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BACKGROUND: Treatment of cystic fibrosis (CF) has been revolutionized by the use of cystic fibrosis transmembrane conductance regulator (CFTR) protein modulators such as elexacaftor/tezacaftor/ivacaftor (ETI) triple therapy. Prior studies support a role for type 2 (T2) inflammation in many people with CF (PwCF) and CF-asthma overlap syndrome (CFAOS) is considered a separate clinical entity. It is unknown whether initiation of ETI therapy impacts T2 inflammation in PwCF. We hypothesized that ETI initiation decreases T2 inflammation in PwCF. METHODS: A single center retrospective chart review was conducted for adult PwCF. As markers of T2 inflammation, absolute eosinophil count (AEC) and total immunoglobulin E (IgE) data were collected longitudinally 12 months prior to ETI therapy initiation and 12 months following therapy initiation. Multivariable analyses adjusted for the age, gender, CFTR mutation, disease severity, inhaled steroid use, and microbiological colonization. RESULTS: There was a statistically significant reduction (20.10%, p < 0.001) in 12-month mean total IgE following ETI initiation; this change remained statistically significant in the multivariate model. The longitudinal analysis demonstrated no change in AEC following therapy initiation. CONCLUSION: This study demonstrates that there is a statistically significant percent reduction in mean total IgE but no change in AEC following ETI initiation. ETI may lead to decreased antigen and superantigen load in the airway as a result of improved mucociliary clearance and these changes may drive the decline in total IgE, without influencing the epigenetic drivers of eosinophilic inflammation. Further studies are warranted to determine the underlying mechanism of ETI impact on T2 inflammation and possible role for asthma immunomodulator therapy post ETI initiation in CFAOS.
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BACKGROUND AND PURPOSE: Racial and socioeconomic disparities in the incidence, treatment, and outcomes of acute ischemic stroke exist and have been described. We aimed to characterize disparities in the use of endovascular thrombectomy in a nationally representative analysis. MATERIALS AND METHODS: Discharge data from the Nationwide Inpatient Sample between 2006 and 2016 were queried using validated International Classification of Disease codes. Patients admitted to US hospitals with acute ischemic stroke were included and stratified on the basis of race, income, and primary payer. Trends in endovascular thrombectomy use, good outcome (discharge to home/acute rehabilitation), and poor outcome (discharge to skilled nursing facility, hospice, in-hospital mortality) were studied using univariate and multivariable analyses. RESULTS: In this analysis of 1,322,162 patients, endovascular thrombectomy use increased from 53/111,829 (0.05%) to 3054/146,650 (2.08%) between 2006 and 2016, respectively. Less increase was observed in black patients from 4/12,733 (0.03%) to 401/23,836 (1.68%) and those in the lowest income quartile from 10/819 (0.03%) to 819/44,984 (1.49%). Greater increase was observed in the highest income quartile from 18/22,138 (0.08%) to 669/27,991 (2.39%). Black race predicted less endovascular thrombectomy use (OR = 0.79; 95% CI, 0.72-0.86). The highest income group predicted endovascular thrombectomy use (OR = 1.24; 95% CI, 1.13-1.36) as did private insurance (OR = 1.30; 95% CI, 1.23-1.38). High income predicted good outcome (OR = 1.10; 95% CI. 1.06-1.14), as did private insurance (OR = 1.36; 95% CI, 1.31-1.39). Black race predicted poor outcome (OR = 1.33; 95% CI, 1.30-1.36). All results were statistically significant (P < .01). CONCLUSIONS: Despite a widespread increase in endovascular thrombectomy use, black and low-income patients may be less likely to receive endovascular thrombectomy. Future effort should attempt to better understand the causes of these disparities and develop strategies to ensure equitable access to potentially life-saving treatment.
