19 June 2019

Using powerful radiation tools Assistant Professor at the LEO Foundation Center for Cutaneous Drug Delivery at Copenhagen University Kathryn Browning is trying to uncover the molecular mechanisms of the skin barrier. Her goal is ambitious: To create mixtures of lipids, which can be used as models of the human skin.

“It is an interesting area to be in because everyone knows somebody with skin diseases. My cousin had atopic dermatitis as an infant. He was given steroid creams and had to be covered head to toe in bandages every night. He was such an angry child, he was in pain constantly, and people didn’t know how to fix it. It was not a good childhood and it would have huge effect, if you could even understand one tiny part of how to deliver drugs or how to help with the itch and the inflammation,” Kathryn Browning explains.

In 2018 she was granted DKK 2.23 million from the LEO Foundation for a project, aiming to develop models of the skin barrier of healthy as well as diseased skin. (see more about the project below)

The curious kid on the block

Have you ever wondered, why you do not swell up like a sponge, when you go for a swim? Or why Bedouins living in the desert do not just evaporate? After all humans are almost 70 pct. water. Most people, however, do not ask themselves these kinds of questions. But British-born Kathryn Browning does. Already from her childhood she was constantly questioning the physics of the world.

“I just loved science and I liked to ask a lot of questions” she says.

She was probably one of those curious children, who ask in average 73 questions every day, as a study from 2017 shows. Most children stop asking questions when they grow up. She didn’t. Instead, Kathryn Browning has made her inquisitiveness part of her professional career as a biophysicist, and now she is working as Assistant Professor at the LEO Foundation Center for Cutaneous Drug Delivery in Copenhagen.

When Kathryn Browning did her first degree in chemistry, she didn’t dream of going into pharmaceutics. She did a PhD at Cambridge University in earth sciences, studying rocks and minerals in the ground and their interaction with oil, gas and water. However, the rocks led her into biology, as she discovered the powerful tool of neutron reflection. Visible light has a limitation, that it cannot be used to examine layers, that are thinner than the wavelength of light. But a beam of neutrons can be used to investigate the properties of very thin layers.

“I just fell in love with surface science and, when I was using neutron reflection, it became clear to me that biology is definitely the way to go – it is a really interesting topic, because there are so many things you can look at, which are interesting in terms of structure, surface and analysis.”

It is all about loving or hating water

Now in Copenhagen Kathryn Browning is investigating the fundamental aspects of chemistry and physics of the skin barrier layer, the “stratum corneum” in her quest to develop models of the layer.

The barrier is only 0,05 millimeters, consisting of multiple layers, and is often described as “bricks and mortar”. The “bricks” being dead skin cells, and the “mortar” a lipid matrix, composed of ceramides, free fatty acids and cholesterol.

For a drug to pass the barrier it must find its way through the lipid structure. It is all about a delicate balance between water loving and water hating properties.

As ceramides are a major lipid constituent of the skin barrier, these lipids are of special interest. The ceramides can self-assemble into multilamellar structures, and it seems that the tail length of the molecules is crucial to the structures.

“Some of the molecule chains are very long and very hydrophobic, and they make the barrier a lot stronger. But we don’t know exactly why and how. So, we focus on finding out, why these ceramides are so important to the whole structure, and to understand the exact bonding between them, that makes these structures form.”

Already Kathryn Browning and her team have realized, how important the right ceramides are to the whole structure.

“We did some work recently, where we just changed one of the molecules – there is only one tiny difference between the two of them -, and I was surprised just how important the exact molecules are. One tiny part of the molecule makes the structure form or not form. So really the exact composition is key to making this structure. “

There are more than 300 ceramides in the skin, and the researchers have access to four or five of the most abounded, which are commercially available.

Lipids for the models are extracted from pigskin

An important component of the research in skin diseases is to compare healthy skin to skin with various diseases, searching for changes to the molecules and structures that are characteristics of the diseases, making them leakier and less able to act as an intact barrier. And also, to investigate the penetration and interactions of drugs targeted to skin disease. The research could end up in new methods for development and testing of drugs as well as delivery methods of drugs that can be targeted specifically for the disease they aim to treat.

