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The myelin-associated protein Nogo-A is among the most potent neurite growth inhibitors in the adult CNS. Recently, Nogo-A expression was demonstrated in a number of neuronal subpopulations of the adult and developing CNS but at present, little is known about the expression of Nogo-A in the nigrostriatal system, a brain structure severely affected in Parkinson’s disease (PD). The present study sought to characterize the expression pattern of Nogo-A immunoreactive (ir) cells in the adult ventral mesencephalon of control rats and in the 6-hydroxydopamine (6-OHDA) rat model of PD. Immunohistochemical analyses of normal adult rat brain showed a distinct expression of Nogo-A in the ventral mesencephalon, with the highest level in the substantia nigra pars compacta (SNc) where it co-localized with dopaminergic neurons. Analyses conducted 1 week and 1 month after unilateral striatal injections of 6-OHDA disclosed a severe loss of the number of Nogo-A-ir cells in the SNc. Notably, at 1 week after treatment, more dopaminergic neurons expressing Nogo-A were affected by the 6-OHDA toxicity than Nogo-A-negative dopaminergic neurons. However, at later time points more of the surviving dopaminergic neurons expressed Nogo-A. In the striatum, both small and large Nogo-A-positive cells were detected. The large cells were identified as cholinergic interneurons. Our results suggest yet unidentified functions of Nogo-A in the CNS beyond the inhibition of axonal regeneration and plasticity, and may indicate a role for Nogo-A in PD. [ABSTRACT FROM AUTHOR]

Abstract: Shedding of syncytiotrophoblast microparticles (MPs) from placenta to maternal blood occurs in normal pregnancy and is enhanced during preeclampsia (PE). The syncytiotrophoblast synthesizes plasminogen activator inhibitors (PAIs) which regulate fibrinolysis, as well as soluble forms of the fms-like tyrosine kinase (sFlt-1) and endoglin, which exert anti-angiogenic actions. An increase in the ratio of PAI-1/PAI-2 and elevated levels of sFlt-1 and sEng in maternal serum are linked to placental damage and maternal endothelial cell dysfunction in PE. The goal of the current study was to determine whether MPs released to maternal perfusate during dual perfusion contain these factors associated with placental pathophysiology in PE. Initially, high levels of alkaline phosphatase activity and Annexin V binding were found in MPs isolated by sequential centrifugation of maternal perfusates at 10,000 and 150,000×g(10 K and 150 K MPs), indicating their plasma membrane origin. ELISA revealed the presence of these factors at the following relative levels: Eng>PAI-2⋙PAI-1>sFlt-1. Based on comparisons of their concentration in perfusates, MPs, and MP-free 150 K supernatants, we determined that MPs constitute a significant portion of Eng released by placenta. Flow cytometric analysis of 10 K MPs supported the levels of expression found by ELISA and indicated that Eng and PAI-2 were almost exclusively localized to the surface of MPs, a site with biological potential. These results indicate that MPs shed from the syncytial surface express factors which may alter the fibrinolytic and angiogenic balance at the maternal–fetal interface and play a role in the pathophysiology of PE. [Copyright &y& Elsevier]

Abstract: Plasminogen activator inhibitors (PAIs) play critical roles in regulating cellular invasion and fibrinolysis. An increase in the ratio of PAI-1/PAI-2 in placenta and maternal serum is suggested to result in excessive intervillous fibrin deposition and placental infarction in pregnancies complicated by preeclampsia (PE) and intrauterine growth restriction (IUGR). In the current study we used dual (maternal and fetal) perfusion of human term placentas to examine the release of PAIs to the intervillous space. ELISA revealed a significant time-dependent increase in total PAI-1 levels in maternal perfusate (MP) between 1 and 7h of perfusion. Conversely, PAI-2 levels decreased resulting in a 3-fold increase in the PAI-1/PAI-2 ratio in MP. Levels of PAI-1, but not PAI-2, in placental tissue extracts increased during perfusion. In perfusions carried out with xanthine and xanthine oxidase (X+XO), compounds used to generate reactive oxygen species (ROS), no time-dependent increase in total PAI-1 levels was observed. In addition, X+XO treatment promoted a 3-fold reduction in active PAI-1 levels in MP, indicating that ROS decrease PAI-1 release to MP. The finding of a time-dependent change in patterns of PAI expression and response to ROS indicates the utility of dual perfusion as a model to dissect mechanism(s) promoting aberrant fibrinolysis in pregnancies complicated by PE and IUGR. [Copyright &y& Elsevier]

