[Endoscopic management of broncho-pleural fistula in a patient with acute respiratory distress syndrome after pneumonectomy]. / Prise en charge endoscopique d'une fistule bronchopleurale chez un patient en syndrome de détresse respiratoire aiguë après pneumonectomie.

We report the management of endobronchial a patient admitted to the ICU for respiratory distress in the consequences of an surgical recovery of his left pneumonectomy complicated by bronchopleural fistula as part of a bronchial carcinoma non-small cell type adenocarcinoma. Endobronchial treatment by gluing of the fistula may be an alternative to surgery. We discuss its indication in the treatment of bronchial fistula.

Fine mapping of the FecL locus influencing prolificacy in Lacaune sheep.

In the Lacaune sheep population, two major loci influencing ovulation rate are segregating: FecX and FecL. The FecX(L) mutation is a non-conservative substitution (p.Cys53Tyr) in BMP15 that prevents the processing of the protein. Using a statistical approach, FecL has been shown to be an autosomal major gene. A full genome scan localized the FecL locus on sheep chromosome 11. Fine mapping reduced the interval containing FecL to markers BM17132 and FAM117A, corresponding to a synteny block of 1.1 megabases on human chromosome 17, which encompasses 20 genes. The expression of 16 genes from this interval was observed in tissues of the reproductive axis, but expression was not affected in homozygous FecL(L) females. In this interval, a unique haplotype was associated with the FecL(L) mutation. This particular haplotype could be predicted by the DLX3:c.*803A>G SNP in the 3' UTR sequence of the DLX3 gene. This SNP provided accurate classification of animals (99.5%) as carriers or non-carriers of the mutation and therefore maybe useful in marker assisted selection. A synergistic action of FecL(L) and FecX(L) mutations on both ovulation rate and litter size was demonstrated. Until now, all the Fec genes identified in sheep belong to the bone morphogenetic protein (BMP) system. Based on the human orthologous region, none of the 20 genes in the FecL region corresponds to known molecules in the BMP system. The identification of the FecL(L) mutation could lead to the discovery of a new pathway involved in the regulation of ovulation rate.

A deletion in the bone morphogenetic protein 15 gene causes sterility and increased prolificacy in Rasa Aragonesa sheep.

Bone morphogenetic protein 15 (BMP15) is a member of the transforming growth factor beta superfamily, is specifically expressed in oocytes and is essential for sheep prolificacy. Reported mutations in this gene cause increased ovulation rate and infertility in a dosage-sensitive manner. In this work, a new naturally occurring mutation in the BMP15 gene from the ovine Rasa Aragonesa breed is described. This mutation is a deletion of 17 bp that leads to an altered amino acid sequence and introduces a premature stop codon in the protein. Highly significant associations (P < 0.0001) were found between the estimated breeding value for prolificacy and the genotype of BMP15 in Rasa Aragonesa animals with high and low breeding values for this trait. As for other mutations in BMP15, this new mutation is associated with increased prolificacy and sterility in heterozygous and homozygous ewes respectively.

Prolificacy genes in sheep: the French genetic programmes.

It has been demonstrated that variations in litter size or ovulation rate in different breeds of sheep can be associated with the segregation of several major genes. This set of natural mutants constitutes a valuable resource to determine key points in the biochemical pathways controlling the development of ovarian follicles. The French genetic programmes were devised to identify two of these genes: the Booroola (FecB) and Lacaune genes. The FecB prolific mutation corresponds to a non-conservative mutation (Q249R) in the intracellular kinase-signalling domain of the bone morphogenetic protein receptor type IB (BMPR-IB) gene. The Lacaune gene is situated on ovine chromosome 11. Positional cloning is currently in progress to identify the relevant gene and mutation. A similar approach, limited to linkage testing of candidate genes, is proposed to classify the different prolificacy genes in sheep.

Long-term results of intrathoracic chemohyperthermia (ITCH) for the treatment of pleural malignancies.

