Abnormal Blood Stem Cells are the Cause of Leukemia

Cancer is, to a large degree, a disease of stem cells. When stem cells acquire particular mutations, they lose their controls on cell division and begin to divide uncontrollably. Several different studies have established that several types of cancers result from abnormal stem cells. Blood cancers, for example, form when stem cells accrue rare genetic mutations, according a new study. This discovery overturns the traditional view that blood cancers can originate from any blood cell, and it could conceivably help to prevent relapses in leukemia patients.

Stanford University researchers have identified the origins of leukemia. They used so-called “next-generation sequencing” techniques and various other methods to identify rare, pre-cancerous, blood stem cells in six individuals with acute myeloid leukemia. After identifying these pre-cancerous cells, they compared the genetic sequences from the pre-cancerous blood stem cells to the sequences of the same chromosomal regions from the patients’ leukemia-plagued stem cells. This analysis revealed the exact order of rare mutations that blood stem cells accrued in order to become cancerous.

Stanford hematologist Ravi Majeti, co-lead author of the study, commented: “I’m surprised that we identified the clonal hierarchy that led to leukemia in five of the six cases. I didn’t think we’d find that amount of evidence of pre-leukemia stem cells.”

Scientists have suspected for the last few decades that cancer stem cells, and in particular leukemia stem cells, lead to cancer. In 2005, a Stanford pathologist named Irving Weissman added a twist to this idea when he proposed that normal blood stem cells become cancerous stem cells by accumulating rare mutations. Weissman’s hypothesis suggested that leukemia originated in blood stem cells. Weissman’s hypothesis makes sense of a simple fact; blood stem cells live much longer than regular blood cells, which only live up to a few weeks at most. A few weeks is simply not enough time, to acquire the number of rare mutations necessary to become cancerous. Since blood stem cells are capable of self-renewal, they survive in the body throughout the lifetime of an organism. Unfortunately, such a hypothesis, despite its great explanatory power, is very difficult to directly test, and, therefore, has remained controversial.

The best way to test Weissman’s hypothesis is to identify the protein-coding mutations in several acute myeloid leukemias, and then isolate and analyze the rare, pre-cancerous stem cells to determine which, of the leukemia mutations were present in those pre-cancerous stem cells.

In addition to their sequencing approach, this team also used high-throughput flow cytometry to identify markers specific to a patient’s healthy blood cell-making stem cells versus their leukemia stem cells in order to isolate the very rare populations of pre-cancerous stem cells.

These techniques were pioneered by Thomas Snyder, who is a chief scientist at ImmuMetrix and co-lead author of this paper. Snyder worked as a post-doctoral researcher in the laboratory of Stanford bioengineer Stephen Quake when this he collaborated on this study. Together, Quake and Snyder developed those techniques to sort and study the genomes of each individual cell. “It is only when you can look at a single cell and determine its genotype that you can conclusively show the early stages in the evolution of the cancer,” said Snyder.

StemCells, Inc. Human Neural Stem Cells Restore Memory in Models of Alzheimer’s Disease

StemCells, Inc., a Newark, California-based company has announced preclinical data that demonstrates that its proprietary human neural stem cell line restored memory and enhanced synaptic function in two animal models that are relevant to Alzheimer’s disease (AD). They presented these data at the Alzheimer’s Association International Conference 2012 in Vancouver, Canada.

In this study, neuroscientists from University of California, Irvine transplanted a neural stem cell line called HuCNS-SC, a proprietary stem cell line made by StemCells and is a purified human neural stem cell line, into a specific region of the brain, the hippocampus in laboratory animals. These injections improved the memories of two different types of laboratory animal that act as AD-significant models. The hippocampus is a portion of the brain that is critically important to the control of memory, and unfortunately, it is severely affected by AD. Specifically, hippocampal synaptic density is reduced in AD and these reductions in synaptic connections are highly correlated with memory loss. After injections of HuCNS-SCs, the animals showed increased synaptic density and improved memory after the cells had been transplanted. Importantly, these results did not require reduction in beta amyloid or tau that accumulate in the brains of patients with AD and account for the pathological hallmarks of the disease.

This research study resulted from collaboration between Frank LaFerla, Ph.D., who is the Director of the University of California, Irvine (UCI) Institute for Memory Impairments and Neurological Disorders (UCI MIND), and Chancellor’s Professor, Neurobiology and Behavior in the School of Biological Sciences at UCI, and Matthew Blurton-Jones, Ph.D., Assistant Professor, Neurobiology and Behavior at UCI.

“This is the first time human neural stem cells have been shown to have a significant effect on memory,” said Dr. LaFerla. “While AD is a diffuse disorder, the data suggest that transplanting these cells into the hippocampus might well benefit patients with Alzheimer’s. We believe the outcomes in these two animal models provide strong rationale to study this approach in the clinic and we wish to thank the California Institute of Regenerative Medicine for the support it has given this promising research.”

Stephen Huhn, M.D., FACS, FAAP, Vice President and Head of the CNS Program at StemCells Inc, added, “While reducing beta amyloid and tau burden is a major focus in AD research, our data is intriguing because we obtained improved memory without a reduction in either of these pathologies. AD is a complex and challenging disorder. The field would benefit from the pursuit of a diverse range of treatment approaches and our neural stem cells now appear to offer a unique and viable contribution in the battle against this devastating disease.”

A New Muscle Disease Target: Myostatin

A protein found in the muscular system known as myostatin has piqued the interest of muscle researchers who focus on muscular disorders.

Myostatin is a heavily-studied protein and has a function that is common to many different types of animals (cows, sheep, humans, & mice). Myostatin inhibits the growth of muscles so that they do not overgrow and capitalize on the nutrients of the developing embryo or body. Mice that carry mutations in the gene that encodes myostatin have twice the muscle mass of normal mice.  Beef cattle with mutations in the myostatin gene are heavily muscled and highly sought after for restaurants.  Piedmontese and Belgium Blue beef cows example of cattle with mutations in the myostatin gene and are very heavily muscled.  This makes it a potential drug target for people who suffer from degenerative diseases that affect muscles.

Myostatin is also known as growth differentiation factor 8 (GDF-8) and it is secreted by muscle cells and circulates throughout the bloodstream.  It controls the growth of muscles by binding to a receptor on the surface of muscle cells called the “activin type II receptor.”

There is, however, a sizable debate as to which muscles are controlled by myostatin. Fortunately, as new study by Carnagie’s Chen-Ming Fan and Christoph Lepper narrows down the target muscles of myostatin.

Skeletal muscles possess a stem cell population called satellite cells that are activated by muscular injury. After muscle injury, for example, by lifting heavy weights, cause the satellite cells to divide and fuse to existing muscle fibers. Does myostain affect satellite cells or the muscle fibers?

According to work by Fan and Se-Jin Lee, who is a former Carnagie Staff Associate and is currently at the Johns Hopkins University Medical School, muscle growth caused by the inhibition of myostatin is not due to the incorporation of satellite cells into muscle fibers. Their work used genetic and pharmacological means to show this. For example, they knocked our the gene that encodes the receptor for myostatin (Acvr2b) they found that the mice had increased muscle mass that was not due to increased satellite cell activity.  Therefore, their conclusions are rather solid.

Now these results might have implications for the possible use of myostatin as a clinical target. The reason for this is that diseases that cause muscle degeneration tend to produce symptoms later in life because a patient’s satellite cells regenerate the muscle until their populations are eventually depleted. Inhibiting myostatin through pharmacological means could benefit patients over time, since they would tend to spare the satellite cell population by growing muscle without satellite cells.

According to Fan, “More work is needed to determine whether these findings are applicable to various conditions , such as exercise, injury and sarcopenia – degenerative loss of muscle mass associated with aging. However, our findings initially indicate that many different diseases affecting the muscular system could potentially be responsive to drugs that inhibit myostatin and thus promote muscle growth, without regard to the status of the muscle stem cell pool.”

Embryonic Stem Cell Lines Derived from Embryos Frozen for 18 Years

How long do human embryos survive in cryostorage? To be completely honest, no one truly knows. According to the Planer PLC Group, a cryopreservation company, a baby was born from an embryo that had frozen for 16 years at their institution. However, it is possible that embryos might live even longer in cryostorage. Furthermore, one study that examined more than 11,000 cryopreserved human embryos determined that the length of time for which the embryo was frozen had no significant effect on post-thaw survival for in vitro fertilisation (IVF) or oocyte donation cycles, or for embryos frozen at the pronuclear or cleavage stages. This study also showed that the duration of storage did not have any significant effect on clinical pregnancy, miscarriage, implantation, or live birth rate, whether from IVF or oocyte donation cycles (Riggs R, Mayer J, Dowling-Lacey D, Chi TF, Jones E, Oehninger S (November 2008). “Does storage time influence postthaw survival and pregnancy outcome? An analysis of 11,768 cryopreserved human embryos”. Fertil. Steril. 93 (1): 109–15).

However, some embryos do not survive the freezing process. Also, some embryos that are frozen are very low-quality embryos that have an exceedingly low probability of ever making a baby. Since these embryos are of very little value from a reproductive standpoint, they might be of use to stem cell biologists who want to make embryonic stem cells from them. Several studies have shown that low quality embryos are excellent sources of material for embryonic stem cells. For example, Lerou PH, et al., Human embryonic stem cell derivation from poor-quality embryos.Nat Biotechnol. 2008 Feb;26(2):212-4. In this paper Daley’s lab derived embryonic stem cell lines at rates comparable to the rates of embryonic stem cell derivation with high-quality embryos. Another paper by Shetty R and Inamdar MS, “Derivation of human embryonic stem cell lines from poor quality embryos,” in Methods Mol Biol. 2012;873:151-61, Indian researchers derived embryonic stem cell lines from low-quality embryos.

A Chinese laboratory has also used low quality embryos that were discarded by fertility clinics. 166 poor quality embryos gave rise to 4 new embryonic stem cell lines in this paper (see Lui W, et al., J Genet Genomics. 2009 Apr;36(4):229-39). Therefore, this practice is well established.

What is questionable is whether or not the embryo is actually dead. Remember, even though grade III embryos are not desirable because they show lower rates of implantation, they still give rise to live births occasionally. For example, one study showed that grade iV embryos (worse than grade III) gave pregnancy rates of 18.2%. Therefore, these studies are being done with low-grade embryos – not embryos that are clinically dead.

