In-vivo microvascular imaging of early squamous-cell cancer of the esophagus by confocal laser endomicroscopy.

We present the case of a 63-year-old man with an early esophageal squamous-cell cancer that was visualized and analyzed by fluorescein-aided confocal laser endomicroscopic imaging. Our findings indicate that endomicroscopy is readily able to provide images of microvessels and to differentiate between normal and abnormal tumor vessels. Endomicroscopy therefore has the potential to characterize microvascular changes in early squamous-cell cancer, suggesting that this technique might have major implications for optimizing treatment strategies in modern endoscopy.

Attempted endoscopic en bloc resection of mucosal and submucosal tumors using insulated-tip knives: a pilot series.

BACKGROUND AND STUDY AIMS: Endoscopic mucosal resection (EMR) of early gastrointestinal cancers has been shown to be effective in treating mucosal malignancies, but en bloc resection (where the entire tumor is removed in one piece) is often not achieved using conventional cap EMR. Other techniques, developed in Japan, include the application of different types of knife such as the insulated-tip instrument. We report our preliminary experience of the use of this knife, in conjunction with other techniques, in attempting en bloc resection of early mucosal cancers and adenomas and in the removal of submucosal tumors (SMTs) of the upper gastrointestinal tract. PATIENTS AND METHODS: A total of 37 patients (26 men, 11 women, age range 53 - 86) were included in the study; 23 patients had 24 mucosal lesions amenable to EMR, and 14 patients had SMTs shown on endosonography to spare the muscularis propria. Lesions were located in the esophagus (n = 13), the stomach (n = 24), and the duodenum (n = 1); 40 % of the mucosal lesions were 20 mm or larger (mean size 18mm), whereas the mean size of the submucosal lesions was 23 mm. After submucosal saline injection, circumcision and dissection of the mucosal lesions was attempted with the aim of achieving en bloc resection. For SMTs, cap mucosectomy of the overlying mucosa was done first, and the tumors were then freed using saline injection, and finally resected using snare polypectomy. RESULTS: The strict aim of the study, i. e. complete tumor removal in a single piece, was achieved in only 25 % of the mucosal lesions (some failures were due to unrecognized submucosal infiltration) and 36 % of the SMTs. When a more liberal definition of success was assumed, this rate increased to 65 % for mucosal lesions (piecemeal, no tumor found at surgery or follow-up endoscopy with biopsy) and 79 % for SMTs (piecemeal). No severe complications necessitating surgery or leading to major morbidity occurred. However, clinically significant complications were found in six patients (minor perforation managed conservatively (n = 1), severe pain without perforation (n = 1), bleeding requiring reintervention (n = 3), and aspiration (n = 1)). CONCLUSIONS: Although we are convinced that methods of achieving en bloc resection of mucosal cancers and SMTs must be pursued, the insulated-tip knife in conjunction with conventional endoscopes still has limitations. Innovative endoscope design (double-channel scopes) as well as the development of new accessories will help to overcome the current limitations and further promote endoscopic tumor resection.

The intracellular location of two aminoacyl-tRNA synthetases depends on complex formation with Arc1p.

In yeast, two aminoacyl-tRNA synthetases, MetRS and GluRS, are associated with Arc1p. We have studied the mechanism of this complex formation and found that the non-catalytic N-terminally appended domains of MetRS and GluRS are necessary and sufficient for binding to Arc1p. Similarly, it is the N-terminal domain of Arc1p that contains distinct but overlapping binding sites for MetRS and GluRS. Localization of Arc1p, MetRS and GluRS in living cells using green fluorescent protein showed that these three proteins are cytoplasmic and largely excluded from the nucleus. However, when their assembly into a complex is inhibited, significant amounts of MetRS, GluRS and Arc1p can enter the nucleus. We suggest that the organization of aminoacyl-tRNA synthetases into a multimeric complex not only affects catalysis, but is also a means of segregating the tRNA- aminoacylation machinery mainly to the cytoplasmic compartment.

Biogenesis of the signal recognition particle (SRP) involves import of SRP proteins into the nucleolus, assembly with the SRP-RNA, and Xpo1p-mediated export.

