Human Stem Cell Research in Europe

New Report Calls for Sustained Public Endorsement and Funding for Human Stem Cell Research

Wednesday, 06 November 2013

A strategic report from the European Science Foundation examines the key scientific questions for human stem cell research in the context of the rapidly emerging field of regenerative medicine. In parallel to the potential new treatments for incurable diseases resulting from stem cell research, heated ethical and legal debates have arisen across the world. This report presents a comparative view of the legislative framework on human stem cell research across Europe and provides a selection of success stories in frontier research and clinical trials that underpin the advances achieved in Europe to date.

In recent years, international research on regenerative medicine and stem cells has yielded some promising results and even greater expectations in society. In this medical field, human embryonic stem cells could be applied in a variety of ways, for example to identify new compounds for drug development, or as cell-based therapies. The potential to use human stem cells to repair or replace tissue or organ functions lost through age, disease, damage or birth defects may raise ethical issues that must be considered integrally with any research. Europe is currently witnessing developments and debates that will impact regulation and public funding of stem cell research and innovation for years to come.

The report observes that Europe plays a leading role in regenerative medicine research, with most countries featuring legislative frameworks that are globally favourable to human stem cell research. The 30 countries' position on human stem cell research was grouped into five broad categories; very permissive, permissive with restrictions, restrictive by default, very restrictive and unlegislated. The report found that 63% of the countries fell into the first two categories.

"Europe has a valuable track record in the area of stem cell research. The report highlights the need to continue to fund this research so that its full potential can be realised." said Professor Stig Slørdahl, Dean of the Faculty of Medicine at Norwegian University of Science and Technology, who chaired the report.

The authors recommend that sustained public endorsement and funding need to continue in order for further research to be carried out and public-private partnerships to develop, bringing safe and innovative therapies to the market, with a potential benefit to millions of patients worldwide.

Dr Vanessa Campo-Ruiz, ESF Science Officer to the Chief Executive and lead author of the report commented:

"We hope this report may help to inform future policy and funding decisions across Europe and thus contribute to ensure this continent's scientific leadership, social welfare and economic growth."

About The European Science Foundation

The European Science Foundation coordinates collaboration in research, networking, and funding of international research programmes, as well as carrying out strategic and science policy activities at a European level. Its members are 67 national research funding and performing organisations, learned societies and academies in 29 countries.

Source: European Science Foundation 

Contact: Emma Knott

Reference:

Human Stem Cell Research and Regenerative Medicine: Focus on European policy and scientific contributions

(available online, pdf)

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Pancreatic Stem Cells Isolated from Mice

Pancreatic Stem Cells Isolated from Mice

Tuesday, 17 September 2013

Scientists have succeeded in growing stem cells that have the ability to develop into two different types of cells that make up a healthy pancreas. The research team led by Dr. Hans Clevers of the Hubrecht Institute, The Netherlands, have isolated and grown stem cells from the pancreases of mice using a 3-D culture system previously developed by the scientists. The results, which are reported in The EMBO Journal, could eventually lead to ways to repair damaged insulin-producing b-cells or pancreatic duct cells.

Cell signalling molecules known as Wnts and a protein called Lgr5 are essential to produce adult stem cells that can be coaxed to grow and divide rapidly. However, these signalling pathways and molecules are inactive in the adult pancreas.

"We have found a way to activate the Wnt pathway to produce an unlimited expansion of pancreatic stem cells isolated from mice," Clevers said.

"By changing the growth conditions we can select two different fates for the stem cells and generate large numbers of either hormone-producing b-cells or pancreatic duct cells."

"This work is still at a very early stage and further experiments are needed before we can use such an approach for the culture of human cells but the results are a promising proof-of-concept," he added.

In the study, the pancreases of mice were altered in a way that makes duct cells proliferate and differentiate. Some cells in this new population were stem cells that were capable of self-renewal. The scientists were able to culture these cells to give rise to large numbers of pancreatic cells or tiny clumps of tissue referred to as organoids.

Therapeutic strategies for pancreatic disease have been hampered by a lack of cell culture systems that allow scientists to grow replacement tissue in a test tube or on a dish. Alternative approaches such as tissue transplantation are limited by the scarcity of donors and the possibility of tissue rejection. The new work offers access to an unlimited supply of pancreatic stem cells that would be beneficial for the development of new therapeutic interventions for pancreatic diseases like diabetes.

The next steps for the scientists will include further refinement of the cell culture methods developed in this study and investigation of ways to extend the approach to human pancreatic cells.

Source: European Molecular Biology Organization 

Contact: Barry Whyte

Reference:

Unlimited in vitro expansion of adult bi-potent pancreas progenitors through the Lgr5/R-spondin axis

Meritxell Huch, Paola Bonfanti, Sylvia F. Boj, Toshiro Sato, Cindy J. M. Loomans, Marc van de Wetering, Mozhdeh Sojoodi, Vivian S.W. Li, Jurian Schuijers, Ana Gracanin, Femke Rignalda, Harry Begthel, Johan H. van Es, Eelco de Koning, Robert G.J. Vries, Harry Heimberg and Hans Clevers

The EMBO Journal, 17 September 2013, doi:10.1038/emboj.2013.204

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Variations in Gene Expression

EMBL-EBI researchers present global map of human gene expression
Friday, 09 April 2010

Just like members of an orchestra are active at different times although playing the same piece of music, every cell in our body contains the same genetic sequence but expresses this differently to give rise to cells and tissues with specialised properties.

By integrating gene expression data from an unprecedented variety of human tissue samples, Alvis Brazma and his team at the European Bioinformatics Institute, an outstation of the European Molecular Biology Laboratory (EMBL), and their collaborators have for the first time produced a global map of gene expression. The full analysis behind this unique view of the genetic activities determining our appearance, function and behaviour is published today in Nature Biotechnology.

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This image, representing the first-ever global map of gene expression, shows the 5,372 samples as dots colour-coded for the six major clusters identified by comparing gene expression profiles. The left and right panels of the figure are projections of the same three-dimensional shape viewed from two different perspectives. Credit: Brazma/EMBL.

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The analysis used data collected from 163 laboratories worldwide involving 5,372 human samples from various tissues, cell types and diseases. Most transcriptomics experiments compare gene expression in only a few cell types or conditions and although technically challenging, integrating this data on a large-scale has created a new way for scientists to explore gene expression. The analysis is visualised as a map subdividing the human gene expression space into six distinct major groups or 'continents'.

