Human Neural Stem Cells Survive Long-term when Transplanted into Primate Brain

Human Neural Stem Cells Survive Long-term when Transplanted into Primate Brain

Wednesday, 23 April 2014

A team of researchers in Korea who transplanted human neural stem cells (hNSCs) into the brains of nonhuman primates and assessed cell survival and differentiation after 22 and 24 months found that the hNSCs had differentiated into neurons at 24 months and did not cause tumours.

The study will be published in a future issue of Cell Transplantation but is currently freely available on-line.

The hNSCs were labelled with magnetic nanoparticles to enable them to be followed by magnetic resonance imaging (MRI). They did not use immunosuppressant’s. According to the researchers, their study is the first to evaluate and show the long-term survival and differentiation of hNSCs without the need for immunosuppression.

The researchers concluded that hNSCs could be of "great value" as a source for cell replacement and gene transfer for the treatment of Parkinson's disease, Huntington's disease, Alzheimer's disease, amyotrophic lateral sclerosis (ALS), spinal cord injury and stroke.

"Stroke is the fourth major cause of death in the US behind heart failure, cancer, and lower respiratory disease," said study co-author Dr. Seung U. Kim of University of British Columbia Hospital's department of neurology in Canada.

"While tissue plasminogen activator (tPA) treatment within three hours after a stroke has shown good outcomes, stem cell therapy has the potential to address the treatment needs of those stroke patients for whom tPA treatment was unavailable or did not help."

Dr. Kim and colleagues in Korea grafted magnetic particle-labelled hNSCs into the brains of laboratory primates and evaluated their performance to assess their survival and differentiation over 24 months. Of particular interest was determining their ability to differentiate into neurons and to determine whether the cells caused tumorigenesis.

"We injected hNSCs into the frontal lobe and the putamen of the monkey brain because they are included in the middle cerebral artery (MCA) territory, which is the main target in the development of the ischemic lesion in animal stroke models," commented Dr. Kim.

"Thus, research on survival and differentiation of hNSCs in the MCA territory should provide more meaningful information to cell transplantation in the MCA occlusion stroke model."

The researchers said that they chose NSCs for transplantation because the existence of multipotent NSCs "has been known in developing rodents and in the human brain with the properties of indefinite growth and multipotent potential to differentiate" into the three major CNS cell types – neurons, astrocytes and oligodendrocytes.

"The results of this study serve as a proof-of-principle and provide evidence that hNSCs transplanted into the non-human primate brain in the absence of immunosuppressant’s can survive and differentiate into neurons," wrote the researchers.

"The study also serves as a preliminary study in our planned preclinical studies of hNSC transplantation in non-human primate stroke models."

"The absence of tumours and differentiation of the transplanted cells into neurons in the absence of immunosuppression after transplantation into non-human primates provides hope that such a therapy could be applicable for use in humans." said Dr. Cesar V. Borlongan, Prof. of Neurosurgery and Director of the Center of Excellence for Aging & Brain Repair at the University of South Florida.

"This is an encouraging study towards the use of NSCs to treat neurodegenerative disorders".

Source: Cell Transplantation Center of Excellence for Aging and Brain Repair

Contact: Robert Miranda

Reference:

Long-term survival and differentiation of human neural stem cells in nonhuman primate brain with no immunosuppression 

Lee, S-R.; Lee, H. J.; Cha, S-H.; Jeong, K-J.; Lee, Y. J.; Jeon, C-Y.; Yi, K. S.; Lim, I.; Cho, Z-H.; Chang, K-T.; Kim, S. U.

Cell Transplant. Appeared or available online: January 29, 2014

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Autologous Transplantation Shows Promising Results for iPS Cell Therapy in Parkinson's Disease

First confirmation in primates

Sunday, 29 September 2013

Autologous and allogeneic transplantation of

iPSC-derived neural cells. Credit: Stem Cell

Reports, Morizane et al. 

A research team led by Professor Jun Takahashi and Assistant Professor Asuka Morizane at the Center for iPS Cell Research and Application (CiRA) at Kyoto University, Japan, has carried out a study to compare the impact of immune response in autologous transplantation (transplantation of cells from the subject's own body) and allogeneic transplantation (transplantation of cells from a different individual of the same species). The researchers used cynomolgus monkeys to carry out transplantation into the brain of neural cells derived from iPS cells. Autologous transplantation was found to produce almost no immune reaction and to result in viable neural cells. By contrast, allogeneic transplantation provoked immune reaction by microglia and lymphocytes.

Parkinson's disease is a progressive and intractable disease of the nervous system in which the loss of dopaminergic neurons in the brain leads to reduced dopamine production, resulting in limb tremor, stiffness causing difficulty in movement, and other symptoms. The therapies applied up till now, based on drugs or electrode treatment, may improve symptoms but have proved unable to halt the depletion of dopaminergic neurons. Hopes have therefore become focused on a therapy with the more radical approach of replacing the lost neural cells through cell transplantation, thereby promoting the formation of new neural pathways to restore brain function. Human iPS cells are looked to as a potential source of the transplant cells.

This is an immunostaining of primate iPSC-

derived neurons on day 39. Green colors shows

dopaminergic neural cells. Credit: Courtesy of

Dr. Asuka Morizane. 

