Significant expression of IL-17 receptor in myeloma cells. (A) Total cell lysates were prepared from MM cells, and separated by electrophoresis on 5% to 20% polyacrylamide gradient gels. Samples were probed with antisera to IL-17 receptor and GAPDH as indicated. (B) RNA was isolated from purified primary MM cells and qPCR was performed as described in “Quantitative PCR for IL-17 and IL-17R expression.” Results are presented as relative expression value. *P < .05. (C) Paraffin-embedded tissue sections from MM patients (N = 10) were stained using anti–IL-17R antibody as described in “Immunohistochemistry” and evaluated using a Nikon transmitted light microscope. The majority of the MM cells are positively stained with IL-17R receptor antibody in 7 of 10 patients. Two representative stained sections are shown at 200× magnification. (D) Myeloma cell lines were stained with isotype control antibody (top panel) or anti–IL-17R antibody (bottom panel) and analyzed by confocal microscopy. One representative cell line of 4 experiments is shown at 640× magnification. (E) MM cell lines (N = 3) were cultured alone or cocultured with BMSCs with or without IL-17 in the presence or absence of anti–IL-17R antibody. Proliferation was measured by 3H-thymidine incorporation after 3 days. Data are presented as percentage proliferation in the presence of IL-17 or IL-17R antibody compared with control and shown as mean plus or minus SEM. *P < .05.
