Journal: Communications biology
Article Title: Small EV in plasma of triple negative breast cancer patients induce intrinsic apoptosis in activated T cells.
doi: 10.1038/s42003-023-05169-3
Figure Lengend Snippet: Fig. 5 TEX-induced apoptosis of activated human CD8 + T cells is not blocked by neutralizing Abs or specific inhibitors. a Apoptosis (%) induced in CD8 + T cells by TEX1 or TEX2 (50 µg/mL) was not significantly blocked by the neutralizing Abs used or the TGF-β inhibitor. b Representative flow cytometry for apoptosis of activated CD8 + T cells co-incubated with TEX1 in the presence of blocking Abs. c Inhibition of apoptosis by various neutralizing Abs of activated CD8 + T cells co-incubated with sEV from TNBC Pts (25 µg/mL). The pretreatment of TNBC-pts sEV with protein kinase (PK) or heat (HI) significantly reduced but did not eliminate T cell apoptosis. d The pretreatment of CD8 + T cells or activated primary CD4 + T cells with the mix of blocking Abs specific for PD1, TRAIL, Fas and anti-CTLA4 (used at the f.c. of 10, 10, 10 and 20 µg/mL, respectively) failed to reduce apoptosis induced by TEX 1 (50 µg/mL). In the blocking assays, primary human T cells were incubated with different blocking Abs and the TGF-β inhibitor. All Abs were used at the f.c. of 10 µg/mL, except for anti-CTLA4 Abs which were used at the f.c. of 20 µg/mL. TGF-β inhibitor was used at the f.c. of 50 nM. Blocking was performed for 30 min before co-incubation for 6 h with TEX or sEV from plasma of TNBC-Pts or HDs. Apoptosis was evaluated by Annexin-V binding assays. The data presented in (a) and (d) are means ± SD from 3 independent experiments.
Article Snippet: Human metastatic Triple Negative Breast Cancer Cell Lines (TNBC-CL) MDA-MB-231 (CL 1) and MDA-MB-436 (CL 2) as well as a non-malignant cell line, HaCaT (immortalized human keratinocytes), were obtained from ATCC including the authentication certificate (genomic profiling) and were cultured in Dulbecco’s modified Eagle’s medium (Gibco Fisher Scientific).
Techniques: Cytometry, Incubation, Blocking Assay, Inhibition, Clinical Proteomics, Binding Assay
ATCC is a verified supplier 
ATCC manufactures this product 
![Fig. 4 Immunoregulatory proteins expressed on the surface of recipient immune cells and on TEX or sEV of malignant and non-malignant origins. a The heatmap presenting mean RFI values for proteins expressed on the surface of activated primary CD4 + T, CD8 + T, B and NK cells. b The heatmap presenting mean RFI values for proteins found on the surface of TEX1, TEX2, HaCaT sEV, TNBC Pt-sEV and HD-sEV. c An image of a T cell interacting with various TEX (shown as blue vesicles) that carry immunoregulatory proteins on the surface membrane. TEX binding to complementary receptors expressed by the T cell initiate immunoregulatory signals which result in immune downregulation [(-)Ireg] and/or immune stimulation. The sum of these simultaneously delivered signals will determine whether TEX mediate immune suppression or immune stimulation in a recipient T cell. Note that a single sEV might carry multiple signaling proteins on its surface membrane.](https://pub-med-unpaywalled-images-cdn.bioz.com/pub_med_ids_ending_with_2121/pm37542121/pm37542121__page6_image1.jpg)
