Increase of poly(L-lysine) uptake but not fluid phase endocytosis in neuraminidase pretreated Madin-Darby canine kidney (MDCK) cells

The uptake of Poly(L-lysine) conjugates in cultured cells has been used as a model for non-specific adsorptive endocytosis of cationic macromolecules. To study the effect of glycocalyx desialylation on the uptake of cationic macromolecules in epithelial cells, Madin-Darby canine kidney (MDCK) cell monolayers were treated with neuraminidase and incubated with Poly(L-lysine) conjugates. Neuraminidase pretreatment of MDCK cells resulted in a 40% increase in the uptake of Poly(L-lysine) whereas trypsin pretreatment did not. Neuraminidase pretreatment did not increase the endocytosis of fluid phase markers in MDCK cells, nor the uptake of Poly(L-lysine) in L929 fibroblasts. These results suggest that the negative charges, rather than the glycoprotein matrices of glycocalyx, play an important role in the control of the uptake of cationic macromolecules in epithelial cells.     

Development and validation of a bioanalytical method for the determination of the cholecystokinin type-1 (CCK(1)) receptor antagonist dexloxiglumide in human plasma

A sensitive bioanalytical method for the measurement of dexloxiglumide, a new selective and potent cholecystokinin type-1 (CCK(1)) receptor antagonist, in plasma, is reported. The method is based on reversed-phase liquid chromatography with ultraviolet absorption detection. Samples are extracted under acidic conditions into an organic solvent, and following evaporation, reconstitution and centrifugation stages, the supernatant is injected on to an ODS column with detection at 244 nm. The method has been validated over the concentration range 0.2-20 microgram/ml, 0.2 microgram/ml being the lower limit of quantification. The overall precision and accuracy (expressed as relative error) of the method was less than 6.1 and 2.3%, respectively. Dexloxigulmide was shown to be stable in plasma when stored at -20 degrees C for at least 200 days. The method is suitable for studying the pharmacokinetics of dexloxiglumide in man.     

Immunohistochemical analysis of pRb2/p130, VEGF, EZH2, p53, p16(INK4A), p27(KIP1), p21(WAF1), Ki-67 expression patterns in gastric cancer

Although the considerable progress against gastric cancer, it remains a complex lethal disease defined by peculiar histological and molecular features. The purpose of the present study was to investigate pRb2/p130, VEGF, EZH2, p53, p16(INK4A), p27(KIP1), p21(WAF1), Ki-67 expressions, and analyze their possible correlations with clinicopathological factors. The expression patterns were examined by immunohistochemistry in 47 patients, 27 evaluated of intestinal-type, and 20 of diffuse-type, with a mean follow up of 56 months and by Western blot in AGS, N87, KATO-III, and YCC-2, -3, -16 gastric cell lines. Overall, stomach cancer showed EZH2 correlated with high levels of p53, Ki-67, and cytoplasmic pRb2/p130 (P < 0.05, and P < 0.01, respectively). Increased expression of EZH2 was found in the intestinal-type and correlated with the risk of distant metastasis (P < 0.05 and P < 0.01, respectively), demonstrating that this protein may have a prognostic value in this type of cancer. Interestingly, a strong inverse correlation was observed between p27(KIP1) expression levels and the risk of advanced disease and metastasis (P < 0.05), and a positive correlation between the expression levels of p21(WAF1) and low-grade (G1) gastric tumors (P < 0.05), confirming the traditionally accepted role for these tumor-suppressor genes in gastric cancer. Finally, a direct correlation was found between the expression levels of nuclear pRb2/p130 and low-grade (G1) gastric tumors that was statistically significant (P < 0.05). Altogether, these data may help shed some additional light on the pathogenetic mechanisms related to the two main gastric cancer histotypes and their invasive potentials.     