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Isquemia Encefálica , Procedimientos Endovasculares , Accidente Cerebrovascular Isquémico , Accidente Cerebrovascular , Isquemia Encefálica/cirugía , Humanos , Factores Socioeconómicos , Accidente Cerebrovascular/cirugía , Trombectomía , Resultado del TratamientoRESUMEN
BACKGROUND: Complete pathological resection of locally advanced and recurrent anorectal cancer is considered the most important determinant of survival outcome. Involvement of the retropubic space with cancer threatening or involving the penile base poses specific challenges due to the potential for margin involvement and blood loss from the dorsal venous plexus. In the present study we evaluate a new transperineal surgical approach to excision of anterior compartment organs involved or threatened by cancer which facilitates exposure and visualisation of the bulbar urethra and the deep vein of the penis caudal to the retropubic space and penile base. METHODS: A retrospective study was performed on male patients with tumour extension into the penile base treated at our institution using the transperineal surgical approach. Descriptive data for patient demographics, radiology, operative details, postoperative histology, complications and outcomes were collated. RESULTS: Ten male patients with tumour extension into the penile base were identified. Two patients had recurrent anal cancer, 6 had locally advanced primary rectal cancer and 2 had recurrent rectal cancer. All patients had exenterative surgery with excision of the penile base utilising the transperineal approach. All patients had R0 resection. No local recurrence developed after a median follow up period of 15 months. CONCLUSIONS: The transperineal approach to the penile base and retropubic space allows for high rates of R0 resection margin status with direct visualisation of the dorsal venous plexus, thereby minimising blood loss. In our experience, this technique is the preferred approach to excision of cancers threatening and involving the penile base and also for most male patients requiring total pelvic exenteration.
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Neoplasias del Ano/cirugía , Recurrencia Local de Neoplasia/cirugía , Exenteración Pélvica/métodos , Pene/cirugía , Neoplasias del Recto/cirugía , Adulto , Anciano , Neoplasias del Ano/patología , Pérdida de Sangre Quirúrgica , Humanos , Masculino , Márgenes de Escisión , Persona de Mediana Edad , Pene/patología , Perineo/cirugía , Neoplasias del Recto/patología , Estudios Retrospectivos , Resultado del TratamientoRESUMEN
AIM: Ovarian metastases from gastrointestinal tract malignancies have been considered an ominous finding with poor prognosis. The aim of this project was to determine the impact on survival, and potential cure, when cytoreductive surgery (CRS) and hyperthermic intraperitoneal chemotherapy (HIPEC) are combined to treat peritoneal malignancy in women with Krukenberg tumours. METHOD: A retrospective analysis of prospectively collected data between January 2010 and July 2015. Female patients undergoing complete CRS (macroscopic tumour removal) and HIPEC for pseudomyxoma peritonei (PMP) of appendiceal origin, or colorectal peritoneal metastases (CPM) were included. Survival was estimated using the Kaplan-Meier method and survival rates compared using the log-rank test. RESULTS: In total, 889 patients underwent surgery for peritoneal malignancy, of whom 551 were female. Of these, 504/551 (91%) underwent complete CRS and HIPEC. Overall, 405/504 (80%) had at least one involved ovary removed either during CRS and HIPEC or at their index prereferral operation. Three hundred and fifty-two patients (87%) had an appendiceal tumour and 53 (13%) had CPM. At a median follow up of 40 months, overall survival (OS) did not differ significantly between patients with or without ovarian involvement in women with a primary low-grade appendiceal tumour or CPM. In women with high-grade primary appendiceal pathology, OS was significantly lower in patients with ovarian metastases compared with those without ovarian involvement. CONCLUSION: Women with ovarian metastases from low-grade appendiceal tumours or colorectal cancer treated with CRS and HIPEC have similar survival rates to patients without ovarian metastases. Long-term survival and cure is feasible in patients amenable to complete tumour removal.
