Thrombopoietin and chemokine mRNA expression in patient post-chemotherapy and in vitro cytokine-treated marrow stromal cell layers

CD34(+) cells and megakaryocyte progenitors were lower in marrow from patients after hematological recovery from the first cycle of 5-fluorouracil, leucovorin, adriamycin, cyclophosphamide (FLAC) chemotherapy plus PIXY321 (GM-CSF/interleukin 3; IL-3 hybrid) than in FLAG + GM-CSF or pre-FLAC marrows, Marrow stromal layers, an in vitro model of the marrow microenvironment. express a combination of stimulatory and inhibitory factors that modulate hematopoietic progenitor cell proliferation and differentiation, The TaqMan assay and quantitative reverse transcriptase-polymerase chain reaction were used to measure monocyte chemoattractant protein-1 (MCP-1). melanoma stimulatory growth activity. and monokine inducible by interferon-gamma (Mig) (inhibitory. chemokines for primitive or megakaryocyte progenitors) mRNA levels in in vitro PIXY and GM-CSF-treated and patient post-FLAC marrow stromal layers, Chemokine mRNA was increased after in vitro GM-CSF and to a lesser extent after PIXY treatment. MCP-1 mRNA levels were Fivefold higher in FLAG + PIXY than in FLAC + GM-CSF layers, and Mig mRNA was elevated in FLAG + GM-CSF layers. Thrombopoietin (TPO), insulin-like growth factor I (TGF-I), and IGF-II (stimulatory factors) for primitive and megakaryocyte progenitors mRNA were also measured. TPO mRNA levels were 30% lower in GR-CSF and PIXY-pretreated than in control layers with no decrease in IGF mRNA, TPO mRNA in stromal layers of patients who developed grade 3 thrombocytopenia (platelets < 20 x 10(9)/l) during the third cycle of FLAC was only 24% of levels in stromal layers of marrow from other post-FLAG patients, Results demonstrate that patient and in vitro treatment had modulatory effects on TPO and chemokine mRNA expression in marrow stromal layers.