As I have noted before in a previous post, defining death in an embryo is difficult to do, but when there are far more dead cells in the embryo than live ones, the chance of the embryo giving rise to a baby becomes so low as to be impossible. If 60% of the cells in the embryos are dead, then the embryo is usually defined as clinically dead. Such an embryo, if it has not experienced early developmental arrest, can be a reasonable source of embryonic stem cells, according to work from the Daley lab

With this in mid, there is a paper from a research group at Chulalongkorn University and Chilalongkorn Memorial Hospital, Bangkok, Thailand, that shows that embryonic stem cells can be successfully made from embryos that had been frozen for 18 year. This paper shows that even embryos that have been frozen for almost two decades can still yield embryonic stem cells.

Evaluations of these embryonic stem cell lines revealed that they were as pluripotent as similar lines derived from embryos that had only been frozen for a few years.

Jane Taylor, a collaborator in this paper from the MRC Centre for Regenerative Medicine at the University of Edinburgh, Scotland, said, “The importance of this study is that is it identifies an alternative source for generating new embryonic stem lines, using embryos that have been in long-term storage.”

These frozen embryos, if they were not clinically dead, were still human beings. They merely needed to be implanted into a mother’s womb in order to execute their intrinsic developmental program that implants itself into the mother’s womb. By using these embryos to derive embryonic stem cell lines, their lives were ended. All other arguments that try to downgrade the essential status of these embryos because they are too young, too small, in the wrong place or too different from an adult rely upon accidental qualities of the embryo. That is, qualities that are temporary and not integral to the essence of the embryo. Its essence is that of a human being. When it grows larger, it is still a human being and the fact that it executes its intrinsic developmental program to do so merely demonstrates its human essence. The same can be said about it appearance, and age.

Location is an even more problematic criterion by which to disqualify the embryo from the human race.

Therefore, these embryos were destroyed and their human lives, killed. Surely there is a better way to do regenerative science that both respects the value of human life and creates technologies to heal us. Interested? Read the other posts on this blog.

Todd Akins and Pregnancy as a Result of Rape

Republican candidate for U.S. Senator from the state of Missouri, Todd Akin, really stuck his foot in his mouth during an interview on the Jaco Report on Fox. After he stated that abortion should be legal to save the life of the mother, the host asked if it should also be legal in the case of rape.

Akin responded: “People always want to try and make that as one of those things, well, how do you slice this particularly tough sort of ethical question. It seems to me, first of all, that from what I understand from doctors, that’s really rare. If it is a legitimate rape, the female body has ways to try and shut that whole thing down. But let’s assume maybe that didn’t work or something. I think there should be some punishment, but the punishment should be of the rapist and not attacking the child.”

Akin issued an apology but the damage is already done. His statement was poorly worded and garbled. He probably meant to refer to a forcible rape, which is also known as an assault rape as opposed to a date rape. He was probably trying to make this distinction since there have been cases whereby women who become pregnant from consensual intercourse have later claimed rape. His wording failed to properly clarify what he meant.

Even worse was his statement that ” the female body has ways to try and shut that whole thing down.” Again I think he was trying to refer to the physical trauma experienced by a woman when she is raped. Stress and emotional factors can alter a woman’s menstrual cycle. In order to get pregnant, and stay pregnant the body of the woman must produce a complex mix of hormones. This hormone production is under the control of the brain and the part of the brain that controls reproductive hormones (the limbic system) is easily influenced by emotions. An assault rape certainly qualifies as great emotional trauma. Such trauma can radically upset ovulation, fertilization, implantation and even the nurturing of a pregnancy.

Having said all that, women do get pregnant from assault rapes. Approximately 1 in 15 women who are raped will get pregnant from it (see Statistics on Sexual Violence Against Women: A Criminological Study, 1990). Another article by Holmes, Resnick, Kilpatrick, and Best (Rape-related pregnancy: estimates and descriptive characteristics from a national sample of women) from the American Journal of Obstetrics and Gynecology (1996 Aug;175(2):320-4; discussion 324-5), finds that the national rate of rape pregnancies is 5.0% per rape among victims of reproductive age (aged 12 to 45). This rate is higher because some women who are raped are too old or too young to become pregnant from the rape. Nationally, there were an estimated 32,101 rape pregnancies each year. Only 11.7% of rape victims received immediate medical attention after the assault, and 47.1% received no medical attention related to the rape. 32.2% kept the infant, 50% underwent abortion and 5.9% placed the infant for adoption. 11.8% had a spontaneous abortion.

Thus the statistics show that pregnancy as a result of an assault or forcible rape does occur frequently enough so that pro-life politicians, thinkers and workers must take it seriously. The simple fact is that the baby should not pay the price of his or her life for the crimes of the father. That is the crux of the pro-life position. Abortion as a that ends the life of a baby who is the product of a rape still ends the life of a baby who had nothing to do with the crime still kills a baby. Had Akin put it this way, then he would not have stuck his foot in his mouth the way he did.

There are complications with forcing the woman to be a life-support system for a baby she did not wish to conceive, but the fact still remains that a baby’s life hangs in the balance. In the scheme of things, it seems to me that having the woman bear the brunt of the pregnancy is the lesser of two evils and saving the life of the baby is a greater good.  Trying to be cute about it will only get you in trouble and mark you as ignorant and insensitive to women.

Using Junk Biology to Promote “Uselessness” of Men

Wesley Smith over at Secondhand Smoke notes the New York Times op-ed by an actual biology professor, Greg Hampikian, who argues that men are unimportant to the propagation of the human species.

The column says: “Think about your own history. Your life as an egg actually started in your mother’s developing ovary, before she was born; you were wrapped in your mother’s fetal body as it developed within your grandmother.”

He continues: “After the two of you left Grandma’s womb, you enjoyed the protection of your mother’s prepubescent ovary.”

Smith notes the rather obvious truth that neither you nor I were never an egg. Neither were we ever a sperm. Our lives began at the completion of fertilization and at that point, a new person – you – were created. This kind of biological claptrap is being written by someone who should know better, and is used to argue that men are just playthings for women. This is beyond ridiculous; it is asinine.

If that wasn’t bad enough, here comes more: ” Then, sometime between 12 and 50 years after the two of you left your grandmother, you burst forth and were sucked by her fimbriae into the fallopian tube. You glided along the oviduct, surviving happily on the stored nutrients and genetic messages that Mom packed for you. Then, at some point, your father spent a few minutes close by, but then left. A little while later, you encountered some very odd tiny cells that he had shed. They did not merge with you, or give you any cell membranes or nutrients — just an infinitesimally small packet of DNA, less than one-millionth of your mass.”

The sperm gave a packet of DNA, a spindle and the biochemical signals to metabolically activate the embryo and get it dividing. These are all vital contributions to the beginning of human life. Hampikian’s abuse of biology is inexcusable.

See Smith’s post here.

Highly Efficient Method for Converting Blood Stem Cells into Induced Pluripotent Stem Cells Without Viruses

A research group from Johns Hopkins University has designed a protocol that reliably converts stem cells from umbilical cord blood into a primitive stem cell state. From this primitive state, these cells can differentiate into any other type of cell in the body.

This paper was published in the August 8th issue of Public Library of Science (PLoS), and serves as the second publication in an ongoing effort to efficiently and consistently convert umbilical cord blood stem cells and other types of stem cells into stem cells that are usable for use in clinical and research settings in place of human embryonic stem cells, according to Elias Zambidis, M.D., Ph.D., who is an assistant professor of oncology and pediatrics at the Johns Hopkins Institute for Cell Engineering and the Kimmel Cancer Center.

Zambidis said: “Taking a cell from an adult and converting it all the way back to the way it was when that person was a 6-day-old embryo creates a completely new biology toward our understanding of how cells age and what happens when things go wrong, as in cancer development.”

The first paper that is sometimes designated ‘Chapter One‘ of this work was published last spring in PLoS One. In this paper, Zambidis’ group described the successful use of a method that safely transformed several different types of human pluripotent stem cells into heart muscle cells. In the latest experiments, Zambidis and his colleagues describe methods that convert umbilical cord blood stem cells into induced-pluripotent stem cells (iPS), which are adult or fetal cells reprogrammed to an embryonic like state.

According to Zambidis, he and his team developed a “super-efficient, virus-free” method for making iPS cells. This overcomes some troubling difficulties for those scientists who work with iPS cells; namely, the vast inefficiency of making iPS cells from adult cells and the use of mutation-causing viruses to introduce those genes into adult cells required to convert adult cells into iPS cells. Generally, out of hundreds of blood cells, only one or two typically revert into iPS cells. However, with Zambidis’ method, 50-60% of blood cells were engineered into iPS cells.

To circumvent the use of viruses to deliver genes, Zambidis’ team used plasmids, or small circles of DNA that replicate briefly inside cells and then degrade. By using plasmids, the cells receive the genes required to drive adult cells into the iPS state, but because these genes are only required transiently, the plasmids do their job and then go away. Therefore, the production of mutations by viral DNAs that insert themselves into the host cell genome is not a problem with this method from Zambidis’ laboratory.

In order to introduce the genes into the cells, Zambidis’ team used a technique called electroporation. They treated the umbilical cord blood cells with the plasmids and then delivered an electrical pulse to the cells, which made tiny holes in the surface through which the plasmids could slip to the cell interior. Once inside, the plasmids triggered the cells to revert to a more primitive cell state. After genetic engineering, the blood cells were also given an additional new step in which they were stimulated with their natural bone-marrow environment. To do this, the Johns Hopkins team took some of the treated cells in a dish alone, and cultured them together with irradiated bone-marrow cells.

When iPS cells made from umbilical cord blood were compared to iPS cells made from hair cells and from skin cells, they found that the most superior iPS cells came from those made from blood stem cells treated with just four genes and cultured with the bone marrow cells. These cells reverted to a primitive stem cell state within seven to 14 days. Their techniques also successfully converted blood stem cells from adult bone marrow and from circulating blood into iPS cells.

In ongoing studies, Zambidis and colleagues are testing the quality of their newly formed iPS cells. They are also interested in the ability of these iPS cells to differentiate into other cell types, as compared with iPS cells made by other methods. These efficient methods to produce virus-free iPS cells will hopefully speed research to develop stem cell therapies that use nearly all cell types, and may provide a more accurate picture of cell development and biology.

First Human Study Using Dental Stem Cells

This is an old paper, but it is still a good read.