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Adenocarcinoma Mucinoso/secundario , Antineoplásicos/administración & dosificación , Neoplasias del Apéndice/patología , Neoplasias Colorrectales/patología , Neoplasias Ováricas/secundario , Neoplasias Peritoneales/secundario , Neoplasias Peritoneales/terapia , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Procedimientos Quirúrgicos de Citorreducción , Supervivencia sin Enfermedad , Femenino , Humanos , Hipertermia Inducida , Infusiones Parenterales , Estimación de Kaplan-Meier , Persona de Mediana Edad , Estudios Retrospectivos , Tasa de Supervivencia , Adulto JovenRESUMEN
Convection-enhanced delivery (CED) is a promising technique that generates a pressure gradient at the tip of an infusion catheter to deliver therapeutics directly through the interstitial spaces of the central nervous system. It addresses and offers solutions to many limitations of conventional techniques, allowing for delivery past the blood-brain barrier in a targeted and safe manner that can achieve therapeutic drug concentrations. CED is a broadly applicable technique that can be used to deliver a variety of therapeutic compounds for a diversity of diseases, including malignant gliomas, Parkinson's disease, and Alzheimer's disease. While a number of technological advances have been made since its development in the early 1990s, clinical trials with CED have been largely unsuccessful, and have illuminated a number of parameters that still need to be addressed for successful clinical application. This review addresses the physical principles behind CED, limitations in the technique, as well as means to overcome these limitations, clinical trials that have been performed, and future developments.
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Encefalopatías/tratamiento farmacológico , Sistemas de Liberación de Medicamentos/instrumentación , Sistemas de Liberación de Medicamentos/métodos , Barrera Hematoencefálica/metabolismo , Neoplasias Encefálicas/tratamiento farmacológico , Catéteres , Ensayos Clínicos como Asunto , Convección , Glioma/tratamiento farmacológico , Humanos , Enfermedad de Parkinson/tratamiento farmacológicoRESUMEN
PURPOSE: Oxaliplatin is increasingly becoming the chemotherapeutic drug of choice for the treatment of peritoneal malignancies using cytoreductive surgery and hyperthermic intraperitoneal chemotherapy (CRS-HIPEC). Oxaliplatin is unstable in chloride-containing media, resulting in the use of 5% dextrose as the carrier solution in these procedures. Exposure of the peritoneum to 5% dextrose during perfusion times varying from 30 min to 90 min is associated with serious hyperglycemias and electrolyte disturbances. This can result in significant postoperative morbidity and mortality. In order to find out whether safer, chloride-containing carrier solutions can be used, we report the results of in-vitro analysis of oxaliplatin stability in both chloride-containing and choride-deficient carrier solutions and discuss the implications for oxaliplatin-based CRS-HIPEC procedures. METHODS: 5 mg of oxaliplatin was added to 50 mL of various carrier solutions at 42 °C: 5% dextrose, 0.9% sodium chloride, Ringer lactate, Dianeal(®) PD4 glucose 1.36% solution for peritoneal dialysis and 0.14 M sterile phosphate buffer pH 7.4. Samples were collected at standardized intervals and oxaliplatin concentration was determined using a stability indicating high-performance liquid chromatographic method, coupled to an UV detector (HPLC-UV); oxaliplatin degradation products were identified using HPLC-mass spectometry. RESULTS: In 5% dextrose, oxaliplatin concentration remained stable over a 2-hour period. Increasing chloride concentrations were associated with increasing degradation rates; however, this degradation was limited to <10% degradation after 30 min (the standard peritoneal perfusion time in most clinical CRS-HIPEC protocols) and <20% degradation after 120 min at 42 °C. In addition, oxaliplatin degradation was associated with the formation of its active drug form [Pt(dach)Cl2]. CONCLUSIONS: The use of chloride-containing carrier solutions for oxaliplatin does not relevantly affect its concentrations under the tested in-vitro conditions. Chloride seems to promote formation of the active cytotoxic drug form of oxaliplatin and therefore could enhance its cytotoxic effect. These data show that more physiological, chloride-containing carrier solutions can be used safely and effectively as a medium for oxaliplatin in CRS-HIPEC procedures.
