Imprinted H19 oncofetal RNA is a candidate tumour marker for hepatocellular carcinoma.

AIMS/BACKGROUND: To study the expression of the H19 gene in hepatocellular carcinoma. H19 is an imprinted, maternally expressed gene, which is tightly linked, both physically and functionally, to the paternally expressed insulin-like growth factor 2 (IGF II). IGF II is known to be involved in liver carcinogenesis. H19 was first discovered in the fetal mouse liver to be under the same regulatory genes as alpha fetoprotein (alpha FP), a widely used tumour marker for hepatocellular carcinoma. METHODS: Using both radioactive and non-radioactive in situ hybridisation, the expression of the H19 gene was compared with the presence of alpha FP, as demonstrated by immunohistochemistry, in 18 cases of hepatocellular carcinoma. RESULTS: H19 expression was present in 13 of 18 cases, whereas staining for alpha FP was positive in only nine of 18 cases. Concordance was found in 12 of 18 tumours (66.7%). In general, the staining pattern for H19 was more diffuse than the immunohistochemical staining for alpha FP. CONCLUSIONS: The addition of a non-radioactive in situ hybridisation assay for H19 RNA to the panel of tumour markers used for the histopathological and cytological diagnosis of hepatocellular carcinoma might be useful.     

Pre-eclampsia is not associated with changes in the levels of regulatory T cells in peripheral blood

PROBLEM: The acceptance of the semi-allogeneic fetus within the maternal environment requires tolerance mechanisms not fully characterized yet. Normal pregnancy is known to be associated with a Th2 profile. Furthermore, regulatory T cells (Tregs) were proposed to regulate the Th2/Th1 balance at early stages of pregnancy. Treg may avoid the shift to a Th1 profile, thus preventing miscarriage. Accordingly, spontaneous abortion is characterized by a Th1 dominance and diminished levels of Treg. The major aim of the present work was to investigate if pre-eclampsia, a late immunological complication of pregnancy, is characterized by similar hallmarks. METHOD OF STUDY: We measured the surface antigens CD4, CD25, CD8 and CTLA4 in peripheral blood from patients suffering from pre-eclampsia (n = 8) and age-matched patients undergoing normal pregnancies (n = 9) by four-color flow cytometry. RESULTS: We were not able to find any significant differences in the levels of CD4(+), CD25(+), CD8(+), CTLA4, CD4(+)/CD25(+), CD4(+)/CD25(bright), CD4(+)/CTLA4, CD25(+)/CTLA4, CD4(+)/CD25(+)/CTLA4, CD8(+)/CD25(+), CD8(+)/CTLA4 or CD8(+)/CD25(+)/CTLA4 cell subsets. CONCLUSIONS: Our data confirm comparable number of Tregs during pre-eclampsia and normal pregnancy in peripheral blood. Other regulatory mechanisms might be involved during late pregnancy.     

Quantitative analysis of granule cell axons and climbing fiber afferents in the turtle cerebellar cortex

The turtle cerebellar cortex is a single flat sheet of gray matter that greatly facilitates quantitative analysis of biotylinated dextran amine labeled granule cell and olivocerebellar axons and Nissl-stained granule and Purkinje neurons. On average, ascending granule cell axons are relatively thicker than their parallel fiber branches (mean±SD: 0.84±0.17 vs 0.64±0.12 µm, respectively). Numerous en passant swellings, the site of presynaptic contact, were present on both ascending and parallel fiber granule cell axons. The swellings on ascending axons (1.82±0.34 µm, n=52) were slightly larger than on parallel fibers (1.43±0.24 µm, n=430). In addition, per unit length (100 µm) there were more swellings on ascending axons (11.2±4.2) than on parallel fibers (9.7±4.2). Each parallel fiber branch from an ascending axon is approximately 1.5 mm long. Olivocerebellar climbing fiber axons followed the highly tortuous dendrites of Purkinje cells in the inner most 15–20% of the molecular layer. Climbing fibers displayed relatively fewer en passant swellings. The spatial perimeter of climbing fiber arbors (area) increased 72% from anteriorly (1797 µm²) to posteriorly (3090 µm²) and 104% from medially (1690 µm²) to laterally (3450 µm²). Differences in the size and spacing of en passant swellings on granule cell axons suggest that ascending axons may have a functionally more significant impact on the excitability of a limited number of radially overlying Purkinje cells than the single contacts by parallel fiber with multiple orthogonally aligned Purkinje cell dendrites. The spatially restricted distribution of climbing fibers to the inner most molecular layer, the paucity of en passant swellings, and different terminal arbor areas are enigmatic. Nevertheless, these finding provide important anatomical information for future optical imaging and electrophysiological experiments.     