November 12th 2009 the first clinical study involving human dental stem cells was published in the journal European Cells and Materials journal.

This study examined patients with impacted wisdom teeth who also had bone loss (resorption) at the site of impaction. Such a bone defect does not repair on its own after the wisdom teeth are removed. Therefore, the researchers used a mixture of dental pulp stem cells harvested from the patient’s non-impacted, upper wisdom teeth and placed them onto a “scaffold” made of collagen sponge. They then used this mixture to fill in the injured areas that remained after the impacted teeth were removed from the lower jaw. The area in the upper jaw served as a control, or comparison, since no dental stem cells were used in that region.

Three months after treatment, the bone had completely regenerated at the injury site and the periodontal tissue had been restored. In the seven patients who returned for one-year follow-up examinations, optimal bone regeneration was observed. The investigators concluded that dental stem cells embedded onto a collagen sponge scaffold can completely restore bone defects in the human jaw. Furthermore, these cells have the potential to repair and/or regenerate tissues and organs.

Before the publication of this paper, jaw defects had been repaired using dental stem cells in an animal model, but never in humans. In fact, no dental stem cell therapies have ever been used in human patients. This bone grafting study is very exciting for the future promise of dental stem cell therapies. It does not matter if the dental stem cells come from a dental stem cell bank, such as the National Dental Pulp Laboratory, or individuals who wish to preserve their own or their children’s pulp in order to have a source of stem cells that they might be able to put to use for future medical needs.

The Legal Case Against Lance Armstrong: Drug Doping.

Unless you are living under a rock, you have probably heard or read that the United States Anti-Doping Agency (USADA) has stripped American cyclist Lance Armstrong of his seven Tour de France victories and all his victories and awards for the last 14 years. In the eyes of the USADA, Armstrong is a cheater.

The facts remain that Armstrong is probably the most heavily tested athlete in the world, and has submitted to over 500 official blood and urine and other types of tests. It is also a fact that never once has Armstrong failed an OFFICIAL drug test, even though there have been unofficial tests that did indicate that he took performance-enhancing drugs (PEDs). The case against Armstrong is entirely circumstantial and relies on the testimonies of convicted dopers who rode with Armstrong or on the testimonies of those who worked with him. On his radio show, conservative commentator Rush Limbaugh said there was no evidence that Armstrong used PEDs. That’s not exactly right. There is no direct evidence from drug tests that Armstrong used PEDs, but the fact remains that there is a strong circumstantial case against him. In this entry, I thought I might bring that evidence to my readers.

The investigation into Armstrong falls under the aegis of the US Food and Drug Administration and is directed by agent Jeff Novitsky. It begins when Armstrong initiated his quest to be an Olympic athlete at the age of 15 (ca 1990). He began his career as a triathlete, but switched to cycling.

In 1990, two teammates of Armstrong, Greg Stock and Erich Kaiter claim in a 20o0 lawsuit against USA Cycling that their coaches regularly administered steroids to them in 1990 and that the side effects of these steroid injections damaged their health and cut their athletic careers short. USA Cycling settled this lawsuit in 2006 by paying each rider $250,000.  Since these two athletes were teammates of Armstrong’s and strongly allege that they were given steroids by their coaches, and since Armstrong was coached by the same people, it is likely that Armstrong received at least some steroid treatments as well during this time.  Whether or not he received them knowingly or unknowingly is difficult to ascertain.

During 1990-2000, Lance Armstrong was tested more than two dozen times. His blood was sent to the UCLA laboratory of the famous antidoping scientist Donald Caitlin. Caitlin is somewhat of a legend in the anti-doping world, since he designed methods to distinguish between synthetic and natural testosterone. Caitlin also discovered the illicit drug Tetrahydrogestrinone (THG), which is an extremely potent anabolic steroid that can induce muscle growth with just a few drops under the tongue, and was distributed by the Bay Area Laboratory Co-Operative (BALCO) to many different athletes, including baseball player Barry Bonds, and sprinters Marion Jones and Tim Montgomery. Caitlin served as an expert witness in cases brought by the USADA and in federal investigations of the BALCO, which was also led by Novitsky.

In May 1999, USA Cycling sent Caitlin a formal request for test results from a cyclist who was identified only by a code. The code, however, was for a cyclist named Lance Armstrong. Of these tests, three tests, June 23, 1993, July 7, 1994, and June 4, 1996 show rather high ratios of testosterone to epitestosterone. Epitestosterone is a natural variant chemical form of testosterone. In most healthy males, the ratio testosterone to epitestosterone is around 1:1, but in athletes, this ratio can vary. For this reason, the highest ratio allowed by most regulations is 4:1, since the highest recorded ratio in a non-doping athlete is 5.25 to 1 (very rare). Taking oral testosterone or testosterone-like drugs will raise the level of testosterone but not epitestosterone. Therefore, a high testosterone to epitestosterone ratio in a blood test is almost certainly (though not always) indicative of taking anabolic steroids. Armstrong’s test ratios were 9 to 1, 7.6 to 1, and 6.5 to 1.

Why wasn’t Armstrong sanctioned at this time? Caitlin apparently retested the sampled to verify the high ratios and the second tests were normal. Thus the high levels could not be officially verified. Several drug testing experts find this very odd that three high tests for the same athlete could not be confirmed, but the simple fact remains, that Caitlin was not able to officially document high testosterone levels in Armstrong’s blood during 1990-2000. Some think that something funny is going on. Caitlin has been described by some as one of Armstrong’s greatest fans, and some have asserted, with little proof, that Caitlin would have gladly faked the test for Armstrong’s sake. This is a very weighty accusation that is being made without any evidence. It seems it is impossible to say what happened with any certainty.

The next witness is Stephanie McIlvain, who is a marketing representative for Oakley sunglasses. She provided Armstrong with all the latest eye wear and clothing from her company and was an Armstrong confidant. When Armstrong was diagnosed with stage III testicular cancer on October, 1996, McIlvain was by his bedside after his surgery 25 days later. Around this time, while sitting in a hospital conference room in Indianapolis, watching a football game, two physicians came into the room and directly asked Armstrong if he had ever used PEDs. According to the sworn testimonies of Armstrong’s former teammate, Frankie Andreu and his wife Betsy Andreu, Armstrong responded to the doctors by admitting that he had taken PEDs that the drugs he had taken were growth hormone, cortisone, anabolic steroids, testosterone, and the hormone erythropoietin (made by the kidneys, it boosts the quantity of red blood cells in the bloodstream thus increasing oxygen delivery to the tissues).

In their testimonies, Armstrong and McIlvain deny that Armstrong said such a thing and Armstrong even denied that physicians had ever asked him such a question. However, in a phone conversation with three-time Tour de France winner Greg LeMond that was apparently secretly taped by LeMond, McIlvain admitted that Armstrong had said exactly what Betsy Andreu had recounted in her testimony. McIlvain allegedly said, “You know I was in that room. I heard it, you know.” She also added, “So many people protesting him that it is just sickening, you know.” In statements to Sports Illustrated, McIlvain said that she stands by her court testimonies.

The next stretch of witnesses are mostly former teammates of Armstrong’s. Stephen Swart was a member of the 1995 Tour de France Motorola team with Lance Armstrong. According to Swart, Armstrong instigated the use of EPO (erythropoietin) to raise their hematocrits (the percentage of blood composed of red blood cells). EPO had been banned by the International Olympic Committee in 1990. According to Swart and Andreu, who were teammates of Armstrong’s, Lance Armstrong was the leader of the cycling team and he determined which riders stayed or were replaced. According to Swart, Armstrong suggested that they all take EPO and it was the force of Armstrong’s personality that persuaded the team to take EPO.

Swat admits to using EPO while he was riding with Motorola, and he told about a small hemotological analyzer that was brought into the hotel room every night to test their hematocrits. This would tell them how much EPO to use, since too much EPO can crowd the blood with so many red blood cells that the thickened blood will spontaneously clot within the blood vessels, thus setting off a heart attack or a stroke.

A normal hematocrit for a healthy male ranges from 42 to 52. Athletes will tend to have higher hematocrits, but a hematocrit of over 50 will typically get you banned from cycling for 15 days. Swart recalls on July 17, 1995, his hematocrit was 48, but Armstrong’s hematocrit was 54-56.

Tests to detect EPO in the blood rely on differences between recombinant EPO and native EPO. Native EPO is decorated with all kinds of sugar groups, but the recombinant EPO has a very different combination of sugar groups attached to it. By detecting these aberrant forms of EPO in the blood, labs can detect EPO in a blood test. This test was developed by scientists at the French national anti-doping laboratory (LNDD) in 2000. The World Anti-Doping Agency (WADA) endorsed this test, and introduced it to detect pharmaceutical EPO, since it could distinguish exogenous EPO from the natural hormone normally present in an athlete’s urine. Swart said that he never saw Armstrong inject himself with EPO and he never saw Armstrong give anyone of the team EPO. If this is the case, then where did the EPO come from?

In the mid-1990s, Armstrong commenced his relationship with the Italian physician Michele Ferrari. Ferrari has publicly defended the use of EPO by athletes, even though he has also denied doping athletes. Armstrong officially ended his relationship with Ferrari in 2004 after Ferrari was convicted of sport fraud by an Italian court.

Nevertheless, former Armstrong teammate and convicted doper Floyd Landis, Dr. Ferrari was “matter-a-fact about” doping. According to Landis, Ferrari helped Landis transfuse his own blood to boost his performance. He said that Ferrari was professional and calculated about his work.

Landis also said that Ferrari was concerned after Armstrong’s cancer diagnosis because he thought that his doping regime might have brought on Armstrong’s cancer. According to Landis: “I remember when we were on a training ride in 2002. Lance told me that Ferrari had been paranoid that he had helped cause the cancer and become more conservative after that.”

Landis also recalls that Armstrong and his cycling team always flew on private charters because there were less stringent customs checks as private airports. However, according to Landis, in 2003, at the airport in St. Moritz, Switzerland, customs agents stopped Armstrong and searched his duffel bag. There were drugs with labels in Spanish and syringes in the duffel bag. Armstrong asked someone in the contingent to tell the customs officers that the syringes and bottles were vitamin-injecting gear. The customs officers, says Landis, “looked at us sideways but let us through.” Armstrong denies that any of this occurred.

