Hemoglobin has been studied extensively in red blood cells – it acts to carry oxygen and carbon dioxide through the body. When it was discovered inside the oocyte it initiated a wave of questions and a new research direction. What role is hemoglobin playing inside the oocyte and what affect does this have on fertility? Dr Hannah Brown, a Postdoctoral Researcher in the Early Development Group started investigating this on a short six month contract in 2012 and this project has continued to evolve and present new questions ever since. With 1 in 6 couples experiencing clinical infertility, does the presence of hemoglobin provide a new avenue to understand the biology at play? Hannah and colleagues have made some significant findings in this area, but there is still plenty more to uncover.
Hemoglobin is the protein molecule in red blood cells that carries oxygen from the lungs to the body’s tissues and returns carbon dioxide from the tissues back to the lungs. Research into hemoglobin has been conducted for decades in red blood cells, but little is known about this protein anywhere else in the body.
When Associate Professor Jeremy Thompson discovered hemoglobin in oocytes he became fascinated with understanding why this was, and offered Dr Hannah Brown a six-month contract to look into the significance of this finding. Until this point, Hannah had been living in Europe and the US for 4 years working as a postdoc and jumped at the opportunity to return home to continue her research.
“I was ready to return home to Australia after postdoccing at Baylor College of Medicine in Houston. I was interested in learning more about Jeremy’s discovery and this opportunity was a great fit with my previous research in ovarian biology.”
“I intended to work on this project for six months and decide on my next step from there. However, the longer I spent researching the role of hemoglobin in the oocyte, the more fascinated I became and I realised there is much more to the picture and much more to uncover,” said Hannah.
In the ovary, the level of internal hemoglobin continues to rise in the oocyte and its surrounding cells, otherwise known as the cumulus oocyte complex, as it matures. The ovary is very unique in terms of its oxygen biology, and the oocyte grows in an environment that has little access to blood vessels and direct oxygen.
When Hannah replicated the process of oocyte maturation in culture during In Vitro Maturation (IVM), the hemoglobin levels inside the oocytes reduced and appeared to ‘turn off’.
“The complication with this project is that we know very little about the role of hemoglobin outside of red blood cells. However, given that hemoglobin is really important for carrying oxygen we think this is the main role its playing inside the oocyte, and it is especially important given the possible low oxygen environment of the ovary and reproductive tract,” explained Hannah.
Hannah and colleagues looked into how and when hemoglobin was produced in the ovary while it’s preparing for ovulation. They found the two components – alpha and beta hemoglobin were present – and these were likely to form functional hemoglobin.
Both components dramatically increase at the time leading up to ovulation, just prior to when the oocyte travels through the fallopian tube to meet the sperm.
“We know a lot of critical processes occur during this window of time and the hemoglobin levels follow this same pattern,” said Hannah.
In response to the reduced levels of hemoglobin in the oocytes undergoing IVM, Hannah added hemoglobin into the culture media of mouse oocytes, and found that the oocytes could take up the added hemoglobin.
To test whether this improved the quality of the oocytes and could lead to better embryos, Hannah fertilised these oocytes and after four days she observed there were more blastocysts in the groups treated with the form of hemoglobin that can carry oxygen.
“This observation was very exciting. We’d made a small, but significant, improvement to the oocytes and resulting embryos,” explained Hannah.
The level of oxygen the oocyte and embryo is cultured in is very important to its development – high levels can cause free radicals that may lead to damaged DNA, proteins and membranes. Knowing this, Hannah added hemoglobin to the oocytes at different oxygen concentrations: 20% oxygen (atmospheric level), 5% oxygen and 2% oxygen.
“Only when we added the hemoglobin at 20% and 2% did we see an improvement. The oocyte didn’t respond at 5% oxygen, and this is a continuing trend we are observing.”
“To really understand the significance, we need to uncover what the hemoglobin is doing – what its natural role is – and we’re looking at biological systems such as metabolism that require high levels of oxygen to help us find the answer. Perhaps hemoglobin can bind nitric oxide which is very important during oocyte maturation, or perhaps it acts as an anti-oxidant,” said Hannah.
Hannah works within the Early Development Group led by A/Prof Thompson. As a group, they explore the metabolic and epigenetic consequences of environmental stress on the earliest stages of embryo development. Additionally, they are developing new technologies and interventions to measure changes and establish optimal environments.
“We are a well-rounded collaborative team that utilises multi-disciplinarily approaches to answer early development questions on the nano-scale.”
“As 1 in 6 couples experience clinical infertility, Hannah’s research into hemoglobin is another progression of our IVM techniques and I believe it will provide further improvement for treating women with fertility problems in the near future.”
“We have a wonderful collaboration with Cook Medical, our technology development partner and with Vrije Universiteit Brussel (VUB) and The University of New South Wales, our clinical and research partners to develop IVM into a practical alternative choice for women seeking to solve their infertility,” explained A/Prof Thompson.
It is this practical application that provides Hannah with the motivation to continue her research.
“I believe IVM will be a feasible procedure for fertility preservation for cancer patients (as an example). We would be able to remove their oocytes and freeze these before they begin cancer treatments,” explained Hannah.
“Additionally, women undergoing IVF are required to inject hormones that are both expensive and have side effects. If we can fully understand the biological pathways of oocyte and embryo development we will be able to improve IVM techniques, and provide a much cheaper alternative for couples experiencing infertility.”
The next step for the team is to develop improved and relevant models to fully explore the workings inside the fallopian tubes to be able to understand the impact on the oocyte.
“I believe over the next five years there will be significant technological improvements and we’ll be able to see in real time what is happening inside the fallopian tubes,” said Hannah.
As well as a Research Leader in the Robinson Research Institute, A/Prof Thompson is a Chief Investigator for the ARC Centre of Excellence for Nanoscale BioPhotonics (CNBP). This Centre is developing technologies to probe the complex and dynamic nano-environments within living organisms using light.
“IVM is a new technique being used in fertility clinics but the success rates have been low. Our work as part of the CNBP and with our partners will help us to more closely mimic the natural pathways inside the ovary and reproductive tract to improve assisted reproductive technologies (ART) – leading to greater pregnancy success,” said Hannah.