Laser microdissection of actinomycin D segregated nucleoli

Nucleoli of tissue culture cells were segregated into their fibrillar (light) and granular (dark) components by treatment with actinomycin D. Following this segregation, the cells were treated with quinacrine hydrochloride, an agent which selectively sensitizes the nucleoli to argon laser light. The actinomycin D-segregated, quinacrine-sensitized nucleolar components (dark and light) were selectively irradiated with the laser microbeam and subsequent uridine uptake assayed. The data indicate that selective damage to the light (fibrillar) area is generally more damaging than damage to the dark (granular) area. These results support the idea that DNA is closely associated with the nucleolar fibrillar component.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/22301/1/0000745.pd

Quantitative near-infrared spectroscopy of cervical dysplasia in vivo

The aims of this study were: (i) to quantify near-infrared optical properties of normal cervical tissues and high-grade squamous intra-epithelial lesions (H-SIL); (ii) to assess the feasibility of differentiating normal cervical tissues from H-SIL on the basis of these properties; and (iii) to determine how cervical tissue optical properties change following photodynamic therapy (PDT) of H-SIL in vivo. Using the frequency domain photon migration technique, non-invasive measurements of normal and dysplastic ecto-cervical tissue optical properties, i.e. absorption (μa) and effective scattering coefficients, and physiological parameters, i.e. tissue water and haemoglobin concentration, percentage oxygen saturation (%SO2), were performed on 10 patients scheduled for PDT of histologically-proven H-SIL. Cervix absorption and effective scattering parameters were up to 15% lower in H-SIL sites compared with normal cervical tissue for all wavelengths studied (674, 811, 849, 956 nm). Following PDT, all μa values increased significantly, due to elevated tissue blood and water content associated with PDT-induced hyperaemia and oedema. Tissue total haemoglobin concentration ([TotHb]) and arterio-venous oxygen saturation measured in H-SIL sites were lower than normal sites ([TotHb]: 88.6 ± 35.8 μmol/l versus 124.7 ± 22.6 μmol/l; %SO2: 76.5 ± 14.7% versus 84.9 ± 3.4%

Intrauterine light delivery for photodynamic therapy of the human endometrium

Photodynamic therapy is currently being evaluated as a minimally invasive procedure for endometrial ablation not requiring anaesthesia. Light penetration depths at 630, 660 and 690 nm and the optimal configuration of intrauterine light-diffusing fibres were determined in 14 human uteri to assist in the design of a light intrauterine device. Post-menopausal ex-vivo uteri showed a significantly lower light penetration depth than pre-menopausal uteri. With a single central diffusing fibre inserted, the fluence rate measured in the uterine wall at the most remote point of the cavity decreased to 1.1 ± 0.4% of that measured at closest proximity, whereas it decreased to only 40.0 ± 9.0% with three fibres. Distension of the uterine cavity with 2 ml of an optically clear fluid increased the fluence rate at the fundus between the fibres at a depth of 2 mm by a factor of 4. We conclude that in normal-sized pre-menopausal uterine cavities, three diffusing fibres will deliver an optical dose above the photodynamic threshold level at a depth of 4 mm, even in the most remote areas, in <30 min without causing thermal damage. For distorted and elongated cavities, either slight distension of the cavity or the insertion of a fourth diffusing fibre is require

Uterus and endometrium: Regeneration processes in rabbit endometrium: a photodynamic therapy model

The origin and process of regeneration in rabbit endo metrium was evaluated following photodynamic epithelial destruction using topically applied anilnolevulinic acid (ALA). Selective destruction of endometrial epithelium was performed using photodynamic therapy (PDT). ALA was diluted to 200 mg/ml dextran 70 shortly prior to adininistra tion. A volume of 1.2 ml was injected into the left uterus. Intrauterine illumination (wavelength 630 nm, light dose 40-80 J/cm2was performed 3 h after drug administration. Tissue morphology was evaluated by light and scanning electron microscopy 1, 3, 7 and 28 days post-treatment (three animals at each dine-point). Regeneration of the endometriwn following epithelial ablation by PDT was fully activated after 24 h and was completed after 72 h. Endometrial surface generation occurred by proliferation, originating primarily in deeper regions of the glands. Findings from our morphological follow-up study support the origin of endometrial regeneration being mainly from undifferentiated stem cells and residual glandular epitheliu

Photodynamic destruction of endometrial tissue with topical 5-aminolevulinic acid in rats and rabbits

