Cell free hemoglobin in the fetoplacental circulation: a novel cause of fetal growth restriction?

Cell free hemoglobin impairs vascular function and blood flow in adult cardiovascular disease. In this study, we investigated the hypothesis that free fetal hemoglobin (fHbF) compromises vascular integrity and function in the fetoplacental circulation, contributing to the increased vascular resistance associated with fetal growth restriction (FGR). Women with normal and FGR pregnancies were recruited and their placentas collected freshly postpartum. FGR fetal capillaries showed evidence of erythrocyte vascular packing and extravasation. Fetal cord blood fHbF levels were higher in FGR than in normal pregnancies ( P < 0.05) and the elevation of fHbF in relation to heme oxygenase-1 suggests a failure of expected catabolic compensation, which occurs in adults. During ex vivo placental perfusion, pathophysiological fHbF concentrations significantly increased fetal-side microcirculatory resistance ( P < 0.05). fHbF sequestered NO in acute and chronic exposure models ( P < 0.001), and fHbF-primed placental endothelial cells developed a proinflammatory phenotype, demonstrated by activation of NF-κB pathway, generation of IL-1α and TNF-α (both P < 0.05), uncontrolled angiogenesis, and disruption of endothelial cell flow alignment. Elevated fHbF contributes to increased fetoplacental vascular resistance and impaired endothelial protection. This unrecognized mechanism for fetal compromise offers a novel insight into FGR as well as a potential explanation for associated poor fetal outcomes such as fetal demise and stillbirth.-Brook, A., Hoaksey, A., Gurung, R., Yoong, E. E. C., Sneyd, R., Baynes, G. C., Bischof, H., Jones, S., Higgins, L. E., Jones, C., Greenwood, S. L., Jones, R. L., Gram, M., Lang, I., Desoye, G., Myers, J., Schneider, H., Hansson, S. R., Crocker, I. P., Brownbill, P. Cell free hemoglobin in the fetoplacental circulation: a novel cause of fetal growth restriction?

The impact of a human IGF-II analog ([Leu27]IGF-II) on fetal growth in a mouse model of fetal growth restriction.

Enhancing placental insulin-like growth factor (IGF) availability appears to be an attractive strategy for improving outcomes in fetal growth restriction (FGR). Our approach was the novel use of [Leu(27)]IGF-II, a human IGF-II analog that binds the IGF-II clearance receptor IGF-IIR in fetal growth-restricted (FGR) mice. We hypothesized that the impact of [Leu(27)]IGF-II infusion in C57BL/6J (wild-type) and endothelial nitric oxide synthase knockout (eNOS(-/-); FGR) mice would be to enhance fetal growth and investigated this from mid- to late gestation; 1 mg·kg(-1)·day(-1) [Leu(27)]IGF-II was delivered via a subcutaneous miniosmotic pump from E12.5 to E18.5. Fetal and placental weights recorded at E18.5 were used to generate frequency distribution curves; fetuses <5th centile were deemed growth restricted. Placentas were harvested for immunohistochemical analysis of the IGF system, and maternal serum was collected for measurement of exogenously administered IGF-II. In WT pregnancies, [Leu(27)]IGF-II treatment halved the number of FGR fetuses, reduced fetal(P = 0.028) and placental weight variations (P = 0.0032), and increased the numbers of pups close to the mean fetal weight (131 vs. 112 pups within 1 SD). Mixed-model analysis confirmed litter size to be negatively correlated with fetal and placental weight and showed that [Leu(27)]IGF-II preferentially improved fetal weight in the largest litters, as defined by number. Unidirectional (14C)MeAIB transfer per gram placenta (System A amino acid transporter activity) was inversely correlated with fetal weight in [Leu(27)]IGF-II-treated WT animals (P < 0.01). In eNOS(-/-) mice, [Leu(27)]IGF-II reduced the number of FGR fetuses(1 vs. 5 in the untreated group). The observed reduction in FGR pup numbers in both C57 and eNOS(-/-) litters suggests the use of this analog as a means of standardizing and rescuing fetal growth, preferentially in the smallest offspring.

Integration of computational modeling with membrane transport studies reveals new insights into amino acid exchange transport mechanisms.

