Elevated metallothionein-bound cadmium concentrations in urine from bladder carcinoma patients, investigated by size exclusion chromatography-inductively coupled plasma mass spectrometry

Cadmium is discussed as being involved in the development of transitional cell carcinoma (TCC) of the bladder and can be observed in urine of these patients. Investigations of urinary samples from bladder cancer patients and normal controls were carried out with special emphasis on metallothionein (MT)-bound cadmium.Compounds that are constituents of urine were separated in urine samples by means of size exclusion chromatography and cadmium was monitored continuously with a hyphenated inductively coupled plasma mass spectrometry (ICP-MS) system. MT-bound cadmium was quantified by peak area integration, taking into account the intensity of the rhodium signal which was added continuously before ICP-MS detection.The obtained results show that urinary cadmium is predominantly bound to the observed MT-fraction. The median of the MT-bound cadmium concentration in the control group was found to be 0.8 μg L⁻¹ whereas the cancer group has a median of 1.8 μg L⁻¹. The variance of the data in the cancer group is much higher than in the controls. However, the urinary MT-bound cadmium is significantly elevated in the cancer group; odds-ratio test: 7.11 (95% C.I.: 1.89-26.80), taking into account the total protein content.Due to the fact that only one main cadmium-containing fraction was observed, there is no necessity to separate the MT-fraction before cadmium determination in urine samples in future studies.     

A Radioimmunoassay for Norethindrone. II. Plasma Levels in Hypertensive and Nomotensive Oral Contraceptive Users

Abstract

A sensitive radioimmunoassay for measurement of norethindrone (N) in blood plasma has been developed. The coefficient of variation within the assay and between assays was 7 and 13.5% respectively. The procedural blanks were negligible, and recovery was approximately 81.8%. Blood plasma levels of N in 20 normotensive and 6 hypertensive women who were ingesting oral contraceptives (OCs) were measured, and the results indicate that the subjects who became hypertensive while taking OCs had significantly higher levels of N compared with normotensive OC users (p < .001).     

Highly Sensitive Sandwich Enzyme Immunoassay for Human Thyroid-Stimulating Hormone (hTSH) in Serum Using Monoclonal Anti-hTSH β-Subunit IgG1-Coated Polystyrene Balls and Polyclonal Anti-Human Chorinic Gonadotropin Fab′-Horseradish Peroxidase Conjugate

Abstract

A highly sensitive sandwich enzyme immunoassay for human thyroid-stimulating hormone (hTSH) in serum is described. Monoclonal anti-hTSH β-subunit IgG₁-coated polystyrene balls were incubated with serum samples and subsequently with polyclonal anti-human chorionic gonadotropin Fab¹-horseradish peroxidase conjugate. Neither hTSH nor anti-hTSH serum was required except for a small amount of hTSH as a standard. The detection limit of hTSH was 0.1 nU/tube or 0.005 mU/1 for most serum samples except for those from pregnant women, since there was no cross-reaction with 209 IU/1 of human follicle-stimulating hormone and 439 IU/1 of human luteinizing hormone when 20 μ1 of serum sample was used. Serum hTSH levels in normal subjects and Graves¹ disease were 1.22 ± 0.70 (SD) mU/1 (range: 0.26–2.65 mU/1) and 0.080 ± 0.062 (SD) mU/1 (range: < 0.005–0.17 mU/1), respectively.     

Fetal hematopoietic stem cells express MFG-E8 during mouse embryogenesis

The milk fat globule-EGF-factor 8 protein (MFG-E8) has been identified in various tissues, where it has an important role in intercellular interactions, cellular migration, and neovascularization. Previous studies showed that MFG-E8 is expressed in different cell types under normal and pathophysiological conditions, but its expression in hematopoietic stem cells (HSCs) during hematopoiesis has not been reported. In the present study, we investigated MFG-E8 expression in multiple hematopoietic tissues at different stages of mouse embryogenesis. Using immunohistochemistry, we showed that MFG-E8 was specifically expressed in CD34⁺ HSCs at all hematopoietic sites, including the yolk sac, aorta-gonad-mesonephros region, placenta and fetal liver, during embryogenesis. Fluorescence-activated cell sorting and polymerase chain reaction analyses demonstrated that CD34⁺ cells, purified from the fetal liver, expressed additional HSC markers, c-Kit and Sca-1, and that these CD34⁺ cells, but not CD34⁻ cells, highly expressed MFG-E8. We also found that MFG-E8 was not expressed in HSCs in adult mouse bone marrow, and that its expression was confined to F4/80⁺ macrophages. Together, this study demonstrates, for the first time, that MFG-8 is expressed in fetal HSC populations, and that MFG-E8 may have a role in embryonic hematopoiesis.     

