Overexpression of Nrf2 in Mesenchymal Stem Cells Promotes Migration and Invasion of B- Acute Lymphoblastic Leukemia Cells

Background and objective:B -Acute lymphoblastic leukemia (B-ALL) is one of the most difficult cancers to treat in adults, especially recurrence and drug resistance are still the biggest challenges for B-ALL. In recent years, a new concept has been proposed, that is, the interaction between bone marrow microenvironment and leukemic cells can reduce the sensitivity of leukemic cells to chemotherapy. The components of bone marrow microenvironment (BMM) have been shown to mediate the occurrence and development of leukemia. There is growing evidence that leukemic cells teach the bone marrow microenvironment, change its function and establish leukemia protective interactions with stromal cells and immune cells. Bone marrow mesenchymal stem cells (Bone marrow mesenchymal stem cells, BM-MSCs) are stromal cells in leukemia microenvironment, which can promote the progression of leukemia under certain conditions. Nuclear factor erythroid related factor 2 (Nrf2) is an important transcription factor and a regulator of cellular antioxidant response, which regulates the expression of genes encoding antioxidant enzymes to prevent various oxidative changes. It has been confirmed that the high expression of Nrf2 in leukemic cells is related to drug resistance and poor prognosis of leukemia. However, the interaction mechanism of Nrf2 overexpression in MSCs on B-cell acute lymphoblastic leukemia (B-ALL) cells is not clear. We will for the first time propose the mechanism of Nrf2 overexpression in MSCs on the migration and invasion of B-ALL leukemic cells.

Methods :Bone marrow nucleated cells were detected in 16 patients with complete remission (B-ALL/CR), 12 patients with relapsed/refractory (B-ALL/R/R) ALL and 10 normal controls. The expression of Nrf2 was detected byreal-time PCR and Western Blot. In vitro cell experiments, real-time PCR and Western Blot were used to detect the expression of Nrf2 in mesenchymal stem cells (MSCs) alone and in the co-culture of MSCs and cell lines (Nalm-6/RS4:11) express the situation Mouse model experiment in vivo, transwell chamber experiment in vitro, three-dimensional (3D) cell culture model, lentivirus transfection, WB and PCR were used to explore the possible mechanism of Nrf2 overexpression in mesenchymal stem cells on the migration and invasion of B-ALL cells.

Results: In clinical samples, the expression of Nrf2 in B-ALL-R/R group was significantly higher than that in ALL-CR group and normal control group, suggesting that the expression of Nrf2 was closely related to the recurrence and drug resistance of ALL. In vitro cell experiment, the expression level of Nrf2 in MSCs co-cultured with cell line (NALM-6/RS4:11) was higher than that in MSC cultured alone(P < 0.05). By regulating the overexpression of Nrf2 in MSCs, it can significantly promote the growth, proliferation, migration and invasion of leukemia cells, and promote the expression of CXCR4 in tumor tissues. We further regulated the expression of Nrf2 in MSCs. The results showed that it significantly promoted the migration and invasion of NALM-6/RS4:11 cells. We concluded that the overexpression of Nrf2 in MSCs promotes the migration and invasion of leukemic cells by activating the activity of SDF-1-CXCR4 axis. In vivo experiments, we used tail vein injection of MSCs and B-ALL leukemia cells to explore the protective effect of Nrf2 overexpression in mesenchymal stem cells on leukemia cells and promote extramedullary infiltration of leukemia. The results showed that the infiltration degree of extramedullary organs of mice after combined infusion of two kinds of cells was significantly higher than that of leukemia cells alone, and the survival time of mice was significantly shortened after combined infusion. After statistical treatment, the difference was statistically significant (P < 0.05).

Conclusion:The expression of Nrf2 in MSCs may be a key component in mediating the interaction between MSCs and ALL cells. This result may provide a new entry point for deepening all disease progression mechanisms and finding more promising targeted therapies.On the other hand, activation of PI3K/AKTERK1/2 signaling pathway in B-ALL cells further promoted the evolution of B-ALL. This may provide new ideas for findingpotential therapeutic targets for B-ALL extramedullary infi ltration, and help to further solve the current problem of B-ALL relapse and refractory.