Adrian Watson

Onset of Maternal Arterial Blood Flow and Placental Oxidative Stress : A Possible Factor in Human Early Pregnancy Failure

The aim was to measure changes in the oxygen tension within the human placenta associated with onset of the maternal arterial circulation at the end of the first trimester of pregnancy, and the impact on placental tissues. Using a multiparameter probe we established that the oxygen tension rises steeply from <20 mmHg at 8 weeks of gestation to >50 mmHg at 12 weeks. This rise coincides with morphological changes in the uterine arteries that allow free flow of maternal blood into the placenta, and is associated with increases in the mRNA concentrations and activities of the antioxidant enzymes catalase, glutathione peroxidase, and manganese and copper/zinc superoxide dismutase within placental tissues. Between 8 to 9 weeks there is a sharp peak of expression of the inducible form of heat shock protein 70, formation of nitrotyrosine residues, and derangement of the mitochondrial cristae within the syncytiotrophoblast. We conclude that a burst of oxidative stress occurs in the normal placenta as the maternal circulation is established. We speculate that this may serve a physiological role in stimulating normal placental differentiation, but may also be a factor in the pathogenesis of pre-eclampsia and early pregnancy failure if antioxidant defenses are depleted.

Variations in expression of copper/zinc superoxide dismutase in villous trophoblast of the human placenta with gestational age

This study investigated expression of the key antioxidant enzyme copper/zinc superoxide dismutase in the villous trophoblast of the human placenta at different gestational ages from 8 weeks (last menstrual period) to term. Immunostaining for the enzyme was observed in the cytotrophoblast cells at all stages. Staining was generally absent from the syncytiotrophoblast at 8 weeks, except for small isolated areas close to the basal surface. The size and location of these areas suggested they were the result of recent cytotrophoblastic fusion. By 10 weeks, examples were more frequent and diffuse staining throughout most of the syncytiotrophoblast was observed at 12 weeks. The intensity of the immunostaining within the syncytiotrophoblast continued to increase until 14 weeks, by which time it matched generally that within the cytotrophoblast cells. A similar pattern of staining was observed within term material. These results are entirely consistent with the hypothesis that the oxygen tension within the intervillous space is low throughout the first trimester of pregnancy. They support the idea that an effective maternal circulation to the human placenta is only established at the start of the second trimester.

Changes in concentration, localization and activity of catalase within the human placenta during early gestation

Using villous tissue from accurately dated gestational age placentae, this study identified significant changes in the protein concentration, enzyme activity and localization of catalase, an enzyme responsible for the intracellular metabolism of hydrogen peroxide, during the first and early second trimester of pregnancy. Enzyme activity was found to increase approximately threefold between weeks 6 and 17, with the greatest increase between 12 and 17 weeks. Immunostaining of tissue sections was supportive of these findings, demonstrating a progressively stronger signal between weeks 6 and 17. Immunostaining also demonstrated that the main cell types expressing catalase were the cytotrophoblast cells as well as a subset of the stromal cells. Between 13-17 weeks gestation, however, it was possible to detect catalase within the syncytiotrophoblast also, although with a much reduced intensity of staining. At the ultrastructural level, immunogold labelling of catalase clearly showed that staining was predominately compartmentalized within peroxisomes, although non-peroxisomal staining was also seen. Immunoreactivity also demonstrated, via morphological identification, that the stromal cells containing detectable levels of catalase were placental macrophages (Hofbauer cells). These results are in agreement with the proposal that the placenta exists in a physiologically low oxygen environment during the early part of gestation. In this environment oxidative activity of the sort resulting in the generation of hydrogen peroxide would presumably be suppressed, thereby limiting the requirement for catalase until oxygen tension begins to rise.

A MICROSCOPICAL STUDY OF WOUND REPAIR IN THE HUMAN PLACENTA

In order to fulfill its many functions as the selective interface between maternal and fetal circulations it is imperative that the human placenta remains intact and in good operational order. That damage of some sort occurs during its short but extremely active life seems inevitable given the dynamic environment in which the placenta exists, and evidence has accumulated that disruption is indeed a regular event. The implications of such damage, one could speculate, may impact on functions such as transport and hormone secretion as well as mutual protection against attack by maternal and fetal immune systems. Consequently, it would seem a theoretical necessity for discontinuities in the placenta surface to be repaired as soon as possible. We have used a combination of ex vivo observation, in vitro modelling, immunohistochemistry and correlative microscopy to provide evidence for a wound response in the placenta and to begin dissecting the detail of how this may operate. Evidence for small lesions caused by fusion and subsequent tearing of the syncytiotrophoblast in vivo, as well as plugging of such wounds by underlying cells is shown. We also identify a putative role for migratory cytotrophoblasts in the healing of larger scale injuries and demonstrate that certain molecules, common to wound repair in other tissues, appear to be involved in placenta repair also. Taken together these results clearly show that the human placenta is capable of a degree of self‐maintenance by activating what appears to be an endogenous wound healing mechanism. Microsc. Res. Tech. 42:351–368, 1998.

AN IN VITRO MODEL FOR THE STUDY OF WOUND HEALING IN FIRST TRIMESTER HUMAN PLACENTA

Abstract.The placenta operates as a vital interface between the mother and fetus. In addition to facilitating fetal nourishment, it acts as a barrier both to potentially deleterious agents and to contact between their two immune systems. As a consequence, damage to the placenta, even on a relatively small scale, could be very dangerous to the fetus. Therefore, wound repair mechanisms are likely to be of great importance in ensuring that an intact placental barrier is re-established as soon as possible. By use of an in vitro method for injuring and subsequently culturing small pieces of first trimester villous tissue, we have observed a number of indications that a wound response is initiated. Pronounced expression of transforming growth factor-β1, heavy infiltration of macrophages and late deposition of tenascin in the region of the wound all provide good evidence of some form of healing activity. Furthermore, we have noted that these indicators are suggestive of ’adult-type’ rather than ’fetal-type’ repair processes.

Evidence for oxygen-derived free radical mediated damage to first trimester human trophoblast in vitro

Summary We have found that, despite considerable experimentation with conditions, we are unable to maintain first trimester syncytiotrophoblast in a viable state in organ culture (conventional conditions i.e. in buffered culture medium supplemented with serum, maintained at 37°C/21% oxygen/5% carbon dioxide). Evidence for syncytiotrophoblastic degeneration includes: parallel release of LDH and HCG, widespread NΣ-Dansyl-1-Lysine uptake, indicating cell death, and extensive vacuolation observed via electron microscopy. These results suggest syncytial death occurs within hours of culture initiation. Despite this the cytotrophoblasts and stromal cell populations showed good viability for a number of days as shown by BrdU incorporation. When investigating the effect of oxygen tension on tissue survival we found that lipid peroxidation was considerably increased under ambient (21%) rather than low (2.5%) oxygen conditions. Furthermore, keeping early villous tissue at 2.5% oxygen reduced the rate of HCG as well as LDH release and also improved the ultrastructural appearance of the syncytiotrophoblast. We subsequently discovered that mitochondrial activity within the syncytiotrophoblast declined to virtually zero within six hours of cultureat 21% O2 but was retained for this period at 2.5% O2. Second trimester tissue, however, showed mitochondrial activity following 6 hours of culture at 21% O2. These results suggest that the syncytiotrophoblast is acutely sensitive to oxygen during the first trimester but becomes more resistant by 14 weeks gestational age, probably due to an increase in the capacity of the mechanisms which defend cells against oxidative stress.