Uptake of system L amino acid substrates into isolated placental plasma membrane vesicles in the absence of opposing side amino acid (zero-trans uptake) is incompatible with the concept of obligatory exchange, where influx of amino acid is coupled to efflux. We therefore hypothesized that system L amino acid exchange transporters are not fully obligatory and/or that amino acids are initially present inside the vesicles. To address this, we combined computational modeling with vesicle transport assays and transporter localization studies to investigate the mechanisms mediating [(14)C]L-serine (a system L substrate) transport into human placental microvillous plasma membrane (MVM) vesicles. The carrier model provided a quantitative framework to test the 2 hypotheses that l-serine transport occurs by either obligate exchange or nonobligate exchange coupled with facilitated transport (mixed transport model). The computational model could only account for experimental [(14)C]L-serine uptake data when the transporter was not exclusively in exchange mode, best described by the mixed transport model. MVM vesicle isolates contained endogenous amino acids allowing for potential contribution to zero-trans uptake. Both L-type amino acid transporter (LAT)1 and LAT2 subtypes of system L were distributed to MVM, with L-serine transport attributed to LAT2. These findings suggest that exchange transporters do not function exclusively as obligate exchangers.

Uterine vasculature remodeling in human pregnancy involves functional macrochimerism by endothelial colony forming cells of fetal origin.

The potency of adult-derived circulating progenitor endothelial colony forming cells (ECFCs) is drastically surpassed by their fetal counterparts. Human pregnancy is associated with robust intensification of blood flow and vascular expansion in the uterus, crucial for placental perfusion and fetal supply. Here, we investigate whether fetal ECFCs transmigrate to maternal bloodstream and home to locations of maternal vasculogenesis, primarily the pregnant uterus. In the first instance, endothelial-like cells, originating from mouse fetuses expressing paternal eGFP, were identified within uterine endothelia. Subsequently, LacZ or enhanced green fluorescent protein (eGFP)-labeled human fetal ECFCs, transplanted into immunodeficient (NOD/SCID) fetuses on D15.5 pregnancy, showed similar integration into the mouse uterus by term. Mature endothelial controls (human umbilical vein endothelial cells), similarly introduced, were unequivocally absent. In humans, SRY was detected in 6 of 12 myometrial microvessels obtained from women delivering male babies. The copy number was calculated at 175 [IQR 149-471] fetal cells per millimeter square endothelium, constituting 12.5% of maternal vessel lumina. Cross-sections of similar human vessels, hybridized for Y-chromosome, positively identified endothelial-associated fetal cells. It appears that through ECFC donation, fetuses assist maternal uterine vascular expansion in pregnancy, potentiating placental perfusion and consequently their own fetal supply. In addition to fetal growth, this cellular mechanism holds implications for materno-fetal immune interactions and long-term maternal vascular health.

Adapting in vitro dual perfusion of the human placenta to soluble oxygen tensions associated with normal and pre-eclamptic pregnancy.

For decades, superoxic ex vivo dual perfusion of the human placental lobule has been used as a model to study the physiology and metabolism of the placenta. The aim of this study was to further develop the technique to enable perfusion at soluble oxygen concentrations similar to those in normal pregnancy (normoxia) and in pre-eclampsia (PE; hypoxia). Our design involved reducing the mean soluble oxygen tension in the maternal-side intervillous space (IVS) perfusate to 5-7% and <3% for normoxia and hypoxia, respectively, while providing a more ubiquitous delivery of perfusate into the IVS, using 22 maternal-side cannulae. We achieved quasi-steady states in [O2](fetal venous (soluble)), which were statistically different between the two adaptations at t=150 to t=240 min of dual perfusion (2.1, 1.2, 2.8 and 0.4, 0.0, 1.5%; median, 25th, 75th percentiles, n=20 and 24 readings in n=5 and n=6 lobules, normoxic and hypoxic perfusion, respectively; P<0.001, Mann-Whitney U-test). Lactate dehydrogenase (LDH) levels in fetal and maternal venous outflow perfusates were unaffected by the adaptations. There was also no difference in tissue lactate release between the two adaptations. Glucose consumption from the fetal circulation and maternal-side 'venous' pyruvate release were higher under normoxic conditions, indicative of a greater metabolic flux through glycolysis. Furthermore, there was greater release of the hypoxic-sensitive marker, macrophage inflammatory protein-1α, into the maternal venous perfusate in the hypoxic model. Also, during hypoxic perfusion, we found that fetal-side venous placental growth factor (PlGF) levels were higher compared with normoxic perfusion. We conclude that these ex vivo adapted methods of placental perfusion provide a means of studying aspects of placental metabolism in relation to normal oxygenation and hypoxia-associated pregnancy disease.