Analysis of vertebrate eye movements following intravitreal drug injections. III. Spontaneous nystagmus is modulated by the GABAa receptor

1. Turtle eye movements were recorded in response to horizontal motion of patterned stimuli and intravitreal injections of selective GABAergic drugs by using a contact lens search-coil technique. Similar to results from rabbit and cat, injection of picrotoxin into the turtle's eye results in a spontaneous horizontal nystagmus, with its slow-phase movement in a temporal-to-nasal direction with respect to the injected eye. In contrast, there were no prominent vertical eye movements in response to either horizontal optokinetic stimuli or drug injections. 2. Injections of bicuculline or bicuculline methyl iodide (BMI), which selectively block the GABAa receptor, had effects similar to those of picrotoxin. The GABAa agonist muscimol, on the other hand, blocked optokinetic nystagmus (OKN). Furthermore, combinations of these drugs demonstrated competitive interactions between the agonists and antagonists. 3. The threshold dose for the eye-movement effects of each drug was ascertained with the use of a radioactive calibration procedure. Tritiated inulin was injected into the vitreous. After 1 h, ocular components were assayed for radioactivity. Then, by the use of an estimate of vitreal/retinal dilution, the retinal concentrations of these threshold doses were calculated. The computed threshold retinal concentrations of the GABAa drugs were found to be in the low micromolar range. 4. These results are discussed in terms of the directionally sensitive (DS) processing which occurs in the retina, and the output of retinal DS cells to pathways involved in oculomotor control of retinal image stabilization. It is known that intravitreal application of picrotoxin makes DS retinal ganglion cells lose their selectivity for any one direction. Based on the effect of picrotoxin on OKN, it is argued that DS retinal cells provide a major input to oculomotor subsystems involved in the stabilization of gaze. Furthermore, these intravitreal drug effects on OKN are selective for GABAa drugs, suggesting that GABAa receptors play a major role in DS processing in the retina.     

Distinct and overlapping roles of CXCR5 and CCR7 in B-1 cell homing and early immunity against bacterial pathogens

CXC chemokine receptor (CXCR)5 and CC chemokine receptor (CCR)7 are the major chemokine receptors required for B cell homing and microenvironmental localization during antigen-independent and -dependent B cell differentiation. Here, we show markedly decreased B-1 B cell numbers in the peritoneal cavity of CXCR5-/- and CXCR5-/-CCR7-/- double-deficient mice paralleled by reduced antigen-induced phosphorylcholine-specific immunoglobulin (Ig)M responses after intraperitoneal (i.p.) administration of streptococcal antigen. CCR7-/- mice also revealed a partial reduction in peritoneal B-1 cell numbers combined with a reduced humoral response to i.p. injected bacterial antigen. However, opposite roles of CXCR5 and CCR7 were observed when the frequency of peritoneal B-2 cells was analyzed. CXCR5-/- mice almost completely lacked B-2 cells, whereas CCR7 deficiency engendered an increase in peritoneal B-2 cells. In addition, CCR7-/- mice had enhanced, splenic IgM+ plasma cell responses, whereas the extrafollicular B cell response of the CXCR5-/- mice was not significantly altered compared with wild-type controls. Thus, the two chemokine receptors exert divergent forces at multiple levels of the innate immune response. CXCR5 plays a dominant role in peritoneal B-1 B cell homing and body cavity immunity, but both chemokine receptors are needed for a proportional peritoneal B-2 cell homing and balanced development of an early splenic B cell response.     

Vascular endothelial growth factor, epidermal growth factor and fibroblast growth factor-4 and -10 stimulate trophoblast plasminogen activator system and metalloproteinase-9

Trophoblast invasion, accompanied by degradation of extracellular matrix, is crucial to normal pregnancy development, whereas shallow placental invasion and implantation likely plays a role in the subsequent development of pre-eclampsia. The growth factors vascular endothelial growth factor (VEGF), epidermal growth factor (EGF) and fibroblast growth factor (FGF) are placental growth factors that activate degradation of extracellular matrix. We determined the effect of VEGF, EGF, FGF-2, FGF-4 and FGF-10 on the plasminogen activator system of first trimester cytotrophoblasts cultured in vitro. We studied the activity of urokinase plasminogen activator (uPA), its inhibitor plasminogen activator inhibitor-1 (PAI-1), and 92 kDa gelatinase-B (matrix metalloproteinase-9, MMP-9), using protein gel and reversed gel zymography. The expression pattern of FGF-4 and FGF-10 in human placental sections was determined by immunohistochemistry. FGF-4 was expressed in first trimester villi stroma, primarily in endothelial cells. FGF-10 expression was localized to first trimester extravillous trophoblasts. VEGF, EGF, FGF-4 and FGF-10, but not FGF-2, stimulate the activity of trophoblast uPA, PAI-1 and MMP-9. These results support the hypothesis that specific growth factors modulate the invasive potential of trophoblasts, and therefore may play an important role in early placental development. Our findings may contribute to the understanding of the pathophysiology of diseases associated with shallow placentation, such as pre-eclampsia.     

