Broader expression of the mouse platelet factor 4‐cre transgene beyond the megakaryocyte lineage

Abstract

Summary Background Transgenic mice expressing cre recombinase under the control of the platelet factor 4 (Pf4) promoter, in the context of a 100‐kb bacterial artificial chromosome, have become a valuable tool with which to study genetic modifications in the platelet lineage. However, the specificity of cre expression has recently been questioned, and the time of its onset during megakaryopoiesis remains unknown. Objectives/Methods To characterize the expression of this transgene, we used double‐fluorescent cre reporter mice. Results In the bone marrow, Pf4‐cre‐mediated recombination had occurred in all CD42‐positive megakaryocytes as early as stage I of maturation, and in rare CD42‐negative cells. In circulating blood, all platelets had recombined, along with only a minor fraction of CD45‐positive cells. However, we found that all tissues contained recombined cells of monocyte/macrophage origin. When recombined, these cells might potentially modify the function of the tissues under particular conditions, especially inflammatory conditions, which further increase recombination in immune cells. Unexpectedly, a subset of epithelial cells from the distal colon showed signs of recombination resulting from endogenous Pf4‐cre expression. This is probably the basis of the unexplained colon tumors developed by Apcflox/flox;Pf4‐cre mice, generated in a separate study on the role of Apc in platelet formation. Conclusion Altogether, our results indicate early recombination with full penetrance in megakaryopoiesis, and confirm the value of Pf4‐cre mice for the genetic engineering of megakaryocytes and platelets. However, care must be taken when investigating the role of platelets in processes outside hemostasis, especially when immune cells might be involved.

Cre/lox Studies Identify Resident Macrophages as the Major Source of Circulating Coagulation Factor XIII-A

“…FXIII-A.Flox mice were crossed with mice expressing cre recombinase in megakaryocytes and certain macrophage populations (Pf4-cre 15,16 ) or exclusively in myeloid cells (LysM-cre 14 and CD11b-cre 13 ). In Pf4-cre.Flox mice, plasma FXIII-A activity was reduced to 23±3% ( P <0.001; Figure 1B).…”

“…This result further supports a macrophage origin for FXIII-A because a stop/flox study has established that Pf4-cre expression occurs within resident tissue macrophages in addition to megakaryocytes. 16 Notably, Pf4-cre-mediated recombination within macrophages is mosaic, 16 potentially accounting for incomplete depletion of plasma FXIII-A. The FXIII-A-expressing cells in brain, heart, and aorta have been previously identified as macrophages, 19,20 and consistent with this, we show (1) that in heart, as previously demonstrated in skin, 26 FXIII-A partially co-localizes with the M2 macrophage marker CD163 and (2) that in heart, brain, and aorta, Pf4-cre depleted FXIII-A mRNA.…”

“…14 The Pf4-cre was designed to delete in the platelet lineage, 15 but a recent reporter mouse study has also shown Pf4-cre expression within resident macrophages. 16 …”

FAK regulates platelet extravasation and tumor growth after antiangiogenic therapy withdrawal

“…FAK was absent only in megakaryocytes and platelets. However, recent work by Pertuy et al (57) showed that the Pf4-Cre transgene might have a broader expression beyond the megakaryocytic lineage. Pertuy et al found that 0.3% of CD45 + leukocytes as well as a small percentage of intestinal epithelial cells showed expression of Pf4.…”

The contribution of mouse models to the understanding of constitutional thrombocytopenia

“…Almost all recombination in the megakaryocytic lineage has been obtained by using the Pf4-Cre system developed by Radek Skoda's group. 2,3 And finally, "knock-in mice" allow the introduction of point mutations or insertions/deletions through homologous recombination at the locus of interest. These models faithfully reproduce the mutations present in humans and represent the best approach to mimic the pathology.…”