Landis won the Tour de France in 2006, but was stripped of his victory after his testosterone to epitestosterone levels were shown to be close to 11 to 1. Landis sent emails to USA Cycling in April and May of 2011, accusing Armstrong of doping while he and Landis were members of the US Postal Service Cycling team. In May 2011, Landis filed a whistle-blower lawsuit against Armstrong under the US False Claims Act. This Act allows citizens who possess evidence that someone has defrauded the US Government to sue on behalf of the Government. Armstrong has responded by labeling Landis as “a person with zero credibility.”

Finally, Armstrong’s bike mechanic and personal assistant from 2002 to 2004, Mike Anderson recalls several instances that are potentially incriminating if true. According to Anderson, Armstrong sent him to Girona, Spain to remove all traces of his ex-wife Kristin before Armstrong arrived with his girlfriend Sheryl Crow. Anderson said that in the bathroom cabinet of this apartment was a white box labeled “ANDRO.”

According to papers filed in 2005, Anderson sued Armstrong for allegedly backing out of their business deal that included Armstrong helping Anderson build a bike shop. Anderson’s lawyers wrote that the substance in the while box was “androstenin or something very close to that.” This piqued the interest of Novitsky at the FDA who was sure that the substance in the white box was androstenedione, the compound that was found in baseball player Mark McGwire’s locker. The International Olympic Committee banned androstenedione in 1997. Armstrong denies ever having taken androstenedione, and without the box or testing its contents, it seems impossible to know what was ever in it.

The Anderson-Armstrong lawsuit also alleges that shortly after winning his 6th Tour de France in 2004, Anderson received a phone call from a friend of Armstrong’s that USADA drug testers had shown up at the Armstrong ranch in Dripping Springs, Texas, and Armstrong was not there. Athletes subject to random drug testing must inform the USADA of their location and missing three random tests within an 18-month period constitutes a violation.

Anderson asserts that he was involved in an attempt to fool USADA agents by keeping watch over the USADA agents in their white SUV while a friend of Armstrong’s drove Lance’s black Suburban from the airport terminal to the ranch to make it look as though Armstrong had been there all along and the USADA agents simply could not raise him for whatever reason.

The two Armstrong friends who allegedly collaborated with Anderson in this cover-up, Derek Russey and John Korioth, deny that this ever happened that and Anderson fabricated the entire tale. USADA still seems to think that Anderson is telling the truth.

Is Armstrong guilty? USADA certainly thinks so. The case is largely circumstantial, but the cumulative case is somewhat strong, but the evidence is all indirect. The testimonies are largely from athletes who are admitted dopers, which certainly taints their credibility. Landis, for example, was also found guilty of fraud. His credibility is certainly low.

However, why would Betsy Andreu lie about what she heard Armstrong say? Her tenacity about this whole thing has bought her little more than tremendous grief from Armstrong’s supporters. Her motive for lying seems low. Likewise, the conversation between LeMond and McIlvair, if it is true, would seem to corroborate the stories of Betsy Andreu and her husband.

It seems to me that at least at some point in his career, Armstrong did use PEDs. Whether he was a serial abuser is difficult to say. Whether all seven of his Tour de France victories were due to PED use is also unproven, in my opinion. It also seems to me that Armstrong has had relationships with noted dopers, such as Dr. Ferrari.

Did Ferrari help Armstrong dope? We know that he helped Landis dope, and it seems highly unlikely that Armstrong would have had a relationship with a physician such as Ferrari unless he wanted Ferrari to help him dope. Therefore, it seems likely to me that at some point in his career, Armstrong did use PEDs. When and where? I do not know and I do not think the USADA knows either. I would say that stripping him of the last 14 years of his career seems more than a little extreme. I would say that it looks to me as though they want to make an example of Armstrong.

If Armstrong truly did all he did while doping, many of the riders he beat were also doping. Thus it seems difficult to assign an unfair advantage to Armstrong during this time. Secondly, if Armstrong was a chronic serial doper, he found a certifiable way to beat the hundreds of tests he was subjected to. He has never officially tested positive for PEDs, which is a testimony to the difficulty of detecting PEDs in the blood of athletes who dope and do so smartly. It shows in some ways how far behind the detection technology lags when it comes to catching up with smart dopers.

For those interested in this subject, please read Chris Cooper’s excellent book, Run, Swim, Throw, Cheat:  The Science Behind Drugs in Sport (Oxford University Press, 2012).

Decreased Oxygen Increases Stem Cell Survival When Treating Muscular Dystrophy


In order to increase the survival of stem cells in culture, research from Purdue University has shown that controlling the amount of oxygen to which the stem cells are exposed, can significantly increase the effectiveness of stem cell-based procedures to treat an often fatal form of muscular dystrophy.

Mutations in the dystrophin gene cause Duchenne muscular dystrophy (DMD). DMD causes the constant breakdown of muscles, and the gradual depletion of muscle-based stem cells that repair the damage and progressive muscle wasting that characterize this disease. Healthy stem cells divide and repair damaged muscles or other tissues. This is the basis of stem cell therapy, which is the implantation of healthy stem cells to repair degenerating or injured tissues. Stem cell therapies have shown promise against DMD and other neurodegenerative diseases, but a major caveat of these treatments is that few of the implanted stem cells survive the procedure.

Shihuan Kuang, a Purdue assistant professor of animal sciences, and Weiyi Liu, a postdoctoral research associate, have shown that the survival of implanted muscle stem cells is increased by as much as fivefold in a mouse model if the cells are cultured under oxygen levels similar to those found in human muscles.

“Stem cells survive in a microenvironment in the body that has a low oxygen level,” Kuang said. “But when we culture cells, there is a lot of oxygen around the petri dish. We wanted to see if less oxygen could mimic that microenvironment. When we did that, we saw that more stem cells survived the transplant.”

According to Liu, when stem cells are grown in higher oxygen levels, they adapt to those levels in order to acclimate to their surroundings. Therefore, when these same cells are injected into muscles with lower oxygen levels, they die from a lack of sufficient oxygen (in layman’s terms, they suffocate).

However, when the cells are grown in lower oxygen levels, they survive when transplanted into muscles. As Liu said, “By contrast, in our study the cells become used to the host environment when they are conditioned under low oxygen levels prior to transplantation.”

Their experiments were done in mice, and when transplanted into the skeletal muscles of DMD mice, Kuang and Liu saw more stem cells survive the transplants. Furthermore, those stem cells retained their ability to duplicate themselves.

“When we lower the oxygen level, we can also maintain the self-renewal process,” Kuang said. “If these stem cells self-renew, they should never be used up and should continue to repair damaged muscle.”

These studies were published in the journal Development, and this protocol shows promise as a strategy to increase the effectiveness of stem cell therapy for DMD patients. DMD occurs in about one in 3,500 boys, and symptoms being at about 3-5 years old. DMD confines almost all patients to wheelchairs by their 20s, is often fatal as muscles that control the abilities to breathe and eat deteriorate.

For future studies, Kuang’s lab will examine those signaling pathways within stem cells that are affected by low oxygen levels. They will determine if human muscle stem cells are similarly regulated by environmental oxygen.

This research was funded by the The National Institutes of Health, the Muscular Dystrophy Association and the U.S. Department of Agriculture funded the research.

NCKU study: Nanomaterials help heart to heal

In advanced countries, heart disease is the major cause of death. In Taiwan, there are about 2 million patients with heart disease every year, and 400,000 deaths, mostly due to coronary artery disease, which leads to heart failure.

A research team led by Patrick C. H. Hsieh, an associate professor and cardiac surgeon at the National Cheng Kung University (NCKU) in Taiwan has discovered a new way to regenerate blood vessels by using nanofibers and a growth factor known as vascular endothelial growth factor (VEGF).

This new technology will help endogenous stem cells regenerate the heart and blood vessels and might be a promising approach to cure heart diseases. The president of NCKU, Hwung-Hweng Hwung, praised this research team’s achievements when he called Hsieh is “a good professor dedicated to scientific research that is beneficial to society.” According to Hsieh, NCKU will assist Hsieh’s team transfer this technology from the laboratory to the clinic in the hope that this novel treatment will be available for people with heart diseases.

Hsieh’s study combines tissue engineering, nanotechnology and controlled protein delivery in order to induce endogenous stem cells to regenerate blood vessels and improve cardiac function.

The growth factor, VEGF, is one of the main players for the growth and formation of blood vessels (see Ahluwalia A, Tarnawski AS. Critical role of hypoxia sensor–HIF-1α in VEGF gene activation. Implications for angiogenesis and tissue injury healing. Curr Med Chem. 2012;19(1):90-7). Even though VEGF has been used in clinical settings as treatment for cardiovascular diseases, it has not worked terribly well (see Simón-Yarza T, et al., Theranostics. 2012;2(6):541-52). However, when VEGF was combined with nanomaterial, the results were quite different.

According to Hsieh: “The combination of nanomaterial with VEGF works well for the nanofibers to create a favorable microenvironment in the heart for recruiting stem cells.”

These experiments were done in rats and pigs, but the implantation of VEGF with led to the growth of newly formed blood vessels, which improved heart function without harmful side effects.

The laboratory animals were treated immediately after heart attacks were induced. Unfortunately, in human patients, this would not be possible. Instead this treatment might be effective if given in the first week after a heart attack, when stem-cell activity in the bone marrow and heart are still high.

This study was published by Science Translational Medicine on Wednesday, and the journals Science and Nature reported the NCUK study and noted it for its potential importance.

The Protein That Keeps Stem Cells Stem Cells

Stem cells have the ability to differentiate into a wide variety of distinct cell types. How do they do it? The answer to this question is certainly one of the Holy Grails of stem cell research. Fortunately, a study from a laboratory at the University of Michigan Medical School team has elucidated one of the main mechanisms by which this happens.

An article published in the journal Cell Stem Cell by stem cell researcher Yali Dou, Ph.D., and her team focuses on the role played by a protein called Mof. Apparently Mof in preserves the undifferentiated state of stem cells and also primes them to become specialized cells in mice.

The Mof protein plays a role in “epigenetic” regulation of gene expression. Mof is an enzyme that modifies histone proteins in a very specific manner. Histones, you might remember, are proteins that package DNA into chromatin. Previous studies have shown that the chromatin in embryonic stem cells is very relaxed and open, which makes gene expression relatively easy in embryonic stem cells. Mof acts as a “histone H4 lysine 16-specific acetyltransferase.” In other words, Mof adds acetic acid groups (CH3COO-) to a particular amino acid (lysine 16) of histone protein H4. The addition of a negatively-charged chemical group such as acetic acid to the positively-charged histone proteins neutralizes the positive charges of the histone proteins, which decreases the affinity of histone proteins for DNA, since DNA is negatively charged. Mof, therefore, modifies the proteins that assemble DNA into chromatin and opens up that DNA to the gene expression machinery.

