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Msx homeobox genes critically regulate embryo implantation by controlling paracrine signaling between uterine stroma and epithelium.

Nallasamy S, Li Q, Bagchi MK, Bagchi IC - PLoS Genet. (2012)

Bottom Line: The mammalian Msx homeobox genes, Msx1 and Msx2, encode transcription factors that control organogenesis and tissue interactions during embryonic development.We observed overlapping expression of these factors in uterine epithelial and stromal compartments of pregnant mice prior to embryo implantation.Increased canonical Wnt signaling in the stromal cells activated β-catenin, stimulating the production of a subset of fibroblast growth factors (FGFs) in these cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Comparative Biosciences, University of Illinois Urbana-Champaign, Urbana, Illinois, United States of America.

ABSTRACT
The mammalian Msx homeobox genes, Msx1 and Msx2, encode transcription factors that control organogenesis and tissue interactions during embryonic development. We observed overlapping expression of these factors in uterine epithelial and stromal compartments of pregnant mice prior to embryo implantation. Conditional ablation of both Msx1 and Msx2 in the uterus resulted in female infertility due to a failure in implantation. In these mutant mice (Msx1/2(d/d)), the uterine epithelium exhibited persistent proliferative activity and failed to attach to the embryos. Gene expression profiling of uterine epithelium and stroma of Msx1/2(d/d) mice revealed an elevated expression of several members of the Wnt gene family in the preimplantation uterus. Increased canonical Wnt signaling in the stromal cells activated β-catenin, stimulating the production of a subset of fibroblast growth factors (FGFs) in these cells. The secreted FGFs acted in a paracrine manner via the FGF receptors in the epithelium to promote epithelial proliferation, thereby preventing differentiation of this tissue and creating a non-receptive uterus refractory to implantation. Collectively, these findings delineate a unique signaling network, involving Msx1/2, Wnts, and FGFs, which operate in the uterus at the time of implantation to control the mesenchymal-epithelial dialogue critical for successful establishment of pregnancy.

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Enhanced proliferation in the uterine epithelium and lack of receptivity in Msx1d/dMsx2d/d mice.A. Immunohsitochemical localization of Ki67 in the uterine sections of Msx1f/fMsx2f/f (left panel, a and c) and Msx1d/dMsx2d/d (right panel, b and d) mice on day 4 of pregnancy. Panels a and b indicate lower magnification (20×) and c and d indicate higher magnification (40×). L and G indicate luminal epithelium and glandular epithelium respectively. B. Real-time PCR was performed to analyze the expression of glandular factors, Lif, Foxa2 and Spink3 in uteri of Msx1f/fMsx2f/f and Msx1d/dMsx2d/d mice on day 4 of pregnancy. The level of Ck18 was used as internal control to normalize gene expression. The data are represented as the mean fold induction ± SEM, ***p<0.0001. C. Transmission electron microscopy of uterine sections obtained from Msx1f/f Msx2f/f (left panel, a and b) and Msx1d/dMsx2d/d (right panel, c and d) mice on day 4 of pregnancy. Panels a and c indicate lower magnification (5Kx) and b and d indicate higher magnification (30Kx). D. Immunohistochemical analysis of Muc-1 expression in the uterine sections of Msx1f/fMsx2f/f (upper panel) and Msx1d/dMsx2d/d (lower panel) mice on day 1 (a and d), day 4 (b and e) and day 5 (c and f) of pregnancy. L indicates luminal epithelium.
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pgen-1002500-g004: Enhanced proliferation in the uterine epithelium and lack of receptivity in Msx1d/dMsx2d/d mice.A. Immunohsitochemical localization of Ki67 in the uterine sections of Msx1f/fMsx2f/f (left panel, a and c) and Msx1d/dMsx2d/d (right panel, b and d) mice on day 4 of pregnancy. Panels a and b indicate lower magnification (20×) and c and d indicate higher magnification (40×). L and G indicate luminal epithelium and glandular epithelium respectively. B. Real-time PCR was performed to analyze the expression of glandular factors, Lif, Foxa2 and Spink3 in uteri of Msx1f/fMsx2f/f and Msx1d/dMsx2d/d mice on day 4 of pregnancy. The level of Ck18 was used as internal control to normalize gene expression. The data are represented as the mean fold induction ± SEM, ***p<0.0001. C. Transmission electron microscopy of uterine sections obtained from Msx1f/f Msx2f/f (left panel, a and b) and Msx1d/dMsx2d/d (right panel, c and d) mice on day 4 of pregnancy. Panels a and c indicate lower magnification (5Kx) and b and d indicate higher magnification (30Kx). D. Immunohistochemical analysis of Muc-1 expression in the uterine sections of Msx1f/fMsx2f/f (upper panel) and Msx1d/dMsx2d/d (lower panel) mice on day 1 (a and d), day 4 (b and e) and day 5 (c and f) of pregnancy. L indicates luminal epithelium.

