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Myostatin propeptide expands primitive CML cells
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Figure 4. MSTNpp stimulation activates STAT5 and SMAD2/3 in chronic myeloid leukemia (CML) cells. (A) Schematic illustration of the RNA-sequencing experiment; CD34+ CD38low chronic phase CML cells from five individual patient samples were sorted and cultured for 3 hours (h) and 24 h with or without MSTNpp prior to RNA- extraction and sequencing. (B) Gene set enrichment assay (GSEA) showing upregulation of STAT5 and TGF-b1 target genes after 3 h MSTNpp stimulation. (C) Histograms showing activation of SMAD2/3 and STAT5 after 15 minutes stimulation with MSTNpp in CD34+ chronic phase CML cells, compared to unstimulated con- trol. Unstained cells in gray, unstimulated cells in blue, and MSTNpp stimulated cells in red. (D) Median fluorescence intensity (MFI) of pSTAT5 with or without MSTNpp stimulation. Paired samples from six individual CD34+ chronic phase CML patient samples are shown. ***P<0.001. NES: normalized enrichment score; FDR: false discovery rate.
the same patients (Figure 3D). All cell fractions tested expressed the MSTN gene, with the highest expression in MSC. Although the expression levels were approximately 20x lower in MSC than in skeletal muscle, these data sug- gest that the MSC in the BM are another source of MSTNpp, possibly contributing to the growth and sur- vival of CML cells.
To evaluate whether the MSTNpp levels are different in leukemic and healthy individuals, we performed a MSTNpp specific ELISA on plasma from CML patients and healthy individuals (Figure 3E), as well as on plasma from healthy BM aspirates. All samples showed detectable levels of MSTNpp, but there was no significant
difference between MSTNpp levels in CML patients ver- sus normal individuals. Moreover, the concentrations of MSTNpp varied greatly depending on the patient/donor, with levels ranging from ~0.1-400 ng/mL. As CML cells respond to MSTNpp even at 1 ng/mL (Figure 2A), the MSTNpp concentrations in the plasma from CML patients are within physiologically relevant levels for the leukemic cells
MSTNpp stimulation activates STAT5 and SMAD2/3 in chronic myeloid leukemia cells
To search for cellular programs and signaling pathways transcriptionally regulated by MSTNpp in primitive CML
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