Effects of vitamin E and C on placental oxidative stress: an in vitro evidence for the potential therapeutic or prophylactic treatment of preeclampsia.

Preeclampsia (PE) is a multisystem disorder that remains a major cause of maternal and foetal morbidity and death. To date, no treatment has been found that prevents the development of the disease. Endothelial dysfunction is considered to underlie its clinical manifestations, such as maternal hypertension, proteinuria and edema; and oxidative stress has been increasingly postulated as a major contributor to endothelial dysfunction in PE. A large body of research has investigated the potential role of antioxidant nutrients in the prevention of PE in women at high increased risk of the disease. Therefore, the present study was primary designed to assess the potential benefit of antioxidant supplementation on markers of placental oxidative stress in an in vitro model of PE, since we previously found that endothelin-1 (ET-1) is able to trigger the placental secretion of stress molecules. In this regard, we evaluated the effects of vitamin C, vitamin E and N-acetylcysteine (NAC), alone or in combination, in placental villi culture after exposure to ET-1. The effect of antioxidant nutrients on trophoblast cells proliferation and vitality was also evaluated. The results obtained suggest that in a pathophysiological condition, such as PE, the deleterious effect of reactive oxygen species may be counteract by an antioxidant therapy, and there is the need to investigate the optimum dosing and timing of antioxidants administration, since an inappropriate antioxidant treatment in pregnant women may have deleterious consequences, reducing placental cells proliferation until to cell death.

Endothelin-1 triggers placental oxidative stress pathways: putative role in preeclampsia.

CONTEXT: Preeclampsia (PE) is a disorder that occurs only during pregnancy. The placenta has a controlling role in this condition. Recent literature suggests that the oxidative stress is a component of PE and plays a main role in the link between decreased placental perfusion and the impaired function of maternal endothelium. OBJECTIVE: Because the human placenta expresses endothelin-1 (ET-1) and its circulating levels are high in pregnancies complicated with PE, the present study investigated the role of ET-1 on placental oxidative stress pathways. DESIGN: Human placental explants, JEG-3, and primary cytotrophoblast cells were cultured with increasing ET-1 concentrations for 6 and 24 h. SETTING: The study was conducted at tertiary clinical care centers in Siena and Padova, Italy. INTERVENTIONS: Human placental explants, JEG-3, and primary cytotrophoblast cells were used to test ET-1 effect. MAIN OUTCOME MEASURE(S): The main outcome measure was ET-1 mRNA and its receptor mRNAs, type A and B, detection by RT-PCR. The common markers of oxidative stress [malondialdehyde (MDA), glutathione (GSH), glutathione disulfide (GSSG), ascorbic acid (AA)] as well as cell proliferation and vitality were measured after stimulation periods. RESULTS: ET-1 inhibits cell proliferation and vitality and triggers oxidative stress in the human placenta by altering the balance between oxidant (increased MDA levels) and antioxidant (decreased GSH, GSSG, and AA) forces in favor of oxidation. CONCLUSIONS: Because MDA damages endothelial cells, whereas GSH, GSSG, and AA protect them, we postulate that ET-1 may be one of the key links between primary placental disorders and the systemic endothelial dysfunction of PE.

Reduced maternal plasma urocortin concentrations and impaired uterine artery blood flow at human mid pregnancy.

OBJECTIVE: Urocortin is a placental neuropeptide belonging to the family of corticotropin-releasing factors (CRFs), playing a role in the uteroplacental blood flow regulation through the binding to specific CRF receptors. Since CRF receptors are expressed in the uterine vascular bed of pregnant rats, and because urocortin has a relaxant effect on uterine vasculature, we evaluated mid-gestation plasma urocortin levels in women with impaired blood flow through uterine arteries. METHODS: Maternal plasma urocortin was assayed by specific radioimmunoassay and uterine artery resistance index (RI) by Doppler evaluation at 22-24 weeks' gestation in 57 healthy pregnant women, of which 29 showed a monolateral or bilateral uterine artery notch. Statistical analysis was performed by one-way analysis of variance (ANOVA), followed by post-hoc Tukey test for multiple comparison and Pearson correlation coefficient test. RESULTS: The mean uterine artery RI was significantly (P <.001) higher in women with a notch than healthy controls. Mean +/- SEM maternal plasma urocortin levels were significantly (P <.001) lower in women with unilateral (52.03 +/- 3.25 pg/mL) or bilateral (47.01 +/- 4.16 pg/mL) uterine artery notch than in healthy control pregnant women (84.01 +/- 3.5 pg/mL). While no difference was found in urocortin levels between patients with unilateral or bilateral uterine artery notch, urocortin concentrations inversely correlated with the mean RI (Pearson r = -0.7318; 95% confidence interval -0.8334 to -0.5822; P <.0001). CONCLUSIONS: The present findings suggest that reduced levels of circulating urocortin are associated with increased uterine artery resistances and support the hypothesis that urocortin may regulate uterine artery tone at mid gestation.

