1,999 research outputs found
Transferring; mapping; cloning of powdery mildew resistance gene of Haynaldia villosa and its utilization in common wheat
Physical mapping of a powdery mildew resistance related gene Hv-S/TPK by FISH with a TAC clone in wheat
Dissertação de mestrado integrado em Medicina (Hematologia), apresentado á Faculdade de Medicina da Universidade de Coimbra.A Policitemia Vera (PV) é uma doença clonal de etiologia desconhecida, na maior parte dos casos, que envolve a célula estaminal progenitora hematopoiética multipotencial. É uma neoplasia mieloproliferativa crónica (NMP) que se caracteriza pela expansão das três linhas celulares hematopoiéticas: eritróide, granulocítica e megacariocítica, com predomínio da primeira, de modo independente dos mecanismos normais de regulação da eritropoiese. Além disso, as células têm aspecto morfológico normal, a fibrose medular é pouco significativa e os níveis de eritropoietina (Epo) são habitualmente normais a baixos. Além da hipercelularidade medular com sobreprodução de uma ou de todas as linhas celulares, a doença cursa com hematopoiese extramedular, hiperviscosidade, propensão para complicações como trombose ou hemorragia e risco de desenvolvimento de mielofibrose ou transformação em leucemia aguda.
A descrição relativamente recente da associação de uma mutação no gene JAK2, localizado no cromosoma 9p24, com as doenças mieloproliferativas clássicas negativas para BCR-ABL, como a PV, veio permitir avanços significativos na compreensão da patofisiologia deste grupo de doenças hematológicas. A mutação provoca uma alteração do aminoácido V (valina) para F (fenilalanina) na posição 617 (JAK2V617F). De acordo com os dados publicados, a frequência da detecção da mutação JAK2V617F em doentes com PV é de cerca de 95%.
A proteína JAK2 é uma tirosina cinase citoplasmática, que se encontra associada ao domínio intracelular dos receptores de citocinas (como a Epo e trombopoietina - Tpo), e de factores de crescimento, essenciais para a função destes receptores. A mutação da JAK2 conduz à activação constitutiva dos receptores, independente da ligação à respectiva citocina e/ou hipersensibilidade a factores de crescimento, com consequente activação de múltiplas vias de sinalização intracelulares como a JAK/STAT (Janus Kinase/Signal Transductor and activator of transcription), a PI3K (fosfatidilinositol 3 cinase) e a MAPK (proteína cinase activadora de mitose), envolvidas na transformação e proliferação dos progenitores hematopoiéticos. Por outro lado, as células evidenciam alteração na diferenciação terminal e resistência à apoptose in vitro que poderá estar relacionada com o aumento da expressão da proteína anti-apoptótica Bcl-XL.
Além dos avanços no diagnóstico, a detecção da mutação JAK2V617F tem contribuido para melhorar a classificação e a terapêutica dos doentes com PV. Deste modo, o conhecimento dos mecanismos moleculares envolvidos na PV tem levado os investigadores à descoberta de novos fármacos dirigidos ao defeito molecular, permitindo novas abordagem terapêuticas mais eficazes e provavelmente de menor toxicidade.
Este trabalho procura fazer uma revisão sobre o actual conhecimento da caracterização molecular e clínica da PV e quais as suas implicações no diagnóstico e abordagem terapêutica desta NMP.Polycythemia Vera (PV) is a clonal disease of unknown etiology, which often involves the pluripotential hematopoietic stem cell. This disease integrates the family of chronic myeloproliferative neoplasm (MPN) and is characterized by the growth of the three hematopoietic celular lineages: granulocytic, megakaryocytic and erythroid, with predominance of the last one and regardless the normal mechanisms of erythropoiesis regulation. Moreover, cells have normal morphological aspect, bone marrow shows slight fibrosis and the levels of erythropoietin (Epo) usually vary from normal to low. Besides marrow hypercellularity with overproduction of one or all the celular lineages, the disease courses with extramedullary hematopoiesis, hyperviscosity, leading to complications such as thrombosis or bleeding and risk of transformation to myelofibrosis or acute leukemia.
Recently it has been described the association between the mutation in the JAK2 gene, located on chromosome 9p24, with the classic myeloproliferative disorders BCR-ABL negative, such as PV, which has brought significant advances in the understanding of the pathophysiology of this group of hematologic malignancies. The mutation causes a change of amino acid V (valine) to F (phenylalanine) at position 617 (JAK2V617F). According to published data, the frequency of JAK2V617F mutation detected in patients with PV is about 95%.
