DETERMINING THE LABELING INDEX IN AUTORADIOGRAPHY

A method is proposed for estimating the labeling index in autoradiographs. It is based on a model for the frequency distribution of grain counts for those cells which have few grains. The method does not require extensive grain counting and provides reasonable estimates even when the background level is high. It is particularly suited for experiments where tumor cells, labeled in vivo, are, autoradiographed after being cultured in vitro.     

The effect of DL-propranolol on γ-aminolevulinic acid synthetase activity and urinary excretion of porphyrins in allylisopropylacetamide-induced experimental porphyria

Experimental porphyria was induced in rats by allylisopropylacetamide. DL-Propranolol, a β-adrenergic-receptor blocking agent, significantly reduced the elevated urinary excretion of δ-aminolevulinic acid, porphobilinogen and total porphyrins. DL-Propranolol also partially prevented the increased activity of δ-aminolevulinic acid synthesis in liver homogenates of allylisopropylacetamide-treated rats. It had no effect on the above parameters in normal rats. These findings support the hypothesis that δ-aminolevulinic acid exists in two forms, a constitutive and an inducible one.In order to examine whether the action of the drug was caused by its membrane effect. D-propranolol and quinidine sulphate were used in similar sets of experiments. These drugs had no effect on the abnormal porphyrin metabolism of allylisoprpyl-acetamide-treated rats, indicating that the results obtained with DL-propranolol were not due to its membrane action.     

Inhibitory effect of membrane active compounds on the uptake of 14C-alpha-aminoisobutyric acid (AIB) in cultured chick embryo liver cells.

High concentrations of beta-adrenoceptor blocking drugs with membrane active properties and of the membrane active compounds quinidine and lidocaine inhibit the uptake of ∝-aminoisobutyric acid by chick embryo liver cells in culture. Beta-adrenoceptor blockers without membrane active properties were without effect. These results are in accordance with previous findings which showed partial inhibition of incorporation of amino acids into proteins caused by membrane active drugs in this system.     

Maternal factors in onset of Huntington disease.

Analyses of father-offspring and mother-offspring similarity in onset age suggest that nuclear genes account for a significant portion of the modification of onset age in Huntington disease. The effects of non-nuclear modifiers are supported by the finding that the offspring of affected women have significantly older mean ages of onset than offspring of affected men irrespective of the onset age in the parent. The absence of increased father-daughter similarity indicates that modification is not X-linked. The absence of reproductive advantage for late-onset individuals and the absence of a multigenerational maternal-lineage effect suggest that the modifying effect of the sex of the affected parent occurs in a single parental generation. Offspring of affected women with onset between ages 35 and 49 had a significantly older mean onset age than their mothers. This suggests that a protective effect may be conferred upon the offspring of affected women.     

Mechanism of aluminium-induced porphyrin synthesis in bacteria

In previous studies, aluminium was found to retard bacterial growth and enhance porphyrin formation in Arthrobacter aurescens RS-2. The aim of this study was to establish the mechanism of action of aluminium which leads to increased porphyrin production. Cultures of Arthrobacter aurescens RS-2 were incubated in the absence and presence of 0.74 mm aluminium. After 6 and 24 h of incubation, various parameters of the haem biosynthetic pathway were determined. After 6 h of incubation with aluminium, the activities of the enzymes aminolevulinate synthase (ALAS), aminolevulinate dehydratase (ALAD), porphobilinogen deaminase (PBGD) and uroporphyrinogen decarboxylase (UROD) were increased by 120, 170, 190 and 203%, respectively, while that of ferrochelatase (FC) was found to be unchanged. However, after 24 h of incubation, no change in the activities of ALAS and ALAD was noted, while an about 2-fold increase in PBGD and UROD activities were observed. FC activity was decreased by 63%. It was concluded that aluminium exerts its effect by inducing the enzymes PBGD and UROD rather than by a direct or indirect effect on ALAS. Its effect on the final step in the haem biosynthetic pathway is discussed.     

A Novel Anti-Tumor Inhibitor Identified by Virtual Screen with PLK1 Structure and Zebrafish Assay

Polo-like kinase 1 (PLK1), one of the key regulators of mitosis, is a target for cancer therapy due to its abnormally high activity in several tumors. Plk1 is highly conserved and shares a nearly identical 3-D structure between zebrafish and humans. The initial 10 mitoses of zebrafish embryonic cleavages occur every∼30 minutes, and therefore provide a rapid assay to evaluate mitosis inhibitors including those targeting Plk1. To increase efficiency and specificity, we first performed a computational virtual screen of∼60000 compounds against the human Plk1 3-D structure docked to both its kinase and Polo box domain. 370 candidates with the top free-energy scores were subjected to zebrafish assay and 3 were shown to inhibit cell division. Compared to general screen for compounds inhibiting zebrafish embryonic cleavage, computation increased the efficiency by 11 folds. One of the 3 compounds, named I2, was further demonstrated to effectively inhibit multiple tumor cell proliferation in vitro and PC3 prostate cancer growth in Xenograft mouse model in vivo. Furthermore, I2 inhibited Plk1 enzyme activity in a dose dependent manner. The IC₅₀ values of I2 in these assays are compatible to those of ON-01910, a Plk1 inhibitor currently in Phase III clinic trials. Our studies demonstrate that zebrafish assays coupled with computational screening significantly improves the efficiency of identifying specific regulators of biological targets. The PLK1 inhibitor I2, and its analogs, may have potential in cancer therapeutics.     

The effect of plant growth-promoting rhizobacteria on the growth of rice (Oryza sativa L.) cropped in southern Brazilian fields

Background and Aims

Several strains of rhizobacteria may be found in the rhizospheric soil, on the root surface or in association with rice plants. These bacteria are able to colonize plant root systems and promote plant growth and crop yield through a variety of mechanisms. The objectives of this study were to isolate, identify, and characterize putative plant growth-promoting rhizobacteria (PGPR) associated with rice cropped in different areas of southern Brazil.

Methods

Bacterial strains were selectively isolated based on their growth on three selective semi-solid nitrogen-free media. Bacteria were identified at the genus level by PCR-RFLP 16S rRNA gene analysis and partial sequencing methodologies. Bacterial isolates were evaluated for their ability to produce indolic compounds and siderophores and to solubilize phosphate. In vitro biological nitrogen fixation and the ability to produce 1-aminocyclopropane-1-carboxylate deaminase were evaluated for each bacterial isolate used in the inoculation experiments.

Results

In total, 336 bacterial strains were isolated representing 31 different bacterial genera. Strains belonging to the genera Agrobacterium, Burkholderia, Enterobacter, and Pseudomonas were the most prominent isolates. Siderophore and indolic compounds producers were widely found among isolates, but 101 isolates were able to solubilize phosphate. Under gnotobiotic conditions, eight isolates were able to stimulate the growth of rice plants. Five of these eight isolates were also field tested in rice plants subjected to different nitrogen fertilization rates.

Conclusions

The results showed that the condition of half-fertilization plus separate inoculation with the isolates AC32 (Herbaspirillum sp.), AG15 (Burkholderia sp.), CA21 (Pseudacidovorax sp.), and UR51 (Azospirillum sp.) achieved rice growth similar to those achieved by full-fertilization without inoculation, thus highlighting the potential of these strains for formulating new bioinoculants for rice crops.