ABSTRACT
Background: In the last years, laparoscopy has been progressively introduced in the management of advanced- stage ovarian cancer (AOC) not only to evaluate tumour resectability, but also to perform primary or interval minimally invasive debulking surgery in selected patients. During laparoscopic debulking for AOC, the need to change the surgical field to treat disease in the upper abdomen can be a time-consuming procedure. Objective: To demonstrate feasibility, safety and effectiveness of laparoscopic approach to remove bulky para- aortic nodes in AOC with a 30-degree 3D-endoscope without repositioning the laparoscopic surgical field. Materials and Methods: A 51-year-old woman was referred to our centre due to AOC with bulky para-aortic nodes (7 cm polylobate mass at CT-scan). The narrated surgical video article demonstrates the surgical steps for laparoscopic removal of bulky para-aortic nodes with a 30-degree 3D-endoscope, maintaining the vision from the upper abdomen perpendicular to the main axis of the vascular structures for the whole duration of the surgery ("top-bottom" view), without repositioning surgical field. Main Outcomes measures: Complete laparoscopic excision of disease was achieved. Results: Post-operative course was uneventful. Patient recovered from surgery and was able to start adjuvant chemotherapy within 30 days from surgery. Conclusions: Repositioning the surgical field to perform para-aortic dissection can be a time-consuming procedure during laparoscopic debulking for ovarian cancer. Laparoscopic removal of bulky para-aortic nodes with a 30-degree 3D-endoscope and "top-bottom view" is feasible, safe and effective.
ABSTRACT
BACKGROUND: According to the European Society for Medical Oncology/ European Society of Gynaecological Oncology/European Society for Radiotherapy and Oncology (ESMO/ESGO/ESTRO) Consensus Conference, the role of preoperative risk groups (RGs) in endometrial cancer (EC) is to direct surgical nodal staging. We compared diagnostic accuracy and economic impact of three work-up strategies to identify RGs. METHODS: A retrospective multicentre study including patients with early-stage EC. The three different work-up strategies were as follows:-Mondovì Hospital: transvaginal ultrasonography, pelvic magnetic resonance imaging (MRI); frozen section examination of the uterus in case of imaging discordance. High-risk patients underwent abdominal computed tomography.-Gemelli Hospital: transvaginal ultrasonography, MRI, One-Step Nucleic Acid Amplification (OSNA) of sentinel lymph node (SLN); frozen section examination of the uterus in case of imaging discordance.-Negrar Hospital: positron emission tomography (PET), frozen section examination of the uterus and of SLN. For statistical purposes patients were assigned, preoperatively and postoperatively, to two groups: group A (high-risk) and group B (not high-risk). RESULTS: Three hundred eighty-five patients were included (93 Mondovì, 215 Gemelli, 77 Negrar). Endometrial biopsy errors led to 47.3% misclassifications. Test accuracy of Mondovì, Gemelli and Negrar strategies was 0.83 (95%CI 0.734-0.901), 0.95 (95%CI 0.909-0.975) and 0.94 (95%CI 0.866-0.985), respectively. Preoperative work-up mean cost per patient in group A was 514.5 at Mondovì, 868.5 at Gemelli, and 1212.8 at Negrar hospital (p-value < 0.001), while in group B was 378.8 at Mondovì, 941.2 at Gemelli, and 1848.4 at Negrar hospital (p-value < 0.001). CONCLUSIONS: In our study, work-up strategies with more relevant economic impact showed a better diagnostic accuracy. Upcoming guidelines should specify recommendations about the gold standard work-up strategy, including the role of SLN.
ABSTRACT
Retinal gene therapy with adeno-associated viral (AAV) vectors is safe and effective in humans. However, the limited cargo capacity of AAV prevents their use for therapy of those inherited retinopathies (IRs) due to mutations in large (>5 kb) genes. Viral vectors derived from adenovirus (Ad), lentivirus (LV) and herpes virus (HV) can package large DNA sequences, but do not target efficiently retinal photoreceptors (PRs) where the majority of genes responsible for IRs are expressed. Here, we have evaluated the mouse retinal transduction profiles of vectors derived from 16 different Ad serotypes, 7 LV pseudotypes and from a bovine HV. Most of the vectors tested transduced efficiently the retinal pigment epithelium. We found that LV-GP64 tends to transduce more PRs than the canonical LV-VSVG, albeit this was restricted to a narrow region. We observed more extensive PR transduction with HdAd1, 2 and 5/F35++ than with LV, although none of them outperformed the canonical HdAd5 or matched the extension of PR transduction achieved with AAV2/8.
