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• W4385667099 abstract "Background: Bidirectional interactions exist between Chronic Lymphocytic Leukemia (CLL) cells and the cells of their microenvironment (ME): CLL cells indeed received several stimuli such as BCR stimulation but they can also interact with the surrounding cells by the release of extracellular vesicles (EVs). EVs are small double membrane particles (from 100nm to 1µm) carrying proteins, DNA and several types of RNA such as microRNAs. EV exchange is today considered as a new way of cell communication. Aims: We hypothesize that BCR stimulation could alter EV content and that CLL-EVs could manipulate immune ME cells such as monocytes and stimulate their differentiation into nurse-like cells (NLC), well known in CLL to support leukemic cell survival. Methods: The microRNA profile of CLL-EVs isolated by ultracentrifugation from leukemic cells cultured with/without BCR stimulation (n=4) was performed by NGS and validated by qPCR (n=25). CLL-EVs integration was monitored using PKH67 and Memglow dyes. After treating purified monocytes with CLL-EVs (Mono+CLL-EVs), the morphology, the cytokine profile, specific microRNA and mRNA expression and their ability to support CLL cell survival were analyzed and compared to those of NLC. Results: NGS highlighted 31 microRNAs differentially expressed between BCR stimulated/unstimulated conditions and qPCR confirmed the increase of 4 of them (n=25): miR-146a-5p (fold: +15.1, p<0.0001), miR-132-3p (fold: +42.1, p<0.0001), miR-155-5p (fold: +3.3, p=0.003) and miR-17-3p (fold: +3.1, p=0.0135) in EVs. While the expression of miR-150 was not affected in CLL EVs after BCR stimulation, miR-150-5p was the most abundant microRNA in CLL EVs. After 24h of incubation of CLL PBMC with CLL-EVs, 76% of monocytes were PKH67+ indicating they were the first cell type to integrate CLL-EVs while they only represent 3.5% of all PBMC (P=0.0019, n=6). Although, Mono+CLL-EVs display a fibroblast morphology similar to M2 macrophages. After 24h of incubation with CLL-EVs, miR-155-5p, miR-146a-5p, miR-132-3p were increased in monocytes (fold: +4.8, +4.5; +5.6, respectively; P<0.05, n = 9) similarly to what happens in the differentiation of monocytes in NLC (fold: +5.6, +27.4, +12, respectively, P<0.01, n=10). Moreover, target of these microRNAs such as S1PR3 or KLF4 were decreased in Mono+CLL-EVs, in NLC and in monocytes transfected with miRNAs mimic (Mono+mimic) (P<0.05, n=11). In addition, genes that were increased (MMP9, SPP1) or decreased (CCR2) after NLC differentiation display a similar trend in Mono+CLL-EVs and in Mono+mimic (n=10, P<0.05). Multiplex immunoassay analysis (n=3) highlighted the increase of IL-10 (fold: +18.6), IL-8 (+6.1), IL-6 (+488.4), BAFF (+18.6), CCL2 (+4.9) and SDF-1α (+13.9) in Mono+CLL-EVs produced in BCR stimulated conditions. These cytokines have already been described as increased in NLC or supporting CLL cell survival. Finally, CLL cells incubated with Mono+CLL-EVs showed a decreased apoptosis compared to co-culture with untreated monocytes (decrease of 18%±3.1, P=0.0156, n=6) highlighting the protective effect of these EV-transformed monocytes. Summary/Conclusion We showed that BCR stimulation modifies CLL-EVs microRNA content. The addition of EVs on monocytes lead to a shape change, an increase of several microRNAs, a decreased expression of some of their targets, an increase of several cytokines and an increased ability to rescue CLL cells from spontaneous apoptosis. All together, our data indicate that CLL-EVs induce the differentiation of monocytes into NLC supporting CLL cell survival. Keywords: Microvesicles, Microenvironment, B cell chronic lymphocytic leukemia, Monocyte" @default.
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• W4385667099 date "2023-08-01" @default.
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• W4385667099 title "P610: EXTRACELLULAR VESICLES FROM CHRONIC LYMPHOCYTIC LEUKEMIA CELLS INDUCE THE DIFFERENTIATION OF MONOCYTES INTO NURSE-LIKE CELLS" @default.
• W4385667099 doi "https://doi.org/10.1097/01.hs9.0000969344.41762.57" @default.
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