From Diamond Light Source, this is the Diamond Podcast.
Meera Senthilingam: Welcome to the Diamond Light Source Podcast, with me, Meera Senthilingam. This month we’re moving out to the clinic to see how Diamond’s beamlines are being used by Clinicians Nationwide, finding out how x-rays are being used to identify the potential risks of metal-on-metal hip replacements as well as for finding biological markers for faster diagnosis of cancer.
Josep Sule-Suso: And then we can see differences maybe in the fat content or in the proteins or in the DNA and we’re using these differences, which we call markers, for in the future to diagnoses cancer at the single cell level.
Meera: Josep Sule-Suso will be explaining how his team are looking into finding these bio-markers in lung cancer cells later in the show when we also investigate the causes of pre-term labour.
Anna David: Most women deliver at term, in other words at 40 weeks of pregnancy, around that time. But about 10% of women deliver at less than 37 weeks and that does seem to be on the increase and we really know very little about why that happens.
Meera: Anna David will be giving us an insight into how the analysis of amniotic membranes could maybe one day help us to predict who is likely to go into pre-term labour. So all that insight into the clinic plus all the latest news and events from Diamond coming us in this June edition of the Diamond Light Podcast.
Meera: The users of Diamond Light Source come from a wide range of scientific disciplines. One recent discipline is Medicine, where clinicians are using Diamond’s X-rays on samples donated by patients in order to research a variety of medical problems. This could offer great contributions to future medical advances, so this month’s podcast is focussing on the range of clinical research taking place using the synchrotron; the first being the work of obstetrician Anna David, from University College, London. Anna is trying to find out more about the causes of preterm labour by studying the structures of amniotic membranes - the thin wall of the amniotic sac in which foetuses develop in the womb. I went along to University College Hospital in London where Anna is based to find out more about how she is looking into this.
Anna David: Most women deliver at term, in other words at 40 weeks of pregnancy, around that time. But about 10% of women deliver at less than 37 weeks and that does seem to be on the increase and we really know very little about why that happens. The problems with babies that deliver pre term that many of them who deliver very early end up having problems with brain development; lung diseases and we know that later on in life they end up having cardio-vascular disease and hypertension, so there’s a big research effort to try to find out why women deliver pre-term.
Meera: How common are pre-term labours, how often do they happen?
Anna: Pre-term labours probably happen in about 10 % of women during pregnancy. We know that in about 4 out of 10 cases the membranes surrounding the baby rupture early and many of those women go into labour, fairly soon after the membranes rupture.
Meera: What are thought to be the main causes then?
Anna: Well in some women there does appear to be infection. We know that women who get an infection get inflammation of the membrane and are more likely to rupture their membranes. There are other reasons, for instance, women who have heavy bleeding early on in pregnancy. The blood seems to weaken the membrane in some way and causes them to rupture early and lastly, stretching the membrane, for example in twins or triplet pregnancies, or in women who have extra fluid around the baby, something called polyhydramnios, seems to put extra pressure on the membranes and causes them to rupture pre-term.
Meera: In order to understand why this happens, you’ve set about looking at the actual structure of the amniotic membrane and looking into what changes in the structure to cause these ruptures?
Anna: The amniotic membrane is a sort of bag that carries the baby. The baby is surrounded by fluid and there is a sac around the baby and usually the placenta is near the top of the uterus and then the membranes are around the baby and down near the bottom of the uterus, down near the cervix or the neck of the womb. There are actually 2 layers to this sack; there’s the Amnion which is the internal membrane and this is a single layer of epithelial cells and there’s the Chorion which is the outer layer of membrane. Now the Amnion is made up of collagen fibres and it’s the collagen fibres which seem to bring the tensile strength to the membrane because the amnion is actually incredibly strong. Now we know during pregnancy, at the end of pregnancy, the amnion alters its structure, in the place down near the cervix. There is a zone of altered morphology down near the cervix which is the area where the membranes are much more likely to rupture.
Meera: Now to set the scene a little bit about how this membrane is structured, we’ve got a placenta here in front of us. So tell me more about this placenta, it’s a little bit bigger than I thought it was going to be!
