Long non-coding RNA CASC2 regulates osteoblasts matrix mineralization.

Long non-coding RNAs (lncRNAs) are master regulators of gene expression and have recently emerged as potential innovative therapeutic targets. The deregulation of lncRNA expression patterns has been associated with age-related and noncommunicable diseases in the bone tissue, including osteoporosis and tumors. However, the specific role of lncRNAs in physiological or pathological conditions in the bone tissue still needs to be further clarified, for their exploitation as therapeutic tools. In the present study, we evaluate the potential of the lncRNA CASC2 as a regulator of osteogenic differentiation and mineralization. Results show that CASC2 expression is decreased during osteogenic differentiation of human bone marrow-derived Mesenchymal Stem/Stromal cells (hMSCs). CASC2 knockdown, using small interfering RNA against CASC2 (siCASC2), increases the expression of the late osteogenic marker Bone Sialoprotein (BSP), but does not impact ALP staining level nor the expression of early osteogenic transcripts, including RUNX2 and OPG. Although siCASC2 does not impact hMSC proliferation nor apoptosis, it promotes the mineralization of hMSC cultured under osteogenic-inducing conditions, as shown by the increase of calcium deposits. Mass spectrometry-based proteomic analysis revealed that 89 proteins are regulated by CASC2 at late osteogenic stages, including proteins associated with bone diseases or anthropometric and musculoskeletal traits. Specifically, the Cartilage Oligomeric Matrix Protein (COMP) is highly enhanced by CASC2 knockdown at late stages of osteogenic differentiation, at both transcriptional and protein level. On the other hand, inhibition of COMP impairs osteoblasts mineralization as well as the expression of BSP. The results indicate that lncRNA CASC2 regulates late osteogenic differentiation and mineralization in hMSC via COMP and BSP. In conclusion, this study suggests that targeting lncRNA CASC2 could be a potential approach for modulating bone mineralization.

Circulating microRNAs Correlate with Multiple Myeloma and Skeletal Osteolytic Lesions.

Multiple myeloma (MM) is the second most frequent hematological disease and can cause skeletal osteolytic lesions. This study aims to evaluate the expression of circulating microRNAs (miRNAs) in MM patients and to correlate those levels with clinicopathological features, including bone lesions. A panel of miRNAs associated with MM onset and progression, or with bone remodeling, was analyzed in the plasma of 82 subjects (47 MM patients; 35 healthy controls). Results show that miR-16-5p, miR-20a-5p, and miR-21-5p are differently expressed between MM patients and healthy controls. Receiver operating characteristic analyses indicate that their combined expression has potential as a molecular marker (Area Under the Curve, AUC of 0.8249). Furthermore, significant correlations were found between the analyzed miRNAs and disease stage, treatment, ß2 microglobulin, serum albumin and creatinine levels, but not with calcium levels or genetic alterations. In this cohort, 65.96% of MM patients had bone lesions, the majority of which were in the vertebrae. Additionally, miR-29c-3p was decreased in patients with osteolytic lesions compared with patients without bone disease. Interestingly, circulating levels of miR-29b-3p correlated with cervical and thoracic vertebral lesions, while miR-195-5p correlated with thoracic lesions. Our findings suggest circulating miRNAs can be promising biomarkers for MM diagnosis and that their levels correlate with myeloma bone disease and osteolytic lesions.

Targeting p53 for Melanoma Treatment: Counteracting Tumour Proliferation, Dissemination and Therapeutic Resistance.