There is no standard treatment for patients with pleural malignancies. The aim of this prospective study was to investigate the toxicity and long-term results of a multimodality treatment consisting of surgery and intrathoracic chemohyperthermia (ITCH) for the treatment of patients with pleural malignancies. From January 1990 to August 2000, 24 patients with mesothelioma (n=17), fibrosarcoma (n=3), pleural adenocarcinoma (n=3) and thymoma (n=1) were included. The mesothelioma stages were T1 or T2 in 10 cases, and T3 or T4 in seven cases. After cytoreductive surgery, ITCH was carried out for over 60 min, at inflow temperatures less than 45 degrees C, either with mitomycin C (n=7) or cisplatin (n=5) or both (n=12). One patient died from major thoracic air leaks after major decortication and pleurectomy. Seven patients had complications, one pleural clotting necessitating reoperation. After a median follow-up of 89 months, the overall 1-year and 5-year survival rates were 74 and 27%, respectively. For T1 and T2 mesothelioma patients, the median survival was 41.3 months, and for T3 and T4 tumours, it was 4.5 months (P=0.001). The fibrosarcoma patients are alive with no evidence of recurrence at 24, 43 and 54 months. In the conclusion, the combination of surgery with ITCH with mitomycin and/or cisplatin is relatively safe. This procedure may offer unexpected long-term survival in a selected group of patients (T1 and T2 mesothelioma patients and fibrosarcoma patients).

The Booroola mutation in sheep is associated with an alteration of the bone morphogenetic protein receptor-IB functionality.

The hyperprolificacy phenotype of Booroola ewes is due to the presence of the FecB(B) allele at the FecB locus, recently identified as a single amino acid substitution (Q249R) in the bone morphogenetic protein (BMP) type-IB receptor (BMPR1B), and is associated with a more precocious differentiation of ovarian granulosa cells (GCs). To evaluate the consequences of the Booroola mutation on BMPR1B functions, the action of ligands of the transforming growth factor-beta (TGFbeta)/BMP family that act through (growth and differentiation factor-5, BMP-4) or independently of (activin A, TGFbeta-1) BMPR1B were studied on primary cultures of GCs from homozygous FecB(+) and FecB(B) ewes. All the tested TGFbeta/BMP family ligands inhibited progesterone secretion by FecB(+) GCs. Those inhibitory effects were lower for GCs from preovulatory (5-7 mm diameter) than from small antral follicles (1-3 mm diameter). The presence of the Booroola mutation was associated with a 3- to 4-fold (P<0.001) decreased responsiveness of GCs from FecB(B) compared with FecB(+) small follicles to the action of BMPR1B ligands. In contrast, TGFbeta-1 and activin A had similar inhibitory effects on progesterone secretion by GCs from FecB(+) and FecB(B) small follicles. No difference between genotypes was observed with GCs from preovulatory follicles. In transfection experiments with HEK-293 cells, co-expression of FecB(+) BMPR1B and BMPR2 resulted in a 2.6-fold (P<0.01) induction of the activity of a BMP-specific luciferase reporter construct by BMP-4. Interestingly, no response to BMP-4 was observed when cells were transfected with the FecB(B) form of the BMPR1B receptor. Overall, these data strongly suggest that the Q249R mutation is associated with a specific alteration of BMPR1B signaling in hyperprolific Booroola ewes.

Different levels of human intervention in domestic rabbits: effects on genetic diversity.

The effects of human interaction on domestic rabbits were evaluated through the analysis of animals (up to 267) belonging to fancy breeds (22), a commercial breed (1), and selected strains (2). Microsatellite loci and mtDNA polymorphism revealed that the genetic pool of domestic rabbits studied only originated from that available in France. The good conservation of the original diversity was probably ensured through the multiplicity of samplings from wild populations. Selected strains, because of the breeding strategy, keep a fairly high level of diversity compared to other breeds.

