Label-free multiphoton microscopy of lipids in oocytes, eggs and early embryos
PhD Research
Intake: October 2012
Funding:
The studentship is fully-funded for 3 years, i.e. includes the payment of tuition fees at the Home/EU rate and a tax free maintenance stipend. Non-EU applicants are welcomed but successful applicants would have to self-fund the difference between the Home/EU and International Fee.
Primary Supervisor: Dr Paola Borri (BorriP@cf.ac.uk),
Secondary Supervisors: Dr Karl Swann (SwannK1@cf.ac.uk) (School of Physics); Prof Wolfgang Langbein (LangbeinWW@cf.ac.uk) (School of Physics)
Project details:
Efficient metabolism is essential for early development of mammalian oocytes and embryos. The metabolic rate of an early embryo is also a key indicator of viability. Whilst most studies of metabolism in oocytes and embryos have looked at exogenous substrates (e.g. glucose) it is clear that endogenous lipid metabolism is also significant [1]. In particular studies in mice and pigs have shown that the metabolism of lipids (beta-oxidation) is one of the critical cytoplasmic events occurring during oocyte maturation, and that it affects the success of pre-implantation development [1,2]. Pig oocytes are most often used for such studies since they contain very large lipid droplets [1]. Mouse oocytes are a better model for human oocytes since both contain much smaller lipid droplets. However, the lipids droplets in mouse oocytes are poorly characterized. In addition, no continuous imaging studies of lipid droplets in oocytes have been made because fluorescent lipid labels are poorly retained in organelles and sustained fluorescent excitation light impairs development.
Coherent Antistokes Raman Scattering (CARS) microscopy has recently emerged as a new multiphoton microscopy technique which overcomes the need of fluorescent labelling and yet retains biomolecular specificity. This technique has proven to be very successful in imaging lipid droplets [3]. In our Biophotonics group we have built a CARS microscope equipped with a heated stage and environmental control to enable long term imaging of mammalian cells.
This project will exploit CARS microscopy to visualise unstained lipid droplets in mouse oocytes and pre-implantation embryos. This will be done with living oocytes undergoing maturation or fertilization, as well as with early embryos. Lipid droplets will be studied alongside other important features of oocytes such as chromosomal configuration, redox state or intracellular calcium levels. Pig oocytes will be used for comparison since they contain the largest lipid droplets for an oocyte. We shall also investigate the metabolism in mouse oocytes and early embryos using deuterium labelled lipids [4]. The work promises to establish new methods for non-invasive assessment of oocyte or embryo quality which could be used in clinical IVF or animal reproductive technologies.
References
[1] Sturney RG et al (2009) Role of fatty acids in energy provision during oocyte maturation and early embryo development. Reprod. Dom Anim 44, (Suppl 1) 50-58
[2] Dunning K.R., et al. (2010) Beta-oxidation is essential for mouse oocyte developmental competence and early embryo development. Biology of Reproduction 83, 909-918
[3] Israel Rocha-Mendoza, et al, “Differential coherent anti-Stokes Raman scattering microscopy with linearly chirped femtosecond laser pulses” Optics Letters 34, 2258 (2009).
[4] Xie XS, et al. (2006). Living cells as test tubes. Science 312, 228–230.
How to apply:
To fill an application form and for further information see: http://www.cardiff.ac.uk/presidents/bioimaging/index.html