Mentions: A hallmark of the receptive state of normal pregnant uterus is the cessation of epithelial cell proliferation prior to implantation [1]–[4], [22]. Therefore, in Msx1f/fMsx2f/f mice, immunostaining of Ki67, a cell proliferation marker, was undetectable in the uterine luminal and glandular epithelium on day 4 of pregnancy (Figure 4A, panels a and c). However, uterine sections of Msx1d/dMsx2d/d mice exhibited robust immunostaining for Ki67 in the luminal and glandular epithelia (Figure 4A, panels b and d), indicating persistent epithelial cell proliferation on day 4 in the absence of Msx1 and Msx2. Previous studies indicated that the ability of the glandular epithelium to undergo differentiation and produce factors, including leukemia inhibitory factor (LIF) [23], Foxa2 [24], and Spink3 [25], is critical for implantation. As shown in Figure 4B, the expression of these factors was drastically reduced in uteri deficient of Msx1 and Msx2. Collectively, these findings indicated that persistent proliferation of luminal and glandular epithelia results in impaired epithelial transition from a proliferative to a non-proliferative state that allows proper differentiation. This impairment is a major contributor to implantation failure in Msx1d/dMsx2d/d mice.


Msx homeobox genes critically regulate embryo implantation by controlling paracrine signaling between uterine stroma and epithelium.

Nallasamy S, Li Q, Bagchi MK, Bagchi IC - PLoS Genet. (2012)

Enhanced proliferation in the uterine epithelium and lack of receptivity in Msx1d/dMsx2d/d mice.A. Immunohsitochemical localization of Ki67 in the uterine sections of Msx1f/fMsx2f/f (left panel, a and c) and Msx1d/dMsx2d/d (right panel, b and d) mice on day 4 of pregnancy. Panels a and b indicate lower magnification (20×) and c and d indicate higher magnification (40×). L and G indicate luminal epithelium and glandular epithelium respectively. B. Real-time PCR was performed to analyze the expression of glandular factors, Lif, Foxa2 and Spink3 in uteri of Msx1f/fMsx2f/f and Msx1d/dMsx2d/d mice on day 4 of pregnancy. The level of Ck18 was used as internal control to normalize gene expression. The data are represented as the mean fold induction ± SEM, ***p<0.0001. C. Transmission electron microscopy of uterine sections obtained from Msx1f/f Msx2f/f (left panel, a and b) and Msx1d/dMsx2d/d (right panel, c and d) mice on day 4 of pregnancy. Panels a and c indicate lower magnification (5Kx) and b and d indicate higher magnification (30Kx). D. Immunohistochemical analysis of Muc-1 expression in the uterine sections of Msx1f/fMsx2f/f (upper panel) and Msx1d/dMsx2d/d (lower panel) mice on day 1 (a and d), day 4 (b and e) and day 5 (c and f) of pregnancy. L indicates luminal epithelium.
© Copyright Policy
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getmorefigures.php?uid=PMC3285581&req=5

