Purification and characterization of MAR1. A mitochondrial associated ribonuclease from Leishmania tarentolae

A relatively thermostable 22-kDa endoribonuclease (MAR1) was purified more than 10,000-fold from a mitochondrial extract of Leishmania tarentolae and the gene cloned. The purified nuclease has a Km of 100-145 +/- 33 nM and a Vmax of 1.8-2.9 +/- 2 nmol/min, depending on the RNA substrate, and yields a 3'-OH and a 5'-phosphate. Cleavage was limited to several specific sites in the substrate RNAs tested, but cleavage of pre-edited RNAs was generally independent of the addition of cognate guide RNA. The MAR1 gene was expressed in Escherichia coli or in L. tarentolae cells, and the recombinant protein was affinity-purified. The cleavage specificity of the recombinant enzyme from L. tarentolae was identical to that of the native enzyme. The single copy MAR1 gene maps to an 820-kilobase pair chromosome and contains an open reading frame of 579 nucleotides. The 18-amino acid N-terminal sequence shows characteristics of an uncleaved mitochondrial targeting sequence. Data base searching revealed two homologues of MAR1 corresponding to unidentified open reading frames in Caenorhabditis elegans (GenBankTM accession number Z69637) and Archaeoglobus fulgidus (GenBankTM accession number AE000943). The function of MAR1 in mitochondrial RNA metabolism in L. tarentolae remains to be determined.     

Osteopenia in children surviving brain tumours

Mature and post-translational precursor gastrin forms are growth factors for colorectal tumours. The immunogen Gastrimmune is composed of the amino terminus of gastrin-17 linked to diphtheria toxoid and raises antibodies in situ which neutralise amidated and glycine-extended gastrin-17. The aim of the study was to determine the effect of treatment with 5-fluorouracil(5-FU)/leucovorin on the antibody titres induced by Gastrimmune and the effect of combination therapy on the growth of the rat colon tumour DHDK12. Gastrimmune was administered to rats s.c. at 3 weekly intervals. The rat colon tumour line DHDK12 was injected into the abdominal wall of BDIX rats. Combinations of 5-FU/leucovorin were injected i.v. on days 1, 3 and 5, with the cycle repeated every 4 weeks. Antibody titres were measured by an ELISA technique. Antibody titres were followed for 40 weeks after Gastrimmune (500 microg.ml(-1)) immunization, with titres peaking between 10 and 20 weeks after a single immunisation and falling by week 30. At termination, no effect was observed on either the histological appearance of the gastro-intestinal tract or the proliferation of the colonic mucosa. Pre- and post-treatment with 5-FU/leucovorin (30 mg.kg(-1)) had no effect on the kinetics and level of antibody response to Gastrimmune. Gastrimmune (200 microg.ml(-1)) and 5-FU/leucovorin combinations (12.5 and 20 mg.kg(-1)) increased the therapeutic effects on the in vivo growth of DHDK12 tumors when compared to the agents given singly. Gastrimmune immunisation may be a therapeutic option for the treatment of colorectal cancer in combination with 5-FU/leucovorin.     

Visualisation of extensive water ribbons and networks in a DNA minor-groove drug complex

The crystal structure is reported of a complex between an ethyl derivative of the minor-groove drug furamidine and the dodecanucleotide duplex d(CGCGAATTCGCG)2, which has been refined to 1.85 A resolution and an R factor of 16.6% for data collected at -173 degreesC. An exceptionally large number (220) of water molecules have been located. The majority of these occur in the first coordination shell of solvation. There are extensive networks of connected waters, both in the major and minor grooves. In particular, there are 21 water molecules associated with the minor-groove drug, via hydrogen bonds from the four charged nitrogen atoms. One cluster of four waters is situated in the groove itself; the majority are on the outer edge of the groove, and serve to bridge between the outward-directed drug nitrogen atoms and backbone phosphate oxygen atoms. These bridges are both intra- and inter-strand, with the net effect that the outer edge of the drug molecule is covered by ribbons of water molecules.     

LDL particle size in subjects with previously unsuspected coronary heart disease: relationship with other cardiovascular risk markers

PURPOSE: We report a retrospective series of 850 patients treated by external irradiation for carcinoma of the eyelid at Institut Curie and we compare our results with other techniques: brachytherapy and surgery. METHODS AND MATERIALS: Eight hundred fifty patients were treated by external radiotherapy for carcinoma of the eyelid. None of these patients have been previously treated. All the patients were classified according to the TNM classification of (UICC). We distinguished five histological types and five clinical groups according to the site of the skin tumor. Three modalities of external radiotherapy were used: contact therapy, conventional radiotherapy, and electrontherapy. We reviewed the clinical files of the 850 patients who went regularly at follow-up visits. RESULTS: We report the 5-year survival results--alive with no evidence of disease: 72%; alive with progression: 2%; died from tumor progression: 0.5%; died from intercurrent disease: 19.5%; and lost to follow-up: 5%. The 5-year local control rate was 97.5%. We observed 45 failures--lymph node, metastatic, and local--and emphasize this last group by presenting the results of treatment of these local failures. We studied the complications of treatment: 2.3% of corneal complications, 2% of cataracts, and 1.4% of serious ocular complications. CONCLUSIONS: Our results concerning local failures and loss of the eye are comparable to those reported for other techniques involving brachytherapy or surgery. Overall, external radiotherapy is a safe and effective treatment, as it ensures a high local control rate and provides perfectly satisfactory functional and esthetic results. It seemed particularly useful to report this series in that few publications are available on this subject that, nevertheless, constitutes a topical issue.     