Intra-uterine growth restriction is associated with increased apoptosis and altered expression of proteins in the p53 pathway in villous trophoblast.

Intrauterine growth restriction (IUGR) affects 3-8% of pregnancies and is associated with altered cell turnover in the villous trophoblast, an essential functional cell type of the human placenta. The intrinsic pathway of apoptosis, particularly p53, is important in regulating placental cell turnover in response to damage. We hypothesised that expression of proteins in the p53 pathway in placental tissue would be altered in IUGR. Expression of constituents of the p53 pathway was assessed using real-time PCR, Western blotting and immunohistochemistry. p53 mRNA and protein expression was increased in IUGR, which localised to the syncytiotrophoblast. Similar changes were noted in p21 and Bax expression. There was no change in the expression of Mdm2, Bak and Bcl-2. The association between altered trophoblast cell turnover in IUGR and increased p53 expression is reminiscent of that following exposure to hypoxia. These observations provide further insight into the potential pathogenesis of IUGR. Further research is required to elicit the role and interactions of p53 and its place in the pathogenesis of IUGR.

IGF2 actions on trophoblast in human placenta are regulated by the insulin-like growth factor 2 receptor, which can function as both a signaling and clearance receptor.

Insulin-like growth factor 2 (IGF2) enhances proliferation and survival of human first-trimester cytotrophoblasts (CTB) by signaling through the insulin-like growth factor 1 receptor (IGF1R). However, the role of the IGF2 receptor (IGF2R) in regulating trophoblast kinetics is unclear: It could act as a clearance receptor for trafficking excess ligand to lysosomes for degradation and/or directly mediate IGF2 signaling. We used an IGF2R knockdown strategy in BeWo cells and placental villous explants to investigate trophoblast proliferation and survival in response to stimulation by IGF. Both IGF1 and IGF2 significantly (P < 0.001) increased mitosis and reduced apoptosis in serum-starved BeWo cells. Small interfering RNA (siRNA)-mediated knockdown of IGF2R further enhanced IGF2-stimulated mitosis (P < 0.01), and IGF2-mediated rescue of apoptosis (P < 0.001) in these cells. Leu(27)IGF2, an IGF2 analogue that binds to IGF2R but not IGF1R, also protected IGF2R-expressing BeWo cells from apoptosis but did not increase mitosis. IGF treatment of term placental villous explants with reduced syncytial expression of IGF2R increased CTB proliferation (P < 0.001) and decreased apoptosis (P < 0.01) compared to untreated controls. Moreover, IGF2-mediated rescue of CTB apoptosis was significantly greater than that in tissue with normal IGF2R expression. Leu(27)IGF2 promoted mitogenesis and survival only in explants with intact IGF2R expression. Given that altered CTB turnover is observed in pregnancies complicated by fetal growth restriction, the development of strategies to manipulate the IGF2R signaling axis in the syncytiotrophoblast may provide a therapeutic avenue for treating this condition.

Immunomodulatory Therapy Reduces the Severity of Placental Lesions in Chronic Histiocytic Intervillositis.

Chronic histiocytic intervillositis (CHI) is a rare, but highly recurrent inflammatory placental lesion wherein maternal macrophages infiltrate the intervillous space. Pregnancies with CHI are at high risk of fetal growth restriction, miscarriage or stillbirth. Presently, the diagnosis can only be made after histopathological examination of the placenta. Given its proposed immunological etiology, current treatments include aspirin, heparin, and immunomodulatory agents. However, the rationale for these medications is largely based upon small case series and reports as there is a lack of larger studies investigating treatment efficacy. Therefore, this study sought to determine whether inclusion of immunomodulatory medications was effective at reducing the severity of lesions and improving pregnancy outcomes in subsequent pregnancies. Thirty-three women with a history of CHI in at least one pregnancy (index case) were identified retrospectively through medical records. Twenty-eight participants presented with a first subsequent pregnancy and a further 11 with a second subsequent pregnancy at a specialist clinic for pregnancy after loss. Data on maternal demographics, medical history, medication, pregnancy outcome, and placental pathology was collected and compared between pregnancies. Twenty-seven (69%) subsequent pregnancies were treated with at least one or both of prednisolone and hydroxychloroquine. Inclusion of at least one immunomodulatory agent in treatment regimen resulted in an almost 25% increase in overall livebirth rate (61.5 vs. 86.2%). In women treated with immunomodulatory medication a greater proportion of placentas had reduced severity of lesions compared to those treated without (86.7 vs. 33.3%, respectively). A reduction in CHI severity was associated with a 62.3% improvement in livebirth rate compared to those where severity remained unchanged in relation to the index case. These data provide preliminary evidence that the use of immunomodulatory medication in the management of CHI improves histopathological lesions and the chance of livebirth in subsequent pregnancies. Due to CHI's rarity and ethical and feasibility issues, randomized controlled trials in affected women are challenging to conduct. As a result, collaboration between centers is required in future to increase study sample sizes and elucidate the mechanisms of hydroxychloroquine and prednisolone in reducing pathology.

