RESUMO
Intermittent infusions of inotropes have been used with increasing frequency for patients with advanced heart failure (HF). The use of intermittent inotrope infusions remains controversial for several reasons. However, the effect, either positive or negative, of intermittent inotropes in refractory unstable HF patients is not well studied. While prior experience with chronic oral inotropes in stable, advanced HF raises substantial concerns, it may not be directly applicable to a critical evaluation of intermittent inotrope infusions in unstable patients. Further study demonstrating the safety and efficacy of this approach in these patients is imperative. The uncertainties surrounding this therapy, mandate the selection of appropriate candidates be made with substantial care. We report three patients with advanced HF despite conventional therapy, in whom intermittent inotropes were considered. These cases illustrate that this therapy can be avoided in many patients by acceleration of conventional medications, patient education, and aggressive follow up intermittent. However, outpatient inotrope infusions may be helpful in highly selected patients with refractory, unstable HF.
RESUMO
The gene encoding the beta subunit of Bacillus subtilis RNA polymerase was isolated from a lambda gt11 expression library using an antibody probe. Gene identity was confirmed by the similarity of its predicted product to the Escherichia coli beta subunit and by mapping an alteration conferring rifampicin resistance within the conserved rif coding region. Including the rif region, four colinear blocks of sequence similarity were shared between the B. subtilis and E. coli beta subunits. In E. coli, these conserved blocks are separated by three regions that either were not conserved or were entirely absent from the B. subtilis protein. The B. subtilis beta gene was part of a cluster with the order rplL (encoding ribosomal protein L7/L12), orf23 (encoding a 22,513-dalton protein that is apparently essential for growth), rpoB (beta), and rpoC (beta'). This organization differs from the corresponding region in E. coli by the inclusion of orf23. Experiments using promoter probe vectors and site-directed mutagenesis located a major rpoB promoter overlapping the 3'-coding region of orf23, 250 nucleotides upstream from the beta initiation codon. Thus, the B. subtilis rpoB region differs from its E. coli counterpart in both genetic and transcriptional organization.
Assuntos
Bacillus subtilis/enzimologia , Bacillus subtilis/genética , RNA Polimerases Dirigidas por DNA/biossíntese , RNA Polimerases Dirigidas por DNA/genética , Genes Bacterianos , Regiões Promotoras Genéticas , Sequência de Aminoácidos , Sequência de Bases , Sequência Conservada , Escherichia coli/enzimologia , Escherichia coli/genética , Biblioteca Gênica , Genótipo , Substâncias Macromoleculares , Dados de Sequência Molecular , Família Multigênica , Mutagênese Sítio-Dirigida , Fases de Leitura Aberta , Homologia de Sequência de Aminoácidos , Especificidade da Espécie , Transcrição GênicaRESUMO
Bacillus subtilis sigma-B is an alternate sigma factor implicated in controlling stationary-phase gene expression. We characterized the genetic organization and regulation of the region containing the sigma-B structural gene (sigB) to learn which metabolic signals and protein factors govern sigma-B function. sigB lay in an operon with four open reading frames (orfs) in the order orfV-orfW-sigB-orfX, and lacZ gene fusions showed that all four frames were translated in vivo. Experiments with primer extension, S1 nuclease mapping, and lacZ transcriptional fusions found that sigB operon transcription initiated early in stationary phase from a site 32 nucleotides upstream of orfV and terminated 34 nucleotides downstream of orfX. Fusion expression was abolished in a strain carrying an in-frame deletion in sigB, suggesting that sigma-B positively regulated its own synthesis, and deletions in the sigB promoter region showed that sequences identical to the sigma-B-dependent ctc promoter were essential for promoter activity. Fusion expression was greatly enhanced in a strain carrying an insertion mutation in orfX, suggesting that the 22-kilodalton (kDa) orfX product was a negative effector of sigma-B expression or activity. Notably, the genetic organization of the sigB operon was strikingly similar to that of the B. subtilis spoIIA operon, which has the gene order spoIIAA-spoIIAB-spoIIAC, with spoIIAC encoding the sporulation-essential sigma-F. The predicted sequence of the 12-kDa orfV product was 32% identical to that of the 13-kDa SpoIIAA protein, and the 18-kDa orfW product was 27% identical to the 16-kDa SpoIIAB protein. On the basis of this clear evolutionary conservation, we speculate these protein pairs regulate their respective sigma factors by a similar molecular mechanism and that the spoIIA and sigB operons might control divergent branches of stationary-phase gene expression.