Melanoma is the deadliest form of skin cancer, primarily due to its high metastatic propensity and therapeutic resistance in advanced stages. The frequent inactivation of the p53 tumour suppressor protein in melanomagenesis may predict promising outcomes for p53 activators in melanoma therapy. Herein, we aimed to investigate the antitumor potential of the p53-activating agent SLMP53-2 against melanoma. Two- and three-dimensional cell cultures and xenograft mouse models were used to unveil the antitumor activity and the underlying molecular mechanism of SLMP53-2 in melanoma. SLMP53-2 inhibited the growth of human melanoma cells in a p53-dependent manner through induction of cell cycle arrest and apoptosis. Notably, SLMP53-2 induced p53 stabilization by disrupting the p53-MDM2 interaction, enhancing p53 transcriptional activity. It also promoted the expression of p53-regulated microRNAs (miRNAs), including miR-145 and miR-23a. Moreover, it displayed anti-invasive and antimigratory properties in melanoma cells by inhibiting the epithelial-to-mesenchymal transition (EMT), angiogenesis and extracellular lactate production. Importantly, SLMP53-2 did not induce resistance in melanoma cells. Additionally, it synergized with vemurafenib, dacarbazine and cisplatin, and resensitized vemurafenib-resistant cells. SLMP53-2 also exhibited antitumor activity in human melanoma xenograft mouse models by repressing cell proliferation and EMT while stimulating apoptosis. This work discloses the p53-activating agent SLMP53-2 which has promising therapeutic potential in advanced melanoma, either as a single agent or in combination therapy. By targeting p53, SLMP53-2 may counteract major features of melanoma aggressiveness.

TNF-alpha-induced microglia activation requires miR-342: impact on NF-kB signaling and neurotoxicity.

Growing evidences suggest that sustained neuroinflammation, caused by microglia overactivation, is implicated in the development and aggravation of several neurological and psychiatric disorders. In some pathological conditions, microglia produce increased levels of cytotoxic and inflammatory mediators, such as tumor necrosis factor alpha (TNF-α), which can reactivate microglia in a positive feedback mechanism. However, specific molecular mediators that can be effectively targeted to control TNF-α-mediated microglia overactivation, are yet to be uncovered. In this context, we aim to identify novel TNF-α-mediated micro(mi)RNAs and to dissect their roles in microglia activation, as well as to explore their impact on the cellular communication with neurons. A miRNA microarray, followed by RT-qPCR validation, was performed on TNF-α-stimulated primary rat microglia. Gain- and loss-of-function in vitro assays and proteomic analysis were used to dissect the role of miR-342 in microglia activation. Co-cultures of microglia with hippocampal neurons, using a microfluidic system, were performed to understand the impact on neurotoxicity. Stimulation of primary rat microglia with TNF-α led to an upregulation of Nos2, Tnf, and Il1b mRNAs. In addition, ph-NF-kB p65 levels were also increased. miRNA microarray analysis followed by RT-qPCR validation revealed that TNF-α stimulation induced the upregulation of miR-342. Interestingly, miR-342 overexpression in N9 microglia was sufficient to activate the NF-kB pathway by inhibiting BAG-1, leading to increased secretion of TNF-α and IL-1ß. Conversely, miR-342 inhibition led to a strong decrease in the levels of these cytokines after TNF-α activation. In fact, both TNF-α-stimulated and miR-342-overexpressing microglia drastically affected neuron viability. Remarkably, increased levels of nitrites were detected in the supernatants of these co-cultures. Globally, our findings show that miR-342 is a crucial mediator of TNF-α-mediated microglia activation and a potential target to tackle microglia-driven neuroinflammation.

miR-99a in bone homeostasis: Regulating osteogenic lineage commitment and osteoclast differentiation.