The continents emerged by grouping samples with similar gene expression signatures. This established the identity of the six groups: brain; muscle; hematopoietic (blood related); healthy and tumour solid tissues; cell lines derived from solid tissues; and partially differentiated cells. By visualising these subsets in 3D, comparisons can be made on the degree of similarity in the gene expression profiles on each grouping. For example, analysis of the continents showed that cell lines are usually more similar to each other than to their tissue of origin.

A new bioinformatics service allowing anyone to explore this expression map has been developed by the European Bioinformatics Institute as part of the ArrayExpress Gene Expression Atlas resource.

Reference:
A global map of human gene expression
Lukk, M., Kapushesky, M., Nikkilä, J., Parkinson, P., Goncalves, A., Huber, W., Ukkonen, E. & Brazma, A.
Nature Biotechnology, 8 April 2010, doi:10.1038/nbt0410-322
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Movies for the Human Genome

EMBL scientists identify the genes involved in cell division in humans
Thursday, 01 April 2010

Name a human gene, and you'll find a movie online showing you what happens to cells when it is switched off. This is the resource that researchers at the European Molecular Biology Laboratory (EMBL) in Heidelberg, Germany, and their collaborators in the Mitocheck consortium are making freely available, as the result of a study in which they have identified the genes involved in mitosis – the most common form of cell division – in humans. Published today in Nature, their work begins to unravel the molecular workings of one of the most fundamental processes of life: how one cell becomes two.

 "Without mitosis, nothing happens in life, really", says Jan Ellenberg, who led the study at EMBL, "and when mitosis goes wrong, you get defects like cancer."

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Normal Cell Division. When no genes are silenced, cell division occurs normally, with each cell giving rise to two. This video was generated within the Mitocheck consortium, whose dataset containing more videos can be found online. Credit: Thomas Walter/EMBL.

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Of the 22,000 genes in each human cell, almost 600 play a part in mitosis, Ellenberg and colleagues found. To uncover which genes are involved in this process, the scientists developed a new method using high-throughput imaging of living cells. They silenced, or inactivated, each of the 22,000 human genes one by one in a different set of cells, and filmed those cells for 48 hours under a microscope. This generated almost 200,000 time-lapse movies of mitosis. Having a person – or even a group of people – process such vast amounts of information would be almost impossible, so the scientists created a new computer program that analyses the footage and automatically detects what characteristic defects cells display, and in what order. By grouping genes with similar effects – for instance, genes which when inactivated led to cells with 2 nuclei instead of one, after division – they were able to identify genes involved in mitosis, which they confirmed with further experimental assays.

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Silencing Gene Hinders Cell Division. When a gene called OGG1 is silenced, cell division starts normally, but then the daughter-cells are unable to separate from each other. The result: individual cells (green) with more than one nucleus (red). Credit: Thomas Walter & Jutta Bulkescher / EMBL.

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"The end result is that we now have a very rich resource for the scientific community, as we're making all the movies and all the analysis data freely available online," Ellenberg emphasises.

"Scientists can go to the website, type in the name of their favourite gene, and watch what happens when it is silenced; they can find out what other genes have similar effects – all in a few mouse clicks, instead of months or years of work in the lab!"

But mitosis is not solved yet, the scientists say. They have yet to uncover exactly how these genes act at the molecular level – a task that will be tackled by a follow-up project called Mitosys. All data from this follow-up work will also be made freely available online, creating what Ellenberg describes as a 'one-stop-shop' for mitosis research.

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This image of a dividing cell is composed of several microscopy images of human cells in which different individual genes were silenced. The images are placed according to genes’ effects: images for genes that affect chromosomes make up the chromosomes (red), while the mitotic spindle (green) is composed of images for genes that affect it. Credit: Thomas Walter & Jutta Bulkescher / EMBL.

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In the mean time, the new methodology the EMBL scientists developed to silence all of an organism's genes in a fast and systematic manner is itself proving a boon to the scientific community.

"A year after we developed these new siRNA microarrays," says Rainer Pepperkok, who led the method's development at EMBL, "they're already in use by over 10 research groups from across Europe."

The current study looked at HeLa cells, a widely studied line of cancer cells. Now that they have narrowed the search from a daunting 22,000 to a more manageable 600 genes, the scientists would like to investigate how these same genes act in other cancers and in healthy cells, as such comparisons could help to identify markers which could be used for diagnosis or to help make better-informed treatment decisions.

The study was carried out as part of the Mitocheck consortium, coordinated by Jan-Michael Peters at the Research Institute of Molecular Pathology in Vienna, Austria, and the data is available at http://www.mitocheck.org/.

References:
Phenotypic profiling of the human genome by time-lapse microscopy reveals cell division genes
Beate Neumann, Thomas Walter, Jean-Karim Hériché, Jutta Bulkescher, Holger Erfle, Christian Conrad, Phill Rogers, Ina Poser, Michael Held, Urban Liebel, Cihan Cetin, Frank Sieckmann, Gregoire Pau, Rolf Kabbe, Annelie Wünsche, Venkata Satagopam, Michael H. A. Schmitz, Catherine Chapuis, Daniel W. Gerlich, Reinhard Schneider, Roland Eils, Wolfgang Huber, Jan-Michael Peters, Anthony A. Hyman, Richard Durbin, Rainer Pepperkok & Jan Ellenberg
Nature, 1 April 2010, doi:10.1038/nature08869

Systematic Localization and Purification of Human Protein Complexes Identifies Chromosome Segregation Proteins
Hutchins et al.
Science Express, published online 1 April 2010
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RNA Research Strategy for Europe Takes Shape

