2 Aims of the work

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Mammalian pregnancy is a parabiotic union of two genetically different individuals, the fetus and the mother. It is now known that some degree of systemic or uterine inflammation is necessary for both normal implantation and pregnancy, but if this inflammation becomes too excessive it can cause pregnancy complications such as abortion. Heme oxygenase-1 (HO-1), the enzyme responsible for the degradation of free heme, plays a key role in inflammatory processes. Viewing pregnancy mainly as an inflammatory process let us hypothesized that HO-1 may play an important role in pregnancy. Therefore, the main aim of this work was to analyze the role of HO-1 in the different processes related to pregnancy by means of functional studies employing in vivo as well as in vitro models.

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The present work is divided didactically in 4 parts: in vivo up-regulation of HO-1, upregulation of HO-1 in T cells, effect of HO-1 in the survival and differentiation of trophoblast cells and analysis of the effect of a partial or total loss of Hmox1 on implantation and fertility. The objectives of each part are indicated below:

I. In vivo up-regulation of HO-1

Since it is postulated that a systemic up-regulation of HO-1 is enough to prevent the rejection of allografts, and because the fetus is normally considered as a semi-allogeneic allograft, the aims of this part of the work were:

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  1. To specifically up-regulate HO-1 in mice undergoing abortion through adenoviral gene transfer.
  2. To determine whether this up-regulation improves pregnancy outcome.
  3. To investigate possible mechanisms by which HO-1 would exert its action.

II. Up-regulation of HO-1 in T cells

Considering the cytoprotective characteristics of the HO-1 molecule, the aim of this second part of the work was to generate T cells over-expressing HO1, and to test their potential in the inhibition and modulation of the immune response in vitro.

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The fulfilment of these aims included:

  1. Generation of modified T-lymphocytes over-expressing HO-1 (THO-1-lymphocytes).
  2. Analysis of the function of the generated THO-1-lymphocytes in vitro.

III. Effect of HO-1 on the survival and differentiation of trophoblast cells

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The use of the trophoblast stem cell line Rcho-1 helps to elucidate the mechanisms necessary for the differentiation of these cells into giant trophoblast cells, and therefore to understand the mechanisms underlying placentation. The aims of this part of the study were:

  1. To analyze the effect of an HO-1 inducer (CoPPIX) or inhibitor (ZnPPIX) on the survival of the trophoblast stem cell line.
  2. To analyze the ability of the Rcho-1 cells to differentiate under the influence of CoPPIX or ZnPPIX, in order to determine the role of HO-1 during trophoblast cell differentiation.

IV. Analysis of the effect of partial or total loss of Hmox-1 on implantation and ferti l ity

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As animals deficient in Hmox1 are not able to yield progeny and the mating of mice heterozygous for Hmox-1 does not yield the expected Mendelian rate, further aims of this work were:

  1. To analyze the pregnancy outcome in different combinations of Hmox1 +/+ , Hmox1 +/- or Hmox1 -/- mice in order to determine whether Hmox1 expression is important at the maternal or paternal side or in both.
  2. To determine whether the lack of Hmox1 has any effect on the fertility of the mice by performing in vitro fertilization.
  3. To analyze whether Hmox1 is necessary in the oocytes, in the maternal uterus, or in both, for implantation to occur.


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