Abstract: Normal placentation involves the development of an utero-placental circulation following the migration of the extravillous cytotrophoblasts into the decidua and invasion of the spiral arteries, which are thereby transformed into large vessels of low resistance. Given the documented role of the receptor tyrosine kinase EphB4 and its ligand ephrin-B2 in the establishment of the embryonal vascular network, we hypothesized that these molecules are also instrumental in the development of the human placenta. Monitoring the expression during placental development revealed that in first trimester and term placentae both molecules are equally expressed at the RNA level. In contrast, the protein levels were significantly reduced during gestation. Immunohistochemistry revealed a distinct localization of the EphB4 and ephrin-B2 proteins. EphB4 was predominantly expressed in the villous syncytial trophoblast layer and in a subset of intravillous capillaries. Prominent expression was also observed in the extravillous cytotrophoblast giant cells. In contrast, ephrin-B2 expression was detected in the villous cytotrophoblast and syncytial trophoblast cell layers, as well as initially in all intravillous capillaries. Strong expression was also observed in extravillous anchoring cytotrophoblast cells. Hypoxia is a major inducer of placental development. In vitro studies employing trophoblast-derived cell lines revealed that predominantly ephrin-B2 expression is induced by hypoxia, however, in an Hif-1α independent manner. These experiments suggest that EphB4 and ephrin-B2 are instrumental in the establishment of a functional placental structure and of the utero-placental circulation. [Copyright &y& Elsevier]

Objective: C-reactive protein (CRP) is a marker of systemic inflammation. Recently, it has been shown that CRP is present in amniotic fluid and fetal urine, and that elevated levels are associated with adverse pregnancy outcome. However, the precise source of amniotic fluid CRP, its regulation, and function during pregnancy is still a matter of debate. The present in vivo and in vitro studies were designed to investigate the production of CRP in human placental tissues. Material and methods: Ten paired blood samples from peripheral maternal vein (MV), umbilical cord artery (UA) and umbilical vein (UV) were collected from women with elective caesarean sections at term. The placental protein accumulation capacity of hCG, hPL, leptin and CRP was compared with the dual in vitro perfusion method of an isolated cotyledon of human term placentae and quantified by ELISA. Values for accumulation (release) were calculated as total accumulation of maternal and fetal circuits normalized for tissue weight and duration of perfusion. For gene expression, RNA was extracted from placental tissue and reverse transcribed. RT-PCR and real-time PCR were performed using specific primers. Results: The median (range) CRP level was significantly different between UA and UV [50.1ng/ml (12.1–684.6) vs. 61ng/ml (16.9–708.1)]. The median (range) difference between UV and UA was 9.3ng/ml (2.2–31.6). A significant correlation was found between MV CRP and both UA and UV CRP levels. Median (range) MV CRP levels [2649ng/ml (260.1–8299)] were 61.2 (6.5–96.8) fold higher than in the fetus. In vitro, the total accumulation rates (mean±SD) were 31±13 (mU/g/min, hCG), 1.16±0.19 (μg/g/min, hPL), 4.71±1.91 (ng/g/min, CRP), and 259±118 (pg/g/min, leptin). mRNA for hCG, hPL and leptin was detectable using conventional RT-PCR, while CRP mRNA could only be demonstrated by applying real-time RT-PCR. In the perfused tissue the transcript levels for the four proteins were comparable to those detected in the native control tissue. Conclusions: Our results demonstrate that the human placenta produces and releases CRP mainly into the maternal circulation similarly to other analyzed placental proteins under in vitro conditions. Further studies are needed to explore the exact role of placental CRP during pregnancy. [Copyright &y& Elsevier]