Prevalidation of in vitro continuous flow exposure systems as alternatives to in vivo inhalation safety evaluation experimentations: Outcome from MAAPHRI-PCRD5 research program

Diesel engine emission aerosol-induced toxicity patterns were compared using both in vitro (organotypic cultures of lung tissue) and in vivo experimentations mimicking the inhalation situation with continuous aerosol flow exposure designs. Using liquid media resuspended diesel particles, we show that toxic response pattern is influenced by the presence of tensioactive agent in the medium which alter particle-borne pollutant bioavailability. Using continuous aerosol exposure in vitro, we show that with high sulfur fuel (300ppm) in the absence of oxidation catalysis, particulate matter was the main toxic component triggering DNA damage and systemic inflammation, while a very limited oxidant stress was evidenced. In contrast, with ultra-low sulfur fuel in the presence of strong diesel oxidation catalysis, the specific role of particulate matter is no longer evidenced and the gas phase then becomes the major component triggering strong oxidant stress, increased NO(2) being the most probable trigger. In vivo, plasma tumor necrosis factor alpha (TNFalpha), lung superoxide dismutase (SOD), catalase and glutathione peroxidase (GPx) activity levels varied in agreement with in vitro observations. Diesel emission treatment with oxycat provokes a marked systemic oxidant stress. Again NO(2) proved to account for a major part of these impacts. In conclusion, similar anti-oxidant responses were observed in in vitro and in vivo experiments after diesel emission aerosol continuous flow exposures. The lung slice organotypic culture model-exposed complex aerosol appears to be a very valuable alternative to in vivo inhalation toxicology experimentations in rodents.     

Effects of antipsychotic drugs on neurogenesis in the forebrain of the adult rat.

The generation of new cells in the adult mammalian brain may significantly modify pathophysiological processes in neuropsychiatric disorders. We examined the ability of chronic treatment with the antipsychotic drugs (APDs) olanzapine and haloperidol to increase the number and survival of newly generated cells in the prefrontal cortex (PFC) and striatal complex of adult male rats. Animals were treated with olanzapine or haloperidol for 3 weeks and then injected with 5-bromo-2'-deoxyuridine (BrdU) to label mitotic cells. Half of the animals continued on the same APD for two more weeks after BrdU challenge, with the other half receiving vehicle during this period. Olanzapine but not haloperidol significantly increased both the total number and density of BrdU-labeled cells in the PFC and dorsal striatum; no effect was observed in the nucleus accumbens. Continued olanzapine treatment after the BrdU challenge did not increase the survival of newly generated cells. The newly generated cells in the PFC did not express the neuronal marker NeuN. Despite the significant increase in newly generated cells in the PFC of olanzapine-treated rats, the total number of these cells is low, suggesting that the therapeutic effects of atypical APD treatment may not be due to the presence of newly generated cells that have migrated to the cortex.     

First clinical experience with the premounted balloon-expandable serpentine stent: Acute angiographic and intermediate-term clinical results

The beStent-Artist coronary stent is a newly developed, stainless steel coronary stent with a serpentine tubular design and terminal stent markers, premounted on a semicompliant balloon. During this pilot evaluation we aimed to test the acute clinical and angiographic results, short-term (30 days) and 6-month clinical results. A total of 57 stents were used to treat 43 lesions in 40 patients. Deployment strategy included predilatation, stent deployment, balloon repositioning to match the distal end of the balloon to the distal stent marker, and subsequent 12–14 atm postdilatation. There were two cases of stent dislodgment, but no procedural complications. In four cases, stent recrossing with another balloon was necessary. In two of these cases, distal dissections were observed and treated with another stent. The minimal lumen diameter (MLD) increased from 0.84 ± 0.52 mm at baseline to 2.7 ± 0.62 mm at the end of the procedure (a corresponding decrease in diameter stenosis from 78.6 > 16.4 to 18.2 ± 10.7%). The acute gain was 1.89 ± 0.61mm. No adverse events occurred by 30 days. During six months, 7/40 (18.5%) of patients required target vessel revascularization due to in-stent restenosis. In summary, the premounted beStent-Artist can be delivered and deployed with favorable immediate results and high success rate with favorable long-term recurrent event rates.Cathet. Cardiovasc. Intervent. 46:249–253, 1999. © 1999 Wiley-Liss, Inc.