Figure 7

Decreasing tamoxifen dose or frequency of administration can reduce off-target hematological toxicity but it also decreases on-target Cre-lox recombination efficiency (a) WT and R26CreERT2 received intraperitoneal injection of tamoxifen at 75 mg/kg, 37.5 mg/kg or 18.75 mg/kg for 3 consecutive days from P9 to P11 or only one intraperitoneal injection of tamoxifen at 75 mg/kg at P9. Animals were euthanized at P19 and blood was analyzed. Data are represented as mean ± SEM of n = 4–12 animals per group. Two-way analysis of variance was used to analyze differences between WT and R26CreERT2. ****p-value ≤ 0.0001. (b) Bmp10fl/fl and R26CreERT2 Bmp10fl/fl mice received intraperitoneal injection of tamoxifen at 75 mg/kg, 37.5 mg/kg or 18.75 mg/kg for 3 consecutive days from P9 to P11 or only one intraperitoneal injection of tamoxifen at 75 mg/kg at P9. Animals were euthanized at P19, and Bmp10 mRNA level was quantified in cardiac tissue (right atria) and liver tissue. Data are represented as mean ± SEM of n = 5–10 animals per group. Two-way analysis of variance was used to analyze differences between Bmp10fl/fl and R26CreERT2 Bmp10fl/fl. *p-value ≤ 0.05; **p-value ≤ 0.01; ***p-value ≤ 0.001; ****p-value ≤ 0.0001. (c) Cre recombinase is fused to a modified ligand-binding domain of the estrogen receptor (ERT2 LBD). Cre ERT2 LBD transgene is inserted in the Rosa26 (R26) locus, allowing ubiquitous expression. The resulting protein, CreERT2, is restricted to the cytoplasm in the absence of tamoxifen. In the presence of tamoxifen or its metabolites (T), CreERT2 is translocated into the nucleus. Once in the nucleus, CreERT2 excises the target DNA sequence flanked by loxP sites (on-target effect) but it also induces DNA damage (off-target effect), independently of the targeted Cre-lox recombination.