"Consensus Conference" organised by ESF Sunday, 08 March 2009 Research into RNA, a molecule found in every cell of our bodies, could lead to remarkable advances in the treatment of diseases such as cancer and diabetes, a meeting organised by the European Science Foundation was told. The conference, held the institute of Parasitología y Biomedicina "López-Neyra", CSIC in Granada, Spain, on 23 February 2009, was part of an ESF initiative to develop a coherent strategy for RNA research in Europe in recognition of the potential of RNA to result in new approaches to treating human diseases. For many years it was believed that RNA's sole function in cells was to transmit genetic information from DNA during the manufacture of proteins – the cell's workhorse molecules. However, in recent years it has become clear that RNA has many functions that are more sophisticated and that there are more types of RNA than previously known. The field exploded into activity with the discovery in 1998 by US researchers Andrew Fire and Craig Mello of a phenomenon called RNA interference, meaning that genes can be 'silenced' by RNA. This discovery, for which Fire and Mello were awarded the Nobel Prize in 2006, revolutionised the way scientists think about how genetic information is controlled in cells, and has opened the possibility of using gene silencing as a therapy where rogue genes cause disease. "Research into RNA has great promise for both basic science and biotechnology and medicine," said the meeting's chairperson, Professor Lars Thelander of Umeå University in Sweden. "Most pharmaceutical companies now have RNA projects, but the field is still in its early days and it could be another ten years before we see products appearing in the clinics." Professor Thomas Cech of the Howard Hughes Medical Institute in the US told the meeting how he discovered that RNA could also act as a biological catalyst – something that it was previously thought was the preserve of proteins representing a wonderful example of the versatility of RNA function. The discovery gave rise to new ideas about how life on Earth might have started and resulted in Professor Cech being awarded a Nobel Prize in 1989. The Granada "Consensus Conference" was organised by ESF as part of a 'Forward Look' entitled 'RNA World: a new frontier in biomedical research' aimed at developing a strategy for research in RNA over the next ten years. Three earlier workshops had examined various aspects of RNA research to identify where gaps in our knowledge lie and what is required to plug these gaps and fulfil the promise that RNA holds. Forward Looks are a key part of ESF's work, examining important areas of science and technology in consultation with leading scientists and policy makers to develop a strategic framework for research. A Forward Look report on RNA research is due to be published later this year, detailing the scientific questions that need to be answered and giving politicians and policy makers the information they need when deciding where to direct research funding to ensure that Europe remains globally competitive in this key area of emerging science. ......... ZenMaster

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The Beauty of Cell Division

The Beauty of Cell Division Sunday, 07 December 2008 Cell division is one of the most fundamental processes of life. It explains how one cell can give rise to an organism of several million cells, it determines the shape of different life forms and it underpins our body's capacity to heal when injured. Often we only notice how important cell division really is when it goes wrong and results in cancer or other diseases. But apart from being crucial for biology, cell division is also a very beautiful process as these images taken by Joël Beadouin in the group of Jan Ellenberg at the European Molecular Biology Laboratory show. The pictures were taken through a confocal microscope and show the different steps of cell division in a human cell magnified approximately 650 times.

Credit: Joël Beadouin, EMBL

From left to right: When not dividing, the genetic material of a cell (DNA in blue) is found loosely in the nucleus, where a second copy of it is made in preparation for division. At the onset of division, the DNA condenses into distinct chromosomes, the nucleus breaks down and protein filaments called microtubules (green) form a spindle apparatus. The spindle aligns the chromosomes in the middle of the cell and in the next step pulls a copy of each chromosome towards the opposite poles of the cell. After this division of the genetic material, or mitosis, is completed, the rest of the cell divides. A band formed by another type of protein filaments, called actin (red), squeezes the mother cell in half to create two identical daughters. The whole process takes approximately 90 minutes. The images were taken as part of Mitocheck, an international research project funded by the European Commission that tries to identify and characterize all the genes necessary for cell division in human cells. ......... ZenMaster

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A Little Wine Boosts Omega-3 in the Body

Researchers find a novel mechanism for a healthier heart Friday, 05 December 2008 Results from the European study IMMIDIET show that moderate wine intake is associated with higher levels of omega-3 fatty acids considered as protective against coronary heart disease Moderate alcohol intake is associated with higher levels of omega-3 fatty acids in plasma and red blood cells. This is the major finding of the European study IMMIDIET that will be published in the January issue of the American Journal of Clinical Nutrition, an official publication of the American Society for Nutrition. The study suggests that wine does better than other alcoholic drinks. This effect could be ascribed to compounds other than alcohol itself, representing a key to understand the mechanism lying behind the heart protection observed in moderate wine drinkers. The IMMIDIET study examined 1,604 citizens from three geographical areas: south-west London in England, Limburg in Belgium and Abruzzo in Italy. Thanks to a close cooperation with General Practitioners of these areas, all participants underwent a comprehensive medical examination, including a one year recall food frequency questionnaire to assess their dietary intake, alcohol consumption included. Omega-3 fatty acids, mainly derived from fish, are considered as protective against coronary heart disease and sudden cardiac death, thus their high blood concentration is definitely good for our health. Now European researchers found that moderate alcohol drinking acts like a 'trigger', boosting the amount of omega-3 fatty acids in our body. "Several studies have shown that moderate alcohol consumption, including wine, is associated with protection against coronary heart disease and ischemic stroke,” says Romina di Giuseppe, lead author of the study, from the Research Laboratories at Catholic University of Campobasso. “Although the mechanisms are not completely defined, there was some evidence that alcohol intake might influence the metabolism of essential polyunsaturated fatty acids, as omega-3. That is exactly what we found in our population study. People drinking moderate amounts of alcohol, one drink a day for women and two for men, had higher concentration of omega-3 fatty acids in plasma and red blood cells independently of their fish intake". However important these results appear to be, the best is yet to come. Researchers from Catholic University of Campobasso, in Italy, and from University of Grenoble, in France, turned their attention on the variety of alcoholic beverages consumed in order to see whether the high levels of omega-3 fatty acids detected might be ascribed to alcohol itself or to other substances. "From our previous studies we know that association between wine drinking and increased concentration of omega-3 fatty acids have been observed,” says Michel de Lorgeril, from the University of Grenoble, partner of the IMMIDIET project and co-leader of the study. “Nevertheless, it was not possible to separate the effects of wine from those of beer or spirits. Our study of 3 populations with different dietary habits and different consumption of alcoholic beverages types allowed us to explore this aspect." "Analysis carried out on different alcoholic beverages,” argues Licia Iacoviello coordinator of the IMMIDIET study at Catholic University of Campobasso, “showed that the association between alcohol and omega-3 fatty acids was present in both wine drinkers and beer or spirits drinkers. However, the association was stronger between wine drinking and omega-3 fatty acids levels. This suggests that components of wine other than alcohol is associated with omega-3 fatty acids concentration. We may guess this effect can be ascribed to polyphenols". Polyphenols are naturally occurring compounds contained in a different variety of food and beverages, such as wine. Due to their strong antioxidant activity, they are able to reduce oxidation processes caused by free radicals. "We consider these data to be a major finding," de Lorgeril concludes, "opening a new window in the field of cardiovascular prevention. Beyond the alcohol issue, our results raise crucial questions regarding the effects of polyphenols on lipids (both in blood and cell membranes) and possibly of lipids on polyphenols". About The IMMIDIET study: Funded by the European Union under Key Action 1: Food, Nutrition and Health QLK1-CT-2000-00100, IMMIDIET aims to acquire fundamental knowledge in the field of cardiovascular disease, especially regarding the interaction between genetics and lifestyle. At the core of the study, there is an important episode of Italian migration: Belgium, a country that became the new home for thousands of Italians, mostly from the Abruzzo region, who came to work in the mines. Many of those emigrants did not come back to Italy but remained in their new country. Some of them married a Belgian partner. Their genes remained the same, of course, but how much "Italy" is still there in their diet? And how much did they transmit it to their spouses? Moreover, how many Italian emigrants assimilate dietary habits of the country in which they were guests? In this framework, the role of genetic factors and lifestyle can be assessed to explore new ways in prevention of cardiovascular diseases. To carry on the research, married couples have been recruited in three European areas: South-East London in England, Limburg in Belgium and Abruzzo in Italy. In the first phase of the study the couples involved were formed by people from the same area, Italians married with Italians (in the Abruzzo region), Belgians married with Belgians (in the Limburg area) and English married with English (in the South-East part of London)". The second phase of IMMIDIET recruited mixed Italian–Belgian couples to understand if, acquiring dietary habits from Abruzzo, the Belgian partner changed his own risk regarding heart diseases. ......... ZenMaster

