Effects of essential divalent metals on carcinogenicity and metabolism of nickel and cadmium

Interactions between the physiologically essential metals calcium, magnesium, and zinc and the carcinogenic metals nickel and cadmium were investigated to help elucidate the mechanisms of action of the carcinogenic metals. Bioassay studies revealed several significant findings, including: (1) the ability of magnesium and calcium to inhibit nickel-induced elevation of pulmonary adenoma incidence in strain A mice; (2) the ability of magnesium, but not of calcium, to prevent cadmium-induced subcutaneous sarcoma formation; and (3) the ability of magnesium, but not of calcium, to inhibit nickel-induced muscle tumor formation. Biochemical studies indicated a direct relationship between the antitumorigenic potential of magnesium and the capacity of this metal to: (1) inhibit nickel and cadmium uptake by the target tissues in vivo; (2) inhibit nickel-induced disturbances in DNA synthesis in vivo; (3) inhibit nuclear and cytosolic uptake of nickel by the target tissue cells in vivo; and (4) inhibit nickel and cadmium binding to DNA in vitro. Calcium, which in most cases did not prevent carcinogenesis, had no consistent influence on the uptake of carcinogenic metals or their biochemical effects in the target tissues. Magnesium and zinc, but not calcium, were also found to attenuate the acute toxic effects of nickel, indicating a possible correlation between prevention of acute effects and reduction in tumorigenicity. Zinc, which antagonizes cadmium tumorigenicity in the rat testis, was found to reduce markedly cadmium uptake into isolated testicular interstitial cells. Also, zinc was found to inhibit strongly cadmium binding to DNA in vitro.     

Nitrogen mineralization ofSesbania sesban used as green manure for lowland rice in Sri Lanka

Sesbania sesban was evaluated as green manure crop for lowland rice in the Dry Zone of Sri Lanka. The legume was grown during a fallow period before lowland rice (Oryza sativa) and ploughed under just before transplanting. Weight loss and nitrogen content in litterbags containing leaves, stems and roots of the legume were monitored. Comparisons were made between rice yields from 20 m² plots after green manuring in combination with different nitrogen fertilizer levels (0, 2.4, 4.8 and 7.2 gm⁻²) and nitrogen fertilizer (9.6 gm⁻²) alone. Above-ground biomass ofS. sesban was 440 gm⁻² (dry wt) when ploughed under after 84 days growth. N-content in leaves, stems and roots was 3.76%, 0.41% and 0.73%, respectively. This gave a N-input fromS. sesban of 9.2 gm⁻² (8.3 g from above-ground parts and 0.9 g from roots). The corresponding K and P inputs were 7.3 and 0.6 gm⁻² respectively. The nitrogen rich leaves, which contained 88% of the nitrogen in the above-ground parts, decomposed and released its nitrogen much more rapidly than the stems and roots. After only four days the leaves had released 5.3 g Nm⁻² and after 14 days they had released 6.4 g Nm⁻². The highest rice yield (505 gm⁻²) was obtained usingS. sesban and 4.8 gm⁻² of N-fertilizer. The yields with only N-fertilizer or onlyS. sesban were 442 gm⁻² and 396 gm⁻², respectively. Due to the rapid decomposition of the nitrogen rich leaves,S. sesban did not behave as a slow release fertilizer. Thus, it is not necessary to apply nitrogen fertilizers as a basal dose.     

Method for kinetic analysis of drug-induced cell cycle perturbations.

A method is described for quantitative study of the flux of cells through the cell cycle phases in in vitro systems perturbed by chemicals, such as chemotherapeutic agents. The method utilizes cell count and the flow cytometric technique of bromodeoxyuridine (BrdUrd) labeling, according to an optimized strategy. Cells are exposed to BrdUrd during the last minutes of drug treatment and fixed for analysis at 0, 1/3Ts, 2/3Ts, Ts, and Tc + TG1 recovery times, where Ts, TG1, Tc are the mean durations of phases S and G1 and of the whole cycle of control cells. As an example of application of the proposed procedure, a kinetic study of the effect of 1-(2-chloroethyl)-1-nitrosourea (CNU) on the L1210 cell cycle is described. Simple data analysis, requiring only a pocket calculator, showed that cells in phases G1 and G2M at the end of a 1 h treatment with 1 microgram/ml CNU were fully able to leave these phases but were destined to remain blocked in the following G2M phase (G1 for a minority of them). We also found that cells initially in S phase were slightly delayed in completing their S phase and that 50% of them remained temporarily blocked in the subsequent G2M phase, irrespective of their position in the S phase.     

