Figure 7.

Paired blood exchange (PBE) demonstrates that CCR2⁺ macrophages observed at UUO-7 are derived from donor monocytes. (A) Schemata of experimental strategy for PBE to track fate of immune cells recruited to the kidney. One day after UUO, whole blood exchange was performed between pairs of C57BL/6 and Ly5.1 mice (n=4) and pairs were euthanized either 2 or 7 days after UUO. (B) Representative flow cytometry plots of circulating CD45⁺ cells from pairs of mice pre- and immediately post-blood exchange, illustrating approximately 40% of circulating cells were derived from donors after the exchange (CD45.1⁺CD45.2⁺ cells in C57BL/6; CD45.2^+/+ cells in Ly5.1 mice). (C) Percentage of circulating immune cells derived from paired donor over the experimental time course (n=4 pairs immediately post-PBE and at UUO-2; n=2 pairs at UUO-7). (D) Illustrative flow cytometry plots mapping donor cells (red) and recipient cells (gray) to the CD11b⁺F4/80^Lo monocyte and CD11b⁺F4/80^Hi macrophage gates, the monocyte waterfall in obstructed and contralateral kidneys at 2 days post-UUO, and the expression of CCR2 and MHCII 7 days post-UUO. (E) Average number of donor cells mapping to the CD11b⁺F4/80^Lo monocyte and CD11b⁺F4/80^Hi macrophage gates in obstructed kidneys at 2 and 7 days after UUO. (F) The expression of CCR2 (mean fluorescent intensity, MFI) on the donor cells compared with the host cells from the CD11b⁺F4/80^Hi macrophage gate in obstructed kidneys 7 days after UUO. n=4 per group. *P<0.05 by Mann–Whitney test.