Nucleologenesis in the Caenorhabditis elegans Embryo

In the Caenorhabditis elegans nematode, the oocyte nucleolus disappears prior to fertilization. We have now investigated the re-formation of the nucleolus in the early embryo of this model organism by immunostaining for fibrillarin and DAO-5, a putative NOLC1/Nopp140 homolog involved in ribosome assembly. We find that labeled nucleoli first appear in somatic cells at around the 8-cell stage, at a time when transcription of the embryonic genome begins. Quantitative analysis of radial positioning showed the nucleolus to be localized at the nuclear periphery in a majority of early embryonic nuclei. At the ultrastructural level, the embryonic nucleolus appears to be composed of a relatively homogenous core surrounded by a crescent-shaped granular structure. Prior to embryonic genome activation, fibrillarin and DAO-5 staining is seen in numerous small nucleoplasmic foci. This staining pattern persists in the germline up to the ∼100-cell stage, until the P4 germ cell divides to give rise to the Z2/Z3 primordial germ cells and embryonic transcription is activated in this lineage. In the ncl-1 mutant, which is characterized by increased transcription of rDNA, DAO-5-labeled nucleoli are already present at the 2-cell stage. Our results suggest a link between the activation of transcription and the initial formation of nucleoli in the C. elegans embryo

Uncoupled Embryonic and Extra-Embryonic Tissues Compromise Blastocyst Development after Somatic Cell Nuclear Transfer

Somatic cell nuclear transfer (SCNT) is the most efficient cell reprogramming technique available, especially when working with bovine species. Although SCNT blastocysts performed equally well or better than controls in the weeks following embryo transfer at Day 7, elongation and gastrulation defects were observed prior to implantation. To understand the developmental implications of embryonic/extra-embryonic interactions, the morphological and molecular features of elongating and gastrulating tissues were analysed. At Day 18, 30 SCNT conceptuses were compared to 20 controls (AI and IVP: 10 conceptuses each); one-half of the SCNT conceptuses appeared normal while the other half showed signs of atypical elongation and gastrulation. SCNT was also associated with a high incidence of discordance in embryonic and extra-embryonic patterns, as evidenced by morphological and molecular “uncoupling”. Elongation appeared to be secondarily affected; only 3 of 30 conceptuses had abnormally elongated shapes and there were very few differences in gene expression when they were compared to the controls. However, some of these differences could be linked to defects in microvilli formation or extracellular matrix composition and could thus impact extra-embryonic functions. In contrast to elongation, gastrulation stages included embryonic defects that likely affected the hypoblast, the epiblast, or the early stages of their differentiation. When taking into account SCNT conceptus somatic origin, i.e. the reprogramming efficiency of each bovine ear fibroblast (Low: 0029, Med: 7711, High: 5538), we found that embryonic abnormalities or severe embryonic/extra-embryonic uncoupling were more tightly correlated to embryo loss at implantation than were elongation defects. Alternatively, extra-embryonic differences between SCNT and control conceptuses at Day 18 were related to molecular plasticity (high efficiency/high plasticity) and subsequent pregnancy loss. Finally, because it alters re-differentiation processes in vivo, SCNT reprogramming highlights temporally and spatially restricted interactions among cells and tissues in a unique way

Docetaxel and prednisone with or without lenalidomide in chemotherapy-naive patients with metastatic castration-resistant prostate cancer (MAINSAIL): a randomised, double-blind, placebo-controlled phase 3 trial

Background Patients with metastatic castration-resistant prostate cancer have few treatment options. We investigated the safety and efficacy of lenalidomide, an immunomodulatory agent with anti-angiogenic properties, in combination with docetaxel and prednisone in chemotherapy-naive patients with metastatic castration-resistant prostate cancer. Methods In this randomised, double-blind, placebo-controlled, phase 3 study, we randomly assigned chemotherapy-naive patients with progressive metastatic castration-resistant prostate cancer in a 1: 1 ratio to receive docetaxel (75 mg/m(2)) on day 1 and prednisone (5 mg twice daily) on days 1-21 and either lenalidomide (25 mg) or placebo once daily on days 1-14 of each 21 day treatment cycle. Permuted block randomisation was done with an interactive voice response system and stratified by Eastern Cooperative Oncology Group performance status, geographic region, and type of disease progression. Clinicians, patients, and investigators were masked to treatment allocation. The primary endpoint was overall survival. Efficacy analysis was by intention to treat. Patients who received at least one dose of study drug were included in the safety analyses. This study is registered with ClinicalTrials.gov, number NCT00988208. Findings 1059 patients were enrolled and randomly assigned between Nov 11, 2009, and Nov 23, 2011 (533 to the lenalidomide group and 526 to the control group), and 1046 patients received study treatment (525 in the lenalidomide group and 521 in the placebo group). At data cutoff (Jan 13, 2012) after a median follow-up of 8 months (IQR 5-12), 221 patients had died: 129 in the lenalidomide group and 92 in the placebo group. Median overall survival was 17.7 months (95% CI 14.8-18.8) in the lenalidomide group and not reached in the placebo group (hazard ratio [HR] 1.53, 95% CI 1.17-2.00, p= 0.0017). The trial was subsequently closed early due to futility. The number of deaths that occurred during treatment or less than 28 days since the last dose were similar in both groups (18 [3%] of 525 patients in the lenalidomide group vs 13 [2%] of 521 patients). 109 (21%) patients in the lenalidomide group and 78 (15%) in the placebo group died more than 28 days from last dose, mainly due to disease progression. At least one grade 3 or higher adverse event was reported in 381 (73%) of 525 patients receiving lenalidomide and 303 (58%) of 521 patients receiving placebo. Grade 3-4 neutropenia (114 [22%] for lenalidomide vs 85 [16%] for placebo), febrile neutropenia (62 [12%] vs 23 [4%]), diarrhoea (37 [7%] vs 12 [2%]), pneumonia (24 [5%] vs five [1%]), dyspnoea (22 [4%] vs nine [2%]), asthenia (27 [5%] vs 17 [3%]), and pulmonary embolism (32 [6%] vs seven [1%]) occurred more frequently in the lenalidomide group than in the placebo group. Interpretation Overall survival with the combination of lenalidomide, docetaxel, and prednisone was significantly worse than with docetaxel and prednisone for chemotherapy-naive men with metastatic, castration-resistant prostate cancer. Further research with this treatment combination is not warranted