Ansiedade de separação materna e adaptação psicossocial ao pré-escolar

Diversos estudos parecem indicar a ansiedade de separação materna como um importante factor que influencia a adaptação psicossocial da criança e consequentemente o seu desenvolvimento social. No presente estudo pretendemos analisar e compreender a relação entre a ansiedade de separação materna e a qualidade da adaptação psicossocial de crianças em idade pré-escolar. Os participantes foram 168 crianças com idades compreendidas entre os 3 e os 4 anos e as suas respectivas mães. A Escala de Ansiedade Materna foi administrada às mães, permitindo identificar os sentimentos e atitudes maternas face à separação. As educadoras de infância preencheram o questionário Adaptação Psicossocial da Criança com vista à obtenção de dados relativos à qualidade da adaptação psicossocial das crianças. Os resultados indicam que a ansiedade de separação materna influencia a qualidade da adaptação psicossocial da criança. Assim, as crianças cujas mães foram identificadas como possuindo uma elevada ansiedade de separação mobilizaram mais comportamentos reveladores de uma baixa competência social. Esperamos com este estudo ter podido contribuir para uma melhor compreensão da relação entre a ansiedade de separação materna e a adaptação psicossocial da criança, bem como alertar para a importância da implementação de estratégias preventivas cada vez mais precoces e eficazes. ------ ABSTRACT ------ Several studies seem to indicate maternal separation anxiety as an important factor that influences the child psychosocial adaptation and consequently child social development. The purpose of the present study is to understand and analyse the relationship between the maternal separation anxiety and the quality of preschooler's psychosocial adaptation. This study included 168 children with ages between 3 and 4 years old. The Maternal Separation Anxiety Scale was administered to their mothers, allowing to identify the maternal feelings and attitudes concerning separation. Teachers filled out the Preschoolers Psychosocial Adaptation questionnaire in order to obtain information related to children psychosocial adaptation. The results indicated that maternal separation anxiety influences the quality of child psychosocial adaptation. Mothers with high maternal separation anxiety have children that used behaviors that reveled a low social competence

Molecular and Functional Characterization of Human Intestinal Organoids and Monolayers for Modeling Epithelial Barrier

BACKGROUND: Patient-derived organoid (PDO) models offer potential to transform drug discovery for inflammatory bowel disease (IBD) but are limited by inconsistencies with differentiation and functional characterization. We profiled molecular and cellular features across a range of intestinal organoid models and examined differentiation and establishment of a functional epithelial barrier. METHODS: Patient-derived organoids or monolayers were generated from control or IBD patient-derived colon or ileum and were molecularly or functionally profiled. Biological or technical replicates were examined for transcriptional responses under conditions of expansion or differentiation. Cell-type composition was determined by deconvolution of cell-associated gene signatures and histological features. Differentiated control or IBD-derived monolayers were examined for establishment of transepithelial electrical resistance (TEER), loss of barrier integrity in response to a cocktail of interferon (IFN)-γ and tumor necrosis factor (TNF)-α, and prevention of cytokine-induced barrier disruption by the JAK inhibitor, tofacitinib. RESULTS: In response to differentiation media, intestinal organoids and monolayers displayed gene expression patterns consistent with maturation of epithelial cell types found in the human gut. Upon differentiation, both colon- and ileum-derived monolayers formed functional barriers, with sustained TEER. Barrier integrity was compromised by inflammatory cytokines IFN-γ and TNF-α, and damage was inhibited in a dose-dependent manner by tofacitinib. CONCLUSIONS: We describe the generation and characterization of human colonic or ileal organoid models capable of functional differentiation to mature epithelial cell types. In monolayer culture, these cells formed a robust epithelial barrier with sustained TEER and responses to pharmacological modulation. Our findings demonstrate that control and IBD patient-derived organoids possess consistent transcriptional and functional profiles that can enable development of epithelial-targeted therapies

Organoids as a biomarker for personalized treatment in metastatic colorectal cancer: drug screen optimization and correlation with patient response

