Study On The Interaction Between Osteoblasts And Leukemia Cells In Leukemia Microenvironment

Objective:Osteoblast cells play an important role in bone marrow niche. The interaction between osteoblast and leukemia cells promotes the leukemogenesis. Such interaction includes the effect of osteoblast on the leukemia cells and the remodeling of osteoblast induced by leukemia cells. In order to identify the mechnism of the interaction between osteoblast and leukemia cells and further demonstrat the effect of abnormal leukemic niche on the leukemogenesis and the development of leukemia, we investigat the role of osteoblast in the leukemia cells’survival and apoptosis and the effect of leukemia cells on the osteoblast.Methods:Leukemia cells from AML1-ET09a-Racl and MLL-AF9 mouse leukemia models and the osteoblast cell lines MC3T3-E1 were used for our study. The ratio of GFP+ leukemia cells co-cultured with or without osteoblast for 4 days were detected with FACS, which represents the survival level. In addition, the apoptosis level of leukemia cells was detected by Flow Cytometry through PI and Annexin V labelling Real-Time-PCR was utilized to detect the mRNA level of TPO、N-cadherin、OPN、Angl in osteoblast which separated from leukemic mice. Finally, we detect the TPO protein level in the bone marrow plasma of AML patients through ELISA technology.Result:The GFP+ cells ratio of AE9a-Racl leukemia cells co-cultured with osteoblast cell lines MC3T3-E1 is higher significantly than that of AE9a-Racl leukemia cells cultured alone. The apoptotic level of AE9a-Racl leukemia cells cultured alone (79%±2.3%) are significant higher than that of AE9a-Racl leukemia in co-culture system (20%±1.9%). Western blot shows that activation level of apoptotic factor in AE9a-Racl leukemia cells co-culltured with MC3T3-E1 at 4 days is lower than control. We get the similar results using MLL-AF9 leukemia cells co-cultured with MC3T3-E1 cells. RT-PCR result shows that TPO、N-cadherin mRNA levels in primary osteooblast separated from leukemic mice is higher than that from normal mice. The Angl mRNA level of osteooblast from leukemia mice is lower. Finally, ELISA experiment shows that the TPO expression level in BM plasma of AML patients is higher than that in normal donors. Furthermore, The TPO expression level in AML patients with AML1-ETO/MLL-AF9 fusion genes are also significantly higher than that in normal donors.Coculsion:Osteoblast cell can support the survival and inhibit the apoptosis of leukemia cells. Leukemia cells can influence the functions of osteoblast on microenvironment associated cytokines production In addition, the TPO expression levels in bone marrow plasma of AML patients are higher than that in normal donors.