BACKGROUND: The tight coupling between osteoblasts and osteoclasts is essential to maintain bone homeostasis. Deregulation of this process leads to loss and deterioration of the bone tissue causing diseases, such as osteoporosis. MicroRNAs are able to control bone-related mechanisms and have been explored as therapeutic tools. In this study, we investigated the potential of miR-99a-5p to modulate osteogenic differentiation, osteoclastogenesis, and the osteoblasts-osteoclasts crosstalk. METHODS: To achieve this goal, human primary Mesenchymal Stem/Stromal Cells (MSC) were differentiated into osteoblasts and adipocytes, and miR-99a-5p expression was evaluated by RT-qPCR. Knockdown and overexpression experiments were conducted to modulate miR-99a-5p expression in MC3T3 cells. Cell proliferation and cell death/apoptosis were evaluated by resazurin assay and flow cytometry, respectively. Proteomic analysis was used to identify the miR-99a-5p regulatory network, and ELISA to evaluate OPG levels in the cell culture supernatant. Conditioned media from MC3T3-transfected cells was used to culture RAW 264.7 cells and the effect on osteoclast differentiation was assessed. Human primary monocytes were isolated to induce osteoclastogenesis and evaluate miR-99a-5p expression. Finally, levels of miR-99a-5p were modulated in RAW 264.7 cells to understand the impact on osteoclastogenesis. RESULTS: The results show that miR-99a-5p is significantly downregulated during the early stages of human primary MSCs osteogenic differentiation and during MC3T3 osteogenic differentiation. On the other hand, miR-99a-5p levels are increased during the initial stages of adipogenic differentiation. Inhibition of miR-99a-5p in MC3T3 pre-osteoblastic cells promoted osteogenic differentiation, whereas its overexpression suppressed the levels of osteogenic specific genes (Runx2 and Alpl), as well as mineralization, with no effect on proliferation or apoptosis. Proteomic analysis of miR-99a-5p-transfected cells showed that numerous proteins known to be involved in cell differentiation were altered, including osteogenic differentiation markers and extracellular matrix proteins. While inhibition of miR-99a-5p increased the Tnfrsf11b (OPG encoding gene)/Tnfsf11 (RANKL encoding gene) mRNA expression ratio, in addition to increasing OPG secretion, miR-99a-5p overexpression resulted in the opposite effect. The cell culture supernatant of miR-99a-5p-inhibited MC3T3 cells impaired the osteoclastogenic potential of RAW 264.7 cells by decreasing the number of multinucleated cells and reducing the expression of osteoclastogenic markers. Interestingly, miR-99a-5p expression is increased during osteoclasts differentiation, both in human primary monocytes and RAW 264.7. These results show that miR-99a-5p per se is a positive regulator of osteoclastogenic differentiation. CONCLUSIONS: Globally, our findings show that miR-99a-5p inhibition promotes the commitment into osteogenic differentiation, impairs osteoclastogenic differentiation, and control bone cells communication. Ultimately, it supports miR-99a-5p as a target candidate for future miRNA-based therapies for bone diseases associated with bone remodeling deregulation.

The Systemic Immune Response to Collagen-Induced Arthritis and the Impact of Bone Injury in Inflammatory Conditions.

Rheumatoid arthritis (RA) is a systemic disease that affects the osteoarticular system, associated with bone fragility and increased risk of fractures. Herein, we aimed to characterize the systemic impact of the rat collagen-induced arthritis (CIA) model and explore its combination with femoral bone defect (FD). The impact of CIA on endogenous mesenchymal stem/stromal cells (MSC) was also investigated. CIA induction led to enlarged, more proliferative, spleen and draining lymph nodes, with altered proportion of lymphoid populations. Upon FD, CIA animals increased the systemic myeloid cell proportions, and their expression of co-stimulatory molecules CD40 and CD86. Screening plasma cytokine/chemokine levels showed increased tumor necrosis factor-α (TNF-α), Interleukin (IL)-17, IL-4, IL-5, and IL-12 in CIA, and IL-2 and IL-6 increased in CIA and CIA+FD, while Fractalkine and Leptin were decreased in both groups. CIA-derived MSC showed lower metabolic activity and proliferation, and significantly increased osteogenic and chondrogenic differentiation markers. Exposure of control-MSC to TNF-α partially mimicked the CIA-MSC phenotype in vitro. In conclusion, inflammatory conditions of CIA led to alterations in systemic immune cell proportions, circulating mediators, and in endogenous MSC. CIA animals respond to FD, and the combined model can be used to study the mechanisms of bone repair in inflammatory conditions.

SLMP53-2 Restores Wild-Type-Like Function to Mutant p53 through Hsp70: Promising Activity in Hepatocellular Carcinoma.

Half of human cancers harbor TP53 mutations that render p53 inactive as a tumor suppressor. In these cancers, reactivation of mutant p53 (mutp53) through restoration of wild-type-like function constitutes a valuable anticancer therapeutic strategy. In order to search for mutp53 reactivators, a small library of tryptophanol-derived oxazoloisoindolinones was synthesized and the potential of these compounds as mutp53 reactivators and anticancer agents was investigated in human tumor cells and xenograft mouse models. By analysis of their anti-proliferative effect on a panel of p53-null NCI-H1299 tumor cells ectopically expressing highly prevalent mutp53, the compound SLMP53-2 was selected based on its potential reactivation of multiple structural mutp53. In mutp53-Y220C-expressing hepatocellular carcinoma (HCC) cells, SLMP53-2-induced growth inhibition was mediated by cell cycle arrest, apoptosis, and endoplasmic reticulum stress response. In these cells, SLMP53-2 restored wild-type-like conformation and DNA-binding ability of mutp53-Y220C by enhancing its interaction with the heat shock protein 70 (Hsp70), leading to the reestablishment of p53 transcriptional activity. Additionally, SLMP53-2 displayed synergistic effect with sorafenib, the only approved therapy for advanced HCC. Notably, it exhibited potent antitumor activity in human HCC xenograft mouse models with a favorable toxicological profile. Collectively, SLMP53-2 is a new mutp53-targeting agent with promising antitumor activity, particularly against HCC.

