Allergic reactions happen when the immune system overreacts to the presence of normally harmless substances known as allergens, such as pollen, mould, or dust mites. Allergic asthma can be triggered by inhaling these allergens, leading to swelling and tightening of the airways and breathing difficulties.
Asthma and allergy affect millions of people worldwide and place a significant burden on the healthcare economy. Asthma, for example, kills three people a day in the UK alone.
Human focused solutions are needed for this uniquely human condition
Most experiments to understand allergies have been carried out using mice, by exposing them to allergens, then extracting cells from their bone marrow or lungs for subsequent analysis. The complex immune responses at play during an allergic reaction however, are uniquely human and can’t be accurately reflected in animals. For some people, existing treatments do not work or fail so new drug targets are urgently needed. A human-focused model to shed light on the molecular and cellular mechanisms underlying allergic asthma would offer a much more effective and relevant avenue of investigation to achieve this.
Understanding cell messaging in allergic responses
Cells communicate with other cells, their environment and within themselves using messages sent via signalling molecules such as hormones as well as through physical changes that may influence how a cell behaves, moves or grows. This cell signalling process is essential to maintaining life and any faults that develop can lead to disease. An improved understanding of the messaging during allergic reactions could enable researchers to disrupt it, changing the way cells behave and revealing new treatment targets.
Understanding the role of ‘Hedgehog’ genes in allergies
Dr Anna Furmanski (now based at Oxford university), completed a pilot study at the University of Bedfordshire, focusing on mimicking cell signalling. Her aim was to use a human-based model to shed light on the messages passed to, and between white blood cells called eosinophils, which may drive adverse immune responses to harmless allergens.
Dr Furmanski investigated a signalling molecule called Hedgehog (Hh), its effects on eosinophils and its potential role in driving allergic reactions. This molecule is named after fruit flies that lack Hh genes and consequently resemble hedgehogs. Humans have three versions of this gene, one of which is aptly called ‘Sonic’.
Growing eosinophils in the lab
Dr Furmanski cultured (grew) eosinophils in the lab from human stem cells – cells capable of being induced to form a number of different cells. She used a chemical which enabled her to coax the stem cells to form eosinophils. Dr Furmanski also used animal-free growth supplements to culture the cells, in contrast to the traditional method used in labs whereby animal-derived biomaterials such as foetal bovine serum (derived from blood drawn from bovine foetuses in slaughterhouses) are used.
Dr Furmanski tested the effectiveness of two different animal-free growth supplements and determined that one of these, when used in conjunction with human serum (blood without cells and clotting molecules), enabled her to successfully culture (grow) animal-free human eosinophils for the first time.
Dr Furmanski confirmed this using molecular biology analyses and a stain specific to eosinophils, which enabled her to visualise and confirm their identity with a microscope.
Mimicking cell signalling
To understand the role of Sonic Hedgehog (Hh), Dr Furmanski grew eosinophils with and without human Sonic Hh protein present in the growth media. She then analysed five genes of interest which have important known roles in Sonic Hh signalling and are key players in cell growth and movement.
Dr Furmanski showed that when the cells were grown with Sonic Hh protein, these genes were ‘highly expressed’ meaning that a lot of proteins encoded by these genes were produced by the cells.
This demonstrated for the first time, that Hh signalling could be triggered in eosinophils. Of particular note, was a highly expressed gene called EPO, which has been implicated in airway tissue damage in asthma. A gene called TGF-beta was also highly expressed and which codes for a chemical that is known to increase the likelihood of changes in the airway during asthma, including fibrosis (scarring) and inflammation.
These findings show that the Sonic Hh signalling pathway represents a potential therapeutic target. If the signalling to eosinophils could be interrupted, this could reduce the respiratory damage that happens in asthma.
As well as investigating the genes involved in eosinophil signalling, Dr Furmanski also tested whether eosinophils migrated towards a chemical signal, which is important as this is how eosinophils move from the blood and into the lungs in allergic asthma.
She found that eosinophils grown with Sonic Hh, migrated to a lesser degree towards a chemical signal (called Interleukin-5) than has been seen in animal experiments. Dr Furmanski therefore plans to carry out further investigations into the movement of eosinophils to determine if there are key differences between human eosinophils and those derived from stem cells which could be influencing these findings.
The impact of this research
The next stage of this research would be to optimise the model further to ensure as many stem cells as possible specialise into eosinophils and that this process is controlled and consistent. Ultimately, this would pave the way for a widely adopted animal-free model to study allergic reactions that replaces animals.