Dou is an associate professor of pathology and biological chemistry, and her lab has studied Mof for several years. Modification of histone proteins by MOF seem to act as tiny beacons and guide the gene expression machinery to the right place so that proper gene expression can occur. Without these tiny Mof-synthesized beacons, the stem cells lose their undifferentiated state and undergo differentiation.

“Simply put, Mof regulates the core transcription mechanism – without it you can’t be a stem cell,” says Dou. “There are many such proteins, called histone acetyltransferases, in cells – but only MOF is important in undifferentiated cells.”

Dou’s lab has also examined a protein called WDR5 that is another histone-modifying enzyme. WDR5 is a component of a complex that makes additional changes to histone proteins. One of these changes involves the addition of a methyl group (-CH3) to histone protein H3, specifically lysine 4. The addition of this methyl group is strongly associated with the activation of gene expression. WDR5 is one of the four components that make up the enzyme that plasters this methyl group onto H3. However, Dou’s research and the data generated in her lab shows that Mof controls the addition of the methyl group on H3 by the complex of which WDR5 is part. Without the function of Mof, WDR5 does not add its methyl group, and the gene expression machinery does not get the message to express particular genes. The genes expressed by this process, are those responsible for stem-ness (the undifferentiated state). As explained by Dou: “Without Mof, embryonic stem cells lost their self-renewal capability and started to differentiate,” she explains.

The new findings may have particular importance for work on induced pluripotent stem cells (iPSCs), which do not come from an embryo, but are made from adult tissue.

IPCS research is an extremely promising technology, since it would allow a patient to be treated with stem cells made from their own tissue. Unfortunately, the way iPSCs are typically made overexpresses genes in cells that can cause cancer when overexpressed in other types of cells.

Dou says that further work on Mof might allow researchers to make iPSCs without overexpressing such dangerous genes. However, further research is needed.

Dou plans to continue to work on how Mof marks chromatin in order to keep parts of the genome readily accessible to the gene expression machinery. Once a stem cell starts to differentiate, or become a certain specialized type of cell, parts of the DNA close up and become less accessible. Many scientific teams have studied this “selective silencing” and those factors that direct stem cells to differentiate by expressing only certain genes. Few labs, however, have examined those factors that broadly regulate DNA transcription to preserve stem-ness.

“Mof marks the areas that need to stay open and maintains the potential to become anything,” Dou explains. Its crucial role in many species is hinted at by the fact that the gene to make Mof has the same sequence in fruit flies and mice.

“If you think about stem cell biology, the self-renewal is one aspect that makes stem cells unique and powerful, and the differentiation is another,” says Dou. “People have looked a lot at differentiation to make cells useful for therapy in the future – but the stem cell itself is actually pretty fascinating. So far, Mof is the only histone acetyltransferase found to support the stemness of embryonic stem cells.”

Resveratrol Protects Heart-Based Stem Cells from Malfunctioning in Diabetic Mice

Resveratrol is a compound found in red wine, and other foods. This remarkable compound has powerful health benefits that are just now being more completely understood.

This present report tested the ability of resveratrol to affect the hearts of diabetic mice. Some studies have shown that high blood glucose levels cause endothelial progenitor cells (EPCs) to fall asleep and those that do not go to sleep are sluggish and display impaired proliferation, adhesion, and migration capacities (see Orlandi A, et al., (2010) Basic Res Cardiol 105: 703–712 & Balestrieri ML, et al. (2008) Biochim Biophys Acta 1784: 936–945). This results in decreased blood vessel construction. New evidence also shows that cardiac stem/progenitor cells (CSPCs) play a role in the heart problem found in diabetics (see Rota M,et al. (2006) Circ Res 99: 42–52). Hyperglycemia-induced CSPC damage negatively affects cardiac structure and function in diabetics.

In this publication, Francesca Delucchi and colleagues in the laboratory of Donatella Stilli at the Dipartimento Biologia Evolutiva e Funzionale, Università di Parma, Parma, Italy, determined if early administration of resveratrol could improve heart muscle function. They were also interested in determining if resveratrol protected the progenitor cell pool or simply improved the environment within the heart.

In this study, 128 adult Wistar rats (n = 128) with drug-induced type-1 diabetes were either untreated (D group; n = 54) or subjected to administration of trans-resveratrol (i.p. injection: 2.5 mg/Kg/day; DR group; n = 64). Twenty-five rats constituted the control group (C).

After 1, 3 or 8 weeks of hyperglycemia, they evaluated heart function and heart muscle function. They also determined how mush inflammation was occurring in the heart. Finally, they cultured CSPCs to determine the effects of environment on them.

The results were rather interesting. In diabetic animals not treated with resveratrol, cardiac function was maintained during the first 3 weeks of hyperglycemia, but the ventricles were already enlarging and the CSPCs were also dying. Signs of ventricular abnormalities appeared after 8 weeks of diabetes.

Administration of resveratrol reduced atrial CSPC loss, and succeeded in preserving the functional abilities of CSPCs and mature cardiac cells. In addition, resveratrol administration improved cardiac environment by reducing inflammation. It also decreased ventricular remodeling in diabetic hearts.

In culture, CSPCs from the resveratrol-treated mice prevented the release of pro-inflammation molecules from co-cultured heart muscle cells.  The same cells from non-treated diabetic mice released a great deal of pro-inflammatory molecules.  the resveratrol-treated cells quelled the release of inflammation-promoting molecules and, consequently, the production of molecules called “Reactive oxygen species,” which are also known as “free radicals.”  Fewer free radicals caused less cell damage and consequently less inflammation.

These findings indicate that resveratrol might play a support role as a so-called adjuvant therapeutic option in diabetic heart failure prevention.

The Ajuba Gene Controls Stem Cells During Heart Development

Heart development is a very complex and multistep process that requires interactions from several different cell types, and also has many crucial stages where things can go wrong. Unfortunately, this is precisely what happens many times and many children are born with congenital heart defects. According to the March of Dimes, of the 10,830 babies born in the US each day, approximately 87 of them have some kind of congenital heart defect (roughly 0.8% of all babies born). This means that about one-half million children in the U.S. have some form of cardiac problem if we exclude high blood pressure. According to the American Heart Association, there are approximately one million people alive with congenital heart defects today. (AHA) Also, heart defects can be part of a wider pattern of birth defects. For example,. more than one-third of children with Down syndrome have heart defects.

Finding those genes that control the crucial steps in heart development can help cardiologists screen for children with congenital heart problems and devise better treatments and even cures for them.

To this end, a German research group had identified a gene called “Ajuba” that regulates stem cell activity in the heart during embryonic development.

First, let’s review a little about how the heart develops in humans. The process of gastrulation, which begins around day 14 converts the single-layer-thick embryo into a three-layered-thick structure. Gastrulation is a complex process and takes several days, but by day 15, a group of cells have moved to the front of the embryo, just under the upper layer of the embryo. These cells that have moved to the front form a horseshoe-shaped structure are called the “cardiogenic mesoderm.” Cardiogenic mesoderm just means “the stuff that will form the heart.” The horseshoe-shaped cells roll up into a tube and immediately begin to beat. The horseshoe also contains an upper tube and a lower tube. The upper tube is called the “Primary heart field” and the tube below is called the “secondary heart field.” The two tubes are not isolated, but are connected at three points.  For pictures see here.

For the nest three days, the primary cardiac field expands and bulges out. and this bulge will form the left ventricle. The rest of the heart will be formed by the secondary heart field. The top of the secondary heart field that was attached to the primary heart field becomes the outflow vessels that empties the ventricle and the lower part of the secondary field expands to form two inflow vessels and the atrium, which will become the upper chambers of the heart.

During the 4th week of development, the heart does an intricate bit of developmental ballet known as “heart looping.” First the left ventricle enlarges and becomes a kind of spherical chamber (days 21-22). The right ventricle is just slightly visible next to the left ventricle. Then the atrium elongates mightily and the right ventricle enlarges. The rapid growth of the atrium swings the attachment between the left ventricle upwards and to the left. The atrium and left ventricle continue to grow from days 26-28 and a division between the left and right atrium begins to form and the division between the left and right ventricle also becomes more obvious.

By day 34, a septum forms between the right and left atrium. This process begins with a primary septum that forms but then dies off at the top. A secondary septum forms as a ridge between the two atria on the top edge, in the back. This ridge grows and fuses with a tissue called the endocardial cushions, which are the beginnings of the heart valves. Thus the primary septum is incomplete at its top and the secondary septum is incomplete at the bottom. These incomplete septa allow blood flow between the two sides of the heart. Since there are no operational lungs at this time in development, this set up works rather well. Thus, these two septa stay that way until birth when the baby takes her first breath. The increased pressure on the left side of the heart pushes the two septa together and they fuse.  For pictures see here.

The ventricles also divide by this time. From days 38 – 46, the ventricular septum grows from the bottom of the ventricle to the top of it, dividing the ventricles into left and ventricles

About day 35, the outflow tract, which is undivided at this point, begins to be divided to eventually form the aorta and the pulmonary artery. The division occurs because swellings organized into a spiral all along the outflow tract wall grow into the tract until they ingrowths from the two sides meet, dividing the outflow tract. By day 56, the two outflow tracts are completely divided.

During these events, specific genes are expressed in various parts of the heart, inducing specific patterns of growth and differentiation. For example, a gene called Hand1 is localized specifically to the outer curvature of the presumptive left ventricle and developing outflow tract, and also at lower levels in the outer curvature of the right ventricle. Mutations in Hand1 in humans can cause hypoplasia (poor growth) of the left ventricle (HLHS).

Now that we have a basic sense of how the heart develops, we can see that this finding expands our knowledge of heart development in the following way: When the primary and secondary heart fields are growing, specific genes prevent the secondary field from growing as fast as the primary field. A signaling molecule called retinoic acid (RA) (derived from vitamin A) signals to the secondary field to grow more slowly, and the Ajuba gene is the target for retinoic acid signaling. The Ajuba protein prevents the activity of a protein called Isl-1. With reduced Isl-1 activity in the secondary field, the heart-specific stem cells do not grow as fast as those in the primary field.