Genetically Engineered-MSC Therapies for Non-unions, Delayed Unions and Critical-size Bone Defects.

The normal bone regeneration process is a complex and coordinated series of events involving different cell types and molecules. However, this process is impaired in critical-size/large bone defects, with non-unions or delayed unions remaining a major clinical problem. Novel strategies are needed to aid the current therapeutic approaches. Mesenchymal stem/stromal cells (MSCs) are able to promote bone regeneration. Their beneficial effects can be improved by modulating the expression levels of specific genes with the purpose of stimulating MSC proliferation, osteogenic differentiation or their immunomodulatory capacity. In this context, the genetic engineering of MSCs is expected to further enhance their pro-regenerative properties and accelerate bone healing. Herein, we review the most promising molecular candidates (protein-coding and non-coding transcripts) and discuss the different methodologies to engineer and deliver MSCs, mainly focusing on in vivo animal studies. Considering the potential of the MSC secretome for bone repair, this topic has also been addressed. Furthermore, the promising results of clinical studies using MSC for bone regeneration are discussed. Finally, we debate the advantages and limitations of using MSCs, or genetically-engineered MSCs, and their potential as promoters of bone fracture regeneration/repair.

S-MiRAGE: A Quantitative, Secreted RNA-Based Reporter of Gene Expression and Cell Persistence.

Nondestructive measurements of cell persistence and gene expression are crucial for longitudinal research studies and for prognostic assessment of cell therapies. Here we describe S-MiRAGE, a platform that utilizes small secreted RNA molecules as sensitive and quantitatively accurate reporters of cellular processes. S-MiRAGE allows cellular numbers or gene expression to be measured from culture media or from biofluids. We show that multiple S-MiRAGE reporters can be multiplexed, and demonstrate the utility of S-MiRAGE by monitoring the differentiation status of human embryonic stem cells in vitro and tumor growth in a mouse model in vivo.

Cancer-associated rs6983267 SNP and its accompanying long noncoding RNA CCAT2 induce myeloid malignancies via unique SNP-specific RNA mutations.

The cancer-risk-associated rs6983267 single nucleotide polymorphism (SNP) and the accompanying long noncoding RNA CCAT2 in the highly amplified 8q24.21 region have been implicated in cancer predisposition, although causality has not been established. Here, using allele-specific CCAT2 transgenic mice, we demonstrate that CCAT2 overexpression leads to spontaneous myeloid malignancies. We further identified that CCAT2 is overexpressed in bone marrow and peripheral blood of myelodysplastic/myeloproliferative neoplasms (MDS/MPN) patients. CCAT2 induces global deregulation of gene expression by down-regulating EZH2 in vitro and in vivo in an allele-specific manner. We also identified a novel non-APOBEC, non-ADAR, RNA editing at the SNP locus in MDS/MPN patients and CCAT2-transgenic mice. The RNA transcribed from the SNP locus in malignant hematopoietic cells have different allelic composition from the corresponding genomic DNA, a phenomenon rarely observed in normal cells. Our findings provide fundamental insights into the functional role of rs6983267 SNP and CCAT2 in myeloid malignancies.

miR-195 inhibits macrophages pro-inflammatory profile and impacts the crosstalk with smooth muscle cells.

