RNA metabolism is the primary target of formamide in vivo

The synthesis, processing and function of coding and non-coding RNA molecules and their interacting proteins has been the focus of a great deal of research that has boosted our understanding of key molecular pathways that underlie higher order events such as cell cycle control, development, innate immune response and the occurrence of genetic diseases. In this study, we have found that formamide preferentially weakens RNA related processes in vivo. Using a non-essential Schizosaccharomyces pombe gene deletion collection, we identify deleted loci that make cells sensitive to formamide. Sensitive deletions are significantly enriched in genes involved in RNA metabolism. Accordingly, we find that previously known temperature-sensitive splicing mutants become lethal in the presence of the drug under permissive temperature. Furthermore, in a wild type background, splicing efficiency is decreased and R-loop formation is increased in the presence of formamide. In addition, we have also isolated 35 formamide-sensitive mutants, many of which display remarkable morphology and cell cycle defects potentially unveiling new players in the regulation of these processes. We conclude that formamide preferentially targets RNA related processes in vivo, probably by relaxing RNA secondary structures and/or RNA-protein interactions, and can be used as an effective tool to characterize these processes

Abordagem da artroplastia total do joelho no Brasil: estudo transversal

CONTEXT AND OBJECTIVE: Total knee arthroplasty (TKA) has evolved particularly since the 1970s, with improvements in implants and surgical instruments, and has thus become an effective intervention for treating knee arthrosis. Many studies have presented rates of satisfactory clinical and radiological results greater than 90%, from follow-ups of over ten years. Nevertheless, despite scientific evidence showing the efficacy of TKA, the approaches taken present controversies in certain respects. The objective of this study was to evaluate how the Brazilian orthopedists deal with TKA, with investigation of the main aspects of this procedure. DESIGN AND SETTING: Cross-sectional survey conducted during the 39th Brazilian Congress of Orthopedics and Traumatology, in São Paulo, Brazil, in November 2007. METHODS: We applied a questionnaire to orthopedists registered at the congress. The questionnaire was randomly distributed and participation was voluntary; 858 completed questionnaires were included in the analysis. RESULTS: Most of the Brazilian orthopedists were members of SBOT and worked in the southeastern region. They used imported cemented implants through an anterior access route centered on the patella, with replacement of the joint surface of the patella and preservation of the posterior cruciate ligament. They did not have experience with simultaneous bilateral TKA. Postoperatively, they used antibiotics and suction drains for 48 hours. There was no consensus regarding prophylaxis for venous thromboembolism or the frequency of the main complications. CONCLUSION: The majority of Brazilian orthopedists work in the southeastern region of the country and agree about the main aspects of the approaches towards TKA.CONTEXTO E OBJETIVO: A artroplastia total do joelho (ATJ) evoluiu sobremaneira desde os anos 70, com melhora dos implantes e do instrumental cirúrgico, tornando-se uma intervenção efetiva para o tratamento da artrose do joelho. Muitos estudos apresentam resultados clínicos e radiológicos satisfatórios superiores a 90% no acompanhamento acima de 10 anos. Apesar das evidências científicas sobre sua eficácia da ATJ, a sua abordagem apresenta controvérsias em alguns aspectos. O objetivo do estudo foi avaliar como o ortopedista brasileiro aborda a ATJ e os principais aspectos técnicos na realização deste procedimento. TIPO DE ESTUDO E LOCAL: Estudo transversal, realizado durante o 39º Congresso Brasileiro de Ortopedia e Traumatologia em São Paulo, Brasil, em novembro de 2007. MÉTODOS: Aplicamos um questionário aos ortopedistas inscritos no congresso. A distribuição foi aleatória com adesão voluntária. Foram incluídos 858 questionários para análise. RESULTADOS: A maioria dos Ortopedistas Brasileiros são membros da SBOT e atua na região sudeste. Usam o implante importado, cimentado, por via de acesso anterior centrada na patela, com substituição da superfície articular da patela e preservação do ligamento cruzado posterior e não tem experiência com a artroplastia total bilateral simultânea. No pós-operatório utilizam antibióticos e dreno de sucção por 48 horas. Não houve consenso quanto à profilaxia para tromboembolismo venoso e frequência das principais complicações. CONCLUSÃO: A maioria dos ortopedistas brasileiros trabalha na região sudeste e concorda quanto aos principais aspectos da abordagem da ATJ.Universidade Federal de São Paulo (UNIFESP) Department of Orthopedics and TraumatologyUniversidade Federal de São Paulo (UNIFESP) Department of Orthopedics and Traumatology Orthopedist and Head of the Knee GroupUNIFESP, Department of Orthopedics and TraumatologyUNIFESP, Department of Orthopedics and Traumatology Orthopedist and Head of the Knee GroupSciEL