Anna: Well it is quite a big thing. A placenta is an amazing structure, it feeds the baby for 40 weeks then suddenly around the time of the end of pregnancy you go into labour and the placenta is delivered. As you can see there are 2 layers to the membrane, the inner layer is really very fine, we can peel off the outer chorion, and the inner membrane is really fine. If I push my finger through the membrane, you really have to push it through quite a lot and it still hasn’t ruptured. You can imagine that this baby is surrounded by, sometimes up to a couple of litres of fluid and yet it still doesn’t rupture.
Meera: So it’s actually incredibly strong and it’s very thin, so it’s basically transparent. I’m pushing my finger right through it now and there’s no sign of it bursting or giving way at all!
Anna: If you stretch the membrane, actually the tensile strength of the membrane, the amnion membrane, increases until it reaches a threshold above which it ruptures. So it is a very clever piece of biological machinery which prevents us going into labour before we should do.
Meera: It seems then that the amniotic membrane really can take a lot of stress. How are you setting about looking at the structure of this and understand what happens in order for this to burst early?
Anna: Well we’re looking at the major component of the amnion which is the collagen fibres. Nobody has really looked how the collagen fibres function – how they’re spaced out, how they’re aligned, how dense they are within the membranes and then how this correlates to how the membranes rupture.
Meera: Which beamline are you using at Diamond and how does this enable you to visualize the structure of the collagen within these membranes?
We’re using the beamline called I22 which is the non-crystalline diffraction beamline.
The x-rays go through the amniotic membrane, we line up the amnion in the x-ray beam, and what happens is as the x-rays hit the collagen fibres in the amnion, some of them are scattered by those collagen fibres and we can work out by the amount of scatter, the alignment of the collagen fibres, how closely spaced they are and how many there are within the membrane.
Meera: What have you been able to visualize so far, how is the collagen structured within this particular area?
Anna: Well, we’ve found that the collagen is actually structured in quite an ordered fashion. So we’ve looked initially at ‘term’ membranes from women who have delivered on time and what we’ve found is that there is a difference in the way that the collagen is aligned in membranes that are sampled from near the zone of altered morphology, near the neck of the womb where it ruptures and membranes that are further apart. So in the membranes near the cervix, the collagen is more lined up, the fibrils are more lined up, and they’re also slightly more spaced apart in comparison to membranes that are near the placenta and you might hypothesise that this would actually increase the risk of the membranes rupturing in that area.
Meera: Have you been able to look at samples of membranes from people that have ruptured pre-term?
Anna: We’ve put the membranes in the beamline and we’ve found that in comparison to ‘term’ membranes the collagen fibres are more lined up so we think there may be some difference in the pre-term compared to the term membranes.
Meera: Are there any thoughts to what this predisposition to this could be or what the causes of this could be? Or how do you essentially use this to predict who may rupture early in the future?
Anna: Well we’re basically looking at how the different factors we know increase your chance of rupturing early, how they affect the collagen structure in the amniotic membrane, so we’re going to be doing some experiments in the next beamline by incubating the membranes overnight with blood, or with bacterial products like lipo-polysaccharide that are related to infection that perhaps might break down the collagen or actually repetitively stretching the membranes and then seeing how those factors alter the collagen’s structure in the amnion; and then we may be able to say more able why those factors affect structure. The other things is that we know there are genetic predispositions to rupturing the membranes early and it may be that these are related to the collagen’s structure – but obviously a lot more work is needed on that and the synchrotron facility is really the only place that we can do this type of work.
So lots more to find out in order to predict who may be prone to pre-term labour. That was Anna David from University College, London. Now moving away from the womb and on to diseases that affect you later on in life, one of which is cancer. There are many forms of cancer affecting millions of people globally each year, many can be treated if caught early enough so any diagnostic method that can help identify cancerous cells in the early stages could make a dramatic difference. Dr Josep Sule-Suso is an oncologist at the University of Keele and he’s using Diamond's infrared beamline
to study lung cancer cells in hope of finding markers that could spot cancer cells early on in their tracks. I spoke to Josep to discover how he’s doing this.