Macrophages are a main component of atherosclerotic plaques. Recent studies suggest that pro-inflammatory M1 macrophages are pro-atherogenic while M2 macrophages promote plaque stability. Moreover, toll-like receptor signalling pathways are implicated in atherosclerotic plaque formation, evolution and regression. We propose microRNAs as key regulators of these processes. In this context, our goal is to promote inflammation resolution using miR-195 to reduce M1-like macrophage polarization and to evaluate the molecular mechanisms underlying such effect, as well as to explore the functional consequences for smooth muscle cell recruitment. Human primary macrophages were differentiated from peripheral blood monocytes and stimulated with LPS or IL-10 to promote M1 or M2c polarization, respectively. miR-195 levels were upregulated in M2c macrophages compared with M1 macrophages. In THP-1 macrophages stimulated with LPS and IFN-γ, results show that TLR2 levels were reduced by miR-195 overexpression compared with scrambled control. In addition, phosphorylated forms of p54 JNK, p46 JNK and p38 MAPK were decreased by miR-195 in macrophages following M1 stimulation. Moreover, miR-195 significantly decreased levels of IL-1ß, IL-6 and TNF-α pro-inflammatory cytokines in the supernatants of M1-stimulated macrophage cultures. At the functional level, results from smooth muscle cell recruitment and migration models showed that miR-195 impairs the capacity of M1 macrophages to promote smooth muscle cells migration. In conclusion, miR-195 is involved in macrophage polarization and inhibits TLR2 inflammatory pathway mediators. Moreover, miR-195 impairs the effect of macrophages on smooth muscle cells recruitment capacity and migration profile. Thus, miR-195 might be used as a new potential tool to promote inflammation resolution in cardiovascular research.

Systemic Delivery of Bone Marrow Mesenchymal Stem Cells for In Situ Intervertebral Disc Regeneration.

Cell therapies for intervertebral disc (IVD) regeneration presently rely on transplantation of IVD cells or stem cells directly to the lesion site. Still, the harsh IVD environment, with low irrigation and high mechanical stress, challenges cell administration and survival. In this study, we addressed systemic transplantation of allogeneic bone marrow mesenchymal stem cells (MSCs) intravenously into a rat IVD lesion model, exploring tissue regeneration via cell signaling to the lesion site. MSC transplantation was performed 24 hours after injury, in parallel with dermal fibroblasts as a control; 2 weeks after transplantation, animals were killed. Disc height index and histological grading score indicated less degeneration for the MSC-transplanted group, with no significant changes in extracellular matrix composition. Remarkably, MSC transplantation resulted in local downregulation of the hypoxia responsive GLUT-1 and in significantly less herniation, with higher amounts of Pax5+ B lymphocytes and no alterations in CD68+ macrophages within the hernia. The systemic immune response was analyzed in the blood, draining lymph nodes, and spleen by flow cytometry and in the plasma by cytokine array. Results suggest an immunoregulatory effect in the MSC-transplanted animals compared with control groups, with an increase in MHC class II+ and CD4+ cells, and also upregulation of the cytokines IL-2, IL-4, IL-6, and IL-10, and downregulation of the cytokines IL-13 and TNF-α. Overall, our results indicate a beneficial effect of systemically transplanted MSCs on in situ IVD regeneration and highlight the complex interplay between stromal cells and cells of the immune system in achieving successful tissue regeneration. Stem Cells Translational Medicine 2017;6:1029-1039.

The miR-143/miR-145 cluster and the tumor microenvironment: unexpected roles.

miR-143 and miR-145 have been widely described as tumor suppressors. Recent findings suggest these microRNAs (miRNAs) have a critical role that affects the stroma rather than epithelial cells. Understanding the part played by the miR-143/miR-145 cluster in the tumor microenvironment is essential for the development of future miRNA-related therapies.

miR-195 in human primary mesenchymal stromal/stem cells regulates proliferation, osteogenesis and paracrine effect on angiogenesis.

Mesenchymal Stromal/Stem Cells (MSC) are currently being explored in diverse clinical applications, including regenerative therapies. Their contribution to regeneration of bone fractures is dependent on their capacity to proliferate, undergo osteogenesis and induce angiogenesis. This study aimed to uncover microRNAs capable of concomitantly regulate these mechanisms. Following microRNA array results, we identified miR-195 and miR-497 as downregulated in human primary MSC under osteogenic differentiation. Overexpression of miR-195 or miR-497 in human primary MSC leads to a decrease in osteogenic differentiation and proliferation rate. Conversely, inhibition of miR-195 increased alkaline phosphatase expression and activity and cells proliferation. Then, miR-195 was used to study MSC capacity to recruit blood vessels in vivo. We provide evidence that the paracrine effect of MSC on angiogenesis is diminishedwhen cells over-express miR-195. VEGF may partially mediate this effect, as its expression and secreted protein levels are reduced by miR-195, while increased by anti-miR-195, in human MSC. Luciferase reporter assays revealed a direct interaction between miR-195 and VEGF 3´-UTR in bone cancer cells. In conclusion, our results suggest that miR-195 regulates important mechanisms for bone regeneration, specifically MSC osteogenic differentiation, proliferation and control of angiogenesis; therefore, it is a potential target for clinical bone regenerative therapies.