Histamine-induced regulation of IL-2 synthesis in man: Characterization of two pathways of inhibition

Histamine at molar concentrations ranging from 10⁻⁵ to 10⁻⁷ exerted an inhibitory effect upon IL-2 synthesis by peripheral blood mononuclear cells in normal man; two pathways of inhibition are described. The first pathway was found to alter the T4 lymphocytes which, in the system used in this study, synthesized nearly 90 % of the total IL-2 production and had no suppressive activity. This suggests that histamine can act at the level of IL-2-producing cells. The second pathway of inhibition was related to induction of suppressor cells. Peripheral blood lymphocytes pre-incubated for 1 h with histamine 10⁻⁵–10⁻⁷ M inhibited the IL-2 synthesis of normal autologous lymphocytes in a co-culture system. This activity was radio-resistant (1200 r) and mediated by T8 lymphocytes. These two pathways of inhibition were mediated by the specific interaction of histamine with H1- and H2-receptor-bearing mononuclear cells.     

Photodynamic Effects of Hypericin on Lipid Peroxidation and Antioxidant Status in Melanoma Cells

Photodynamic-induced cytotoxicity by hypericin (HYP) was studied on three human melanoma cell lines: one pigmented cell line (G361) and two amelanotic cell lines (M18 and M6). No significant variation in the rate of uptake and in the maximum level of HYP incorporation for the different cells was observed. In the dark, no cytotoxicity was observed in the range0–10^?6M HYP for the three cell lines. Amelanotic cells were found to be more sensitive than pigmented cells to irradiation of HYP with visible light (Λ > 590 nm). In addition, for the three cell lines HYP-induced photocytotoxicity was found to be drug-dose and light-dose dependent. Under the conditions used, thiobarbituric acid-reacting substances (TBARs) were significantly increased in amelanotic cells after irradiation (P < 0.0001). By contrast, the amount of TBARS remained unchanged in pigmented cells. Antioxidant defenses including enzymes and glutathione (GSH) were assayed before and after HYP photosensitization. Significantly increased total SOD activity was observed after photosensitization for amelanotic cells (P < 0.05), while glutathione peroxidase (GSHPx) and catalase (Cat) activities but also GSH levels were significantly decreased (P < 0.01). In pigmented cells a significantly increased Cat activity was found (P < 0.05), whereas GSHPx was unaffected after irradiation. It can be inferred that (a) HYP may be an effective PDT agent for melanoma and (b) there is a relationship between melanin content and sensitivity to HYP phototoxicity in human melanoma cells.     

A New Highly Selective and Sensitive Assay for Fluorescence Imaging of .OH in Living Cells: Effectively Avoiding the Interference of Peroxynitrite

A new nonredox fluorescent probe to realize the imaging of hydroxyl radicals (^.OH) in living cells was designed and synthesized. The structure comprised the fluorescent dye boron dipyrromethene (BDP) and a 2,2,6,6‐tetramethyl‐1‐piperidinoxyl (TEMPO) unit. This probe could rapidly respond to ^.OH with a detection limit of 18 pM , and it possessed superior photostability and pH insensitivity. Other reactive oxygen species (ROS) and relevant intracellular components did not interfere. In particular, the important problem of ONOO^? interference was efficiently avoided. An MTT assay proved that the probe was not very cytotoxic. The probe could penetrate into intact cell membranes to selectively detect intracellular ^.OH without causing cellular damage in living mice macrophages, normal human liver cells. and human hepatoma cells. These advantageous characteristics make the fluorescent probe potentially useful as a new candidate to detect ^.OH in broad biosystems.     

The hydrological response of heavy clay grassland soils to rainfall in south‐west England using δ2H

Stable isotopes of water have been previously used in catchment studies to separate rain‐event water from pre‐event groundwater. However, there are a lack of studies at the smaller scale looking at the separation of event water from pre‐event water. This is particularly relevant for heavy clay soil systems through which the movement of water is uncertain but is thought to be rainwater‐dominated. The data presented here were collected at a rural site in the south‐west of England. The historic rainfall at the site was isotopically varied but similar to the global meteoric water line, with annual weighted means of ?37‰ for δ²H and ?5.7‰ for δ¹⁸O and with no seasonal variation. Drainage was sampled from the inter‐flow (surface runoff + lateral through‐flow) and drain‐flow (55 cm deep mole drains) pathways of two 1 ha lysimeters during two rainfall events, which had δ²H values of ?68‰ and ?92‰, respectively. The δ²H values of the lysimeter drainage water suggest that there was no contribution of event water during the first, small discharge (Q) event; however, the second larger event did show isotopic variation in δ²H values negatively related to Q indicating that rainwater was contributing to Q. A hydrograph separation indicated that only 49–58% of the inter‐flow and 18–25% of the drain‐flow consisted of event water. This was surprising given that these soil types are considered retentive of soil water. More work is needed on heavy clay soils to understand better the nature of water movement from these systems. Copyright © 2010 John Wiley & Sons, Ltd.     

