A Promoting Effect Of Spleen On Disseminated Leukemia Cells

Abnormal environment not only takes part in the malignant transformation of hematopoietic cells but also has great impact on the development of hematopoietic malignancies. Leukemia cells arise from a specific environment and compete with normal cells from the beginning of leukemogenesis. Then they induce leukemic environment favorable for the outgrowth of themselves. Both bone marrow (BM) and spleen are involved and other organs are also infiltrated by leukemia cells at late stage. Splenomegaly is more frequently observed in T cell acute lymphoblastic leukemia (T-ALL), and is related to the poor prognosis of patients. However, the mechanism is not clear. In this study, we used the non-radiated Notch1-induced mouse T-ALL model to study the effects of spleen and BM environment on the early residing, malignant phenotype and dissemination of leukemia cells.

The gross change of organs during the development of leukemia was analyzed and the earliest, sustained and a final 12.25±0.6802-time increase in weight could be observed in spleen. Though leukemia cells could be detected in all organs analyzed at late stage, the earliest and sustained increase was detected in spleen but not in BM. Furthermore, two-photon fluorescence microscopy analysis showed that more leukemia cells were found in spleens but not in BM within 3 days post-transplantation. These results suggested that spleen may be more important for the initiation and early development of Notch1 induced T-ALL than any other organs. In vitro transwell experiment showed that more leukemia cells migrated to lower chambers with normal spleen cells than with normal BM cells. The concentration of a panel of cytokines/chemokines in BM, spleen and peripheral blood was screened pre-transplantation or within 3 days post-transplantation. Notably, the concentration of MIP-3β was higher in spleen sample at day 0, and it increased rapidly and kept at high levels within 3 days. Further evidence showed that leukemia cells expressed basal level of CCR7 and MIP-3β, and higher level of them could be observed when leukemia cells were co-cultured with spleen cells but not BM cells. Moreover, transwell experiment confirmed that MIP-3β promoted the migration of leukemia cells. Then we study the effects of spleen environment on leukemia cells. Spleen cells were more potent to stimulate the proliferation of leukemia cells than BM cells. Leukemia cells co-cultured with spleen cells were more potent to migrate in the in vitro migration assay than those co-cultured with BM cells. Leukemia cells freshly isolated from spleen expressed higher level of both MIP-3β and CCR7. When same amount of leukemia cells were injected to mice, spleen-origin leukemia cells caused shorter life span than BM-origin leukemia cells. Therefore, spleen environment promotes the malignant phenotype of leukemia cells. In vivo experiments were then carried out to see whether splenectomy had effect on the survival of leukemia mice. The survival time of mice in either group (splenectomy pre- or post-injection of leukemia cells) was significantly longer than that in the sham group. Furthermore, distinct effects could be observed in mice groups underwent splenectomy at both early stage and late stage post-injection of leukemia cells. Moreover, less severe invasion and destruction of organ structures in thymus, lymph nodes and BM could be observed in two splenectomy groups.

In summary, our work demonstrates that spleen has a promoting effect on disseminated leukemia cells. The highly expressed MIP-3β in the spleen environment recruits leukemia cells. Subsequently, spleen environment promotes the proliferation of leukemia cells, stimulates the expression of CCR7, and enhances the migration ability of leukemia cells. The removal of spleen could prolong the survival of leukemia mice. Therefore, this study provides evidence for an organ specific effect on emerging leukemia cells in vivo and has implications for developing new treatments for leukemia patients.