Chronic histiocytic intervillositis: A breakdown in immune tolerance comparable to allograft rejection?

Chronic histiocytic intervillositis (CHI) is a pregnancy disorder characterized by infiltration of maternal macrophages into the intervillous space of the human placenta, often with accompanying perivillous fibrin deposition. CHI is associated strongly with foetal growth restriction and increased risk of miscarriage and stillbirth. Although rare, affecting 6 in every 10 000 pregnancies beyond 12 weeks' gestation, the rate of recurrence is high at 25%-100%. To date, diagnosis of CHI can only be made post-delivery upon examination of the placenta due to a lack of diagnostic biomarkers, and criteria vary across publications. No treatment options have shown proven efficacy, and CHI remains a serious obstetric conundrum. Although its underlying aetiology is unclear, due to the presence of maternal macrophages and the reported increased incidence in women with autoimmune disease, CHI is hypothesized to be an inappropriate immune response to the semi-allogeneic foetus. Given this lack of understanding, treatment approaches remain experimental with limited rationale. However, there is recent evidence that immunosuppression and antithrombotic therapies may be effective in preventing recurrence of associated adverse pregnancy outcomes. With similarities noted between the pathological features of CHI and acute rejection of solid organ transplants, further investigation of this hypothesis may provide a basis for tackling CHI and other immune-related placental conditions. This review will explore parallels between CHI and allograft rejection and identify areas requiring further confirmation and exploitation of this comparison.

Does altered oxygenation or reactive oxygen species alter cell turnover of BeWo choriocarcinoma cells?

This study assessed the effect of 20 and 6% ambient oxygen (O(2)) or 5-50 micromol/l hydrogen peroxide (H(2)O(2)) on apoptosis, necrosis, proliferation and fusion of BeWo cells. The expression of p53, Mdm2 and Bax was assessed by western blotting. Apoptosis was increased in cells cultured in 6% O(2) tension and 50 micromol/l H(2)O(2) (P < 0.05, P < 0.01 by ADP:ATP ratio). In the same conditions, cell viability as estimated by the MTT assay was decreased (6% O(2) P < 0.01, 50 micromol/l H(2)O(2) P < 0.05). Human chorionic gonadotrophin secretion was decreased by culture in 6%O(2) and 50 micromol/l H(2)O(2) (P < 0.05). Cell fusion was also decreased by treatment with 50 micromol/l H(2)O(2) (P < 0.05). Treatment with 50 micromol/l H(2)O(2) was associated with increased expression of p53 and decreased expression of Mdm2 (P < 0.05). This study provides evidence that BeWo cell turnover is altered following exposure to hypoxia or ROS. It is concluded that BeWo cell culture is an appropriate model for investigating the regulation of trophoblast cell turnover. In addition, these data support a role for p53 in mediating altered trophoblast cell turnover in response to oxidative stress.

Oxygen and the liberation of placental factors responsible for vascular compromise.

