFα from human rheumatoid synovial membrane cultures. Furthermore, apremilast significantly reduced clinical score in both murine models of arthritis over a ten day treatment period and maintained a healthy joint architecture in a dose-dependent manner. Importantly, unlike rolipram, apremilast demonstrated no adverse behavioural effects in naïve mice.ConclusionsApremilast is an orally available PDE4 inhibitor that reduces TNFα production from human synovial cells and significantly suppresses experimental arthritis. Apremilast appears to be a potential new agent for the treatment of rheumatoid arthritis.
Conclusion. We propose that a therapeutic strategy that targets TNFRI while sparing TNFRII has the potential to both inhibit inflammation and promote Treg cell activity, which might be superior to TNF blockade.
Exosomes are a subset of extracellular vesicles essential for cell-cell communication in health and disease with the ability to transport nucleic acids, functional proteins and other metabolites. Their clinical use as diagnostic biomarkers and therapeutic carriers has become a major field of research over recent years, generating rapidly expanding scientific interest and financial investment. Their reduced immunogenicity compared to liposomes or viral vectors and their ability to cross major physiological barriers like the blood-brain barrier make them an appealing and innovative option as biomarkers and therapeutic agents. Here, we review the latest clinical developments of exosome biotechnology for diagnostic and therapeutic purposes, including the most recent COVID-19-related exosome-based clinical trials. We present current exosome engineering strategies for optimal clinical safety and efficacy, and assess the technology developed for good manufacturing practice compliant scaling up and storage approaches along with their limitations in pharmaceutical industry.
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