Success stories
John Evans: A vision of the hormones that create life
John at work. The image on the right is the view down the microscope. Cells from the pituitary gland of a rat. Cells containing 'luteinising hormone' have been dyed red.
The Challenge of Medical Research
The human body is a symphony of delicate processes, so ingenious, complicated and beautiful it is hard for us to conceive. John Evans, a full time researcher in the department of Obstetrics and Gynaecology at the Christchurch School of Medicine and Health Sciences has dedicated his life to revealing its mysteries. He works in the field of reproductive endocrinology, in other words he studies the hormones involved in the reproductive process. The challenge John faces as a medical researcher is to tune into this symphony of intricate interactions, to learn how to introduce just the right amount of the right chemical at the right time to alter a target process without disturbing the fine balance in the rest of the body. John hopes to find ways of both increasing and decreasing fertility by making subtle alterations to the hormone balance. The more precisely the processes are understood the more accurately the target process can be pinpointed. The result being a safer treatment with fewer side effects.
How does nanotechnology help?
Tuning into Nature’s symphony is no small task. The tools and techniques currently used are reaching their limits. As part of the bio-nanotech group in the MacDiarmid Institute John has been working with Dr Maan Alkaisi an electrical engineer from Canterbury University. Maan’s expertise is in nano-fabrication - using the powerful lithography equipment at Canterbury University to make very very small devices. John is familiar with the biological research requirements. With their powers combined they have created the ‘bio-chip’- a tiny platform with microscopic cavities that trap single biological cells. An atomic force microscope (AFM) scanned over the cell’s surface enables high-resolution real time viewing of cell functions. The advantage of using the AFM is that it can image cells in their natural environment, which means they can be kept alive (it doesn’t require a vacuum like electron microscopy). The AFM probe, which touches the surface as it scans in the past tended to move or distort cells making them impossible to image.
From Statistics to Single Cells
Cells can be positioned and held in place so each one can be given an “address” on the biochip. The AFM tip can be programmed to return to the same address an hour after a chemical is added to watch its effect on the cell. Researchers will be able to distinguish between one cell giving a strong signal and many cells that are each giving a weak signal - a revolutionary step from statistics to single cells. Birth of the individual! Renaissance of a cellular sort! The bio-chip may be the instrument John needs to join the endocrinological symphony.
Reproductive Endocrinology
The story begins in the pituitary gland – the master control of the endocrine system. This pea-sized gland situated at the base of the skull secretes hormones that are essential for many of the body’s functions, including the stress response and growth as well as in reproduction.
The Human Brain
The hormones from the pituitary gland that act on the ovary to stimulate the release of an egg are called gonadotrophins. John is focussing on two of these called LH (luteinising hormone) and FSH (follicle stimulating hormone). The pituitary gland is controlled by a part of the brain called the hypothalamus. They are connected by a stalk, down which hormones like GnRH (gonadotrophin-releasing hormone) are sent to trigger the release of other hormones (like LH and FSH) from pituitary cells. GnRH binds to a receptor molecule – a long protein threaded through the pituitary cell membrane (each receptor is only receptive to a few hormones.) This causes changes in the receptor, which activate a G-protein within the cell. The G-protein sets off a chain reaction that finally results in the production and release of LH.
Sometimes, however, LH doesn’t need to be made. It can be stored in granules within the pituitary cell. Granules are made of a membrane that will fuse with the cell membrane. To release the LH you need to tell the granules to get to the surface of the membrane. A pore opens up (as in the picture below) through which the stored LH can escape. This process is called ‘exocytosis’.
Exocytosis
Using atomic force microscopy John will be able to watch as the pores form. He wants to discover how they function, where, when and in what numbers they form and the effect of adding different concentrations of peptides like GnRH at different times. Bio-chip technology makes all this possible.
Rat’s Brains to Bio-chips
How do you get the cells from the rat’s pituitary to the biochip cavities without killing them? John has this down to a fine art. In a pituitary, cells are basically held together with protein. He gets an enzyme that chews up protein and this breaks up the linking strands and separates out the cells. Then you put them in a medium, which has to be compatible with the integrity of the cell. Pure Water is no good. It is absorbed by the cells, which makes them burst. They need a range of chemicals in the surrounding solution to keep them happy: salts, amino acids, glucose, cholesterol ... Choosing a suitable medium is an art in itself. The next step is to position the cells on the platform. This is where Maan comes in. Cells have dielectric properties that are affected by their ionic composition and other cell characteristics. Maan use s the cell’s dielectric properties to move them around by creating electric fields with a system of electrodes laid over the platform surface. The cells' characteristics are different in living, dead and infected cells and thus the dielectric properties will be distinct. These differences can be used to separate the healthy cells from the rest. “Once they’re there (in their allotted cavities) …the cells will in theory be happy for quite some time. Then we can run the AFM over one – take a look – add some GnRH and see if, when and in what conformation pores appear.”
Applications
After leaving the pituitary, gonadotrophins (like LH) make their way to the ovary. Their effect is to stimulate ovulation. Attempts have been made to use this property to modulate fertility. The idea is that giving women injections of GnRH will increase LH emission, which will in turn stimulate ovulation and increase their fertility. Conversely GnRH antagonists can be found that bind to the same receptor protein without triggering LH release. They block the process and have possible use in contraceptives.
Blatantly removing or adding a hormone from the delicate endocrine symphony, however, is a dangerous business. Hormones are multi-taskers. They have different functions all around the body. Oxytocin, for example, is a hormone normally associated with contractions of the uterus during childbirth. However it has been observed to have effects at the ovary and the thymus and even in memory, and also in control of LH! “There’s a fine balance, we’re not quite sure about this. We may be able to develop a peptide-based contraceptive that will require quite small amounts of peptide if it can upset the balance and be targeted at a particular process and if therefore small amounts can be used then the possibility is that the side effects will be lessened… So if we can understand what’s happening in the pituitary then we may be able to target that for either increasing or decreasing fertility. That’s the rationale for beginning to understand what’s going on”
If John and his team succeed the implications will be far reaching; contraceptives and fertility treatment with minimal side effects; non-toxic contraceptives for possum control and a greater appreciation of the intricate endocrine symphony at the heart of life.
