Project description:Bi-directional communication with the microenvironment is essential for homing and survival of cancer cells with implications for disease biology and behaviour. In chronic lymphocytic leukemia (CLL) the role of the microenvironment on malignant cell behaviour is well described. However, how CLL cells engage and recruit nurturing cells is poorly characterised. Here we demonstrate that CLL cells secrete exosomes that are nanovesicles originating from the fusion of multivesicular bodies with the plasma membrane, to shuttle proteins, lipids, microRNAs (miR) and mRNAs to recipient cells. We characterise and confirm the size (50 - 100 nm) and identity of the CLL-derived exosomes by Electron microscopy (EM), Atomic force microscopy (AFM), flow cytometry and western blotting using both exosomes-specific and CLL-specific markers. Incubation of CLL-exosomes, derived either from cell culture supernatants or from patient plasma, with human stromal cells shows that they are readily taken up into endosomes, and induce expression of genes such as c-fos and ATM as well as enhance proliferation of recipient HS-5 cells. Furthermore, we show that CLL exosomes encapsulate abundant small RNAs and are enriched in certain miRs and specifically hsa-miR-202-3p. We suggest that such specific packaging of miR-202-3p impacts on the expression of 'suppressor of fused' (Sufu), a Hedgehog (Hh) signalling intermediate, in the parental CLL cells. Thus, our data show that CLL cells secrete exosomes that alter the transcriptome and behaviour of recipient cells. Such communication with microenvironment is likely to have an important role in CLL disease biology. miR analysis was carried out on 3 exosomal samples and 3 corresponding cellular samples from CLL patients. Exosomes are a discrete population of small (50-100 nm diameter) EVs of endosomal origin with a lipid membrane bilayer and a cup-shaped morphology. We used common reference design which allows visualization of variations between the samples. Each sample equals one array and each sample compared with common reference (Pool). The pool or common reference was a collection of 23 RNA samples isolated from 23 CLL cases. We hypothesised that CLL derived exosomes should contain unique miRs that are reflective of CLL cell content and additionally relevant for disease biology.

			Action	DRS
	Dugez-Exo-SG-ALS1-SD990-01.raw	Raw
	Dugez-Exo-SG-ALS1-SD990-02.raw	Raw
	Dugez-Exo-SG-ALS1-SD990-03.raw	Raw
	Dugez-Exo-SG-ALS1-SD990-04.raw	Raw
	Dugez-Exo-SG-ALS1-SD990-05.raw	Raw

Dataset Information

Proteomic Analysis of Myotube-Secreted Exosomes of Amyotrophic Lateral Sclerosis (ALS) and Healthy subjects

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Optimized method for extraction of exosomes from human primary muscle cells.

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