It is hoped that iPS cells will make it possible to use cells derived from the transplant patients themselves to perform autologous transplantation. If autologous transplantation could allow immune reaction to be avoided, it would also make unnecessary the use of immunosuppressant drugs and avert the risk of side-effects caused by immunosuppression. However, the studies of iPS cell-based autologous transplantation carried out so far, which have used a mouse model, have produced no firm conclusion, with immune reaction observed in some studies but not in others. Moreover, these studies did not involve transplantation of differentiated cells derived from iPS cells in a way that mimicked clinical application. There had thus been no studies directly investigating the effect of autologous transplantation and allogeneic transplantation in primates. This study by Dr. Takahashi's group sought to clarify this area by transplanting dopaminergic neurons prepared from iPS cells into the brains of cynomolgus monkeys and comparing the extent of immune reaction between autologous and allogeneic transplantation.

iPS cells prepared from four cynomolgus monkeys were differentiated into dopaminergic neural cells over a period of 28 days and transplanted into the monkeys' brains, which were observed over a period of approximately three months during which no immunosuppressant were used. The study data show that, in primates, autologous transplantation of iPS cell-derived neural cells produces almost no immune reaction and is superior to allogeneic transplantation in terms of immune reaction control and cell viability.

Source: Center for iPS Cell Research and Application - Kyoto University

Contact: Akemi Nakamura

Reference:

Direct Comparison of Autologous and Allogeneic Transplantation of iPSC-Derived Neural Cells in the Brain of a Nonhuman Primate

Asuka Morizane, Daisuke Doi, Tetsuhiro Kikuchi, Keisuke Okita, Akitsu Hotta, Toshiyuki Kawasaki, Takuya Hayashi, Hirotaka Onoe, Takashi Shiina, Shinya Yamanaka and Jun Takahashi

Stem Cell Reports, 26 September 2013, 10.1016/j.stemcr.2013.08.007

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Adult Cells Transformed into Early-stage Nerve Cells, Bypassing the Pluripotent Stem Cell Stage

Adult Cells Transformed into Early-stage Nerve Cells, Bypassing the Pluripotent Stem Cell Stage

Friday, 03 May 2013

A University of Wisconsin-Madison research group has converted skin cells from people and monkeys into a cell that can form a wide variety of nervous-system cells — without passing through the do-it-all stage called the induced pluripotent stem cell, or iPSC.

Standing at centre, Su-Chun Zhang, professor of

neuroscience in the School of Medicine and Public

Health., talks with his staff as they prepare stem-cell

cultures in the Zhang's research lab at the Waisman

Center at the University of Wisconsin-Madison on

March 8, 2013. Pictured at right are postdoctoral

students Yan Liu, background, and Lin Yao,

foreground. Credit: Photo by Jeff Miller/
UW-Madison.

Bypassing the ultra-flexible iPSC stage was a key advantage, says senior author Su-Chun Zhang, a professor of neuroscience and neurology.

"iPSC cells can generate any cell type, which could be a problem for cell-based therapy to repair damage due to disease or injury in the nervous system."

In particular, the absence of iPSC cells rules out the formation of tumours by pluripotent cells in the recipient, a major concern involving stem cell therapy.

A second advance comes from the virus that delivers genes to reprogram the adult skin cells into a different and more flexible form. Unlike other viruses used for this process, the Sendai virus does not become part of the cell's genes.

Jianfeng Lu, Zhang's postdoctoral research associate at the UW–Madison Waisman Center, removed skin cells from monkeys and people, and exposed them to Sendai virus for 24 hours. Lu then warmed the culture dish to kill the virus without harming the transforming cells. Thirteen days later, Lu was able to harvest a stem cell called an induced neural progenitor. After the progenitor was implanted into new-born mice, neural cells seemed to grow normally, without forming obvious defects or tumours, Zhang says.

Other researchers have bypassed the pluripotent stem cell stage while turning skin cells into neurons and other specialized cells, Zhang acknowledges, but the new research, just published in Cell Reports, had a different goal.

"Our idea was to turn skin cells to neural progenitors, cells that can produce cells relating to the neural tissue. These progenitors can be propagated in large numbers."

The research overcomes limitations of previous efforts, Zhang says. First, the Sendai virus, a kind of cold virus, is considered safe because it does not enter the cell's DNA, and it is killed by heat within 24 hours. (This is quite similar to the fever that raises our temperature to remove cold virus.) Second, the neural progenitors have a greater ability to grow daughter cells for research or therapy. Third, the progenitor cells are already well along the path toward specialization, and cannot become, say, liver or muscle cells after implantation. Finally, the progenitors can produce many more specialized cells.

The neurons that grew from the progenitor had the markings of neurons found in the rear of the brain, and that specialization can also be helpful.

"For therapeutic use, it is essential to use specific types of neural progenitors," says Zhang.

"We need region-specific and function-specific neuronal types for specific neurological diseases."

Progenitor cells grown from the skin of ALS (Lou Gehrig's disease) or spinal muscular atrophy patients can be transformed into various neural cells to model each disease and allow rapid drug screening, Zhang adds.

Eventually, the process could produce cells used to treat conditions like spinal cord injury and ALS.

"These transplantation experiments confirmed that the reprogrammed cells indeed belong to cells of the intended brain regions and the progenitors produced the three major classes of neural cells: neurons, astrocytes and oligodendrocytes," Zhang says.

"This proof-of-principle study highlights the possibility to generate many specialized neural progenitors for specific neurological disorders."

Source: University of Wisconsin-Madison 

Contact: Su-Chun Zhang

Reference:

Generation of Integration-free and Region-Specific Neural Progenitors from Primate Fibroblasts

Jianfeng Lu, Huisheng Liu, Cindy Tzu-Ling Huang, Hong Chen, Zhongwei Du, Yan Liu, Mohammad Amin Sherafat, and Su-Chun Zhang
Cell Reports, 02 May 2013, 10.1016/j.celrep.2013.04.004

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Transplanted Brain Cells in Monkeys Light Up Personalized Therapy

UW study is key step toward treating disease with stem cells

Thursday, 14 March 2013

For the first time, scientists have transplanted neural cells derived from a monkey's skin into its brain and watched the cells develop into several types of mature brain cells, according to the authors of a new study in Cell Reports. After six months, the cells looked entirely normal, and were only detectable because they initially were tagged with a fluorescent protein.

Because the cells were derived from adult cells in each monkey's skin, the experiment is a proof-of-principle for the concept of personalized medicine, where treatments are designed for each individual.

And since the skin cells were not "foreign" tissue, there were no signs of immune rejection — potentially a major problem with cell transplants.

Standing at centre, Su-Chun Zhang, professor of 

neuroscience in the School of Medicine and Public 

Health, talks with his staff as they prepare stem-cell 

cultures in the Zhang's research lab at the Waisman 

Center at the University of Wisconsin–Madison on 

March 8, 2013. Pictured at right are postdoctoral 

students Yan Liu, background, and Lin Yao, 

foreground. Credit: Photo by Jeff Miller.

"When you look at the brain, you cannot tell that it is a graft," says senior author Su-Chun Zhang, a professor of neuroscience at the University of Wisconsin-Madison.

"Structurally the host brain looks like a normal brain; the graft can only be seen under the fluorescent microscope."

“This is the first time I saw, in a nonhuman primate, that the transplanted cells were so well integrated, with such a minimal reaction. And after six months, to see no scar, that was the best part," Marina Emborg says, an associate professor of medical physics at UW-Madison and the lead co-author of the study. "

The cells were implanted in the monkeys "using a state-of-the-art surgical procedure" guided by an MRI image, says Emborg. The three rhesus monkeys used in the study at the Wisconsin National Primate Research Center had a lesion in a brain region that causes the movement disorder Parkinson's disease, which afflicts up to 1 million Americans. Parkinson's is caused by the death of a small number of neurons that make dopamine, a signalling chemical used in the brain.

The transplanted cells came from induced pluripotent stem cells (iPS cells), which can, like embryonic stem cells, develop into virtually any cell in the body. iPS cells, however, derive from adult cells rather than embryos.

In the lab, the iPS cells were converted into neural progenitor cells. These intermediate-stage cells can further specialize into the neurons that carry nerve signals, and the glial cells that perform many support and nutritional functions. This final stage of maturation occurred inside the monkey.

Zhang, who was the first in the world to derive neural cells from embryonic stem cells and then iPS cells, says one key to success was precise control over the development process.

"We differentiate the stem cells only into neural cells. It would not work to transplant a cell population contaminated by non-neural cells."

Another positive sign was the absence of any signs of cancer, says Zhang — a worrisome potential outcome of stem cell transplants.

This neuron, created in the Su-Chun Zhang lab at the 

University of Wisconsin–Madison, makes dopamine, 

a neurotransmitter involved in normal movement. The 

cell originated in an induced pluripotent stem cell, which 

derive from adult tissues. Similar neurons survived and 

integrated normally after transplant into monkey brains 

— as a proof of principle that personalized medicine may 

one day treat Parkinson's disease (Date: 2010). Credit: 

courtesy by Yan Liu and Su-Chun Zhang, Waisman 

Center, University of Wisconsin–Madison. 

"Their appearance is normal, and we also used antibodies that mark cells that are dividing rapidly, as cancer cells are, and we do not see that. And when you look at what the cells have become, they become neurons with long axons [conducting fibres], as we'd expect. They also produce oligodendrocytes that are helping build insulating myelin sheaths for neurons, as they should. That means they have matured correctly, and are not cancerous."

The experiment was designed as a proof of principle, says Zhang, who leads a group pioneering the use of iPS cells at the Waisman Center on the UW-Madison campus. The researchers did not transplant enough neurons to replace the dopamine-making cells in the brain, and the animal's behaviour did not improve.

Although promising, the transplant technique is a long way from the clinic, Zhang adds.

"Unfortunately, this technique cannot be used to help patients until a number of questions are answered: Can this transplant improve the symptoms? Is it safe? Six months is not long enough. And what are the side effects? You may improve some symptoms, but if that leads to something else, then you have not solved the problem."

Nonetheless, the new study represents a real step forward that may benefit human patients suffering from several diseases, says Emborg.

"By taking cells from the animal and returning them in a new form to the same animal, this is a first step toward personalized medicine."

The need for treatment is incessant, says Emborg, noting that each year, Parkinson's is diagnosed in 60,000 patients.

"I'm gratified that the Parkinson's Disease Foundation took a risk as the primary funder for this small study. Now we want to move ahead and see if this leads to a real treatment for this awful disease."

"It's really the first-ever transplant of iPS cells from a non-human primate back into the same animal, not just in the brain," says Zhang.

"I have not seen anybody transplanting reprogrammed iPS cells into the blood, the pancreas or anywhere else, into the same primate. This proof-of-principle study in primates presents hopes for personalized regenerative medicine."

