Effect of verapamil on the aldosterone-stimulating properties of metoclopramide: in vitro and in vivo studies

The role of calcium in the regulation of aldosterone secretion has been recently clarified. Angiotensin II and potassium stimulate aldosterone secretion through a calcium-entry dependent mechanism, while ACTH action is both calcium and cyclic AMP dependent. To establish whether also the so-called aldosterone dopaminergic regulatory system is calcium-dependent we have studied, in vitro and in vivo, the effect of verapamil, a calcium entry blocker agent, on the aldosterone-stimulating properties of the antidopaminergic drug, metoclopramide. In the rat adrenal cells perfusion system, verapamil blocked both angiotensin II and metoclopramide-stimulated aldosterone. This effect on metoclopramide action seems to be present also in vivo in normal subjects: in fact aldosterone response was slightly but significantly reduced after pretreatment with verapamil. In conclusion the results suggest that also the dopaminergic system could regulate aldosterone secretion through calcium-mediated mechanisms.     

The phylogeny of the hominoid primates: a statistical analysis of the DNA-DNA hybridization data

Sibley and Ahlquist compared the single-copy nuclear DNA sequences of the hominoid primates using DNA-DNA hybridization. From this data set they estimated a phylogeny that clusters man and chimpanzees using a distance Wagner procedure. However, no assessment of statistical confidence in this estimated phylogeny was made, despite the fact that their data set contains internal inconsistencies concerning the correct branching order. This paper presents a modification of Pielou's Q- statistic that allows one to make nonparametric tests of phylogenetic relationship from distance data. The results of this analysis indicate that the estimated phylogeny of Sibley and Ahlquist is without statistical significance owing to the internal inconsistencies of the data set. A survey and additional analyses of other types of molecular data indicate that the phylogeny that clusters chimpanzees and gorillas and has the human lineage splitting off earlier is statistically consistent with all the molecular data (including the DNA-DNA hybridization data), whereas the phylogeny estimated by Sibley and Ahlquist can be rejected at the 5% level using the data on restriction- endonuclease sites in the mitochondrial genome.      

Microtubule organization in sperm cells in the pollen tubes ofBrassica oleracea L.

Summary Microtubules tightly cross-linked into bundles are described in the sperm cells ofBrassica oleracea pollen tubes. The sperm cells are lobed and tailed and the microtubule bundles are often located in these parts of the cells. In the present paper we suggest that the cross-linked microtubule organization could determine an intertubular sliding, probably generating a motility system that propels the sperm cells through the tube.Abbreviations GC generative cell - Mfs microfilaments - Mts microtubules - SC sperm cell - VC vegetative cell - VN vegetative nucleus     

Interactions ofCandida albicans with bacteria and salivary molecules in oral biofilms

The yeastCandida albicans coaggregates with a variety of streptococcal species, an interaction that may promote oral colonization by yeast cells.C. albicans andCandida tropicalis are the yeasts most frequently isolated from the human oral cavity and our data demonstrate that both these species bind toStreptococcus gordonii NCTC 7869 while two otherCandida species (Candida krusei andCandida kefyr) do not. Adherence ofC. albicans was greatest when the yeast had been grown at 30° C to mid-exponential growth phase. For 21 strains ofC. albicans there was a positive correlation between the ability to adhere toS. gordonii and adherence to experimental salivary pellicle. Whole saliva either stimulated or slightly inhibited adherence ofC. albicans toS. gordonii depending on the streptococcal growth conditions. The results suggest that the major salivary adhesins and coaggregation adhesins ofC. albicans are co-expressed.     

Mitochondrial DNA is not fragmented during apoptosis.

We have exposed mouse thymocytes and P-815 mastocytoma cells to four different conditions reported to cause apoptosis: 1) incubation in the absence of mitogenic factors; 2) incubation in the presence of dexamethasone; 3) stimulation with external ATP; 4) treatment with high concentrations of the K+ ionophore valinomycin. These treatments caused DNA fragmentation to a varying extent in the two cell types. High stringency hybridization with a cDNA probe specific to a mitochondrial DNA sequence revealed that during apoptosis induced by lack of mitogenic factors, dexamethasone, or extracellular ATP, mitochondrial DNA was not fragmented. On the contrary, valinomycin caused extensive degradation of mitochondrial DNA. These results support the notion that DNA fragmentation during apoptosis is a specific nuclear event and suggest that other agents, such as valinomycin, may act less selectively.     

Monoclonal antibodies to rabbit liver cytochrome P-448

Cytochrome P-448 (mol wt 55,000 Daltons) from rabbit liver was purified to a specific content of 16.6 nmol/mg. Mice were immunised with this preparation, their spleens removed and dissociated lymphocytes hybridised with myeloma cells. Four monoclonal antibodies against cytochrome P-448 were raised and partially characterised. All four antibodies interacted with cytochrome P-448 in intact microsomal fractions and selectively immunoadsorbed cytochrome P-448 from solubilised microsomal preparations. One of the antibodies inhibited benzo[a] pyrene hydroxylase activity in a reconstituted system, one had no effect on activity and two increased activity. The possible applications of such antibodies are discussed.     

