Recent findings have shown that nanovesicles preparations from either primary immune cells culture supernatants or plasma contain immunoglobulins, suggesting that a natural way of antibody production may be through exosome release. To verify this hypothesis, we used the OKT3 hybridoma clone, which produces a murine IgG2a monoclonal antibody used to reduce rejection in patients undergoing organ transplantation. We showed exosome-associated immunoglobulins in hybridoma supernatants, by Western blot, nanoscale flow cytometry and immunocapture-based ELISA. The OKT3-exo was also being able to trigger cytokines production in both CD4 and CD8 T cells. These results show that nanovesicles contain immunoglobulin and could be used for immunotherapy. These data could lead to a new approach to improve the effectiveness of therapeutic antibodies by exploiting their natural property to be expressed on nanovesicle membrane, that probably render them more stable and as a consequence more capable to interact with their specific ligand in the best way.

Cholesterol has been implicated in the clinical progression of breast cancer, a disease that continues to be the most commonly diagnosed cancer in women. Previous work has identified the cholesterol metabolite 27-hydroxycholesterol (27HC) as a major mediator of the effects of cholesterol on breast tumor growth and progression. 27HC can act as an estrogen receptor (ER) modulator to promote the growth of ERα+ tumors, and as a liver X receptor (LXR) ligand in myeloid immune cells to establish an immune-suppressive program. In fact, the metastatic properties of 27HC require the presence of myeloid cells with neutrophils (polymorphonuclear neutrophils; PMNs) being essential for the increase in lung metastasis in murine models. In an effort to further elucidate the mechanisms by which 27HC alters breast cancer progression, we made the striking finding that 27HC promoted the secretion of extracellular vesicles (EVs), a diverse assortment of membrane bound particles that includes exosomes. The resulting EVs had a size distribution that was skewed slightly larger than EVs generated by treating cells with vehicle. The increase in EV secretion and size was consistent across 3 different subtypes: primary murine PMNs, RAW264.7 monocytic cells, and 4T1 murine mammary cancer cells. Label-free analysis of 27HC-EVs indicated that they had a different metabolite composition to those from vehicle-treated cells. Importantly, 27HC-EVs from primary PMNs promoted tumor growth and metastasis in 2 different syngeneic models, demonstrating the potential role of 27HC-induced EVs in the progression of breast cancer. EVs from PMNs were taken up by cancer cells, macrophages, and PMNs, but not T cells. Since EVs did not alter proliferation of cancer cells, it is likely that their protumor effects are mediated through interactions with myeloid cells. Interestingly, RNA-seq analysis of tumors from 27HC-EV-treated mice do not display significantly altered transcriptomes, suggesting that the effects of 27HC-EVs occur early on in tumor establishment and growth. Future work will be required to elucidate the mechanisms by which 27HC increases EV secretion, and how these EVs promote breast cancer progression. Collectively, however, our data indicate that EV secretion and content can be regulated by a cholesterol metabolite, which may have detrimental effects in terms of disease progression, important findings given the prevalence of both breast cancer and hypercholesterolemia.
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Microbial density and diversity increase towards the distal intestine, affecting tissue physiology, metabolism, and function of both immune and nervous systems. Intrinsic enteric–associated neurons (iEAN) continuously monitor and modulate intestinal functions, including nutrient absorption and motility. Through molecular, anatomic and functional approaches, we characterized the influence of the microbiota on iEAN. We found that iEAN are functionally adapted to the intestinal segment they occupy, with a stronger microbiota influence on distal intestine neurons. Chemogenetic characterization of microbiota-influenced iEAN identified a subset of viscerofugal CART+ neurons, enriched in the distal intestine, able to modulate feeding through insulin-glucose levels. Retro- and anterograde tracing revealed that CART+ viscerofugal neurons send axons to the gut sympathetic ganglion and are synaptically connected to the liver and pancreas. Our results demonstrate a region-specific adaptation of enteric neurons and indicate that specific iEAN subsets are capable of regulating host physiology independently from the central nervous system. h4>One Sentence Summary/h4> Microbes impact regionally defined intrinsic enteric neuron translatomes, including a novel CART+ glucoregulatory viscerofugal population.

Generation of secondary alveolar septa occurs primarily after birth in humans and is complete in mice postnatally, when mechanical stresses vary as air space pressure oscillates. Alveolar mesenchymal cells deposit elastic fibers, which limit cell strain; although when the elastic fiber network is incomplete, this function is also served by the intracellular cytoskeleton. Intermediate filament proteins support deformation during cell division and migration, which occur during septal elongation. Because platelet-derived growth factor receptor-α (PDGFRα) signaling is essential for alveolar septation, we hypothesized that neuropilin-1 (NRP1) may link PDGFRα to cytoskeletal deformation. During cell migration, NRP1 links receptor tyrosine kinase signaling to cytoskeletal and focal adhesion remodeling. Therefore, we examined the consequences of nrp1 gene deletion in alveolar mesenchymal cells (myofibroblasts and pericytes). NRP1 depletion reduced the proportion of mesenchymal cells that contain nestin and desmin within the subpopulation that lacked PDGFRα but contained PDGFRβ. Desmin was reduced at alveolar entry rings, air spaces were enlarged, and surface area was reduced after NRP1 depletion. PDGFRα and NRP1 colocalized to membrane lipid rafts, which are known to contain Src kinase. NRP1 depletion reduced alveolar mesenchymal cell migration and PDGF-A-mediated activation of Src kinase, which may limit accumulation of desmin at septal tips (alveolar entry rings). Cooperation between NRP1 and PDGF signaling is required for secondary septation, and manipulation of NRP1 could promote alveolar regeneration without producing fibrosis.