Evidence Of Allopolyploidy In Urochloa Humidicola Based On Cytological Analysis And Genetic Linkage Mapping

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)The African species Urochloa humidicola (Rendle) Morrone & Zuloaga (syn. Brachiaria humidicola (Rendle) Schweick.) is an important perennial forage grass found throughout the tropics. This species is polyploid, ranging from tetra to nonaploid, and apomictic, which makes genetic studies challenging; therefore, the number of currently available genetic resources is limited. The genomic architecture and evolution of U. humidicola and the molecular markers linked to apomixis were investigated in a full-sib F-1 population obtained by crossing the sexual accession H031 and the apomictic cultivar U. humidicola cv. BRS Tupi, both of which are hexaploid. A simple sequence repeat (SSR)-based linkage map was constructed for the species from 102 polymorphic and specific SSR markers based on simplex and double-simplex markers. The map consisted of 49 linkage groups (LGs) and had a total length of 1702.82 cM, with 89 microsatellite loci and an average map density of 10.6 cM. Eight homology groups (HGs) were formed, comprising 22 LGs, and the other LGs remained ungrouped. The locus that controls apospory (apo-locus) was mapped in LG02 and was located 19.4 cM from the locus Bh027.c.D2. In the cytological analyses of some hybrids, bi-to hexavalents at diakinesis were observed, as well as two nucleoli in some meiocytes, smaller chromosomes with preferential allocation within the first metaphase plate and asynchronous chromosome migration to the poles during anaphase. The linkage map and the meiocyte analyses confirm previous reports of hybridization and suggest an allopolyploid origin of the hexaploid U. humidicola. This is the first linkage map of an Urochloa species, and it will be useful for future quantitative trait locus (QTL) analysis after saturation of the map and for genome assembly and evolutionary studies in Urochloa spp. Moreover, the results of the apomixis mapping are consistent with previous reports and confirm the need for additional studies to search for a co-segregating marker.11Brazilian Agricultural Research Corporation (Embrapa)Brazilian National Council for Scientific and Technological Development (CNPq) [478262/2004-3, 502336/2005-6, 482458/2007-0]State of Sao Paulo Research Foundation (FAPESP) [2005/51010-0, 2008/52197-4]Foundation for Science and Technological Development of the State of Mato Grosso do Sul (Fundect)Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior (CAPES)FAPESP [2007/57022-5, 2010/50032-8]Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior (CAPES - EMBRAPA Program)CNPq [304914/2010-0]CNPqConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Promiscuous Binding of Invariant Chain-Derived CLIP Peptide to Distinct HLA-I Molecules Revealed in Leukemic Cells

Antigen presentation by HLA class I (HLA-I) and HLA class II (HLA-II) complexes is achieved by proteins that are specific for their respective processing pathway. The invariant chain (Ii)-derived peptide CLIP is required for HLA-II-mediated antigen presentation by stabilizing HLA-II molecules before antigen loading through transient and promiscuous binding to different HLA-II peptide grooves. Here, we demonstrate alternative binding of CLIP to surface HLA-I molecules on leukemic cells. In HLA-II-negative AML cells, we found plasma membrane display of the CLIP peptide. Silencing Ii in AML cells resulted in reduced HLA-I cell surface display, which indicated a direct role of CLIP in the HLA-I antigen presentation pathway. In HLA-I-specific peptide eluates from B-LCLs, five Ii-derived peptides were identified, of which two were from the CLIP region. In vitro peptide binding assays strikingly revealed that the eluted CLIP peptide RMATPLLMQALPM efficiently bound to four distinct HLA-I supertypes (-A2, -B7, -A3, -B40). Furthermore, shorter length variants of this CLIP peptide also bound to these four supertypes, although in silico algorithms only predicted binding to HLA-A2 or -B7. Immunization of HLA-A2 transgenic mice with these peptides did not induce CTL responses. Together these data show a remarkable promiscuity of CLIP for binding to a wide variety of HLA-I molecules. The found participation of CLIP in the HLA-I antigen presentation pathway could reflect an aberrant mechanism in leukemic cells, but might also lead to elucidation of novel processing pathways or immune escape mechanisms

Towards a Clinically Relevant Lentiviral Transduction Protocol for Primary Human CD34+ Hematopoietic Stem/Progenitor Cells