Thermodynamics of the association of aqueous calcium and hexacyanoferrate ions

Thermodynamic quantities have been obtained for the formation of the ion pairs Ca²⁺Fe(CN)₆ ^4- and Ca²⁺Fe(CN)₆ ^3- in dilute aqueous solution by a potentiometric method employing an ion-exchange (Orion) calcium electrode. The reversibility of the electrode was examined against calomel and silver chloride references and was further tested in a determination of the formation constant for CaSO₄ (aq). The mean enthalpies of ion association of calcium with both hexacyanoferrates were obtained from the temperature variation of equilibrium constants within the range 15 to 35°C. The enthalpies were also independently determined calorimetrically. The results, discussed in terms of the electrostatic model, suggest that there is little penetration, if any, of hydration shells in the ion pairs.     

The effect of ultraviolet light on RNA metabolism in bromouracil deoxyriboside-grown HeLa cells

Abstract— The presence of 5-bromouracil deoxyriboside (BrUdR) in the DNA of HeLa cells has profound effects on RNA metabolism after u.v. irradiation. In normally grown cells 200 ergs/mm² depresses RNA synthesis by about 30 per cent while in BrUdR-grown cells the same exposure to u.v. depresses RNA synthesis by 95 per cent. When BrUdR-grown cells are u.v. -irradiated after being labeled with ³H-uridine, the normal autoradiographic pattern, where label shifts from nucleus to cytoplasm, fails to occur. Also, in lieu of the increase in RNA specific activity that is observed in unirradiated cells for a few hours after 20-min pulse-labeling, there occurs a constant decrease in specific activity after the irradiation.     

Radiometal (169Yb) uptake in normal and tumour cells in vitro: Influence of metabolic cell activity and complex structure

Trivalent radiometal tracers have been used for tumour imaging and metastatic pain palliation. For better understanding their tumour accumulation, basic model studies of uptake of different¹⁶⁹Yb complexes into cultured normal and tumour cells were performed.Whereas the uptake of¹⁶⁹Yb citrate is strongly dependent on the metabolic activity and is not tumour-cell specific, the uptake of¹⁶⁹Yb complexed with aminocarbonic acids (NTA, EDTA, DTPA) does not correlate to the metabolic activities. These complexes are taken up to a greater amount by the tumour cells (by a factor of about 2).Uptake of both complex types leads to a stable association to cellular compounds,¹⁶⁹Yb is not releasable by the strong complexing agent DTPA.Protein binding of the¹⁶⁹Yb complexes shows great influence on their cellular uptake. The bound proportion is no more available for cellular uptake.The results indicate that i) uptake of¹⁶⁹Yb citrate is an active cellular transport process which is not tumour-specific, ii) the¹⁶⁹Yb aminocarbonic acid complexes show a weak favouring by the tumour cells, iii) different from earlier acceptions the Yb complexes studied are not taken up by the cells in protein-bound form. The structure of the Yb complex is decisive for its protein binding and cellular uptake.     

Wellenmechanische absolutrechnung am HeHHe+

The wave function of the HeHHe⁺ molecule has been calculated by means of the GENERAL SCF –MO –LC (LCGO ) PROGRAM SYSTEM, taking all four electrons into account. The calculations were carried out for a number of linear equidistant, linear non-equidistant, and bent nuclear arrangements. The minimum energy of ?5.7930 a.u. was found for a linear equidistant configuration with a He? H distance of 0.939 Å. The corresponding ionization energy was 37.9 eV. An estimation of the energy of formation of HeHHe⁺ from HeH⁺ and He based on SCF -calculations on HeH⁺ and He gave 7.9 kcal/mole. The frequencies of the normal vibrations were calculated.     

