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Extracellular vesicles (EVs) are small, membranous structures involved in intercellular communication. Here, we analyzed the effects of thyroid cancer-derived EVs on the properties of normal thyroid cells and cells contributing to the tumor microenvironment. EVs isolated from thyroid cancer cell lines (CGTH, FTC-133, 8505c, TPC-1 and BcPAP) were used for treatment of normal thyroid cells (NTHY), as well as monocytes and endothelial cells (HUVEC). EVs’ size/number were analyzed by flow cytometry and confocal microscopy. Gene expression, protein level and localization were investigated by qRT-PCR, WB and ICC/IF, respectively. Proliferation, migration and tube formation were analyzed. When compared with NTHY, CGTH and BcPAP secreted significantly more EVs. Treatment of NTHY with cancer-derived EVs changed the expression of tetraspanin genes, but did not affect proliferation and migration. Cancer-derived EVs suppressed tube formation by endothelial cells and did not affect the phagocytic index of monocytes. The number of 6 μm size fraction of cancer-derived EVs correlated negatively with the CD63 and CD81 expression in NTHY cells, as well as positively with angiogenesis in vitro. Thyroid cancer-derived EVs can affect the expression of tetraspanins in normal thyroid cells. It is possible that 6 μm EVs contribute to the regulation of NTHY gene expression and angiogenesis.