Multiple myeloma cells alter the senescence phenotype of bone marrow mesenchymal stromal cells under participation of the DLK1-DIO3 genomic region

Abstract

BackgroundAlterations and senescence in bone marrow mesenchymal stromal cells of multiple myeloma patients (MM-BMMSCs) have become an important research focus. However the role of senescence in the pathophysiology of MM is not clear.MethodsCorrelation between senescence, cell cycle and microRNA expression of MM-BMMSCs (n = 89) was analyzed. Gene expression analysis, copy number analysis and methylation specific PCR were performed by Real-Time PCR. Furthermore, cyclin E1, cyclin D1, p16 and p21 genes were analyzed at the protein level using ELISA. Cell cycle and senescence were analyzed by FACS. MiRNA transfection was performed with miR-485-5p inhibitor and mimic followed by downstream analysis of senescence and cell cycle characteristics of MM-BMMSCs. Results were analyzed by Mann–Whitney U test, Wilcoxon signed-rank test and paired t-test depending on the experimental set up.ResultsMM-BMMSCs displayed increased senescence associated β-galactosidase activity (SA-βGalA), cell cycle arrest in S phase and overexpression of microRNAs. The overexpressed microRNAs miR-485-5p and miR-519d are located on DLK1-DIO3 and C19MC, respectively. Analyses revealed copy number accumulation and hypomethylation of both clusters. KMS12-PE myeloma cells decreased SA-βGalA and influenced cell cycle characteristics of MM-BMMSCs. MiR-485-5p was significantly decreased in co-cultured MM-BMMSCs in connection with an increased methylation of DLK1-DIO3. Modification of miR-485-5p levels using microRNA mimic or inhibitor altered senescence and cell cycle characteristics of MM-BMMSCs.ConclusionsHere, we show for the first time that MM-BMMSCs have aberrant methylation and copy number of the DLK1-DIO3 and C19MC genomic region. Furthermore, this is the first study pointing that multiple myeloma cells in vitro reduce both the senescence phenotype of MM-BMMSCs and the expression of miR-223 and miR-485-5p. Thus, it is questionable whether senescence of MM-BMMSCs plays a pathological role in active multiple myeloma or is more important when cell interaction with myeloma cells is inhibited. Furthermore, we found that MiR-485-5p, which is located on the DLK1-DIO3 cluster, seems to participate in the regulation of senescence status and cell cycle characteristics of MM-BMMSCs. Thus, further exploration of the microRNAs of DLK1-DIO3 could provide further insights into the origin of the senescence state and its reversal in MM-BMMSCs.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1078-3) contains supplementary material, which is available to authorized users.

Multiple myeloma cells promote migration of bone marrow mesenchymal stem cells by altering their translation initiation

“…Previous studies have shown MIR-199b-3p and MIR-125a-3p are major negative regulators of cell migration in cancer models [23,24]. Further studies are warranted to elucidate the function of the microRNAs in the signaling cascade we present and to prove their regulatory role in our model (16). This is compatible with our observation of increased MMcond-MSCs migration.…”

“…Cells were lysed, proteins were extracted, and the Western blot test was preformed, as described elsewhere [16]. The following proteins were detected using rabbit/mouse anti-human: peIF4E (Ser-209)/total eIF4E, peIF4GI(Ser-1108)/total eIF4GI, p4EBP(Ser-65)/total 4EBP, p-mTOR(Ser-2448)/total mTOR, p-MNK (Thr-197/Thr-202)/total MNK (Cell Signaling Technology, Danvers, MA, USA); and tubulin (Sigma-Aldrich).…”

The function of miR-519d in cell migration, invasion, and proliferation suggests a role in early placentation

“…In a study of cervical cancer, miR-519d was found to be significantly upregulated in cervical cancer samples, where it promoted the migration and invasion of HeLa and SiHa cervical cancer cells and the proliferation of cervical cancer cells through targeting Smad 7 [33]. miR-519d is also overexpressed in bone marrow mesenchymal stromal cells of multiple myeloma patients compared to bone marrow mesenchymal stromal cells of donors [34]. …”

Multiple myeloma cells modify VEGF/IL‐6 levels and osteogenic potential of bone marrow stromal cells via Notch/miR‐223

“…In contrast, K562 cells did not induce a significant down‐regulation of miR‐223 possibly accounting for a specific impact of MM cells on MM‐BMMSCs. The counteracting of MM cells on aberrations in MM‐BMMSCs compared to HD‐BMMSCs indicates the excitatory effect of MM cells on MM‐BMMSCs . Overexpression of miR‐223 in MM‐BMMSCs compared to HD‐BMMSCs could result from a senescence‐like state induced by increased activation of stromal cells by MM cells.…”