Subject(s)
Dependovirus/genetics , Herpesvirus 4, Bovine/genetics , Lentivirus/genetics , Retinal Pigment Epithelium/virology , Animals , Dependovirus/classification , Electroretinography , Epithelial Cells/virology , Genetic Vectors/administration & dosage , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Herpesvirus 4, Bovine/classification , Lentivirus/classification , Male , Mice , Mice, Inbred BALB C , Photoreceptor Cells, Vertebrate/metabolism , Retinal Pigment Epithelium/cytology , Transduction, GeneticABSTRACT
Gene therapy with adeno-associated viral (AAV) vectors is limited by AAV cargo capacity that prevents their application to the inherited retinal diseases (IRDs), such as Stargardt disease (STGD) or Usher syndrome type IB (USH1B), which are due to mutations in genes larger than 5 kb. Trans-splicing or hybrid dual AAV vectors have been successfully exploited to reconstitute large gene expression in the mouse retina. Here, we tested them in the large cone-enriched pig retina that closely mimics the human retina. We found that dual AAV trans-splicing and hybrid vectors transduce pig photoreceptors, the major cell targets for treatment of IRDs, to levels that were about two- to threefold lower than those obtained with a single AAV vector of normal size. This efficiency is significantly higher than that in mice, and is potentially due to the high levels of dual AAV co-transduction we observe in pigs. We also show that subretinal delivery in pigs of dual AAV trans-splicing and hybrid vectors successfully reconstitute, albeit at variable levels, the expression of the large genes ABCA4 and MYO7A mutated in STGD and USH1B, respectively. Our data support the potential of dual AAV vectors for large gene reconstitution in the cone-enriched pig retina that is a relevant preclinical model.
Subject(s)
Gene Transfer Techniques , Genetic Therapy , Usher Syndromes/genetics , ATP-Binding Cassette Transporters/genetics , Animals , Dependovirus/genetics , Gene Expression Regulation , Genetic Vectors , Humans , Macular Degeneration/genetics , Macular Degeneration/therapy , Mice , Myosin VIIa , Myosins/genetics , Photoreceptor Cells/metabolism , Photoreceptor Cells/pathology , Stargardt Disease , Sus scrofa , Usher Syndromes/therapyABSTRACT
The aim of this study was to evaluate the risk of non-Hodgkin's lymphoma (NHL) in an adult population residing in an area in northern Italy exposed to industrial air pollution from a big power plant, a coke oven, 2 chemical factories, and some minor plants. The design was a population-based case-control study and information about residential history and the main risk factors for NHL was obtained interviewing 133 cases and 279 controls using a structured questionnaire. Three exposure categories (heavy, moderate, and slight) were defined on the basis of the location of the major facilities with respect to the subject residence. NHL risk was not associated either with location or duration of residence in the heavily polluted area. However, the unavoidable limitations of this study prevent us from drawing definitive conclusions.
Subject(s)
Air Pollutants , Air Pollution/statistics & numerical data , Coke , Environmental Exposure/statistics & numerical data , Lymphoma, Non-Hodgkin/epidemiology , Power Plants/statistics & numerical data , Case-Control Studies , Humans , Italy/epidemiology , Lymphoma, Non-Hodgkin/chemically induced , Risk Assessment , Risk FactorsABSTRACT
La transfusión de sangre alogénica (TSA) no es inocua, y como consecuencia han surgido múltiples alternativas a la misma (ATSA). Existe variabilidad respecto a las indicaciones y buen uso de las ATSA. Dependiendo de la especialidad de los médicos que tratan a los pacientes, el grado de anemia, la política transfusional, la disponibilidad de las ATSA y el criterio personal, estas se usan de forma variable. Puesto que las ATSA tampoco son inocuas y pueden no cumplir criterios de coste-efectividad, la variabilidad en su uso es inaceptable. Las sociedades españolas de Anestesiología y Reanimación (SEDAR), Hematología y Hemoterapia(SEHH), Farmacia Hospitalaria (SEFH), Medicina Intensiva y Unidades Coronarias(SEMICYUC), Trombosis y Hemostasia (SETH) y Transfusiones Sanguíneas (SETS) han elaborado un documento de consenso para el buen uso de la ATSA. Un panel de expertos de las 6sociedades ha llevado a cabo una revisión sistemática de la literatura médica y elaborado el 2013. Documento Sevilla de Consenso sobre Alternativas a la Transfusión de Sangre Alogénica. Solo se contempla las ATSA dirigidas a disminuir la transfusión de concentrado de hematíes. Se definen las ATSA como toda medida farmacológica y no farmacológica encaminada a disminuir la transfusión de concentrado de hematíes, preservando siempre la seguridad del paciente. La cuestión principal que se plantea en cada ítem se formula, en forma positiva o negativa, como: La ATSA en cuestión reduce/no reduce la tasa transfusional». Para formular el grado de recomendación se ha usado la metodología Grades of Recommendation Assessment, Development and Evaluation (GRADE) (AU)
Since allogeneic blood transfusion (ABT) is not harmless, multiple alternatives to ABT (AABT) have emerged, though there is great variability in their indications and appropriate use. This variability results from the interaction of a number of factors, including the specialty of the physician, knowledge and preferences, the degree of anemia, transfusion policy, and AABT availability. Since AABTs are not harmless and may not meet cost-effectiveness criteria, such variability is unacceptable. The Spanish Societies of Anesthesiology (SEDAR), Hematology and Hemotherapy (SEHH), Hospital Pharmacy (SEFH), Critical Care Medicine (SEMICYUC), Thrombosis and Hemostasis (SETH)and Blood Transfusion (SETS) have developed a Consensus Document for the proper use of AABTs. A panel of experts convened by these 6 Societies have conducted a systematic review of the medical literature and have developed the 2013 Seville Consensus Document on Alternatives to Allogeneic Blood Transfusion, which only considers those AABT aimed at decreasing the transfusion of packed red cells. AABTs are defined as any pharmacological or non-pharmacological measure aimed at decreasing the transfusion of red blood cell concentrates, while preserving patient safety. For each AABT, the main question formulated, positively or negatively, is: Does this particular AABT reduce the transfusion rate or not? All the recommendations on the use of AABTs were formulated according to the Grades of Recommendation Assessment, Development and Evaluation(GRADE) methodology (AU)
Subject(s)
Humans , Blood Transfusion, Autologous , Blood Transfusion/methods , Blood Substitutes/therapeutic use , Anemia/therapy , Glycated Hemoglobin/therapeutic use , Fibrinogen/therapeutic use , Practice Patterns, Physicians'ABSTRACT
La transfusión de sangre alogénica (TSA) no es inocua, y como consecuencia han surgido múltiples alternativas a la misma (ATSA). Existe variabilidad respecto a las indicaciones y buen uso de las ATSA. Dependiendo de la especialidad de los médicos que tratan a los pacientes, el grado de anemia, la política transfusional, la disponibilidad de las ATSA y el criterio personal, estas se usan de forma variable. Puesto que las ATSA tampoco son inocuas y pueden no cumplir criterios de coste-efectividad, la variabilidad en su uso es inaceptable. Las sociedades españolas de Anestesiología y Reanimación (SEDAR), Hematología y Hemoterapia (SEHH), Farmacia Hospitalaria (SEFH), Medicina Intensiva y Unidades Coronarias (SEMICYUC), Trombosis y Hemostasia (SETH) y Transfusiones Sanguíneas (SETS) han elaborado un documento de consenso para el buen uso de la ATSA. Un panel de expertos de las 6 sociedades ha llevado a cabo una revisión sistemática de la literatura médica y elaborado el 2013. Documento Sevilla de Consenso sobre Alternativas a la Transfusión de Sangre Alogénica. Solo se contempla las ATSA dirigidas a disminuir la transfusión de concentrado de hematíes. Se definen las ATSA como toda medida farmacológica y no farmacológica encaminada a disminuir la transfusión de concentrado de hematíes, preservando siempre la seguridad del paciente. La cuestión principal que se plantea en cada ítem se formula, en forma positiva o negativa, como: «La ATSA en cuestión reduce/no reduce la tasa transfusional». Para formular el grado de recomendación se ha usado la metodología Grades of Recommendation Assessment, Development and Evaluation (GRADE) (AU)