JAK2 protein is a cytoplasmic tyrosine kinase, which is associated to the intracelular domain of cytokine receptors, such as Epo and thrombopoietin (Tpo), and growth factors which are essential to the function of these receptors. JAK2 mutation leads to the constitutive receptors activation, independent of connection to their cytokine and / or hypersensitivity to growth factors, with consequent activation of multiple intracellular signaling pathways such as JAK / STAT (Janus Kinase / Signal transducer and transcription activator), the PI3K (phosphatidylinositol 3 kinase) and MAPK (Mitogen-activated protein), involved in the transformation and proliferation of hematopoietic progenitors. Moreover, the cells show changes in terminal differentiation and resistance to in vitro apoptosis which is possibly related to the increasing expression of anti-apoptotic protein Bcl-XL.
In addition to the advances in diagnosis, detection of JAK2V617F mutation has contributed to the improvement of classification and treatment in patients with PV. Thus, knowledge of the molecular mechanisms involved in PV has led investigators to the discovery of new drugs targeting molecular defects, allowing new therapeutic approach more efficient and probably less toxic.
The aim of this article is to review the current knowledge of clinical and molecular characterization of PV, and its implications on the diagnosis and therapeutic approach of this myeloproliferative disorder
Physical mapping of a powdery mildew resistance related gene Hv-S/TPK by FISH with a TAC clone in wheat
waviCGH: a web application for the analysis and visualization of genomic copy number alterations
waviCGH is a versatile web server for the analysis and comparison of genomic copy number alterations in multiple samples from any species. waviCGH processes data generated by high density SNP-arrays, array-CGH or copy-number calls generated by any technique. waviCGH includes methods for pre-processing of the data, segmentation, calling of gains and losses, and minimal common regions determination over a set of experiments. The server is a user-friendly interface to the analytical methods, with emphasis on results visualization in a genomic context. Analysis tools are introduced to the user as the different steps to follow in an experimental protocol. All the analysis steps generate high quality images and tables ready to be imported into spreadsheet programs. Additionally, for human, mouse and rat, altered regions are represented in a biological context by mapping them into chromosomes in an integrated cytogenetic browser. waviCGH is available at http://wavi.bioinfo.cnio.es
Development of a PNA Probe for Fluorescence In Situ Hybridization Detection of Prorocentrum donghaiense
Prorocentrum donghaiense is a common but dominant harmful algal bloom (HAB) species, which is widely distributed along the China Sea coast. Development of methods for rapid and precise identification and quantification is prerequisite for early-stage warning and monitoring of blooms due to P. donghaiense. In this study, sequences representing the partial large subunit rDNA (D1–D2), small subunit rDNA and internal transcribed spacer region (ITS-1, 5.8S rDNA and ITS-2) of P. donghaiense were firstly obtained, and then seven candidate DNA probes were designed for performing fluorescence in situ hybridization (FISH) tests on P. donghaiense. Based on the fluorescent intensity of P. donghaiense cells labeled by the DNA probes, the probe DP0443A displayed the best hybridization performance. Therefore, a PNA probe (PP0443A) analogous to DP0443A was used in the further study. The cells labeled with the PNA probe displayed more intensive green fluorescence than that labeled with its DNA analog. The PNA probe was used to hybridize with thirteen microalgae belonging to five families, i.e., Dinophyceae, Prymnesiophyceae, Raphidophyceae, Chlorophyceae and Bacillariophyceae, and showed no visible cross-reaction. Finally, FISH with the probes PP0443A and DP0443A and light microscopy (LM) analysis aiming at enumerating P. donghaiense cells were performed on the field samples. Statistical comparisons of the cell densities (cells/L) of P. donghaiense in the natural samples determined by FISH and LM were performed using one-way ANOVA and Duncan's multiple comparisons of the means. The P. donghaiense cell densities determined by LM and the PNA probe are remarkably higher than (p<0.05) that determined by the DNA probe, while no significant difference is observed between LM and the PNA probe. All results suggest that the PNA probe is more sensitive that its DNA analog, and therefore is promising for the monitoring of harmful algal blooms of P. donghaiense in the future
Anti-müllerian hormone is not associated with cardiometabolic risk factors in adolescent females
<p>Objectives: Epidemiological evidence for associations of Anti-Müllerian hormone (AMH) with cardiometabolic risk factors is lacking. Existing evidence comes from small studies in select adult populations, and findings are conflicting. We aimed to assess whether AMH is associated with cardiometabolic risk factors in a general population of adolescent females.</p>
<p>Methods: AMH, fasting insulin, glucose, HDLc, LDLc, triglycerides and C-reactive protein (CRP) were measured at a mean age 15.5 years in 1,308 female participants in the Avon Longitudinal Study of Parents and Children (ALSPAC). Multivariable linear regression was used to examine associations of AMH with these cardiometabolic outcomes.</p>
