How Gurdon Institute’s Dr Emma Rawlins uses organoids to unravel secrets of lung development
How does the human embryo build a lung? And how do our bodies maintain and repair this complex organ from day to day?
The answers to these questions could help researchers devise new regenerative therapies for the millions of people living with lung disease.
At the Wellcome Trust / Cancer Research UK Gurdon Institute in Cambridge, Dr Emma Rawlins is focused on the innovative use of organoids – mini versions of an organ in a dish – to unravel the secrets of human lung development.
“The big picture is the burden of chronic lung disease throughout the world,” she explains to the Cambridge Independent. “The British Lung Foundation estimates that one in seven people in the UK has some sort of lung condition, which ranges from asthma to full-blown chronic obstructive pulmonary disease (COPD) or lung fibrosis.
“What we are interested in is how you normally build, maintain and repair your lungs in order to contribute towards the tissue maintenance and repair we need if we are going to cure some of these people.
“If you have a chronic lung disease like COPD you are gradually losing your gas exchange surface. If you have lung fibrosis, your gas exchange surface will gradually be getting clogged by all these horrible fibrotic cells that stop you breathing properly. There are no real cures for these. It’s a real unmet need.”
While there are treatments for both COPD and idiopathic pulmonary fibrosis (IPF), they can only slow down their progression. The impact of COPD is highly variable, and is influenced by whether a patient smokes, but average life expectancy for someone with IPF is around three to five years.
“The big concept in my work is whether we can learn from what normally happens in the embryo and in the day-to-day maintenance in order to regenerate a diseased lung. That’s a massive challenge,” says Dr Rawlins.
But pioneering techniques are now being used to help meet this challenge.
“For years, we’ve used mice,” explains Dr Rawlins. “It’s still a very valid model. Mouse embryos have many similarities to humans. They have the same overall structure. But there are differences in cell types – and size and time are the two biggest things you can’t get around.”
A mouse lung has a total capacity of about 1ml, compared to the average human lung capacity of 6,000ml. And while our gestation period is nine months, for mice it is just 19-21 days. These differences have prompted the search for closer equivalents to study.
“In the UK we are in a very privileged position in that we have access to human embryonic material,” says Dr Rawlins.
“They have research nurses, who deal with informed consents and permissions, to collect these embryos from aborted tissue.”
The tissue bank was established in 1999 to provide human embryonic and fetal material from 26 days to 20 weeks of development for use by academic researchers.
“It makes some people very uncomfortable, but you have to remember that human tissue is typically discarded as medical waste – it would be incinerated,” says Dr Rawlins.
“These are the most well-used embryos of any species anywhere. Material can be sent all over the UK and further afield, as long as the appropriate ethics and governance regulations are in place, to make the maximum possible use of every sample. Anything that can’t be used fresh by a lab is frozen and banked for future use.
“It’s an amazing resource.”
Dr Rawlins has typically used this embryonic material for comparing genetic results between human and mouse cells. Now, it can also be used for something more revolutionary.
“With new organoid techniques, we can start to grow cells from human embryos and carry out functional genetics on cells in a dish.
“Since we worked out how to do this a couple of years ago, that’s really the direction my work has gone in,” she says.
Specialised stem cells known as progenitors, which only exist in the embryonic lung, are used in the lab to create what might be described as the beginnings of a human lung.
“In the embryo, these cells build the lung. They make the beautiful branched structure,” says Dr Rawlins. “For every tissue, you have to have a group of cells that make the right number of divisions to get to the right size and the right number of cell types. They do that in such an organised way.”
In the human embryo, these stem cells divide, are differentiated into different cell types and create the shape and structure of the lung in a process known as morphogenesis.
“What we can model in a dish at the moment is the cell divisions and the types of cells that they make,” says Dr Rawlins. “What will be the challenge is the morphogenesis.
“Instead of growing organoids or balls of cells, which are very useful and very beautiful, could we actually grow something that starts to look like a tiny lung – part of a tube or part of a gas exchange surface? It becomes an engineering challenge in addition to a biological challenge.
“There are a lot of different cell types in the lung and what we are studying very actively is the mechanisms that control that differentiation process. We can manipulate the organoids and ask all sorts of very interesting questions.
“The short-term goal is to take our organoids, which are immature and embryonic, and get them to something that resembles, at least in cell types, a neonatal lung. This is the most tractable initial question but it’s really important.
