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Oxidative stress as déterminants of age depedent changes in regenerative properties of muscle satellite cells
Università degli studi di Chieti
Dept Neuroscience Imaging and Clinical Sciences
Type of Research Project
- Basic science
What is the background of the project?
In humans aging is a complex process that determines many physical and metabolic alterations correlated to the accumulation of oxidative damage in different tissues. Sarcopenia is an age-related non-pathological condition that includes a progressive loss of mass and strength in skeletal muscle, associated with a decline in the fibers’ functional capability. This condition could be correlated to abnormal reactive oxygen species (ROS) accumulation with consequent fiber oxidative damage. This complex situation is not only evident in mature muscle fibers but also in muscle resident satellite cells (involved in fiber damage repairing) in which some functional parameters seem to be modified.
What is the aim of the project?
The aim of this project is to study the relationship between ROS production and satellite cell activation during muscle using in vitro cell culture systems from human skeletal muscle of young and sarcopenic subjects.
What techniques and methods are used?
Satellite cell populations will be isolated from muscle fragments obtained from needle-biopsies. ROS levels will be measured in satellite cells derived from our sample, using a fluorescent probe, H2DCFDA (2 ', 7'-dichlorodihydrofluorescein diacetate), and by the rate of reduction of NBT (nitroblue tetrazolium) which produces formazan, quantified spectrophotometrically at 550 nm. The capability to differentiate of these cells will be evaluated Fusion Index, MF20 immunostaining and Western Blot. Furthermore will be evaluated the expression of genes involved during myogenesis.
What is the role of the student?
- The student will mainly observe
- The student will observe the practical experiments but will be highly involved in the analysis of the results
- The tasks will be done under supervision
What are the tasks expected to be accomplished by the student?
During the period of stay in the laboratory, the student will follow all the procedures relating to the sterilization of the material necessary for cell culture procedures, the preparation of culture media and the maintenance of cell cultures, including medium changes, transplants and cell counts. He/she will learn to use bench top centrifuges in order to pellet cells and separate the supernatant. The student will follow spectrophotometric and spectrofluorimetric procedures to determine the ROS levels. The student will follow procedures for determining the amount of protein, extraction of RNA and subsequent electrophoresis and RT-PCR (Reverse transcriptase-polymerase chain reaction). Following the experiments, data analysis will follow.
Will there be any theoretical teaching provided (preliminary readings, lectures, courses, seminars etc)
During the experimentation the student will be informed in detail of what is being done and why in order to have both the technique and the objective of the experiment understood
What is expected from the student at the end of the research exchange? What will be the general outcome of the student?
- The student will prepare a scientific report
What skills are required of the student? Is there any special knowledge or a certain level of studies needed?
Knowledge of cell biology and theoretical bases of molecular biology
Are there any legal limitations in the student’s involvement
Type of students accepted
This project accepts: - Medical students - Graduated students (less than 6 months)
- Superoxide Anion Production and Bioenergetic Profile in Young and Elderly Human Primary Myoblasts; M.Marrone; R.M.L. La Rovere; S.Guarnieri; E.S. Di Filippo; G. Monaco; T. Pietrangelo; G. Bultynck; S. Fulle; R. Mancinelli (PubMed; Oxidative Medicine and Cellular Longevity; 2018 Jul 24;2018:2615372. doi: 10.1155/2018/2615372)
- Myomir dysregulation and reactive oxygen species in aged human satellite cells; Di Filippo ES; Mancinelli R; Pietrangelo T; La Rovere RM; Quattrocelli M; Sampaolesi M; Fulle S. (PubMed; Biochemical and biophysical research communications; 2016 Apr 29;473(2):462-70. doi: 10.1016/j.bbrc.2016.03.030)
- Oxidative stress and muscle homeostasis; Musarò A. ; Fulle S; Fanò G. (PubMed; Current opinion in clinical nutrition and metabolic care; 2010 May;13(3):236-42. doi: 10.1097/MCO.0b013e3283368188)
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