Auto preenchimento da ficha clínica no rastreamento do câncer de colo de útero: percepções da mulher / Auto fill in clinical sheet no trace of cervical cancer: women's perceptions

Objetivo: descrever a percepção das mulheres frente ao auto preenchimento de um novo modelo de ficha clínica da consulta de enfermagem no controle do câncer do colo de útero. Método: Estudo qualitativo, com 20 mulheres, em uma unidade de atenção primária à saúde de um município da Zona da Mata Mineira em 2012 e 2013. Resultados: O estudo revelou a adesão de 100% de autopreenchimento da ficha clínica. A idade das participantes variou de 20 a 67 anos, com média de 43 anos, 60% destas eram casadas. Grande maioria relatou uma percepção positiva em relação ao autopreenchimento da ficha clínica, tendo esta adesão de 100%. Conclusão: Devido à grande aceitação do autopreenchimento da ficha clínica na consulta de rastreamento do câncer do colo do útero, esta se tornou uma estratégia para estimular a adesão a este atendimento. O que demonstra a necessidade de incorporar novas práticas para aumentar a adesão das mulheres que não realizam esta consulta, seja por aspectos subjetivos relacionados a sentimentos negativos ou devido a exposição da sua história, do seu corpo e de sua vida sexual...

Objective: to describe the perception of women regarding a new self-reported medical record model for nurse appointments for cervical cancer control. Method: qualitative study with 20 women in a primary health care unit in a town in Zona da Mata, Minas Gerais, Brazil, from 2012 to 2013. Results: the study revealed a 100% adoption of self-reported medical records. The age of participants ranged from 20 to 67 years, mean 43 years, 60% of them married. A vast majority reported a positive perception of self-reporting in medical records, with a 100% adherence. Conclusion: Due to the high acceptance of self-reporting in medical records at cervical cancer screening appointments, it has become a strategy to encourage the adoption of this service. This demonstrates the need to incorporate new practices to increase the participation of women who do not seek cervical cancer screening appointments, either due to subjective aspects related to negative feelings or because of exposure of their history, body, and sex life. Descriptors: Clinical record; Uterine cervical neoplasms; Women's health; Primary care nursing...

Objetivo: describir la percepción de las mujeres frente al relleno automático de un nuevo modelo de ficha clínica de consulta de enfermería en el control del cáncer cervicouterino. Método: estudio cualitativo con 20 mujeres en una unidad de atención primaria a la salud en un municipio de la Zona da Mata de Minas Gerais, 2012 a 2013. Resultados: el estudio reveló la adhesión del 100% al relleno automático de las fichas clínicas. La edad de las participantes estuvo entre 20 a 67 años, con el promedio de 43 años, el 60% de ellas eran casadas. La inmensa mayoría reportó una percepción positiva en relación al relleno automático de las fichas clínicas, con adhesión del 100%. Conclusión: Debido a la gran aceptación del relleno automático de la ficha clínica en la consulta de rastreo del cáncer de cuello uterino, esto se ha convertido en una estrategia para fomentar la adhesión a este servicio. Esto demuestra la necesidad de incorporar nuevas prácticas para aumentar la adhesión de las mujeres que no llevan a cabo esta consulta, ya sea por aspectos subjetivos relacionados con los sentimientos negativos o debido a la exposición de su historia, su cuerpo y su vida sexual.Descriptores: Ficha clínica; Neoplasias del cuello uterino; Salud de la mujer; Enfermería de atención primaria...