A novel method to isolate the common fraction of two DNA samples: hybrid specific amplification (HSA).

Hybrid specific amplification (HSA) is a novel simple method elaborated in order to isolate the common fraction of two DNA samples while avoiding the background due to repeated sequences. The method is based on the suppressive PCR principle, associated with a Cot1 pre-hybridization step. In recent work we demonstrated that hyperprolificity observed in Booroola ewes is associated with a mutation in the bone morphogenetic protein receptor IB gene (BMPR-IB). We applied HSA between ovarian cDNA and DNA from four BAC clones containing BMPR-IB in order to test for the presence of other genes expressed in ovary and to isolate additional BMPR-IB exon sequences. Of the 460 clones obtained, none contained repeated sequences. We successfully obtained 37 clones representing the major part of BMPR-IB coding sequence, together with 5'- and 3'-UTR sequences. Here we have successfully applied HSA to a particular tissue, but it should be possible to trap the common fraction of two DNA samples, whatever their nature.

Mutation in bone morphogenetic protein receptor-IB is associated with increased ovulation rate in Booroola Mérino ewes.

Ewes from the Booroola strain of Australian Mérino sheep are characterized by high ovulation rate and litter size. This phenotype is due to the action of the FecB(B) allele of a major gene named FecB, as determined by statistical analysis of phenotypic data. By genetic analysis of 31 informative half-sib families from heterozygous sires, we showed that the FecB locus is situated in the region of ovine chromosome 6 corresponding to the human chromosome 4q22-23 that contains the bone morphogenetic protein receptor IB (BMPR-IB) gene encoding a member of the transforming growth factor-beta (TGF-beta) receptor family. A nonconservative substitution (Q249R) in the BMPR-IB coding sequence was found to be associated fully with the hyperprolificacy phenotype of Booroola ewes. In vitro, ovarian granulosa cells from FecB(B)/FecB(B) ewes were less responsive than granulosa cells from FecB(+)/FecB(+) ewes to the inhibitory effect on steroidogenesis of GDF-5 and BMP-4, natural ligands of BMPR-IB. It is suggested that in FecB(B)/FecB(B) ewes, BMPR-IB would be inactivated partially, leading to an advanced differentiation of granulosa cells and an advanced maturation of ovulatory follicles.

Characterization of FSH-regulated genes isolated by mRNA differential display from pig ovarian granulosa cells.

The present authors have isolated FSH-regulated genes from primary granulosa cell cultures with or without Follicle Stimulating Hormone (FSH) treatment using mRNA differential display. mRNA differential display consists of amplification of partial sequences of cDNAs (150-400 bp) corresponding to 3' ends of cellular messenger RNAs, and thus, generates 3' expressed sequence tags (3' ESTs). Five thousand cDNA bands were examined, among which the present authors have isolated and sequenced 16 different FSH-regulated products. These sequences were compared with those available in databases. Three of the sequences showed similarity to identified genes from other species (bovine NADH dehydrogenase subunit 4, Xenopus chromosome sequence-associated polypeptide E and transformation-sensitive protein IEF SSP) and four others with human ESTs. Regulation of the corresponding genes has been checked by RT-PCR since most of these are expressed at a low level. FSH-regulation was confirmed for 12 mRNAs (four down- and eight up-regulated). The present authors have also mapped 12 of these ESTs on porcine chromosomes regions using a somatic cell hybrid panel.

Regional localisations of VIM, HSD3b, ACTA1 and PGM1 in pigs.

The comparative map between human and pig has progressed rapidly over the past 2 years. Nevertheless, some points still need to be clarified, particularly the correspondences between human chromosome 10 (HSA10) and porcine chromosome 10 (SSC10) and between human chromosome 1 (HSA1) and porcine chromosomes. The gene codings for vimentin (VIM) carried by HSA10 and three genes carried by HSA1 (hydroxy delta 5 steroid dehydrogenase 3 beta: HSD3B: alpha actin 1: ACTA1: and phosphoglucomutase 1: PGM1) were selected and the regional localisations on pig chromosomes were determined using a well-characterised somatic cell hybrid panel.