Archaeobotanical data and crop storage evidence from an early Bronze Age 2 burnt house at Arslantepe, Malatya, Turkey

Excavations on the southwest area at Arslantepe, Malatya, Turkey, by far the largest tell on the Malatya plain from the 5th millennium to the Neo-Hittite age, revealed an important change in the settlement patterns during the two main levels of the VI C Period of the site (Early Bronze Age 2, 2750–2500 cal b.c.). The latter level corresponds to a village founded on neatly shaped terraces in a layout which lasted for centuries, well into the following Early Bronze Age 3. This continuity was not broken even by violent fires that at times destroyed some houses, producing a huge quantity of charred plant remains, which comprised fruits, seeds and wood charcoal. The archaeobotanical data so far obtained from the EB2 house A607, the richest one in macro-remains, on which efforts have been concentrated first, provides much data about the use of the surrounding land. Charcoal of Quercus (deciduous oaks) (85%) followed by Populus (poplar) (9%) are dominant among wood remains, while Hordeum (barley) (70%) is the dominant crop found, followed by Cicer (chickpea) (17%). The crop storage methods were investigated by mapping the positions of charred fruits and seeds both according to the grid system and in comparison to the layout of facilities (grinding stone, hearths, oven) and the distribution of pottery (jars, bowls, pots) in order to detect where the crops were kept and the ways in which they were stored, processed, and used. The house facilities and furniture suggest that the house was a multifunctional place, which included storage space, but which was limited to household needs. The new archaeobotanical investigation so far carried out on the burnt house A607 suggests some implications on the degree of agriculture, on crop storage and on food processing practises and also gives information on the natural landscape surrounding the site.     

Fungal Bioweathering of Mimetite and a General Geomycological Model for Lead Apatite Mineral Biotransformations

Fungi play important roles in biogeochemical processes such as organic matter decomposition, bioweathering of minerals and rocks, and metal transformations and therefore influence elemental cycles for essential and potentially toxic elements, e.g., P, S, Pb, and As. Arsenic is a potentially toxic metalloid for most organisms and naturally occurs in trace quantities in soil, rocks, water, air, and living organisms. Among more than 300 arsenic minerals occurring in nature, mimetite [Pb₅(AsO₄)₃Cl] is the most stable lead arsenate and holds considerable promise in metal stabilization for in situ and ex situ sequestration and remediation through precipitation, as do other insoluble lead apatites, such as pyromorphite [Pb₅(PO₄)₃Cl] and vanadinite [Pb₅(VO₄)₃Cl]. Despite the insolubility of mimetite, the organic acid-producing soil fungus Aspergillus niger was able to solubilize mimetite with simultaneous precipitation of lead oxalate as a new mycogenic biomineral. Since fungal biotransformation of both pyromorphite and vanadinite has been previously documented, a new biogeochemical model for the biogenic transformation of lead apatites (mimetite, pyromorphite, and vanadinite) by fungi is hypothesized in this study by application of geochemical modeling together with experimental data. The models closely agreed with experimental data and provided accurate simulation of As and Pb complexation and biomineral formation dependent on, e.g., pH, cation-anion composition, and concentration. A general pattern for fungal biotransformation of lead apatite minerals is proposed, proving new understanding of ecological implications of the biogeochemical cycling of component elements as well as industrial applications in metal stabilization, bioremediation, and biorecovery.     