Conduta de profissionais durante a consulta de rastreio do câncer de mama e útero / Professional conduct during breast and uterine/cervical cancer screening consultations / Conducta de profesionales durante consulta de rastreo del cáncer de mama y útero

Objetivou-se avaliar a conduta de profissionais quanto às ações de anamnese e exame físico realizadas na consulta de rastreio de câncer de mama e de colo de útero na atenção primária à saúde. Estudo transversal, descritivo, de análise quantitativa realizado em um município de Minas Gerais, MG, Brasil. Para avaliação, aplicou-se um questionário estruturado às mulheres que compareceram à consulta com profissional enfermeiro ou médico, no período de dezembro 2012 a janeiro de 2013. O instrumento de pesquisa, contém perguntas sociodemográficas e referentes às ações de anmnese, exame físico e ginecológico, conforme recomendações do Ministério da Saúde. Foram entrevistadas cinquenta mulheres, com média de 40 anos de idade. Os resultados evidenciaram ações de anamnese, exame físico e ginecológico incompletos em todas as consultas. Para promoção integral da atenção à saúde da mulher faz-se necessário o cumprimento das recomendações do Ministério da Saúde.

Aimed to evaluate the behavior of professionals regarding checking medical history (anamneses) and the physical examinationperformed in screening consultation for breast and cervical cancer in primary health care. This is a descriptivecross-sectional study of quantitative analysis, and was conducted in Minas Gerais, MG, Brazil. For evaluation purposes, astructured questionnaire was answered by women who attended a consultation with nurses or doctors, from December2012 to January 2013. The research instrument contained sociodemographic questions and questions related to medicalhistory (anamneses), physical and gynecological examination, as recommended by the Ministry of Health. Fifty women wereinterviewed, averaging 40 years of age. The results showed incomplete anamnesis, physical and gynecological examinationin all consultations. For full promotion of women’s health attention, it is necessary to comply with Ministry of Health recommendations.

El objetivo fue evaluar la conducta de profesionales cuanto a las acciones de anamnesis y examen físico realizados en consultade rastreo de cáncer de mama y de cuello uterino en la atención primaria a la salud. Estudio transversal, descriptivo,cuantitativo realizado en una ciudad en Minas Gerais, MG, Brasil. Para evaluación, se aplicó cuestionario estructurado a lasmujeres que fueron a la consulta con el profesional enfermero o médico, de diciembre de 2012 a enero de 2013. El instrumentode investigación tuvo preguntas sociodemográficas y de acciones de anmnesis, examen físico y ginecológico, segúnrecomendación del Ministerio de la Salud. Cincuenta mujeres fueron entrevistadas, con promedia de 40 años de edad. Losresultados señalaron acciones de anamnesis, examen físico y ginecológico incompletos en todas las consultas. Para promocióntotal de la atención de salud de la mujer, es necesario cumplir con las recomendaciones del Ministerio de la Salud.

Therapeutic synergy between microRNA and siRNA in ovarian cancer treatment.

UNLABELLED: Development of improved RNA interference-based strategies is of utmost clinical importance. Although siRNA-mediated silencing of EphA2, an ovarian cancer oncogene, results in reduction of tumor growth, we present evidence that additional inhibition of EphA2 by a microRNA (miRNA) further "boosts" its antitumor effects. We identified miR-520d-3p as a tumor suppressor upstream of EphA2, whose expression correlated with favorable outcomes in two independent patient cohorts comprising 647 patients. Restoration of miR-520d-3p prominently decreased EphA2 protein levels, and suppressed tumor growth and migration/invasion both in vitro and in vivo. Dual inhibition of EphA2 in vivo using 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine (DOPC) nanoliposomes loaded with miR-520d-3p and EphA2 siRNA showed synergistic antitumor efficiency and greater therapeutic efficacy than either monotherapy alone. This synergy is at least in part due to miR-520d-3p targeting EphB2, another Eph receptor. Our data emphasize the feasibility of combined miRNA-siRNA therapy, and will have broad implications for innovative gene silencing therapies for cancer and other diseases. SIGNIFICANCE: This study addresses a new concept of RNA inhibition therapy by combining miRNA and siRNA in nanoliposomal particles to target oncogenic pathways altered in ovarian cancer. Combined targeting of the Eph pathway using EphA2-targeting siRNA and the tumor suppressor miR-520d-3p exhibits remarkable therapeutic synergy and enhanced tumor suppression in vitro and in vivo compared with either monotherapy alone.

