The ceramide pathway is involved in the survival, apoptosis and exosome functions of human multiple myeloma cells in vitro

Abstract

Multiple myeloma (MM) is characterized by the clonal proliferation of malignant plasma cells and refractoriness to traditional therapies. It has been shown that exosomes are involved in modulating the progression and the metastasis of cancers through microRNAs (miRs). Ceramide is a type of sphingolipid; the ceramide pathway of exosomal secretion has been shown to affect the apoptosis of cancer cells. But the role of this pathway in MM cell function, exosome function and miR regulation remains unknown. In this study, we showed that C6 ceramide (an exogenous ceramide supplement, 1.25-40 μmol/L) dose-dependently inhibited the proliferation and promoted the apoptosis in human MM OPM2 cell line, which were associated with elevated caspase 3/9 and PARP cleavage. We also found that C6 ceramide (5-20 μmol/L) dose-dependently stimulated exosome secretion and increased exosomal levels of tumor-suppressive miRs (miR 202, miR 16, miR 29b and miR 15a). Of note, exosomes from C6 ceramide-treated OPM2 cells could influence the proliferation and apoptosis of the recipient OPM2 cells, which correlated with increased tumor-suppressive exosomal miRs. In contrast, GW4869 (a ceramide inhibitor, 5-20 μmol/L) exerted the opposite effects on the regulation of MM function, exosome secretion and miR levels in MM exosomes. However, exosomes from GW4869-treated OPM2 cells had no effect on these miRs and the survival of targeted OPM2 cells. Taken together, our findings reveal that the ceramide pathway modulates MM survival, probably directly via the caspase pathway and indirectly via exosomal miR mechanisms.

C6-ceramide treatment inhibits the proangiogenic activity of multiple myeloma exosomes via the miR-29b/Akt pathway

“…Particle size and concentration analysis. The analyses of size and concentration were in consistent with our previous studies [15,20] . In details, exosomes were resuspended in PBS, the particle size and concentration were measured by using the Nano Tracking System 300 (NTA300, Malvern Instruments, UK).…”

“…C6 ceramide (C6-cer) is a short-chain, cell-permeable analog of ceramide that has been utilized in preclinical studies to mimic the effects of endogenous ceramide [16] . We have previously showed that C6-cer treated MM exosomes could inhibit the proliferation of MM cells, indicating the antitumor role of C6-cer by exosomes [15] . However, whether C6-cer treated MM cell-derived exosomes could affect angiogenesis remains largely unknown.…”

“…It has been found that miR-29b is highly associated with the proliferation and survival of MM cells [17] . C6-cer has been shown to increase the exosomal levels of some tumor-suppressive miRs (miR-29b, miR-202, and miR-15a/16) [15] . Furthermore, it has been shown that miR-29b can be used as high effective anti-cancer therapeutic agent by simultaneously exert the effects including both anti-angiogenesis and antitumorigenesis via regulating the Akt pathway [19] .…”

Role of sphingolipids in the biogenesis and biological activity of extracellular vesicles

“…As in A, before n-SMase action SM stabilizes membrane structure, while Cer production by n-SMase induces spontaneous negative curvature to the membrane of endosomes, thus promoting the formation of internal vesicles inside MVBs. Subsequent in vitro works reported that, while exogenous cell permeable C6 Cer dose-dependently increases the number of exosomes released from multiple myeloma cells (51), pharmacological or genetic block of n-SMase inhibited packaging of the prion protein into exosomes (52) and reduced exosome release among others in HEK cells (53), T cells (54), N2a cells (55), astrocytes (56), microglia (57), macrophages (58), hepatocytes (59), and multiple myeloma cells (51). Also, in vivo, in the brain and serum of 5XFAD mice, a transgenic model of Alzheimer's disease (AD), the n-SMase inhibitor, GW4869, decreased exosome concentration (60).…”

Potential Therapeutic Roles of Exosomes in Multiple Myeloma: A Systematic Review

“…Apart from miRNAs and proteins, there are other exosome-associated treatment methods for MM [ Table 1]. For instance, C6 ceramide, an exogenous ceramide supplement, was found to dosedependently stimulate exosome secretion and elevate the exosome levels of some tumor-suppressive miRNAs, such as miRNA-202, miRNA-16, miRNA-29b, and miRNA-15a, thus increasing the apoptosis and inhibiting the proliferation of recipient MM cells, indicating that it is a target for MM treatment [32]. Exosomes also show anti-tumor immune effects.…”