A first catalog of genes involved in pig ovarian follicular differentiation.

As a first step toward the characterization of genetic expression in pig ovaries, we have selected 238 clones by differential hybridization from a pig granulosa cell cDNA library, using probes prepared from RNA extracted from either untreated or FSH-treated cells and, in order to generate expressed sequence tags (ESTs), we have performed 3' and 5' single-pass sequencing of these clones. Sequences of the 3' end of the 167 clones that produced informative sequence data were first compared with each other, revealing a redundancy level of 21%. Sequences from the 136 unique clones were analyzed for similarities with sequence data included in Genbank and EMBL databases. Among these unique clones, 54 (40%) matched significantly with sequences from either Genbank of EMBL: 4 with known genes in pig, 35 matched with previously reported human genes, and 15 with other mammalian genes. Eighty-two clones (60%) showed no significant match with any gene or DNA sequence in the Genbank and EMBL databases and thus may represent new pig transcripts.

[Long-term outcome of 72 patients surgically treated for stage II non-small cell bronchial cancer, between 1982 and 1989]. / Devenir à long terme de 72 patients opérés d'un cancer bronchique non à petites cellules, de stade II, entre 1982 et 1989.

We report a 72 patients trial, who had surgical treatments for non small cell lung cancer, stage II (T1N1, T2N1). In our retrospective study, the overall 5-years survival is 44%, with a 24-month median survival. 45% of the patients have recurrence mainly due to distant metastasis. State II appears to be an heterogeneous group. As shown in other studies, the presence of hilar nodes (N1H) seems to be linked with a pejorative outcome. In our series, the survival associated with Lobar N1 (N1L) disease is the same as the survival of Hilar N1 disease, but the initial sites of recurrence differ. The interest of a postchirurgical treatment is controversial. The postoperative radiotherapy reduces the local recurrence without increasing the survival. The chemotherapy treatment is debatable and several studies are under way. We reviewed the different causes of death. The appearance of second cancer in the cured patients is very frequent.

Assignment of markers by using polymerase chain reaction on pools of swine flow-sorted chromosomes.

Gene chromosomal assignment can be realized not only by somatic hybrid panels but also by spot-blot hybridization or polymerase chain reaction (PCR) of flow-sorted chromosomes. We propose a swine chromosome assignment strategy by PCR amplification on pooled chromosomal DNA, which allows assignment despite possible chromosomal contamination during sorting. Each pool contains three different chromosomes, each chromosome being present in one or two pools. We present concordant results obtained for eight markers already mapped to different swine chromosomes and we assign the somatostatin gene to chromosome 13, a new marker in the pig genome.

Denaturing gradient gel electrophoresis detection of polymorphism in a PCR fragment of sheep epidermal growth factor gene.

The ovine map is not yet well-developed, which represents a problem when looking for markers of a region of interest in sheep. A means of circumventing this is to use comparative mapping. In this study primers were determined using consensus sequences for the epidermal growth factor gene of humans, rats and mice, and an ovine epidermal growth factor gene fragment was amplified by polymerase chain reaction (PCR). A new set of specific ovine primers was chosen to study the polymorphism of this DNA fragment by denaturing gradient gel electrophoresis. Eighty-four individuals belonging to seven sheep breeds were studied with this technique and four alleles were detected. The heterozygosity rate was 0.57. Family analysis showed mendelian inheritance of the alleles. Usually, genetic analysis of type-I loci used in the comparative mapping is based on the detection of restriction fragment length polymorphisms in sheep DNA using cDNA probes from other species. Our work shows that another method, based on PCR and denaturing gradient gel electrophoresis techniques, can be efficiently used.

Synteny conservation between parts of human chromosome 4q and bovine and ovine chromosomes 6.