O cuidado de enfermagem vivenciado por mulheres mastectomizadas / Nursing care experienced by women with mastectomies

O cuidado é a essência da enfermagem, ele atua em todas as ações gerenciais e assistenciais. O objeto deste estudo é "o cuidado de enfermagem na vivência das mulheres mastectomizadas", tendo como objetivos identificar e analisar a vivência do cuidado de enfermagem pelas mulheres submetidas à mastectomia. Para o desenvolvimento deste trabalho utilizou-se pesquisa qualitativa do tipo exploratório descritiva. A coleta de dados foi realizada através de entrevistas, utilizando um roteiro semi-estruturado. Os sujeitos do estudo foram nove mulheres na faixa etária a partir dos 35 anos de idade que fazem acompanhamento em um hospital universitário de ensino localizado na Zona da Mata Mineira. As informações foram analisadas, surgindo as seguintes categorias: 1) O cuidado indispensável; 2) A superioridade dos procedimentos técnicos na vivência do cuidado; 3) O "descuidado". Verificou-se que mesmo as mulheres que tinham um relato mais positivo do atendimento, não obtiveram um cuidado integral e humanizado pela equipe de enfermagem, tendo sido identificada a necessidade desse ser especializado a essas mulheres. Esses profissionais devem oferecer cuidado que agrupa técnica, ciência e humanização, fornecer todas as orientações, respeitar as necessidades e o nível de entendimento para, na perspectiva do autocuidado, reabilitar as mulheres mastectomizadas.

Detection of the Epstein-Barr virus in blood and bone marrow mononuclear cells of patients with aggressive B-cell non-Hodgkin's lymphoma is not associated with prognosis.

The Epstein-Barr virus (EBV) is associated with a large spectrum of lymphoproliferative diseases. Traditional methods of EBV detection include the immunohistochemical identification of viral proteins and DNA probes to the viral genome in tumoral tissue. The present study explored the detection of the EBV genome, using the BALF5 gene, in the bone marrow or blood mononuclear cells of patients with diffuse large B-cell lymphomas (DLBCL) and related its presence to the clinical variables and risk factors. The results show that EBV detection in 21.5% of patients is not associated with age, gender, staging, B symptoms, international prognostic index scores or any analytical parameters, including lactate dehydrogenase (LDH) or ß-2 microglobulin (B2M). The majority of patients were treated with R-CHOP-like (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisolone or an equivalent combination) and some with CHOP-like chemotherapy. Response rates [complete response (CR) + partial response (PR)] were not significantly different between EBV-negative and -positive cases, with 93.2 and 88.9%, respectively. The survival rate was also similar in the two groups, with 5-year overall survival (OS) rates of 64.3 and 76.7%, respectively. However, when analyzing the treatment groups separately there was a trend in EBV-positive patients for a worse prognosis in patients treated with CHOP-like regimens that was not identified in patients treated with R-CHOP-like regimens. We conclude that EBV detection in the bone marrow and blood mononuclear cells of DLBC patients has the same frequency of EBV detection on tumoral lymphoma tissue but is not associated with the risk factors, response rate and survival in patients treated mainly with immunochemotherapy plus rituximab. These results also suggest that the addition of rituximab to chemotherapy improves the prognosis associated with EBV detection in DLBCL.

Decoy activity through microRNAs: the therapeutic implications.

INTRODUCTION: microRNAs (miRNAs), small noncoding RNAs, are deregulated in several diseases including cancer. miRNAs regulate gene expression at a posttranscriptional level by binding to 5'UTR, coding regions or 3'UTR of messenger RNAs (mRNA), inhibiting mRNA translation or causing mRNA degradation. The same miRNA can have multiple mRNA targets, and the same mRNA can be regulated by various miRNAs. AREAS COVERED: Recently, seminal contributions by several groups have implicated miRNAs as components of an RNA-RNA language that involves cross-talk between competing endogenous RNAs through a decoy mechanism. We review the studies that described miRNAs as players in a biological decoy activity. miRNAs can either be trapped by competing endogenous RNAs or interact with proteins that have binding sites for mRNAs. EXPERT OPINION: The miRNA decoy functions have implications for the design of therapeutic approaches in human diseases, including specific ways to overcome resistance to drug therapy and future miRNA-based clinical trials design.