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Antineoplásicos/química , Cloruros/química , Compuestos Organoplatinos/química , Antineoplásicos/uso terapéutico , Procedimientos Quirúrgicos de Citorreducción , Estabilidad de Medicamentos , Hipertermia Inducida , Compuestos Organoplatinos/uso terapéutico , Oxaliplatino , Neoplasias Peritoneales/tratamiento farmacológico , Neoplasias Peritoneales/cirugía , Neoplasias Peritoneales/terapia , SolucionesRESUMEN
BACKGROUND: Colorectal cancer peritoneal metastasis (CPM) confers an exceptionally poor prognosis, and traditional treatment involving systemic chemotherapy (SC) is largely ineffective. Cytoreductive surgery (CRS) combined with hyperthermic intraperitoneal chemotherapy (HIPEC) is increasingly advocated for selected patients with CPM; however, opinions are divided because of the perceived lack of evidence, high morbidity, mortality, and associated costs for this approach. As there is no clear consensus, the aim of this study was to compare outcomes following CRS+HIPEC vs SC alone for CPM using meta-analytical methodology, focusing on survival outcomes. Secondary outcomes assessed included morbidity, mortality, quality of life (QOL), and health economics (HE). METHODS: An electronic literature search was conducted to identify studies comparing survival following CRS+HIPEC vs SC for CPM. The odds ratio (OR) was calculated using the Mantel-Haenszel method with corresponding 95% confidence intervals (CI) and P-values. Heterogeneity was examined using the Q-statistic and quantified with I(2). The fixed-effect model (FEM) was used in the absence of significant heterogeneity. For included studies, 2- and 5-year survival was compared for CRS+HIPEC vs SC alone. RESULTS: Four studies (three case-control, one RCT) provided comparative survival data for patients undergoing CRS+HIPEC (n=187) vs SC (n=155) for CPM. Pooled analysis demonstrated superior 2-year (OR 2.78; 95% CI 1.72-4.51; P=0.001) and 5-year (OR 4.07; 95% CI 2.17-7.64; P=0.001) survival with CRS+HIPEC compared with SC. Mortality ranged from 0 to 8%. No data were available for the assessment of QOL or HE. CONCLUSIONS: Although limited by between-study heterogeneity, the data support the assertion that in carefully selected patients, multimodal treatment of CPM with CRS+HIPEC has a highly positive prognostic impact on medium- and long-term survival compared with SC alone. There is a paucity of comparative data available on morbidity, QOL, and HE.
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Antineoplásicos/uso terapéutico , Neoplasias Colorrectales/terapia , Procedimientos Quirúrgicos de Citorreducción , Hipertermia Inducida , Neoplasias Peritoneales/secundario , Neoplasias Colorrectales/patología , Neoplasias Colorrectales/cirugía , Terapia Combinada , Humanos , Neoplasias Peritoneales/terapiaRESUMEN
BACKGROUND: Cytoreductive surgery and hyperthermic intraperitoneal chemotherapy (CRS-HIPEC) is the preferred treatment of peritoneal carcinomatosis (PC) of colorectal carcinoma. Patients with positive lymph node status have worse survival after CRS-HIPEC, which is probably due to higher rates of systemic failure. In this study, we analysed the effect of administration and timing of systemic chemotherapy on the outcome of lymph node positive colorectal carcinoma patients treated with CRS-HIPEC. PATIENTS AND METHODS: A prospective database was reviewed to identify lymph node positive patients with PC treated with CRS-HIPEC within 1 year after primary tumour diagnosis between 2004 and 2012. Medical history of the patients was studied for the administration of perioperative systemic chemotherapy and follow-up. Outcome parameters were progression-free survival (PFS), overall survival (OS) and pattern of recurrence. RESULTS: Seventy-three patients treated with CRS-HIPEC for PC from lymph node positive colorectal carcinoma were identified. Fourteen patients received pre-CRS-HIPEC chemotherapy only, 32 patients underwent post-CRS-HIPEC chemotherapy only, 9 patients received chemotherapy both pre- and post-CRS-HIPEC and 16 patients did not receive any systemic chemotherapy. Of the 47 patients who did not receive pre-CRS-HIPEC chemotherapy, 11 (23%) did not receive any chemotherapy due to major postoperative complications. PFS and OS were significantly higher in patients who received systemic chemotherapy (PFS: median 15 versus 4 months, P = 0.024; OS: median 30 versus 14 months, P = 0.015), although this difference was attenuated after adjustment for major complications. Different chemotherapy timings did not differ significantly in either survival or recurrence patterns. CONCLUSIONS: In patients with PC from lymph node positive colorectal carcinoma, perioperative systemic chemotherapy is associated with increased OS and PFS, although this difference may be partly explained by the occurrence of major postoperative complication; with no evidence of difference in PFS, OS and systemic recurrence rate by timing of systemic chemotherapy.
