Splitting cell adhesiveness into independent measurable parameters by comparing ten human melanoma cell lines.

The concept of cell adhesiveness was analyzed by looking for correlations between the adhesive behavior and measurable biological properties of different cell populations. Ten established lines of melanoma cells were assayed for passive deformability (by micropipet aspiration), active spreading (by measuring the height/diameter ratio after incubation on different surfaces), density and mobility of concanavalin A binding sites (by quantitative analysis of fluorescence microscopic images), spontaneous and concanavalin A-mediated agglutination (by measuring the number of cell conjugates resisting calibrated shearing forces), and binding to glass capillary tubes (with a quantitative assay of binding strength). Forty-four different parameters were thus measured, and each set of determinations was repeated 2 or 3 t at different days on each cell line. Analysis of variance was performed to assess the capacity of each parameter to discriminate between different lines. Correlations between different parameters were studied in order to understand a possible influence of cell intrinsic properties on the behavior of individual cells. The following conclusions were suggested by experimental data 1. Cell spreading ability, resistance to slow deformation within a micropipette and ability to form shear-resistant bonds, are independent properties. It is therefore suggested that different mechanisms rule the cell deformations on time scales of several minutes, tens of seconds, and fractions of a second. 2. Cell spreading ability may effectively influence binding strength only when adhesive stimuli are low, since in this case, cell stiffness is likely to impair the formation of extensive contact areas. 3. Individual cells may display marked heterogeneity within a given population, that emphasizes the danger of using averaged parameters to predict rare events (such as metastasis formation). 4. The most useful parameters to discriminate between different cell lines were, spreading ability and shear-resistant lectin agglutination, and substrate adhesion. It is concluded that cell adhesion is influenced by several measurable cellular properties that may display independent variations. The importance of a given parameter depends on the conditions of bond formation and rupture.     

Spatially periodic discrete contact regions in polylysine-induced erythrocyte—Yeast adhesion

Cell-cell adhesion occurs when human erythrocytes and yeast cells are suspended together in suprathreshold concentrations of polylysine in saline. The threshold polycation concentration for adhesion depends on cell concentration and decreases with increasing polycation molecular weight. The threshold concentration was similar for erythrocyte-erythrocyte adhesion and for yeast-erythrocyte adhesion. Transmission electron micrographs show that the erythrocytes adhere to yeast as if to engulf the cell. The regions of close contact between the erythrocyte membrane and the yeast cell walls are spatially discrete. The contact separation distance for the asymmetric erythrocyte-yeast adhesion is very similar to that (0.83 μm) observed when polylysine-induced adhesion occurs in the symmetrical erythrocyte-erythrocyte system. The spacing is attributed to the growth of a squeezing wave as an interfacial instability, on the intercellular aqueous layer. Freeze-fracture electron microscopy of cells that were not fixed during preparation for microscopy confirms the discrete nature of contacts between polylysine treated erythrocytes.     

Enhancement of the kidney Cd burden by SH-containing chelating agents

The accumulation of Cd in the kidneys is enhanced markedly if chelating agents that contain SH-groups like 2,3 dimercaptopropanol (BAL) are injected immediately after the metal. This is not a transient effect but persists for more than 3 d. It is less pronounced at higher chelate doses or when the pH of urine is increased. Our experiments indicate that chelating agents, which form unstable complexes at acid pH and are able to pass through the cell membrane, will cause metal accumulation in the kidneys.     

Cell orientation induced by extracellular signals

Cells like fibroblasts and osteoblasts are oriented by different extracellular guiding signals like an electric field, a bent surface, and a periodically stretched surface. An automatic controller is responsible for the cell alignment. The controller contains both a deterministic and a stochastic signal. The following machine properties were determined: (1) The angle dependence of the cellular signal transformer is cos 2(psi 0 - psi). (2) The set point of the automatic controller is psi 0 = +/- 90 degrees. The cells like to orient their long axis perpendicular to the direction of the applied guiding signal. (3) The signal transformer measures the extracellular signal in a quadratic fashion. The cells cannot register the sign of the guiding field. (4) The stochastic signal in the automatic controller can be quantified by a characteristic time (approximately 130 min for fibroblasts). (5) The extracellular signal is registered in cell-made standards (ratio of the deterministic and stochastic signal equals one): 0.3 +/- 0.05 V/mm for human fibroblasts (electric field) and 85 +/- 3 microns for human fibroblasts and osteoblasts (cyclindrically bent surface). (6) The lag-time in the signal transduction system of fibroblasts is approximately 4 min.     

Characterization of a nickel resistant mouse cell line

Nickel is a potent carcinogen and, at high concentrations, is toxic to mammalian cells. The effects associated with nickel exposure are well-documented but its mechanism of action in the cell has not yet been fully described. In order to understand the metabolic fate of nickel in mammalian cells, a variant cell population has been selected that continues to grow and divide in the presence of nickel chloride concentrations that are toxic to the parental cell line (Balb/c-3T3 mouse fibroblasts). Nickel resistance is not caused by altered uptake of nickel from the medium or increased clearance from the cells and is not associated with changes in metallothionein expression. Compared to the normal cells, the nickel resistant cells have a decreased number of chromosomes and numerous centromeric fusions. The expression of some proteins and the distribution of nickel bound by various proteins are altered in the nickel resistant cells. Preliminary results indicate that the nickel resistant phenotype may be transferred by genomic DNA-mediated transfection into a recipient NIH-3T3 cell line. Current investigations are directed at identifying a gene responsible for nickel resistance.     