Ajuba-deficient animals show an increased pool of Isl1(+) heart stem cells and dramatically increased numbers of heart muscle cells at the inflow and outflow tissues. This puts heart muscle cells where they should not be and the heart’s coordinated development is compromised.

Gergana Dobreva, the head of the Bad Nauheim-based Max Planck Institute, and the senior author of this paper said of the Ajuba-deficient zebrafish, “In almost all the investigated [zebra]fish we observed a dramatic enlargement of the heart. If Ajuba is absent, there is clearly no other switch that finally silences the Isl-1 controlled part of cardiac development.”

What are the clinical applications of this study? Dobreva thinks: “Once we understand how cardiac development is regulated, we will also be more familiar with the causes of congenital heart defects and will consequently be able to consider therapeutic approaches.”

Dobreva continued: “One possibility would be to optimize the production of replacement cells from embryonic or artificially produced stem cells in the laboratory. Silencing Ajuba in these cells might enhance their development into functional cardiac muscle cells. Sufficient replacement cells for treating patients could be cultured in this way.”

Finally, another possibility would be to temporarily silence Ajuba in the adult heart and let it repair itself. Further studies are in the works

New Brain Stem Cell And Higher Cortical Functions

Neuroscientists at the Scripps Research Institute in La Jolla, California have identified a new stem cell population in the brain that might differentiate into those neurons responsible for higher thinking. Also, by culturing these neurons in the laboratory, scientists might be able to design better treatments for those cognitive disorders, such as schizophrenia and autism that result from abnormal connections among particular brain cells.

This new research also illustrated how neurons in the uppermost layers of the cerebral cortex form during embryonic development of the brain.

Senior author of this work, Ulrich Mueller, professor and director of the Dorris Neuroscience Center at Scripps Research, commented: “The cerebral cortex is the seat of higher brain function, where information gets integrated and where we form memories and consciousness. If we want to understand who we are, we need to understand this area where everything comes together and forms our impression of the world.”

Previously, scientists thought that all cortical neurons, whether they occupied the lower or upper layers of the brain, were derived from the same stem cell; a cell called the radial glial cell (RGC). The fate of neurons were thought to result from when they were born with the earliest neurons migrating only a little and staying close to where they were born (lower layers), and later born neurons migrating further from where they were born (uppermost layers).

Mueller’s research team, however, has identified a neural stem cells that specifically gives rise to neurons that make the upper layers of the cerebral cortex, regardless of the time or place of birth.

Santos Franco, a senior research associate in the Mueller Laboratory said, “Advanced functions like consciousness, thought, and creativity require a lot of different neuronal cell types and a central question has been how all this diversity is produced in the cortex. Our study shows this diversity already exists in the progenitor cells.”

According to Mueller: “The [older] model was that there is a stem cell in the center of the ball that generated the different types of neurons in successive waves. What we now show is that there are at least two different populations of RGCs and potentially more.”

Franco used a mouse strain that he had constructed in which he could track upper-layer neurons as they were born and as they migrated. A marker gene called Cux2 is only expressed by upper-layer neurons, and Franco used an enzyme from bacterial viruses called the Cre protein to flip on a red-glowing protein when Cux2 is expressed.

To their surprise, a population of RGCs flipped on Cux2 at the earliest time of their development (embryonic day 9-10). The problem is that no upper layer neurons exist at this early time in development, which means that these cells are programmed to form upper layer neurons even though no such tissue exists at this time. Non-Cux-2-expressing neurons became lower layer neurons.

Culturing Cux-2-expressing neurons in the laboratory they formed the types of neurons normally found in the upper layer of the brain. Likewise, non-Cux2-expressing neurons formed other types of neurons normally found in the lower layers of the brain.

During development, Cux2-positive stem cells proliferate and self-renew before they differentiate into neurons. Does the birthday of the neuron determine it’s eventual developmental fate? To determine if this is the case, Mueller and his colleagues used a molecule called TCF4 to force premature differentiation of the Cux2-expressing cells. Even under these conditions, the Cux2-expressing cells still formed upper layer neurons.

Thus regardless of their birth date or location of their birth, they still form upper layer neurons. As Mueller puts it, these RGCs have some intrinsic property that determined their cell fate from the start.

This RGC subset is responsible for the huge proliferation of cells required to generate the larger upper-layer cortex found in the brains of primates. With bigger brains, however, comes the risk of disorders from upper-layer neuron connection abnormalities. TO date, researchers have only managed to generate lower-layer neurons from stem cells in the laboratory. According to Mueller, “The opens a door now to try to make the upper-layer neurons, which are frequently affected in psychiatric disorders.”

Creating Cartilage

Cartilage is the shock absorber of the body. It allows two bones to move past each other without deleterious effects. Today, we walk on paved streets and carpeted buildings with stiff floors. Our cartilage takes a constant beating and as we age, it has a tougher and tougher time bouncing back.

As we age, the daily wear and tear eventually grinds this tissue down and movement can become painful and tedious. Osteoarthritis affects over 27 million Americans and it can cause pain, stiff joints, cracking sounds, inflammation and bone spurs.

Cartilage formation depends upon the activity of one cell type – the chondrocyte. With advancing age, chondrocytes divide less and less and eventually, they fall behind making new cartilage to repair the defects generated by everyday wear and tear. In the long-term, chondrocytes can respond to stress by simply dying-off.

Orthopedic specialists consider cartilage regeneration the holy grail of orthopedic medicine. Since people are living longer, orthopedists have taken to cleaning out damaged joints with arthroscopic surgery, braces to stabilize a wobbly gait, and artificial knees and hips to replace heavily damaged joints.

Now, stem cell technology has given the hope of actually re-making new cartilage in aging,arthritic patients. Bio-engineers are working hard to crack the nut of cartilage production. They have identified prominent proteins required to turn stem cells into chrondrocytes, and have also designed three-dimensional scaffolds upon which stem cells will grow and eventually make cartilage. Cartilage-forming cells seem to behave normally if they are constantly surrounded by molecules found in normal cartilage. Also the scaffolding derived from cartilage seems to provide many of the molecular prompts for cartilage behavior.

Much of this is still in the experimental phase, the “stem cell strategy” for re-synthesizing cartilage seems to be one of the best possibilities and clinical trials are in the works in Norway, Spain, Iran, Malaysia, and other places too.

Tissue engineering had its start in the 1970s, and then it comes to cartilage, it has certainly had its ups and downs. For example, the type of cartilage found at the ends of long bones is known as “hyaline cartilage” because of its slippery feel, glassy look, elastic properties and smooth texture. Hyaline cartilage is a terrific weight-bearing cap. Nevertheless attempts to make cartilage at joints has resulted in the production of “fibrocartilage.”

For example, surgeons have often treated arthritic joints by cleaning out bone spurs, scar tissue, and then drilling small holes into the ends of the bones. This causes stem cells to move into the joint and make new cartilage, but they make fibrocartilage instead of hyaline cartilage. Fibrocartilage is found at the place where our pelvic bones join, our intervertebral discs, and our jaw joint. It does not have the ability to resist impact forces the way hyaline cartilage does. Therefore, it erodes quickly at joints. John Sandy, a biochemist at Rush University Medical Center, Chicago put it this way, “Those stem cells that come out are confused. They’re not getting the right signals….So they hit the middle road.” In a study that examined microfracture surgery, two-thirds of athletes who had the procedure showed good results, but only half of those were able to play at their original level for several years.

Chondrocyte transplantation has also been attempted. A Cambridge, MA biotech company called Genzyme has an off-the-shelf product known as Carticel that takes thousands of live chondrocytes from healthy cartilage elsewhere in the body and cultures them to expand their numbers. The expanded chondrocytes are then injected into the affected site. This is known as autologous chondrocyte implantation and this procedure has outperformed microfracture surgery, at least in some studies. Unfortunately, some patients need follow-up surgery, and a nine-year study found that 50% of patients did not improve at all (see Christopher M. Revell, and Kyriacos A. Athanasiou, Success Rates and Immunologic Responses of Autogenic, Allogenic, and Xenogenic Treatments to Repair Articular Cartilage Defects. Tissue Eng Part B Rev. 2009 March; 15(1): 1–15). According to Wan-Ju Li, a tissue engineer at the University of Wisconsin, Madison, chondrocytes lose their ability to form cartilage if they are grown for too many generations in culture.

Can other cells be used to form cartilage? The answer is a clear “yes.” Stem cells from cartilage, tendons, and synovial membranes (the sac that surrounds the joint) can all form cartilage, as can stem cells from fat, and umbilical cord.

The next question is, “how do we coax these stem cells into making cartilage?” What works in culture dishes in the laboratory may not work inside a living joint, but certainly, getting it to work in the laboratory is the first place to start. Several compounds have been found that are definitely pro-cartilage molecules. These include a growth factor called TGF-beta, which jumps starts stems into the cartilage-forming program. However, as John Sandy explains, TGF-beta does not work alone because it will tend to drive cells to form fibrocartilage rather than hyaline cartilage.

The other growth factor needed to turn stem cells into hyaline cartilage-making chondrocytes is fibroblast growth factor-2 (FGF-2; see Andrew M. Handorf and Wan-Ju Li, Fibroblast Growth Factor-2 Primes Human Mesenchymal Stem Cells for Enhanced Chondrogenesis PLoS One 2011; 6(7): e22887). FGF-2 turns on a transcription factor in stem cells called Sox9 which switches on the production of type 2 collagen and aggrecan (two cartilage-specific proteins).

Other booster compounds that increase the cartilage-making profiles of stem cells include a synthetic molecule called kartogenin (Kristen Johnson, et al., A Stem Cell–Based Approach to Cartilage Repair. Science 11 May 2012: Vol. 336 no. 6082 pp. 717-721). Kartogenin inhibits a stem cell protein called filamin A and this unleashes all kinds of cartilage-specific processes in the stem cells. Kartogenin has taken the cartilage camp by storm, and Joan Marini of the National Institutes of Health in Bethesda, MD and Antonella Forlino of the University of Pavia in Italy wrote in the June 28 edition of the New England Journal of Medicine: “Stimulating the differentiation of one’s own stem cells by means of an easily deliverable chemical compound would be more advantageous than using conventional drilling and microfracture techniques.”

