Selective in vivo growth of lymphocyte function- associated antigen-1-positive murine myeloma cells. Involvement of function-associated antigen-1-mediated homotypic cell-cell adhesion.

Abstract

Lymphocyte function-associated antigen-1 (LFA-1) expression on multiple myeloma cells and its potential role in myeloma biology have been the subject of conflicting literature reports. In this study we used the 5T experimental mouse model to analyze the involvement of LFA-1 in myeloma cell bone marrow homing, survival, and growth. The 5T33MM vitro (5T33MMvt) myeloma line was used. LFA-1 and intercellular adhesion molecule-1 (ICAM-1) expression were analyzed by flow cytometry. A small molecule antagonist of LFA-1/ICAM interactions, BIRT 377, was used to block LFA-1 in vitro. Transendothelial migration was assessed by measuring migration through Transwells coated with bone marrow endothelial cells. Immediate in vivo homing was analyzed by tracing 51Cr-labeled cells. Invert microscopic cell counting was used to analyze homotypic cell adhesion. Cell cycle analysis was used to analyze apoptosis. S+G(2)/M phase analysis and 3H-thymidine incorporation were used to assess proliferation. Cells were separated into LFA-1(+) and LFA-1(-) fraction by magnetic activated cell sorting. 5T33MMvt cells had a heterogeneous LFA-1 expression and all cells were positive for the LFA-1 ligand ICAM-1. LFA-1 inhibition with BIRT 377 did not affect transendothelial migration of the 5T33MMvt cells; however, it did result in cell cluster scattering, indicating LFA-1 involvement in homotypic cell-cell adhesion. No effect was observed on apoptosis, but the percentage of cells in S+G(2)/M phase was decreased by 39%. 3H-thymidine incorporation confirmed this effect on 5T33MMvt cell proliferation (38% reduction). When 5T33MMvt cells were injected into animals, all myeloma cells isolated at the end stage of the disease were LFA-1(+) in contrast to the situation before injection. LFA-1(+) and LFA-1(-) MM cells had similar in vivo bone marrow homing capacities. Mice injected with LFA-1(+) 5T33MMvt cells developed myeloma (5/5) within 12 weeks after injection. In contrast, LFA-1(-) recipients did not develop the disease (0/5), even 1 year after tumor inoculation. Our data suggest that LFA-1-mediated homotypic cell-cell adhesion is involved in myeloma cell proliferation and raises the possibility that this interaction may have a crucial role in in vivo myeloma cell growth. LFA-1 does not appear to play a role in the bone marrow homing of these cells.