The influence of different flow velocities on tumor cell survival in micropore filters

Earlier studies have shown a lower degree of lodgement and early survival of tumor cells in muscle than in liver after infusion via the femoral artery and portal vein, respectively. A possible explanation to this difference might be that the tumor cells are mechanically destroyed, and thus die more rapidly in muscle because they enter this capillary network at a much higher flow velocity. In the present study, the effect on early tumor cell (rat fibrosarcoma) survival of a high and a low flow velocity/deformation rate was evaluated in micropore (5 μm) filters, using isotope (Cr⁵¹) technique. These experiments were combined with scanning electron microscopic (SEM) analyses. The filter experiments showed no significant differences in the rate of cell death in the filters between tumor cells subjected to high or low deformation rates, and there were no qualitative differences in tumor cell appearance in the SEM study. It is, therefore, concluded that the difference in tumor cell lodgement and survival between muscle and liver is not primarily caused by differences in the rate of cell deformation upon entry of the organ capillary network.     

Transformation of human kidney epithelial cells to tumorigenicity by nickel(II) and V-HA-RAS oncogene

Nickel is a toxic metal of environmental concern that has been found to be carcinogenic in man and animals. Primary human kidney (NHKE) cells were immortalized or rescued from senescence after exposure to NiSO₄. The cell lines (IHKE) displayed abnormal karyotype and anchorage independent growth was observed. However, none of the IHKE cells produced tumor upon injection into athymic nude mice. Transfer of the v-Ha-ras oncogene into IHKE cells induced conversion of the immortalized cells into cell lines (THKE) that were tumorigenic when transplanted into athymic nude mice. Ha-ras DNA was present in the transformed cell lines and expressed at high level.     

A new technique for the study of erythrocyte survival by double labeling and use of a well-type Ge detector

A double label procedure with⁵⁷Co and⁵⁸Co has been developed for detailed in vivo studies of erythrocyte survival. A well-type Ge detector is used in the measurements. The activities necessary for these experiments are very low, and the associated dose received by the test persons can be neglected.     

The distribution of cadmium within the human prostate

Five normal prostates from autopsies of humans aged 61-76 yr were divided into 2 x 24 topographically well-defined pairs of slices that were analyzed for cadmium (Cd) and examined histologically. The corresponding kidney cortex concentrations were also determined. The concentrations in ng Cd/g wet wt were in the range of 50-500 in the prostates and 8,000-39,000 in the kidneys with good mutual correlation. Large variations in Cd concentrations within the prostates were found. The concentrations were highest at the base (near the bladder) and lowest at the apex of the gland. Furthermore, large variations within horizontal layers were found, and this variation was not correlated to the histological amount of stroma or glands.     

The effect of silicon (Si) on lipid parameters in blood serum and arterial wall

The influence of silicon treatment on the levels of lipid parameters of blood serum and aortic wall was studied in rats. The concentrations of total cholesterol, phospholipids, triglycerides, HDL-cholesterol, LDL-cholesterol, and HDL-phospholipids were measured in sera of rats receiving per os a soluble, inorganic silicon compound--sodium metasilicate nonahydrate (Na2SiO3.9H2O)--dissolved in the drinking water. In the aortic tissue levels of total cholesterol, triglycerides and phospholipids were estimated. An increase in HDL-cholesterol and HDL-phospholipid concentrations, with a simultaneous decrease of LDL-cholesterol and triglyceride levels, was observed in the sera of the tested group. The levels of total cholesterol and phospholipids in the sera, as well as the concentrations of lipids in the aortic walls, showed no significant differences. The results obtained could provide evidence for the existence of an additional mechanism of silicon antiatheromatous action, concerning the modification of activity of enzymatic systems involved in lipids metabolism.     

Determination of selenium in duplicate diets of residents of Pinhel,Portugal, by neutron activation

A rapid cyclic instrumental neutron activation analysis (CINAA) method has been used to determine the selenium content of 27 duplicate diet samples from each of the 27 districts surrounding Pinhel, Portugal. The accuracy and precision of the CINAA method have been evaluated by analyzing certified reference materials and observed to be within ±5–10% for samples containing at least 40 ppb of selenium. The detection limit has been found to vary between 26–42 ppb selenium depending on the sample composition. The average daily dietary intake has been calculated as 37 μg of selenium per day.     

Multielement determination in rice,wheat, and barley by instrumental neutron activation analysis

INAA has been used for the determination of Na, Mg, Al, Cl, K, Sc, Cr, Mn, Fe, Co, Cu, Zn, As, Se, Br, Rb, Sr, Mo, and W in grains of rice, wheat, and barley, which were collected from different plant fields in Iraq. Samples and standards were irradiated in the IRT-5000 reactor, at neutron fluxes of 2 × 10¹³ cm⁻²·s⁻¹ and 3.2 × 10¹¹ cm⁻²·s⁻¹. Interferences of photopeaks with each other were considered, and reaction interferences were calculated and determined experimentally. Accuracy of our method was assessed by the analysis of IAEA standards Wheat Flour and Bovine liver. A good agreement has been achieved between the present results and recommended values. The precision and detection limit were determined for all elements in all types of grain.     