Another pro-chondrocyte protein is vimentin, which helps cells assume a round shape. According to Ricky Tuan, tissue engineer at the University of Pittsburg, vimentin pushes bone marrow stem cells into nice round cells that look like chondrocytes. Even more vimentin pushes the cells to make type 2 collagen (cartilage-specific; see Bobick B. E., Tuan R. S., Chen F. H. (2010) The intermediate filament vimentin regulates chondrogenesis of adult human bone marrow-derived multipotent progenitor cells. J. Cell. Biochem 109, 265–276).

Thus with the right blend of compounds, good hyaline cartilage can be made, but according to Ming Pei, an orthopedic surgeon and cell biologist at West Virginia University in Morgantown, making proper hyaline cartilage probably comes down to using the right stem cell. Pei thinks that stem cells from the synovial membrane have an advantage over other stem cells when it comes to cartilage making because of a substance they make.

Pei’s research team mixed the matrix made by synovial stem cells with FGF-2 in a low-oxygen environment. When they added other synovial stem cells those cells ramped up their cartilage making capabilities (Pei M, He F, Kish VL. Expansion on extracellular matrix deposited by human bone marrow stromal cells facilitates stem cell proliferation and tissue-specific lineage potential. Tissue Eng Part A. 2011 Dec;17(23-24):3067-76). These conditions seem to provide a safe place or stem cells to become chondrocytes and make cartilage. Such safe places for stem cells are called “stem cell niches.” Pei’s niche seems to be optimized for stem cells to make cartilage.

In order to get the chondrocytes to make cartilage that sticks together, they need to be in a niche that closely enough resembles their native niche. To mimic this niche, Li and Tuan started to build synthetic scaffolds that they could seed with stem cells. Such matrices definitely improved cartilage production by stem cells. According to Pei, “It’s easy to fabricate and there’s no batch-to-batch difference.”

Li notes that the polymer is designed to degrade after six-twelve months in the body and they have all the strength and mechanical properties to keep the stem cells together until they make a matrix of their own. In 2009, Tuan and Li tested their scaffold by seeding it with human stem cells so that it would create a patch that was inserted into pigs that had suffered cartilage damage. They used two types of cells to seed the matrices – stem cells and mature chondrocytes. After implanting the matrices, those that had been seeded with stem cells make hyaline cartilage, but those seeded with mature chondrocytes made fibrocartilage, after six months. Li reported, “It was glassy cartilage with good mechanical properties.”

As an alternative scaffold, scientists are also using scaffolds made from cartilage procured from cadavers. According to Pei, natural cartilage scaffolds have advantages that synthetic scaffolds do not have, such as chondrocyte-inducing molecules embedded in them.

Other laboratories are using matrices made from fibrin (the stuff blood clots are made from) and seeding with platelets, which are rich in TGF-beta. Doctors at Cairo University in Egypt seeded this scaffold with stem cells that were then implanted into the knees of five patients, all of whom reported improvements after one year.

Regardless of the exact scaffold that is used, Li is buoyantly confident that a stem cell-based strategy will result in making cartilage in the joints of aging patients.

If a treatment is found for osteoarthritis, the next question becomes, “when should the treatment be offered?” David Felson, a rheumatologist at Boston University School of Medicine. notes that knee injuries increase the likelihood a person will suffer from osteoarthritis sixfold. Felson’s research seems to indicate that such injuries “probably account for a great majority of osteoarthritis.”

Early detection is probably not practical, since most people ignore their injuries. Some patients can have bones rubbing together long before they start to experience the pain of osteoarthritis.

However, perhaps chemical markers can help detect the early signs of joint trouble. Carla Scanzello, who works at Rush University Medical Center as a rheumatologist reported that inflammatory molecules that gradually destroy cartilage leave chemical tells that can be detected and might provide a way to detect the early signs of joint damage before symptoms appear (See Carla R. Scanzello, et al., Synovial inflammation in patients undergoing arthroscopic meniscectomy: molecular characterization and relationship with symptoms. Arthritis Rheum. 2011 February; 63(2): 391–400).

Stem cell treatments might also reduce the number of patients who need artificial joint replacements. An artificial hip or knee can last 10-15 years. IF you are older, that is usually not a problem, if you are younger, that becomes a problem. According to Li, there are technical problems to be worked out, but the largest hurdles have been largely conquered, what remains is largely engineering questions. His goal is to eventually make joint replacement a thing of the past and turn orthopedic surgeons into stem cell scientists.

Epigenetic Alterations Are Linked to Cell Fate Decisions in Mesenchymal Stem Cells

Stem cell scientists at the University of California, Los Angeles (UCLA) have discovered that the activity of particular enzymes that modify the structure of chromatin is linked to the differentiation of mesenchymal stem cells into particular cell types.

A study in the laboratory from the laboratory of Cun-Yu Wang, who is a professor at the UCLA School of Dentistry, determined that even though the DNA sequences of the genomes of stem cells are not altered, the structure of their DNA is. Furthermore, these alterations to the DNA structure are inherited by the progeny of those cells. Changes in DNA that affect the appearance or behavior of cells that are not changes in the sequence of DNA are known as epigenetic changes and these epigenetic changes can greatly affect the differentiation of stem cells.

Wang’s research team discovered that two enzymes, KDM4B and KDM6B, promote the differentiation of mesenchymal stem cells (MSCs) into bone cells. KDM4B and KDM6B are “histone demethylases,” which is a fancy way of saying that they remove specific chemical groups known as “methyl groups” (—CH3) from histone proteins. Histone proteins help package DNA into a compact structure known chromatin. Chemical modification of these histones influences the nature of that chromatin. The addition of acetyl groups (-CH2-COO-) tends to make the chromatin rather loose and easily accessed by gene expression machinery. However, the addition of methyl groups tends to tighten the chromatin up and shut down gene expression. Demethylation or removing methyl groups from histones tends to take tight chromatin and loosen it up so that new gene expression is possible.

The conversion of MSCs into bone cells is stimulated by a group of growth factors called “Bone Morphogen Proteins” (BMPs). To this end, Wang and his colleagues used BMP-4 & -7 to induce the bone fate in MSCs. They discovered that BMP-4 & -7 induces expression of histone demethylases KDM4B and KDM6B. Then they determined the targets of these enzymes within MSCs. They found that KDM4B and KDM6B remove methyl groups from Histone H3 (specifically, H3K27me3 and H3K9me3). These epigenetic alterations mark the cells for a bone-making fate.

To more fully nail down the function of these histone demethylases, Wang and his crew depleted KDM4B or KDM6B in human MSCs. They found that reduction of these enzymes greatly reduced bone-cell differentiation and increased fat cell differentiation. KDM6B increases the expression of HOX genes expression by removing H3K27me3, which promotes bone differentiation. KDM4B jacked-up the expression of a gene called DLX by removing H3K9me3. DLX suppresses fat cell differentiation.

Wang and his colleagues found that in mice, the presence of H3K27me3 and H3K9me3 in MSCs are very highly correlated with an increase in osteoporosis or the aging of bone marrow in mice. Importantly, Wang’s colleagues showed that concentrations of H3K27me3- and H3K9me3-positive MSCs in mouse bone marrow were significantly elevated in female mice that had had their ovaries removed. In these mice, bone deposition was reduced and fat-making was highly active. The fact that BMP-4/7 signaling culminates in the removal of these methyl groups is an indication that KDM4B and KDM6B may represent novel therapeutic targets for metabolic bone disease.

Wang cautioned: “Through our recent discoveries on the lineage decisions of human bone marrow stem cells, we may be more effective in utilizing these stem cells for regenerative medicine for bone diseases such as osteoporosis, as well as for bone reconstruction. However, while we know certain genes must be turned on in order for the cells to become bone-forming cells, as opposed to fat cells, we have only a few clues as to how these genes are switched on.”

However, with respect to his work with older mice, Dr. Wang was more sanguine: “Interestingly, in our aged mice, as well as osteoporotic mice, we observed a higher amount of silencing histone methyl groups which were normally removed by the enzymes KDM4B and KDM6B in young and healthier mice. And since these enzymes can be easily modified chemically, they may become potential therapeutic targets in tissue regeneration and treatment for osteoporosis.”

Dr. No-Hee Park, the dean of the UCLA School of Dentistry said this about Wang’s findings: “The discovery that Dr. Wang and his team have made has considerable implications for craniofacial bone regeneration and treatment for osteoporosis. As a large portion of our population reaches an age where osteoporosis and gum disease could be major health problems, advancements in aging-treatment are very valuable.”

Update: Induced Pluripotent Stem Cells Made from Alzheimer’s Patients

Earlier this year, I reported that scientists had made induced pluripotent stem cells iPSCs from the skin cells of Alzheimer’s patients in order to differentiate them into neurons and study the effects of Alzheimer’s disease on the function of those neurons.  An update on this research is now available from talks presented at the Alzheimer’s Association International Conference.

Current mouse models of Alzheimer’s disease use animals in which particular genes known to play a role in the onset and pathology of Alzheimer’s disease are overexpressed in the brains of the animals.  For example 3xTg-AD mice overexpress a mutant version of the beta-amyloid precursor protein (beta-APP), a mutant version of the presenilin gene (PS1M146V), and a mutant version of the tau protein (tauP301L).  Al three of these alleles play important roles in Alzheimer’s disease (AD) etiology.  For example tauP301L is the most common mutation found in the tau protein-encoding gene associated with neurodegenerative diseases.  Mice that mice that overexpress the mutant human tauP301L have, in their brains, neurofibrillary tangles (NFTs), neuronal cell losses and memory disturbances (see see Wakasaya Y., et al., J. Neurosci 2011 89(4):576-84).  The PS1M146V mutation in humans is responsible for one of the more aggressive forms of early onset AD.  Finally, the double Swedish mutation of the beta-APP protein gene (Lys to Asn at residue 595 plus Met to Leu at position 596) increases production of amyloid protein, the material found in the plaques in the brains of AD, 6-8 times.  The 3XTg-AD mouse show much of the pathology found in the brains of AD patients, and have been useful in AD research.

However, according to Alzheimer’s Association scientist William Thies:  “Current animal models of Alzheimer’s are highly engineered to express elements of the disease, and, while valuable for research, incompletely represent how the disease form and progresses in people.  In order to develop better therapies and eventually prevent Alzheimer’s, we need better, more accurate animal and cellular models of the disease.  This newly reported research (i.e., the production of iPSCs from AD patients) is a significant step forward in that direction.”