Source: University of Wisconsin-Madison 

Contact: Su-Chun Zhang

Reference:

Induced Pluripotent Stem Cell-Derived Neural Cells Survive and Mature in the Nonhuman Primate Brain

Marina E. Emborg, Yan Liu, Jiajie Xi, Xiaoqing Zhang, Yingnan Yin, Jianfeng Lu, Valerie Joers, Christine Swanson, James E. Holden, Su-Chun Zhang

Cell Reports, 14 March 2013, 10.1016/j.celrep.2013.02.016

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Chimeric Macaques Produced for the First Time

OHSU research produces the world's first primate chimeric offspring 
Thursday, 05 January 2012

Newly published research by scientists at Oregon Health & Science University provides significant new information about how early embryonic stem cells develop and take part in formation of the primate species. The research, which took place at OHSU's Oregon National Primate Research Center, has also resulted in the first successful birth of chimeric monkeys — monkeys developed from stem cells taken from two separate embryos. The research will be published this week in the online edition of the journal Cell and will be published in a future printed copy of the journal.

Chimeric macaques. Credit:

Oregon Health & Science
University.

The research was conducted to gain a better understanding of the differences between natural stem cells residing in early embryos and their cultured counterparts called embryonic stem cells. This study also determined that stem cell functions and abilities are different between primates and rodents.

Here's more information about the early primate stem cells that were studied: The first cell type was totipotent cells — cells from the early embryo that have the ability to divide and produce all of the differentiated cells in the placenta and the body of organism. These were compared with pluripotent cells — cells derived from the later stage embryo that have only the ability to become the body but not placenta.

In mice, either totipotent or pluripotent cells from two different animals can be combined to transform into an embryo that later becomes a chimeric animal. However, the current research demonstrated that for reasons yet unknown, chimeric animals can only develop from totipotent cells in a higher animal model: the rhesus macaque. OHSU showed this to be the case by successfully producing the world's first primate chimeric offspring, three baby rhesus macaques named Roku, Hex and Chimero.

"This is an important development — not because anyone would develop human chimeras — but because it points out a key distinction between species and between different kind of stem cells that will impact our understanding of stem cells and their future potential in regenerative medicine," explained Shoukhrat Mitalipov, Ph.D., an associate scientist in the Division of Reproductive and Developmental Sciences at ONPRC.

"Stem cell therapies hold great promise for replacing damaged nerve cells in those who have been paralyzed due to a spinal cord injury or for example, in replacing dopamine-producing cells in Parkinson's patients who lose these brain cells resulting in disease. As we move stem cell therapies from the lab to clinics and from the mouse to humans, we need to understand what these cells do and what they can't do and also how cell function can differ in species."

Source: Oregon Health & Science University

Contact: Jim Newman
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Genomic Fault Zones Come and Go

The fragile regions in mammalian genomes thought to play a key role in evolution go through a 'birth and death' process
Wednesday, 01 December 2010

The fragile regions in mammalian genomes that are thought to play a key role in evolution go through a "birth and death" process, according to new bioinformatics research performed at the University of California, San Diego. The new work, published in the journal Genome Biology on November 30, could help researchers identify the current fragile regions in the human genome – information that may reveal how the human genome will evolve in the future.

"The genomic architecture of every species on Earth changes on the evolutionary time scale and humans are not an exception. What will be the next big change in the human genome remains unknown, but our approach could be useful in determining where in the human genome those changes may occur," said Pavel Pevzner, a UC San Diego computer science professor and an author on the new study. Pevzner studies genomes and genome evolution from a computational perspective in the Department of Computer Science and Engineering at the UC San Diego Jacobs School of Engineering.


According to research performed at UC

San Diego, the fragile regions in mammalian

genomes that are thought to play a key role

in evolution go through a "birth and death"

process. The graphic above is from the new

Genome Biology paper in which the findings

are outlined. Credit: Pavel Pevzner/

Max Alekseyev.

The fragile regions of genomes are prone to "genomic earthquakes" that can trigger chromosome rearrangements, disrupt genes, alter gene regulation and otherwise play an important role in genome evolution and the emergence of new species. For example, humans have 23 chromosomes while some other apes have 24 chromosomes, a consequence of a genome rearrangement that fused two chromosomes in our ape ancestor into human chromosome 2.
This work was performed by Pevzner and Max Alekseyev – a computer scientist who recently finished his Ph.D. in the Department of Computer Science and Engineering at the UC San Diego Jacobs School of Engineering. Alekseyev is now a computer science professor at the University of South Carolina.

Turnover Fragile Breakage Model
"The main conclusion of the new paper is that these fragile regions are moving," said Pevzner.

In 2003, Pevzner and UC San Diego mathematics professor Glen Tesler published results claiming that genomes have "fault zones" or genomic regions that are more prone to rearrangements than other regions. Their "Fragile Breakage Model" countered the then largely accepted "Random Breakage Model" – which implies that there are no rearrangement hotspots in mammalian genomes. While the Fragile Breakage Model has been supported by many studies in the last seven years, the precise locations of fragile regions in the human genome remain elusive.

The new work published in Genome Biology offers an update to the Fragile Breakage Model called the "Turnover Fragile Breakage Model." The findings demonstrate that the fragile regions undergo a birth and death process over evolutionary timescales and provide a clue to where the fragile regions in the human genome are located.

Do the Math: Find Fragile Regions
Finding the fragile regions within genomes is akin to looking at a mixed up deck of cards and trying to determine how many times it has been shuffled.