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Carcinoma/tratamiento farmacológico , Neoplasias Colorrectales/tratamiento farmacológico , Recurrencia Local de Neoplasia/tratamiento farmacológico , Neoplasias Peritoneales/tratamiento farmacológico , Adulto , Anciano , Carcinoma/patología , Quimioterapia del Cáncer por Perfusión Regional , Neoplasias Colorrectales/patología , Neoplasias Colorrectales/cirugía , Terapia Combinada , Supervivencia sin Enfermedad , Femenino , Humanos , Hipertermia Inducida , Estimación de Kaplan-Meier , Ganglios Linfáticos/efectos de los fármacos , Ganglios Linfáticos/patología , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia/patología , Recurrencia Local de Neoplasia/cirugía , Atención Perioperativa , Neoplasias Peritoneales/patología , Neoplasias Peritoneales/cirugíaRESUMEN
AIM: To compare outcome of women with ovarian metastasis who underwent cytoreductive surgery and hyperthermic intraperitoneal chemotherapy (CRS-HIPEC) to outcome of women without ovarian metastasis who underwent CRS-HIPEC. METHODS: A prospective CRS-HIPEC database was searched to identify women with surgically treated colorectal carcinoma between 2000 and 2012. Patients with ovarian metastasis were identified and patients with peritoneal carcinomatosis but without ovarian metastasis were included as control cases. RESULTS: 75 patients with macroscopic ovarian metastasis underwent CRS-HIPEC with curative intent, while 50 female patients without ovarian metastasis were identified who underwent CRS-HIPEC. Patients with ovarian metastasis more often had a primary appendiceal tumour and had a more extensive intra-abdominal tumour load compared to patients without ovarian metastases. Median follow-up time was 45 months (95% confidence interval (CI): 37-53 months). Overall survival (OS) did not differ significantly between the two groups with a median OS in the ovarian metastasis group of 40 months (95% CI 26-54) compared to 64 months (95% CI 17-111, P = 0.478) in the non-ovarian metastasis group. Recurrence patterns did not differ significantly between groups (p = 0.183). CONCLUSIONS: Patients with ovarian metastasis of colorectal and appendiceal origin who underwent CRS-HIPEC had similar outcome compared to patients without ovarian metastasis. Given the findings of high coincidence of peritoneal metastases with ovarian metastases and ovarian metastases not being an independent factor for survival after CRS-HIPEC, this procedure should be recommended for patients with peritoneal metastases and ovarian metastases of colorectal and appendiceal carcinoma.
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Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Neoplasias del Apéndice/patología , Carcinoma/tratamiento farmacológico , Carcinoma/cirugía , Neoplasias Colorrectales/patología , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/cirugía , Neoplasias Peritoneales/tratamiento farmacológico , Neoplasias Peritoneales/cirugía , Adulto , Anciano , Carcinoma/metabolismo , Quimioterapia Adyuvante , Quimioterapia del Cáncer por Perfusión Regional/métodos , Bases de Datos Factuales , Supervivencia sin Enfermedad , Femenino , Estudios de Seguimiento , Humanos , Hipertermia Inducida , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Neoplasias Ováricas/metabolismo , Cavidad Peritoneal , Neoplasias Peritoneales/metabolismo , Estudios Prospectivos , Estudios Retrospectivos , Resultado del TratamientoRESUMEN
A 35-year-old man presented with progressive infected necrosis of the second digit of his right hand, caused by exposure to hydrogenfluoride.
