Fig. 7
Gfi1 overexpression in MM.1S enhances bone loss in the MM-bearing mice and MM-induced OCL formation. Plasma Igλ concentrations were measured by ELISA, 1 week (1w) and 7 weeks (7w) after the intratibial injection of MM cells and at the time of harvest (8w) in both MM.1S Gfi1 o/e- (N = 7) and EV-injected mice (N = 6) (a). X-ray of three tibiae from an EV and two Gfi1 o/e-injected animals displaying similar plasma Igλ levels (b). Fixed tibiae were scanned using a μCT system (vivaCT 40, Scanco Medical). Representative μCT images demonstrating osteocytic lesions in the tibias of MM.1S Gfi1 o/e- and MM.1S EV-bearing mice compared with uninjected tibia (c). The bone tissue was measured by relative volume of calcified tissue (BV/TV) (N = 7) and the trabecular number (Tb.N.) (N = 6) in mice injected IT with (105) Gfi1 o/e or EV MM.1S cells, respectively. Data are shown as mean ± SEM (d). Representative microscopic images of bone sections stained with TRAP to identify osteoclasts (arrows) on cancellous bone of MM.1S Gfi1 o/e- and EV-bearing mice. Magnification × 20 (on the left) and representative areas (black squares) are magnified on the right (× 40). The bar graph on the right shows OCL surface normalized to bone surface (Oc.S/BS) from MM.1S Gfi1 o/e- (N = 6) and EV-bearing mice (N = 4). Data are shown as mean ± SEM (e). Mouse OCL precursors, obtained from murine bone marrow after stimulation with MCSF (10 ng/ml) and RANKL (50 ng/ml), were co-cultured with MM.1S EV and MM.1S Gfi1 o/e myeloma cells for 72 h and then TRAP stained. The bar graph represents the OCL number expressed as percentages normalized to EV controls. Values are mean ± SD; *p < 0.05. Representative microscopic images of TRAP-positive OCL after 72 h exposure to either MM.1S EV (left panel) or MM.1S Gfi1 o/e (right panel) showing increase number and size of OCL in cultures exposed to MM.1S Gfi1 o/e MM cells (f)