RESUMEN
OBJECTIVE: Despite the resurgence in regional flap use, otolaryngology resident regional flap experience has been incompletely studied. We sought to characterize United States (US) otolaryngology resident exposure to, and perceptions of, supraclavicular flaps (SCFs), submental flaps (SMFs), and other regional flaps. METHODS: An online survey was disseminated every two weeks to 106 US otolaryngology residency program directors for distribution to residents within their programs between August and October 2016. 121 surveys were returned of which 106 were sufficiently completed and eligible for data analysis. RESULTS: Among residents with adequate responses, 52 were postgraduate year (PGY) 1-3 (junior) residents and 54 were PGY 4-7 (senior) residents. Senior residents participated in more pectoralis major flaps (mean: 8.1, 95%-CI: 6.5-9.8) compared to SCFs (mean: 1.5, 95%-CI: 1.0-2.0, pâ¯<â¯0.001) and SMFs (mean: 0.7; 95%-CI: 0.4-1.0, pâ¯<â¯0.001). Among senior residents exposed to SCFs, SMFs and pectoralis flaps, more individuals judged pectoralis major flaps as successful or very successful (96.2%, 95%-CI: 91.1-100%), compared to SCFs (64.3%, 95%-CI: 46.5-82.0%; pâ¯<â¯0.001) and SMFs (63.2%, 95%-CI: 41.5-84.8%; pâ¯=â¯0.001). CONCLUSIONS: Senior otolaryngology residents were exposed to fewer SCFs and SMFs compared to pectoralis major flaps. Resident perception that SCFs and SMFs were not as successful as pectoralis major flaps should be investigated further.
Asunto(s)
Competencia Clínica , Educación de Postgrado en Medicina/métodos , Colgajos Tisulares Libres/trasplante , Otolaringología/educación , Procedimientos de Cirugía Plástica/educación , Intervalos de Confianza , Femenino , Colgajos Tisulares Libres/clasificación , Rechazo de Injerto , Supervivencia de Injerto , Neoplasias de Cabeza y Cuello/cirugía , Humanos , Internado y Residencia/métodos , Masculino , Colgajo Miocutáneo/trasplante , Evaluación de Necesidades , Encuestas y Cuestionarios , Estados UnidosRESUMEN
1. Benzodiazepines (BZ) and barbiturates both potentiate chloride currents through GABA(A) receptors to enhance inhibition. However, unlike barbiturates BZ do not impair autonomic control of heart rate. We hypothesised that BZ might not significantly potentiate GABAergic transmission in the caudal nucleus of the solitary tract (cNTS), which is critically important for mediating the baroreceptor reflex. 2. In rat brain slices the BZ agonists chlordiazepoxide and midazolam (2 and 50 microM) did not significantly enhance currents evoked by GABA in voltage-clamped cNTS neurones. Chlordiazepoxide (50 microM) reversibly increased electrically evoked IPSPs in 5/10 rostral NTS (rNTS) neurones but only in 2/10 cNTS neurones. Pentobarbitone (50-100 microM) was effective in enhancing GABA(A)-mediated responses in all NTS neurones. An inverse BZ agonist, methyl 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM; 1 or 10 microM), failed to depress GABA-induced currents in the cNTS. 3. Microinjections of midazolam (10 and 100 microM solutions) into the cNTS did not affect the baroreceptor reflex (P > 0.2) while pentobarbitone (100 microM) significantly and reversibly depressed it (gain decrease to 53 +/- 11 % of control, P < 0.01). 4. Reverse transcriptase polymerase chain reaction revealed the presence of alpha(1), alpha(2), beta(2), beta(3) and gamma(2) GABA(A) receptor subunit mRNA in the cNTS. No alternatively spliced variants of the alpha(1)- and gamma(2)-subunits were revealed. Moreover, GABA(A) epsilon-unit mRNA was found in both the cNTS and rNTS as two alternatively spliced transcripts. 5. Immunocytochemical analysis revealed numerous GABA(A) epsilon-subunit-positive neurones within the cNTS with significantly fewer epsilon-subunit-positive cells in the rNTS. 6. As incorporation of the epsilon-subunit in recombinant GABA(A) receptors may confer BZ insensitivity we propose that the paucity of BZ actions in the cNTS is due to a high level of epsilon-subunit expression. This is the first demonstration of a possible physiological impact of the epsilon-subunit on native GABA(A) receptors.