Assuntos
Bacillus subtilis/genética , RNA Polimerases Dirigidas por DNA/genética , Genes Bacterianos , Óperon , Fator sigma/genética , Sequência de Aminoácidos , Bacillus subtilis/enzimologia , Bacillus subtilis/crescimento & desenvolvimento , Sequência de Bases , Deleção Cromossômica , Elementos de DNA Transponíveis , RNA Polimerases Dirigidas por DNA/metabolismo , Dados de Sequência Molecular , Mutação , Mapeamento por Restrição , Homologia de Sequência do Ácido Nucleico , Transcrição GênicaRESUMO
We began an analysis of rpoF, the gene encoding the cryptic, 37,000-dalton minor sigma factor (sigma-37) of Bacillus subtilis RNA polymerase. Using antibody raised against sigma-37 holoenzyme to probe a lambda gt11 expression vector library, we isolated a 901-base-pair EcoRI fragment that expressed the COOH-terminal half of sigma-37 fused to lacZ. We used this fragment as a hybridization probe to isolate the entire rpoF gene and additional flanking sequences. Identity of the cloned gene was confirmed by the size and immunological reaction of its product expressed in Escherichia coli and, after DNA sequencing, by the homology of its predicted product (264 residues; 30,143 daltons) with other sigma factors. The DNA sequence also suggested that rpoF may lie in a gene cluster. Upstream of rpoF was an open reading frame that would encode a protein of 17,992 daltons; this frame overlapped the rpoF-coding sequence by 41 base pairs. Immediately following rpoF was a reading frame that would encode a protein of at least 20,000 daltons; expression of this region may be translationally coupled to that of rpoF. By plasmid integration and PBS1 transduction, we found the chromosomal locus of rpoF linked to ddl and dal at 40 degrees on the B. subtilis map and near no known lesions affecting growth regulation or development. Further, an rpoF null mutation resulting from gene disruption had no effect on cell growth or sporulation in rich medium, suggesting that sigma-37 may partly control a regulon not directly involved in the sporulation process.
Assuntos
Bacillus subtilis/genética , Cromossomos Bacterianos/análise , RNA Polimerases Dirigidas por DNA/genética , Genes Bacterianos , Genes , Fator sigma/genética , Fatores de Transcrição/genética , Sequência de Aminoácidos , Bacillus subtilis/enzimologia , Sequência de Bases , Clonagem Molecular , Enzimas de Restrição do DNA , Mutação , PlasmídeosRESUMO
Amniotic fluid was obtained from 85 women during the last trimester of gastation and analyzed for cortisol by a radioimmunoassay procedure and for lecithin/sphingomyelin (L/S) ratios by a combined thin-layer chromatography densitometer scanning technique. A total of 114 samples were examined. Cortisol values ranged from 38 to 438 ng. per milliliter; L/S ratios ranged from 0.3 to 9.2. Comparison of cortisol levels with L/S ratios by multiple regression analysis gave an "r" value of 0.371. From less than 32 weeks' gestation to 41 or 42 weeks there was an increase in cortisol levels from 139 plus or minus 124 to 290 plus or minus 78 ng. per milliliter whereas the L/S ratios increased from 1.8 plus or minus 2.3 to 3.9 plus or minus 2.0. These data indicate that there is no good correlation between cortisol and L/S ratios in the samples of amniotic fluid analyzed.