Since allogeneic blood transfusion (ABT) is not harmless, multiple alternatives to ABT (AABT) have emerged, though there is great variability in their indications and appropriate use. This variability results from the interaction of a number of factors, including the specialty of the physician, knowledge and preferences, the degree of anemia, transfusion policy, and AABT availability. Since AABTs are not harmless and may not meet cost-effectiveness criteria, such variability is unacceptable. The Spanish Societies of Anesthesiology (SEDAR), Hematology and Hemotherapy (SEHH), Hospital Pharmacy (SEFH), Critical Care Medicine (SEMICYUC), Thrombosis and Hemostasis (SETH) and Blood Transfusion (SETS) have developed a Consensus Document for the proper use of AABTs. A panel of experts convened by these 6 Societies have conducted a systematic review of the medical literature and have developed the 2013 Seville Consensus Document on Alternatives to Allogeneic Blood Transfusion, which only considers those AABT aimed at decreasing the transfusion of packed red cells. AABTs are defined as any pharmacological or non-pharmacological measure aimed at decreasing the transfusion of red blood cell concentrates, while preserving patient safety. For each AABT, the main question formulated, positively or negatively, is: "Does this particular AABT reduce the transfusion rate or not?" All the recommendations on the use of AABTs were formulated according to the Grades of Recommendation Assessment, Development and Evaluation (GRADE) methodology (AU)
Subject(s)
Humans , Male , Female , Transplantation, Homologous/instrumentation , Transplantation, Homologous/methods , Transplantation, Homologous , Cost-Benefit Analysis/organization & administration , Cost-Benefit Analysis/standards , Cost-Benefit Analysis , Evaluation of the Efficacy-Effectiveness of Interventions , Anesthesiology/methods , Transplantation, Homologous/standards , Transplantation, Homologous/trends , 50303 , Anesthesiology/organization & administration , Anesthesiology/standards , Erythrocyte Transfusion/trends , Erythrocyte TransfusionABSTRACT
Since allogeneic blood transfusion (ABT) is not harmless, multiple alternatives to ABT (AABT) have emerged, though there is great variability in their indications and appropriate use. This variability results from the interaction of a number of factors, including the specialty of the physician, knowledge and preferences, the degree of anemia, transfusion policy, and AABT availability. Since AABTs are not harmless and may not meet cost-effectiveness criteria, such variability is unacceptable. The Spanish Societies of Anesthesiology (SEDAR), Hematology and Hemotherapy (SEHH), Hospital Pharmacy (SEFH), Critical Care Medicine (SEMICYUC), Thrombosis and Hemostasis (SETH) and Blood Transfusion (SETS) have developed a Consensus Document for the proper use of AABTs. A panel of experts convened by these 6 Societies have conducted a systematic review of the medical literature and have developed the 2013 Seville Consensus Document on Alternatives to Allogeneic Blood Transfusion, which only considers those AABT aimed at decreasing the transfusion of packed red cells. AABTs are defined as any pharmacological or non-pharmacological measure aimed at decreasing the transfusion of red blood cell concentrates, while preserving patient safety. For each AABT, the main question formulated, positively or negatively, is: « Does this particular AABT reduce the transfusion rate or not?¼ All the recommendations on the use of AABTs were formulated according to the Grades of Recommendation Assessment, Development and Evaluation (GRADE) methodology.
Subject(s)
Blood Transfusion/standards , Complementary Therapies , Humans , Patient Safety , Surgical Procedures, OperativeABSTRACT
Since allogeneic blood transfusion (ABT) is not harmless, multiple alternatives to ABT (AABT) have emerged, though there is great variability in their indications and appropriate use. This variability results from the interaction of a number of factors, including the specialty of the physician, knowledge and preferences, the degree of anemia, transfusion policy, and AABT availability. Since AABTs are not harmless and may not meet cost-effectiveness criteria, such variability is unacceptable. The Spanish Societies of Anesthesiology (SEDAR), Hematology and Hemotherapy (SEHH), Hospital Pharmacy (SEFH), Critical Care Medicine (SEMICYUC), Thrombosis and Hemostasis (SETH) and Blood Transfusion (SETS) have developed a Consensus Document for the proper use of AABTs. A panel of experts convened by these 6 Societies have conducted a systematic review of the medical literature and have developed the 2013 Seville Consensus Document on Alternatives to Allogeneic Blood Transfusion, which only considers those AABT aimed at decreasing the transfusion of packed red cells. AABTs are defined as any pharmacological or non-pharmacological measure aimed at decreasing the transfusion of red blood cell concentrates, while preserving patient safety. For each AABT, the main question formulated, positively or negatively, is: "Does this particular AABT reduce the transfusion rate or not?" All the recommendations on the use of AABTs were formulated according to the Grades of Recommendation Assessment, Development and Evaluation (GRADE) methodology.