Impaired uterine artery blood flow at mid gestation and low levels of maternal plasma corticotropin-releasing factor.

OBJECTIVE: Corticotropin-releasing factor (CRF) is a placental neuropeptide that plays a role in the control of uteroplacental blood flow regulation. Because CRF has a relaxant effect on uterine vasculature in pregnant rats, we aimed to evaluate mid-gestation plasma CRF levels in women with impaired uterine artery blood flow. METHODS: Maternal plasma CRF was assayed by specific radioimmunoassay, and uterine artery resistance index (RI) was assessed by Doppler evaluation at 22-24 weeks' gestation in 55 healthy pregnant women, of whom 24 showed a unilateral or bilateral uterine artery notch, reflecting resistance. Statistical analysis was performed by the Kruskal-Wallis test followed by the post hoc Dunn's test and the Spearman rank test. RESULTS: The mean uterine artery RI was significantly (P <.001) higher in women with a notch than in healthy controls. Mean +/- standard error of the mean maternal plasma CRF levels were significantly lower in women with a unilateral (168.45 +/- 27.5 pg/mL; P <.01) or bilateral (186.07 +/- 34.5 pg/mL; P <.001) uterine artery notch than in healthy control pregnant women (375.06 +/- 21.77 pg/mL). Although no difference was found in CRF levels between patients with a unilateral or bilateral uterine artery notch, a significant inverse correlation was found between the mean RI and maternal plasma CRF levels (Spearman r = -0.6540; 95% confidence interval, -0.7865, -0.4640; P <.001). CONCLUSION: Reduced levels of circulating CRF were associated with increased uterine artery resistance, which supports the hypothesis that CRF may regulate uterine artery tone at mid gestation.

Paracrine regulation of endometrial function: interaction between progesterone and corticotropin-releasing factor (CRF) and activin A.

Under the influence of ovarian steroid hormones, endometrial cells aer able to produce a wide variety of growth factors and peptide hormones that area believed to promote: (1) physiological growth and differentiation during the endometrial cycle; (2) decidualization, an essential preparative event for establishment of pregnancy; and (3) pathological growth and differentiation in endometriosis and cancer. Among the local factors produced by the human endometrium, corticotropin-releasing factor (CRF) and activin A have been evaluated in terms of localization and effects. CRF is a neuropeptide expressed by the epithelial and stromal cells of the human endometrium in increasing amounts from the endometrial proliferative to the secretory phase. CRF expression also increases in the pregnant endometrium, from early in the pregnancy until term. CRF-type 1 receptor mRNA is only expressed by stromal cells. Progesterone induces CRF gene expression and release from decidualized cells and CRF decidualizes cultured stromal endometrial cells. Urocortin, a CRF-related peptide, has been identified in endometrial epithelial and stromal cells, and its function is still under investigation. Activin A is a growth factor expressed in increasing amounts throughout endometrial phases by both epithelial and stromal cells. This growth factor is secreted into the uterine cavity with higher levels in the secretory phase. Maternal decidua expresses activin A mRNA in increasing amounts from early pregnancy until term. Human endometrium also expresses activin-A receptors and follistatin, its binding protein. Activin A decidualizes cultured human endometrial stromal cells (an effect reversed by follistatin) and modulates embryonic trophoblast differentiation and adhesion. Activin A is expressed in endometriosis and endometrial adenocarcinoma.

Differentiation state affects morphine induced cell regulation in neuroblastoma cultured cells.

Neuroblastoma (NB) is the most common extracranial solid cancer in childhood and the most common cancer in infancy. Our purpose was to investigate in vitro how cancer cell survival occurs in presence of morphine in undifferentiated and differentiated SHSY-5Y human neuroblastoma cultured cell line. Exposure of differentiated cells to morphine dose-dependently induced apoptosis in these cells through c-Jun N-terminal kinase (JNK)/caspase pathway. Otherwise, morphine induced activation for mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) pathway, caused positive regulation of cell survival in undifferentiated cells. Therefore, cell differentiation state bimodally affects the cellular regulation activity triggered by morphine in isolated cultured neuroblastoma cells raising concerns about the application of morphine to this type of cancer patients.