Abstract Purpose In Vivo EPID dosimetry, is a time-consuming procedure. This paper reports a time workload analysis of SOFTDISO software (SW) (Best Medical Italy) obtained in our center. Methods the time workload analysis was subdivided into two phases: the first regard pre-clinical mounting in particular the creation of communication DICOM nodes ( t links) , the SW commissioning time ( t commissioning) and the radiology technician training time ( t training) ; the second refer to clinical routine. This last phase was divided in four steps: single patient's loading times, t load , (import patient from TPS to SW, t TPS-SW , and from TPS to EPID management device, t TPS - EPID) ; single patient's daily synchronization time on SW, t synchro ; single patient's setting time, t set-up ; time of troubles, t troubles . In our center the TPS was Pinnacle 9.10 (Philips) and EPID device was IViewGT (Elekta). Results The pre-clinical phase required about three weeks where a multi-disciplinary team (medical physicist, product specialist and system administrator) has worked to interface all devices, the medical physicist has performed the SW commissioning measure and along with the product specialist they trained the staff. For a single patient by the clinical times shows in Table 1 we can calculate the daily time, t daily , equal to (240 ± 135) s and the insertion time, t load , equal to (180 ± 30) s. This time analysis was performed overall 30 patient. The daily workload was estimated from 10 of 20 patients treated per day who undergo to in vivo dosimetry checks and was equal to 45 min. Conclusions After a month a medium sized radiotherapy center gets these results with a dedicated medical physicist and a radiology technician. Additional clinical time is required to discuss the out of tolerance value provided by the SW. This time was estimated in 1 h per week. [ABSTRACT FROM AUTHOR]

Abstract: In the current study perfusions of an isolated cotyledon of term placenta using standard medium were compared to medium containing xanthine plus xanthine oxidase (X+XO), which generates reactive oxygen species (ROS). A time-dependant increase in the levels of different cytokines (TNF-α, IL-1ß, IL-6, IL-8 and IL-10) was observed between 1 and 7h with more than 90% of the total recovered from the maternal compartment with no significant difference between the 2 groups. For 8-iso-PGF2α 90% of the total was found in the fetal compartment and a significantly higher total release was seen in the X+XO group. Microparticles (MPs) isolated from the maternal circuit were identified by flow cytometry as trophoblastic sheddings, whereas MPs from the fetal circuit were predominantly derived from endothelial cells. More than 90% of the total of MPs was found in the maternal circuit. The absolute amount of the total as well as the maternal fraction were significantly higher in the X+XO group. Immunohistochemistry (IHC) of the perfused tissue revealed staining for IL-1β of villous stroma cells, which became clearly more pronounced in experiments with X+XO. Western blot of tissue homogenate revealed 2 isoforms of IL-1β at 17 and 31kD. In X+XO experiments there was a tendency for increased expression of antioxidant enzymes in the tissue. Western blot of MPs from the maternal circuit showed increased expression of antioxidant enzymes in the X+XO group and for IL-1β only the 17kD band was detected. In vitro reperfusion of human placental tissue results in mild tissue injury suggestive of oxidative stress. In view of the increased generation of ROS in perfused tissue with further increase under the influence of X+XO, the overall manifestation of oxidative stress remained rather mild. Preservation of antioxidant capacity of human placental tissue could be a sign of integrity of structure and function being maintained in vitro by dual perfusion of an isolated cotyledon. The observed changes resemble findings seen in placentae from preeclampsia. [Copyright &y& Elsevier]

Abstract: Aim of this study was to analyse the relationship between the plasma levels of polyphenols and the antioxidant activity of red and white wine. Twenty healthy subjects (HS) were randomly allocated to drink 300ml of red (n =10) or white n =10 wine for 15 days. Ten HS who refrained from any alcohol beverage for 15 days were used as control. Urinary PGF-2α-III, a marker of oxidative stress and plasma levels of polyphenols were measured. Urinary PGF-2α-III significantly fell in subjects taking wine with a higher percentage decrease in subjects given red wine (−38.5±6%, p <0.001) than in those given white wine (−23.1±6%). Subjects taking red wine had higher plasma polyphenols than those taking white wine (1.9±0.6μM versus 1.5±0.33μM, p <0.001). Plasma polyphenols were inversely correlated with urinary PGF2α (r =0.77, p <0.001). No changes of urinary isoprostanes were observed in subjects who refrained from wine intake. In vitro study demonstrated that only a mixture of polyphenols, all in a range corresponding to that found in human circulation, inhibited LDL oxidation and PKC-mediated NADPH oxidase activation. Such inhibitory effects were more marked using the concentrations of polyphenols detected in human circulation after red wine intake. This study shows that red wine is more antioxidant than white wine in virtue of its higher content of polyphenols, an effect that may be dependent upon a synergism among polyphenols. [Copyright &y& Elsevier]