Research On The Regulation Of BLyS Expression Levels Through MAPK Signaling Pathway In Multiple Myeloma

Objective B Lymphocyte stimulator (BLyS) promote the proliferation and survival of MM cells, involving in the occurrence and development of multiple myeloma (MM). The mitogen-activated protein kinase (MAPK) signaling pathway is considered to be a positive regulator of tumor initiation, progression, and maintenance, and the most characteristic function of MAPK is regulating the gene expression at transcriptional level. So the study is undertaken to discover the activation status of MAPK pathway, and identify whether there is interaction between MAPK signaling pathway and BLyS expression. Preliminarily study the role of MAPK signaling pathway in the up-regulation of BLyS expression levels induced by INF-γ.Methods We examined BLyS gene expression, localization and function in multiple myeloma cells (KM3and U266) and PBMCs of MM patients by RT-PCR, Western blot, fluorescence immunocytochemical method and WST-1cell proliferation assay. Western blot was used to verify the activation status of MAPK signaling pathway in MM cell lines and PBMCs of MM patients and healthy controls. The influence of MAPK signaling pathway on cell proliferation and apoptosis were also detected by WST-1cell proliferation assay and Western Blot. Western Blot analysis the impact of MAPK activator and inhibitor targeting JNK on BLyS expression levels. Building BLyS gene silenceing fragment (BLyS-SiRNA) to transfect MM cells. Western Blot detected the activation of MAPK signaling pathway after treated by BLyS-SiRNA or recombinant human BLyS (rhBLyS). Real time quantitative PCR (RFQ-PCR) detected the effect of MAPK signaling pathway inhibitor on the expression BLyS receptor BCMA in MM cells; The change of BLyS expression after stimulation by INF-y or co-stimulation by INF-y and MAPK inhibitor were also detected by Western Blot.Results BLyS gene was expressed in MM cell lines and PBMCs of MM patients. BLyS protein was located in the plasma-membrane of MM cells and promoted the proliferation of MM cells. In addition to the expression of total protein ERK, JNK, p38, activated protein p-JNK was also expressed in MM cell lines; MAPK pathway inhibitor targeting JNK SP600125restrained the proliferation and survival of MM cells, and promoted the apoptosis of MM cells. JNK signaling pathway inhibitor SP600125could down-regulate the expression of BLyS, and JNK pathway activator anisomycin could up-regulate the expression of BLyS. Recombinant BLyS promoted the expression of p-JNK, while BLyS SiRNA treatment restrained the expression of p-JNK. MAPK signaling pathway inhibitor could up-regulated the expression of BLyS receptor BCMA. The expression levels of p-JNK and BLyS were up-regulated by IFN-y in MM cells, and SP600125could partially offset the up-regulation of IFN-y.Conclusion JNK pathway was activated in MM cells, and its activation could promote the proliferation and inhibit the apoptosis of MM cells; The activated degree of JNK pathway and the expression level of BLyS was positively correlated; JNK signaling pathway play an important role in the up-regulation of BLyS expression levels induced by INF-γ; JNK activation and BLyS expression in MM cells may form a positive feedback loop that promote the survival and proliferation of MM cells.