Page 53 - Haematologica May 2020

EGR1 regulation of human BMSC
which have been reported to promote cell proliferation, differentiation and survival.33-35 Furthermore, experiments using different ROS inhibitors showed that ROS produc- tion in BMSC was mostly affected by oxidative pathway changes, and to a lesser extent by p38 pathway alterations or 4-ABAH induced inhibition of myeloperoxidase (MPO).
In addition to the role of EGR1 in regulating key BMSC functions in vitro as demonstrated here, we observed that EGR1 expression levels were significantly different in dif- ferent (patho)physiological situations. These expression data were generated with prospectively isolated BMSC and can therefore be considered to accurately reflect the in vivo situation. We found that EGR1 was significantly
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Figure 6. Increased proliferation in EGR1 knockdown bone marrow mesenchymal stromal cells (BMSC) is mediated by reactive oxygen species (ROS). (A) Biological process annotations for down-regulated proteins in EGR1 knockdown cells were identified using the DAVID Bioinformatics Resources 6.8. (B) Heatmap of down-reg- ulated proteins related to oxidative-reduction processes in EGR1 knockdown cells versus controls. NT: non-transfected cells; scr ctr: scramble control. (C) Representative FACS histogram of ROS levels in EGR1 knockdown cells compared with scramble control. (D) Fold change of ROS in EGR1 knockdown cells in com- parison with scramble control. Data are shown as mean fluorescence intensity (MFI) fold change±standard deviation (SD) (n=3). ROS MFI levels relative to dimethyl sulfoxide (DMSO) controls (E), percentage of dividing cells expressed as fold change compared to DMSO controls (F), and colony-forming units-fibroblast (CFU-F) fre- quencies (G) in EGR1 knockdown cells after ROS inhibitor treatment. Data are shown as mean±SD (n=3). NAC: N-acetylcysteine. *P<0.05.
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