Synthesis and antitumor activity of new silver(I)-N-heterocyclic carbene complexes

Abstract

In this study, two novel benzimidazole-based N-heterocyclic carbene ligands (1a-b) and their silver(I) complexes (2a-b) were synthesized. All new compounds were characterized by FT-IR, LC-MS, ¹H NMR, and ¹³C NMR spectroscopies. The in vitro antitumor activities of NHC ligands (1a-b) and their silver(I) complexes (2a-b) against DU-145 human prostate cancer cells, MDA-MB-231 and MCF-7 human breast cancer cells and L-929 (normal cells adipose from mouse) were also determined using MTT analysis for 24?h, 48?h, and 72?h. The results showed that while NHC ligands did not have in vitro antitumor activity on MCF-7, MDA-MB-231 and DU-145 cells, Ag(I)-NHC complexes have in vitro antitumor activities. The in vitro antitumor activity of 2a was found to be lower than that of 2b. Ag(I)-NHC complexes were observed to have higher IC₅₀ values for non-cancerous cell lines than cancer cells.     

Introduction of Adonis aestivalis as a new source of effective cytotoxic cardiac glycoside

Cardiac glycosides are used for treatment of irregular heartbeats, cardiac arrhythmia and congestive heart failures. In this research, digitoxin as a cardiac glycoside was identified and isolated for the first time in the world from Adonis aestivalis and investigated for its cytotoxic activity against cervical cancer cell (HeLa) lines and human lymphocytes by MTT test. Digitoxin extracted from the aerial parts of the plant collected from west of Iran and purified by column and thin layer chromatographic techniques. The structure of isolated cardiac glycoside was identified by IR, ¹H NMR and ¹³C NMR methods and so the presence of digitoxin was established. The half maximal inhibitory concentration values for cervical cancer and lymphocyte cells were obtained to be 5.62 and 412.94 μg/mL. The results of this study introduced the new resource of digitoxin which has considerable cytotoxic effects against HeLa cancer cells but did not damage normal human lymphocyte cells.     

Synthesis,characterization, and cytotoxic activity studies of new N4O complexes derived from 2-({3-[2-morpholinoethylamino]-N3-([pyridine-2-yl]methyl) propylimino} methyl)phenol

A new unsymmetrical five-coordinate Schiff base ligand (HL) with an N₄O donor set ( 2 ) has been prepared by condensation of N1-(2-morpholinoethyl)-N1-([pyridine-2-yl]methyl)propane-1,3-diamine with 2-hydroxy-benzaldehyde. Metal complexes [ML]ⁿ⁺ (M = Zn²⁺, Cd²⁺, Mn²⁺, Cu²⁺, Ni²⁺, Ag⁺, Fe³⁺, and Co²⁺ ( 3–10 ) were synthesized by the reaction of the ligand and metal salts in ethanol. The resulting products were characterized by elemental analyses, infrared, ¹H and ¹³C nuclear magnetic resonance spectra (in the case of Cd and Zn complexes), UV–Vis, electrospray ionization-mass spectrometric, and conductivity measurements. The structure of the complexes [ZnL](ClO₄) ( 3 ), [CdL](ClO₄) ( 4 ), and [CuL](ClO₄) ( 7 ) has been determined by single-crystal X-ray diffraction analysis. The metal complexes were determined to have a distorted trigonal bipyramidal (Zn and Cd) or a distorted square pyramidal (Cu) geometry. The cytotoxic potential of each compound (1–10) against MCF-7 and MDA-MB-231 (breast cancer cells), PC-3 (prostate cancer cells), and WI-38 human normal lung fibroblast cells was evaluated using the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) assay. Compounds 1, 2, and 10 did not display any activity toward any cell line tested. None of the compounds except compound 8 was cytotoxic toward PC-3. Compounds 4 and 8 showed the highest cytotoxic activity against the MCF-7 and MDA-MB-231 cell lines. Because compounds 3, 6, and 9 have similar half-maximal inhibitory concentration values against cancer cells and normal cells, these compounds displayed poor selectivity between cancer and normal cells. More importantly, it was observed that compound 5 acts differently toward different types of cell lines. For example, it displays lower cytotoxicity against the WI-38 normal cell line than it does against the MDA-MB-231 cell line.     

An immunoradiometric assay for human follicle stimulating hormone using a monoclonal antibody coupled to magnetisable cellulose

A simple immunoradiometric assay for human follicle stimulating hormone (hFSH) was developed using a pair of monoclonal antibodies obtained from commercial sources. The system developed makes use of a capture antibody covalently coupled to magnetisable cellulose, which is a more economical and stable immunosorbent as compared to the other solid phases. The detector antibody is labeled with¹²⁵I using the chloramine-T oxidation method and purified by gel filtration. After initial cross-matching of the capture and detector antibodies, various assay parameters have been optimised. This assay does not show any significant cross reactivity with homologous hormones. A number of serum samples from men and women from reproductive age group was screened and compared with another commercially available kit (r=0.98). Sensitivity of the assay is 1.4 mIU/ml, interassay variation is <5% and intraassay variation around 15%. The assay is reproducible and sensitive enough for regular estimation of serum hFSH and is relatively inexpensive.     