Anti-tumor activity of the methanolic extracts of Salvia menthifolia

In the present research we investigated the anti-proliferative activity of Salvia menthifolia Ten. (formerly Salvia menthaefolia), Lamiaceae, on a glioblastoma cell line, since up to date poor therapeutic results have been reported for treatment of malignant glioblastoma. Methanol extracts from different anatomical parts of S. menthifolia were tested on DBTRG-05MG cell line by MTT assay. The most active primary stems extract was also evaluated for apoptosis induction. Results confirmed the anti-tumor property of all the organs and demonstrated that the primary stems extract induced apoptosis after 4 h with the highest values of DNA fragmentation after 6 to 24 h. Some extracts were also HPLC analyzed for polyphenols, althought activities could be due also to other constituents and to synergistic interactions. Rosmarinic acid, caffeic acid, luteolin-7-O-glucosyde and quercitrin were found in all the extracts. The good performance revealed for S. menthifolia towards this extremely aggressive human glioblastoma cell line confirms that the genus Salvia is a natural source of anti-tumor agents though there are great differences among the various species.

In vitro antiproliferative effect of six Salvia species on human tumor cell lines.

This study was designed to examine the in vitro antiproliferative activity of the methanol crude extracts of six Salvia species: Salvia dominica L. leaves, Salvia lanigera Desf. aerial parts, Salvia menthaefolia Ten. roots, Salvia palaestina Benth. aerial parts, Salvia sclarea L. roots and Salvia spinosa L. aerial parts. Extracts were screened for their possible antitumoral activity by MTT test on nine human cancer cell lines: glioblastoma (DBTRG-05MG, T98G, U-87MG), colorectal adenocarcinoma (WiDr and HT-29), prostate adenocarcinoma (MDA Pca2b), choriocarcinoma (JEG-3), endometrium adenocarcinoma (HEC-1A) and B lymphoblast (CIR). IC(50) values were determined for only five extracts and ranged from 90 to 400 microg/mL approximately. Salvia menthaefolia extract exhibited marked antiproliferative activity against all tumor cell lines showing lower IC(50) values, while S. spinosa, S. sclarea and S. dominica extracts showed a degree cytotoxic activity dependent on the cell line type. Finally S. palaestina extract revealed a moderate antiproliferative effect only against three cell lines. Salvia lanigera extract displayed toxic activity at all concentrations tested. The results strengthen the evidence that the genus Salvia could be considered a natural resource of potential antitumor agents.

Identification of genes regulated by interleukin-1beta in human endometrial stromal cells.

Interleukin-1beta (IL-1b) is an important immune regulatory factor that in human endometrium plays a role in both menstruation and implantation in the event of pregnancy. It promotes inflammatory-like processes and also stimulates tissue remodelling. We present a cDNA microarray study documenting the major effects of IL-1beta on gene expression in stromal cells from human endometrium. Endometrial stromal cells from five normal healthy women at the mid secretory phase were cultured with or without IL-1beta at 50 and 500 pg/ml for 48 h. cDNA microarrays were used to compare the levels of gene expression in total RNA isolated from cells stimulated with IL-1beta. These cDNA arrays were produced containing 15 164 sequence-verified clones, which included genes known to be important in angiogenesis, immune modulators, apoptosis, cell signalling, extra-cellular matrix (ECM) remodelling and cell cycle regulation. Genes which were regulated by IL-1beta were identified by analysis of the microarray data using the Significance Analysis of Microarrays software package. Upregulated (n = 23) and downregulated (n = 6) different genes were observed, which changed at least 3-fold, at a false discovery rate of less than 2% (P < 0.02). Our results have identified genes regulated by IL-1beta, which are involved in leukocyte recruitment, ECM remodelling and other cellular functions. Changes in three genes, IL-8, colony-stimulating factor 2 and aldoketo reductase family 1 member 1, which were upregulated by IL-1beta, were verified using real-time PCR. Novel functions regulated by IL-1beta in endometrium, including genes involved in free radical protection, and fatty acid metabolism were also identified. These results also provide new insights into the role of IL-1beta in disorders of the endometrium, especially in implantation-related infertility and endometriosis, in which this cytokine plays a major role.

Placental stress factors and maternal-fetal adaptive response: the corticotropin-releasing factor family.

The placenta and its accessory membranes amnion and chorion undertake the role of intermediary barriers and active messengers in the maternal-fetal dialog. They synthesize, metabolize, and serve as target to numerous hormones that regulate maternal and fetal physiology during pregnancy. Among these factors, corticotropin-releasing factor (CRF) has been one of the more investigated in the last decade. Increasing evidence indicates that in the event of acute or chronic metabolic, physical, or infectious stress, maternal or fetal physiologic and pathologic conditions may influence placental secretion of CRF. The current opinion is that the placenta actually takes part in a stress syndrome by releasing CRF, which may help to influence uterine perfusion, maternal metabolism, fluid balance, and possibly uterine contractility, thereby protecting the fetus from a hostile environment.