Optimization of tamoxifen-induced Cre activity and its effect on immune cell populations
Optimization of tamoxifen-induced Cre activity and its effect on immune cell populations
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DOI：
10.1038/s41598-020-72179-0
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发表时间：
2020-09-17
影响因子：
4.6
通讯作者：
Creusot, Remi J.
中科院分区：
文献类型：
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作者：
Donocoff, Rachel S.;Teteloshvili, Nato;Creusot, Remi J.
三苯氧胺（TAM）诱导的Cre重组酶系统是研究基因功能的重要工具，当早期切除或过度表达可导致发育缺陷或胚胎致死时。然而，对于TAM体内给药的最佳途径和剂量仍缺乏共识。在这里，我们评估剂量和交付TAM的Cre激活免疫细胞亚群的评估纵向和空间使用转基因小鼠普遍表达的Cre/ER和Cre诱导的荧光报告YFP。在比较了两种TAM递送方法（腹膜内与口服管饲）和不同剂量后，我们发现连续五天口服施用3 mg TAM提供了最大的报告基因诱导，体内不良反应最小。TAM的血清水平在开始治疗后1周达到峰值，然后缓慢下降，与给药和给药方法无关。特定组织（肝、脾、淋巴结和胸腺）中的TAM浓度也取决于递送方法和剂量。Cre诱导在髓系细胞和B细胞中最高，在T细胞中显著较低，双阳性胸腺细胞对TAM的反应显著较高。除了建立TAM的最佳剂量和给药外，我们的研究还揭示了当使用Cre/ER模型时，Cre在不同细胞免疫群体中的不同活性。
Tamoxifen (TAM) inducible Cre recombinase system is an essential tool to study gene function when early ablation or overexpression can cause developmental defects or embryonic lethality. However, there remains a lack of consensus on the optimal route and dosage of TAM administration in vivo. Here, we assessed dosage and delivery of TAM for activation of Cre in immune cell subsets assessed longitudinally and spatially using transgenic mice with ubiquitously expressed Cre/ER and the Cre-inducible fluorescent reporter YFP. After comparing two TAM delivery methods (intraperitoneal versus oral gavage) and different doses, we found that 3 mg of TAM administered orally for five consecutive days provides maximal reporter induction with minimal adverse effects in vivo. Serum levels of TAM peaked 1 week after initiating treatment then slowly decreased, regardless of dosing and delivery methods. TAM concentration in specific tissues (liver, spleen, lymph nodes, and thymus) was also dependent on delivery method and dose. Cre induction was highest in myeloid cells and B cells and substantially lower in T cells, and double-positive thymocytes had a notably higher response to TAM. In addition to establishing optimal dose and administration of TAM, our study reveals a disparate activity of Cre in different cell immune populations when using Cre/ER models.