Subject(s)
Bloodless Medical and Surgical Procedures/standards , Humans , Practice Guidelines as TopicABSTRACT
OBJECTIVES: Reconstructive surgery plays an important role in cosmetic and functional results of major excisional surgery performed as a treatment for invasive vulvar cancer. Traditional techniques -- gracilis myocutaneous o rectus abdominis flaps -- have several limits. We describe here a different surgical approach that we have used since 1998 in an effort to obtain better results in vulvar reconstruction. METHODS: From January 1998 to June 2007, thirty three patients who underwent excisional radical surgery for invasive vulvar tumors, were treated with vulvar reconstruction using the gluteal fold fascio-cutaneous local flap. Flaps were designed along the gluteal fold in adequate length and size. They were oval or triangular in shape depending on the defect they were supposed to cover. The flaps -- which always included the fascial layer -- were raised up to identify a perforator branch of the internal pudendal artery and then harvested as an island flap to achieve better mobility. RESULTS: We had no major complications, only two patients presented marginal necrosis and eight patients experienced significant seromas. Advantages over the alternative techniques included reduced dimensions of scars, absence of flap liponecrosis, no need of modifying patient's position on the surgical table, and very limited blood loss. CONCLUSIONS: We conclude that gluteal fold flap offers excellent cosmetic and functional results with a low complication rate. Therefore we support the gluteal fold flap as a valid surgical option whenever reconstruction is needed after radical excision of vulvar neoplasms.
Subject(s)
Plastic Surgery Procedures/methods , Surgical Flaps , Vulvar Neoplasms/surgery , Aged , Aged, 80 and over , Carcinoma, Squamous Cell/surgery , Female , Gynecologic Surgical Procedures/methods , Humans , Middle Aged , Neoplasm Recurrence, Local/surgeryABSTRACT
BACKGROUND: When preparing for fertilization, oocytes undergo meiotic maturation during which structural changes occur in the endoplasmic reticulum (ER) that lead to a more efficient calcium response. During meiotic maturation and subsequent fertilization, the actin cytoskeleton also undergoes dramatic restructuring. We have recently observed that rearrangements of the actin cytoskeleton induced by actin-depolymerizing agents, or by actin-binding proteins, strongly modulate intracellular calcium (Ca2+) signals during the maturation process. However, the significance of the dynamic changes in F-actin within the fertilized egg has been largely unclear. METHODOLOGY/PRINCIPAL FINDINGS: We have measured changes in intracellular Ca2+ signals and F-actin structures during fertilization. We also report the unexpected observation that the conventional antagonist of the InsP(3) receptor, heparin, hyperpolymerizes the cortical actin cytoskeleton in postmeiotic eggs. Using heparin and other pharmacological agents that either hypo- or hyperpolymerize the cortical actin, we demonstrate that nearly all aspects of the fertilization process are profoundly affected by the dynamic restructuring of the egg cortical actin cytoskeleton. CONCLUSIONS/SIGNIFICANCE: Our findings identify important roles for subplasmalemmal actin fibers in the process of sperm-egg interaction and in the subsequent events related to fertilization: the generation of Ca2+ signals, sperm penetration, cortical granule exocytosis, and the block to polyspermy.
Subject(s)
Actin Cytoskeleton/physiology , Calcium Signaling/physiology , Cytoskeleton/physiology , Fertilization/physiology , Sperm-Ovum Interactions/physiology , Actin Cytoskeleton/drug effects , Actin Cytoskeleton/metabolism , Actins/metabolism , Animals , Calcium Signaling/drug effects , Cytoskeleton/drug effects , Depsipeptides/pharmacology , Female , Fertilization/drug effects , Heparin/pharmacology , Male , Protein Multimerization/drug effects , Spermatozoa/physiology , Starfish/physiology , Zygote/metabolism , Zygote/physiologyABSTRACT
Intermediate Care Units are created for patients who predictably have low risk of requiring therapeutic life support measures but who require more monitoring and nursing cares than those received in the conventional hospitalization wards. Previous studies have demonstrated that Intermediate Care Units may promote hospital care grading, allowing for better classification in critical patients, improving efficacy and efficiency of the ICUs and thus decreasing costs and above all mortality in the conventional hospitalization wards. This document attempts to group the currently existing knowledge that served as a base for the consensus meeting on the application of them in the establishment of future ICUs in our hospital setting.
