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

Alkaline phosphatase attenuates LPS-induced early pregnancy defects.

<p>Mice treated with vehicle, MPLA, bIAP, LPS or bIAP + LPS were analyzed for: % of mice with embryo implantation sites (EISs; A); Number of EISs (B); Wet weight of individual EIS (C). Morphological appearance (D) and histological analysis (E) of day 8 uteri from mice treated with vehicle, MPLA, bIAP, LPS or bIAP plus LPS. Bovine IAP (bIAP; 150 units) was administered intravenously (tail vein) 5 min before LPS (100 μg) injection. Mice in bIAP and bIAP + LPS groups received an additional injection of bIAP (i.p., 5 units) on day 5 (1800 h) and days 6 and 7 (0900 h) of pregnancy. * Results are presented as mean ± SEM. p<0.05 (ANOVA followed by Tukey test).</p

Primers used for gene cloning and qRT-PCR.

<p>Primers used for gene cloning and qRT-PCR.</p

Expression of <i>MyD88</i> and <i>Trif</i> mRNAs in the early pregnant uterus.

<p>Uterine horns or implantation sites were collected from gestational days 1 to 8. (A) The levels of <i>MyD88</i> and <i>Trif</i> mRNA were determined by qRT-PCR, normalized to the housekeeping gene <i>Rpl7</i> and presented as the mean ± SD of three separate experiments (*,^#p < 0.05; one-way ANOVA and Tukey’s test). Cell specific expression of <i>MyD88</i> (B) and <i>Trif</i> (C) mRNAs was determined 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>MyD88</i> or <i>Trif</i> mRNA accumulations. bl, blastocyst; em, embryo; ge, glandular epithelium; le, luminal epithelium; pdz, primary decidual zone; s, stroma; sdz, secondary decidual zone.</p

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

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