Background: Hematopoietic stem cells (HSC), in particular mobilized peripheral blood stem cells, represent an attractive target for cell and gene therapy. Efficient gene delivery into these target cells without compromising self-renewal and multipotency is crucial for the success of gene therapy. We investigated factors involved in the ex vivo transduction of CD34 + HSCs in order to develop a clinically relevant transduction protocol for gene delivery. Specifically sought was a protocol that allows for efficient transduction with minimal ex vivo manipulation without serum or other reagents of animal origin. Methodology/Principal Findings: Using commercially available G-CSF mobilized peripheral blood (PB) CD34 + cells as the most clinically relevant target, we systematically examined factors including the use of serum, cytokine combinations, prestimulation time, multiplicity of infection (MOI), transduction duration and the use of spinoculation and/or retronectin. A self-inactivating lentiviral vector (SIN-LV) carrying enhanced green fluorescent protein (GFP) was used as the gene delivery vehicle. HSCs were monitored for transduction efficiency, surface marker expression and cellular function. We were able to demonstrate that efficient gene transduction can be achieved with minimal ex vivo manipulation while maintaining the cellular function of transduced HSCs without serum or other reagents of animal origin. Conclusions/Significance: This study helps to better define factors relevant towards developing a standard clinical protocol for the delivery of SIN-LV into CD34 + cells

Regulatory T cells and their role in rheumatic diseases: a potential target for novel therapeutic development

Regulatory T cells have an important role in limiting immune reactions and are essential regulators of self-tolerance. Among them, CD4+CD25high regulatory T cells are the best-described subset. In this article, we summarize current knowledge on the phenotype, function, and development of CD4+CD25high regulatory T cells. We also review the literature on the role of these T cells in rheumatic diseases and discuss the potential for their use in immunotherapy

Using viral vectors as gene transfer tools (Cell Biology and Toxicology Special Issue: ETCS-UK 1 day meeting on genetic manipulation of cells)

In recent years, the development of powerful viral gene transfer techniques has greatly facilitated the study of gene function. This review summarises some of the viral delivery systems routinely used to mediate gene transfer into cell lines, primary cell cultures and in whole animal models. The systems described were originally discussed at a 1-day European Tissue Culture Society (ETCS-UK) workshop that was held at University College London on 1st April 2009. Recombinant-deficient viral vectors (viruses that are no longer able to replicate) are used to transduce dividing and post-mitotic cells, and they have been optimised to mediate regulatable, powerful, long-term and cell-specific expression. Hence, viral systems have become very widely used, especially in the field of neurobiology. This review introduces the main categories of viral vectors, focusing on their initial development and highlighting modifications and improvements made since their introduction. In particular, the use of specific promoters to restrict expression, translational enhancers and regulatory elements to boost expression from a single virion and the development of regulatable systems is described

Intraperitoneal drain placement and outcomes after elective colorectal surgery: international matched, prospective, cohort study

Despite current guidelines, intraperitoneal drain placement after elective colorectal surgery remains widespread. Drains were not associated with earlier detection of intraperitoneal collections, but were associated with prolonged hospital stay and increased risk of surgical-site infections.Background Many surgeons routinely place intraperitoneal drains after elective colorectal surgery. However, enhanced recovery after surgery guidelines recommend against their routine use owing to a lack of clear clinical benefit. This study aimed to describe international variation in intraperitoneal drain placement and the safety of this practice. Methods COMPASS (COMPlicAted intra-abdominal collectionS after colorectal Surgery) was a prospective, international, cohort study which enrolled consecutive adults undergoing elective colorectal surgery (February to March 2020). The primary outcome was the rate of intraperitoneal drain placement. Secondary outcomes included: rate and time to diagnosis of postoperative intraperitoneal collections; rate of surgical site infections (SSIs); time to discharge; and 30-day major postoperative complications (Clavien-Dindo grade at least III). After propensity score matching, multivariable logistic regression and Cox proportional hazards regression were used to estimate the independent association of the secondary outcomes with drain placement. Results Overall, 1805 patients from 22 countries were included (798 women, 44.2 per cent; median age 67.0 years). The drain insertion rate was 51.9 per cent (937 patients). After matching, drains were not associated with reduced rates (odds ratio (OR) 1.33, 95 per cent c.i. 0.79 to 2.23; P = 0.287) or earlier detection (hazard ratio (HR) 0.87, 0.33 to 2.31; P = 0.780) of collections. Although not associated with worse major postoperative complications (OR 1.09, 0.68 to 1.75; P = 0.709), drains were associated with delayed hospital discharge (HR 0.58, 0.52 to 0.66; P < 0.001) and an increased risk of SSIs (OR 2.47, 1.50 to 4.05; P < 0.001). Conclusion Intraperitoneal drain placement after elective colorectal surgery is not associated with earlier detection of postoperative collections, but prolongs hospital stay and increases SSI risk

Genetic diversity and population structure analysis of the tropical pasture grass Brachiaria humidicola based on microsatellites, cytogenetics, morphological traits, and geographical origin

