Selective in vivo growth of lymphocyte function- associated antigen-1–positive murine myeloma cells  Kewal Asosingh, Virginie Vankerkhove, Ivan Van Riet, Ben Van Camp, Karin Vanderkerken  Experimental Hematology  Volume 31, Issue 1, Pages 48-55 (January 2003) DOI: 10.1016/S0301-472X(02)00970-0

Figure 1 LFA-1 and ICAM-1 expression on 5T33MMvt cells. (A) Expression of LFA-1 α chain (CD11a). (B) Expression of ICAM-1. Filled histograms illustrate levels of background staining. Histograms of one experiment, representative for three independent stainings, are illustrated. Experimental Hematology 2003 31, 48-55DOI: (10.1016/S0301-472X(02)00970-0)

Figure 2 Involvement of LFA-1 in the homotypic 5T33MMvt cell-cell adhesion. Single cell suspensions of 5T3MMvt cells were incubated with BIRT 377 or 1% DMSO. Cell-cell cluster formation was analyzed after 18 hours by invert microscopic cell counting. Mean ± SD values of three independent experiments are shown. Experimental Hematology 2003 31, 48-55DOI: (10.1016/S0301-472X(02)00970-0)

Figure 3 Involvement of LFA-1–mediated homotypic cell-cell adhesion in cell cycle progression of 5T33MMvt cells. Cell cycle analysis was performed by staining the nuclei with PI for flow cytometry. Mean ± SD values of three independent analyses are shown. Histograms of one experiment, representative for three, are illustrated. Experimental Hematology 2003 31, 48-55DOI: (10.1016/S0301-472X(02)00970-0)

Figure 4 Involvement of LFA-1–mediated homotypic cell-cell adhesion in 3H-thymidine incorporation. 5T33MMvt cells were incubated in different conditions, as indicated in the figure and pulsed with 3H-thymidine for 18 hours. Data are illustrated as counts per minutes (CMP). Mean ± SD values of quadruplets are shown. This experiment is representative of three independent experiments. Experimental Hematology 2003 31, 48-55DOI: (10.1016/S0301-472X(02)00970-0)

Figure 5 LFA-1 expression on 5T33MMvt-vv cells. 5T33MMvt cells were injected into naive mice, and after tumor take, myeloma cells isolated the BM (5T33MMvt-vv) were analyzed for LFA-1 expression by double staining with LFA-1 α chain (CD11a) and 5T33MM antidiotype antibodies. Upper panel illustrated CD11a expression on 5T33MMvt cells before injection. Lower panel illustrates the expression after in vivo inoculation. Idiotype negative cells are normal hematopoietic cells as revealed by microscopic examination of the cells after May Grunwald Giemsa staining. Data from one animal of five are shown. Experimental Hematology 2003 31, 48-55DOI: (10.1016/S0301-472X(02)00970-0)

Figure 6 Homing kinetics of LFA-1+ and LFA-1− 5T33MMvt cells. The homing of LFA-1+ and LFA-1− 5T33MMvt cells was analyzed by in vivo tracing of 51Cr labeled cells, 18 hours after intravenous injection. Data are expressed as percentage of total radioactivity injected. BM represents the sum of radioactivity recovered from the ribs, vertebrae, fore and hind legs. Mean ± SD values of four mice per group are shown. Experimental Hematology 2003 31, 48-55DOI: (10.1016/S0301-472X(02)00970-0)

Figure 7 In vivo growth of LFA-1+ and LFA-1− 5T33MMvt cells. (A) 5T33MMvt cells were sorted into LFA-1+ and LFA-1− fractions and injected intravenously into naive mice. All LFA-1+ recipients developed myeloma within 12 weeks; in contrast, all LFA-1− remained healthy, even 72 weeks after tumor inoculation. (B) BM isolated from mice injected with LFA-1+ 5T33MMvt cells was positive for 5T33MM idiotype and CD11a. (C) BM from LFA-1− recipients stained with 5T33MM idiotype and was negative for tumor cells. 5T33MMvt were stained in parallel as positive control. Experimental Hematology 2003 31, 48-55DOI: (10.1016/S0301-472X(02)00970-0)

Figure 7 In vivo growth of LFA-1+ and LFA-1− 5T33MMvt cells. (A) 5T33MMvt cells were sorted into LFA-1+ and LFA-1− fractions and injected intravenously into naive mice. All LFA-1+ recipients developed myeloma within 12 weeks; in contrast, all LFA-1− remained healthy, even 72 weeks after tumor inoculation. (B) BM isolated from mice injected with LFA-1+ 5T33MMvt cells was positive for 5T33MM idiotype and CD11a. (C) BM from LFA-1− recipients stained with 5T33MM idiotype and was negative for tumor cells. 5T33MMvt were stained in parallel as positive control. Experimental Hematology 2003 31, 48-55DOI: (10.1016/S0301-472X(02)00970-0)

Figure 7 In vivo growth of LFA-1+ and LFA-1− 5T33MMvt cells. (A) 5T33MMvt cells were sorted into LFA-1+ and LFA-1− fractions and injected intravenously into naive mice. All LFA-1+ recipients developed myeloma within 12 weeks; in contrast, all LFA-1− remained healthy, even 72 weeks after tumor inoculation. (B) BM isolated from mice injected with LFA-1+ 5T33MMvt cells was positive for 5T33MM idiotype and CD11a. (C) BM from LFA-1− recipients stained with 5T33MM idiotype and was negative for tumor cells. 5T33MMvt were stained in parallel as positive control. Experimental Hematology 2003 31, 48-55DOI: (10.1016/S0301-472X(02)00970-0)