Monia Cecati

Placental thrombomodulin expression in recurrent miscarriage

BackgroundEarly pregnancy loss can be associated with trophoblast insufficiency and coagulation defects. Thrombomodulin is an endothelial-associated anticoagulant protein involved in the control of hemostasis and inflammation at the vascular beds and its also a cofactor of the protein C anticoagulant pathway.DiscussionWe evaluate the Thrombomodulin expression in placental tissue from spontaneous recurrent miscarriage and voluntary abortion as controls. Thrombomodulin mRNA was determined using real-time quantitative polymerase chain reaction. Reduced expression levels of thrombomodulin were found in recurrent miscarriage group compared to controls (1.82-fold of reduction), that corresponds to a reduction of 45% (from control group Delta CT) of thrombomodulin expression in spontaneous miscarriage group respect the control groups.SummaryWe cannot state at present the exact meaning of a reduced expression of Thrombomodulin in placental tissue. Further studies are needed to elucidate the biological pathway of this important factor in the physiopathology of the trophoblast and in reproductive biology.

HLA-G and pregnancy adverse outcomes

There is growing evidence that pregnancy complications such as preeclampsia, recurrent pregnancy loss (RPL), IUGR, and premature birth could be associated with abnormal immunologic interactions at the fetal-maternal interface. The restricted expression of HLA-G to the subpopulation of trophoblast cells which invade the uterus has generated much interest. The alternative splicing of HLA-G primary transcript, gives origin to seven isoforms, including both membrane-bound forms (HLA-G1, G2, G3, G4) and soluble forms (sHLA-G: sHLA-G5, G6, G7). sHLA-G consists predominantly of sHLA-G1 after its shedding by metalloproteinases, and secreted sHLA-G5 representing the quantitatively dominating and full-length isoforms. HLA-G expression and HLA-G genetic variations in both the mother and the embryo/fetus may be important for pregnancy outcome. It is also intuitively apparent that a gene with putative immunosuppressive and immunotolerant potential might be functional in both the mother and the embryo/fetus/placenta. Reduced or aberrant HLA-G expression seems to be associated with certain complications of pregnancy, among which preeclampsia and possibly the risk of miscarriage, and that this may be further linked to HLA-G polymorphisms. Most of the studies aimed at assessing the role of HLA-G in pregnant diseases have considered only the maternal genotype and ignored the contribution of the fetus. In this regard, the mother, placenta and the fetus form a synthesis. Therefore, studies on placental diseases should address HLA-G expression and genetic variations also to the fetus/placenta.

The involvement of inflammatory cytokines in the pathogenesis of recurrent miscarriage.

OBJECTIVE To investigate the inflammatory cytokine expression pattern in trophoblastic tissue from women with unexplained recurrent miscarriage (RM). STUDY DESIGN Trophoblasts were obtained during uterine evacuation from 11 women with RM and from 20 healthy pregnant women undergoing elective termination of pregnancy, who served as controls. The array was performed using GEArray Q Series Human Inflammatory Cytokines & Receptors Gene Array HS-015 membranes. Data were confirmed by quantitative real-time PCR. The Mann-Whitney U test was performed for statistical analysis. RESULTS Microarray analysis identified three genes that were differentially expressed between RM patients and controls. We observed significant downregulation of Transforming Growth Factor beta 3 (TGF-β3) and Interleukin 25 (IL-25) (5-fold reduction and 2.5-fold reduction, respectively) and significant upregulation of CD-25, also known as Interleukin 2 receptor alpha (IL-2RA) (7-fold increase) in women with RM compared with controls. The median ΔC(t) of TGF-β3 was 8.2 (interquartile range, 7.67-8.9) in RM patients vs. 5.85 (interquartile range, 5.3-6.09) in controls; the median ΔC(t) of IL-25 was 5.18 (interquartile range, 4.46-5.76) in RM patients vs. 3.85 (interquartile range, 3.6-4.51) in controls, and the median ΔC(t) of CD-25 was 9.62 (interquartile range, 7.81-12.42) in RM patients vs. 12.44 (interquartile range, 11.02-13.86) in controls. DISCUSSION Our results suggest that the immunological and inflammatory regulation mechanisms of the placental environment play a key role in recurrent miscarriage. The observed trophoblast cytokine expression pattern at the maternal-fetal interface confirms the immunotrophic theory, as demonstrated by a switch from a T-helper-1 (Th1) profile to a T-helper-2 (Th2) profile in women who experience recurrent miscarriages.

Unexplained fetal loss: the fetal side of thrombophilia

Carrier status of the fetus for factor V polymorphism or double homozygosity for mutant alleles of the PAI-1 4 G/4 G and MTHFR T677 T polymorphisms must be considered risk factors for intrauterine fetal death. The clinical implications of these data need to be addressed in a prospective study to confirm our preliminary data and to answer the question of whether or not double homozygous individuals should be treated with low molecular-weight heparin and/or low-dose aspirin.

The role of p38α mitogen-activated protein kinase gene in the HELLP syndrome

Mitogen-activated protein kinase (MAPK) p38α was shown to be implicated in the organogenesis of the placenta, and such placental alteration is crucial for the development of hemolysis, elevated liver enzymes, and low platelets (HELLP) syndrome. We aimed to analyze for the first time human placental expression of MAPK p38α in pregnancies complicated by HELLP. The placental expression of MAPK p38α was investigated by semiquantitative polymerase chain reaction using cDNA extracted from placental tissue of 15 pregnancies with HELLP syndrome and 15 gestational age-matched controls. Seven patients with HELLP also had intrauterine fetal growth restriction (IUGR). In placenta from pregnancy complicated by HELLP, the expression of MAPK p38α is significantly decreased compared to the group with normal pregnancy (p < 0.001), while no difference was found between the HELLP and HELLP with IUGR subpopulations. Our study shows for the first time that MAPK p38α is expressed in the human placenta. Pregnancies with placental dysfunction and hypertensive complications are characterized by a significantly decreased expression of MAPK p38α. Our observations suggest that p38 MAPK signaling may be essential in placental angiogenesis and functioning.