BACKGROUND: The inability to predict treatment response of colorectal cancer patients results in unnecessary toxicity, decreased efficacy and survival. Response testing on patient-derived organoids (PDOs) is a promising biomarker for treatment efficacy. The aim of this study is to optimize PDO drug screening methods for correlation with patient response and explore the potential to predict responses to standard chemotherapies. METHODS: We optimized drug screen methods on 5-11 PDOs per condition of the complete set of 23 PDOs from patients treated for metastatic colorectal cancer (mCRC). PDOs were exposed to 5-fluorouracil (5-FU), irinotecan- and oxaliplatin-based chemotherapy. We compared medium with and without N-acetylcysteine (NAC), different readouts and different combination treatment set-ups to capture the strongest association with patient response. We expanded the screens using the optimized methods for all PDOs. Organoid sensitivity was correlated to the patient's response, determined by % change in the size of target lesions. We assessed organoid sensitivity in relation to prior exposure to chemotherapy, mutational status and sidedness. RESULTS: Drug screen optimization involved excluding N-acetylcysteine from the medium and biphasic curve fitting for 5-FU & oxaliplatin combination screens. CellTiter-Glo measurements were comparable with CyQUANT and did not affect the correlation with patient response. Furthermore, the correlation improved with application of growth rate metrics, when 5-FU & oxaliplatin was screened in a ratio, and 5-FU & SN-38 using a fixed dose of SN-38. Area under the curve was the most robust drug response curve metric. After optimization, organoid and patient response showed a correlation coefficient of 0.58 for 5-FU (n = 6, 95% CI -0.44,0.95), 0.61 for irinotecan- (n = 10, 95% CI -0.03,0.90) and 0.60 for oxaliplatin-based chemotherapy (n = 11, 95% CI -0.01,0.88). Median progression-free survival of patients with resistant PDOs to oxaliplatin-based chemotherapy was significantly shorter than sensitive PDOs (3.3 vs 10.9 months, p = 0.007). Increased resistance to 5-FU in patients with prior exposure to 5-FU/capecitabine was adequately reflected in PDOs (p = 0.003). CONCLUSIONS: Our study emphasizes the critical impact of the screening methods for determining correlation between PDO drug screens and mCRC patient outcomes. Our 5-step optimization strategy provides a basis for future research on the clinical utility of PDO screens

Geolocation with respect to persona privacy for the Allergy Diary app - a MASK study

Background: Collecting data on the localization of users is a key issue for the MASK (Mobile Airways Sentinel network: the Allergy Diary) App. Data anonymization is a method of sanitization for privacy. The European Commission's Article 29 Working Party stated that geolocation information is personal data. To assess geolocation using the MASK method and to compare two anonymization methods in the MASK database to find an optimal privacy method. Methods: Geolocation was studied for all people who used the Allergy Diary App from December 2015 to November 2017 and who reported medical outcomes. Two different anonymization methods have been evaluated: Noise addition (randomization) and k-anonymity (generalization). Results: Ninety-three thousand one hundred and sixteen days of VAS were collected from 8535 users and 54,500 (58. 5%) were geolocalized, corresponding to 5428 users. Noise addition was found to be less accurate than k-anonymity using MASK data to protect the users' life privacy. Discussion: k-anonymity is an acceptable method for the anonymization of MASK data and results can be used for other databases.Peer reviewe

Synthesis Of Carbon Nanotubes And Nanofibers By Thermal Cvd On Sio2 And Al2o3 Support Layers.

In this work, catalytic thermal chemical vapor deposition method, using a mixture of methane and hydrogen at atmospheric pressure in a horizontal tubular quartz furnace, was used to grow carbon nanostructured materials. Silicon wafers with SiO2 or Al2O3 layers were used as support for thin nickel film deposition used as catalyst. It has been shown that the interaction between catalysts and substrates is of critical importance for carbon nanotube growth. However, this mechanism is not completely understood. Here, the interaction between catalyst nickel film and two different oxide layers supported on silicon wafers was studied as well as the influence of both support systems (SiO2/Si and Al2O3/Si) on the carbon nanostructures growth at different temperatures and process running times. The substrates were characterized by atomic force microscopy and the carbon nanostructured materials were studied by Raman spectroscopy, high resolution scanning and transmission electron microscopy. At higher temperatures it was observed a high density of carbon nanotubes grown over Al2O3 support layer when compared to SiO2 support layer showing a different behavior for Ni catalyst on each of the substrates. A quite different Ni catalyst behavior was observed at lower temperatures due to the formation of carbon nanofibers instead of carbon nanotubes on both substrates.94143-5