What we’re trying to do is we’re using the Diamond synchrotron the beamline B22
to study single cancer cells, some of them are lung cancer, some of them are normal cells. And the fact we use the synchrotron is because it allow us to look inside the cell, smaller areas in the cell and then we can see differences maybe in the fat content, or in the proteins or in the DNA and we are using these differences, which we call markers, for in the future to diagnose cancer at a single cell level.
Meera: How do you set about using infra red, how does it enable you to see more about what is going on inside the cell?
Josep: Infra red, especially using the synchrotron, allows me to look different parts within the cell, to look at what the cells are made of.
Meera: What are the advantages of using a synchrotron and how do transfer the information you find here, back to the lab to look into it further?
Josep: The advantage of using a synchrotron is the gives you a very bright infra red beam which allows you to study differences not just one single cell, but areas within the cell. This is very difficult to do with what we call bench top spectrometers – instruments you can have in the pathology lab for instance. So the idea is to find out these differences using the synchrotron and once we got these differences, these markers, then we can use these bench top spectrometers, which can be placed in pathology departments in hospitals, then to characterise these cells that are abnormal but not cancer yet.
Meera: What have you been able to see so far? What are the key differences between a cancerous call and a normal one?
Josep: This is a pilot study so we still need to do a lot of work but primary work shows that we are seeing differences in the protein concentration and the structure of some of the proteins and also the DNA.
Meera: You have some samples in front of us now. Can you tell me a bit more about what’s on these slides, what are we looking at?
Josep: These slides come from patients with lung cancer, and contain a small amount of tissue that contains both cancer cells and normal cells. So we take these samples to the synchrotron and we shine the infrared light on to each individual cell trying to find these markers.
Meera: How is this going to help the situation, what are the current problems when it come to lung cancer and diagnosing it?
Josep: Presently if we had someone who we suspect may have lung cancer, they would have a biopsy, usually a bronchoscopy - where they put the tube along the nose or the mouth down to the main windpipe and try to get a small piece of tissue and from there the pathologist, the doctor who looks at the sample would tell if it is cancer or not. However, sometimes the samples are too small or they contain a very small number of cells and the pathologist will say that some cells are abnormal but I’m not quite sure if they are cancer. In these situations, the patients have to undergo further biopsies with all the risks and side effects for them, delaying the treatment and also increasing the costs for the NHS, so this is the problem.
Meera: So your hopeful method will allow them to look at single cells and therefore diagnose a bit earlier or a bit more specifically.
Josep: Yes, that is the theory - to be able to use infrared light to look at those cells that are considered abnormal and then based on the data we obtain from infra red light to be able to say yes, they are cancer, and the patient can start treatment quicker.
Meera: So how much earlier do you think it might be possible to catch the cancer?
Josep: I don’t think is a matter of catching the cancer earlier, it is about helping patients start treatment earlier, a matter of 2-3 weeks difference, I think that’s what we are aiming at here rather than catching the cancer earlier.
Meera: You’ve mentioned you’ve got some of the slides here, so this one in particular – could you just tell me what’s on here and what are the differences? At the moment there are 3 samples really on there I think and what are the differences between these?
Josep: These samples are 3 cuts from the same piece of tissue at different levels and they show part of lung tissue which is normal and also part of lung cancer. We’ll use the infrared beamline to study just one of them and we’ll look at cells that are within the normal part of the lung and within the cancer.
Meera: How would you then summarise the stage which you’ve currently reached with the study and what the next steps are hoped to be?
Josep: Well at the moment we’ve started this study which is a pilot study. We’ve tried to include 50 patients and from each patient to get a sample of tissue with cancer and a sample without cancer. Based on the data obtained on this pilot study we can show there are differences between lung cancer cells and normal cells using the infrared beams. The next step would be to include loads of patients and I’m talking hundreds of patients if not thousands that would help us to confirm if these markers do indeed exist.
Meera: Would it be possible to scale this up and look out to different types of cancers?
Josep: Yes, we’ve started with lung cancer as this is the main area of my work, but I can imagine that this could be extended to many other types of cancer as well.
Meera: That was Dr Josep Sule-Suso from the University of Keele.
Voice-over: Did you know it took Dorothy Hodgkin 12 years to solve the structure of Insulin? If she were to do this today on one of Diamond’s MX Beamlines, it would only take 15 minutes!
