Identification of microRNAs for the early diagnosis of colorectal cancer (CRC)

Introduction: microRNAs (miRNAs) are small non-coding RNAs involved in the development of various cancers. Quantitative real-time PCR (qPCR) is the assay commonly used to investigate miRNA expression and qPCR-low-density arrays are the most used technique for both identification and validation of modulated miRNAs. One crucial pre-processing step for miRNA analysis is data normalization, aimed at reducing nonbiological sources of variation [1]. This process would allow to identify a small set of miRNAs to be used for data normalization in subsequent validation studies [2]. Materials and methods: we analyzed the expression levels of 381 human miRNAs on TaqMan Array MicroRNA Card A v.2 (Applied Biosystems), on a cohort of 60 plasma samples (38 precancerous lesions/cancer and 22 without lesion) from individuals enrolled in the CRC screening program of the Milan Local Health Authority that underwent colonoscopy at our Institute (INT) after a positive fecal occult blood test (FIT+). Starting from these data, we developed and applied a data-driven normalization method able to identify a small set of reference miRNAs to use for data normalization in the subsequent validation studies [2,3]. Briefly, by considering the miRNAs expressed in all the samples, the relative expression of each miRNA was first computed according to their mean expression value [4] and the best subset of miRNAs that resemble this value was selected [2,3]. Results and discussion: we identified 4 housekeeping miRNAs suitable for data normalization and 7 miRNAs significantly different in subjects with precancerous/cancerous lesions versus subjects without lesions. We also identified 4 miRNAs related to presence of initial adenoma, one linked to advanced adenoma and 8 to presence of cancerous lesion. We are now constructing of a Custom TaqMan Array Cards including the identified miRNAs and some other miRNAs of interest, i.e those haemolysis-related and miR-378 [5], with the aim of validating these biomarkers on a prospective cohort of 120 FIT+ subjects that underwent colonoscopy at INT. Conclusion: the adopted strategy allowed the identification of reference miRNAs to be used for data normalization and the identification of modulated miRNAs that will be validated in larger prospective series. Acknowledgements: This work was supported by grants from Associazione Italiana per la Ricerca sul Cancro (AIRC) (Grants No.10529 and No. 12162 to MA Pierotti)

Histological characteristics of the corpus luteum of Nelore cows in the first, second and third trimester of pregnancy

Foram avaliadas características morfológicas do corpo lúteo de 48 vacas Nelore gestantes obtidos de abatedouros. Os ovários com o corpo lúteo foram coletados, identificados e divididos em três grupos, considerando o estágio da gestação determinado pelo tamanho do feto: Grupo I - onze animais com gestação até 90 dias; Grupo 2 - vinte animais com gestação de 90 a 180 dias, e Grupo 3 - 17 animais com gestação de 180 a 261 dias. Todos os corpos lúteos foram dissecados, submetidos a processamento histológico e avaliados utilizando microscopia de luz. As características morfológicas das células luteais esteroidogênicas não mudou durante a gestação. Porém, foi observado um aumento de tecido conjuntivo, fibroblastos e matriz extracelular durante o final da gestação. Células em degeneração foram observadas em todos os períodos da gestação, mas com maior intensidade no fim do terceiro trimestre. Grânulos foram observados após a coloração com Tricrômico de Gomory e Xylidine Ponceau, caracterizados como grânulos de proteína. Nenhuma explicação foi encontrada na literatura para coloração de grânulos pelo Tricrômico de Gomory