In this graphic, the coloured marks represent

positions of the putative fragile regions in the

human genome. The Turnover Fragile Breakage

Model suggests that these regions likely form

(still active) fragile regions in the human genome.

The graphic above is from the new Genome

Biology paper in which the Turnover Fragile

Breakage Model (TFBM) is presented. According

to the TFBM, the fragile regions in mammalian

genomes that are thought to play a key role in

evolution go through a "birth and death" process.

Credit: Max Alekseyev/Pavel Pevzner.

 Looking at a genome, you may identify breaks, but to say it is a fragile region, you have to know that breaks occurred more than once at the same genomic position.

"We are figuring out which regions underwent multiple genome earthquakes by analyzing the present-day genomes that survived these earthquakes that happened millions of years ago. The notion of rearrangements cannot be applied to a single genome at a single point in time. It's relevant when looking at more than one genome," said Pevzner, explaining the comparative genomics approach they took.

"It was noticed that while fragile regions may be shared across different genomes, most often such shared fragile regions are found in evolutionarily close genomes. This observation led us to a conclusion that fragility of any particular genomic position may appear only for a limited amount of time. The newly proposed Turnover Fragile Breakage Model postulates that fragile regions are subject to a 'birth and death' process and thus have limited lifespan," explained Alekseyev.

The Turnover Fragile Breakage Model suggests that genome rearrangements are more likely to occur at the sites where rearrangements have recently occurred – and that these rearrangement sites change over tens of millions of years. Thus, the best clue to the current locations of fragile regions in the human genome is offered by rearrangements that happened in our closest ancestors – chimpanzee and other primates.

Pevzner is eagerly awaiting sequenced primate genomes from the Genome 10K Project. Sequencing the genomes of 10,000 vertebrate species – including 100s of primates – is bound to provide new insights on human evolutionary history and possibly even the future rearrangements in the human genome.

"The most likely future rearrangements in human genome will happen at the sites that were recently disrupted in primates," said Pevzner.

Work tied to the new Turnover Fragile Breakage Model may also be useful for understanding genome rearrangements at the level of individuals, rather than entire species. In the future, the computer scientists hope to use similar tools to look at the chromosomal rearrangements that occur within the cells of individual cancer patients over and over again in order to develop new cancer diagnostics and drugs.

Source: University of California at San Diego
Contact: Daniel Kane

Reference:
Comparative genomics reveals birth and death of fragile regions in mammalian evolution
Max A Alekseyev, Pavel A Pevzner
Genome Biology 2010, 11:R117 (30 November 2010), doi:10.1186/gb-2010-11-11-r117
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Transposable Elements in Human Genetic Diseases and Human Development

Jumping elements, some of which cause genetic diseases, become incorporated in the genome at different stages of human development 
Friday, 30 April 2010

The density of transposable (jumping) elements between sex chromosomes in primates may have important consequences for the studies of human genetic diseases, say Penn State University researchers. Erika Kvikstad, a 2009 Penn State Ph.D. graduate in genetics, and Kateryna Makova, an associate professor of biology at Penn State, used a statistical regression method to study the genomes of the human, chimpanzee, macaque, and orang-utans. They concluded that there is a strong sex-chromosome bias in the distribution of transposable elements, and providing insights about whether these non-coding, but important, DNA elements integrate themselves specifically into the male germline or female germline, or integrate themselves into the genome during the early stages of embryogenesis. Their study will be published in the May 2010 issue of the scientific journal Genome Research.

According to Kvikstad, now a postdoctoral scholar at the Université Claude Bernard Lyon 1 in Lyon, France, the team chose to study primates because of the importance of human evolution, human disease, and the "unique availability of a very detailed description of the human genome – more so than any other mammalian genome." The strides made in sequencing the human and other primate genomes have made this research possible only in the last decade. Makova, one of the researchers who contributed to the analysis of the macaque and chimpanzee genomes, notes that the sequence of the orang-utan genome used in the Penn State study has not yet been published. Makova received special permission to use the orang-utan data set in her study.

The team looked specifically at the densities of transposable elements, which are snippets of DNA capable of moving about, replicating themselves, and inserting copies within the genome. The classes of transposable elements are further distinguished by being short or long interspersed nuclear elements – SINEs and LINEs. Kvikstad and Makova looked at one SINE family – Alu sequences, which are about 300 base pairs long, – and one LINE family – L1 sequences, which can be thousands of bases long.

"The transposable elements that we chose to study, Alus and L1s, are significant because they are abundant," says Kvikstad.

"They comprise about a third of the primate genome. They are actively moving around in the genome via a copy-and-paste retro-transposition mechanism, so they can create new variation; for example, human diseases and cancers."

Even more importantly, Kvikstad points out:

"These transposable elements are highly abundant on the sex chromosomes – X and Y – which mean they could be evolving uniquely because of the unusual nature of sex-chromosome transmission. The Y is paternally inherited, so it resides in the male germline only; the X spends two-thirds of its time in the female germline and one-third in the male germline. If there are germline-specific differences in the activity of transposable elements, for example, we should see clues to these differences in their sex-chromosome distributions."

The team's findings surprised Kvikstad and Makova.

"Even after we corrected for regional genomic effects, we still observed a very strong sex-chromosome bias in distributions of transposable elements," says Makova.

"This finding clearly indicates that there are biases according to which elements integrate into the genome. There also are differences between these two classes of elements. Our study suggests that Alus probably integrate mostly in the male germline, while L1s integrate in both male and female germ lines, or they might integrate in early embryogenesis."

