Loss of syndecan-1 and cell-surface HS reduces the in vitro growth of MM resulting from increased apoptosis. L363 and RPMI-8226 MM control cells (TetR) and L363 and RPMI-8226 MM cells containing inducible shRNA, targeting either syndecan-1 (shSYN1) or EXT1 (shEXT1a, b, and c), were incubated with (+dox) or without doxycycline (−dox) for 5 days before each experiment to allow for optimal knockdown of the target genes. (A-B) Expression of syndecan-1 (A) or cell-surface HS (B) measured by FACS using antibodies BB4 (Serotec) and 10E4 (Seikagaku America), respectively. The expression level of the untreated samples was normalized to 100%. Bars represent the mean ± SD of at least 5 independent experiments. (C-D) The growth rate of L363-TetR, -shSYN1, and -shEXT1a, b, and c MM cells (C) or RPMI-TetR and -shEXT1a MM cells (D) was analyzed over a 10-day culture period in the presence of 10% FCS. The growth curves represent the mean ± SD of 3 to 5 independent experiments (in triplicate). (E) The percentages of apoptotic cells were measured for the different L363 MM cell lines after 3 days of culture. Apoptotic cells were determined as annexin V⁺/TO-PRO⁻. Necrotic, TO-PRO single-positive cells, were excluded. *P ≤ .05; **P ≤ .01; ***P ≤ .001.