Molecular and functional profiling of apical versus basolateral small extracellular vesicles derived from primary human proximal tubular epithelial cells under inflammatory conditions

Abstract

Proximal tubular epithelial cells (PTEC) are central players in inflammatory kidney diseases. However, the complex signalling mechanism/s via which polarized PTEC mediate disease progression are poorly understood. Small extracellular vesicles (sEV), including exosomes, are recognized as fundamental components of cellular communication and signalling courtesy of their molecular cargo (lipids, microRNA, proteins). In this study, we examined the molecular content and function of sEV secreted from the apical versus basolateral surfaces of polarized human primary PTEC under inflammatory diseased conditions. PTEC were cultured under normal and inflammatory conditions on Transwell inserts to enable separate collection and isolation of apical/basolateral sEV. Significantly increased numbers of apical and basolateral sEV were secreted under inflammatory conditions compared with equivalent normal conditions. Multi‐omics analysis revealed distinct molecular profiles (lipids, microRNA, proteins) between inflammatory and normal conditions for both apical and basolateral sEV. Biological pathway analyses of significantly differentially expressed molecules associated apical inflammatory sEV with processes of cell survival and immunological disease, while basolateral inflammatory sEV were linked to pathways of immune cell trafficking and cell‐to‐cell signalling. In line with this mechanistic concept, functional assays demonstrated significantly increased production of chemokines (monocyte chemoattractant protein‐1, interleukin‐8) and immuno‐regulatory cytokine interleukin‐10 by peripheral blood mononuclear cells activated with basolateral sEV derived from inflammatory PTEC. We propose that the distinct molecular composition of sEV released from the apical versus basolateral membranes of human inflammatory PTEC may reflect specialized functional roles, with basolateral‐derived sEV pivotal in modulating tubulointerstitial inflammatory responses observed in many immune‐mediated kidney diseases. These findings provide a rationale to further evaluate these sEV‐mediated inflammatory pathways as targets for biomarker and therapeutic development.

Extracellular Vesicles from Uterine Aspirates Represent a Promising Source for Screening Markers of Gynecologic Cancers

“…), which cannot always be clearly distinguished by size, density, morphology and other physical characteristics [ 21 , 42 ]. Furthermore, a great diversity in molecular composition is shown even for EVs with similar physical characteristics and originating from the same cells [ 43 , 44 , 45 ].…”

miR-6869-5p Transported by Plasma Extracellular Vesicles Mediates Renal Tubule Injury and Renin-Angiotensin System Activation in Obesity

“…Protein samples from PTECs (miR-6869-5p-transfected and EV-treated cells) were analyzed by Western blotting, as previously described ( 19 , 33 , 34 ). We used the following primary antibodies against several EV-characteristic markers: CD9 (#ab92726, Abcam, Cambridge, MA, USA), CD81 (#sc-7637, 1:200, Santa Cruz Biotechnology, USA), CD63 (#ab59479, Abcam, 1:1,000), AGT (AF3156, R&D Systems, Minneapolis, MN, 1:1,000), ACE2 (AF333, R&D Systems, 1:1,000), ACE (AF929, R&D Systems, 1:1,000), AT1 (MAB102441, R&D Systems, 1:1,000), KIM-1 (NBP1-76701SS, Novus, 1:1,000), and NGAL (AF1757-SP, R&D Systems, 1:1,000).…”

Vectorial Release of Human RNA Viruses from Epithelial Cells

“…The apical exosomes of MDCK cells polarized in culture are enriched in HSP70, GPR5C and CD63 while the basolateral exosomes have more TSG101, CD81 and CD9 [25,26]. This depends on the cell type since CD81 is enriched in the extracellular vesicles secreted from the apical side of proximal tubular epithelial cells [27].…”