It has been long suggested that placental hypoxia could play a role in PM-associated FGR but direct evidence for such a role has been lacking. We tested this hypothesis by looking for molecular evidence of placental hypoxia in PM.
Cellular response to hypoxia is orchestrated by the transcription factor HIF-1, which modifies the expression of several genes, in particular angiogenic factors. Using a very reliable and sensitive quantitative real time RT-PCR approach, we could not associate any clinical or biological features of PM with a transcription profile characteristic of a molecular response to hypoxia when addressing the whole placental tissue.
The absence of transcriptional evidence for placental hypoxia in PM was further substantiated by the lack of association between the clinical and biological features of PM and expression patterns of hypoxia-specific markers as evidenced by immunohistochemistry. The staining patterns obtained were similar to those previously described [27
]. When we used a previously validated scoring process [35
] to quantify the expression of these markers, we found no evidence for a hypoxia-specific expression pattern in any of the groups. Because only 2 samples were classified as LBW, we also used an alternative definition of LBW as a birth weight lower than the 5th
centile of a normal foetal growth curve [40
] to compare appropriate (n=32) and small (n=7) for gestational age infants and showed that they had similar IHC profiles (data not shown).
Taken together, these results strongly argue against a role for placental hypoxia in PM pathogenesis.
In contrast to a recent study that showed that PM was associated with an increase in sFlt-1 mRNA levels in normotensive primigravidae and in malaria-infected placentae with intervillositis compared with control placentae [41
], we did not find any difference in sFlt-1 RNA levels between the different groups when addressing either the whole tissue or specifically the SCT. Differences in the recruitment criteria (we focused on normotensive women, and included all gravidities, rather than just primigravidae) and in the technical approaches used (the Flt-1 antibody we used only recognizes the membrane-bound form, unlike the ones the authors used which binds both membrane-bound and soluble forms) could explain these conflicting results. Moreover, each study was relatively small (around 60 women), and suffers from some limitations in power. Further investigation of the interactions between malaria and hypertension and pre-eclampsia in first pregnancies may be warranted.
Since all the samples in our cohort presented similar transcription and expression profiles, it was possible that they could all be hypoxic as a consequence of labour or tissue handling. However, since we included only placentae from children with a normal Apgar score, labour-induced acute hypoxia is unlikely. Moreover, since HIF-1α transcript levels have been shown to change very rapidly after the onset of cellular hypoxia, if all samples had suffered from hypoxia as a consequence of labour or tissue handling, all would have expressed similar levels of HIF-1α transcript. But we did find a difference in HIF-1α SCT transcript levels between malaria-infected placentae and controls, which further rules out the possibility of a general labour-induced hypoxia.
The increased HIF-1α transcript levels in the SCT of malaria-infected placentae could reflect local inflammation [42
]. Indeed, hypoxia-induced HIF-1α up-regulation is thought to occur at the level of protein stability (post-transcriptional). Thus, higher HIF-1α transcript levels in the syncytium of malaria-infected placentae are likely to be a consequence of local inflammation that could be both the underlying cause of the molecular and protein changes noted, and the driving force behind malaria-associated LBW. Because similar HIF-1α transcripts levels were found in malaria cases in the presence or absence of monocyte infiltrates, which are markers of chronic inflammation, the increase in HIF-1α transcript levels might be due to a more acute inflammation or to the presence of infected erythrocytes. Testing of this hypothesis requires further study.
We could not associate any of the biological or clinical features of PM with a transcription or expression profile characteristic of a response to hypoxia. This argues against a role for placental hypoxia in PM-associated FGR pathogenesis. Although the role of placental hypoxia in the pathogenesis of FGR has been well described, particularly in animal models [43
], it is not universally implicated in FGR. For example, in several studies there was no difference in the expression of VEGF between normal and FGR-affected placentae [44
Our study does not rule out the possibility of fetal hypoxia, which could decrease fetal growth. Kingdom and Kaufmann described a situation leading to fetal hypoxia in the absence of placental hypoxia when the fetoplacental perfusion is inadequate. This is known as post-placental hypoxia [47
]. However, one study did not find evidence of FGR-associated fetal hypoxia [47
]. Thus, assessment of placental and fetal blood flow throughout pregnancy [48
] combined with arterial gas analysis of cord blood appears to be essential to adequately address the impact of a potential fetal hypoxia on intra-uterine growth during PM.