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Tissue Engineering for Transplanting from Own Stem Cells II

Use of several types of adult stem cells grow new trachea Thursday, 20 November 2008 The first tissue-engineered trachea (windpipe), utilising the patient's own stem cells, has been successfully transplanted into a young woman with a failing airway. The bioengineered trachea immediately provided the patient with a normally functioning airway, thereby saving her life. These remarkable results provide crucial new evidence that adult stem cells, combined with biologically compatible materials, can offer genuine solutions to other serious illnesses. In particular, the successful outcome shows it is possible to produce a tissue-engineered airway with mechanical properties that permit normal breathing and which is free from the risks of rejection seen with conventional transplanted organs. The patient has not developed antibodies to her graft, despite not taking any immunosuppressive drugs. Lung function tests performed two months after the operation were all at the better end of the normal range for a young woman. The pan-European team from the universities of Barcelona, Bristol, Padua and Milan report on this pioneering work in an article published early online and in an upcoming edition of The Lancet. The loss of a normal airway is devastating, but previous attempts to replace large airways have met serious problems. The 30-year-old mother of two, suffering from collapsed airways following a severe case of TB, was hospitalised in March 2008 with acute shortness of breath rendering her unable to carry out simple domestic duties or care for her children. The only conventional option remaining was a major operation to remove her left lung, which carries a risk of complications and a high mortality rate. Based on successful laboratory work previously performed by the team, and given the urgency of the situation, it was proposed that the lower trachea and the tube to the patient's left lung (bronchus) should be replaced with a bioengineered airway based on the scaffold of a human trachea. A seven-centimetre tracheal segment was donated by a 51-year-old transplant donor who had died of cerebral haemorrhage. Spain has a policy of assumed consent for organ donation. Using a new technique developed in Padua University, the trachea was de-cellularised over a six-week period so that no donor cells remained. Stem cells were obtained from the recipient's own bone marrow, grown into a large population in Professor Martin Birchall's lab at the University of Bristol, and matured into cartilage cells (chondrocytes) using an adapted method originally devised for treating osteoarthritis by Professor Anthony Hollander at the University of Bristol. The donor trachea was then seeded with chondrocytes on the outside, using a novel bioreactor which incubates cells, developed at the Politecnico di Milano, Italy, allowing them to migrate into the tissue under conditions ideal for each individual cell type. In order to replicate the lining of the trachea, epithelial cells were seeded onto the inside of the trachea using the same bioreactor. Four days after seeding, the graft was used to replace the patient's left main bronchus. Professor Paolo Macchiarini of the University of Barcelona performed the operation in June 2008 at the Hospital Clínic, Barcelona. Professor Macchiarini, lead author on the paper, said: "We are terribly excited by these results. Just four days after transplantation the graft was almost indistinguishable from adjacent normal bronchi. After one month, a biopsy elicited local bleeding, indicating that the blood vessels had already grown back successfully". Martin Birchall, Professor of Surgery at the University of Bristol, added: "Surgeons can now start to see and understand the very real potential for adult stem cells and tissue engineering to radically improve their ability to treat patients with serious diseases. We believe this success has proved that we are on the verge of a new age in surgical care". Anthony Hollander, Arthritis Research Campaign Professor of Rheumatology and Tissue Engineering at the University of Bristol, concurred: "This successful treatment manifestly demonstrates the potential of adult stem cells to save lives". The patient, Claudia Castillo, a young woman from Colombia but now living in Spain, had no complications from the operation and was discharged from hospital on the tenth post-operative day. She has remained well since and has a normal quality of life. She is able to care for her children, walk up two flights of stairs and occasionally go out dancing in the evenings. She said: "Above all I would like to thank Dr. Macchiarini and his medical team who did the research, for the time and dedication they devoted to my case to make sure that everything turned out alright." Reference: Clinical transplantation of a tissue-engineered airway Paolo Macchiarini, Philipp Jungebluth, Tetsuhiko Go, M Adelaide Asnaghi, Louisa E Rees, Tristan A Cogan, Amanda Dodson, Jaume Martorell, Silvia Bellini, Pier Paolo Parnigotto, Sally C Dickinson, Anthony P Hollander, Sara Mantero, Maria Teresa Conconi, Martin A Birchall The Lancet, Early Online Publication, 19 November 2008, doi:10.1016/S0140-6736(08)61598-6 See also: Tissue Engineering for Transplanting from Own Stem Cells I CellNEWS - Thursday, 20 November 2008 ......... ZenMaster

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Tissue Engineering for Transplanting from Own Stem Cells I

First tracheal transplant without immunosuppression Thursday, 20 November 2008 Summary:

• Tissue engineering has made possible this doubly innovative operation – the first trachea transplant and the first tissue transplant to be performed without the need for immunosuppression.
• Professor Paolo Macchiarini, Head of the thoracic surgery department of Hospital Clínic of Barcelona, has led the basic research and the international team formed by the universities of Bristol, Padua and Milan, who contributed to this success.
• The transplanted tissue is a hybrid from a donor that was re-populated with stem and epithelial cells from the recipient. Five months later, Claudia Castillo, who required the operation to save a lung following tuberculosis, is in perfect health.