Asunto(s)
Benzodiazepinas/farmacología , Receptores de GABA-A/efectos de los fármacos , Núcleo Solitario/efectos de los fármacos , Animales , Ansiolíticos/farmacología , Barorreflejo/efectos de los fármacos , Carbolinas/farmacología , Clordiazepóxido/farmacología , Electrofisiología , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Femenino , Moduladores del GABA/farmacología , Inmunohistoquímica , Técnicas In Vitro , Iontoforesis , Masculino , Midazolam/farmacología , Pentobarbital/farmacología , ARN Mensajero/biosíntesis , Ratas , Ratas Wistar , Receptores de GABA-A/biosíntesis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ácido gamma-Aminobutírico/farmacologíaRESUMEN
We have characterized the DSox14 gene, a new member of the family of transcription factors related to the mammalian sex determining factor, SRY. It contains two exons and the intron is large for Drosophila at 2.8 kb. The encoded protein consists of 691 amino acids (72 kDa) and includes an HMG box domain, which is closely related to the mouse Sox4 DNA binding domain. Expression of the DSox14 HMG box domain in vitro shows that it binds the sequence AACAAT with a K(d) of 190 nM, generating a bend angle of 48.6 degrees. At higher protein concentrations, a second HMG box binds at the recognition sequence, increasing the bend angle by 5 degrees. DSox14 is variably expressed throughout development as three alternative transcripts but not at all during the 1st and 2nd larval instars. The several mRNA transcripts are produced primarily from different transcriptional start sites. Analysis of the expression of DSox14 mRNAs during early development shows that they are maternally contributed at a low level and ubiquitously expressed during embryogenesis. The widespread pattern of expression suggests that DSox14 affects a large number of target genes.
Asunto(s)
Proteínas de Drosophila , Drosophila melanogaster/genética , Proteínas del Grupo de Alta Movilidad/genética , Secuencia de Aminoácidos , Animales , Sitios de Unión , Northern Blotting , Clonación Molecular , ADN/química , ADN/genética , ADN Complementario/química , ADN Complementario/genética , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Embrión no Mamífero/metabolismo , Femenino , Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Proteínas del Grupo de Alta Movilidad/metabolismo , Hibridación in Situ , Masculino , Datos de Secuencia Molecular , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factores de Transcripción SOXB2 , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de AminoácidoRESUMEN
About 400 distinct mutations have been defined in the BRCA1 gene, and these are spread fairly evenly through the 5592 bp of coding DNA. This circumstance presents a formidable challenge for mutation screening. Apart from total direct sequencing, the preferred screening method has been single-strand conformation polymorphism (SSCP) gels, with a smaller input from constant denaturant gradient electrophoresis (CDGE), heteroduplex (HD) analysis, and mismatch cleavage. The protein truncation test (PTT) was used early in BRCA1 mutation screening but has not been widely adopted, perhaps because a straightforward analysis of the whole BRCA1 gene requires working with RNA and all its perceived problems. The present work was undertaken to assess the practicality of using the PTT under routine conditions for the screening of long genes such as BRCA1 that are not highly expressed in lymphocytes. We conclude that, provided RNA preparation is carried out effectively and consistently, the PTT approach has significant advantages over other methodologies such as SSCP gels.
Asunto(s)
Genes BRCA1 , Mutación , Secuencia de Bases , Estudios de Cohortes , Cartilla de ADN , Exones , Humanos , Polimorfismo Conformacional Retorcido-Simple , Biosíntesis de Proteínas , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The glial cell line-derived neurotrophic factor (GDNF)-family of neurotrophic factors consisted until recently of three members, GDNF, neurturin, and persephin. We describe here the cloning of a new GDNF-family member, neublastin (NBN), identical to artemin (ART), recently published (Baloh et al., 1998). Addition of NBN/ART to cultures of fetal mesencephalic dopamine (DA) neurons increased the number of surviving tyrosine hydroxylase (TH)-immunoreactive neurons by approximately 70%, similar to the maximal effect obtained with GDNF. To investigate the neuroprotective effects in vivo, lentiviral vectors carrying the cDNA for NBN/ART or GDNF were injected into the striatum and ventral midbrain. Three weeks after an intrastriatal 6-hydroxydopamine lesion only about 20% of the nigral DA neurons were left in the control group, while 80-90% of the DA neurons remained in the NBN/ART and GDNF treatment groups, and the striatal TH-immunoreactive innervation was partly spared. We conclude that NBN/ART, similarly to GDNF, is a potent neuroprotective factor for the nigrostriatal DA neurons in vivo.