Connective Tissue Growth Factor in Regulation of RhoA Mediated Cytoskeletal Tension Associated Osteogenesis of Mouse Adipose-Derived Stromal Cells

Background: Cytoskeletal tension is an intracellular mechanism through which cells convert a mechanical signal into a biochemical response, including production of cytokines and activation of various signaling pathways. Methods/Principal Findings: Adipose-derived stromal cells (ASCs) were allowed to spread into large cells by seeding them at a low-density (1,250 cells/cm 2), which was observed to induce osteogenesis. Conversely, ASCs seeded at a high-density (25,000 cells/cm 2) featured small cells that promoted adipogenesis. RhoA and actin filaments were altered by changes in cell size. Blocking actin polymerization by Cytochalasin D influenced cytoskeletal tension and differentiation of ASCs. To understand the potential regulatory mechanisms leading to actin cytoskeletal tension, cDNA microarray was performed on large and small ASCs. Connective tissue growth factor (CTGF) was identified as a major regulator of osteogenesis associated with RhoA mediated cytoskeletal tension. Subsequently, knock-down of CTGF by siRNA in ASCs inhibited this osteogenesis. Conclusions/Significance: We conclude that CTGF is important in the regulation of cytoskeletal tension mediated AS

Deciphering the pathogenesis of tendinopathy: a three-stages process

Our understanding of the pathogenesis of "tendinopathy" is based on fragmented evidences like pieces of a jigsaw puzzle. We propose a "failed healing theory" to knit these fragments together, which can explain previous observations. We also propose that albeit "overuse injury" and other insidious "micro trauma" may well be primary triggers of the process, "tendinopathy" is not an "overuse injury" per se. The typical clinical, histological and biochemical presentation relates to a localized chronic pain condition which may lead to tendon rupture, the latter attributed to mechanical weakness. Characterization of pathological "tendinotic" tissues revealed coexistence of collagenolytic injuries and an active healing process, focal hypervascularity and tissue metaplasia. These observations suggest a failed healing process as response to a triggering injury. The pathogenesis of tendinopathy can be described as a three stage process: injury, failed healing and clinical presentation. It is likely that some of these "initial injuries" heal well and we speculate that predisposing intrinsic or extrinsic factors may be involved. The injury stage involves a progressive collagenolytic tendon injury. The failed healing stage mainly refers to prolonged activation and failed resolution of the normal healing process. Finally, the matrix disturbances, increased focal vascularity and abnormal cytokine profiles contribute to the clinical presentations of chronic tendon pain or rupture. With this integrative pathogenesis theory, we can relate the known manifestations of tendinopathy and point to the "missing links". This model may guide future research on tendinopathy, until we could ultimately decipher the complete pathogenesis process and provide better treatments

VEGF, VEGFR-1, and CTGF cell densities in tendon are increased-with cyclical loading: An in vivo tendinopathy model

Tendon injuries can occur in athletes and workers whose tasks involve repetitive, high-force hand activities, but the early pathophysiologic processes of tendinopathy are not well known. The purpose of this animal study was to evaluate the effects of cyclical tendon loading on the densities of cells producing growth factors such as vascular endothelial growth factor (VEGF), its receptor, vascular endothelial growth factor receptor 1 (VEGFR-1), and connective tissue growth factor (CTGF) in the Flexor Digitorum Profundus (FDP) tendon at the epicondyle. The FDP muscles of nine New Zealand rabbits were electrically stimulated to contract repetitively for 80 h of cumulative loading over 14 weeks. The contralateral limbs served as controls. The tendons at the medial epicondyle insertion sites were harvested, and sections were immunostained with antibodies directed against VEGF, VEGFR-1, or CTGF. Positive-staining cells were counted in six regions of interest: three along the enthesis, and three corresponding regions 1500 microns distal to the enthesis. VEGF (p = 0.0001), VEGFR-1 (p = 0.046), and CTGF (p = 0.0001) cell densities were increased in the tendon of the loaded limb compared to the nonloaded limb. In addition, regional differences in VEGF, VEGFR-1, and CTGF cell densities were found. VEGF, VEGFR-1, and CTGF are increased in tendon experiencing cyclical loading and may play a role in the early vascular changes in the progression to tendinosis