After 4 years of going from consultation to consultation, Claudia Castillo finally found a solution to her respiratory problems. The young Colombian woman suffered from a cough that took a long time to be diagnosed as tuberculosis. She arrived at Hospital Clínic of Barcelona with complications and there, she met Professor Paolo Macchiarini, Head of the Thoracic Surgery Department, who led the international team that made possible the first trachea transplant and the first tissue transplant without immunosuppression. She underwent an operation on the upper part of the trachea but nothing could be done to repair the blockage in the left lung. The infection had led to a severe collapse just before the branch of the trachea and this obstruction prevented air from reaching the lung. The only treatment option at the time involved removing the affected lung. As the young mother of two children, removing the lung would have considerably reduced quality of life for Claudia Castillo. In March 2008, her situation worsened to the point where she was unable to carry out domestic chores or look after her children, so intervention became urgent. In June, after obtaining authorization from the ethics committee of Hospital Clínic of Barcelona and from the Catalan Transplant Organization (OCATT), the first trachea transplant and the first tissue transplant of any kind without immunosuppression took place. The study, published online on Wednesday by the journal The Lancet, with Professor Paolo Macchiarini as the principal author, together with his colleagues Dr. Philip Jungebluth, Dr. Tetsuhiko Go and Dr. Jaume Martorell, presents the details of this transplant – the first treatment alternative for treating the collapsed trachea that the patient was suffering from. The technique consists of depleting the trachea to be transplanted of the donor's cells and repopulating it with cells from the recipient before the operation. Thus, thanks to tissue bioengineering, the donor trachea becomes a hybrid that the recipient's body identifies as its own, thereby making immunosuppression unnecessary. The transplant and most of the processes involved were carried out at Hospital Clínic of Barcelona, but this would have been impossible without the collaboration of the University of Bristol (UK), the University of Padua (Italy) and the University of Milan (Italy). Professor Paolo Macchiarini led the prior basic research. The process of preparing the trachea requires many cycles of washing to eliminate all the donor cells – many more than those suggested by the basic research. The tissue was a 7-cm segment of trachea from a 51-year-old donor who had died from brain haemorrhage. The team of Dr. Maria T. Conconi at the University of Padua (Italy) confirmed that, after 25 washing cycles, the trachea treated at Hospital Clínic was free from donor antigens – the molecules that would cause the tissue to be rejected by the recipient. Meanwhile, at the University of Bristol, the teams of Professor Martin Birchall and Professor Anthony Hollander cultivated the recipient's cells that would later be introduced into the trachea. These cells were epithelial cells taken from the trachea and cartilage cells (chondrocytes), differentiated from stem cells taken from the patient's bone marrow. This technique was initially designed to treat cases of osteoarthritis. Back at Hospital Clínic, the team of Professor Paolo Macchiarini introduced these cells into the trachea using a bioreactor designed by the team of Dr. Sandra Mantero at the University of Milan. The epithelial cells were inserted into the inner surface of the trachea and the chondrocytes covered the outer surface. The donor tissue thus became a hybrid very similar to new tissue from the patient herself. The operation was performed 4 days later at Hospital Clínic, where the thoracic surgery team extracted the damaged section of trachea and replaced it with the new trachea. This pioneering operation was not without question marks but if anything had gone wrong, it would have been changed to a lung-resection operation – the classical treatment choice. Thanks to the skill of the surgeons and the huge international effort, the operation was a success. Five months later, the lung that had been so long out of use was providing normal respiration. This innovation in biomedicine and surgery may become an alternative for diseases of the upper airways, such as congenital deformities or primary tumours, which cannot currently be treated using conventional surgical techniques. The clinical application of stem cell cultures and the prevention of the problems deriving from immunosuppression are a milestone in the history of transplantation. There are already some cases being studied that may benefit from the new technique and research continues into improving the process. If all goes well, Claudia Castillo will be just the first patient to benefit from a new advance led by researchers from Hospital Clínic of Barcelona. Reference: Clinical transplantation of a tissue-engineered airway Paolo Macchiarini, Philipp Jungebluth, Tetsuhiko Go, M Adelaide Asnaghi, Louisa E Rees, Tristan A Cogan, Amanda Dodson, Jaume Martorell, Silvia Bellini, Pier Paolo Parnigotto, Sally C Dickinson, Anthony P Hollander, Sara Mantero, Maria Teresa Conconi, Martin A Birchall The Lancet, Early Online Publication, 19 November 2008, doi:10.1016/S0140-6736(08)61598-6

See also: Tissue Engineering for Transplanting from Own Stem Cells II CellNEWS - Thursday, 20 November 2008 ......... ZenMaster

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Man's Best Friend Recruited in the Hunt for Disease Genes

Man's Best Friend Recruited in the Hunt for Disease Genes Thursday, 16 October 2008 For centuries man has had a uniquely close relationship with dogs – as a working animal, for security and, perhaps most importantly, for companionship. Now, dogs are taking on a new role – they are helping in the hunt for genetic mutations that lead to diseases in humans. "Dogs get very similar diseases to humans," said Kerstin Lindblad-Toh of Institute of Medical Biochemistry and Microbiology, Uppsala University in Sweden and the Broad Institute of MIT and Harvard, Cambridge, Massachusetts. "If you ask a dog owner what sort of conditions their pets get, they will say cancer, allergies, eye diseases." Lindblad-Toh was speaking at the European Science Foundation's 3rd Functional Genomics Conference, held in Innsbruck, Austria, on 1-4 October. Functional genomics describes the way in which genes and their products, proteins, interact together in complex networks in living cells. If these interactions are abnormal, diseases can result. The Innsbruck meeting brought together more than 450 scientists from across Europe to discuss recent advances in the role of functional genomics in disease. Many canine diseases could share the same genetic basis in humans and dogs, Lindblad-Toh told the conference, and because dogs have been bred into clear isolated populations – the different breeds – it is often easier to detect a genetic flaw that leads to a disease than it is in humans. Once the rogue gene has been found in the dog, it could make it easier look for mutations in the same gene in man. "For example we have found genetic mutation that results in a condition called day blindness that can affect dachshunds," Lindblad-Toh said. A similar condition can arise in humans, and analysis of the mutated protein in the dog is providing new information about the disease in man. The team is also looking at genes associated with cancer of the blood vessels to which golden retrievers are prone. A new European consortium has been set up called LUPA, where twenty veterinary schools from 12 countries spread across Europe will work together to collect 10,000 DNA samples from purebred dogs, comparing healthy animals with those affected by similar diseases as human. The analysis of the genome of affected dogs compared to healthy ones of the same breed will lead to the identification of genes implied in the mechanisms of these diseases. The four-year project aims initially to pinpoint genetic markers for dog diseases and help to reduce the high level of inherited disease in purebred dogs. The identification of these genes implied in disease development will help to understand the mechanisms and pathways of the pathology. For example in Sweden, more than one-third of English Springer Spaniels are diagnosed with mammary tumours, analogous to breast cancers in humans. An increased risk for malignant mammary tumours has been reported also in other breeds, including Cocker Spaniels, German Shepherds and Boxers, suggesting that these breeds may carry genetic risk factors for this type of cancer. If the genes implicated in the disease can be singled out this could provide a new opportunity to improve prevention, diagnosis and treatment of human breast cancer. "We want to find a lot of risk factors and bring them back to human patients over the next few years," Lindblad-Toh said. Lupa was the female wolf (Canis lupus) that according to Roman mythology was nurturing the twins Romulus and Remus, founders of Rome. ......... ZenMaster