“If we can get them to resemble the neonatal lung, we can really study in detail the process of maturation, from embryo to neonate. That would be massive.”
Lungs continue to develop in the womb until around 36 weeks, meaning babies that are born prematurely can have respiratory issues.
“Doctors have amazing interventions but you still end up with a smaller lung,” says Dr Rawlins. “Can we improve that and can we find biomarkers for lung maturity to guide doctors?”
This could help medics determine when a premature baby is ready to come off an incubator.
“It’s terribly distressing – I have a friend who went through this,” says Dr Rawlins. “The baby was relatively healthy but was born a few weeks early by Caesarian section because the mother had been so sick.
“The doctors know that being on an incubator damages the lung so they tried four times to get her off it, and three times she had to go back on. To identify some markers will be a challenge but would be amazing.”
To stimulate the embryonic lung stem cells to differentiate into different cells, the researchers change the culture medium in which they are kept, altering hormones and growth factors, but they want to improve their maturity.
“We get a certain amount of differentiation. But we think that’s not enough,” says Dr Rawlins. “We are trying – this is all brand new – to provide other neighbouring cells.
“At the moment, we are growing epithelial cells, which form the gas exchange surface of the lung, but they have a really close relationship with the blood vessels, so we are asking ‘Can we mimic that relationship in the dish and will that improve the maturation? Do the cells need to talk to one another?’
“But we are also starting to think about the more mechanical forces. In the embryo, the lungs are already making breathing movements. Do you need those to differentiate the cells?
“The final aspect is the change in circulating hormones as you approach birth. Can we mimic some of that in our culture dish?”
The stem cells themselves can self-renew, providing an ongoing resource for the researchers.
“If we get the progenitor cells to make more copies of themselves, we can keep them forever. We can freeze-store them and gene-edit them,” says Dr Rawlins.
“But then once we switch to differentiation conditions, at the moment we can only keep that going for a few weeks.”
The work has helped to identify the progenitors in the human lung and some new signalling pathways that control them.
“We are pushing at the boundaries now,” says Dr Rawlins. “What we are starting to work on is whether we can apply what we’re learning to lung regeneration.
“We have to do this in a mouse model at the moment. We are asking whether we can take the cells that we have growing as organoids and put them into an adult lung and, as a cell therapy, will they integrate and form a functional gas exchange area?
“We don’t know the answer – it’s work in progress.”
But Dr Rawlins admits she is “not terribly keen” on this regeneration approach using cell transplants.
“The practicalities of growing the cells and keeping them clean, and the genetic matching, are too difficult to make it very realistic,” she says.
“What I’m more excited about is that if we know how we build the structure in the embryo and we know the cues that maintain these stem cells in adults, can we reactivate some of our embryonic cues?”
Dr Rawlins and a colleague in the Cambridge Stem Cell Institute are now growing adult human lung organoids in culture, as well as embryonic lung organoids, to compare how they are regulated.
“What really attracts me about reusing embryonic pathways is that embryonic development is usually a very self-limiting process. You build it and then you stop. That would be crucial to activating any of this in an adult,” she says.
In adults, lung stem cells do very little from day to day until we are struck down by an upper respiratory tract infection.
“This is fascinating. If you have a cold or flu, your secretory cells make lots of snot, and you get an injury to your epithelium. Lots of cells then proliferate. Your stem cells respond and over time repair it.
“Amazingly, there are several documented cases now of people having had a big bit of their lung cut off to remove a cancer and, over the course of several years, they have regrown pretty much all of the gas exchange surface.
“So the stem cells are there and can act, but day-to-day there is very little going on. It’s a remarkably flexible system, which is modelled extremely well in the mouse.”
Understanding cell differentiation in the human lung could also have implications for treating lung cancer.
“You can think of a cancer as a group of undifferentiated cells,” explains Dr Rawlins. “If you could force them to differentiate into a very benign cell type, because we’ve worked out these mechanisms from the embryo, that would be very useful.”
It is thought that there are two key factors in other diseases, such as COPD.
“One is where embryonic development hasn’t gone perfectly,” says Dr Rawlins. “A person may have had enough gas exchange capacity to last into adult life but, as it starts to decline, they notice more because they didn’t have enough to start with.
“The second factor is having repeated injuries, especially if they are a chronic smoker. The stem cells have repeatedly responded and are just exhausted.”
Understanding the role of our progenitors and stem cells, and using organoids to probe the development and maintenance of the human lung, could prove crucial in the long-term battle against such conditions.