Abstract
BackgroundThe canonical Wnt signaling is concurrently important for osteoblast differentiation and myeloma cell proliferation. Its activation in myeloma cells and its inhibition in osteoblasts and their progenitors have been identified in the previous studies. Osteoblast progenitors and myeloma cells from a myeloma patient share the same bone marrow (BM) microenvironment, but respond differently to DKK-1 secreted by myeloma cells. The mechanisms remain unclear.MethodsPrimary multiple myeloma (MM) cells were isolated from BM mononuclear cells of 12 MM patients. Human bone marrow stromal cells (SCs) were obtained from BM adherent cells of these MM patients and 10 healthy donors. The mRNA expression levels of DKK-1 binding receptor LRP5/6 and Kremen1/2 (Krm1/2) were analyzed by Real-time PCR in human myeloma cell line (HMCL) RPMI-8226, NCI-H929, U266, LP-1, CZ-1, KM-3, Sko-007, primary myeloma cells and SCs from 12 MM patients and SCs from 10 healthy donors. The binding capability of DKK-1 binding receptors to DKK-1 on primary myeloma cells and SCs was detected by flow cytometry assay.ResultsThe mRNA expression levels of DKK-1 binding receptor LRP5/6 and Krm1/2 in SCs from patients with MM were significantly higher than those in myeloma cells and in SCs from healthy donors. The binding capability to DKK-1of DKK-1 binding receptors on SCs from MM patients was obviously higher than those on myeloma cells and SCs from healthy donors by flow cytometry assay. Similar to the effects of coculture with rhDKK1, coculture of SCs from healthy donors with myeloma cells in the presence or absence of a Transwell insert did up-regulate SCs' mRNA levels of LRP5/6 and Krm1/2, and down-regulate their mRNA levels of β-catenin.ConclusionCompared with myeloma cells, the SCs from MM patients overexpress DKK-1 binding receptors LRP5/6 and Krm1/2 in response to DKK-1 secreted by myeloma cells, which results in intracellular Wnt signaling inhibition. Our study provides a novel insight into mechanisms of myeloma associated osteolytic lesions.
Highlights
The canonical Wnt signaling is concurrently important for osteoblast differentiation and myeloma cell proliferation
Expression of DKK-1 protein by human MM cells We examined the expression of DKK-1 protein in 6 human myeloma cell line (HMCL), and 5 freshly purified primary CD138+ myeloma cells and 4 stromal cells (SCs) from patients with MM by western blotting
Influences of MM cells on the mRNA expression of lipoprotein receptor-related protein 5/6 (LRP5/6), Krm1/2 and b-catenin in SCs from healthy donors in coculture The mRNA expressions of LRP5/6, Krm1/2 and b-catenin were detected in SCs from healthy donors after 24-72 h of coculture with RPMI-8226 or NCI-H929 in the presence or absence of a Transwell insert
Summary
The canonical Wnt signaling is concurrently important for osteoblast differentiation and myeloma cell proliferation. Its activation in myeloma cells and its inhibition in osteoblasts and their progenitors have been identified in the previous studies. Osteoblast progenitors and myeloma cells from a myeloma patient share the same bone marrow (BM) microenvironment, but respond differently to DKK-1 secreted by myeloma cells. Multiple myeloma (MM) is one of hematological malignancies characterized by bone marrow (BM) infiltration of monoclonal plasma cells and progression of osteolytic bone lesions [1]. The deficit of bone formation is attributable to osteoblastic dysfunction resulting from the proliferation of myeloma cells in the BM. The mechanisms by which myeloma cells affect the formation and function of osteoblasts are still under investigation. Increasing evidences suggest that Wnt/b-catenin signaling activation has been linked to many human malignancies, including hematologic malignancies. In comparison with normal B-cell populations, MM cells express high levels of b-catenin, including the stabilized unphosphorylated form which functions in activating T-cell factor/lymphoid enhancer factor (TCF/LEF)-mediated transcription of target genes, stimulating the proliferation of MM cells [7]
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