Newt cells secrete extracellular vesicles with therapeutic bioactivity in mammalian cardiomyocytes

Abstract

Newts can regenerate amputated limbs and cardiac tissue, unlike mammals which lack broad regenerative capacity. Several signaling pathways involved in cell proliferation, differentiation and survival during newt tissue regeneration have been elucidated, however the factors that coordinate signaling between cells, as well as the conservation of these factors in other animals, are not well defined. Here we report that media conditioned by newt limb explant cells (A1 cells) protect mammalian cardiomyocytes from oxidative stress-induced apoptosis. The cytoprotective effect of A1-conditioned media was negated by exposing A1 cells to GW4869, which suppresses the generation of extracellular vesicles (EVs). A1-EVs are similar in diameter (~100–150 nm), structure, and share several membrane surface and cargo proteins with mammalian exosomes. However, isolated A1-EVs contain significantly higher levels of both RNA and protein per particle than mammalian EVs. Additionally, numerous cargo RNAs and proteins are unique to A1-EVs. Of particular note, A1-EVs contain numerous mRNAs encoding nuclear receptors, membrane ligands, as well as transcription factors. Mammalian cardiomyocytes treated with A1-EVs showed increased expression of genes in the PI3K/AKT pathway, a pivotal player in survival signaling. We conclude that newt cells secrete EVs with diverse, distinctive RNA and protein contents. Despite ~300 million years of evolutionary divergence between newts and mammals, newt EVs confer cytoprotective effects on mammalian cardiomyocytes.

Extracellular Vesicles in the Development of Cancer Therapeutics

“…This includes endosomal sorting complexes required for transport (ESCRT) and tetraspanins necessary for intraluminal vesicle formation, sphingomyelinase to generate ceramides vital for intraluminal vesicles’ formation and sorting, and Rab27a and Rab27b critical for cellular endosomal trafficking [ 55 , 71 , 73 ]. In an early effort to suppress sEVs’ biogenesis, GW4869, a sphingomyelinase inhibitor, was used, which reduced ceramide generation and inhibited sEV formation [ 74 ]. Furthermore, attenuation of neutral sphingomyelinase 2 (nSMase2) in breast cancer cells by a knockdown approach reduced sEV formation and attenuated sEV-associated miR-210 transfer, leading to the suppression of cancer cell metastasis in vitro and in a xenograft mouse model [ 75 ].…”

Multiplex profiling identifies clinically relevant signalling proteins in an isogenic prostate cancer model of radioresistance

“…Following transfer membranes were blocked using 5% w/v milk protein and probed using primary antibodies directed against PARP1 (#sc-8007; 1:100, Santa Cruz, UK 56 ), AR (#sc-377546, 1:250, Santa Cruz), p53 (#sc-126, 1:500, Santa Cruz, UK 57 ), Notch-3 (#ab23426, 1:250, Abcam, UK 58 ), HES-1 (#ab71559, 1:1000, Abcam 59 ) or YB-1 (#ab12148, 1:2000, Abcam 60 ). After washing, the membrane was incubated with horseradish peroxidase conjugated goat secondary antibody (sc-2020, 1:1000, Santa Cruz 61 ) or Anti-Rabbit IgG HRP Linked (#7074, 1:1000, Cell Signalling 62 ) antibodies. Following incubation with the primary and secondary antibodies, a detection reagent luminol (SC-2048, Santa Cruz) was applied to blots and chemiluminescence images were then developed using a Fujifilm LAS-4000 luminescent image analyser.…”

Exosome in Cardiovascular Diseases: A Complex World Full of Hope

“…A curious and unusual study was performed by Middleton and colleagues, who took a cue from newts, as they can regenerate lost organs and tissues, including the heart, and analyzed the compatibility of newt EVs with mammalian cells [60]. They extracted EVs form newt myogenic precursor cells (A1) and, after seeing that, in many ways (size, morphology, content, surface agents, and GW4869 sensitivity), they were similar to mammalian ones, they treated mammalian CMs with these vesicles.…”

“…They extracted EVs form newt myogenic precursor cells (A1) and, after seeing that, in many ways (size, morphology, content, surface agents, and GW4869 sensitivity), they were similar to mammalian ones, they treated mammalian CMs with these vesicles. Gene expression analysis indicated that treated CMs were more resistant to oxidative stress and, thus, had enhanced cell survival, due to the activation of the Protein kinase B (AKT) pathway [60]. This evidence makes A1 EXOs and their content interesting candidates for therapeutic studies in cardiovascular fields.…”