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Unpicking the Complexity of Human Disease

Impressive advances in our understanding of the genetic basis of disease were outlined at the 3rd ESF Functional Genomics Conference in Innsbruck, Austria Wednesday, 15 October 2008 The mysteries of the human genome are slowly being revealed – but the more we uncover the more complicated the picture becomes. This was one key message to emerge from the European Science Foundation's 3rd Functional Genomics Conference held in Innsbruck, Austria, on 1-4 October. Functional genomics describes the way in which genes and their products, proteins, interact together in complex networks in living cells. If these interactions are abnormal, diseases can result. "The human genome is just a string of letters which has to be interpreted so that we can understand the function of the genes," said Mike Taussig of Babraham Bioscience Technologies in Cambridge, UK, who organised the conference, which focused on the role of functional genomics in disease. More than 450 scientists from across Europe were told of new developments in research ranging from pinpointing genes involved in diabetes and cancer to using the genetic sequences of different breeds of dog to throw light on human diseases. "What we have tried to do is bring together genome knowledge as it is now, the work which has been done, what it means in functional terms and where it affects our susceptibility to disease," Taussig said. "Conferences like these are important because we try to cover a broad field – there are so many aspects of genomics that it would be impossible to encompass everything in a single lecture. It is very useful for researchers who want to improve their general view of the field, because when you are immersed in one specialty you often do not appreciate how well connected everything is." Dr Patrik Kolar, head of the unit for genomics and systems biology in the European Commission's research directorate, said: "Functional genomics and systems biology is an important and integral part of our health research programme because an understanding of these basic biological processes has huge potential and real applications for understanding disease, and when you understand disease you can design new drugs." "This kind of conference is one of the things that brings together the European community in functional genomics and I am really happy to see that most of the participants are from collaborative projects funded though our Framework programme," Dr Kolar added. Professor Mark McCarthy of the University of Oxford in the UK, who is searching for genes involved in type 2 diabetes, illustrated the unexpected complexity of the role of genes in disease. Here, so-called genome-wide scans, which compare the genetic profiles of healthy people with those who have the disease, have so far revealed around 20 individual gene mutations that can be present in people with type 2 diabetes. However, these variants explain only a small proportion of people's overall susceptibility to the condition. "If you look at the variants we are finding from really large sample sizes, the effects are pretty small," McCarthy told the conference. "For diabetes, weight and age are still better predictors of risk than the gene profile. So, on the one hand we are happy that we have found more signals that we might have imagined, but on the other hand, we are disappointed because we are explaining so little of the variance. There is much work to be done to turn these association signals into function and mechanism." Mutations in cancer genes have also turned out to be far more complicated than people might have first suspected. Professor Mike Stratton, head of the Cancer Genome Project at the Wellcome Trust Sanger Institute in Cambridge, UK, led the team that mapped and identified the high-risk breast cancer susceptibility gene BRCA2. His group is searching for particular types of gene mutations in cancer cells, and is revealing new insights into a class of mutation called rearrangement, where one gene breaks and is fused to another. This rearrangement process could result in the creation of a rogue protein that promotes cancer. Until recently, it has been difficult to study rearrangement mutations because technologies have been lacking. "For many years we have wanted a screen which would allow us to extract rearranged parts of the cancer genome and make a catalogue," Stratton told conference delegates. Techniques have now been developed that are allowing researchers to pinpoint rearrangements and look at them in detail. "New sequencing technologies are enabling us to look at a much larger number of rearrangements, allowing us to do genome-wide screens to identify fusion genes that could be cancer genes," Stratton said. "It turns out there is a lot of complexity in these rearrangements that we would not anticipated before we started." For example it is becoming clear that while there are many more rearrangement mutations than people first thought, the majority of these seem to be effectively harmless, or 'passenger' mutations. The significant mutations are the 'drivers', and these are much more elusive to track down. Meanwhile Dr Kerstin Lindblad-Toh of Uppsala University in Sweden and the Broad Institute of MIT and Harvard, Cambridge, Massachusetts, is analysing gene mutations in different breeds of dog to throw light on human diseases. Because dogs have been reared as distinct breeds with clear isolated populations, it is often easier to detect a genetic flaw than it is in humans, and dogs are susceptible to many similar diseases that occur in man. Professor Olli Kallioniemi's team at the Institute for Molecular Medicine in Finland is working on innovative ways to discover the effects of small strands of RNA, called small interfering RNAs (siRNAs), which can 'silence' genes and are showing promise in the fight against diseases such as cancer. The Finnish researchers have developed new high-throughput techniques for testing thousands of different siRNAs on living cells in one go. "This is a new cell array screening platform which we think has great potential for showing real utility in biological experiments," Kallioniemi told the meeting. Professor Patrik Brundin of Lund University in Sweden leads a team that is learning how best to repair the brain of people with neurodegenerative disorders such as Parkinson's disease, where a part of the brain called the substantia nigra degenerates, leading to slow movement and tremors. Brundin told the meeting that his university has over the past 20 years transplanted brain tissue into 18 patients with Parkinson's, whose condition improved markedly and remained stable for many years. However, certain unexplained side effects have arisen, including uncontrollable movements. "Nigral transplants have clearly worked well in select cases, but the technique needs refinement and is difficult to perform in large series of patients," Brundin said. One key issue is a safe and sustainable supply of tissue, and embryonic stem cells could hold promise. However, Brundin warned that many hurdles remain to be overcome. "Today some people are saying that you can do this with stems cells, but stem cell transplantation to the brain is currently science fiction and should remain so for the moment – there are many challenges before we can do clinical trials." In all, leading scientists, from Europe and the US, gave twelve key lectures at the meeting. In addition, there were more than 40 symposia, with topics ranging from the role of proteins in ageing to new ways to disable viruses that cause influenza. ......... ZenMaster

