M-CSF and GM-CSF Regulation of STAT5 Activation and DNA Binding in Myeloid Cell Differentiation is Disrupted in Nonobese Diabetic Mice
Figure 1
Flow cytometric analysis of STAT5 phosphorylation in in vitro myeloid differentiation. (a) Bone marrow cultures (1 million cells/mL)
from NOD and C57BL/6 control mice were differentiated in culture with either, (b)
1000?U/mL GM-CSF plus 2g/mL
anti-M-CSF blocking antibodies -M),
or (c) with 500?U/mL M-CSF with 2g/mL
anti-GM-CSF blocking antibodies -GM),
for 48 hours at 3/5.
Cultures treated with medium only (“media
only” in (a), “0” in (b) and (c))
or with medium supplemented only with antibodies to block M-CSF or GM-CSF (ANTI-M-CSF, ANTI-GM-CSF, respectively, in (a)) were run in parallel with the
cytokine-stimulated cultures. In addition, separate aliquots of bone marrow
cells were cultured in M-CSF or GM-CSF alone without blocking antibodies (“no” in (b) and (c)). At 48 hours, cells were
stained with anti-STAT5Ptyr-FITC and anti-CD11b-PE antibodies for analysis by
intracellular immune-histochemical flow cytometry. Strain and treatment
regiments for each pair of cultures are listed on the X-axis. The percentage of
STAT5Ptyr+ in CD11b+ cells is given on the Y-axis. Graphs are representative of
3–9 separate runs
of each treatment. The p-values indicated were derived from Mann-Whitney
U test, Student’s t-test, or one-way ANOVA analyses, as appropriate, for
the sample group comparisons indicated by brackets.
