Figure 3.
Figure 3. Combined targeting of the IL-6R/STAT3 pathway and the MEK1,2/ERK1,2 module induces apoptosis of INA-6 cells even in the presence of BMSCs. (A) INA-6 cells transfected with expression plasmids for siRNAs against ERK1 and ERK2 (pSU-ERK1 + 2), or control plasmid (pSU), were cultured in medium with IL-6 (□) or in the presence of BMSCs (▦), and treated with or without 50 μg/mL Sant7. Apoptosis was assayed after 3 days and survival of the respective pSU-transfected samples was set as 100%. Sant7 induced strong cell death in medium and knockdown of ERK1,2 had no effect. In the presence of BMSCs, myeloma cells were protected from the apoptotic effect of Sant7, and blockade of both the IL-6R/STAT3 and the MAPK pathways was required for apoptosis. (B) INA-6 cells transfected with an expression plasmid for an siRNA against STAT3 (pSU-STAT3) or control plasmid (pSU), cultured as described in panel A and treated with or without 50 μM PD98059. In the presence of BMSCs, the combination of both drugs (ie, blockade of the IL-6R/STAT3 and the MAPK pathways) was required for apoptosis. (C) INA-6 cells cultured as described in panel A and treated with 50 μM PD98059 and/or 50 μg/mL Sant7. The effects of both pharmacologic inhibitors mimicked those of the siRNAs that abrogate the respective pathway. (D) INA-6 cells cultured in medium with IL-6 (top) or in the presence of BMSCs (bottom) without (□) or with (▪) 50 μM PD98059, and with increasing concentrations of Sant7. Increasing concentrations of Sant7 alone were sufficient to kill the cells in medium, but the combination of both pathway blockers was required to achieve the same result in the presence of BMSCs. (E) Expression of STAT3 protein that is derived from a sequence no longer recognizable by the siRNA (plasmid pS3R) largely blocked the detrimental effects of STAT3 siRNA expression (□). Cell death induced through IL-6 depletion was not affected (). All data shown are derived from at least 3 independent experiments. Statistical analysis (A-C) revealed very significant (**) or no (n.s. = not significant) differences as indicated. Error bars denote the range of values derived from at least 3 independent experiments.

Combined targeting of the IL-6R/STAT3 pathway and the MEK1,2/ERK1,2 module induces apoptosis of INA-6 cells even in the presence of BMSCs. (A) INA-6 cells transfected with expression plasmids for siRNAs against ERK1 and ERK2 (pSU-ERK1 + 2), or control plasmid (pSU), were cultured in medium with IL-6 (□) or in the presence of BMSCs (▦), and treated with or without 50 μg/mL Sant7. Apoptosis was assayed after 3 days and survival of the respective pSU-transfected samples was set as 100%. Sant7 induced strong cell death in medium and knockdown of ERK1,2 had no effect. In the presence of BMSCs, myeloma cells were protected from the apoptotic effect of Sant7, and blockade of both the IL-6R/STAT3 and the MAPK pathways was required for apoptosis. (B) INA-6 cells transfected with an expression plasmid for an siRNA against STAT3 (pSU-STAT3) or control plasmid (pSU), cultured as described in panel A and treated with or without 50 μM PD98059. In the presence of BMSCs, the combination of both drugs (ie, blockade of the IL-6R/STAT3 and the MAPK pathways) was required for apoptosis. (C) INA-6 cells cultured as described in panel A and treated with 50 μM PD98059 and/or 50 μg/mL Sant7. The effects of both pharmacologic inhibitors mimicked those of the siRNAs that abrogate the respective pathway. (D) INA-6 cells cultured in medium with IL-6 (top) or in the presence of BMSCs (bottom) without (□) or with (▪) 50 μM PD98059, and with increasing concentrations of Sant7. Increasing concentrations of Sant7 alone were sufficient to kill the cells in medium, but the combination of both pathway blockers was required to achieve the same result in the presence of BMSCs. (E) Expression of STAT3 protein that is derived from a sequence no longer recognizable by the siRNA (plasmid pS3R) largely blocked the detrimental effects of STAT3 siRNA expression (□). Cell death induced through IL-6 depletion was not affected (). All data shown are derived from at least 3 independent experiments. Statistical analysis (A-C) revealed very significant (**) or no (n.s. = not significant) differences as indicated. Error bars denote the range of values derived from at least 3 independent experiments.

Close Modal
This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal