Physical performance and clinical outcomes in dialysis patients: a secondary analysis of the EXCITE trial.

Background/Aims: Scarce physical activity predicts shorter survival in dialysis patients. However, the relationship between physical (motor) fitness and clinical outcomes has never been tested in these patients. Methods: We tested the predictive power of an established metric of motor fitness, the Six-Minute Walking Test (6MWT), for death, cardiovascular events and hospitalization in 296 dialysis patients who took part in the trial EXCITE (ClinicalTrials.gov Identifier: NCT01255969). Results: During follow up 69 patients died, 90 had fatal and non-fatal cardiovascular events, 159 were hospitalized and 182 patients had the composite outcome. In multivariate Cox models - including the study allocation arm and classical and non-classical risk factors - an increase of 20 walked metres during the 6MWT was associated to a 6% reduction of the risk for the composite end-point (P=0.001) and a similar relationship existed between the 6MWT, mortality (P<0.001) and hospitalizations (P=0.03). A similar trend was observed for cardiovascular events but this relationship did not reach statistical significance (P=0.09). Conclusions: Poor physical performance predicts a high risk of mortality, cardiovascular events and hospitalizations in dialysis patients. Future studies, including phase-2 EXCITE, will assess whether improving motor fitness may translate into better clinical outcomes in this high risk populatio

Survey on retinopathy of prematurity (ROP) in Italy

This study aims to investigate the incidence and the relative risk factors of retinopathy of prematurity (ROP) and posterior-ROP (P-ROP): ROP in Zone I and posterior Zone II, as well as to analyze the occurrence of surgical treatment of ROP and to evaluate the short term outcome of the disease in Italy

Replacement of Vegetative ς(A) by Sporulation-Specific ς(F) as a Component of the RNA Polymerase Holoenzyme in Sporulating Bacillus subtilis

Soon after asymmetric septation in sporulating Bacillus subtilis cells, ς(F) is liberated in the prespore from inhibition by SpoIIAB. To initiate transcription from its cognate promoters, ς(F) must compete with ς(A), the housekeeping sigma factor in the predivisional cell, for binding to core RNA polymerase (E). To estimate the relative affinity of E for ς(A) and ς(F), we made separate mixtures of E with each of the two sigma factors, allowed reconstitution of the holoenzyme, and measured the concentration of free E remaining in each mixture. The affinity of E for ς(F) was found to be about 25-fold lower than that for ς(A). We used quantitative Western blotting to estimate the concentrations of E, ς(A), and ς(F) in sporulating cells. The cellular concentrations of E and ς(A) were both about 7.5 μM, and neither changed significantly during the first 3 h of sporulation. The concentration of ς(F) was extremely low at the beginning of sporulation, but it rose rapidly to a peak after about 2 h. At its peak, the concentration of ς(F) was some twofold higher than that of ς(A). This difference in concentration cannot adequately account for the replacement of ς(A) holoenzyme by ς(F) holoenzyme in the prespore, and it seems that some further mechanism—perhaps the synthesis or activation of an anti-ς(A) factor—must be responsible for this replacement

Functional assay for BRCA1: mutagenesis of the COOH-terminal region reveals critical residues for transcription activation

The breast and ovarian cancer susceptibility gene product BRCA1 is a tumor suppressor but its precise biochemical function remains unknown. The BRCA1 C-terminus acts as a transcription activation domain and germ-line cancer-predisposing mutations in this region abolish transcription activation whereas benign polymorphisms do not. These results raise the possibility that loss of transcription activation by BRCA1 is crucial for oncogenesis. Therefore, identification of residues involved in transcription activation by BRCA1 will help understand why particular germ-line missense mutations are deleterious and may provide more reliable presymptomatic risk assessment. The BRCA1 C-terminus (aa 1560–1863) consists of two BRCT (BRCA1 C-terminal) domains preceded by a region likely to be nonglobular. We combined site-directed and random mutagenesis followed by a functional transcription assay in yeast. First, error prone PCR-induced random mutagenesis generated eight unique missense mutations causing loss of function, six of which targeted hydrophobic residues conserved in canine, mouse, rat and human BRCA1. Second, random insertion of a variable pentapeptide cassette generated 21 insertion mutants. All pentapeptide insertions N-terminal to the BRCT domains retained wild-type activity whereas insertions in the BRCT domains were, with few exceptions, deleterious. Third, site-directed mutagenesis was used to characterize five known germ-line mutations and to perform deletion analysis of the C-terminus. Deletion analysis revealed that the integrity of the most C-terminal hydrophobic cluster (I1855, L1854, and Y1853) is necessary for activity. We conclude that the integrity of the BRCT domains is crucial for transcription activation and that hydrophobic residues may be important for BRCT function. Therefore, the yeast-based assay for transcription activation can be used successfully to provide tools for structure-function analysis of BRCA1 and may form the basis of a BRCA1 functional assay

Cleavage/polyadenylation factor IA associates with the carboxyl-terminal domain of RNA polymerase II in Saccharomyces cerevisiae

The carboxyl-terminal domain (CTD) of the largest subunit of RNA polymerase II plays an important role in transcription and processing of the nascent transcript by interacting with both transcription and RNA processing factors. We show here that the cleavage/polyadenylation factor IA of Saccharomyces cerevisiae directly contacts CTD. First by affinity chromatography experiments with yeast extracts we demonstrate that the Rna15p, Rna14p, and Pcf11p subunits of this complex are associated with phosphorylated CTD. This interaction is confirmed for Rna15p by yeast two-hybrid analysis. Second, Pcf11p, but not Rna15p, is shown to directly contact phosphorylated CTD based on in vitro binding studies with recombinant proteins. These findings establish a direct interaction of cleavage/polyadenylation factor IA with the CTD. Furthermore, a quantitative analysis of transcription run-on performed on temperature-sensitive mutant strains reveals that the lack of either functional Rna14p or Pcf11p affects transcription termination more severely than the absence of a functional Rna15p. Moreover, these data reinforce the concept that CTD phosphorylation acts as a regulatory mechanism in the maturation of the primary transcript