Clues to apoptosis pathway involvement in hemolysis, elevated liver enzyme, and low platelet (HELLP) syndrome and intrauterine growth restriction (IUGR)

Objective: The neurotrophin family comprises molecules involved in growth, differentiation, survival, regeneration, normal functions of the neuronal system, and in angiogenesis. We have investigated the expression pattern of neurotrophic signaling molecules in pregnancies complicated by elevated liver enzyme, and low platelet (HELLP) syndrome and intrauterine growth restriction (IUGR). Methods: Placentas from normal and pathological pregnancies were collected. Macroarray analysis was performed and the data were confirmed by real-time PCR. Results: Real-time PCR analyses (pathological vs. normal pregnancies) confirmed a significant down-regulation for IL-6, STAT3α, STAT3β, and Bcl-2. The expression of Mcl-1 isoform 1 (long) was significantly increased. Conclusions: We suggest that decreased expression of IL-6 could mean that abnormalities in the immunological system function involve inflammatory cytokines other than IL-6 in examined pathological pregnancies. The STAT3α and STAT3β down-regulation lead to a marked reduction of cellular transcriptional activity. Decreased expression of IL-6 is associated with a down-regulation of Bcl-2 but not of Mcl-1 isoform 1, suggesting that these two antiapoptotic proteins may function independently and that Mcl-1 may have a distinct role in controlling apoptotic pathway.

Placental expression of endothelial and inducible nitric oxide synthase and nitric oxide levels in patients with HELLP syndrome

OBJECTIVE To determine placental gene expression of endothelial and inducible nitric oxide synthases and measure nitric oxide levels in patients with hemolysis, elevated liver enzyme levels, and low platelet count syndrome. STUDY DESIGN Preterm placentas were obtained from 15 patients with hemolysis, elevated liver enzyme levels, and low platelet count syndrome and 30 controls matched for age, parity, and gestational age. mRNA levels were evaluated by real-time polymerase chain reaction, whereas nitric oxide and peroxynitrite production was measured by a commercially available kit. RESULTS Placental gene expression of inducible nitric oxide and endothelial nitric oxide synthases were significantly lower in the hemolysis, elevated liver enzyme levels, and low platelet count syndrome group than in controls, whereas nitric oxide and peroxynitrite production were significantly higher in hemolysis, elevated liver enzyme levels, and low platelet count syndrome compared with controls. CONCLUSION The reduced endothelial nitric oxide and inducible nitric oxide synthases gene expression in women with hemolysis, elevated liver enzyme levels, and low platelet count syndrome may indicate extreme placental dysfunction that is unable to compensate the endothelial derangement and the related hypertension. The higher nitric oxide formation found in hemolysis, elevated liver enzyme levels, and low platelet count syndrome placentas could be explained as a counteraction to the impaired fetoplacental perfusion, typical of the syndrome.

Potential Role of Placental Klotho in the Pathogenesis of Preeclampsia

The aim of this study was to analyze the placental expression and allele status of promoter region of Klotho in association with preeclampsia, which represents the most common hypertensive disease of pregnancy. Klotho mRNA and protein levels were determined using real-time PCR and Western blot, respectively, in placental tissue samples obtained from 34 patients affected with preeclampsia and 34 controls. A PCR-based genotyping analysis was carried out in the promoter region of Klotho gene. Moreover, expression levels of pluripotency markers, Nanog and Oct4, and telomere length were assessed using real-time PCR. Klotho mRNA and protein levels were reduced in preeclamptic placentas compared with controls. −744delA single-nucleotide polymorphism was significantly associated with preeclampsia. In pathological placentas, there was a downregulation of pluripotency markers and a reduced telomere length. This study is the first to evaluate the placental expression level of Klotho in association with preeclampsia. Further analyses will clarify its role in the pathogenesis of this pregnancy hypertensive disorder.

PP035. Placental klotho protein in preeclampsia: A possible link to long term outcomes

INTRODUCTION An aging-suppressor gene, klotho, is a candidate factor for vascular disease because its deficiency leads to impaired endothelium-dependent vasodilation and impaired angiogenesis. Although klotho protein is predominantly expressed in the kidney, it is detected in a limited number of other tissues, such as the placenta, ovary, prostate gland, and small intestine. This protein is involved in several metabolic pathways such as calcium and phosphate homeostasis, the insulin-like growth factor 1 (IGF-1), apoptosis, angiotensin-II-induced events in the kidney and oxidative stress. OBJECTIVES The aim was to assess the expression of the klotho gene in the placenta from pregnancies affected by severe preeclampsia. METHODS Placentas were collected from normal pregnancies (n=12) and pregnancies complicated by preeclampsia (n=12), matched for gestational age. Klotho mRNA and protein were determined using real-time quantitative polymerase chain reaction (PCR) and Western blot, respectively. RESULTS Real-Time PCR analyses demonstrated a significant (p=0.005) 83% down-regulation of Klotho in patients with Preeclampsia versus Controls. Results of Western Blot agreed with those from Real-Time PCR. CONCLUSION Klotho mRNA expression in the placenta is decreased in preeclamptic pregnancies. Given its role in cardiovascular disease in aging, it may link preeclamptic mothers and their offsprings to long term cardiovascular outcomes.

NNMT (nicotinamide N -methyltransferase)

Review on NNMT (nicotinamide N-methyltransferase), with data on DNA, on the protein encoded, and where the gene is implicated.