RESUMO
BACKGROUND: Spinal anaesthesia, the most common form of anaesthesia for caesarean section, leads to sympathetic blockade and profound maternal hypotension resulting in adverse maternal and neonatal outcomes. Hypotension, nausea and vomiting remain common but until the publication of the National Institute of Health and Care Excellence (NICE) 2021 guidance, no national guideline existed on how best to manage maternal hypotension following spinal anaesthesia for caesarean section. A 2017 international consensus statement recommended prophylactic vasopressor administration to maintain a systolic blood pressure of >90% of an accurate pre-spinal value, and to avoid a drop to <80% of this value. This survey aimed to assess regional adherence to these recommendations, the presence of local guidelines for management of hypotension during caesarean section under spinal anaesthesia, and the individual clinician's treatment thresholds for maternal hypotension and tachycardia. METHODS: The West Midlands Trainee-led Research in Anaesthesia and Intensive Care Network co-ordinated surveys of obstetric anaesthetic departments and consultant obstetric anaesthetists across 11 National Health Service Trusts in the Midlands, England. RESULTS: One-hundred-and-two consultant obstetric anaesthetists returned the survey and 73% of sites had a policy for vasopressor use; 91% used phenylephrine as the first-line drug but a wide range of recommended delivery methods was noted and target blood pressure was only listed in 50% of policies. Significant variation existed in both vasopressor delivery methods and target blood pressures. CONCLUSIONS: Although NICE has since recommended prophylactic phenylephrine infusion and a target blood pressure, the previous international consensus statement was not adhered to routinely.
Assuntos
Anestesia Obstétrica , Raquianestesia , Cesárea , Hipotensão , Vasoconstritores , Humanos , Feminino , Gravidez , Adulto , Hipotensão/etiologia , Raquianestesia/efeitos adversos , Anestesia Obstétrica/efeitos adversos , Reino Unido , Inquéritos e Questionários , Vasoconstritores/administração & dosagem , Vasoconstritores/efeitos adversosRESUMO
The purpose of this study was to determine whether the observed synergistic interaction between cisplatin and valinomycin (VM) in human ovarian carcinoma is the result of mitochondrial DNA (mtDNA) damage. A polymerase chain reaction (PCR)-based method was used to quantitate the lesion frequencies produced by cisplatin, VM and/or drug combination in a 1.1 kbp segment of mtDNA and a 0.536 kbp segment of the nuclear-located beta-globin gene in human ovarian CaOV-3 carcinoma cells. Our data indicates that the nuclear DNA (nDNA) received more cisplatin-induced damage at doses of 25 microM or less than did mtDNA. At higher cisplatin doses (50 microM or more), however, the damage was relatively equal in both segments. VM alone produced little or no damage on mtDNA, yet a significant amount of damage was detected within nDNA. However, when 1 microM VM was used in combination with low doses of cisplatin (0-40 microM), extensive mtDNA damage was detected as compared with the absence of detectable damage on nDNA. In mtDNA, the lesion frequency was 5.45 lesions/10 kb/10 microM cisplatin in the presence of 1 microM VM, whereas no detectable lesions were induced by cisplatin alone. This drug combination produced no detectable damage on DNA, indicating that cisplatin-induced mtDNA damage could be the basis for the observed synergistic interaction with VM. These results also correlate well with our recent in vivo study with the nude mice model of human ovarian cancer treated with a cisplatin/liposomal VM drug combination. Furthermore, this report shows evidence for the role of mitochondria and mtDNA as alternative targets for drug action in cancer therapy.
Assuntos
Cisplatino/farmacologia , Dano ao DNA , DNA Mitocondrial/efeitos dos fármacos , DNA de Neoplasias/efeitos dos fármacos , Neoplasias Ovarianas/metabolismo , Sequência de Bases , Núcleo Celular/metabolismo , Sinergismo Farmacológico , Feminino , Humanos , Dados de Sequência Molecular , Sondas de Oligonucleotídeos , Distribuição de Poisson , Reação em Cadeia da Polimerase , Células Tumorais Cultivadas , Valinomicina/farmacologiaRESUMO
Acute isolates of Coxiella burnetii possess a 36-kbp plasmid termed QpH1. DNA hybridizations show that QpH1 contains approximately 6-kbp region of DNA which is not present in the QpRS plasmid from chronic isolates. This QpH1-specific region of DNA contains the contiguous EcoRI fragments G, E, and D. The GED region was found to possess seven open reading frames (ORF's) coding for proteins ranging from 5.5 to 42.3 kDa in molecular mass when subcloned and expressed in vitro. Summing the predicted ORF's accounts for 95% of the GED coding potential. E. coli expression produced a stable 42.3-kDa protein from the pHIN19 subclone of GED. The ORF of the 42.3-kDa protein, termed cbhE', has been localized on GED by both in vitro transcription/translation and DNA sequencing. The cbhE' gene is estimated as 1142 bp in length with a putative promoter region of TCAACT (-35)-N16-TAAAAT (-10)-N14-AGAAGGA (Shine-Dalgarno)-N10-ATG.
