Ontogeny of Tissue-Resident Macrophages

Abstract

The origin of tissue-resident macrophages, crucial for homeostasis and immunity, has remained controversial until recently. Originally described as part of the mononuclear phagocyte system, macrophages were long thought to derive solely from adult blood circulating monocytes. However, accumulating evidence now shows that certain macrophage populations are in fact independent from monocyte and even from adult bone marrow hematopoiesis. These tissue-resident macrophages derive from sequential seeding of tissues by two precursors during embryonic development. Primitive macrophages generated in the yolk sac (YS) from early erythro-myeloid progenitors (EMPs), independently of the transcription factor c-Myb and bypassing monocytic intermediates, first give rise to microglia. Later, fetal monocytes, generated from c-Myb(+) EMPs that initially seed the fetal liver (FL), then give rise to the majority of other adult macrophages. Thus, hematopoietic stem cell-independent embryonic precursors transiently present in the YS and the FL give rise to long-lasting self-renewing macrophage populations.

Keywords: C-Myb; erythro-myeloid progenitors; fetal liver; hematopoiesis; hematopoietic stem cells; macrophages; monocytes; yolk sac.

Figure 1

Fetal hematopoiesis. Primitive, transient definitive, and definitive waves of fetal hematopoiesis sequentially generate progenitors able to seed the fetal liver. Primitive hematopoiesis starts at E7.0 in the blood islands of the extra-embryonic yolk sac (YS) and generates erythro-myeloid progenitors (EMPs). Early EMPs initially express CD41 and later, CSF-1R, a signature of myeloid/macrophage commitment. Concomitant to the establishment of the blood circulation at E8.5, the YS hemogenic endothelium (HE) generates late EMPs expressing C-Myb. At approximately E9.0, the intra-embryonic mesoderm generates additional HE and emerging progenitors with lymphoid potentials (LMPs) without long-term reconstitution (LTR) capacity. These C-Myb⁺ EMPs and LMPs constitute the so-called transient definitive wave. Finally, hematopoietic stem cells (HSCs) with LTR activity emerge from the main HE situated in the aorta-gonad-mesonephros (AGM) regions and in the placenta.

Figure 2

Primitive hematopoiesis and yolk sac macrophage ontogeny. Early EMPs emerge in the YS around E7.5 before establishment of the blood circulation. They express CD41 and CSF-1R and are independent of the transcription factor C-Myb. Upon establishment of the blood circulation around E8.5, EMPs differentiate into primitive macrophages as well as primitive erythrocytes and granulocytes. Primitive macrophages seed all fetal tissues, in particular the head where they will give rise to future brain microglia that are able to continuously self-renew throughout adulthood. EMPs seeding the fetal liver briefly expand to generate a local macrophage population, likely important for sustaining enucleation of primitive erythrocytes passing through the sinusoid prior to the establishment of definitive hematopoiesis and the generation of fetal monocyte-derived macrophages in the fetal liver.

Figure 3

Fetal monopoiesis and macrophage ontogeny. At E8.5 as the blood circulation is established, the YS HE, possibly in conjunction with other hemogenic sites, generates EMPs expressing CD41 and c-Myb. These late EMPs seed the fetal liver around E9.5 where they expand rapidly to give rise to CSF-1R⁺ myeloid progenitors that are able to generate fetal monocytes through cMoP intermediates at E12.5. Fetal monocytes then spread via the blood circulation to all tissues, with the exception of the brain, which is isolated by the establishment of the blood–brain barrier at approximately E13.5. In the tissues, fetal monocytes begin to differentiate into macrophages, progressively outnumbering the previously established primitive macrophages. These fetal monocyte-derived macrophages maintain the capacity to self-renew throughout adulthood in certain tissues, such as the liver or the lung, where they will not be replaced by adult BM-derived monocytes.