Meera: You’re listening to the Diamond Light Source Podcast and this month we’re entering the clinical world to find out how Hospital based clinicians are using Diamond’s Light source to learn more about a variety of medical issues and still to come… we’ll be finding out why metal-on-metal hip replacements could be causing side effects in some patients and how clinicians are trying to identify the possible causes. Before that, let’s join Sarah Bucknall from Diamond’s Communication team for a round up of the latest news and events from Diamond.
Sarah Bucknall: The latest news from Diamond is really exciting. We’ve had approval for funding for Phase 3 of Diamond Light Source which basically means 10 more beamlines added to the facility. At the moment we’re drawing to the end of phase 2 where we’ll have 22 operational beamlines by 2012 and over the next 7 years we’re going to add 10 beamlines so we’ll extend capabilities and extend scientific output.
Meera: So what makes this phase 3? How is this going to differ from the beamlines that are finishing just now on phase 2?
Sarah: Well we go out to our scientific community and we ask them what they want from Diamond, from that we draw up proposals for these beamlines that we want to add on. The next 10 beamlines are really varied, they’ll be for a wide range of scientific disciplines. One of the beamlines will provide high resolution 3D images of biological samples, so they’ll allow scientists to look inside the cell at a scale of a few millionths of a millimetre and that would be useful, for example, with the analysis of bone calcification or radiation damage of cells and basically further our knowledge of diseases and develop new therapies. And moving into a different area, we could also branch out into food research, allowing scientist to understand the viscosity.
Meera: So that will result in an even wider range of science taking place at Diamond, speaking of which, there have been some recent papers published in a key journal – the Journal Science - recently, what where these about?
Sarah: Well there was one recently which will help with the development of Industrial Catalysts. Researchers from the Universities of Manchester and Cardiff have succeeded in engineering crystals that are able to maintain a porous structure which means they can carry out chemical reactions within that crystal structure. Previously the pores have been maintained by solvents and as soon as you wash those solvents away, the structure collapses.
Meera: So they’re focusing on making more stable catalysts and they’ve been getting their ideas from nature, from enzymes in fact
Sarah: Yes they’ve taken their initiative from enzymes which are basically nature’s catalysts and the design of the new crystal is such that is can exist happily in water-based environments. So we’re talking about Industrial catalysts that don’t need to exist in a chemical environment and that are able to trap gas molecules and basically carry out those chemical reactions.
Meera: Well it’s always important to improve the catalysts used in our industries, but now moving away from chemistry and onto biology, some key findings about a specific protein have been found out.
Yes this is a really recent paper in Science, researchers from the Universities of Leeds, Oxford and Imperial College, London have determined the 3-D structure of a single transporter protein in basically all its 3 main structural states. This protein is responsible for getting essential chemicals into the cell and it sits within the membrane of a cell. Some of the scientist used the Membrane Protein Laboratory
that’s based at Diamond to map out the structure of this protein.
Meera: How has the protein been found to work then?
Sarah: Well this group have been looking at this for 10 years. They mapped out the first 2 states of the protein and they published those results a couple of years ago and they’ve finally completed the puzzle by publishing the 3rd state and what it is if you can imagine this protein that sits in the membrane of a cell a cavity is open facing away from the cell where the chemicals can go in to that cavity. It then locks up, so this is the 2nd state, and it then opens up a cavity inside the cell which releases the chemical so that is the 3rd state. So now they have got a really detailed idea of how the chemicals are transported inside the cell.
Meera: How can this info be used, what are the potential applications for it?
Sarah: Well one of their unexpected findings is that this is the case for many transporter proteins whereas they thought it was more of a stand alone. Now that they’ve got detailed knowledge of the mechanism, then can hopefully look at new drug developments, altering the delivery of compounds into a cell is a potential benefit for treating illnesses. For example, if there are conditions where particular chemicals are lacking and need a boost, they can use this mechanism to help. One example would be diabetes where they would need more glucose, or somebody with depression needed more serotonin.
Meera: Some pretty important findings there, but stepping away from the research, you’ve got some quite exciting things coming up at the Royal Society there.
