α5β1, αVβ3 and the platelet-associated integrin αIIbβ3 coordinately regulate adhesion and migration of differentiating mouse trophoblast cells

AbstractDuring blastocyst implantation, interaction between integrins on the apical surface of the trophoblast and extracellular matrix (ECM) in the endometrium anchors the embryo to the uterine wall. Strong adhesion of the blastocyst to fibronectin (FN) requires integrin signaling initiated by exogenous fibronectin. However, it is not known how integrin signaling enhances blastocyst adhesion. We present new evidence that the integrin, αIIbβ3, plays a key role in trophoblast adhesion to fibronectin during mouse peri-implantation development. Trafficking of αIIb to the apical surface of the trophoblast increased dramatically after blastocysts were exposed to fibronectin, whereas other fibronectin-binding integrins, α5β1 and αVβ3, were resident at the apical surface before ligand exposure. Functional comparisons among the three integrins revealed that ligation of α5β1 most efficiently strengthened blastocyst fibronectin-binding activity, while subsequent trophoblast cell migration was dependent primarily on the β3-class integrins. In vivo, αIIb was highly expressed by invasive trophoblast cells in the ectoplacental cone and trophoblast giant cells of the parietal yolk sac. These data demonstrate that trafficking of αIIb regulates adhesion between trophoblast cells and fibronectin as invasion of the endometrium commences

Alkaline phosphatase protects lipopolysaccharide-induced early pregnancy defects in mice.

Excessive cytokine inflammatory response due to chronic or superphysiological level of microbial infection during pregnancy leads to pregnancy complications such as early pregnancy defects/loss and preterm birth. Bacterial toxin lipopolysaccharide (LPS), long recognized as a potent proinflammatory mediator, has been identified as a risk factor for pregnancy complications. Alkaline phosphatase (AP) isozymes have been shown to detoxify LPS by dephosphorylation. In this study, we examined the role of alkaline phosphatase (AP) in mitigating LPS-induced early pregnancy complications in mice. We found that 1) the uterus prior to implantation and implantation sites following embryo implantation produce LPS recognition and dephosphorylation molecules TLR4 and tissue non-specific AP (TNAP) isozyme, respectively; 2) uterine TNAP isozyme dephosphorylates LPS at its sites of production; 3) while LPS administration following embryo implantation elicits proinflammatory cytokine mRNA levels at the embryo implantation sites (EISs) and causes early pregnancy loss, dephosphorylated LPS neither triggers proinflammatory cytokine mRNA levels at the EISs nor induces pregnancy complications; 4) AP isozyme supplementation to accelerate LPS detoxification attenuates LPS-induced pregnancy complications following embryo implantation. These findings suggest that a LPS dephosphorylation strategy using AP isozyme may have a unique therapeutic potential to mitigate LPS- or Gram-negative bacteria-induced pregnancy complications in at-risk women

Basement membrane ultrastructure and component localization data from uterine tissues during early mouse pregnancy

Basement membranes (BMs) are specialized extracellular scaffolds that provide architecture and modulate cell behaviors in tissues, such as fat, muscle, endothelium, endometrium, and decidua. Properties of BMs are maintained in homeostasis for most adult tissues. However, BM ultrastructure, composition, and localization are rapidly altered in select uterine tissues that are reprogrammed during pregnancy to enable early maternal-embryo interactions. Here, our data exhibit both static and dynamic BMs that were tracked in mouse uterine tissues during pre-, peri-, and postimplantation periods of pregnancy. The data exhibit spatial-temporal patterns of BM property regulation that coincide with the progression of adapted physiology. Further interpretation and discussion of these data in this article are described in the associated research article titled, “Embryo implantation triggers dynamic spatiotemporal expression of the basement membrane toolkit during uterine reprogramming” (C.R. Jones-Paris, S. Paria, T. Berg, J. Saus, G. Bhave, B.C. Paria, B.G. Hudson, 2016) [1]

Effects of LPS on uterine <i>MyD88</i>, <i>Trif</i>, <i>Tnfα</i>, <i>Il6</i> and <i>Il1β</i> mRNA expressions.

<p>Relative expression of <i>MyD88</i> (*p < 0.05) and <i>Trif</i> (*p < 0.05) mRNAs in ovariectomized uteri (A) and day 5 EISs (C), respectively. Relative expression of <i>Tnfα</i> (*p < 0.05), <i>Il6</i> (^#p < 0.05) and <i>Il1β</i> (⁺p < 0.05) mRNA in ovariectomized uteri (B) and day 5 EISs (D), respectively. All uteri were collected 6 h after vehicle, LPS (100 μg) or MPLA (100 μg) injection. Data were normalized to <i>Rpl7</i> and expressed as mean ± SD (n = 4), and one-way ANOVA followed by Tukey’s test, was used for statistical analysis.</p

Accumulation of bIAP isozyme in the decidua following its systemic injection.

<p>Alkaline phosphatase (AP) histochemical staining in sections from the liver and embryo implantation site (EIS) from either vehicle or bIAP-treated day 7 pregnant mice. AP staining in the absence (left 4 panels) or presence (right 4 panels) of levamisole in the EIS (top, 40X) and liver (bottom, 100X) from the bIAP-treated mouse. pv = portal vein</p

Expressions of <i>Alpl</i> mRNA and alkaline phosphatase (AP) activity in the early pregnant uterus.

<p>(A) The levels of <i>Alpl</i> mRNA were determined by qRT-PCR. Results were normalized to the housekeeping gene <i>Rpl7</i> and presented as the mean ± SD of three separate experiments, with different letters (a, b, c) indicating statistical difference (p < 0.05; one-way ANOVA and Tukey’s test). (B) Cell-type specific localization of <i>Alpl</i> mRNA was determined by <i>in situ</i> hybridization (n = 3). Inserts show higher magnification (200X) of <i>Alpl</i> mRNA accumulations. (C) AP activity patterns in the periimplantation uterus (n = 6). (D) Levamisole and L-phenylalanine were used as inhibitors of tissue-nonspecific AP (TNAP) and tissue-specific AP (TSAP), respectively. (E) TNAP activity localization in cells of the deciduum and deciduomata (n = 5). The embryo-induced decidua (deciduum) was collected on day 6 of pregnancy. The oil-induced decidua (deciduomata) was collected 48 h after intrauterine instillation of oil in day 4 pseudopregnant mouse. bl, blastocyst; em, embryo; ge, glandular epithelium; le, luminal epithelium; pdz, primary decidual zone; s, stroma; sdz, secondary decidual zone.</p

Localization of <i>Cd14</i> (A) and <i>Tlr4</i> (B) mRNAs in the early pregnant uterus.

<p>Uterine horns or implantation sites were collected from gestational days 1 to 8 for mRNA localization by <i>in situ</i> hybridization. Dark-field photographs were representative of three experiments. No hybridization signals were observed in sections hybridized with sense probes. Inserts show higher magnification (200X) of <i>Cd14</i> or <i>Tlr4</i> mRNA accumulations. bl, blastocyst; em, embryo; ge, glandular epithelium; le, luminal epithelium; pdz, primary decidual zone; s, stroma; sdz, secondary decidual zone.</p