Circadian Rhythms of Rat Pancreatic Acinar Cells

On the basis of the circadian oscillations of the rat's exocrine pancreatic function and previous reports on concomitant ultrastructural changes in the pancreatic tissue, we analysed stereologically the circadian rhythmicity in the structure of this organ. Twenty-four male Wistar rats, four and a half months old, were singly housed two months before the experiment in a lighting regimen LD=12:12, constant environmental temperature and relative humidity, with food and water ad libitum. The experiment was performed in winter. The rats were randomly divided into 6 balanced groups and killed under ether anesthesia at 6 equidistant time points in 24 hours. The pancreatic tissue was fixed in glutaraldehyde and osmium and embedded in Epon. 1 µm thick sections were examined by light microscopy for the evaluation by stereological methods of: a) volume fractions of the different parenchymal components of the exocrine pancreas; b) surface fractions of acinar cell faces; c) size distribution of acinar cell nuclei, their number per unit tissue volume and their mean diameter. Single cosinor method analysis of the data demonstrated statistically significant circadian rhythms for the volume fraction of the cytoplasm of acinar cells and the volume fractions of pancreatic acini and acinar cells. The volume fraction of the cytoplasm of the rat pancreatic acinar cells undergoes circadian oscillations with the highest values at the end of the light span; this rise precedes the well-known physiological nocturnal surge of pancreatic digestive enzymes. Our findings further support the hypothesis of a close relationship between pancreatic cell structure and its function.     

The full oncogenic activity of Ret/ptc2 depends on tyrosine 539, a docking site for phospholipase Cgamma.

RET/PTC oncogenes, generated by chromosomal rearrangements in papillary thyroid carcinomas, are constitutively activated versions of proto-RET, a gene coding for a receptor-type tyrosine kinase (TK) whose ligand is still unknown. RET/PTCs encode fusion proteins in which proto-RET TK and C-terminal domains are fused to different donor genes. The respective Ret/ptc oncoproteins display constitutive TK activity and tyrosine phosphorylation. We found that Ret/ptcs associate with and phosphorylate the SH2-containing transducer phospholipase Cgamma (PLCgamma). Two putative PLCgamma docking sites, Tyr-505 and Tyr-539, have been identified on Ret/ptc2 by competition experiments using phosphorylated peptides modelled on Ret sequence. Transfection experiments and biochemical analysis using Tyr-->Phe mutants of Ret/ptc2 allowed us to rule out Tyr-505 and to identify Tyr-539 as a functional PLCgamma docking site in vivo. Moreover, kinetic measurements showed that Tyr-539 is able to mediate high-affinity interaction with PLCgamma. Mutation of Tyr-539 resulted in a drastically reduced oncogenic activity of Ret/ptc2 on NIH 3T3 cells (75 to 90% reduction) both in vitro and in vivo, which correlates with impaired ability of Ret/ptc2 to activate PLCgamma. In conclusion, this paper demonstrates that Tyr-539 of Ret/ptc2 (Tyr-761 on the proto-RET product) is an essential docking site for the full transforming potential of the oncogene. In addition, the present data identify PLCgamma as a downstream effector of Ret/ptcs and suggest that this transducing molecule could play a crucial role in neoplastic signalling triggered by Ret/ptc oncoproteins.     

Cholesterol Conjugated Uniform and Gapmer Phosphorothioate Oligonucleotides Targeted Against PKC-α and C-raf Gene Expression

Abstract

In vitra and in vivo antitumor activity of phosphorothioate antisense oligonucleotides targeted against two protein kinases within the mitogen-activated protein (MAP) kinase signaling cascade has been well documented by ISIS 3521/CGP 6412XA (targeted against PKC-α protein) and ISIS 5132KGP69846A (targeted against C-rwfl kinase). For both of these compounds, cationic lipid formulations are necessary to observe any pharmacological activity in cell culture. In contrast, in vivo functional delivery of phosphorothioate oligonucleotides to cells in tissues does not appear to be a prohlem. These oligonucleotides have demonstrated reduction in either PKC-α or C-raf gene expression in tissues or human tumor xenografts following systemic administration.     