Improving the spatial orientation of human teeth using a virtual 3D approach

Since teeth are resistant to decomposition processes, they provide important and at times unique sources of information about fossil humans. Fortunately, dental remains reflect significant evolutionary changes. These changes make a very important and often exclusive contribution to the definition of new taxa or the attribution of fossil specimens to existing taxa.The traditional approach to dental morphometric analyses usually focuses on the recording of several measures of the tooth with calipers, especially the two basic crown diameters (buccolingual and mesiodistal). However, since these measures do not adequately represent the complex morphology of the tooth, 2D images and 3D digital models of dental morphology have been used. For both types of analysis, the possibility of correctly comparing homologous teeth depends on the adoption of a common orientation system. The lack of such a system makes it difficult to compare the results of different studies.Here we describe a new method for orienting teeth specifically devised for the upper and lower first molar (M1). Samples of unworn maxillary (n = 15) and mandibular (n = 15) first molars of modern humans were scanned with a Roland Picza 3D digitizer. The 3D virtual models were used to compare our new orientation method with those proposed in the literature. The new orientation system, which meets a geometric criterion, is based on three points identified on the cervical line and ensures acceptable repeatability of the spatial positioning and orientation independent of the shape and wear of the first molar under investigation. This orientation system is a first step toward the creation of a virtual set of hominid and fossil human first molars, which will allow us to make comparisons via a sophisticated and noninvasive approach. This pilot study also provides guidelines to extend the new methodology to the other types of teeth.     

Preliminary pharmacokinetic study of ibuprofen enantiomers after administration of a new oral formulation (ibuprofen arginine) to healthy male volunteers

The pharmacokinetics of ibuprofen enantiomers were investigated in a crossover study in which seven healthy male volunteers received single oral doses of 800 mg racemic ibuprofen as a soluble granular formulation (sachet) containing L-arginine (designated trade name: Spedifen®), 400 mg (-)R-ibuprofen arginine or 400 mg (+)S-ibuprofen arginine. Plasma levels of both enantiomers were monitored up to 480 minutes after drug intake using an enantioselective analytical method (HPLC with ultraviolet detection) with a quantitation limit of 0.25 mg/l. Substantial inter-subject variability in the evaluated pharmacokinetic parameters was observed in the present study. After (+)S-ibuprofen arginine, the following mean pharmacokinetic parameters ±SD were calculated for (+)S-ibuprofen: t_max 28.6 ± 28.4 min; C_max 36.2 ± 7.7 mg/l; AUC 86.4 ± 14.9 mg · h/l; t½ 105.2 ± 20.4 min. After (-)R-ibuprofen arginine, the following mean pharmacokinetic parameters were calculated for (+)S-ibuprofen and (-)R-ibuprofen, respectively: t_max 90.0 ± 17.3 and 50.5 ± 20.5 min; C_max 9.7 ± 3.0 and 35.3 ± 5.0 mg/l; AUC 47.0 ± 17.2 and 104.7 ± 27.7 mg · h/l; t_½ 148.1 ± 63.6 and 97.7 ± 23.3 min. After racemic ibuprofen arginine, the following mean pharmacokinetic parameters were calculated for (+)S- and (-)R-ibuprofen, respectively: t_max 30.7 ± 29.1 and 22.9 ± 29.8 min.; C_max 29.9 ± 5.6 and 25.6 ± 4.4 mg/l; AUC 105.1 ± 23.0 and 65.3 ± 15.0 mg · h/l; t_½ 136.6 ± 20.7 and 128.6 ± 45.0 min. T_max values of S(+)- and (-)R-ibuprofen after a single dose of 400 mg of each enantiomer did not differ significantly from the corresponding parameters obtained after a single dose of 800 mg of racemic ibuprofen arginine, indicating that the absorption rate of (-)R- and (+)S-ibuprofen is not different when the two enantiomers are administered alone or as a racemic compound. An average of 49.3 ± 9.0% of a dose of the (-)R-ibuprofen arginine was bioinverted into its antipode during the study period (480 minutes post-dosing). The percent bioinversion during the first 30 minutes after (-)R-ibuprofen arginine intake averaged 8.1 ± 3.9%. The mean AUC of (+)S-ibuprofen calculated after 800 mg racemic ibuprofen arginine (105.1 ± 23.0 mg · h/l) was lower than the mean AUC value obtained by summing the AUCs of (+)S-ibuprofen after administration of 400 mg (+)S-ibuprofen arginine and 400 mg (-)R-ibuprofen arginine (133.4 ± 26.6 mg · h/l). In conclusion, the administration of Spedifen® resulted in very rapid absorption of the (+)S-isomer (eutomer) with t_max values much lower than those observed for this isomer when conventional oral solid formulations such as capsules or tablets of racemic ibuprofen are administered. This characteristic is particularly favourable in those conditions in which a very rapid analgesic effect is required. Chirality 9:297–302, 1997. © 1997 Wiley-Liss, Inc.     

