The Release of Exosomes from Astrocytes in Response to Fatty Acid Alters the Metabolism of Proopiomelanocortin (POMC) Neurons

Abstract

Exosomes are extracellular micro-vesicles with a complex content that include a vast heterogeneity of molecules such as growth factors, cytokines and RNAs, including micro-RNAs (mi-RNAs) that affect target cells. Neurons are influenced by neighboring astrocytes through the release of exosomes amongst other signals. We postulated that the metabolic status is communicated by hypothalamic astrocytes via exosomes to neighboring POMC neurons to modify their functions in the promotion of energy expenditure and satiety. With this aim, cultures of primary hypothalamic astrocytes were treated with both palmitic or oleic acid (PA or OA; 0.5 mM) for 24 hours. Exosomes were isolated and purified from the media culture and used to treat (1.25 or 2.50 µg/mL) a POMC neuronal cell line for 24 hours. Exosomes released in response to PA (E-PA) or (E-OA) did not change the expression of the markers of inflammation [interleukin (IL)-6] and Endoplasmic Reticle stress (CHOP) comparing to exosomes released in response to vehicle (E-V) or with no exosomes (control). Next-Generation miRNA sequencing analysis was performed to reveal modifications in miRNAs contained in exosomes. The content of miR-199a-3p and miR-145-5p was higher in E-PA compared to E-V. POMC neurons transfected with a mimetic of miR-199a-3p (1.5 pmol) increased insulin-like growth factor 1 receptor (IGF1r) protein levels (p < 0.05), as well as POMC expression (ns). Moreover, levels of mTOR and p70S6k, reported targets of miR-199a-3p, were decreased (p < 0.05, both). Mimetic overexpression of miR- 145-5p reduced POMC expression (p < 0.001) and protein levels of insulin receptor substrate 1 (IRS1; p < 0.001), which is a known target of this miRNA. To determine modifications in cellular metabolism in the POMC neurons in response to exosomal astrocytes, Seahorse Cell Mito Stress test was performed. The mitochondrial spare respiratory capacity of neurons was increased (p < 0.0001) in response to both doses (1.25 or 2.50 µg/mL) of E-PA and E-OA, with the maximal respiration (p < 0.0001) also increasing with E-PA or E-OA (both doses) compared to E-V or control. Our results suggest that astrocytes may communicate directly with neurons via exosomes, and that the exosomes content is modulated by the source cell in response to the nutritional environment. The messages contained in exosomes from astrocytes can directly modulate the levels of cellular receptors and factors involved in cell proliferation, protection, metabolism, and nutrient sensing. Specific miRNAs participate in these processes as well as on the neuropeptide expression in targeted neurons. In addition, cellular respiration is affected in POMC neurons treated with exosomes from astrocytes in response to fatty acid in a way that suggests that cells are preparing for a possible respiratory stress by increasing their spare respiratory capacity and maximal respiration.