Maternal endothelial activation in pre-eclampsia is attributed to the release of unknown factors from a hypoperfused placenta. To further characterize these factors, we have used a serum-free placental villous explant culture model and investigated the effect of the liberated soluble factors produced on human endothelial cell cultures. Term placental villous explants from uncomplicated pregnancies were cultured for 4 days in 20, 6 or 1% O2 to mimic placental hyperoxia, normoxia and hypoxia. Medium collected from viable explants was applied to cultured human uterine microvascular endothelial cells. Medium conditioned by hypoxic explants caused a significant decrease in endothelial cell ATP levels and mitochondrial dehydrogenase activity, suggestive of a reduced metabolic rate. An additional reduction in mitochondrial membrane potential and increased endothelial cell death occurred as the oxygen concentration to which explants had been exposed decreased. Effects of the hypoxic explant medium were also seen ex vivo in a wire myography model of myometrial artery function, with increased vasoconstriction and attenuated vasodilation following exposure to hypoxic explant medium. These results suggest that hypoxia (1% O2) may stimulate the release of soluble factors from the placenta, which have an adverse effect on endothelial cell metabolism and mitochondrial integrity in vitro. These potentially pathogenic factors are now being characterized.

Uterine spiral artery remodeling involves endothelial apoptosis induced by extravillous trophoblasts through Fas/FasL interactions.

OBJECTIVE: Invasion of uterine spiral arteries by extravillous trophoblasts in the first trimester of pregnancy results in loss of endothelial and musculoelastic layers. This remodeling is crucial for an adequate blood supply to the fetus with a failure to remodel implicated in the etiology of the hypertensive disorder preeclampsia. The mechanism by which trophoblasts induce this key process is unknown. This study gives the first insights into the potential mechanisms involved. METHODS AND RESULTS: Spiral arteries were dissected from nonplacental bed biopsies obtained at Caesarean section, and a novel model was used to mimic in vivo events. Arteries were cultured with trophoblasts in the lumen, and apoptotic changes in the endothelial layer were detected after 20 hours, leading to loss of endothelium by 96 hours. In vitro, coculture experiments showed that trophoblasts stimulated apoptosis of primary decidual endothelial cells and an endothelial cell line. This was blocked by caspase inhibition and NOK2, a FasL blocking antibody. NOK2 also abrogated trophoblast-induced endothelial apoptosis in the vessel model. CONCLUSIONS: Extravillous trophoblast induction of endothelial apoptosis is a possible mechanism by which the endothelium is removed, and vascular remodeling may occur in uterine spiral arteries. Fas/FasL interactions have an important role in trophoblast-induced endothelial apoptosis.

Excessive stimulation of poly(ADP-ribosyl)ation contributes to endothelial dysfunction in pre-eclampsia.

1. Pre-eclampsia is a serious pregnancy disorder associated with widespread activation of the maternal vascular endothelium. Recent evidence implicates a role for oxidative stress in the aetiology of this condition. 2. Reactive oxygen species, particularly superoxide anions, invokes endothelial cell activation through many pathways. Oxidant-induced cell injury triggers the activation of nuclear enzyme poly(ADP-ribose) polymerase (PARP) leading to endothelial dysfunction in various pathophysiological conditions (reperfusion, shock, diabetes). 3. We have studied whether the loss of endothelial function in pre-eclampsia is dependent on PARP activity. Endothelium-dependent responses of myometrial arteries were tested following exposure to either plasma from women with pre-eclampsia or normal pregnant women in the presence and absence of a novel potent inhibitor of PARP, PJ34. Additional effects of plasma and PJ34 inhibition were identified in microvascular endothelial cell cultures. 4. In myometrial arteries, PARP inhibition blocked the attenuation of endothelium-dependent responses following exposure to plasma from women with pre-eclampsia. In endothelial cell cultures, plasma from pre-eclamptics induced measurable oxidative stress and a concomitant increase in PARP activity and reduction in cellular ATP. Again, these biochemical changes were reversed by PJ34. 5. These results suggest that PARP activity plays a pathogenic role in the development of endothelial dysfunction in pre-eclampsia and promotes PARP inhibition as a potential therapy in this condition.

The influence of oxygen and tumor necrosis factor-alpha on the cellular kinetics of term placental villous explants in culture.