Booroola Merino sheep are characterized by a high ovulation rate attributable to the presence of the FecB allele of the FEC gene. This gene, which has been assigned to sheep chromosome 6, is linked to the gene for EGF, which in man is located on the long arm of chromosome 4 (HSA 4q). To increase the number of known markers on sheep chromosome 6, we used comparative mapping data from sheep, man, and cattle. In our study, we show a synteny between EGF and the genes PDHA2 and FGF5 (from HSA 4q) and microsatellites ILSTS018, ILSTS090, and ILSTS093 (from bovine chromosome 6) in sheep. We also show that the conservation between HSA 4q and sheep chromosome 6 is disrupted between EGF and FGF2.

Sequence analysis and genetic mapping of porcine chromosome 11 centromeric S0048 marker.

We report the existence of a new family of swine centromeric satellite DNA composed of a 51-bp repeat unit, most specifically found on pig chromosome 11 centromere and with less specificity at the centromeric region of other meta- and submetacentric chromosomes. This satellite DNA family, which has no homologies with the Mc1 and Ac2 families published previously, was named Mc2. We designed a specific primer set for PCR amplification of this centromeric satellite DNA. Specificity of amplification was checked by using a porcine somatic cell hybrid panel and by FISH. Furthermore, the development of a PCR-RFLP marker of Mc2 repetition allowed its genetic mapping on the PiGMaP reference families panel. The centromere of chromosome 11 was thus integrated to the genetic map previously published.

[Spontaneous hydro-pneumothorax disclosing malignant mesothelioma. Apropos of 2 cases]. / Hydro-pneumothorax spontané révélant un mésothéliome malin. A propos de deux cas.

We report the cases of two males who presented with spontaneous complete unilateral pneumothorax with ipisilateral liquid effusion. Neither had a history of previous respiratory disease. In both cases chest tube drainage resulted in recurrence of pneumothorax with chronic illness requiring surgical exploration. The surgery revealed a malignant pleural mesothelioma by histological examination. Thus, spontaneous pneumothorax, particularly with abondant effusion can be a revealing symptom of malignant pleural mesothelioma.

Protein kinase C inhibition of in vitro FSH-induced differentiation in pig granulosa cells.

In granulosa cells, growth factor IGF I plays a major role in both growth and differentiation, acting through an autocrine/paracrine mechanism, and its production is regulated by FSH, via cyclic AMP (cAMP). As protein kinase C is also involved in granulosa cell function, we investigated the possibility that its activation could balance the positive effects of FSH. Using pig granulosa cells cultured in vitro, we studied the effects of protein kinase C activation by tetradecanoylphorbol acetate (TPA) on IGF I mRNA level. We also checked morphological modifications, cAMP production and steroidogenesis at the P450 side chain cleavage mRNA and progesterone levels. Our data demonstrate that protein kinase C activation antagonizes the in vitro FSH-induced differentiation, particularly morphological modifications and accumulation of IGF I mRNA. These inhibitory effects on FSH responses suggest that there could be a balance between protein kinase A and protein kinase C pathways in regulating differentiation in pig granulosa cells.

A new device for the treatment of pleural malignancies: intrapleural chemohyperthermia preliminary report.

The prognosis of malignant pleural tumors remains extremely unfavorable. The aim of this study is to evaluate the combination of intrathoracic intrapleural chemotherapy and intrapleural hyperthermia (ITCH) in these diseases. Under anesthesia, 5 men were studied. After pleurectomy for mesothelioma (3/5) or adenocarcinoma (2/5), ITCH is carried out for over 60 min, either with mitomycin C (4/5) or cisplatin (1/5). No pre- or postoperative death occurred. The maximal pleural temperature is 42.6 degrees C. The blood level of mitomycin C never reached the systemic toxic level. All the patients were discharged from the surgical ward, 3 are still alive 15 months later. Therefore, ITCH appears to be a safe and reliable therapy.