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Dedos/patología , Ácido Fluorhídrico/efectos adversos , Adulto , Dedos/cirugía , Humanos , Masculino , Necrosis/inducido químicamente , Necrosis/cirugíaAsunto(s)
Oftalmopatías/etiología , Lupus Eritematoso Sistémico/complicaciones , Adolescente , Quimioterapia Combinada , Oftalmopatías/patología , Femenino , Glucocorticoides/uso terapéutico , Humanos , Lupus Eritematoso Sistémico/tratamiento farmacológico , Prednisolona/uso terapéutico , Prednisona/uso terapéuticoRESUMEN
Using anther-derived rice (Oryza sativa L.) cell-suspension cultures, we have identified an 18-kD protein that is posttranslationally modified by spermidine and is influenced by endogenous polyamine levels. The posttranslationally modified residue has been identified as the unusual amino acid hypusine [N[epsilon]-(4-amino-2-hydroxybutyl)lysine] by reverse-phase high-performance liquid chromatography and gas chromatography-mass-spectrometry analyses. Differential labeling of the protein with labeled amines provided evidence that the butylamine moiety of spermidine is the immediate precursor of the hypusine residue in the protein. The eukaryotic translation initiation factor 5A (eIF-5A) is the only known mammalian protein that undergoes a similar posttranslational modification with hypusine. The purified 18-kD protein co-electrophoreses with human translational initiation factor eIF-5A in both isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gels. The purified protein from rice stimulated methionyl-puromycin synthesis in vitro, indicating its functional similarity to mammalian eIF-5A. The results presented provide evidence that the posttranslationally modified 18-kD protein from rice containing hypusine is eIF-5A and suggest the conservation of hypusine-containing translation initiation factor eIF-5A in eukaryotes.
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High resolution, magnetic resonance imaging was used to quantitatively study the morphometry of the superior oblique muscles of two patients with superior oblique myokymia, as well as 18 superior oblique muscles of 14 patients with normal superior oblique function. The cross sectional area of each superior oblique muscle was measured at 3-millimeter intervals along the entire muscle length. In both cases of myokymia, the affected superior oblique muscles were significantly smaller than normal (P < .05). These anatomical changes in the superior oblique muscle of patients with myokymia suggest that an antecedent injury to the trochlear nerve has occurred. This injury, even if clinically unapparent, may be the initial event which leads to subsequent development of superior oblique myokymia.
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Fasciculación/diagnóstico , Trastornos de la Motilidad Ocular/diagnóstico , Músculos Oculomotores/patología , Adulto , Lesiones Encefálicas/complicaciones , Carbamazepina/uso terapéutico , Fasciculación/etiología , Fasciculación/terapia , Femenino , Humanos , Imagen por Resonancia Magnética , Masculino , Trastornos de la Motilidad Ocular/etiología , Trastornos de la Motilidad Ocular/terapia , Traumatismos del Nervio TroclearAsunto(s)
Defectos de la Visión Cromática/inducido químicamente , Deferoxamina/efectos adversos , Degeneración Retiniana/inducido químicamente , Anciano , Anemia Refractaria/terapia , Deferoxamina/administración & dosificación , Deferoxamina/uso terapéutico , Femenino , Angiografía con Fluoresceína , Fondo de Ojo , Humanos , Inyecciones SubcutáneasRESUMEN
When rice (Oryza sativa) cell suspension cultures are grown in the presence of [terminal methylenes-(3)H]spermidine, label is incorporated in a single polypeptide with a molecular mass of 18 kilodaltons on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Preincubation of cell cultures with polyamine biosynthesis inhibitors difluoromethylarginine and difluoromethylornithine, resulted in increased incorporation of the label into the 18 kilodalton polypeptide. In cells in which protein synthesis was arrested by cycloheximide, no label was detected in the 18 kilodalton polypeptide, suggesting a requirement for de novo protein synthesis.
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Two forms of the 32 kDa-D1 reaction center protein of photosystem II (PSII), having slightly different mobilities on denaturing polyacrylamide gels, have been resolved in Spirodela oligorrhiza, Glycine max L., Gossypium hirsutum L., Triticum aestivum L., and Zea mays L. The protein band with faster mobility is identified as the 32 kDa-D1 protein, and the less mobile band as a novel form, designated 32*. The two forms are structurally similar based on immunological and partial proteolytic tests. 32* is associated exclusively with the grana and is present in the PSII reaction center. Temporally, 32* appears several hours after the translocation of newly synthesized and processed 32 kDa-D1 protein from the stroma lamellae to the grana. Formation of the 32* is strictly light-dependent under physiological light intensities and correlates with a reciprocal loss of the 32-kDa form. Light induced formation of 32* is inhibited by 3-(3,4-dichlorophenyl)-1,1-dimethylurea but is not coupled to linear electron transport.