Explanted placental fragments may provide a more physiological in vitro model of component cell function than single cell isolates. We have characterized these fragments for cell turnover and have monitored responses from 14 normal placentas under conditions of exogenous TNFalpha and atypical oxygen concentrations (3% and 17%), conditions associated with abnormal pregnancy and an aberrant in utero environment. Explants were assessed for apoptotic morphology, immunolocalization of Mib-1 (a proliferation marker), caspase 3 activity (an apoptosis promoter), lactate dehydrogenase (a necrosis marker), and human chorionic gonadotrophin [hCG, a marker of cytotrophoblast (CT) differentiation]. Consistent with a reduction in hCG, explants under 17% O(2) (with and without TNFalpha) showed a progressive degeneration of syncytiotrophoblast (ST) (days 0-2) followed by a restoration of hCG (days 4-8) localized to newly differentiated but not syncytialized CTs. In 3% O(2), hCG showed the same initial decline but failed to recover thereafter. Proliferation dropped significantly in 17% O(2) but was restored and exaggerated sixfold in 3% O(2). All reductions in hCG were associated with cell death and caspase-3. Early apoptosis was linked with syncytial loss; later apoptosis (days 8-11) was localized to the non-ST. Prolonged exposure to TNFalpha (days 4-11) increased ST apoptosis and necrosis but 3% O(2) had no significant effect. These findings show that placental explants can accommodate many aspects of CT proliferation, differentiation, and ST apoptosis in culture. TNFalpha enhanced ST decline but 3% oxygen (compared with 17%) was associated with reduced CT differentiation and a strong shift towards proliferation. These outcomes may reflect previous morphological changes in compromised pregnancies and confirm a possible role for oxygen and TNFalpha in aberrant trophoblast turnover.

Functional characteristics of chorionic plate placental arteries from normal pregnant women and women with pre-eclampsia.

OBJECTIVES: We aimed to compare placental small artery function from women with pre-eclampsia and normal pregnancy. In particular, we wished to test the hypothesis that these arteries respond differently to an endothelium-dependent vasodilator, to the presence of nitric oxide, and to the presence of cyclic monophosphate nucleotides. METHODS: A small vessel wire myograph was used to study placental arteries (200 to 550 microm). Contractile function was assessed with vasopressin. Relaxation was assessed with the endothelium-dependent vasodilator, bradykinin, and the endothelium-independent vasodilators sodium nitroprusside (a nitric oxide donor) and papaverine (a phosphodiesterase inhibitor). RESULTS: The constrictor response to vasopressin did not differ between patient groups (p=0.79; repeated measures ANOVA). For both normal pregnancy and pre-eclampsia, the response of pre-constricted arteries to the endothelium-dependent vasodilator, bradykinin, was minimal. Vasorelaxation to sodium nitroprusside and papaverine was attenuated in pre-eclampsia compared to normal pregnancy (p=0.03 and p<0.001, respectively; repeated measures ANOVA). CONCLUSIONS: In pre-eclampsia, placental arteries exhibit an attenuated vasodilatory response to nitric oxide.

Preeclampsia is associated with alterations in the p53-pathway in villous trophoblast.

BACKGROUND: Preeclampsia (PE) is characterized by exaggerated apoptosis of the villous trophoblast of placental villi. Since p53 is a critical regulator of apoptosis we hypothesized that excessive apoptosis in PE is mediated by abnormal expression of proteins participating in the p53 pathway and that modulation of the p53 pathway alters trophoblast apoptosis in vitro. METHODS: Fresh placental villous tissue was collected from normal pregnancies and pregnancies complicated by PE; Western blotting and real-time PCR were performed on tissue lysate for protein and mRNA expression of p53 and downstream effector proteins, p21, Bax and caspases 3 and 8. To further assess the ability of p53 to modulate apoptosis within trophoblast, BeWo cells and placental villous tissue were exposed to the p53-activator, Nutlin-3, alone or in combination with the p53-inhibitor, Pifithrin-α (PFT-α). Equally, Mdm2 was knocked-down with siRNA. RESULTS: Protein expression of p53, p21 and Bax was significantly increased in pregnancies complicated by PE. Conversely, Mdm2 protein levels were significantly depleted in PE; immunohistochemistry showed these changes to be confined to trophoblast. Reduction in the negative feedback of p53 by Mdm2, using siRNA and Nutlin-3, caused an imbalance between p53 and Mdm2 that triggered apoptosis in term villous explants. In the case of Nutlin, this was attenuated by Pifithrin-α. CONCLUSIONS: These data illustrate the potential for an imbalance in p53 and Mdm2 expression to promote excessive apoptosis in villous trophoblast. The upstream regulation of p53 and Mdm2, with regard to exaggerated apoptosis and autophagy in PE, merits further investigation.

Endothelial colony-forming cells derived from pregnancies complicated by intrauterine growth restriction are fewer and have reduced vasculogenic capacity.