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Zebrafish Development Tracked Cell by Cell

Provides the first complete developmental blueprint of a vertebrate Thursday, 09 October 2008 Researchers at the European Molecular Biology Laboratory (EMBL) have generated a digital zebrafish embryo - the first complete developmental blueprint of a vertebrate. With a newly developed microscope scientists could for the first time track all cells for the first 24 hours in the life of a zebrafish. The data was reconstructed into a three-dimensional, digital representation of the embryo. The study, published in the current online issue of Science, grants many new insights into embryonic development. Movies of the digital embryo and the underlying database of millions of cell positions, divisions and tracks will be made publicly available to provide a novel resource for research and scientific training. To get from one cell to a complex organism, cells have to divide, travel around the body and arrange intricate shapes and specialized tissues. The best way to understand these dynamic processes is to look at what happens in the first few hours of life in every part of an embryo. While this is possible with invertebrates with a few hundred cells, like worms for example, it has so far been impossible to achieve for vertebrates.

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The montage shows the zebrafish digital embryo (left halves, colours encode movement directions of cells) and the microscopy data (right halves) at different time points in zebrafish development. Credit: Philipp Keller, EMBL.

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"Imagine following all inhabitants of a town over the course of one day using a telescope in space. This comes close to tracking the 10 thousands of cells that make up a vertebrate embryo – only that the cells move in three dimensions," says Philipp Keller. Together with Annette Schmidt he carried out the research in the labs of Jochen Wittbrodt and Ernst Stelzer at EMBL. Two newly developed technologies were key to the scientists' interdisciplinary approach to tracking a living zebrafish embryo from the single cell stage to 20,000 cells: a Digital Scanned Laser Light Sheet Microscope, that scans a living organism with a sheet of light along many different directions so that the computer can assemble a complete 3D image, and a large-scale computing pipeline operated at the Karlsruhe Institute of Technology. Zebrafish is a widely used model organism that shares many features with higher vertebrates. Taking more than 400,000 images per embryo the interdisciplinary team generated terabytes of data on cell positions, movements and divisions that were reassembled into a digital 3D representation of the complete developing embryo. "The digital embryo is like Google EarthTM for embryonic development. It gives an overview of everything that happens in the first 24 hours and allows you to zoom in on all cellular and even sub-cellular details," says Jochen Wittbrodt, who has recently moved from EMBL to the University of Heidelberg and the Karlsruhe Institute of Technology. New insights provided by the digital embryo include: fundamental cell movements that later on form the heart and other organs are different than previously thought and the position of the head-tail body axes of the zebrafish is induced early on by signals deposited in the egg by the mother. The new microscopy technology is also applicable to mice, chickens and frogs. A comparison of digital embryos of these species is likely to provide crucial insights into basic developmental principles and their conservation during evolution.

All movies can be downloaded as Apple QuickTime and DiVX versions on the public digital embryo repository website: http://www.embl-heidelberg.de/digitalembryo. Reference: Reconstruction of Zebrafish Early Embryonic Development by Scanned Light Sheet Microscopy Philipp J. Keller, Annette D. Schmidt, Joachim Wittbrodt, Ernst H. K. Stelzer Science, Published Online October 9, 2008, DOI: 10.1126/science.1162493 ......... ZenMaster

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EU Call for Ban on Animal Cloning for Food

EU Call for Ban on Animal Cloning for Food Wednesday, 03 September 2008 The European Parliament members called for a ban in the EU on the cloning of animals for food supply. MEPs also urged an embargo on imports of cloned animals, their offspring and products derived from these sources. In a resolution adopted by 622 MEPs in favour, 32 against and 25 abstentions, the House calls on the Commission "to submit proposals prohibiting for food supply purposes:

• the cloning of animals,
• the farming of cloned animals or their offspring,
• the placing on the market of meat or dairy products derived from cloned animals or their offspring, and
• the importing of cloned animals, their offspring, semen and embryos from cloned animals or their offspring, and meat or dairy products derived from cloned animals or their offspring."

Animal health problems and risks for the European quality model The text refers to the health, welfare and higher mortality problems of cloned animals and their surrogate dams recently highlighted by several groups of European experts, including the European Food Safety Authority (EFSA) and the European Group on Ethics. MEPs also stress that cloning would significantly reduce genetic diversity within livestock populations, increasing the possibility of whole herds being decimated by diseases to which they are susceptible. In addition, the European Parliament fears that use of cloning techniques would harm the image of the European agricultural model, which is based on product quality, environment-friendly principles and respect for stringent animal welfare conditions. They also point out that Directive 98/58/EC on the protection of farm animals bans natural or artificial procedures which are likely to cause suffering or injury to any of the animals concerned. Currently no products derived from cloned animals are sold in Europe or the rest of the world. However, experts believe that such products could reach the market by 2010. A moratorium on the sale of this type of product, introduced in the USA in 2001, was challenged by the US Food and Drug Administration, which concluded in January that meat and milk from clones of cattle, pigs and goats and their offspring are as reliable as those of traditionally bred animals. European Commission quizzed During their debate yesterday evening, MEPs quizzed the European Commission on its position and its plans as regards animal cloning. "Not only is it a case of food safety, we in Europe believe that we are producing food quality products", EP Agriculture Committee Chairman Neil Parish (EPP-ED, UK) said. "It is also a question of animal welfare and consumer confidence" and there is a "risk of producing less strong and healthy animals". He stressed "we have to look at this seriously". Mr Parish said: "Cloning entails serious health and welfare problems for clones and their surrogate dams; animal health problems come from invasive techniques required to produce a clone; there is the suffering of surrogate dams which carry cloned foetuses, and high levels of ill health and mortality in early life for cloned animals. I call on the Commission to submit proposals prohibiting the cloning of animals in the food supply and the placing of cloned animals on the market in meat and dairy products." Androula Vassiliou, the Commissioner for health and food safety, said that the Commission was closely following scientific developments in this area and "is aware that even though the efficiency of animal cloning has improved over the last years, adverse health effects on animal health and welfare still occur today". The Commission "is now evaluating the necessary steps to be taken" and "takes ethical considerations fully into account", including the opinion of the European Group of Ethics which "advocated that at the moment there are no convincing arguments to justify the production of food from clones and their offspring". "According to global trade rules, imports of food products from third countries might be suspended if they present a serious threat to animal or public health. On the basis of the studies conducted and the opinion of EFSA, the Commission will consider whether restrictions must be imposed", she added. ......... ZenMaster

