Mutant Telomeric Repeats in Yeast Can Disrupt the Negative Regulation of Recombination-Mediated Telomere Maintenance and Create an Alternative Lengthening of Telomeres-Like Phenotype

Some human cancers maintain telomeres using alternative lengthening of telomeres (ALT), a process thought to be due to recombination. In Kluyveromyces lactis mutants lacking telomerase, recombinational telomere elongation (RTE) is induced at short telomeres but is suppressed once telomeres are moderately elongated by RTE. Recent work has shown that certain telomere capping defects can trigger a different type of RTE that results in much more extensive telomere elongation that is reminiscent of human ALT cells. In this study, we generated telomeres composed of either of two types of mutant telomeric repeats, Acc and SnaB, that each alter the binding site for the telomeric protein Rap1. We show here that arrays of both types of mutant repeats present basally on a telomere were defective in negatively regulating telomere length in the presence of telomerase. Similarly, when each type of mutant repeat was spread to all chromosome ends in cells lacking telomerase, they led to the formation of telomeres produced by RTE that were much longer than those seen in cells with only wild-type telomeric repeats. The Acc repeats produced the more severe defect in both types of telomere maintenance, consistent with their more severe Rap1 binding defect. Curiously, although telomerase deletion mutants with telomeres composed of Acc repeats invariably showed extreme telomere elongation, they often also initially showed persistent very short telomeres with few or no Acc repeats. We suggest that these result from futile cycles of recombinational elongation and truncation of the Acc repeats from the telomeres. The presence of extensive 3′ overhangs at mutant telomeres suggests that Rap1 may normally be involved in controlling 5′ end degradation.     

Time-dependent changes in the serum levels of prolactin, nesfatin-1 and ghrelin as a marker of epileptic attacks young male patients

A relationship between hormones and seizures has been reported in animals and humans. Therefore, the purpose of this study was to investigate the association between serum levels of prolactin, nesfatin-1 and ghrelin measured different times after a seizure or non-epileptic event and compared with controls. The study included a total of 70 subjects, and of whom 18 patients had secondary generalized epilepsy (SGE), 16 patients had primary generalized epilepsy (PGE), 16 patients exhibited paroxysmal event (psychogenic) and 20 healthy males were control subjects. The first sample was taken within 5 min of a seizure, with further samples taken after 1, 24, and 48 h so long as the patient did not exhibit further clinically observable seizures; blood samples were taken once from control subjects. Prolactin was measured immediately using TOSOH Bioscience hormone assays. Nesfatin-1 and ghrelin peptides were measured using a commercial immunoassay kit. Patients suffering from focal epilepsy with secondary generalization and primary generalized epilepsy presented with significantly higher levels of serum prolactin and nesfatin-1 and lower ghrelin levels 5 min, 1 and 24 h after a seizure than patients presenting with paroxysmal events (psychogenic) and control subjects; the data were similar but not statistically significant after 48 h. The present study suggests that increased serum prolactin and nesfatin-1 concentrations, decreased ghrelin concentrations could be used as markers to identify patients that have suffered a recent epileptic seizure or other paroxysmal event (psychogenic).     

miRNA-205 suppresses melanoma cell proliferation and induces senescence via regulation of E2F1 protein

MicroRNAs (miRNAs) regulate gene expression by repressing translation or directing sequence-specific degradation of complementary mRNA. Here, we report that expression of miR-205 is significantly suppressed in melanoma specimens when compared with nevi and is correlated inversely with melanoma progression. miRNA target databases predicted E2F1 and E2F5 as putative targets. The expression levels of E2F1 and E2F5 were correlated inversely with that of miR-205 in melanoma cell lines. miR-205 significantly suppressed the luciferase activity of reporter plasmids containing the 3'-UTR sequences complementary to either E2F1 or E2F5. Overexpression of miR-205 in melanoma cells reduced E2F1 and E2F5 protein levels. The proliferative capacity of melanoma cells was suppressed by miR-205 and mediated by E2F-regulated AKT phosphorylation. miR-205 overexpression resulted in induction of apoptosis, as evidenced by increased cleaved caspase-3, poly-(ADP-ribose) polymerase, and cytochrome c release. Stable overexpression of miR-205 suppressed melanoma cell proliferation, colony formation, and tumor cell growth in vivo and induced a senescence phenotype accompanied by elevated expression of p16INK4A and other markers for senescence. E2F1 overexpression in miR-205-expressing cells partially reversed the effects on melanoma cell growth and senescence. These results demonstrate a novel role for miR-205 as a tumor suppressor in melanoma.     

