Immunophenotype profile by flow cytometry reveals different subtypes of extracellular vesicles in porcine seminal plasma

Autor: Isabel Barranco, Alberto Alvarez-Barrientos, Ana Parra, Pablo Martínez-Díaz, Xiomara Lucas, Jordi Roca
Jazyk: angličtina
Rok vydání: 2024
Předmět:
Zdroj: Cell Communication and Signaling, Vol 22, Iss 1, Pp 1-17 (2024)
Druh dokumentu: article
ISSN: 1478-811X
DOI: 10.1186/s12964-024-01485-1
Popis: Abstract Background Porcine seminal plasma (SP) is endowed with a heterogeneous population of extracellular vesicles (sEVs). This study evaluated the immunophenotypic profile by high-sensitivity flow cytometry of eight sEV subpopulations isolated according to their size (small [S-sEVs] and large [L-sEVs]) from four different SP sources, namely three ejaculate fractions (the first 10 mL of the sperm rich fraction [SRF-P1], the remaining SRF [SRF-P2], and the post-SRF [PSRF]) and entire ejaculate (EE). Methods Seminal EVs were isolated using a size exclusion chromatography-based protocol from six SP pools (five ejaculates/pool) of each SP source and characterized using complementary approaches including total protein (BCA™assay), particle size distribution (dynamic light scattering), morphology (transmission electron microscopy), and purity (albumin by Western blot). Expression of CD9, CD63, CD81, CD44 and HSP90β was analyzed in all sEV subpopulations by high-sensitivity flow cytometry according to MIFlowCyt-EV guidelines, including an accurate calibration, controls, and discrimination by CFSE-labelling. Results Each sEV subpopulation exhibited a specific immunophenotypic profile. The percentage of sEVs positive for CD9, CD63, CD81 and HSP90β differed between S- and L-sEVs (P
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