Abstract
The inherent abilities of natural killer (NK) cells to recognize and kill target cells place them among the first cells with the ability to recognize and destroy infected or transformed cells. Cancer cells, however, have mechanisms by which they can inhibit the surveillance and cytotoxic abilities of NK cells with one believed mechanism for this: their ability to release exosomes. Exosomes are vesicles that are found in abundance in the tumor microenvironment that can modulate intercellular communication and thus enhance tumor malignancy. Recently, our lab has found cancer cell exosomes to contain the inhibitor of apoptosis (IAP) protein survivin to be associated with decreased immune response in lymphocytes and cellular death. The purpose of this study was to explore the effect of survivin and lymphoma-derived survivin-containing exosomes on the immune functions of NK cells. NK cells were obtained from the peripheral blood of healthy donors and treated with pure survivin protein or exosomes from two lymphoma cell lines, DLCL2 and FSCCL. RNA was isolated from NK cell samples for measurement by PCR, and intracellular flow cytometry was used to determine protein expression. Degranulation capacity, cytotoxicity, and natural killer group 2D receptor (NKG2D) levels were also assessed. Lymphoma exosomes were examined for size and protein content. This study established that these lymphoma exosomes contained survivin and FasL but were negative for MHC class I-related chains (MIC)/B (MICA/B) and TGF-β. Treatment with exosomes did not significantly alter NK cell functionality, but extracellular survivin was seen to decrease natural killer group 2D receptor (NKG2D) levels and the intracellular protein levels of perforin, granzyme B, TNF-α, and IFN-γ.
Highlights
We evaluated the lymphoma exosomes for several protein markers using flow cytometry and Western blotting
As natural killer (NK) cells are frequently found to be inhibited in the tumor microenvironment (TME), we investigated whether extracellular survivin or lymphoma exosomes containing survivin would modify NK cell function
We found that treatment of NK cells with lymphoma exosomes or extracellular survivin for 24 h did not result in consistent alterations to levels of RNA by either block RT-Polymerase Chain Reaction (PCR) (Figure 6A) or real time qRT-PCR (Figure 6C)
Summary
The active role of the tumor microenvironment (TME) in tumor pathogenesis has recently been gaining intense scrutiny as it can be responsible for delivering signals for clonal expansion [1], drug resistance [2], metastatic migration [3], and immune modulation [4]. Cancer development is not an autonomous process but depends on surrounding nonmalignant components such as fibroblasts, microvasculature, stroma, mesenchymal cells, and immune cells [5,6]. Interaction between tumors and their surrounding environment occurs via direct contact, soluble factors, and material exchange [7]. One important method of cell-to-cell interaction that has received much interest in the preceding decade for communication and signaling is through information transfer between cells via small bilayer lipid vesicles known as exosomes, or more generally, as extracellular vesicles (EVs)
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