The role of cytokines and chemokines in the microenvironment of the blood-brain barrier in leukemia central nervous system metastasis

Abstract

Aim: Central nervous system (CNS) metastasis is a major obstacle in the treatment of leukemia, and the underlying mechanisms of leukemia CNS metastasis are not fully understood. The present study is an investigation of the role of the CNS microenvironment in leukemia CNS metastasis.

Methods: Analog blood-brain barrier (BBB) was set by coculturing human brain microvascular endothelial cells (HBMVECs) and leukemia cells (U937 and IL-60), as well as HBMVECs and sera from leukemia patients, in vitro. The permeability of the HBMVEC monolayer and the levels of tight junction proteins, cytokines and chemokines (C&Ckines) were measured.

Results: The permeability of HBMVECs increased when cocultured with leukemia sera. The expression of C&Ckines was significantly upregulated in HBMVECs cocultured with leukemia sera or leukemia cells, compared to the normal sera (P<0.05, respectively). Specifically, significantly higher levels of vascular endothelial growth factor A (VEGF-A) and matrix metalloprotease 9 (MMP-9) were found in HBMVECs and leukemia cells/sera coculturing systems.

Conclusion: Both leukemia cells and the molecules in leukemia sera play an important role in leukemia CNS metastasis. VEGF-A and MMPs may be the main factors resulting in the degradation of the BBB and inducing the CNS migration of leukemia cells.

Keywords: CNS leukemia; IL-60; U937; chemokine; cytokine.

Figure 1

Human brain microvascular endothelial cells cultured with different media. Note: Magnification: 400×.

Figure 2

Permeability of HBMVECs and concentration of cytokines in different cell culture media, after exposure to different factors, on different days. Notes: M, HBMVEC exposure to culture medium containing sera of AML patients; L, HBMVEC exposure to culture medium with sera of ALL patients; C, HBMVEC exposure to culture medium with sera of healthy controls; HL60, HBMVEC exposure to culture medium with HL60 cells; and U937, HBMVEC exposure to culture medium with U937 cells. Abbreviations: ALL, acute lymphoblastic leukemia; AML, acute myeloid leukemia; BSA, bovine serum albumin; HBMVECs, human brain microvascular endothelial cells.

Figure 2

Permeability of HBMVECs and concentration of cytokines in different cell culture media, after exposure to different factors, on different days. Notes: M, HBMVEC exposure to culture medium containing sera of AML patients; L, HBMVEC exposure to culture medium with sera of ALL patients; C, HBMVEC exposure to culture medium with sera of healthy controls; HL60, HBMVEC exposure to culture medium with HL60 cells; and U937, HBMVEC exposure to culture medium with U937 cells. Abbreviations: ALL, acute lymphoblastic leukemia; AML, acute myeloid leukemia; BSA, bovine serum albumin; HBMVECs, human brain microvascular endothelial cells.

Figure 3

ZO-1 expression in HBMVECs. Notes: (A) ZO-1 expression in HBMVECs before exposure to leukemia cells, as determined by immunofluorescence microscopy. (B) ZO-1 expression in HBMVECs after exposure to leukemia cells, as determined by immunofluorescence microscopy. (C) ZO-1 expression in HBMVECs before exposure to leukemia cells, as determined by immunohistochemistry. (D) ZO-1 expression in HBMVECs after exposure to leukemia cells, as determined by immunohistochemistry. ZO-1 immunoreactivity (red staining) was restricted to junctional areas and some punctate staining in the cytoplasm (arrows). Exposure to leukemia cells resulted in a weaker junctional and cytoplasmic pattern of ZO-1 immunoreactivity. DAPI (blue staining) was used to visualize the nuclei. Magnification: 1000×. Abbreviations: HBMVECs, human brain microvascular endothelial cells; ZO-1, Zonula occludens-1; DAPI, 4′,6-diamidino-2-phenylindole.