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Human Ovulation Moment Caught on Camera

Human Ovulation Moment Caught on Camera 
Wednesday, 18 June 2008 

 Fertile women release one or more eggs every month, but until now, only animal ovulation has been recorded in detail. Now, gynaecologist Dr Jacques Donnez spotted the release of a human egg in progress, during a routine hysterectomy. He could film the process in close-up, when the egg was emerging from the ovary. Human eggs are produced by follicles, fluid-filled sacs on the side of the ovary, which, around the time of ovulation, produce a reddish protrusion seen in the pictures. The egg comes from the end of this, surrounded by a jelly-like substance containing supportive cells. The egg itself is only the size of a full-stop, and the whole ovary, which contains many immature eggs, just a couple of inches long. Dr Donnez, from the Catholic University of Louvain, told New Scientist that the pictures would help scientists understand the mechanisms involved in ovulation. He said that some theories had suggested an "explosive" release for the egg, but the ovulation he witnessed took 15 minutes to complete, so the event was progressive.

These images are the first time the event of ovulation in humans has been captured in clear detail. The yellow blob is a protruding egg cell, surrounded by supportive cumulus cells (at the black arrow). The reddish part is the follicle (S), and the pale pink tissue is part of the ovary (F).

Ovulation takes place on the surface of the ovarian tissue.

The egg, surrounded by supportive cumulus cells, is shown emerging from the follicle on the ovary.

After the release from the follicle, the egg travels down the Fallopian tube where it can be fertilised. 
Credit: The above pictures was kindly provided by Prof. Jacques Donnez, at Université Catholique de Louvain, Brussels, Belgium, and Eric Steinmehl, managing editor at “Fertility and Sterility” and the American Society for Reproductive Medicine, Birmingham, Alabama. 

Reference: 
Laparoscopic observation of spontaneous human ovulation 
Jean-Christophe Lousse, M.D., Jacques Donnez, M.D., Ph.D. 
Fertility and Sterility, published online 28 April 2008, doi:10.1016/j.fertnstert.2007.12.049

See also: 
How much is a human egg worth? 
CellNEWS - Wednesday, 21 February 2007 
Most egg cells in a female body die naturally by programmed cell death 
CellNEWS - Tuesday, 24 July 2007 
Back to the question: How much is a human egg worth? 
CellNEWS - Tuesday, 09 October 2007 
.........

ZenMaster

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Heart Derived Stem Cells Develop Into Heart Muscle

A First In Stem Cell Research Wednesday, 23 April 2008 Dutch researchers at University Medical Center Utrecht and the Hubrecht Institute have succeeded in growing large numbers of stem cells from adult human hearts into new heart muscle cells. This is a breakthrough in stem cell research. Until now, it was necessary to use embryonic stem cells to make this happen. The work, which was funded in part by the EU, are published in the latest issue of the journal Stem Cell Research. The stem cells are derived from material left over from open-heart operations. Researchers at UMC Utrecht used a simple method to isolate the stem cells from this material and reproduce them in the laboratory, which they then allowed to develop. The cells grew into fully developed heart muscle cells that contract rhythmically, respond to electrical activity, and react to adrenaline. “We’ve got complete control of this process, and that’s unique,” says principal investigator Prof. Pieter Doevendans. “We’re able to make heart muscle cells in unprecedented quantities, and on top of it they’re all the same. This is good news in terms of treatment, as well as for scientific research and testing of potentially new drugs.” Doevendans will use the cultured heart muscle cells to study things like cardiac arrhythmia (abnormal heart rhythms). Stem cells from the hearts of patients with genetic heart defects can be grown into heart muscle cells in the lab. Researchers can then study the cells responsible for the condition straight away. They can also be used to test new medicines. This could mean that research into genetic heart conditions can move forward at a much faster pace. In the future, new heart muscle cells can likely be used to repair heart tissue damaged during a heart attack. For some time now, it has been known that the heart is a source of stem cells. Although in the past researchers from other countries have succeeded in using these cells to make heart muscle cells, this always required the presence of heart muscle cells from newborn mice or rats in the growth medium. The stem cells discovered by the UMC Utrecht researchers are able to develop on their own. Heart muscle cells can also be made from embryonic stem cells (see e.g. article by Lei Yang et al. below). The disadvantage of this method is that the yield is low, because not all cells develop into muscle cells. Also, the ethical considerations of isolating stem cells from embryos are the subject of controversy. EU support for the research came from the EU-funded SC&CR ('Application and process optimization of human stem cells for myocardium repair') project, which is financed through the 'Life sciences, genomics and biotechnology for health' Thematic Area of the Sixth Framework Programme (FP6) and the Heart Development and Heart Repair project. References: TGF-beta-1 induces efficient differentiation of human cardiomyocyte progenitor cells into functional cardiomyocytes in vitro Marie-José Goumans, Teun P. de Boer, Anke M. Smits, Linda W. van Laake, Patrick van Vliet, Corina H.G. Metz, Tom H. Korfage, K. Peter Kats, Ron Hochstenbach, Gerard Pasterkamp, Marianne C. Verhaar, Marcel A.G. van der Heyden, Dominique de Kleijn, Christine L. Mummery, Toon A.B. van Veen, Joost P.G. Sluijter, Pieter A. Doevendans Stem Cell Research, In Press, Available online 12 March 2008, doi:10.1016/j.scr.2008.02.003 Human cardiovascular progenitor cells develop from a KDR+ embryonic-stem-cell-derived population Lei Yang, Mark H. Soonpaa, Eric D. Adler, Torsten K. Roepke, Steven J. Kattman, Marion Kennedy, Els Henckaerts, Kristina Bonham, Geoffrey W. Abbott, R. Michael Linden, Loren J. Field & Gordon M. Keller Nature , doi:10.1038/nature06894; Published online 23 April 2008 ......... ZenMaster

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