Abstract
The release of exosomes can lead to cell–cell communication. Nutrients such as vitamin D3 and sphingolipids have important roles in many cellular functions, including proliferation, differentiation, senescence, and cancer. However, the specific composition of sphingolipids in exosomes and their changes induced by vitamin D3 treatment have not been elucidated. Here, we initially observed neutral sphingomyelinase and vitamin D receptors in exosomes released from HN9.10 embryonic hippocampal cells. Using ultrafast liquid chromatography tandem mass spectrometry, we showed that exosomes are rich in sphingomyelin species compared to whole cells. To interrogate the possible functions of vitamin D3, we established the optimal conditions of cell treatment and we analyzed exosome composition. Vitamin D3 was identified as responsible for the vitamin D receptor loss, for the increase in neutral sphingomyelinase content and sphingomyelin changes. As a consequence, the generation of ceramide upon vitamin D3 treatment was evident. Incubation of the cells with neutral sphingomyelinase, or the same concentration of ceramide produced in exosomes was necessary and sufficient to stimulate embryonic hippocampal cell differentiation, as vitamin D3. This is the first time that exosome ceramide is interrogated for mediate the effect of vitamin D3 in inducing cell differentiation.
Highlights
Exosomes are a subset of extracellular vesicles universally recognized as natural nanoparticles involved in numerous biologic processes and clinical diseases
In order to test whether physiological concentration of vitamin D3 (VD3) (100 nmol/L), previously used to induce HN9.10 cell differentiation [16,17] was the better concentration, we first set out to study the dose-dependent effect of VD3 on HN9.10 cell viability after 24 h of culture by using MTT assay
Given the interest in exosomes in cell-to-cell communication, it is perhaps surprising that the role of VD3 in exosome lipid composition has not been investigated
Summary
Exosomes are a subset of extracellular vesicles universally recognized as natural nanoparticles involved in numerous biologic processes and clinical diseases. They have specific proteins, lipids, RNA, and DNA compositions that can be transferred from a donor cell to a recipient allowing cell-to-cell communication useful for maintaining cell functions and tissue homeostasis [1]. Numerous cell types are capable of releasing exosomes in different biological fluids [4]. Exosomes influence the fate of the recipient cells [1] Due to their action, they have been considered the main players in different physiopathological conditions [4] and useful diagnostic or prognostic markers [7]. A mass spectrometric proteomics characterization of exosomes purified from the bronchoalveolar lavage fluid of patients with sarcoidosis and healthy control subjects was performed by identifying
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