Long-term culture, genetic manipulation and xenotransplantation of human normal and breast cancer organoids

Organoid technology has revolutionized the study of human organ development, disease and therapy response tailored to the individual. Although detailed protocols are available for the generation and long-term propagation of human organoids from various organs, such methods are lacking for breast tissue. Here we provide an optimized, highly versatile protocol for long-term culture of organoids derived from either normal human breast tissues or breast cancer (BC) tissues, as well as culturing conditions for a panel of 45 biobanked samples, including BC organoids covering all major disease subtypes (triple-negative, estrogen receptor-positive/progesterone receptor-positive and human epidermal growth receptor 2-positive). Additionally, we provide methods for genetic manipulation by Lipofectamine 2000, electroporation or lentivirus and subsequent organoid selection and clonal culture. Finally, we introduce an optimized method for orthotopic organoid transplantation in mice, which includes injection of organoids and estrogen pellets without the need for surgery. Organoid derivation from tissue fragments until the first split takes 7–21 d; generation of genetically manipulated clonal organoid cultures takes 14–21 d; and organoid expansion for xenotransplantation takes >4 weeks

Drug-repurposing screen on patient-derived organoids identifies therapy-induced vulnerability in KRAS-mutant colon cancer

Summary: Patient-derived organoids (PDOs) are widely heralded as a drug-screening platform to develop new anti-cancer therapies. Here, we use a drug-repurposing library to screen PDOs of colorectal cancer (CRC) to identify hidden vulnerabilities within therapy-induced phenotypes. Using a microscopy-based screen that accurately scores drug-induced cell killing, we have tested 414 putative anti-cancer drugs for their ability to switch the EGFRi/MEKi-induced cytostatic phenotype toward cytotoxicity. A majority of validated hits (9/37) are microtubule-targeting agents that are commonly used in clinical oncology, such as taxanes and vinca-alkaloids. One of these drugs, vinorelbine, is consistently effective across a panel of >25 different CRC PDOs, independent of RAS mutational status. Unlike vinorelbine alone, its combination with EGFR/MEK inhibition induces apoptosis at all stages of the cell cycle and shows tolerability and effective anti-tumor activity in vivo, setting the basis for a clinical trial to treat patients with metastatic RAS-mutant CRC

Targeting mutant RAS in patient-derived colorectal cancer organoids by combinatorial drug screening

Colorectal cancer (CRC) organoids can be derived from almost all CRC patients and therefore capture the genetic diversity of this disease. We assembled a panel of CRC organoids carrying either wild-type or mutant RAS, as well as normal organoids and tumor organoids with a CRISPR-introduced oncogenic KRAS mutation. Using this panel, we evaluated RAS pathway inhibitors and drug combinations that are currently in clinical trial for RAS mutant cancers. Presence of mutant RAS correlated strongly with resistance to these targeted therapies. This was observed in tumorigenic as well as in normal organoids. Moreover, dual inhibition of the EGFR-MEK-ERK pathway in RAS mutant organoids induced a transient cell-cycle arrest rather than cell death. In vivo drug response of xenotransplanted RAS mutant organoids confirmed this growth arrest upon pan-HER/MEK combination therapy. Altogether, our studies demonstrate the potential of patient-derived CRC organoid libraries in evaluating inhibitors and drug combinations in a preclinical setting

DCL knockdown results in a delayed NB tumor growth.

<p>The normalized tumor growth is shown for shDCL-2 (A), shDCL-3 (B) and NC (C) tumors developed from the correspondent Dox-inducible NB cells lines injected subcutaneously in BALB/c athymic nude mice. Tumor growth (A-C) was normalized to the maximum tumor size per group. (D) Average of tumor volume 9, 12 and 14 days after injecting the Dox-inducible NB cells. NC, negative control Dox-inducible NB tumors. shDCL-2 and shDCL-3, Dox-inducible NB tumors that express shRNA against DCL. Error bars, s.e.m. *, <i>P</i><0.05; **, <i>P</i><0.01; ***, <i>P</i><0.001.</p