Asunto(s)
Cuerpo Estriado/citología , Dopamina/fisiología , Factores de Crecimiento Nervioso/fisiología , Proteínas del Tejido Nervioso/fisiología , Neuronas/citología , Neuronas/fisiología , Sustancia Negra/citología , Tirosina 3-Monooxigenasa/análisis , Secuencia de Aminoácidos , Animales , Línea Celular , Células Cultivadas , Clonación Molecular , Técnicas de Transferencia de Gen , Vectores Genéticos , Factor Neurotrófico Derivado de la Línea Celular Glial , Humanos , Lentivirus , Ratones , Datos de Secuencia Molecular , Factores de Crecimiento Nervioso/genética , Proteínas del Tejido Nervioso/química , Proteínas del Tejido Nervioso/genética , Fármacos Neuroprotectores , Ratas , Alineación de Secuencia , Homología de Secuencia de AminoácidoRESUMEN
The high mobility group (HMG) I proteins are small, non-histone chromosomal proteins that promote gene activation during development and within rapidly dividing cells. They do so by facilitating enhanceosome formation on inducible genes, via both protein/DNA and protein/protein interactions. The HMG I-C gene is tightly regulated, normally being expressed exclusively during embryonic development. However, HMG I-C expression is also observed frequently in a number of tumor types, and this expression has been shown to contribute to the malignant transformation process. With the aim of dissecting pathways that lead to aberrant expression of HMG I-C in tumor cells, we have analyzed HMG I-C gene regulation in the human hepatoma cell line PLC/PRF/5. One of the two HMG I-C transcripts detected in this cell line originates from a novel downstream initiation site at nucleotide -161 relative to the first methionine. Transcription from the downstream initiation site is mediated by a PRE located between nt -222 and -217. We show here that the Sp1 and Sp3 transcription factors interact with the PRE and transactivate the HMG I-C promoter in a cooperative fashion. This study provides the first characterization of this downstream HMG I-C promoter.
Asunto(s)
Proteínas del Grupo de Alta Movilidad/genética , Proteínas de Neoplasias/genética , Secuencias Reguladoras de Ácidos Nucleicos , Transcripción Genética , Animales , Secuencia de Bases , Carcinoma Hepatocelular/genética , Línea Celular , Secuencia Conservada , Proteínas de Unión al ADN/metabolismo , Drosophila melanogaster/citología , Evolución Molecular , Proteína HMGA2 , Proteínas del Grupo de Alta Movilidad/metabolismo , Humanos , Datos de Secuencia Molecular , Mutagénesis , Proteínas de Neoplasias/metabolismo , Regiones Promotoras Genéticas , ARN Mensajero , Factor de Transcripción Sp1/metabolismo , Factor de Transcripción Sp3 , Factores de Transcripción/metabolismo , Transfección , Células Tumorales CultivadasRESUMEN
The gene on chromosome 12 coding for the human protein HMGI-C has been cloned and partially sequenced. It consists of five exons, the first and last of which include long untranslated regions. The 5' UTR includes a (CA/T)n tract and a polymorphic (CT)n tract. Exons II, III and IV (87, 51 and 33 bp) are dispersed over > 30 kb. Exons I-III separately encode the three basic DNA binding domains ('A-T hooks'), exon IV encodes an 11 amino acid sequence characteristic of HMGI-C and absent from the human HMGI(Y) gene [Friedmann, M., Holth, L. T., Zoghbi, H. Y. and Reeves, R. (1993) Nucleic Acids Res., 21, 4259-4267], whilst exon V encodes the acidic C-terminal domain, which is subject to multiple phosphorylation. The HMGI-C gene is thus a striking example of the separation of functional protein elements into different coding exons.
