Onset of maternal arterial blood flow and placental oxidative stress. A possible factor in human early pregnancy failure

Abstract

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.

Figure 1.

Oxygen-free radicals are produced through the leakage of electrons from electron transport chains in the mitochondria and endoplasmic reticulum on to molecular oxygen. The superoxide anion generated is not freely diffusable through cell membranes and must be dismutated in situ by either Cu/Zn or Mn SOD. Although H₂O₂ is not a free radical it can react with O₂^·− to form the extremely reactive hydroxyl radical. Catalase and GPX must therefore operate in concert with the SODs to keep concentrations at physiological levels.

Figure 2.

A: The Paratrend multiparameter probe has a diameter of 0.5 mm and contains PO₂, PCO₂, pH, and temperature sensors. B: Sample output from the Paratrend probe illustrating the stability of the pH, PCO₂, and PO₂ measurements obtained. In this example, at 60 days of gestation, the probe was first introduced into the extra-embryonic coelom (EEC), and was then withdrawn through the placental chorionic plate (CP) into the placental intervillous space (IVS) and then finally into the endometrium (END) underlying the placenta. The probe rapidly stabilizes in each new environment, and thereafter there is a slight downward trend in the oxygen measurements. This most likely reflects oxygen consumption by the Clark electrode depleting the concentration in the immediate vicinity of the probe.

Figure 3.

Bivariate scattergram of the placental and uterine PO₂ measurements against gestational age. The curves were fitted with the LOWESS technique with a tension of 60. The PO₂ in the placenta rises steeply between 10 to 12 weeks, reflecting the onset of free maternal blood flow into the intervillous space.

Figure 4.

Bivariate scattergrams of the placental activities of the antioxidant enzymes and the concentration of reduced glutathione against gestational age. All showed a significant correlation except for total SOD activity (Table 1) ▶ , and lines were fitted using the LOWESS technique. a: GPX activity, LOWESS tension 70. b: Reduced glutathione concentration, LOWESS tension 70. c: Catalase activity, LOWESS tension 75. d: Total SOD activity.

Figure 5.

Representative Northern blots for the antioxidant enzymes. A significant increase in expression with gestational age was observed for catalase, GPX, and Cu/ZnSOD but not for MnSOD.

Figure 6.

Immunohistochemical localization of inducible Hsp 70 expression using fluorescein isothiocyanate-labeled secondary antibodies in villi at 6 weeks (a), 9 weeks (b), and 13 weeks gestational age (c). d: Negative control and nitrotyrosines with chromogenic substrate at 6 weeks (e), 9 weeks (f), and 13 weeks gestational age (g). h: Negative control. There is strong immunolabeling for both at 9 weeks, particularly in the syncytiotrophoblast (arrows).

Figure 7.

Scattergram of the mean fluorescence intensity of inducible Hsp 70 expression in the syncytiotrophoblast at different gestational ages. Two separate runs were performed, indicated by the symbols. The sections in each run were all immunolabeled together and viewed on the same day with the confocal microscope settings remaining constant. Both runs display a peak of Hsp 70i expression at 8 to 9 weeks, indicating a period of oxidative stress.

Figure 8.

Electron micrographs of syncytiotrophoblastic mitochondria (arrows) at different gestational ages demonstrating the distortion of the cristae and dilatation of the intracristal space that coincides with Hsp 70 expression and the formation of nitrotyrosine residues at 6 weeks (a), 10 weeks (b), 14 weeks (c). d: Scattergram plot of the percentage mitochondrial volume occupied by intracristal space at different gestational ages. The line was fitted using the LOWESS technique with a tension of 66.

Figure 9.

Summary diagram illustrating the origin and possible effects of the syncytiotrophoblastic oxidative stress. Factors that may modulate the degree of stress are indicated by the dashed lines.