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2,4-Dinitrofenol/análogos & derivados , Clorofila/biosíntesis , Gránulos Citoplasmáticos/metabolismo , Proteínas de Plantas/biosíntesis , Plantas/metabolismo , Clorofila/aislamiento & purificación , Oscuridad , Dinitrofenoles/farmacología , Diurona/farmacología , Transporte de Electrón , Herbicidas/farmacología , Cinética , Luz , Complejos de Proteína Captadores de Luz , Peso Molecular , Fragmentos de Péptidos/aislamiento & purificación , Proteínas del Complejo del Centro de Reacción Fotosintética , Complejo de Proteína del Fotosistema II , Proteínas de Plantas/aislamiento & purificaciónRESUMEN
1-Aminocyclopropane-1-carboxylic acid (ACC) synthase (EC 4.4.1.14) is a key enzyme regulating ethylene biosynthesis in higher plants. A monoclonal antibody (mAb T20C) that immunoprecipitates the ACC synthase activity from tomato pericarp tissue extracts revealed that mAb T20C immunodecorates an approximately 67-kDa polypeptide. On isoelectric focusing gels, ACC synthase activity in cell-free preparations was resolved into three distinct activity peaks with pI values 5.3, 7, and 9. mAb T20C specifically recognized the pI 7 form of the enzyme on electrophoretic transfer (Western) blots. When analyzed by sodium dodecyl sulfate gel electrophoresis under reducing conditions, the eluted pI 7 form was confirmed to migrate as a polypeptide of 67 kDa. The 67-kDa pI 7 isoform is a previously undescribed form of ACC synthase.
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Association equilibria have been determined in the ternary system uridyl triplets (T)-ribosomal protein S1 (S)-ribosomes (Rb) depleted of S1 at 6 and 10 mM Mg2+. For 1:1 stoichiometry of reactants, four thermodynamically independent equilibria characterize the ternary system. The binary interaction Rb + T was studied by following the fluorescence quenching of labeled ribosomes by added T. The Rb + T association constant for UpUpUp triplets was 10-20-fold greater than for ApUpG triplets. The interaction Rb + S was studied by following the changes in fluorescence anisotropy when labeled S1 reacted with ribosomes. The remaining two independent equilibrium constants (for S + T and RbT + S) were obtained from fits to observed anisotropy measurements when varying amounts of T were added to a solution of ribosomes and fluorescently labeled S1. This indirect procedure allows one to measure S + T binding, an association that is difficult to determine directly. Over the concentration interval 5-10 mM Mg2+, the association constant for Rb + S increases with the sixth power of [Mg2+], whereas the association constant for S + T decreases approximately 2-fold as Mg2+ is increased from 6 to 10 mM Mg2+. T binds to Rb more tightly at 10 mM than at 6 mM Mg2+. When S1 is bound to Rb, however, at 10 mM Mg2+ the binding constant for T is decreased 10-fold and the Mg2+ dependence is reversed. These interactions can be described in terms of coupling free energies. For the ternary complex, three linearly independent coupling free energies can be written.(ABSTRACT TRUNCATED AT 250 WORDS)
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Oligonucleótidos/metabolismo , Oligorribonucleótidos/metabolismo , Proteínas Ribosómicas/metabolismo , Ribosomas/metabolismo , Sitios de Unión , Escherichia coli/metabolismo , Cinética , Modelos Biológicos , TermodinámicaRESUMEN
We have determined the equilibrium constants for the binding of AEDANS-labelled S1 to S1-depleted 30S and 70S ribosomes. For "tight" ribosomes, the association of S1 increases with the sixth power of Mg2+ concentration, but for 30S subunits and "loose" ribosomes, there is virtually no dependence of the association on Mg2+ over the same concentration range, 2-10 mM in Mg2+. The binding of S1 to 70S ribosomes at 10 mM Mg2+ is stabilized by 2 kcal/mol compared to the binding to 30S subunits. When intact S1 binds to tight ribosomes, the fluorescence anisotrophy is that for virtually complete rotational immobilization. The anisotropies vary considerably with the preparation and treatment of both S1 and ribosomes and these variations are detailed here. The results suggest the linkage of Mg2+-dependent conformational changes in the intact ribosomes, perhaps including rRNA, and the binding of S1.