This bias has implications for understanding and perhaps someday even preventing and treating genetic diseases.

"For us to really understand how the genetic diseases occur, we need to know when the elements integrate – at what point in human development this occurs," says Makova.

"We are studying evolution mostly, but our results are relevant to genetic diseases caused by insertions of transposable elements in the genome. For instance, Alu insertions are known to cause some types of neurofibromatosis, haemophilia, breast cancer, Apert syndrome, cholinesterase deficiency, and complement deficiency."

When transposable elements were first discovered in the 1940s, many in the scientific community labelled them as "junk" DNA.

"I don't think many people agree that they are 'junk' DNA any longer. Many of these elements have function. Alu elements frequently possess the regulatory elements. Both the Alu and L1 elements are often involved in recombination, the phenomenon under which the genome can undergo rearrangement and reshuffling," says Makova.

Kvikstad and Makova spent a year analyzing the primate data. Previously, together with Francesca Chiaromonte, associate professor of statistics at Penn State, they had worked together on a project looking at primate insertions and deletions of a much smaller size, under 30 base pairs.

"We are the first team to look into this much detail at the distribution of transposable elements on human sex chromosomes," Makova says.

Kvikstad points out other important implications of this study.

"In particular, we noted that gene density was not a significant predictor of either Alu or L1 element density, at any evolutionary time point," she says.

"By contrast, density of conserved non-coding DNA or 'most conserved elements' was a strong negative predictor of L1 density – so L1 elements are scarce in regions of the genome that might contain many of these potentially functional non-coding DNAs. This is an important distinction, since previous studies inferring the action of natural selection in shaping the densities of transposable elements relied on gene density as a proxy for natural selection. Our results suggest that the potentially functional DNA residing in these most-conserved elements may be an additional hallmark of natural selection."

Source: The Pennsylvania State University
Contact: Barbara K. Kennedy

Reference:

The (r)evolution of SINE versus LINE distributions in primate genomes: Sex chromosomes are important
Erika M. Kvikstad and Kateryna D. Makova
Genome Res. 2010, 20: 600-613
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Chimp and Human Y Chromosomes Evolving Faster Than Expected

Chimp and Human Y Chromosomes Evolving Faster Than Expected Wednesday, 13 January 2010 Contrary to a widely held scientific theory that the mammalian Y chromosome is slowly decaying or stagnating, new evidence suggests that in fact the Y is actually evolving quite rapidly through continuous, wholesale renovation. By conducting the first comprehensive interspecies comparison of Y chromosomes, Whitehead Institute researchers have found considerable differences in the genetic sequences of the human and chimpanzee Y’s — an indication that these chromosomes have evolved more quickly than the rest of their respective genomes over the 6 million years since they emerged from a common ancestor. The findings are published online this week in the journal Nature. "The region of the Y that is evolving the fastest is the part that plays a role in sperm production," say Jennifer Hughes, first author on the Nature paper and a postdoctoral researcher in Whitehead Institute Director David Page's lab. "The rest of the Y is evolving more like the rest of the genome, only a little bit faster." The chimp Y chromosome is only the second Y chromosome to be comprehensively sequenced. The original chimp genome sequencing completed in 2005 largely excluded the Y chromosome because its hundreds of repetitive sections typically confound standard sequencing techniques. Working closely with the Genome Center at Washington University, the Page lab managed to painstakingly sequence the chimp Y chromosome, allowing for comparison with the human Y, which the Page lab and the Genome Center at Washington University had sequenced successfully back in 2003. The results overturned the expectation that the chimp and human Y chromosomes would be highly similar. Instead, they differ remarkably in their structure and gene content. The chimp Y, for example, has lost one third to one half of the human Y chromosome genes – a significant change in a relatively short period of time. Page points out that this is not all about gene decay or loss. He likens the Y chromosome changes to a home undergoing continual renovation. "People are living in the house, but there's always some room that's being demolished and reconstructed," says Page, who is also a Howard Hughes Medical Institute investigator. "And this is not the norm for the genome as a whole." Wes Warren, Assistant Director of the Washington University Genome Center, agrees. "This work clearly shows that the Y is pretty ingenious at using different tools than the rest of the genome to maintain diversity of genes," he says. "These findings demonstrate that our knowledge of the Y chromosome is still advancing." Hughes and Page theorize that the divergent evolution of the chimp and human Y chromosomes may be due to several factors, including traits specific to Y chromosomes and differences in mating behaviours. Because multiple male chimpanzees may mate with a single female in rapid succession, the males' sperm wind up in heated reproductive competition. If a given male produces more sperm, that male would theoretically be more likely to impregnate the female, thereby passing on his superior sperm production genes, some of which may be residing on the Y chromosome, to the next generation. Because selective pressure to pass on advantageous sperm production genes is so high, those genes may also drag along detrimental genetic traits to the next generation. Such transmission is allowed to occur because, unlike other chromosomes, the Y has no partner with which to swap genes during cell division. Swapping genes between chromosomal partners can eventually associate positive gene versions with each other and eliminate detrimental gene versions. Without this ability, the Y chromosome is treated by evolution as one large entity. Either the entire chromosome is advantageous, or it is not. In chimps, this potent combination of intense selective pressure on sperm production genes and the inability to swap genes may have fuelled the Y chromosome's rapid evolution. Disadvantages from a less-than-ideal gene version or even the deletion of a section of the chromosome may have been outweighed by the advantage of improved sperm production, resulting in a Y chromosome with far fewer genes than its human counterpart. To determine whether this rapid rate of evolution affects Y chromosomes beyond those of chimps and humans, the Page lab and the Washington University Genome Center are now sequencing and examining the Y chromosomes of several other mammals. Reference: Chimpanzee and human Y chromosomes are remarkably divergent in structure and gene content Jennifer F. Hughes, Helen Skaletsky, Tatyana Pyntikova, Tina A. Graves, Saskia K. M. van Daalen, Patrick J. Minx, Robert S. Fulton, Sean D. McGrath, Devin P. Locke, Cynthia Friedman, Barbara J. Trask, Elaine R. Mardis, Wesley C. Warren, Sjoerd Repping, Steve Rozen, Richard K. Wilson, David C. Page Nature, online January 13, 2010, doi:10.1038/nature08700 The fickle Y chromosome: Chimp genome reveals rapid rate of change. Lizzie Buchen Nature 463, 149 (2010), doi:10.1038/463149a ......... ZenMaster

