Primary cultures and maintenance of cell lines. Proliferation analysis methods. Preparation of histo-autoradiographic samples; Setting up of cell suspensions; Immunohistochemical detection of proliferation parameters. Biology and terminology of cultured cells. Molecular basis of cell cycle regulation. Flow cytometric techniques. Molecular mechanisms of protein synthesis control. Embryonic stem cells and iPS. Adult embryonic stem cells: their use in regenerative biomedicine.
Material provided by the professor (ppt slides of the lectures, video files and updated scientific reviews).
Alberts, “Biologia molecolare della cellula”, Zanichelli
«Biologia cellulare e molecolare. Concetti e esperimenti» Gerald Karp-Edises
“Colture Cellulari” Mariantonietta Meloni Aracne
“Culture of Animal Cells” Ian Freshney Wiley
Learning Objectives
The course aims to provide in-depth knowledge of the molecular mechanisms that control fundamental cellular functions such as proliferation and differentiation, focusing in particular on the different types of stem cells (embryonic, adult, induced pluripotent). Students will be able to understand the ways by which the cell modulates its behavior in response to exogenous stimuli, providing details on the molecular mechanisms underlying cell cycle regulation and protein synthesis. The aim of the course is also to provide students with (theoretical and practical) knowledge of methodological approaches for the study of cell biology.
Prerequisites
Courses required: none
Courses recommended: Cytology and Histology, Biochemistry, Molecular Biology and Genetics.
Basic knowledge of molecular biology, Citology and Histology, Biochemistry and Gentics is required.
Teaching Methods
Lectures (5 CFU, 40 hours)
Laboratory (1 CFU, 12 hours)
Teaching supporting material availbale on the Moodle platform.
Further information
Contact hours: Tuesday 12.00 -14.00 by e-mail appointment
Department of Experimental, Clinical and Biomedical Sciences
Viale Morgagni 50-50134 Firenze
Tel. 055 2751232
email : chiara.donati@unfi.it
email: caterina.bernacchioni@unifi.it
Type of Assessment
The exam consists of a final oral test preceded by passing a test of 12 multiple choice questions (at least 10 correct answers). The time provided for the execution of the test is 10 minutes, the answer not given is no longer penalizing for a wrong answer. The final oral exam is divided into three main questions (of which 1 on cell biology techniques) that the student must know how to deal with in an appropriate, exhaustive and correct manner. The multiple choice test is mandatory for access to the oral exam but has no weight as a criterion for the composition of the final grade. During the final oral exam there will also be an interview on the data reported in the laboratory diary filled out during the laboratory experiences.
Course program
Cell theory. Stem cells. Definition and characteristics of stem cells. Potency and self-renewal of stem cells. Embryonic stem cells. Adult stem cells. Transit amplifier progenitors. Epidermal, corneal, hematopoietic and intestinal stem cells. Crucial signaling pathways in the regulation of stem cell proliferation and differentiation: canonical signaling pathway of Wnt /beta catenin and Notch signaling pathway. Skeletal muscle satellite cells. Adult stem cell plasticity. Stem niche. Reproductive and therapeutic cloning. Mesoangioblasts. Reprogramming of somatic cells: iPS cells. Global molecular mechanisms of translational control. Specific mRNA molecular mechanisms of translational control. Cell cycle, mitosis and meiosis. The discovery of MPF: fusion experiments of eukaryotic cells, experiments with animal embryos, with yeasts. Discovery of CDC genes. The cyclin /CDK complex and regulatory mechanisms. MPF and entry into mitosis. Role of proteolysis from APC /cyclosome in late mitosis. Mitogens and mechanisms that control the entry into the cell cycle and the beginning of the S phase in animal cells. Enabling DNA upon duplication. The response to DNA damage, p53, ATM / ATR. Experiments by Howard and Pelc. Methodological approaches for the study of the cell cycle, the flow cytofluorimeter. Methods to evaluate the extent of the phases of the cell cycle. Brief history of cell culture. Advantages and limits of cell cultures. Cell biology in culture, preparation of primary cultures and cell lines / strains. Handling of cell cultures, description of laboratory equipment, micro-environment, sterility and preservation techniques. Contamination. Culture environment: substrates, culture media, temperature and gas phase, cryopreservation. Cell characterization: growth curves, vitality test, quantification and cytotoxicity. Techniques for the study of in vitro and in vivo proliferation. .