Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2015 Jul 24;47(7):e174.
doi: 10.1038/emm.2015.42.

Fetal hematopoietic stem cells express MFG-E8 during mouse embryogenesis

Jaehun Lee  1 , Byung-il Choi  2 , Seo Young Park  3 , Su Yeon An  1 , Jiyou Han  1 , Jong-Hoon Kim  1
Affiliations

Fetal hematopoietic stem cells express MFG-E8 during mouse embryogenesis

Jaehun Lee et al. Exp Mol Med. .

Abstract

The milk fat globule-EGF-factor 8 protein (MFG-E8) has been identified in various tissues, where it has an important role in intercellular interactions, cellular migration, and neovascularization. Previous studies showed that MFG-E8 is expressed in different cell types under normal and pathophysiological conditions, but its expression in hematopoietic stem cells (HSCs) during hematopoiesis has not been reported. In the present study, we investigated MFG-E8 expression in multiple hematopoietic tissues at different stages of mouse embryogenesis. Using immunohistochemistry, we showed that MFG-E8 was specifically expressed in CD34(+) HSCs at all hematopoietic sites, including the yolk sac, aorta-gonad-mesonephros region, placenta and fetal liver, during embryogenesis. Fluorescence-activated cell sorting and polymerase chain reaction analyses demonstrated that CD34(+) cells, purified from the fetal liver, expressed additional HSC markers, c-Kit and Sca-1, and that these CD34(+) cells, but not CD34(-) cells, highly expressed MFG-E8. We also found that MFG-E8 was not expressed in HSCs in adult mouse bone marrow, and that its expression was confined to F4/80(+) macrophages. Together, this study demonstrates, for the first time, that MFG-8 is expressed in fetal HSC populations, and that MFG-E8 may have a role in embryonic hematopoiesis.

PubMed Disclaimer

Figures

Figure 1
Figure 1
Milk fat globule-EGF-factor 8 (MFG-E8) and CD34 expressions in the yolk sac blood island of developing mouse fetuses at embryonic day E10.5. (a) Hematoxylin and eosin (H&E) staining image of E10.5 mouse embryo. Decidua, spongial layer and labyrinth of the placenta are shown outside of the yolk sac surrounding the embryo. (b and c) Enlarged image of the area indicated with ‘*' in a are shown in b. High magnification of the boxed area in b are shown in c. Cells located in the yolk sac blood island are large and have prominent nuclei. (d–f) Immunofluorescent labeling of the blood island in the yolk sac with anti-CD34 and anti-MFG-E8 antibodies. Note that the CD34+ cell co-expressed MFG-E8. The boxed area in f are separately shown in g. White arrowhead indicates CD34− cells. Scale bars in: a=250 μm; b and f=50 μm, and c and g=10 μm.
Figure 2
Figure 2
Milk fat globule-EGF-factor 8 (MFG-E8) and CD34 expression in the aorta, and liver of developing embryos at embryonic day E11.5 (a–d) Hematoxylin and eosin (H&E) staining and immunofluorescent labeling of dorsal aorta and cardinal veins with anti-CD34 and anti-MFG-E8 antibodies. CV, cardinal vein; DA, dorsal aorta. (e–h) H&E staining and immunofluorescent labeling of fetal liver sections with anti-CD34 and anti-MFG-E8 antibodies. Enlarged images of the boxed are shown in the upper right corner in each panels. Scale bar: 50 μm.
Figure 3
Figure 3
Expression of milk fat globule-EGF-factor 8 (MFG-E8) in placental development during pregnancy. (a–d) Hematoxylin and eosin (H&E) staining and immunofluorescent labeling of the developing placenta with anti-CD34 and anti-MFG-E8 antibodies at embryonic day (E)11.5. High magnification images of the boxed areas are shown in the upper right corner of each panels. Scale bar: 50 μm. (e) real-time reverse transcription-polymerase chain reaction (RT-PCR) analysis of MFG-E8 mRNA in the placenta at different days of gestation. (f) quantitative PCR (qPCR) analysis of MFG-E8 mRNA in the placenta at different days of gestation. The expression level was presented as relative expressions (fold changes) over the lowest value (the lowest level was assigned a value of 1) after normalization to β-actin expression.
Figure 4
Figure 4
Expression of milk fat globule-EGF-factor 8 (MFG-E8) in CD34+ cells in the fetal liver. (a and b) Flow cytometric analysis of CD34+ cells in the developing liver before (a) and after (b) purification with anti-CD34 antibody. (c) quantitative PCR (qPCR) analysis of sorted cells for the expression of Cd34. Cd34 expression in the CD34+ fraction was presented as relative expressions (fold changes) over CD34− fraction after normalization to β-actin expression. *P<0.05. (d and e) real-time reverse transcription-polymerase chain reaction (RT-PCR) (d) and qPCR (e) analyses of sorted CD34+ and CD34− fractions for the expression of Mfg-e8. Note that only a CD34+ fraction expresses Mfg-e8. P<0.05. Mfg-e8 expression in the CD34+ fraction was presented as relative expressions (fold changes) over CD34− fraction after normalization to β-actin expression.
Figure 5
Figure 5
Milk fat globule-EGF-factor 8 (MFG-E8) expression in adult bone marrow. (a) Immunohistochemical analysis of bone marrow sections with antibodies against c-kit and MFG-E8. Note that c-kit+ cells are located in the near sinusoids and these cells did not express MFG-E8. S, sinusoid. Enlarged images of the boxed region are shown in the upper right corner of each panel. (b) Expression of MFG-E8 in F4/80+ macrophages in the bone marrow of adult mice. Scale bar: a=50 μm; b=20 μm.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

References

    1. Carrascosa C, Obula RG, Missiaglia E, Lehr HA, Delorenzi M, Frattini M, et al. MFG-E8/lactadherin regulates cyclins D1/D3 expression and enhances the tumorigenic potential of mammary epithelial cells. Oncogene. 2012;31:1521–1532. - PubMed
    1. Ensslin MA, Shur BD. Identification of mouse sperm SED1, a bimotif EGF repeat and discoidin-domain protein involved in sperm-egg binding. Cell. 2003;114:405–417. - PubMed
    1. Ye H, Li B, Subramanian V, Choi BH, Liang Y, Harikishore A, et al. NMR solution structure of C2 domain of MFG-E8 and insights into its molecular recognition with phosphatidylserine. Biochim Biophys Acta. 2013;1828:1083–1093. - PubMed
    1. Andersen MH, Berglund L, Rasmussen JT, Petersen TE. Bovine PAS-6/7 binds alpha v beta 5 integrins and anionic phospholipids through two domains. Biochemistry. 1997;36:5441–5446. - PubMed
    1. Ensslin MA, Shur BD. The EGF repeat and discoidin domain protein, SED1/MFG-E8, is required for mammary gland branching morphogenesis. Proc Natl Acad Sci USA. 2007;104:2715–2720. - PMC - PubMed

Publication types