One problem however is, that it is difficult to get samples from patients. Most research and tests of drugs are made on samples from healthy skin, but this approach does not accurately represent lesioned and diseased stratum corneum. That is why Kathryn Browning wants to create models of the stratum corneum.

For that purpose, a wide variety of skin lipids not commercially available shall be used. Enter pigs. The skin from pigs is very similar to that of humans and is available from the veterinary and medical departments at the university, where students have been practicing their surgical skills. Kathryn Browning has hired a Postdoc, Amalie Ribel-Madsen, who is an expert in analysis of lipids and ceramides, and she is working to extract the lipid fraction from pigskin, and to separate all the different components.

The idea is to have a toolbox of different molecules, which can be mixed together to create a stratum corneum lipid matrix, which looks like a specific disease.

“Maybe it’s a bit of a pipe dream, but I would love, if we could get to the point, that we can make versions of the stratum corneum, that act essentially the same as real skin. You could have a bottle of lipid mixture, that looks like healthy skin and you have a bottle that looks like Atopic dermatitis and one like psoriasis. Instead of testing on people, just take it out of the fridge and spin it to test it. It would be fantastic.”

Beamtime is the best time

The research is still early on but right now Kathryn Browning, together with her team, is preparing four different compositions of lipids with slightly different ceramides. The mixtures are going through a process called spin coating, where the lipids are spun so fast that they spread out and arrange into structures, that are similar to the skin, forming a thin, perfectly smooth and uniform layer.

With the samples, each measuring 8 x 5 centimeter, in their suitcases, Kathryn Browning and her PhD, Liv Sofia Elinor Damgaard, are going to the synchrotron facilities in Oxford shire in England and Grenoble in France this summer to study the layers in detail and to conduct experiments on them.

Getting beamtime for neutron reflection is expensive and acquires a reasoned application, but Kathryn Browning has got all she asked for this year. On this trip she has four days, and for her the beamtime is the best time.

“It is very intense, and it is really exiting to go there. There is a special atmosphere and comradery when we go abroad. It is stressful and tiering, we work 24 hours a day, and you don’t really sleep. But all of the hard work is worth it – when you see things changing in real time and you realize, that you have a good experiment. That is what keeps me coming back. “

Although the time spent at the Synchrotron facility is highly appreciated, Kathryn Browning also is content with her everyday work life at the University.

“It is very inspiring to work at the LEO Foundation Center for Cutaneous Drug Delivery, we are ten colleagues of eight different nationalities, and various fields and backgrounds are represented as well. So, I get a lot of different perspectives on my research.”

When she leaves the university after work Kathryn Browning does not leave her curiosity behind. Everywhere she goes her attention is captured by surfaces and new questions arise in her mind, whether it be the paint on a wall “how do you make pigment stick to a wall?” or a straw in a glass of soda “Why do bubbles form on the straw?”

“There are tons of interesting interfaces everywhere you look, and that is where all the interesting stuff happens!”

The project

“Neutron reflectivity of healthy and atopic dermatitis lesional skin lipid models.”

The aim of the project is to compare drug interaction and penetration in healthy and compromised atopic dermatitis stratum corneum using a variety of biophysical techniques such as neutron reflection. In order to do this, the plan is to isolate and separate a wide range of lipids found in the skin’s barrier layer. These more complex models will allow the researchers to more accurately reflect the differences between the healthy and diseased skin.

The LEO Foundation Center for Cutaneous Drug Delivery

With a grant of DKK 40 million from the Leo Foundation the Center for Cutaneous Drug Delivery was established in 2016 at the Department of Pharmacy, UCPH. The center conducts research on, what happens in and on the skin, when we apply drugs. Particular focus is on the physical-chemical aspects of the interaction between skin and drugs, which is important to the development of new drugs.

What happens in a synchrotron?

A superconducting accelerator accelerates protons to a speed close to the speed of light. Neutrons, the small neutral particles in the atomic nucleus, are knocked out of the atom in the accelerator. Those neutrons are led to different instruments with material samples, that researchers want to analyze. The way the neutrons either penetrate the material or reflect in the material is detected by the instrument’s detectors. The data presented in graphs and numbers can be used to calculate, what the material consists of at the molecular level, and what properties it has. Under a beam of neutrons, you can find out, where atoms sit and what is the structure of materials. And when you know it, you can change it.