<p>Results: AMH values ranged from 0.16–35.84 ng/ml and median AMH was 3.57 ng/ml (IQR: 2.41, 5.49). For females classified as post-pubertal (n = 848) at the time of assessment median (IQR) AMH was 3.81 ng/ml (2.55, 5.82) compared with 3.25 ng/ml (2.23, 5.05) in those classed as early pubertal (n = 460, P≤0.001). After adjusting for birth weight, gestational age, pubertal stage, age, ethnicity, socioeconomic position, adiposity and use of hormonal contraceptives, there were no associations with any of the cardiometabolic outcomes. For example fasting insulin changed by 0% per doubling of AMH (95%CI: −3%,+2%) p = 0.70, with identical results if HOMA-IR was used. Results were similar after additional adjustment for smoking, physical activity and age at menarche, after exclusion of 3% of females with the highest AMH values, after excluding those that had not started menarche and after excluding those using hormonal contraceptives.</p>
<p>Conclusion: Our results suggest that in healthy adolescent females, AMH is not associated with cardiometabolic risk factors.</p>
The diacylglycerol kinase α/Atypical PKC/β1 integrin pathway in SDF-1α mammary carcinoma invasiveness
Diacylglycerol kinase α (DGKα), by phosphorylating diacylglycerol into phosphatidic acid, provides a key signal driving cell migration and matrix invasion. We previously demonstrated that in epithelial cells activation of DGKα activity promotes cytoskeletal remodeling and matrix invasion by recruiting atypical PKC at ruffling sites and by promoting RCP-mediated recycling of α5β1 integrin to the tip of pseudopods. In here we investigate the signaling pathway by which DGKα mediates SDF-1α-induced matrix invasion of MDA-MB-231 invasive breast carcinoma cells. Indeed we showed that, following SDF-1α stimulation, DGKα is activated and localized at cell protrusion, thus promoting their elongation and mediating SDF-1α induced MMP-9 metalloproteinase secretion and matrix invasion. Phosphatidic acid generated by DGKα promotes localization at cell protrusions of atypical PKCs which play an essential role downstream of DGKα by promoting Rac-mediated protrusion elongation and localized recruitment of β1 integrin and MMP-9. We finally demonstrate that activation of DGKα, atypical PKCs signaling and β1 integrin are all essential for MDA-MB-231 invasiveness. These data indicates the existence of a SDF-1α induced DGKα - atypical PKC - β1 integrin signaling pathway, which is essential for matrix invasion of carcinoma cells
Role of cellular senescence and NOX4-mediated oxidative stress in systemic sclerosis pathogenesis.
Systemic sclerosis (SSc) is a systemic autoimmune disease characterized by progressive fibrosis of skin and numerous internal organs and a severe fibroproliferative vasculopathy resulting frequently in severe disability and high mortality. Although the etiology of SSc is unknown and the detailed mechanisms responsible for the fibrotic process have not been fully elucidated, one important observation from a large US population study was the demonstration of a late onset of SSc with a peak incidence between 45 and 54 years of age in African-American females and between 65 and 74 years of age in white females. Although it is not appropriate to consider SSc as a disease of aging, the possibility that senescence changes in the cellular elements involved in its pathogenesis may play a role has not been thoroughly examined. The process of cellular senescence is extremely complex, and the mechanisms, molecular events, and signaling pathways involved have not been fully elucidated; however, there is strong evidence to support the concept that oxidative stress caused by the excessive generation of reactive oxygen species may be one important mechanism involved. On the other hand, numerous studies have implicated oxidative stress in SSc pathogenesis, thus, suggesting a plausible mechanism in which excessive oxidative stress induces cellular senescence and that the molecular events associated with this complex process play an important role in the fibrotic and fibroproliferative vasculopathy characteristic of SSc. Here, recent studies examining the role of cellular senescence and of oxidative stress in SSc pathogenesis will be reviewed
Acute tubulointerstitial nephritis complicating Legionnaires' disease: a case report
<p>Abstract</p> <p>Introduction</p> <p>Legionnaires' disease is recognized as a multi-systemic illness. Afflicted patients may have pulmonary, renal, gastrointestinal tract and central nervous system complications. However, renal insufficiency is uncommon. The spectrum of renal involvement may range from a mild and transient elevation of serum creatinine levels to anuric renal failure requiring dialysis and may be linked to several causes. In our present case report, we would like to draw attention to the importance of the pathological documentation of acute renal failure by reporting a case of a patient with acute tubulointerstitial nephritis complicating Legionnaires' disease.</p> <p>Case presentation</p> <p>A 55-year-old Caucasian man was admitted to our hospital for community-acquired pneumonia complicated by acute renal failure. <it>Legionella pneumophila </it>serogroup type 1 was diagnosed. Although the patient's respiratory illness responded to intravenous erythromycin and ofloxacin therapy, his renal failure worsened, he became anuric, and hemodialysis was started. A renal biopsy was performed, which revealed severe tubulointerstitial nephritis. After initiation of steroid therapy, his renal function improved dramatically.</p> <p>Conclusions</p> <p>This case highlights the importance of kidney biopsies in cases where acute renal failure is a complicating factor in Legionnaires' disease. If the presence of acute tubulointerstitial nephritis can be confirmed, it will likely respond favorably to steroidal treatment and thus irreversible renal damage and chronic renal failure will be avoided.</p
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