pgen-1002500-g004: Enhanced proliferation in the uterine epithelium and lack of receptivity in Msx1d/dMsx2d/d mice.A. Immunohsitochemical localization of Ki67 in the uterine sections of Msx1f/fMsx2f/f (left panel, a and c) and Msx1d/dMsx2d/d (right panel, b and d) mice on day 4 of pregnancy. Panels a and b indicate lower magnification (20×) and c and d indicate higher magnification (40×). L and G indicate luminal epithelium and glandular epithelium respectively. B. Real-time PCR was performed to analyze the expression of glandular factors, Lif, Foxa2 and Spink3 in uteri of Msx1f/fMsx2f/f and Msx1d/dMsx2d/d mice on day 4 of pregnancy. The level of Ck18 was used as internal control to normalize gene expression. The data are represented as the mean fold induction ± SEM, ***p<0.0001. C. Transmission electron microscopy of uterine sections obtained from Msx1f/f Msx2f/f (left panel, a and b) and Msx1d/dMsx2d/d (right panel, c and d) mice on day 4 of pregnancy. Panels a and c indicate lower magnification (5Kx) and b and d indicate higher magnification (30Kx). D. Immunohistochemical analysis of Muc-1 expression in the uterine sections of Msx1f/fMsx2f/f (upper panel) and Msx1d/dMsx2d/d (lower panel) mice on day 1 (a and d), day 4 (b and e) and day 5 (c and f) of pregnancy. L indicates luminal epithelium.
Mentions: A hallmark of the receptive state of normal pregnant uterus is the cessation of epithelial cell proliferation prior to implantation [1]–[4], [22]. Therefore, in Msx1f/fMsx2f/f mice, immunostaining of Ki67, a cell proliferation marker, was undetectable in the uterine luminal and glandular epithelium on day 4 of pregnancy (Figure 4A, panels a and c). However, uterine sections of Msx1d/dMsx2d/d mice exhibited robust immunostaining for Ki67 in the luminal and glandular epithelia (Figure 4A, panels b and d), indicating persistent epithelial cell proliferation on day 4 in the absence of Msx1 and Msx2. Previous studies indicated that the ability of the glandular epithelium to undergo differentiation and produce factors, including leukemia inhibitory factor (LIF) [23], Foxa2 [24], and Spink3 [25], is critical for implantation. As shown in Figure 4B, the expression of these factors was drastically reduced in uteri deficient of Msx1 and Msx2. Collectively, these findings indicated that persistent proliferation of luminal and glandular epithelia results in impaired epithelial transition from a proliferative to a non-proliferative state that allows proper differentiation. This impairment is a major contributor to implantation failure in Msx1d/dMsx2d/d mice.

Bottom Line: The mammalian Msx homeobox genes, Msx1 and Msx2, encode transcription factors that control organogenesis and tissue interactions during embryonic development.We observed overlapping expression of these factors in uterine epithelial and stromal compartments of pregnant mice prior to embryo implantation.Increased canonical Wnt signaling in the stromal cells activated β-catenin, stimulating the production of a subset of fibroblast growth factors (FGFs) in these cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Comparative Biosciences, University of Illinois Urbana-Champaign, Urbana, Illinois, United States of America.

ABSTRACT
The mammalian Msx homeobox genes, Msx1 and Msx2, encode transcription factors that control organogenesis and tissue interactions during embryonic development. We observed overlapping expression of these factors in uterine epithelial and stromal compartments of pregnant mice prior to embryo implantation. Conditional ablation of both Msx1 and Msx2 in the uterus resulted in female infertility due to a failure in implantation. In these mutant mice (Msx1/2(d/d)), the uterine epithelium exhibited persistent proliferative activity and failed to attach to the embryos. Gene expression profiling of uterine epithelium and stroma of Msx1/2(d/d) mice revealed an elevated expression of several members of the Wnt gene family in the preimplantation uterus. Increased canonical Wnt signaling in the stromal cells activated β-catenin, stimulating the production of a subset of fibroblast growth factors (FGFs) in these cells. The secreted FGFs acted in a paracrine manner via the FGF receptors in the epithelium to promote epithelial proliferation, thereby preventing differentiation of this tissue and creating a non-receptive uterus refractory to implantation. Collectively, these findings delineate a unique signaling network, involving Msx1/2, Wnts, and FGFs, which operate in the uterus at the time of implantation to control the mesenchymal-epithelial dialogue critical for successful establishment of pregnancy.

Show MeSH
Related in: MedlinePlus