Bupropion and desipramine increase dopamine transporter mRNA expression in the ventral tegmental area/substantia nigra of rat brain

The homeobox gene Otx2 is a mouse cognate of the Drosophila orthodenticle gene, which is required for development of the brain, rostral to rhombomere three. We have investigated the mechanisms involved in this neural function and specifically the requirement for Otx2 in the visceral endoderm and the neuroectoderm using chimeric analysis in mice and explant recombination assay. Analyses of chimeric embryos composed of more than 90% of Otx2-/- ES cells identified an essential function for Otx2 in the visceral endoderm for induction of the forebrain and midbrain. The chimeric studies also demonstrated that an anterior neural plate can form without expressing Otx2. However, in the absence of Otx2, expression of important regulatory genes, such as Hesx1/Rpx, Six3, Pax2, Wnt1 and En, fail to be initiated or maintained in the neural plate. Using explant-recombination assay, we could further demonstrate that Otx2 is required in the neuroectodem for expression of En. Altogether, these results demonstrate that Otx2 is first required in the visceral endoderm for the induction, and subsequently in the neuroectoderm for the specification of forebrain and midbrain territories.     

Ergonomic working conditions for the nursing team in a cardiology unit]

Aiming at finding out the ergonomics conditions of the nursing personnel in a hospital admission unit, the authors utilized as a methodological presupposition the search for a global experienced work situation through the analysis of some elements as: the man (nursing worker); the work activity (prescribed duty, real activities and work position) and the environment (temperature, noise and illumination). The authors selected the Cardiology admission unit for the site of their research. The results verified that the nursing workers the mentioned unit face problems related to salaries, work installations, relationship, work organization, levels of formation and the activities executed. The authors suggested some recommendations.     

Frequency-stabilized mode-locked solid-state laser system forprecision range-Doppler imaging

We report measurements which show that an actively stabilized cw mode-locked Nd:YLF laser, in combination with a flashlamp-pumped Nd:glass amplifier, can achieve better than 1 cm resolution of distant rotating targets using range-Doppler imaging. To do this, we have produced trains of 50 ps mode-locked pulses with less than 25 kHz peak-to-peak optical frequency broadening and jitter of the laser modes. This frequency stability is achieved by active control of the oscillator cavity length using an external cavity as a reference. Cavity length stabilization can also reduce mode-locked laser timing jitter if the jitter is caused by cavity optical path length changes common to all laser modes. In our laser, however, the active optical-frequency-stabilization did not significantly improve laser pulse timing stability from the approximate 2-ps jitter levels achieved in our passively stabilized cavity. Analysis of the data indicates that a significant fraction of the timing jitter was due to laser cavity path length changes that varied from mode to mode     

Morbillivirus infections in wildlife (in relation to their population biology and disease control in domestic animals)

The three members of the morbillivirus genus that infect wildlife in ecosystems where domestic animals occur are rinderpest, peste des petits ruminants (PPR) and canine distemper. Data on the relative susceptibility of species of the Order Artiodactyla for rinderpest have been obtained from historical records of outbreaks. Rinderpest in wildlife has only occurred in equatorial and eastern Africa since the great pandemic of 1889-1897. The distributions, densities and population dynamics of susceptible species in this region are described. There has only been one recorded outbreak of PPR in wildlife but the possibility of its occurrence in the future now that it is present in many parts of west and eastern Africa is discussed. Wild carnivora are not likely to be important maintenance hosts for canine distemper but the disease is of significance in free-ranging carnivores and particularly in small populations of endangered susceptible wildlife species. It is also of great significance in zoo populations.     

Selection and characterization of Toxoplasma gondii mutants resistant to artemisinin

Toxoplasma gondii infection, like malaria, is sensitive to inhibition by artemisinin (ART). Mechanisms of action for ART in malaria treatment have been proposed, but little is known about its effects in T. gondii infection. To better understand its inhibitory effects on T. gondii, mutants resistant to ART were selected by progressive culture in permissive levels of the drug. Five clonal isolates were established and characterized. The isolates were approximately 65-fold less sensitive to ART than is the parental RH and showed cross-resistance to the ART derivatives dihydroartemisinin and artemether. In addition to ART resistance, 1 clone (C9) formed morphologically unusual parasitophorous vacuoles and another (A2) was avirulent for mice and protected mice from challenge with the wild type. These clonal T. gondii mutant isolates will be useful for the study of not only the mechanism of action of ART but also parasite vacuole biology and virulence factors.