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Human Embryonic Stem Cells Reverse Retinal Degeneration

Human Embryonic Stem Cells Reverse Retinal Degeneration Monday, 12 October 2009 A new study reports that transplanted pigment-containing visual cells derived from human embryonic stem cells (hESCs) successfully preserved structure and function of the specialized light-sensitive lining of the eye (known as the retina) in an animal model of retinal degeneration. The findings, published by Cell Press in the October 2nd issue of the journal Cell Stem Cell, represent an exciting step towards the future use of cell replacement therapies to treat devastating degenerative eye diseases that cause millions of people worldwide to lose their sight. The retinal pigment epithelium (RPE) is a layer of pigmented cells sandwiched between the visual retinal cells, called photoreceptors, and the nourishing blood vessels at the back of the eye. The RPE provides essential support to the retinal photoreceptors and is critical for normal vision. Deterioration of the RPE plays a central role in the progression of diseases such as age-related macular degeneration and sub-types of retinitis pigmentosa. These conditions are associated with a progressive loss of vision that often leads to blindness. "Although there are a variety of therapeutic approaches under development to delay the degenerative process, the grim reality is that many patients eventually lose their sight," explains Dr. Benjamin Reubinoff, a senior author of the study. "Cell therapy to replenish the degenerating RPE cells may potentially halt disease progression." Dr. Reubinoff and Dr. Eyal Banin who led the study, with their colleagues from Hadassah-Hebrew University Medical Center in Jerusalem, developed conditions to guide hESCs to differentiate into functional RPE-like cells in the laboratory. The researchers found that nicotinamide (vitamin B3, NIC) and Activin A, an important growth factor, promoted differentiation of hESCs towards an RPE fate. The hESC-derived RPE-like cells, which could be identified by their characteristic black pigment, exhibited multiple biological properties and genetic markers that define authentic RPE cells. Further, the cells successfully delayed deterioration of retinal structure and function when they were transplanted into an animal model of retinal degeneration caused by RPE dysfunction. Taken together, the results demonstrate that NIC and Activin A promoted the differentiation of hESCs towards an RPE fate. The hESC-derived cells exhibited the defining characteristics associated with RPE and successfully rescued the retina when transplanted into an animal model of retinal degeneration. "Our findings are an important step towards the potential future use of hESCs to replenish RPE in blinding diseases," concludes Dr. Banin. ......... ZenMaster

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Cure for Colour Blindness in Monkeys

