Fetal microchimerism in skin wound healing

Abstract

Skin wound healing is a complex regenerative process involving various cell types. We recently investigated whether fetal microchimeric cells (FMCs) acquired during gestation contribute to maternal wound healing and used fetal microchimerism to investigate the recruitment of distant endothelial progenitor cells in skin wounds. Our study showed that fetal progenitor cells are recruited into maternal wounds and participate in inflammation and angiogenesis. These fetal cells might have beneficial effects in situations of maternal defective healing, and might also modify the adult maternal wound environment toward a scarless fetal-like wound healing.

Keywords: :

• angiogenesis
• fetal stem cells
• inflammation
• microchimerism
• regeneration
• skin
• wound healing

This article refers to:

Skin wound healing is a complex regenerative process involving various cell types and biological systems. It is schematically divided into three overlapping stages: inflammation, tissue synthesis and remodelling.¹ Conflicting data have been published regarding the participation of distant endothelial progenitor cells (EPCs) to the angiogenesis in the wound bed.^2-4 These studies used chimeric mice obtained by transplantation of a tagged bone marrow from a transgenic mouse into a wild type (WT) recipient after a myeloablative treatment usually chemotherapy or irradiation. This conditioning may modify the natural course of the healing process. Natural chimerism occurs in parous and pregnant females as a result of the trafficking of fetal cells to the maternal circulation during pregnancy and their long-term persistence after delivery.^5-8 Fetal microchimeric cells (FMCs) have been identified in larger numbers in pathological conditions, including cancer, inflammatory diseases or reparative processes as compared with normal tissues.^9-15 This recruitment capacity associated with their described plasticity^7,16 has suggested that the study of fetal cell microchimerism can inform on homing and engraftment of host stem cells in response to tissue injury. We recently investigated whether fetal microchimeric cells contributed to maternal wound healing and used this model to investigate the recruitment of distant EPCs in this process.¹⁷ Tracking GFP⁺ fetal cells in WT females, we showed that FMCs were constantly present in high numbers in maternal wounds reaching hundreds per 10⁴ maternal cells. Their numbers dramatically decreased to undetectable levels when the wound healed. Interestingly, in chronic wounds performed on Bleomycin-induced fibrotic skin, FMCs were still present at high levels on late time points after wounding as long as wounds were still open. FMCs were mainly CD45⁺ leukocytes at early stages and vWF⁺ endothelial cells at later stages. This followed the expected sequence of events during skin wound healing.^2,3 In 2/11 mice, we identified vessels entirely constituted of fetal GFP⁺ endothelial cells and showed that they were branching and were connected to maternal circulation on adjacent serial sections. Besides we observed an amplification of circulating GFP⁺ cells mainly CD34⁺CD11b⁻VEGFR2⁻ two days after skin wounding. These findings along with previous reports, including ours, converge toward the presence of fetal EPCs among the transferred FMCs.^11,12,18,19 Two important questions that we addressed were as follows: were FMCs recruited in high numbers de novo or did randomly transferred FMCs proliferate in situ after tissue injury? Did fetal EPCs respond to specific chemotactic and angiogenic signals as their maternal counterparts? To answer these questions, we implanted Matrigel plugs spiked with recombinant VEGFa in the sub-cutis of pregnant mice. By detecting fetal GFP⁺ CD31⁺ endothelial cells in VEGFa-spiked Matrigel, we showed that fetal EPCs were recruited toward an acellular Matrigel plug in response to VEGFa signaling. Thus, FMCs that were detected in maternal skin wounds were, at least partly, recruited de novo in response to specific signals along with their maternal counterparts.¹⁷

The transfer of wild type cells has been reported to improve diabetic chronic wounds in parabiotic mice.²⁰ It is therefore tempting to assume that normal FMCs might have beneficial effects in situations of maternal defective healing where these will display a selective advantage and possibly rescue the affected skin.^20,21 Indeed, we have previously observed the transient healing of a recalcitrant leg ulcer in a sickle cell patient during pregnancy and hypothesized that it might be due to the recruitment of healthy fetal cells from her offspring.²²

We next hypothesized that the presence of fetal cells recruited into normal wounds during pregnancy might modify the adult maternal wound environment toward a scarless fetal-like wound healing.²³ Although a difference in collagen deposition is hard to assess on histology, Collagen 1A1 and TGFβ transcripts were respectively decreased by 34% and 43% in wound beds of pregnant compared with non-pregnant mice (Fig. 1). This decrease did not reach statistical significance. Of course, this might be due to many accompanying physiological changes in pregnant mice. Nevertheless, this raises questions about the advantage of using fetal stem cells instead of adult derived stem cells in cell therapies for wound regeneration.

Figure 1.Collagen1A1 and TGFβ1 transcripts in skin wounds on pregnant and virgin mice. Surgical wounds (5 mm diameter) were performed on pregnant (E10) and virgin mice matched for age (n = 7 for each group). Seven days after wounding, wounds were harvested and mRNA extracted. Real-time PCR was conducted using SYBR^®GREENPCR Master Mix (Applied Biosystems). mRNA values were normalized to the expression level of 18s RNA. Each sample was analyzed in duplicate. Histogram represents means +/− SEM.

In conclusion, the study of fetal microchimerism generates new concepts about the potential of fetal stem cells in therapy.

Abbreviations:
WT	=	wild type
GFP	=	enhanced green fluorescent protein
vWF	=	von Willebrand factor