The three-dimensional structure of the aspartyl protease from the HIV-1 isolate BRU.

The crystal structure of the aspartyl protease encoded by the gene pol of the human immunodeficiency virus (HIV-1, isolate BRU) has been determined to 2.7 A resolution. The enzyme, expressed as an insoluble denatured polypeptide in inclusion bodies of Escherichia coli has been renatured and crystallized. It differs by several amino acid replacements from the homologous enzymes of other HIV-1 isolates. A superposition of the C alpha-backbone of the BRU protease with that of the SF2 protease gives a roots mean square positional difference of 0.45 A. Thus, neither the denaturation/renaturation process nor the amino acid replacements have a noticeable effect on the three-dimensional structure of the BRU protease or on the detailed conformation of the catalytic site, which is very similar to that of other aspartyl proteases.     

Hybridization of DNA from methanogenic bacteria with nitrogenase structural genes (nifHDK)

Summary Using the Southern hybridization technique, homologies were examined between restricted DNA of four methanogenic bacteria (Methanobacterium ivanovi, Methanobacterium thermoautotrophicum, Methanococcus voltae, Methanosarcina barkeri) and the nif (nitrogen fixation) genes of Klebsiella pneumoniae and Anabaena strain 7120. With K. pneumoniae probes, no hybridization was observed with nifA, nifNE, and nifJ but positive results were obtained with the nifHDK genes coding for nitrogenase. Homology was detected, in the four strains, with K. pneumoniae and Anabaena nifH probes. In M. voltae and M. ivanovi, the homology found with nifH was estimated to be about 70% and a weaker hybridization was observed also with nifD and nifK. In M. voltae, the sequence homologous to nifH was found on a 3.0 kbp HindIII fragment and sequences homologous to nifD and nifK on a 3.8 kbp HindIII fragment. The 3.0 kbp fragment was cloned and the region homologous to nifH was localized more precisely. When this fragment was used as a probe against other DNAs, it behaved as a K. pneumoniae and Anabaena nifH probe. The results suggest that the structural genes for nitrogenase may be present in archaebacteria and raise interesting questions regarding their evolution.     

Development of calcium and secretory responses in the human promyelocytic leukemia cell line HL60

We have begun to characterize the development of the excitation-response coupling sequence in the human promyelocytic leukemia cell line HL60. Using the recently developed fluorescent calcium probe quin-2, it was found that DMSO induced myeloid differentiation of the HL60 cells is accompanied by the development of a calcium response to the addition of the chemotactic factors fMet-Leu-Phe and leukotriene B4. The characteristics (time course, concentration dependence, stereospecificity, and metabolic dependence) of the calcium response are extremely similar to those previously described in human neutrophils. These results imply that functional receptors for leukotriene B4 appear in HL60 cells upon the induction of differentiation and also lend strong support to the use of these HL60 cells as a model of human myeloid differentiation. We have also characterized the emergence of a secretory response to fMet-Leu-Phe and leukotriene B4 in cytochalasin B treated HL60 cells. In addition, it is found that differentiation was required for the calcium ionophore A23187 to express its secretory activity toward the HL60 cells. This last set of results implies that differentiation is accompanied by the coordinated appearance of surface receptors and cytoplasmic factors required for the expression of cellular responsiveness.     

Structural Aspects of Saltatory Particle Movement

A variety of cells possess particles which show movements statistically different from Brownian movements. They are characterized by discontinuous jumps of 2–30 µ at velocities of 0.5–5 µ/sec or more. A detailed analysis of these saltatory, jumplike movements makes it most likely that they are caused by transmission of force to the particles by a fiber system in the cell outside of the particle itself. Work with isolated droplets of cytoplasm from algae demonstrates a set of fibers involved in both cytoplasmic streaming and saltatory motion, suggesting that both phenomena are related to the same force-generating set of fibers. Analysis of a variety of systems in which streaming and/or saltatory movement occurs reveals two types of fiber systems spatially correlated with the movement, microtubules and 50 A microfilaments. The fibers in Nitella (alga) are of the microfilament type. In other systems (melanocyte processes, mitotic apparatus, nerve axons) microtubules occur. A suggestion is made, based on work on cilia, that a microtubule-microfilament complex may be present in those cases in which only microtubules appear to be present, with the microfilament closely associated with or buried in the microtubule wall. If so, then microfilaments, structurally similar to smooth muscle filaments, may be a force-generating element in a very wide variety of saltatory and streaming phenomena.