Subject(s)
Critical Care/classification , Intensive Care Units , Critical Care/methods , Critical Care/organization & administration , Critical Care/standardsABSTRACT
The aim of the study was to investigate the variations in prostate cancer prognosis during a period of major diagnostic change, such as the introduction of the prostate-specific antigen (PSA) test. Data were provided by 14 Italian cancer registries (CRs). Incidence and follow-up information was collected for patients diagnosed from 1978 to 1994. Relative survival was computed taking into account incidence period, age, tumour stage and grade at diagnosis. A multivariate analysis was carried out to evaluate the independent simultaneous effect on survival of some prognostic determinants. A large geographical variability was observed: in 1993-1994 Italian survival rates ranged from 76% to 52%, with a north-south gradient. A striking prognostic improvement (up to +27 percentage points) between the late 1980s and the early 1990s occurred in almost all CRs, particularly with regard to younger patients. Multivariate analysis showed a strong influence of incidence period on survival, also after correction by tumour stage. The slowdown of metastatic cancers suggests that the survival improvement could be due both to the introduction of an effective opportunistic screening and to a quantitative change in the application of clinical treatment, even if the effect of the lead-time bias phenomenon has to be taken into account.
Subject(s)
Prostatic Neoplasms/mortality , Prostatic Neoplasms/prevention & control , Age Factors , Aged , Biomarkers, Tumor , Humans , Incidence , Italy/epidemiology , Male , Mass Screening/methods , Middle Aged , Neoplasm Metastasis , Neoplasm Staging , Prognosis , Prostate-Specific Antigen , Prostatic Neoplasms/etiology , Prostatic Neoplasms/pathology , Registries , Survival AnalysisABSTRACT
Glutathione (GSH) and homo-GSH (hGSH) are the major low-molecular weight thiols synthesized in Medicago truncatula. Two M. truncatula cDNAs (gshs1 and gshs2) corresponding to a putative GSH synthetase (GSHS) and a putative hGSH synthetase (hGSHS) were characterized. Heterologous expression of gshs1 and gshs2 cDNAs in an Escherichia coli strain deficient in GSHS activity showed that GSHS1 and GSHS2 are a GSHS and an hGSHS, respectively. Leucine-534 and proline-535 present in hGSHS were substituted by alanines that are conserved in plant GSHS. These substitutions resulted in a strongly stimulated GSH accumulation in the transformed E. coli strain showing that these residues play a crucial role in the differential recognition of beta-alanine and glycine by hGSHS. Phylogenetic analysis of GSHS2 and GSHS1 with other eukaryotic GSHS sequences indicated that gshs2 and gshs1 are the result of a gene duplication that occurred after the divergence between Fabales, Solanales, and Brassicales. Analysis of the structure of gshs1 and gshs2 genes shows they are both present in a cluster and in the same orientation in the M. truncatula genome, suggesting that the duplication of gshs1 and gshs2 occurred via a tandem duplication.
Subject(s)
Gene Duplication , Glutathione Synthase/genetics , Medicago sativa/enzymology , Peptide Synthases/genetics , Amino Acid Sequence , Animals , Chromosome Mapping , DNA, Complementary/analysis , DNA, Plant/analysis , Escherichia coli/genetics , Genes, Plant , Glutathione Synthase/classification , Glutathione Synthase/metabolism , Humans , Medicago sativa/genetics , Molecular Sequence Data , Peptide Synthases/classification , Peptide Synthases/metabolism , Phylogeny , Sequence Homology, Amino Acid , Tandem Repeat SequencesABSTRACT
Reactive oxygen species are produced as an early event in plant defense response against avirulent pathogens. We show here that alfalfa responds to infection with Sinorhizobium meliloti by production of superoxide and hydrogen peroxide. This similarity in the early response to infection by pathogenic and symbiotic bacteria addresses the question of which mechanism rhizobia use to counteract the plant defense response.