Sarah: Yes, we’re taking part in the Royal Society Summer Science Exhibition. This time round it’s a bit of a bigger deal because the Royal Society are celebrating their 350th Anniversary. So the exhibition is 10 days long from 25th June to the 4th July and it’s going to be at London’s South Bank Centre and it’s basically going to be a gathering of lots of interesting and exciting science that’s going on around the country and Diamond will be taking part in that. We have some researchers there who are using Diamond to study Alzheimer’s, there’s a chemist who’s doing research into more efficient hydrogen storage, and also a journey into atomic and molecular research under high pressures and temperatures. So there’s lots coming up and it promises to be a really good event so people just need to keep an eye on our website to find out more.
Meera: That was Sarah Bucknall from Diamond’s Communication team there who’ll be back in the next edition with more news from Diamond. But in the meantime if you’d like to find out more about the research taking place at Diamond, come along to their stand at the Royal Society Summer Exhibition in London this summer from the 25th June to the 4th July. Now moving back to the science taking place at Diamond, this month we’re investigating the world of clinical science and the research taking place with clinicians across the UK. Our last feature for this month’s podcast looks into the hip replacement. Thousands of hip replacements are performed in the UK each year and scientists are constantly looking to improve the materials used to provide longer lasting replacements with maximum bone conservation. A more recent development was the metal-on-metal hip replacement which was also used in hip resurfacing procedures but in some patients these have led to adverse reactions and inflammation around the hip. Dr Alistair Hart is an orthopaedic surgeon based at Imperial College, London, and I met him at his labs at Charing Cross Hospital in London to find out more about the basics of our hip joints and what’s thought to be going wrong with these replacements.
Alistair Hart: Hip joints are ball and socket joints. The ball of the thigh bone, or femur, fits within the socket of the pelvic bone. When this joint is worn out, we call that disease arthritis. The most common type of arthritis is ‘wear and tear’ or ‘osteoarthritis’.
Meera: What are the current options available for people with osteoarthritis or hip problems in general?
Alistair: They take pain killers and try to live with it or if the hip is completely worn out, they undergo hip replacement. We can offer hip replacements that are either just replacing the surfaces of the bone, as in hip resurfacing, or by removing the head of the femur or the thigh bone as well as the socket part of the pelvis and that is a standard hip replacement.
Meera: The standard hip replacement has been around for a long time and the hip resurfacing is a slightly newer technique, what are the different materials available for this hip resurfacing?
Alistair: There is only one type of material suitable for hip resurfacing, that is cobalt and chromium alloy. It is the only material that can be made thin enough to replace the 3mm worn out cartilage and still stick to the bone. It can be made to reline both sides of the joint and encourage a thin film of fluid to separate those two metal surfaces so that the wear of the surfaces is very low. The hope for his resurfacing was that it would outlast standard total hip replacements and that if a second hip replacement was required, there would be more bone conserved for the second hip. Unfortunately recent evidence points towards a higher failure rate following hip resurfacing that flowing standard hip replacements.
Meera: And why are these thought to fail at a faster rate?
Alistair: The likely mechanism of failure involves the metal-on-metal joint couple. If the 2 surfaces are metal against metal then that seems to cause more problems.
Meera: Now we’re just in your lab at the moment and you’ve got a few failed hip joints in front of us that have used the hip resurfacing technique. Could you just take me through this particular sample here? So you’ve just taken one failed hip resurfacing joint out now, tell me a bit more about this one
Alistair: The first one is hip resurfacing and we see that the bone appeared to have been well fixed to both the cup implant and to the head implant.
Meera: The actual movement between the two is reasonably smooth.
Alistair: It does feel smooth and in fact in the body it is even smoother.
Meera: It’s a failed joint essentially, but it doesn’t really look like it so what would have happened for this to have failed?
Alistair: The most common pattern of failure at the moment seems to be unexplained. Patients after only a few years of having the new metal-on-metal hip replacement, develop pain on the hip. Some develop inflammatory lumps when we perform MRI scanning. It does seem to relate to the amount of metal that is released from the surface.
Meera: So the fact this metal on metal does wear, releases nano sized metal particles into the tissue surrounding the patients with hip resurfacing and this could therefore be one of the potential causes of the failure?