Intratumor heterogeneity of biomarker expression in breast carcinomas

Small biopsy samples are used increasingly to assess the biomarker expression for prognostic information and for monitoring therapeutic responses prior to and during neoadjuvant therapy. The issue of intratumor heterogeneity of expression of biomarkers, however, has raised questions about the validity of the assessment of biomarker expression based on limited tissue samples. We examined immunohistochemically the expression of HER-2neu (p185^erbB-2), epidermal growth factor receptor (EGFR), Bcl-2, p53, and proliferating cell nuclear antigen (PCNA) in 30 breast carcinomas using archived, paraffin embedded tissue and determined the extent of intratumor heterogeneity. Each section was divided into four randomly oriented discrete regions, each containing a portion of the infiltrating carcinoma. For each tumor, the entire lesion and four regions were analyzed for the expression of these markers. Scores of both membrane and cytoplasmic staining of HER-2neu and EGFR, scores of cytoplasmic staining of Bcl-2, and scores of nuclear staining of both p53 and PCNA were recorded. The intensity of staining and the proportion of immunostained cells were determined. A semiquantitative immunoscore was calculated by determining the sum of the products of the intensity and corresponding proportion of stained tumor cells. We analyzed both invasive (IDC) and in situ (DCIS) carcinomas. The Wilcoxon signed-rank test was used for paired comparisons between overall and regional immunoscores and between overall and regional percentages of stained cells. Spearman's correlation coefficients were used to assess the level of agreement of overall biomarker expression with each of the regions. Generalized linear models were used to assess overall and pair-wise differences in the absolute values of percent changes between overall and regional expression of biomarkers. For IDCs, there were no statistically significant differences in the expression of the biomarkers in terms of either the percentage of cells staining or the immunoscores when comparing the entire tumor with each region except for the lower EGFR expression of arbitrarily selected region 1 and lower p53 expression of region 1 compared to that of the entire tumor section. For DCIS, there were no statistically significant differences in the expression of the biomarkers between the entire tumor and each region except in PCNA of region 2 compared to that of entire tumor section. Positive correlation of immunoscores was observed between the entire tumor and each region as well as across all four regions for IDC. Similar observations were noted with DCIS except for HER-2neu and PCNA. No statistically significant differences were observed in the absolute values of percent changes of biomarker expression between overall and the four regions for both DCIS and IDC. Therefore, no significant intratumor heterogeneity in the expression of HER-2neu, Bcl-2, and PCNA was observed in IDC. Minor regional variations were observed for EGFR and p53 in IDC. Similarly, no significant regional variation in the expression of markers was observed in DCIS except for PCNA.     

Role of the Dinaric Karst (western Balkans) in shaping the phylogeographic structure of the threatened crayfish Austropotamobius torrentium

1. This study examines phylogeography and phylogeny of the threatened stone crayfish, Austropotamobius torrentium, in order to elucidate the role of the Dinaric Karst geology in shaping the evolutionary history and genetic diversity of aquatic fauna in the western Balkans. Mitochondrial 16S rRNA and COI genes were partially sequenced from 188 and 159 crayfish, respectively, sampled from 70 localities. Phylogenetic relationships were reconstructed using four methods of phylogenetic inference. Divergence times between phylogroups were estimated in a Bayesian framework, and their demographic history was examined using neutrality tests and mismatch distribution analysis. 2. Seven geographically localised phylogroups separated by pronounced genetic gaps were found. Five of them have a distribution range within the northern‐central Dinaric (NCD) region, while the remaining two include populations from the southern Balkans (SB) and central and south‐eastern Europe (CSE). The oldest divergence event separated two NCD lineages from the rest of populations in the Late Miocene or Early Pliocene. Divergences amongst the five NCD phylogroups and SB + CSE occurred in the Pliocene. The most recent split separated SB and CSE phylogroups during the Late Pliocene. For both genes, uncorrected pairwise divergences between most of the phylogroups (4.1–8.7% for COI and 1.6–4.8% for 16S rRNA) were of the same range as, or higher than, some of the interspecific distances previously reported for the genus Austropotamobius. 3. Geographically isolated and deeply divergent cryptic monophyletic phylogroups within A. torrentium in the NCD region arose in the course of intensification of Neotectonic movements during the Pliocene and the beginning of the Pleistocene and the development of karstification that has heavily fragmented the palaeohydrography of the area. The results confirm a gradual north–south expansion of stone crayfish during the pre‐Pleistocene that preceded the rapid northward post‐glacial re/colonisation of central Europe (CSE phylogroup) through the Danube drainage. 4. Austropotamobius torrentium comprises morphologically cryptic but molecularly distinct taxa. Considering the relatively small geographical areas they inhabit, the NCD phylogroups of stone crayfish should be given the highest conservation priority.     

Genomic dissection of pod shattering in common bean: mutations at non‐orthologous loci at the basis of convergent phenotypic evolution under domestication of leguminous species

The complete or partial loss of shattering ability occurred independently during the domestication of several crops. Therefore, the study of this trait can provide an understanding of the link between phenotypic and molecular convergent evolution. The genetic dissection of ‘pod shattering’ in Phaseolus vulgaris is achieved here using a population of introgression lines and next‐generation sequencing techniques. The ‘occurrence’ of the indehiscent phenotype (indehiscent versus dehiscent) depends on a major locus on chromosome 5. Furthermore, at least two additional genes are associated with the ‘level’ of shattering (number of shattering pods per plant: low versus high) and the ‘mode’ of shattering (non‐twisting versus twisting pods), with all of these loci contributing to the phenotype by epistatic interactions. Comparative mapping indicates that the major gene identified on common bean chromosome 5 corresponds to one of the four quantitative trait loci for pod shattering in Vigna unguiculata. None of the loci identified comprised genes that are homologs of the known shattering genes in Glycine max. Therefore, although convergent domestication can be determined by mutations at orthologous loci, this was only partially true for P. vulgaris and V. unguiculata, which are two phylogenetically closely related crop species, and this was not the case for the more distant P. vulgaris and G. max. Conversely, comparative mapping suggests that the convergent evolution of the indehiscent phenotype arose through mutations in different genes from the same underlying gene networks that are involved in secondary cell‐wall biosynthesis and lignin deposition patterning at the pod level.