ObjectiveThe goal of this study was to determine the optimal parameters for photodynamic endometrial destruction with topically applied 5-aminolevulinic acid, a precursor for the endogenous synthesis of the fluorescent photosensitizer protoporphyrin IX.Study design5-Aminolevulinic acid pharmacokinetics were measured in rat and rabbit models by analyzing tissue frozen sections 3 to 12 hours after topical administration. Dose-response studies were conducted for 100 to 400 mg/ml 5-aminolevulinic acid. Photodynamic therapy was performed intraluminally, and tissue morphologic features were evaluated 3 and 7 days after treatment.ResultsPeak fluorescence was observed 3 hours after topical administration. Glandular fluorescence significantly exceeded stromal and myometrial in all studies, particularly for 200 mg/ml 5-aminolevulinic acid. Histologic studies revealed persistent epithelial destruction with minimal regeneration.ConclusionTopical 5-aminolevulinic acid photodynamic therapy can be used for highly effective, long-lasting destruction of endometrial epithelium. However, optical dosimetry can vary, particularly in the rabbit model, and this appears to have an impact on long-term reepithelialization

Online peer assessment in higher education: a systematic review of literature in educational practices

Neste artigo procedemos a uma revisão sistemática da literatura sobre Online Peer Assessment (OPA) no Ensino Superior, mediadas por Tecnologias Digitais de Informação e Comunicação (TIC). Identificamos suas características, centrais e opcionais e mapeamos exemplos práticos (procedimentos e TIC) de OPA que poderão ser transversais, adaptáveis e aplicáveis em diversas unidades curriculares e regimes educacionais. Os resultados apontam para a utilização da OPA como uma estratégia que potencia a “avaliação para a aprendizagem”. Os referenciais teóricos subjacentes, os métodos de avaliação e os tipos de TIC utilizadas indicam seu direcionamento para um maior envolvimento e responsabilidade do aluno na sua aprendizagem. Evidenciamos na literatura que se busca desenvolver essas competências, dando ao aluno oportunidades com alguma regularidade para autoavaliar-se e avaliar seus pares por meio de feedback construtivo. Há também evidências de que dar ou produzir feedback é mais benéfico para a aprendizagem do que apenas recebê-lo, como também é cognitivamente mais exigente e envolve os alunos de forma mais ativa e os direciona ao pensamento crítico e a processos metacognitivos. A partir dos resultados alcançados, consideramos que a OPA, enquanto ferramenta cognitiva, contribui para a construção do conhecimento e para a reflexão sobre a aprendizagem. Um desafio que se coloca diz respeito ao desenvolvimento criativo com foco na diversificação e na inovação das práticas de avaliação no sentido de potenciar aprendizagens e resultados acadêmicos, em atenção às necessidades de aprendizagem que se manifestam frente às expectativas da educação atual e futura e às exigências da sociedade.In this research a systematic review of literature about Online Peer Assessment (OPA) in higher education, mediated by Information and Communication Technology (ICT), was carried out. Optional and central characteristics and its practices (procedures and ICT) were identified and mapping. These procedures and practices may be transversal, adaptable and applied in several curriculum units and educational regimes. The results point to the use of OPA as a strategy that enhances “assessment for learning”. The theoretical framework, the methods of assessment and the kinds of ICT used indicate directions for greater involvement and responsibility from the part of the student in his/her learning. Theoretical framework identifies the need to develop students’ skills, providing them with opportunities for self-assessment and peer assessment on a regular basis through constructive feedback. There is also evidence that to give or receive feedback is more beneficial for learning than just receiving it. It is also cognitively more demanding and directs the students to critical thinking and metacognitive processes. From the results obtained, OPA is seen as a cognitive tool, contributing to the building of knowledge and to reflection about learning. Issues for further reflection are also identified such as the creative development of assessment methods with a focus on diversity and innovation in order to enhance students’ learning and academic results, taking into account their learning needs in face of the expectations of current and future education and the demands of society.CIEC - Centro de Investigação em Estudos da Criança, IE, UMinho (UI 317 da FCT), PortugalFundos Nacionais através da FCT (Fundação para a Ciência e a Tecnologia) e cofinanciado pelo Fundo Europeu de Desenvolvimento Regional (FEDER) através do COMPETE 2020 – Programa Operacional Competitividade e Internacionalização (POCI) com a referência POCI-01-0145-FEDER-007562info:eu-repo/semantics/publishedVersio

Applications of optical trapping to single molecule DNA

This is the final report of a three-year, Laboratory Directed Research and Development (LDRD) project at the Los Alamos National Laboratory (LANL). The project focused on the methodologies required to integrate optical trapping with single molecule detection (SMD) so as to demonstrate high speed sequencing through optical micromanipulation of host substrates, nucleotide cleavage, and single molecule detection. As part of this effort, the new technology of optical tweezers was applied to the confinement and manipulation of microsphere handles containing attached DNA fragments. The authors demonstrated substrate optical trapping in rapid flow streams, the fluorescence excitation and detection of fluorescently labeled nucleotides in an optical trapping system, and the epifluorescent imaging of DNA fragments in flow streams. They successfully demonstrated optical trapping in laminar flow streams and completely characterized the trapping process as functions of fluid flow velocity, chamber dimension, trapping depth, incident laser power, and fluorescence measurement geometry