Alistair: Yes, all hip replacements wear. In fact the metal on metal hip that is functioning releases a million very small nano sized particles every step and as we walk a million steps per year we release a trillion metal particles into the hip joint every year.
Meera: That’s a lot of nano particles being released, what’s thought to be causing the damage as a result of these particles being released?
Alistair: We know that the hip implant is made of cobalt, chromium and molybdenum in the ratio 60:30:7 however there are many possible culprits for the active metal species which causes the inflammation and the failure – for example, is it the metal particles of cobalt, chromium and molybdenum or is it just the cobalt or the chromium or the molybdenum and which type of cobalt, chromium and molybdenum? And do these metal elements recombine into a larger molecule that is the active species?
Meera: That’s a lot questions that need answering. How are you setting about looking into this?
Alistair: When we collected the failed hips, we also collected the tissue surrounding the failed hips. We examined this tissue at the Diamond Light Source for both physical and chemical characteristics. We use a high intensity x-ray beam at the I-18 microfocus beamline at Diamond. It is probably the only method for exactly determining the type of chemical in the tissues.
Meera: Now you have various images and results in your office of what you’ve managed to find using these techniques so I think we’re now heading back to your office in order to see these.
Alistair: Yes, let’s go and look at the chemical maps and the spectroscopy that’s shown us exactly the type of metal in the tissues.
Meera: we’re now in your office and we’ve got various images and graphs set in front of us. What have you been able to find out so far about where these metal nano particles of metal are migrating to or where they’re located.
Alistair: What we did first was use the Diamond x-ray beam was to map the metal within the tissue of over 30 different tissue samples. We found that these contained mainly Chromium. We did not find many areas that had cobalt, chromium and molybdenum in. The majority of areas showed just Chromium, in other words, the alloy had changed from the hip replacement to how the cells in the tissues were incorporating it.
Meera: What kind of chromium did you find?
Alister: One of the abilities of Diamond Light Source was that it was able to tell us exactly the valence or oxidation state the chromium was in. We found that it was in the chromium 3 valence state. There are many types of chromium including chromium 6 which is a cancer-causing chromium. We did not find the chromium 6. This does not exclude that it is never present but it does fit with the clinical data which suggests there aren’t cancers around these hips, but that there are inflammatory lumps. We went on to show that Chromium 3 is actually present, but is present as Chromium-3-phosphate
Meera: What’s the next step from here? What can you do knowing this to deal with the situation?
Alistair: Now we have clues to what agents we can give to cells in a test tube in a controlled experimental environment to see if we can recreate the inflammation that occurs within a human. It may not necessarily be the chromium phosphate as the active agent. We do now know the types of cobalt- molybdenum-chromium we should test and then what we want to do is then take those cells that have been treated experimentally with the active agent and put them back into the Diamond Light Synchrotron to see if we can produce the same images from patients.
Meera: Having done all of this, what is thought to be the potential solution?
Alistair: I don’t actually know the answer to that. The orthopaedic companies are trying many different avenues and that probably illustrates the confusion for example, it’s possible to have a hip replacement that is made of metal-on-metal, ceramic-on-ceramic, metal-on-plastic, but it’s also possible to have them made on metal-on-ceramic combination, there are polyurethane surfaces being used, so it appears we are a long way away from a well accepted, everlasting hip. It is probably better to develop better biocompatibility tests, that is tests that are more realistic, so instead of stifling innovation, we can develop these new types of bearings, these new types of hips, but in a way that’s safer.
Alistair Hart from Imperial College, London. Now that’s it from this edition of the Diamond Light Podcast but do join us again in July when we’ll be bringing you the highlights from the Royal Society Summer Exhibition with a focus on the research that’s being showcased there by Diamond. In the meantime if you have any questions about diamond or research that’s taking place there, the email address is firstname.lastname@example.org
or you can listen to previous editions of the podcast online at www.diamond.ac.uk/podcast
. Or you can also subscribe on iTunes. Thank you to Anna David, Josep Sule-Suso, Alistair Hart and Sarah Bucknall. I’m Meera Senthilingam, thank you for listening and see you next month.