Figure 4

Transition between fetal and adult hematopoiesis. Hemogenic endothelial cells from extra and intra-embryonic hematopoietic tissues generate C-Myb-dependent multipotential progenitors, such as LMPs and pre-HSCs, between E9.0 and E10.5, culminating with the emergence of mature HSCs with long-term reconstitution-bearing potential. CD93 (AA4.1) expression is associated with the emergence of lymphoid potential, whereas Sca-1 is the hallmark of HSCs. These progenitors seed the fetal liver around E10/E11, expanding and giving rise to the various lineages of the hematopoietic system, including fetal monocytes. These late fetal monocytes continue to participate in the tissue-resident macrophage network until hematopoiesis switches completely from the fetal liver to the bone marrow. Once adult hematopoiesis begins to take place in the bone marrow generating monocytes, certain tissues, such as the dermis, heart peritoneum, and the gut, continue to recruit adult monocytes to generate resident macrophages and replace with time the embryonic-derived macrophages.

Figure 5

Timeline of fetal and adult hematopoiesis. The primitive hematopoiesis is initiated in the yolk sac independently of C-Myb activity, and generates early CSF-1R⁺ EMPs that give rise to YS macrophages without monocytic intermediates during a short time window and will establish the brain microglia. The transient definitive hematopoiesis and then the definitive hematopoiesis are both dependent on C-Myb activity and generate progenitors that differentiate in the fetal liver. The transient definitive wave, which include EMPs and then LMPs, give rise in particular to fetal monocytes that seed the tissues prior to birth to establish the self-renewing tissue-resident macrophage network. Although only HSCs, which result from the definitive hematopoiesis, seem to be maintained in the bone marrow in adults, the relative contribution of the transient definitive wave to the adult immune system remains unclear.

Figure 6

Fate-mapping systems. (A) The Runx-iCre fate-mapping model (86) used in our study targets the hemogenic transition (88), hence, labeling specifically progenitors in the process of budding out from the hemogenic endothelium. The Runx1 expression decreases in progenitors once they start to express Vav thus reducing the chances of tagging released progenitors from precedent waves (88). As a consequence, Runx-iCre tagging is restricted to a short time window in the lifespan of a given progenitor and allows a sharp definition of each hematopoietic wave. However, this model also restricts the tagging to only a small fraction of the targeted progenitor wave. (B) Tie2 is expressed in all endothelial cells that constitute the hemogenic endothelia even before the hemogenic transition (89). Thus, all endothelial cells and their progeny (non-hematopoietic and hematopoietic cells) will be labeled after tamoxifen injection using the Tie2-iCre model. As a consequence, an early tamoxifen injection (such as at E7.5) will result in the tagging of all hematopoietic cells emerging before the time of analysis. This will include progenitors from the primitive, the transient definitive, and the definitive waves if, for example, the analysis is done at E11.5. A late injection (such as at E10.5) will restrict the tagging to only the latest hematopoietic stem cells wave as they are just budding from HEs (90). Thus, this model might not be suitable to clearly separate the primitive from the transient definitive waves of hematopoiesis. However, this model could be important to study late HSC progeny as no other progenitors than HSCs emerge from HE after E10.5 (91). (C) C-kit is expressed by all hematopoietic progenitors and does not label endothelial cells that constitute the HEs (89). An early tamoxifen injection (such as at E7.5) will restrict the labeling to early progenitors making suitable the c-kit-iCre model to study the primitive hematopoiesis. However, the FL recruits progenitors of each hematopoietic wave from E8.5 until E11 (79). These progenitors still express c-kit and coexist after seeding the FL during the time necessary for their differentiation (47, 55). A later tamoxifen injection (such as at E9.5) might thus result in the cumulative labeling of undifferentiated primitive and definitive progenitors, including the transient wave of EMPs and LMPs. Thus, such model may not be suitable to resolve the complexity of the different embryonic hematopoietic waves characterized by short time windows of emergence and strong overlapping tendencies. Primitive hematopoietic progenitors are rapidly consumed and the engagement of EMPs and LMPs in FL hematopoiesis reduces the expression of c-kit on their surface. Thus, later tamoxifen injection (such as at E11.5) could restrict the labeling to newly derived HSCs expressing high level of c-kit without labeling precedent progenitor waves (87). Such model might be interesting to study the progeny of late HSCs although the risk of tagging the progeny of EMPs and LMPs or later committed progenitors derived from HSCs remains high and difficult to exclude. Further analysis would be necessary to clarify the potential of such model.