The Geological Roles Played by Microfungi in Interaction with Sulfide Minerals from Libiola Mine,Liguria, Italy

The potential role played by fungi in the weathering of sulfide abandoned mines and waste rock dumps is scarcely investigated, yet. In particular microfungi may produce biofilms that work as sites of metals and minerals precipitation. This study aimed to investigate interactions, bioalteration, and biocorrosion between three microfungi (Trichoderma harzianum group, Penicillium glandicola, P. brevicompactum) isolated from the Libiola sulfide mine (Liguria, Italy) and pyrite-rich mineralizations occurring within the waste rock dumps. After six weeks of incubation, Environmental Scanning Electron Microscope (ESEM) analyses showed how single pyrite crystals were completely corroded and altered by all the selected species. These results represent the first step to establish that fungi play a central role in the biogeochemical cycles of extreme and contaminated sites such as sulfide mines, and that they actively contribute to the evolution of the degraded ecosystem to more harmonized scenery.     

Hydrogen peroxide release from human blood platelets

The release of hydrogen peroxide from human blood platelets after stimulation with particulate membrane-perturbing agents has been determined by fluorescence using scopoletin as the detecting agent. Platelet suspensions containing less than 1 polymorphonuclear leukocyte/10⁸ platelets showed a significant release of hydrogen peroxide (6.11 nmol/10⁹ platelets per 20 min, S.D., 0.26, n=9) after addition of zymosan or latex particles, compared to unstimulated platelets. The release of hydrogen peroxide was only observed when the scopoletin was added to the platelet suspensions during the stimulation. Any attempt to determine hydrogen peroxide release in the supernatant at the end of the incubation with zymosan or latex failed. A NADH-dependent production of hydrogen peroxide was observed by measuring the difference of oxygen uptake in the presence and absence of catalase (500 units), which was not inhibited by potassium cyanide (1 mM). By this method the NADH-dependent cyanide-insensitive peroxide production and release was 6.0 nmol/10⁹ platelets per 20 min from resting platelets (S.D., 2, n=6) vs. 15 nmol/10⁹ platelets per 20 min from stimulated platelets (S.D., 2, n=6).     

First outbreak of food poisoning caused by Salmonella enterica subspecies enterica serovar Berta in Italy

Aims:  To provide an epidemiologic interpretation of a suspected outbreak of food poisoning caused by Salmonella enterica subspecies enterica serovar Berta strains isolated from humans and from the leftovers of the implicated foods (cream, dairy‐based desserts and eggs). Methods and Results:  We have correlated the similarity between the strains through genotyping with Pulsed Field Gel Electrophoresis (PFGE), studying antimicrobial sensitivity patterns and epidemiological investigation. The clonal origin of the outbreak was confirmed by all laboratory tests. PFGE analysis of the restriction profiles obtained with XbaI and SpeI revealed a certainly correlation from the strains isolated from the various sources, while the antimicrobial sensitivity pattern was the same in all cases, with all strains sensitive to all antibiotics tested. Conclusions:  Poor hygiene conditions in the facility concerned, lack of hygiene in food handling, high summer temperatures and positive cultures from asymptomatic staff could all be implicated in the infection, with food being the means through which it spread. Significance and Impact of the Study:  This study describes the first outbreak of food poisoning caused by Salmonella enterica subspecies enterica serovar Berta (Salmonella Berta) reported in Italy. It confirms the importance of correlating epidemiological investigations with genotyping and phenotyping to understand the dynamics of infection.