Asunto(s)
Cromosomas Humanos Par 12 , Exones , Proteínas del Grupo de Alta Movilidad/genética , Factores de Transcripción/genética , Secuencia de Aminoácidos , Secuencia de Bases , Biblioteca Genómica , Proteína HMGA2 , Proteínas del Grupo de Alta Movilidad/química , Humanos , Datos de Secuencia Molecular , Secuencias Repetitivas de Ácidos Nucleicos , Selección Genética , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido Nucleico , Relación Estructura-Actividad , Factores de Transcripción/química , Transcripción GenéticaRESUMEN
A cDNA library prepared from mRNA extracted from immature male gonads of the bivalve mollusc Ensis minor (razor shell) was probed with a 133-bp reverse-transcriptase PCR product corresponding to a segment of the sperm protein EM6 [Giancotti, V., Russo, E., Gasparini, M., Serrano, D., Del Piero, D., Thorne, A. W., Cary, P.D. & Crane-Robinson, C. (1993) Eur. J. Biochem. 136, 509-516]. A single 1.5-kb clone was found to encode both sperm proteins EM1 and EM6. Mass spectrometry was used to define the C-terminus of EM1, and since the N-terminus of EM6 is known from Edman degradation, this showed that the pentapeptide NTNNS must be lost on proteolytic processing. Both EM1 and EM6 contain highly repeated amino acid sequences, suggestive of extended structures. EM1 contains seven tandem repeats of the dipeptide S(K/R), followed by six potential cdc2 phosphorylation sites and seven repeats of the octapeptide KRSASKKR, with occasional K/R substitutions. EM6 contains a globular domain preceded by 17 almost identical uninterupted tandem repeats of the motif KKRSXSRKRSAS, where X is charged. Its C-terminus contains 15 short basic clusters. Assignment of EM1 and EM6 to the established categories of molluscan sperm proteins [PLI, PLII, PLIII, PLIV; Ausio, J. (1992) Mol. Cell. Biochem. 115, 163-172] is discussed.
Asunto(s)
Histonas/metabolismo , Moluscos/metabolismo , Protaminas/metabolismo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , ADN Complementario/genética , ADN Complementario/aislamiento & purificación , Histonas/química , Histonas/genética , Masculino , Espectrometría de Masas , Datos de Secuencia Molecular , Protaminas/química , Protaminas/genética , Conformación Proteica , Espermatozoides/metabolismoRESUMEN
The constitutive DNA from members of four families showing predisposition to breast cancer was amplified by PCR in the region of exons 5, 6, 7 and 8 of the p53 proto-oncogene. Single-strand conformation polymorphism (SSCP) gels were used to compare patient DNA with mutant and wild-type control samples. No cases of anomalous mobility were observed in samples from the susceptible families. The lack of inherited mutations was confirmed for exons 5 and 7 by solid-phase DNA sequencing. The results lend further support to the view that inherited mutations in p53 alleles are not a significant contributor to breast cancer predisposition and it is not, therefore, clinically worthwhile to screen predisposed or potentially predisposed families for germline mutations in the p53 gene.
Asunto(s)
Neoplasias de la Mama/genética , Genes p53/genética , Pruebas Genéticas , Mutación de Línea Germinal/genética , Adulto , Anciano , Secuencia de Bases , ADN de Neoplasias/genética , Exones/genética , Femenino , Humanos , Persona de Mediana Edad , Datos de Secuencia Molecular , Linaje , Reacción en Cadena de la Polimerasa , Polimorfismo Conformacional Retorcido-Simple , Proto-Oncogenes Mas , Análisis de Secuencia de ADNRESUMEN
The HMGI family contains three members: I, Y and I-C. HMGI and HMGY are alternative splicings of the same gene and are essential transcription factors at several genetic loci. HMGI-C is transcribed from a different gene and is observed only in highly transformed cells. This work shows that human I-C is present in a more restricted range of cell types than I/Y and is absent from hemopoietic cells, as noted for mouse I-C. However, high expression in a human hepatoma line allowed the cloning of the cDNA and 812 bp of 5'-untranslated, 330 bp of coding and 58 bp of 3'-untranslated DNA were sequenced. The open reading frame showed 4 amino acid substitutions and one additional amino acid when compared to mouse I-C, none of them in the basic DNA binding motifs.