Assuntos
Coxiella burnetii/genética , DNA Bacteriano/genética , Plasmídeos/genética , Clonagem Molecular , DNA Bacteriano/química , Especificidade da EspécieRESUMO
A gene termed cbhE' was cloned from the QpH1 plasmid of Coxiella burnetii. Expression of recombinants containing cbhE' in vitro and in Escherichia coli maxicells, produced an insert-encoded polypeptide of approx. 42 kDa. The CbhE protein was not cleaved when intact maxicells were treated with trypsin. Hybridizations of total DNA isolated from the six strains of C. burnetii indicate that this gene is unique to C. burnetii strains associated with acute disease, i.e., Hamilton[I], Vacca[II], and Rasche[III]. The cbhE' gene was not detected in strains associated with chronic disease (Biotzere[IV] and Corazon[V]) or the Dod[VI] strain. The cbhE' open reading frame (ORF) is 1022 bp in length and is preceded by a predicted promoter/Shine-Dalgarno (SD) region of TCAACT(-35)-N16-TAAAAT(-10)-N14-AGAAGGA (SD) located 10 nucleotides (nt) before the presumed AUG start codon. The ORF ends with a single UAA stop codon and has no apparent Rho-factor-independent terminator following it. The cbhE' gene codes for the CbhE protein of 341 amino acid (aa) residues with a deduced Mr of 39,442. CbhE is predominantly hydrophilic with a predicted pI of 4.43. The function of CbhE is unknown. No nt or aa sequences with homology to cbhE' or CbhE, respectively, were found in searches of a number of data bases.
Assuntos
Proteínas de Bactérias/genética , Coxiella/patogenicidade , Plasmídeos/genética , Doença Aguda , Sequência de Aminoácidos , Proteínas de Bactérias/biossíntese , Proteínas de Bactérias/química , Sequência de Bases , Clonagem Molecular , Coxiella/genética , Expressão Gênica/fisiologia , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Fases de Leitura Aberta/genética , Regiões Promotoras Genéticas/genética , Febre Q/microbiologiaRESUMO
A highly polymorphic Xenorhabdus luminescens strain was isolated. The primary form of X. luminescens was luminescent and nonswarming and produced a yellow pigment and antimicrobial substances. The primary form generated a secondary form that had a distinct orange pigmentation, was weakly luminescent, and did not produce antimicrobial substances. Both the primary and secondary forms generated a set of colony variants at frequencies that exceeded normal rates for spontaneous mutation. The variant forms include nonswarming and swarming forms that formed large colonies and a small-colony (SC) form. The primary and secondary forms generated their SC forms at frequencies of between 1 and 14% and 1 and 2%, respectively. The SC forms were distinct from their parental primary and secondary forms in colony and cellular morphology and in protein composition. The cellular morphology and protein patterns of the nonswarming and swarming colony variants were all very similar. The DNA fingerprints of all forms were similar. Each SC-form colony reverted at high frequency to the form from which it was derived. The proportion of parental-type cells in the SC-form colonies varied with age, with young colonies containing as few as 0.0002% parental-type cells. The primary-to-secondary switch was stable, but all the other colony forms were able to switch at high frequencies to the alternative colony phenotypes.
RESUMO
The dominant natural form of ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) is composed of large (L) 55-kDa and small (S) 15-kDa subunits. This enzyme (as the L8S8 form) is widely distributed among oxygenic photosynthetic species and among chemosynthetic bacteria. Another form lacking small subunits is found as an L2 dimer in Rhodospirillum rubrum or an L oligomer of uncertain aggregation state from Rhodopseudomonas spharoides. The present article reviews two basically different approaches in cloning the R. rubrum gene for RuBisCO. One results in high level expression of this gene product fused with a limited aminoterminal stretch of ß-galactosidase and the other results in expression of wild-type enzyme in Escherichia coli. Also reviewed are a number of reports of cloning and assembly of the L8S8 enzyme in using E. coli L and S subunit genes from Anacystis nidulans, Anabaena 7120, Chromatium vinosum and Rps. sphaeroides.In vitro oligonucleotide-directed mutagenesis has been applied to the gene for RuBisCO from R. rubrum. In terms of contributing new information to our understanding of the catalytic mechanism for RuBisCO, the most significant replacement has been of lys 166 by a number of neutral amino acids or by arg or his. Results establish that lys 166 is a catalytically essential residue and illustrate the power of directed mutagenesis in understanding structure-function correlates for RuBisCO.Oligonucleotide-directed mutagenesis has also been applied to the first and second conserved regions of the S subunit gene for RuBisCO from A. nidulans. In the latter region, corresponding amino acid changes of trp 55 and trp 58 to phe, singly or together, had little or no effect upon enzyme activity. In contrast, mutagenesis in the first conserved region leading to the following pairs of substitutions: arg10 arg 11 to gly 10 gly11; thr14 phe 15 ser 16 to ala 14 phe 15 ala 16; ser 16 tyr 17 to ala 16 asp 17; or pro 19 pro 20 to ala 19 ala 20, are all deleterious.Advances are anticpated in the introduction and expression of interesting modifications of S (and L) subunit genes in plants. A new method of introducing and expressing foreign genes in isolated etiochloroplasts is identified.
RESUMO
The genes for the large and small subunits of ribulose-1,5-bisphosphate carboxylase/oxygenase from the cyanobacterium Anacystis nidulans were subcloned into plasmid pUC9. After induction, both genes were expressed in Escherichia coli and the subunits were assembled into an active holoenzyme. The enzyme was purified from E. coli to high specific activity and was found to contain equimolar amounts of large and small subunits. The assembly of the hexadecameric ribulose bisphosphate carboxylase/oxygenase in E. coli should provide the basis for studies on the mechanism of assembly and the role of small subunits in catalysis.