CONTEXT: Endothelial colony-forming cells (ECFCs) are the only putative endothelial progenitor cells capable of vasculogenesis, and their dysfunction may represent a risk factor for cardiovascular disease. Intrauterine growth restriction (IUGR) is a pregnancy-related disorder associated with long-term cardiovascular risk. OBJECTIVE: Our objective was to determine whether ECFCs derived from pregnancies complicated by IUGR exhibit altered vasculogenic potential. DESIGN AND SETTING: This was a prospective cohort study; patients were recruited at St. Mary's Hospital, Manchester, United Kingdom. PARTICIPANTS: Twenty-three women with normal pregnancies and 13 women with IUGR-complicated pregnancies at gestational ages above 37 weeks were included. MAIN OUTCOME MEASURES: Vasculogenic capacity of rigorously characterized ECFCs was investigated in vivo by measuring blood vessel formation in collagen/fibronectin gels implanted in mice; proliferative, migratory, and chemotactic abilities were assessed in cell culture. Placental uptake of fetal ECFCs, assessed by differences in arterial and venous cord blood content, was determined by flow cytometry. RESULTS: In vivo, IUGR ECFCs formed fewer blood vessels (P < .001) and capillaries (P = .001) compared with normal pregnancy-derived ECFCs. In culture conditions, IUGR ECFCs had reduced proliferation (P = .01) and migration (P = .007) and diminished chemotactic abilities to stromal cell-derived factor 1 (P = .007) coupled with reduced hypoxia-induced matrix metalloproteinase-2 release (P = .02). Finally, in IUGR pregnancies, the number of ECFCs was lower in arterial cord blood (P = .002) and placental uptake of cells was reduced (P < .001). CONCLUSIONS: ECFCs derived from IUGR cord blood are rarefied and dysfunctional, resulting in diminished vasculogenic potential; this could be a cause of placental dysfunction in IUGR, with long-term postnatal implications for cardiovascular function in offspring.

Endothelial progenitor cells: Their potential role in pregnancy and preeclampsia.

The normal maternal cardiovascular adaptation to pregnancy involves a complex physiologic response to the growing conceptus, including alterations in maternal vascular endothelial cells that contribute to a profound fall in total systemic vascular resistance. There is a large body of evidence that adverse changes in the vascular endothelium underlie the multisystemic maternal manifestations of the hypertensive pregnancy disorder preeclampsia. Our knowledge is incomplete regarding the mechanisms of adaptive endothelial changes of normal pregnancy, and why these changes are attenuated or fail in women who develop preeclampsia. Endothelial progenitor cells (EPCs) are a heterogeneous population of cells that exist in both the fetus and adult. These cells can be mobilized into the circulation by growth factors and can then support the health of the vascular endothelium by several mechanisms. This review highlights some of the current understanding of EPCs, their potential role in pregnancy, and emerging evidence for EPC dysfunction in preeclampsia. We speculate that interference with nitric oxide (NO)-driven mobilization or activity of EPCs in the maternal circulation partially contributes to the widespread endothelial dysfunction underlying the clinical manifestations of preeclampsia. Potential roles of EPCs in the placenta and fetus are also considered.

Placental apoptosis in health and disease.

Apoptosis, programmed cell death, is an essential feature of normal placental development but is exaggerated in association with placental disease. Placental development relies upon effective implantation and invasion of the maternal decidua by the placental trophoblast. In normal pregnancy, trophoblast apoptosis increases with placental growth and advancing gestation. However, apoptosis is notably exaggerated in the pregnancy complications, hydatidiform mole, pre-eclampsia, and intrauterine growth restriction (IUGR). Placental apoptosis may be initiated by a variety of stimuli, including hypoxia and oxidative stress. In common with other cell-types, trophoblast apoptosis follows the extrinsic or intrinsic pathways culminating in the activation of caspases. In contrast, the formation of apoptotic bodies is less clearly identified, but postulated by some to involve the clustering of apoptotic nuclei and liberation of this material into the maternal circulation. In addition to promoting a favorable maternal immune response, the release of this placental-derived material is thought to provoke the endothelial dysfunction of pre-eclampsia. Widespread apoptosis of the syncytiotrophoblast may also impair trophoblast function leading to the reduction in nutrient transport seen in IUGR. A clearer understanding of placental apoptosis and its regulation may provide new insights into placental pathologies, potentially suggesting therapeutic targets.