Asunto(s)
Proteínas del Grupo de Alta Movilidad/genética , Secuencia de Aminoácidos , Secuencia de Bases , Línea Celular , Clonación Molecular , Cartilla de ADN/química , ADN Complementario/genética , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/genética , Expresión Génica , Genes , Proteína HMGA2 , Humanos , Datos de Secuencia Molecular , ARN Mensajero/genética , Alineación de Secuencia , Homología de Secuencia de AminoácidoRESUMEN
The synchronous macroplasmodial growth phase of the slime mould Physarum polycephalum was used to study the in vivo replication of large chromosomal DNA segments. Newly replicated DNA was isolated at various points in S-phase by its preferential association with the nuclear matrix. This DNA was then used to probe cosmid clones of the Physarum genome. The results indicate that certain dispersed repetitive sequences in the genome are coordinately replicated. The observed pattern of replication may be due either to the presence of a replication origin within each repetitive sequence or to the systematic arrangement of these sequences around a replication origin. The latter appears more likely since the repetitive sequences are probably not randomly scattered within the genome.
Asunto(s)
Replicación del ADN , ADN de Hongos/biosíntesis , Genes Fúngicos , Physarum/genética , Secuencias Repetitivas de Ácidos Nucleicos , Núcleo Celular/análisis , Cósmidos , ADN de Hongos/análisis , ADN de Hongos/genética , Desoxirribonucleasas de Localización Especificada Tipo II , Interfase , Hibridación de Ácido Nucleico , Physarum/crecimiento & desarrollo , Physarum/metabolismoRESUMEN
Plasma nicotine three minutes after smoking a cigarette was measured in 10 sedentary workers in mid-morning and five hours later on four typical working days. The average mid-morning level after they had been smoking their usual cigarettes (mean nicotine yield 1-34 ng) was 150-4 nmol/l (24-4 ng/ml) (range 95-6-236-7 nmol/l (15-5-38-4 ng/ml)). Despite great variation between smokersthe mid-morning levels of each smoker were fairly consistent over the four mornings and correlated 0-82 with their carboxyhaemoglobin levels. After continuing to smoke their usual brand or switching to a high-nicotine brand (3-2 mg) average afternoon levels of 185-6 and 180-0 nmol/6 (30-1 and 29-2 ng/ml) respectively were not significantly higher than the morning levels, but after switching to low-nicotine cigarettes (0-14 mg) the plasma nicotine dropped to an average of 52-4 nmol/l (8-5 ng/ml). The changes between morning and afternoon while smoking usual or high-nicotine cigarettes showed marked individual variation. The findings suggest that the plasma nicotine level just after a cigarette depends more on the way the cigarette is smoked than on its nicotine yield or the number which have been smoked over the preceding few hours.
Asunto(s)
Nicotina/sangre , Fumar , Adulto , Conducta , Carboxihemoglobina/análisis , Ritmo Circadiano , Femenino , Humanos , Recién Nacido , Masculino , Persona de Mediana Edad , Nicotina/análisis , Estadística como Asunto , Factores de TiempoRESUMEN
In 10 sedentary workers, smoking as they felt inclined over a five-hour period in the middle of a typical working day, changing to low nicotine cigarettes (<0.3 mg) caused an increase in the number and weight of cigarettes smoked, while changing to high nicotine cigarettes (3.2 mg) caused a decrease (P < 0.01). The average number and weight smoked in five hours for usual, low, and high nicotine brands were respectively 10.6 (6.00 g), 12.5 (6.52 g), and 6.7 (4.19 g). When smoking the usual brand the average blood carboxyhaemoglobin (COHb) increased 1.78% (from 6.38% to 8.16%). But on changing to either high or low nicotine cigarettes the COHb levels instead of increasing, tended to fall (P < 0.01). The average fall of 0.34% while smoking low nicotine cigarettes was due to the low carbon monoxide (CO) yield of these cigarettes, while the fall of 1.04% when smoking high nicotine cigarettes was attributable to reduced consumption. The findings support the view that smoking behaviour is modified to regulate nicotine intake. Besides having low tar and CO yields, the least harmful cigarettes for heavy smokers may be those with a high, rather than low, nicotine yield.