Subject(s)
Medicago sativa/metabolism , Medicago sativa/microbiology , Sinorhizobium meliloti/metabolism , Hydrogen Peroxide/metabolism , Medicago sativa/ultrastructure , Microscopy, Electron , Nitrogen Fixation , Respiratory Burst , Superoxides/metabolism , SymbiosisABSTRACT
In nitrogen-poor soils, rhizobia elicit nodule formation on legume roots, within which they differentiate into bacteroids that fix atmospheric nitrogen. Protection against reactive oxygen species (ROS) was anticipated to play an important role in Rhizobium-legume symbiosis because nitrogenase is extremely oxygen sensitive. We deleted the sodA gene encoding the sole cytoplasmic superoxide dismutase (SOD) of Sinorhizobium meliloti. The resulting mutant, deficient in superoxide dismutase, grew almost normally and was only moderately sensitive to oxidative stress when free living. In contrast, its symbiotic properties in alfalfa were drastically affected. Nitrogen-fixing ability was severely impaired. More strikingly, most SOD-deficient bacteria did not reach the differentiation stage of nitrogen-fixing bacteroids. The SOD-deficient mutant nodulated poorly and displayed abnormal infection. After release into plant cells, a large number of bacteria failed to differentiate into bacteroids and rapidly underwent senescence. Thus, bacterial SOD plays a key protective role in the symbiotic process.
Subject(s)
Fabaceae/microbiology , Fabaceae/physiology , Plants, Medicinal , Rhizobium/physiology , Superoxide Dismutase/physiology , Symbiosis/physiology , Bacterial Proteins/physiologyABSTRACT
This paper examines the survival of elderly European cancer patients, on the basis of the EUROCARE II results. Using Hakulinen and Abeywickrama's method, the relative survival rates at 1 and 5 years from diagnosis were computed by sex and quinquennial age group for the elderly (65-99 years old). Age-standardised rates for the whole elderly group were also calculated. The analysis covered: all malignancies combined, stomach, colon, rectum, pancreas, lung, melanoma, bladder, kidney and non-Hodgkin's lymphomas for both sexes; prostate and larynx for men; and breast, ovary, uterine cervix and corpus for women. Data relating to 701521 cancer patients came from 44 population-based cancer registries in 16 European countries. The relative risks of death (RRs) of older patients (65-99) with respect to middle-aged adults (55-64) were computed by sex and country, for all malignancies only. The most prominent finding was the decrease in survival rates with increasing age for almost all cancer sites. The age-curves of survival rates at 1 year from diagnosis usually had a steeper slope than those at 5 years, particularly in women. This suggests that disease stage at presentation plays an important role in determining survival, particularly in the elderly. Thus, all factors which influence timing diagnosis in the elderly and cause a delay in tumour detection, such as psycho-social factors, access to care, co-morbidities and other clinical features affecting performance status, are very important predictors of prognosis. Very large geographic variations in relative survival rates were found among European countries. The ordering of countries was similar for almost all cancer sites. Western and Central Europe generally had the best survival, followed by Northern countries and by Southern ones (the latter with survival around the European average: 39% in men, 47% in women). The UK had survival rates unexpectedly lower than rates of nearest nations, often below the European average. Eastern countries usually had the lowest rates. In the very elderly patients (over 85 years), an apparent rise in the survival rates was noted, particularly at 5 years from diagnosis and in men. This 'too good' survival is unlikely to be due to real better prognosis, but rather to a selection bias. Countries with this unusual rise are also those registering a high proportion of DCO cases (those cases retrieved by death certificate only) (around 10%) or DCO unavailable. Another 'natural' bias has also to be taken into account: in elderly patients with a very bad prognosis, who are often suffering from other serious co-morbid conditions, cancer diagnoses could be under-notified and not reach at all the data sources commonly monitored by cancer registries.