The signal recognition particle (SRP) targets nascent secretory proteins to the ER, but how and where the SRP assembles is largely unknown. Here we analyze the biogenesis of yeast SRP, which consists of an RNA molecule (scR1) and six proteins, by localizing all its components. Although scR1 is cytoplasmic in wild-type cells, nuclear localization was observed in cells lacking any one of the four SRP "core proteins" Srp14p, Srp21p, Srp68p, or Srp72p. Consistently, a major nucleolar pool was detected for these proteins. Sec65p, on the other hand, was found in both the nucleoplasm and the nucleolus, whereas Srp54p was predominantly cytoplasmic. Import of the core proteins into the nucleolus requires the ribosomal protein import receptors Pse1p and Kap123p/Yrb4p, which might, thus, constitute a nucleolar import pathway. Nuclear export of scR1 is mediated by the nuclear export signal receptor Xpo1p, is distinct from mRNA transport, and requires, as evidenced by the nucleolar accumulation of scR1 in a dis3/rrp44 exosome component mutant, an intact scR1 3' end. A subset of nucleoporins, including Nsp1p and Nup159p (Rat7p), are also necessary for efficient translocation of scR1 from the nucleus to the cytoplasm. We propose that assembly of the SRP requires import of all SRP core proteins into the nucleolus, where they assemble into a pre-SRP with scR1. This particle can then be targeted to the nuclear pores and is subsequently exported to the cytoplasm in an Xpo1p-dependent way.

Arc1p organizes the yeast aminoacyl-tRNA synthetase complex and stabilizes its interaction with the cognate tRNAs.

Eukaryotic aminoacyl-tRNA synthetases, in contrast to their prokaryotic counterparts, are often part of high molecular weight complexes. In yeast, two enzymes, the methionyl- and glutamyl-tRNA synthetases associate in vivo with the tRNA-binding protein Arc1p. To study the assembly and function of this complex, we have reconstituted it in vitro from individually purified recombinant proteins. Our results show that Arc1p can readily bind to either or both of the two enzymes, mediating the formation of the respective binary or ternary complexes. Under competition conditions, Arc1p alone exhibits broad specificity and interacts with a defined set of tRNA species. Nevertheless, the in vitro reconstituted Arc1p-containing enzyme complexes can bind only to their cognate tRNAs and tighter than the corresponding monomeric enzymes. These results demonstrate that the organization of aminoacyl-tRNA synthetases with general tRNA-binding proteins into multimeric complexes can stimulate their catalytic efficiency and, therefore, offer a significant advantage to the eukaryotic cell.

Cloning and characterization of the Schizosaccharomyces pombe tRNA:pseudouridine synthase Pus1p.

Saccharomyces cerevisiae cells that carry deletions in both the LOS1 (a tRNA export receptor) and the PUS1 (a tRNA:pseudouridine synthase) genes exhibit a thermosensitive growth defect. A Schizosaccharomyces pombe gene, named spPUS1, was cloned from a cDNA library by complementation of this conditional lethal phenotype. The corresponding protein, spPus1p, shows sequence similarity to S. cerevisiae and murine Pus1p as well as other known members of the pseudouridine synthase family. Accordingly, recombinant spPus1p can catalyze in vitro the formation of pseudouridines at positions 27, 28, 34, 35 and 36 of yeast tRNA transcripts. The sequence and functional conservation of the Pus1p proteins in fungi and mammalian species and their notable absence from prokaryotes suggest that this family of pseudouridine synthases is required for a eukaryote-specific step of tRNA biogenesis, such as nuclear export.

[ESWL-therapy of pancreatic duct calculi]. / ESWL-Therapie von Pankreasgangsteinen.

In 23 patients suffering from pancreatic duct stones, extracorporeal shock wave lithotripsy (ESWL) was performed in combination with endoscopic sphincterotomy (EST). Calculi-disintegration and resolution of obstruction was achieved in all cases. Completely stone-free ducts were achieved in 7 patients, some peripheral asymptomatic stone material remained in 16. 8 patients became completely asymptomatic, 11 reported a marked reduction of their pain. 15 patients gained weight. No major complications were observed. ESWL combined with EST is a successful non-operative new treatment option in pancreatic stone disease.