Developmental Competence of Vitrified-Warmed Bovine Oocytes at the Germinal-Vesicle Stage is Improved by Cyclic Adenosine Monophosphate Modulators during In Vitro Maturation

Cryopreservation of mature oocytes and embryos has provided numerous benefits in reproductive medicine. Although successful cryopreservation of germinal-vesicle stage (GV) oocytes holds promise for further advances in reproductive biology and clinical embryology fields, reports regarding cryopreservation of immature oocytes are limited. Oocyte survival and maturation rates have improved since vitrification is being performed at the GV stage, but the subsequent developmental competence of GV oocytes is still low. The purpose of this study was to evaluate the effects of supplementation of the maturation medium with cyclic adenosine monophosphate (cAMP) modulators on the developmental competence of vitrified-warmed GV bovine oocytes. GV oocytes were vitrified-warmed and cultured to allow for oocyte maturation, and then parthenogenetically activated or fertilized in vitro. Our results indicate that addition of a cAMP modulator forskolin (FSK) or 3-isobutyl-1-methylxanthine (IBMX) to the maturation medium significantly improved the developmental competence of vitrified-warmed GV oocytes. We also demonstrated that vitrification of GV oocytes led to a decline in cAMP levels and maturation-promoting factor (MPF) activity in the oocytes during the initial and final phases of maturation, respectively. Nevertheless, the addition of FSK or IBMX to the maturation medium significantly elevated cAMP levels and MPF activity during IVM. Taken together, our results suggest that the cryopreservation-associated meiotic and developmental abnormalities observed in GV oocytes may be ameliorated by an artificial increase in cAMP levels during maturation culture after warming.     

Temporal hierarchy of the visual perceptive systems in the Mondrian world.

Our earlier psychophysical work has shown that colour and motion are not perceived at the same time, with colour leading motion by about 50-100 ms. In pursuing this work, we thought it would be interesting to use a more complex colour stimulus, one in which the wavelength composition of the light reflected or emitted from surfaces changes continually, without entailing a change in the perceived colour (colour constancy). We therefore used a Mondrian figure--an abstract multi-coloured scene with no recognizable objects--against which squares (either red or green) moved up and down, changing colour from red to green in various phase differences with the change in direction of motion. The red and green squares changed continually in their spectral characteristics, as did every other patch on the Mondrian. The results showed that colour is still perceived before motion, by about 80 ms.     

Immunological abnormality of peripheral blood B cells in patients with autoimmune thyroid disease

We investigated the response to immunoglobulin G-secreting cells (ISC) by peripheral blood mononuclear cells (PB-MNC) and purified B cells following stimulation with Staphylococcus aureus Cowan 1 (SAC) or with B cell stimulatory factor 2 (interleukin 6: IL-6), using the reverse hemolytic plaque assay in an attempt to clarify the immunological functions of peripheral blood B cells in patients with autoimmune thyroid disease (AITD). ISC response by PB-MNC following stimulation with SAC was significantly decreased in patients in the hyperthyroid state of Graves' disease and Hashimoto's thyroiditis as compared with that of normal controls. The difference in SAC-response was not significant between patients with euthyroid state of Graves' disease and normal controls. ISC response by PB-MNC following stimulation with SAC exhibited a reciprocal relationship to TRAb in patients with Graves' disease. Using purified B cells, some spontaneous ISC response without SAC stimulation was observed in patients in the hyperthyroid state of Graves' disease and Hashimoto's thyroiditis. This spontaneous ISC response was further enhanced by IL-6. These results suggest that in organ-specific autoimmune diseases such as AITD, immunological abnormalities exist in B cells and some B cells are nonspecifically activated in the immunologically active state.     

The Drug Release Study of Ceftriaxone from Porous Hydroxyapatite Scaffolds

Hydroxyapatite (HAP) is an important biomedical material that is used for grafting osseous defects. It has an excellent bioactivity and biocompatibility properties. To isolate hydroxyapatite, pieces of cleaned cattle’s bone were heated at different temperature range from 400°C up to 1,200°C. A reasonable yield of 60.32% w/w HAP was obtained at temperature range from 1,000°C to 1,200°C. Fourier transform infrared spectra and the thermogravimetric measurement showed a clear removal of organic at 600°C as well as an excellent isolation of HAP from the bones which was achieved at 1,000–1,200°C. This was also confirmed from X-ray diffraction of bone sample heated at 1,200°C. The concentration ions were found to be sodium, potassium, lithium, zinc, copper, iron, calcium, magnesium, and phosphate present in bones within the acceptable limits for its role in the bioactivity property of HAP. Glucose powder was used as a porosifier. Glucose was novel and excellent as porogen where it was completely removed by heating, giving an efficient porosity in the used scaffolds. The results exhibited that the ceftriaxone drug release was increased with increasing the porosity. It was found that a faster, higher, and more regular drug release was obtained from the scaffold with a porosity of 10%.