Subject(s)
Neoplasms/mortality , Age Factors , Aged , Aged, 80 and over , Ethnicity , Europe/epidemiology , Female , Humans , Male , Middle Aged , Neoplasms/diagnosis , Neoplasms/epidemiology , Prognosis , Quality of Health Care/standards , Registries , Sex Factors , Survival RateABSTRACT
Sinorhizobium meliloti Rm5000 is an aerobic bacterium that can live free in the soil or in symbiosis with the roots of leguminous plants. A single detectable superoxide dismutase (SOD) was found in free-living growth conditions. The corresponding gene was isolated from a genomic library by using a sod fragment amplified by PCR from degenerate primers as a probe. The sodA gene was located in the chromosome. It is transcribed monocistronically and encodes a 200-amino-acid protein with a theoretical M(r) of 22,430 and pI of 5. 8. S. meliloti SOD complemented a deficient E. coli mutant, restoring aerobic growth of a sodA sodB recA strain, when the gene was expressed from the synthetic tac promoter but not from its own promoter. Amino acid sequence alignment showed great similarity with Fe-containing SODs (FeSODs), but the enzyme was not inactivated by H(2)O(2). The native enzyme was purified and found to be a dimeric protein, with a specific activity of 4,000 U/mg. Despite its Fe-type sequence, atomic absorption spectroscopy showed manganese to be the cofactor (0.75 mol of manganese and 0.24 mol of iron per mol of monomer). The apoenzyme was prepared from crude extracts of S. meliloti. Activity was restored by dialysis against either MnCl(2) or Fe(NH(4))(2)(SO(4))(2), demonstrating the cambialistic nature of the S. meliloti SOD. The recovered activity with manganese was sevenfold higher than with iron. Both reconstituted enzymes were resistant to H(2)O(2). Sequence comparison with 70 FeSODs and MnSODs indicates that S. meliloti SOD contains several atypical residues at specific sites that might account for the activation by manganese and resistance to H(2)O(2) of this unusual Fe-type SOD.
Subject(s)
Bacterial Proteins/genetics , Sinorhizobium meliloti/enzymology , Sinorhizobium meliloti/genetics , Superoxide Dismutase/genetics , Amino Acid Sequence , Bacterial Proteins/isolation & purification , Bacterial Proteins/metabolism , Base Sequence , Chromosome Mapping , Chromosomes, Bacterial , Cloning, Molecular , DNA Primers , Dimerization , Genomic Library , Kinetics , Molecular Sequence Data , Polymerase Chain Reaction , Recombinant Proteins/isolation & purification , Recombinant Proteins/metabolism , Sequence Alignment , Sequence Homology, Amino Acid , Sinorhizobium meliloti/growth & development , Superoxide Dismutase/isolation & purification , Superoxide Dismutase/metabolismABSTRACT
Two catalases, KatA and KatB, have been detected in Sinorhizobium meliloti growing on rich medium. Here we characterize a new catalase gene encoding a third catalase (KatC). KatC activity was detectable only at the end of the stationary phase in S. meliloti growing in minimum medium, whereas KatA activity was found during the exponential phase. Analysis with a katC-lacZ fusion demonstrated that katC expression is mainly regulated at the transcription level. An increase of catalase activity correlating with KatA induction was detected in bacteroids. A dramatic decrease of nitrogen fixation capacity in a katA katC double mutant was observed, suggesting that these catalases are very important for the protection of the nitrogen fixation process.
Subject(s)
Arabidopsis Proteins , Catalase/genetics , Gene Expression Regulation, Bacterial , Plant Proteins/genetics , Sinorhizobium meliloti/genetics , Catalase/biosynthesis , Genes, Bacterial , Medicago sativa/microbiology , Molecular Sequence Data , Mutation , Nitrogen Fixation/genetics , Plant Proteins/biosynthesis , Sinorhizobium meliloti/enzymology , Sinorhizobium meliloti/growth & development , SymbiosisABSTRACT
A gamma-ECS cDNA from Medicago truncatula was isolated using an Arabidopsis thaliana cDNA as probe. The analysis of the amino acid sequence deduced from this cDNA revealed 80% identity with the gamma-ECS from A. thaliana and Brassica juncea and suggested a plastidial localisation for the enzyme. Gamma-ECS activity and high level of GSH were detected in the gamma-ECS-deficient E. coli strain expressing a fusion protein containing the M. truncatula gamma-ECS protein. Southern blot analysis suggests that gamma-ECS is encoded by a small multigenic family in M. truncatula and shows that homologous genes are present in two other leguminous plants, Medicago sativa and Pisum sativum. Gamma-ECS gene expression was analysed by Northern blot in seedlings, plantlets and mature plants.