Copper deficiency and hyperlipoproteinemia induced by a tetramine cupruretic agent in rabbits

The effectiveness of a cupruretic agent, N,N'-bis-(2 amino ethyl)-1,3-propanediamine HCl or 2,3,2-tetramine HCl (TETA), in the induction of copper (Cu) deficiency and the ability of a Cu-deficient diet in the maintenance of the depressed Cu status 10 wk after TETA treatment were examined in this study. In the first experiment, 42 male New Zealand White rabbits, 35 d of age, were randomly divided into three dietary treatments: a copper (Cu)-deficient (2.3 mg Cu/kg diet), a Cu-adequate (13.5 mg Cu/kg diet), and a commercial ration (21.6 mg Cu/kg diet) group. A single oral dose of 100 mg of 2,3,2-tetramine HCl TETA/kg body wt/d were administered to half of the rabbits from each treatment group for 10 d while the remaining rabbits were untreated. In the second experiment, 10 similar rabbits were assigned to three treatments: Cu-deficient plus TETA (n = 4); Cu-adequate plus TETA (n = 3); and Cu-adequate alone (n = 3). The rabbits were fed a TETA dose of 100 mg/d for three 4-d periods over 3 wk, and thereafter maintained on the diets for another 10 wk. Rabbits from the first experiment fed Cu-deficient diet and treated with TETA demonstrated cardiac hypertrophy and markedly reduced plasma and liver Cu concentrations that indicated that the animals were Cu-deficient. Significant elevations (twofold) in low density lipoprotein (LDL) protein, cholesterol, triglyceride, and apolipoprotein B (apo B) concentrations were observed in TETA treated rabbits fed Cu-deficient diet. In the second experiment, the plasma LDL protein level remained elevated, the plasma Cu level was reduced 45%, and the Cu level of the heart when expressed as microgram/g dry tissue was reduced, 10 wk post TETA treatment in rabbits maintained on Cu-deficient diet. Thus, Cu deficiency and hyperlipoproteinemia was rapidly induced by TETA and was still evident 10 wk posttreatment in rabbits maintained on a Cu-deficient diet.     

The in vitro transformation of a rat liver epithelial cell line with chromium

The transformation of a rat liver epithelial cell line under a wide range of doses of chromium was determined by anchorage-independent growth and tumor formation in syngeneic animals. Chronic exposure to low concentrations and brief exposure to high concentrations of hexavalent chromium (K₂CrO₄) transformed the cells, but one dose (1 mM K₂CrO₄, 2h) was clearly optimal in this regard. The cytotoxicity, effects on cell cycle, rates of chromium uptake, and mutagenic activity under the different treatment conditions were evaluated. The results showed that cells could adapt to the presence of chromium under certain treatment conditions, but this was not the case for the optimal transforming dose. Cells treated with chromium above the optimal transforming dose showed evidence of a transient G2 arrest, whereas all lower levels of treatment did not. A low level continuous exposure to chromate was mutagenic, whereas high level short exposures, including the optimal transforming dose, were not. An increase in the amount of protein complexed with isolated nucleic acids was detected in cells following treatment with the optimal transforming dose of chromate. The results indicate that the effects of chromium on this in vitro system vary with dose; and the identification of those events relevant to metal carcinogenesis will require consideration of treatment conditions.     

Mercury and selenium distribution in human kidney cortex

Concentration of mercury and selenium were analyzed in tissue fractions of human kidney cortex samples from seven autopsy cases. Total mercury content ranged between 0.3–9.0 nmol Hg/g wet wt. Between 27–61% of the total mercury was found in the 105,000g supernatant of the tissue homogenate from six cases. In kidney cortex from the seventh case, a deceased dentist with the highest concentration of mercury, only 3% of the total mercury was found in the 105,000g supernatant and about 88% in a SDS-insoluble fraction. In this fraction the molar ratio between mercury and selenium was close to 1∶1. This study supports results from previous animal studies and indicates that mercury in human kidney cortex could be deposited in forms with different solubility. It could be of importance to speciate different forms of mercury in tissues according to solubility and association to selenium when interpretations of mercury concentrations are made.     

A comparison of thermodynamic approaches to predict the adhesion of dairy microorganisms to solid substrata

Four different thermodynamic approaches were compared on their usefulness to predict correctly the adhesion of two fouling microorganisms from dairy processing to various solid substrata. The surface free energies of the interacting surfaces were derived from measured contact angles according to:

Precise large deletions by the PCR-based overlap extension method

The authors describe an efficient method for generating large deletions (>200 nts) of precise length using the PCR-based method of gene splicing by overlap extension (1). This method is technically simple and less time consuming than conventional loop-out mutagenesis techniques requiring preparation of a single-stranded DNA template.