Gene therapy used to treat adult vision disorders involving cone cells Wednesday, 16 September 2009 Researchers from the University of Washington and the University of Florida used gene therapy to cure two squirrel monkeys of colour blindness — the most common genetic disorder in people. Writing online Wednesday in the journal Nature, scientists cast a rosy light on the potential for gene therapy to treat adult vision disorders involving cone cells — the most important cells for vision in people. "We've added red sensitivity to cone cells in animals that are born with a condition that is exactly like human colour blindness," said William W. Hauswirth, Ph.D., a professor of ophthalmic molecular genetics at the UF College of Medicine and a member of the UF Genetics Institute and the Powell Gene Therapy Center. "Although colour blindness is only moderately life-altering, we've shown we can cure a cone disease in a primate, and that it can be done very safely. That's extremely encouraging for the development of therapies for human cone diseases that really are blinding." The finding is also likely to intrigue millions of people around the world who are colour-blind, including about 3.5 million people in the United States, more than 13 million in India and more than 16 million in China. The problem mostly affects men, leaving about 8 percent of Caucasian men in the United States incapable of discerning red and green hues that are important for everyday things like recognizing traffic lights. "People who are colour-blind feel that they are missing out," said Jay Neitz, Ph.D., a professor of ophthalmology at the University of Washington. "If we could find a way to do this with complete safety in human eyes, as we did with monkeys, I think there would be a lot of people who would want it. Beyond that, we hope this technology will be useful in correcting lots of different vision disorders." The discovery comes about 10 years after Neitz and his wife Maureen Neitz, Ph.D., a professor of ophthalmology at the University of Washington and senior author of the study, began training two squirrel monkeys named Dalton and Sam. In addition to teaching the animals, the Neitz research group worked with the makers of a standard vision-testing technique called the Cambridge Colour Test to perfect a way the monkeys could "tell" them which colours they were seeing. The tests are similar to ones given to elementary children the world over, in which students are asked to identify a specific pattern of coloured dots among a field of dots that vary in size, colour and intensity. The researchers devised a computer touch screen the monkeys could use to trace the colour patterns. When the animals chose correctly, they received a reward of grape juice. Likewise, decades were spent by Hauswirth and colleagues at the University of Florida, to develop the gene-transfer technique that uses a harmless adeno-associated virus to deliver corrective genes to produce a desired protein. In this case, researchers wanted to produce a substance called long-wavelength opsin in the retinas of the monkeys. This particular form of opsin is a colourless protein that works in the retina to make pigments that are sensitive to red and green. "We used human DNA’s, so we won't have to switch to human genes as we move toward clinical treatments," said Hauswirth, who is also involved in a clinical trial with human patients to test gene therapy for the treatment of Leber congenital amaurosis, a form of blindness that strikes children. About five weeks after the treatment, the monkeys began to acquire colour vision, almost as if it occurred overnight. "Nothing happened for the first 20 weeks," Neitz said. "But we knew right away when it began to work. It was if they woke up and saw these new colours. The treated animals unquestionably responded to colours that had been invisible to them." It took more than a year and a half to test the monkeys' ability to discern 16 hues, with some of the hues varying as much as 11-fold in intensity. Dalton is named for John Dalton, an English chemist who realized he was colour-blind and published the first paper about the condition in 1798. "We've had Dalton and Sam for 10 years. They are like our children," Neitz said. "This species are friendly, docile monkeys that we just love. We think it is useful to continue to follow them — it's been two years now that they've been seeing in colour, and continuing to check their vision and allowing them to play with the computer is part of their enrichment." With the discovery, the researchers are the first to address a vision disorder in primates in which all photoreceptors is intact and healthy, providing a hint of gene therapy's full potential to restore vision. About 1 in 30,000 Americans have a hereditary form of blindness called achromatopsia, which causes nearly complete colour blindness and extremely poor central vision. "Those patients would be targets for almost exactly the same treatment," Hauswirth said. Even in common types of blindness such as age-related macular degeneration and diabetic retinopathy, vision could potentially be rescued by targeting cone cells, he said. "The major thrust of the study is you can ameliorate if not cure colour blindness with gene therapy," said Gerald H. Jacobs, Ph.D., a research professor of psychology at the University of California, Santa Barbara, who was not involved in the research. "There are still questions about safety, but in these monkeys at least, there were no untoward effects. Those who are motivated to ameliorate their colour defect might take some hope from the findings.” "This is also another example of how utterly plastic the visual system is to change," Jacobs said. "The nervous system can extract information from alterations to photo-pigments and make use of it almost instantaneously." Reference: Gene therapy for red–green colour blindness in adult primates Katherine Mancuso, William W. Hauswirth, Qiuhong Li, Thomas B. Connor, James A. Kuchenbecker, Matthew C. Mauck, Jay Neitz & Maureen Neitz Nature advance online publication 16 September 2009, doi:10.1038/nature08401 ......... ZenMaster

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Monkey Skin Cells Reprogrammed into Stem Cells

What’s good for the mouse is good for the monkey Friday, 05 December 2008 Scientists have successfully created the first induced pluripotent stem (iPS) cell lines from adult monkey skin cells. The research, published by Cell Press in the December issue of the journal Cell Stem Cell, demonstrates that the method of direct reprogramming is conserved among species and may be useful for creation of clinically valuable primate models for human diseases. Although previous work has shown that induction of four key transcription factors can reprogram adult mouse and human skin cells into iPS cells, creation of iPS cells in other species has not been demonstrated. "We sought to generate monkey iPS cells from skin cells isolated an adult male rhesus macaque using the predicted monkey transcription factors OCT4, SOX2, KLF4 and c-MYC," explains Dr. Hongkui Deng from the Key Laboratory of Cell Proliferation and Differentiation at Peking University in Beijing, China. Dr. Deng and colleagues used retroviruses expressing these four factors to infect adult monkey skin cells. This technique led to creation of cells that displayed multiple hallmarks of embryonic stem (ES) cells. Specifically, the cells exhibited physical characteristics associated with ES cells, expressed genes appropriate for ES cells and possessed the ability to develop into multiple types of differentiated cells. These results reveal that monkey iPS cells can be generated using the same four transcription factors that have been used to successfully create mouse and human iPS cells. The work has multiple exciting applications. "As the rhesus macaque is the most relevant primate model for most human diseases, highly efficient generation of monkey iPS cells would allow investigation of the treatment of various diseases in this model," offers Dr. Deng. "In addition, direct reprogramming with the four transcription factors could be a universal strategy for generating iPS cells in other species." Reference: Generation of Induced Pluripotent Stem Cells from Adult Rhesus Monkey Fibroblasts Haisong Liu, Fangfang Zhu, Jun Yong, Pengbo Zhang, Pingping Hou, Honggang Li, Wei Jiang, Jun Cai, Meng Liu, Kai Cui, Xiuxia Qu, Tingting Xiang, Danyu Lu, Xiaochun Chi, Ge Gao, Weizhi Ji, Mingxiao Ding, Hongkui Deng Cell Stem Cell, Volume 3, Issue 6, 587-590, 4 December 2008, doi:10.1016/j.stem.2008.10.014 See also: Guidelines and Techniques for the Generation of Induced Pluripotent Stem Cells Nimet Maherali and Konrad Hochedlinger Cell Stem Cell, Volume 3, Issue 6, 595-605, 4 December 2008, doi:10.1016/j.stem.2008.11.008 ......... ZenMaster

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