Bone Marrow Osteoblast Damage by Chemotherapeutic Agents
Figure 1
Melphalan and VP-16 exposure of HOB increases the level of active TGF-β1.
A) HOB were exposed to melphalan [50 µg/ml] or VP-16 [50 µM] for 24 hours and real time PCR for TGF-β1 completed. B) HOB were exposed to melphalan [100 µg/ml] or VP-16 [100 µM] for 4 hours, fixed and stained for detection of phospho-Smad2 (green), as a read-out of TGF-β1 activity (DAPI stain for nuclei-blue). C) HOB were exposed to melphalan [100 µg/ml], VP-16 [100 µM] or rTGF-β1 [10 ng/ml] with or without SB 431542 [10 µM] for 4 hours and a western blot completed for p-Smad2 and total Smad2. P-Smad2 expression was normalized to total Smad2 expression. D) HOB were exposed to rTGF-β1 [10 ng/ml] for 24 hours and real time PCR completed (left). Additionally, HOB were exposed to rTGF-β1 [10 ng/ml] for 24 hours, the HOB layer was rinsed and new media was added and allowed to condition for 24 hours before being evaluated by ELISA to quantitate the amount of secreted TGF-β1 (right). (NT = No Treatment, Mel = melphalan, VP = VP-16, TGF = rTGF-β1, SB = SB 431542).