Figure 3

CGRP enhances cell proliferation with suppression of ROS production and cell apoptosis in HSPCs under stress conditions. (a) Percentages of BrdU-positive cells in each BM cell fraction (LSK, Lin⁻, BMMNCs, and myeloid cells) of the recipient mice (CD45.1) following transplantation of BM cells (CD45.2) from Ramp1^−/− and WT mice were determined by flow cytometry. Data are shown as means ± SD of three mice. *p < 0.05, **p < 0.01 compared with WT mice (Student’s t test) (b) The homing ability of the transplanted donor BMMNCs (Ramp1^−/− and WT) into the BM of recipient mice was determined by flow cytometry. Total numbers of donor-derived CD45.2⁺ BMMNCs in each recipient mouse are shown. (c) The ROS production in Lin⁻ cells from BMMNCs of the recipient mice transplanted with donor BM cells from Ramp1^−/− or WT mice was determined by flow cytometry using redox-sensitive MitoTracker Orange CMH2TMROS fluorescence staining 16 hours after transplantation. Data are shown as means ± SD of three mice. *p < 0.05 compared with WT mice (Student’s t test). (d) The percentage of apoptotic cells in Lin⁻ cells from BMMNCs of the recipient mice transplanted with donor BM cells from Ramp1^−/− or WT mice were determined by flow cytometry using annexin V staining after transplantation. Data are shown as means ± SD of three mice. *p < 0.05 compared with WT mice (Student’s t test). (e) The percentages and absolute numbers of LSK cells in the Lineage negative population from BMMNCs cultured alone or co-cultured with BM stromal cells in the presence or absence of CGRP^{8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29,30,31,32,33,34,35,36}, as determined by flow cytometery. Data are shown as means ± SD of three mice. *p < 0.05, **p < 0.01 compared with WT mice (Ordinary one-way ANOVA and Turkey’s multiple comparisons test).