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Brachiaria humidicola (Rendle) Schweick. is a warm-season grass commonly used as forage in the tropics. Accessions of this species were collected in eastern Africa and massively introduced into South America in the 1980s. Several of these accessions form a germplasm collection at the Brazilian Agricultural Research Corporation. However, apomixis, ploidy, and limited knowledge of the genetic basis of this germplasm collection have constrained breeding activities. The objectives of this work were to identify genetic variability in the Brazilian B. humidicola germplasm collection using microsatellite markers and to compare the results with information on the following: (1) collection sites of the accessions; (2) reproductive mode and ploidy levels; and (3) genetic diversity revealed by morphological traits. The evaluated germplasm population is highly structured into four major groups. The sole sexual accession did not group with any of the clusters. Genetic dissimilarities did not correlate with either geographic distances or genetic distances inferred from morphological descriptors. Additionally, the genetic structure identified in this collection did not correspond to differences in ploidy level. Alleles exclusive to either sexual or apomictic accessions were identified, suggesting that further evaluation of the association of these loci with apospory should be carried out.539698709Brazilian Agricultural Research Corporation (EMBRAPA) [02.05.2.10.00.02.01, 01.06.1.07.05.06.02]Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Foundation for Science and Technological Development of the State of Mato Grosso do Sul (FUNDECT) [41/100165/2004]Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Brazilian Agricultural Research Corporation (EMBRAPA) [02.05.2.10.00.02.01, 01.06.1.07.05.06.02]CNPq [482458/2007-0, 478262/2004-3, 307430/2007-3]FAPESP [2010/51010-0, 2007/57022-5, 2006/52953-8]Foundation for Science and Technological Development of the State of Mato Grosso do Sul (FUNDECT) [41/100165/2004

Monitorando indicadores de desempenho relacionados ao tempo de assistência da equipe de enfermagem Identificando indicadores de desempeño relativos al tiempo de asistencia del equipo de enfermería Monitoring performance indicators regarding the length of care by the nursing team

Este estudo descritivo objetivou: caracterizar unidades de clínica médica-cirúrgica de um hospital filantrópico, e a equipe de enfermagem lotada nestas unidades; identificar o perfil assistencial da clientela e investigar o tempo de assistência dispensado aos pacientes pela equipe de enfermagem. Os dados foram coletados a partir de documentos dos Departamentos de Pessoal e de Enfermagem, e aplicação de instrumento de classificação de pacientes. Para cálculo das horas de assistência, utilizou-se equação proposta pelo Compromisso com a Qualidade Hospitalar (CQH). Os achados evidenciaram relação de 1:1 profissional de enfermagem/leito e variação de 0,10 a 0,21 enfermeiros/leito. Os pacientes demandaram, predominantemente, cuidados mínimos (47,1% a 79,6%) e intermediários (17,7% a 38,6%) e o tempo médio dispensado pela equipe de enfermagem variou de 4,1 a 5,1 horas. Os valores mostraram que as horas de assistência dispensadas pela equipe de enfermagem estavam insuficientes para atender à complexidade assistencial dos pacientes.<br>Este estudio descriptivo tuvo como objetivos: caracterizar unidades de clínica médico-quirúrgica de un hospital filantrópico y el equipo de enfermería asignado a esas unidades; identificar el perfil asistencial de los pacientes e investigar el tiempo de asistencia dispensado a los pacientes por el equipo de enfermería. Los datos fueron recopilados a partir de documentos de los Departamentos de Personal y de Enfermería, y de la aplicación de un instrumento de clasificación de pacientes. Para el cálculo de las horas de asistencia se utilizó la ecuación propuesta por el Compromiso con la Calidad Hospitalaria (CCH). Los resultados evidenciaron relación de 1:1 profesional de enfermería/lecho y variación de 0,10 a 0,21 enfermeros/lecho. Los pacientes necesitaron, predominantemente, cuidados mínimos (47,1% a 79,6%) e intermedios (17,7% a 38,6%) y el promedio de tiempo dispensado por el equipo de enfermería varió entre 4,1 a 5,1 horas. Los valores mostraron que las horas de asistencia dispensadas por el equipo de enfermería eran insuficientes para atender a la complejidad asistencial de los pacientes.<br>The objectives of the present study were to: characterize medical-surgical clinic units of a philanthropic hospital and the nursing team performing in these units; identify the clientele's care profile, investigate the time length of care provided to the patients by the nursing team. Data were collected from documents of Personnel and Nursing Departments and the application of the classification instrument of patients. To calculate the care hours; the equation proposed by the Hospital Commitment Quality (HCQ) was used. The findings showed relationship of 1:1 nursing professional/bedside and variation from 0.10 to 0.21 nursing/bedside. The patients needed, mainly, minimum (47.1% to 79.6%) and intermediate (17.7% to 38.6%) care, and the average time spent by the nursing team ranged from 4.1 to 5.1 hours. The values showed that the care hours provided by the nursing team were not enough to assist to the patients' care complexity