Figure 4. P2ry12-CreER recombination in non-microglial populations.

(A) Analysis of P2ry12-CreER; Rosa26^Ai14 recombination showed a lack of recombination in oligodendrocytes (OLIG2), astrocytes (SOX9) or neurons (NEUN). (B) Analysis of recombination and coexpression with Cx3cr1-EGFP and CD206 in the spleen, lungs, heart, thymus, intestine and liver of adult P2ry12-CreER; Rosa26^Ai14; Cx3cr1-EGFP mice. (C) Quantification of recombination overlap with Cx3CR1-EGFP (heart, intestine, lungs, spleen, thymus) or CD206 (heart, intestine, lungs). CD206 coexpression in the spleen and thymus was not analyzed due to density of CD206 expression. (D) Flow cytometry and blood smear recombination analysis revealed negligible recombination of blood cells in P2ry12-CreER;Rosa26^Ai14 mice compared to Cx3cr1-EGFP (internal control), or to Cx3cr1-CreER; Rosa26^Ai14 mice. *p=0.0015, Student’s t-test. N = 4 mice for A. and D., N = 3 mice for B and C. Scale bars = 50 µm (A), 100 µm (B,D).

Figure 4—figure supplement 1. P2ry12-CreER recombination in the spleen, liver, lungs and circulating platelets.

(A) P2ry12-CreER recombination in the spleen showed a striking pattern of recombination in the marginal zone of the spleen. (B) Higher magnification of spleen recombination and comparison with Cx3CR1-EGFP expression shows that P2ry12-CreER recombination in the spleen is more restricted to the marginal zone than Cx3CR1-EGFP expression, which is also expressed throughout the red and white pulp. (C) P2ry12-CreER; Rosa26^Ai14 recombination in the adult liver was not seen in perivascular SMA-adjacent Cx3cr1-EGFP cells, but was seen in Kupffer cells. (D) P2ry12-CreER; Rosa26^Ai14 recombination in the lungs was seen primarily around SMA-delimited airways and associated vessels. (E) P2ry12-CreER; Rosa26^Ai14 recombination in platelets was analyzed by blood smears. In PF4-Cre; Rosa26^Ai14 mice, which have well-described platelet recombination, TdTomato+; CD41+ platelets were observed, but no TdTomato expression was observed in P2ry12-CreER; Rosa26^Ai14 platelets. Scale bars = 1 mm (A), 500 µm (B), 100 µm (C, D), 30 µm (E).

Figure 4—figure supplement 2. P2ry12-CreER recombination in embryonic organs.

Pregnant mice were induced with tamoxifen at E13.5, E15.5 and E17.5 and embryos were collected at E18.5. The liver (A), heart (B), intestine (C) and lungs (D) were analyzed for recombination and coexpression with IBA1 or CD206. (E) Quantification of recombination overlaps with IBA1 or CD206 in the liver, heart, intestine and lungs. Error bars = SEM. Scale bars = 50 µm (A–D).