Gold(I) bis(N‐heterocyclic carbene) complexes: Metabolic stability,in vitro inhibition,and genotoxicity

We report the profiling of the metabolic stability, normal cell inhibition, and genotoxicity of the two gold complexes [Au (ⁱPr₂‐bimy)₂]PF₆ ( 1 ) and [Au (Fpyr)(ⁱPr₂‐bimy)]PF₆ ( 2 ), which show strong apoptotic activities in lung cancer cells. Liver microsomal tests revealed that the compounds have a relatively high half‐life compared to midazolam and do not suffer rapid metabolism and in vitro clearance. The cytotoxic potential of these compounds were also relatively weak in normal cells, with higher IC₅₀ values compared to cancer cells, with a 2–60 times difference. The Ames test revealed that the compounds do not give rise to any mutations as well. Overall, the compounds showed stability in liver microsomes, specificity for cancer cells, and a lack of genotoxic potential.     

Radon and thoron concentrations in offices and dwellings of the Gunma prefecture, Japan

Summary Aone year survey of indoor radon and thoron concentrations was carried out in offices and dwellings of the Gunma prefecture, Japan. A passive integrating radon and thoron discriminative monitor was used in the survey. The annual mean radon concentration was 22±14 Bq ^. m^-3, and ranged from 12 to 93 Bq ^. m^-3 among the 56 surveyed rooms. Radon concentration in offices was generally higher than that in the dwellings, with the arithmetic averages of 29 and 17 Bq ^. m^-3, respectively. Radon concentrations were generally lower in the traditional Japanese wooden houses than those houses built with other building materials. Seasonal variation of indoor radon was also observed in this survey. Compared to summer and autumn, radon concentrations were generally higher in spring and winter. The mean value of thoron to radon ratio was estimated to be 1.3, higher values were observed in the dwellings than in the offices. The annual effective dose from the exposure to indoor radon was estimated to be 0.47 mSv after taking the occupancy factors of offices and dwellings into account.     

A discrete dielectrophoresis device for the separation of malaria-infected cells

Malaria is a serious disease caused by Plasmodium parasites that infect red blood cells (RBCs). This paper presents the continuous separation of malaria-infected RBCs (iRBCs) from normal blood cells. The proposed method employed the discrete dielectrophoresis (DEP) in a microfluidic device with interdigitated electrodes. Our aim is to treat a sample having high concentration of cells to realize high throughput and to prevent the clogging of the microchannel with the use of the discrete DEP. The discrete DEP force for deflecting cells in the device was controlled by adjusting the magnitude, frequency, and duty cycle of the applied voltage. The effectiveness of the proposed method was demonstrated by separating the malaria-infected cells in samples having a cell concentration of 10⁶ cells/µl. From experimental results, we determined the enrichment that is needed to enhance the detection in the case of low parasitemia. The enrichment of the infected cells at the device output was 3000 times as high as that of the input containing 1 infected cell to 10⁶ normal cells. Therefore, the proposed method is highly effective and can significantly facilitate the detection of the infected cells for the identification of Malaria patients.     

DECREASED HOST CELL REACTIVATION OF UV-IRRADIATED ADENOVIRUS 5 BY FIBROBLASTS FROM COCKAYNE'S SYNDROME PATIENTS

Abstract— Over a period of 5 years, we performed 29 experiments in which survival curves of UV-irradiated adenovirus were determined using fibroblast strains from 10 normal persons and from 7 persons having Cockayne's syndrome. In all of these, the survival of UV-irradiated adenovirus 5 was less when assayed using monolayers of fibroblasts from Cockayne's syndrome patients than from normal persons. Survival curves using normal fibroblasts were, within error, straight lines on a log survival vs. linear fluence plot. Survival curves obtained using Cockayne's syndrome fibroblasts showed 2 components: an initial sensitive component, reflecting the behavior of approx. 75% of the infected cells, followed by a component having normal sensitivity. In the 28 experiments that were considered reliable, 58 curves were done using Cockayne's fibroblasts, 41 using normal human fibroblasts. Although experimental variation was encountered, there was no individual case in which sensitivity as measured using Cockayne's was equal to (or less than) the sensitivity measured using normal fibroblasts.