CD44 expression in BMC and BM-StrC: (A) Unseparated BMC

Figure 4: HSPC - BM-StrC binding supports HSPC quiescence and retards cell cycle progression: (A-D) BMC / HSPC were CFSE labeled. Cells were incubated for 24 h-72 h. Cell division was evaluated by CFSE-dilution (flow cytometry). (A) CFSE-labeled wt and CD44v6/v7^ko BMC and HSPC were seeded on BSA, HA or BM-StrC coated plates. Proliferation was evaluated after 24 h, 48 h and 72 h; mean percent ± SD (triplicates) of cells that had divided 2 times; significant differences between wt and CD44v6/v7^ko HSPC; (B,C) CFSE-labeled wt and CD44v6/v7^ko HSPC were seeded on BSA, HA, wt or CD446/v7^ko BM-StrC. The mean percent ± SD (triplicates) of HSPC that had not divided or divided 1-time or 2-times after 24 h-72 h of culture and a representative example after 72 h of culture are shown; significant differences between wt versus CD44v6/v7^ko HSPC: s; significant differences depending on the culture condition (BSA versus HA, wt or CD44v6/v7^ko BMStrC): +; (D) the experiment described in (B) was repeated in the presence of anti-panCD44, -CD44v6 or -CD44v; the mean percent±SD (triplicates) of HSPC that had divided 2-times after 24h-72h of culture is shown; significant differences in the presence of anti-CD44: *. (E-H) Cell cycle progression (PI staining) was evaluated in CD44^wt and CD44v6/v7^ko HSPC seeded on BSA, HA, CD44^wt and CD44v6/v7^ko BM-StrC; (E,F) the mean percent ± SD (triplicates) of HSPC in G0, G1/S and G2/M and a representative example are shown; significant differences between CD44^wt and CD44v6/v7^ko HSPC: s; significant differences by culture condition: +; (G;H) the experiment shown in (E) was repeated in the presence of anti-panCD44, -CD44v6 or -CD44v7; the mean percent ± SD (triplicates) of HSPC in G0, G1/S and G2/M and a representative example are shown; significant differences in the presence of anti-CD44: *.
Wt HSPC divide less frequently than CD44v6/v7^ko HSPC. HA and wt BM-StrC promote HSPC quiescence; anti-panCD44 and -CD44v6 drive HSPC into proliferation. CD44v6/v7^ko HSPC are not efficiently protected from proliferation by HA or wt BM-StrC. Furthermore, CD44^wt, but not CD44v6/v7^ko HSPC transition from G1/S to G2/M is retarded by HA and wt BM-StrC. The wt BM-StrC-promoted retardation in cell cycle progression is strongly affected by anti-CD44.