Just another island dwarf? Phenotypic distinctiveness in the poorly known Soemmerring's Gazelle,Nanger soemmerringii (Cetartiodactyla: Bovidae), of Dahlak Kebir Island

The gazelles of Dahlak Kebir are the only population of Nanger soemmerringii Cretzschmar, 1828 living on an island. Little is known on the biology of these animals, except that they are evidently smaller than their conspecifics living on the continent. We took advantage of a recently acquired collection of crania, probably the largest available study sample of the Dahlak Kebir population worldwide, to explore the phenotypic variation of this island endemism. To this aim, we employed state of the art geometric morphometrics techniques and multivariate statistics to compare the insular population with samples of two out of three subspecies of N. soemmerringii from continental Africa. We found that not only is the size of the animal remarkably smaller in Dahlak Kebir gazelles, but their cranial shape is also highly distinctive, and this might be only partly explained by allometry. We also showed that phenotypic variance might have been reduced in the island population, likely as a consequence of genetic bottlenecks. This unique population is part of a species vulnerable to extinction. Our results suggest that the Dahlak Kebir Island gazelles might represent a significant component of its variation and potential for adaptive change and evolution. More information, including molecular data, and an accurate assessment of its taxonomic relevance and conservation status, is urgently needed. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 111 , 603–620.     

Genotyping cell-free tumor DNA in the blood to detect residual disease and drug resistance

DNA fragments released from cancer cells into the blood can be used to generate molecular profiles of tumors. Non-invasive ''liquid biopsies'' can be used to monitor minimal residual disease and detect the emergence of drug resistance.     

Epigenetic reprogramming of breast cancer cells by valproic acid occurs regardless of estrogen receptor status

Estrogen receptors (ERs) are a recognized prognostic factor and therapeutic target in breast cancer. The loss of ER expression relates to poor prognosis, poor clinical outcome and impairs the use of anti-estrogenic treatment. Histone deacetylase inhibitors are candidate drugs for cancer therapy. Among them, valproic acid (VPA) is a long used and safe anti-epileptic drug. We studied the biological consequences of the chromatin remodeling action of VPA in a normal human mammary epithelial cell line and in ERalpha-positive and ERalpha-negative breast cancer cell lines. In these cells and regardless of their ER status, VPA-induced cell differentiation, as shown by increased milk lipids production, decreased expression of the CD44 antigen and growth arrest in the G(0)-G(1) phase of the cell cycle. These effects were accompanied by decreased Rb phosphorylation, hyperacetylation of the p21(WAF1/CIP1) gene promoter and increased p21 protein expression. Only in breast cancer cells, cyclin B1 expression was decreased and the cells accumulated also in G(2). ERalpha expression decreased in ERalpha-positive, increased in ERalpha-negative and was unchanged in normal mammary epithelial cells, as did the expression of progesterone receptor, a physiological ERalpha target. VPA decreased the expression of the invasiveness marker pS2 in ERalpha-positive breast cancer cells, but did not cause its re-expression in ERalpha-negative cells. Overall, these data suggest that in both ERalpha-positive and -negative malignant mammary epithelial cells VPA reprograms the cells to a more differentiated and "physiologic" phenotype that may improve the sensitivity to endocrine therapy and/or chemotherapy in breast cancer patients.     

Two naphthalene degrading bacteria belonging to the genera Paenibacillus and Pseudomonas isolated from a highly polluted lagoon perform different sensitivities to the organic and heavy metal contaminants

Two bacterial strains were isolated in the presence of naphthalene as the sole carbon and energy source from sediments of the Orbetello Lagoon, Italy, which is highly contaminated with both organic compounds and metals. 16S rRNA gene sequence analysis of the two isolates assigned the strains to the genera Paenibacillus and Pseudomonas. The effect of different contaminants on the growth behaviors of the two strains was investigated. Pseudomonas sp. ORNaP2 showed a higher tolerance to benzene, toluene, and ethylbenzene than Paenibacillus sp. ORNaP1. In addition, the toxicity of heavy metals potentially present as co-pollutants in the investigated site was tested. Here, strain Paenibacillus sp. ORNaP1 showed a higher tolerance towards arsenic, cadmium, and lead, whereas it was far more sensitive towards mercury than strain Pseudomonas sp. ORNaP2. These differences between the Gram-negative Pseudomonas and the Gram-positive Paenibacillus strain can be explained by different general adaptive response systems present in the two bacteria.