Amyloid plaque formation is a hallmark of AD, and the present mouse models do not form amyloid plaques in their brains in the same way as the brains of human AD patients.  Furthermore, significant brain cell death does not occur in the present mouse models even though they do occur in human AD patients.

Amyloid plaques form as a result of the processing of beta-APP.  beta-APPis processed in one of two ways.  If an enzyme called alpha-secretase clips beta-APP, it forms a soluble N-terminal fragment (sAPPa) and a C-terminal fragment (CTFa).  The sAPPa protein seems to enhance synapse formation, neurite outgrowth and neuronal survival.  CTFa remains in the membrane and is cleaved by presenilin-containing gamma-secretase to produce a soluble N-terminal fragment (p3) and a membrane-bound C-terminal fragment (AICD, or APP intracellular domain).  AICD might be involved in nuclear signaling.  This mode of beta-APP processing does not produce amyloid protein for plaque formation.


If beta-APP is first cleaved by beta-secretase, it produces a soluble N-terminal fragment (sAPPb) and a membrane-bound C-terminal fragment (CTFb).  CTFb is longer than CTFa, and cleavage of CTFb by gamma-secretase produces AICD and a soluble fragment called amyloid-beta, or Abeta.  If Ab accumulates in the extracellular spaces of the brain, it can aggregate to form amyloid plaques.  Abeta is ‘stickier’ than other APP fragments.  It accumulates bit by bit, into microscopic plaques by means of a multi-step mechanism by which Ab peptides aggregate into oligomers that cluster together to form fibrils with a regular b-sheet structure.  The fibrils adhere to form mats, which clump together with other substances to eventually form plaques.  Ab plaques may kill off brain cells and trigger inflammation, which also kills cells.  Environmental conditions, age, genetic predisposition, and health in general influence of production of Abeta.

Andrew Sproul works as a postdoctoral research fellow at the New York Stem Cell Foundation in the laboratory of Scott Noggle.  Sproul used iPSCs to model AD and reported on his results at this conference.  Sproul used skin cells from AD patients and skin cells from unaffected relatives as controls and then made iPSCs from these skin cells.

“One advantage of this technology is that we get a near infinite supply of disease and control patient stem cells,” noted Sproul.  “Another is that we can then turn the iSPCs into any tissue in the body.  This allows us to investigate the role of various cells in Alzheimer’s disease progression by manipulating the iPSCs for form different types of brain cells (forebrain nerve cells, neural stem cells, glial cells) that we and others believe are involved in Alzheimer’s.”

Researchers made iPSCs from 12 young people with early-onset Alzheimer’s and healthy controls from families that harbored the genetic predisposition for young-onset AD.  The iPSC lines have been tested to ensure that they are pluripotent

“We have made both the control and Alzheimer’s iPSCs into brain cells and have demonstrated that they are electrically active.  These brain cells include forebrain cholinergic neurons (neurons that re;ease acetylcholine as a neurotransmitter)  which are particularly vulnerable in Alzheimer’s disease.”

“We have also begun to use the iPSC-derived neurons and neural stem cells to compare differences in cellular function between people with Alzheimer’s and their unaffected relatives.  For example, we, in conjunction with Dr. Sam Gandy’s group at Mount Sinai School of Medicine, have demonstrated that Alzheimer’s neurons produce more of the toxic form of beta amyloid, the protein fragment that makes up amyloid plaques, though this aspect of the research is preliminary,” Sproul said.

Much of the research in this group is concerned with Presenilin-1.  Mutations in PSM1 are responsible for the most common form of rare, inherited young-onset AD (less than 2% of all cases).  The iPSC platform might provide the best system to drug testing those new compounds that could mitigate the effects of this devastating disease.

Because the majority of AD patients have the sporadic form of AD, scientists plan to expand their research to include large-scale production of iPSCs from people with different forms of AD.

Sproul added, “We have begun to extend this work by collaborating with four different institutions in New York City; the Mount Sinai School of Medicine, Columbia University, New York University, and Rockefeller University.  Over the next few years, we expect to provide substantial insight into Alzheimer’s and valuable tools to help create the next generation of therapeutics.”

ObamaCare and the Coming Two-Tiered Medical System

John C. Goodman has a fabulous article in the Wall Street Journal about the coming hell that awaits the US when ObamaCare kicks in completely.

Most of the provisions of ObamaCare come online on Jan. 1, 2014. Within a decade of this date, approximately 30 million people more people are expected to acquire health plans. Now research has indicate if people get health insurance and if they are under the mistaken illusion that it is free, they begin to go to the doctor for hang nails and other ridiculous things. Therefore, economic studies show that these 30 million extra people will essentially double their use of the health-care system.

The Obama administration is constantly reminding senior citizens that they are entitled to a free annual checkup. Also, in its new woman-centered campaign, women are being told, they will have access to free breast and pelvic exams and even free contraceptives. Yes, once Shangri-La arrives when ObamaCare fully takes effect, we will all be entitled to a long list of preventive services, with no deductible or copayment. If it sounds too good to be true, that’s because it is.

The problem? Where to start. First, our health-care system simply cannot meet this huge increase in demand for primary-care services. While the original ObamaCare bill contained a line item for increased doctor training, this provision was zeroed out before passage to keep down the cost of health reform. This the entire primary care services entitlement will result in nothing less than gridlock of the worse kind.

Since, according to ObamaCare, health insurance must cover the tests and procedures recommended by the U.S. Preventive Services Task Force, doctors will be forced to offer these services. Here’s the catch: According to a study in the American Journal of Public Health (2003), scholars at Duke University calculated that arranging for and counseling patients about all those screenings would require 1,773 hours of the average primary-care physician’s time each year, or 7.4 hours per working day. Given the massive amounts of paperwork ObamaCare is going to generate for physicians, there is no way on this green earth that any doctor, nurse or physician’s assistant in going to be able to perform these services for patients.

Health care professionals will spend time searching for problems and talking about the search, and if screenings turn up a real problem, there will have to be more testing and more counseling. Here’s the bottom line: in order to meet the promise of free preventive care nationwide, every family doctor in America would have to work full-time delivering it, leaving no time for all the other things they need to do.

Now if we apply some very basic economics to this situation, if demand exceeds supply in a normal market, prices rises until they price many people out of the market. However, in this case, as in other developed nations, Americans do not primarily pay for care with their own money for health care. Therefore they pay with something else even more precious and that is time.

According to a 2009 survey by medical consultancy Merritt Hawkins, the average wait to see a new family doctor in this country is just under three weeks. However, in Boston, Massachusetts, which enacted near-universal coverage under Gov Mitt Romney, the wait is about two months.

How long it takes you to see a doctor is a non-price barrier to medical care, and there is substantial evidence that it is even more important in deterring care than the fee the doctor charges, even for low-income patients.

If people cannot find a primary-care physician who will see them in a reasonable length of time, all too often they go to hospital emergency rooms. However, a 2007 study of California in the Annals of Emergency Medicine showed that up to 20% of the patients who entered an emergency room left without ever seeing a doctor, because they got tired of waiting. Under ObamaCare, this situation will certainly worsen dreadfully.

Economics again: if demand exceeds supply, doctors much more flexibility to see whomever, whenever. Therefore, they tend to see those patients first who pay the highest fees. For example, a 2008 New York Times survey of dermatologists uncovered an extensive two-tiered system. Patients in need of services covered by Medicare waited 2-3 weeks to see a doctor, and the appointments were made by answering machine. For Botox and other treatments not covered by Medicare for which patients pay the market price out-of-pocket, appointments to see those same doctors were often available on the same day, and made by live receptionists.

Physicians will increasingly need to jealously protect their time in order to make a proper living. Therefore, patients in plans that pay below-market prices, which include the elderly, disabled on Medicare, low-income families on Medicaid and people with subsidized insurance acquired through the ObamaCare exchanges, will wait the longest. However, the wait time will only get longer and longer as more and more Americans turn to “concierge medicine” for their care.

Concierge medicine differs from region to region and doctor to doctor, but it generally refers to patients who pay doctors to be their agents, rather than the agents of third-party-payers (e.g. insurance companies or government bureaucracies).

As an example, a Medicare patient can pay $1,500 to $2,000 to form a new relationship with a doctor, and this relationship includes same day or next-day appointments. It also usually means that patients can talk with their physicians by telephone and email. The physician helps the patient obtain tests, make appointments with specialists and in other ways negotiate an increasingly bureaucratic health-care system.

This spells trouble for ObamaCare, since a typical primary-care physician has about 2,500 patients, according to a 2009 study by the Centers for Disease Control and Prevention. When that same physician opens a concierge practice, he’ll typically take about 500 patients with him, according to the MDVIP, which is the largest organization of concierge doctors. That’s about all the doctor can handle, given the extra time and attention those patients are going to expect. What about the 2,000 patients left behind? They must find another physician. Therefore, as concierge care grows, the strain on the rest of the system increases.

It is not difficult to predict what happens next. As concierge medicine rapidly grows, every senior and non-senior who can afford one will have a concierge doctor. The rest who cannot afford the cost will not. This we will quickly switch to a two-tiered health-care system, in which the rich get fast, high-quality care, and the poor wait forever for low-quality care.

This will leave us with a vulnerable population that will have less access to care than they had before ObamaCare became law. All because the president and the dunderheads in Congress do not understand that incentives matter or even the first rule of economics.

Mesenchymal Stem Cell Treatment Prevent Post-Fracture Arthritis in Mice

Researchers at Duke University Medical Center have found a promising stem cell therapy that might prevent osteoarthritis after joint injury.

Joint injuries tend to raise the odds of contracting a type of arthritis known as post-traumatic arthritis (PTA). There are no available therapies that delay the onset or progression of arthritis after joint-related injuries.

However, a research team at the Duke University Health System has discovered a promising therapeutic approach for PTA that utilizes a particular type of stem cell known as a mesenchymal stem cell (MSC). MSCs have the ability to quell PTA, since these particular stem cells have the ability to suppress inflammation. Additionally, MSCs have the ability to readily differentiate into cartilage-making, bone-making, fat-making, and smooth muscle-making cells, which makes them prim candidates to regenerate damaged joints, since these stem cells have beneficial properties in other regions of the body.

This therapeutic approach was investigated in mice that had suffered the type of bone fractures (intraarticular fractures) that would usually lead to PTA. Injections of 10,000 MSCs into the joints of mice that had suffered intraarticular fractures decreased inflammation and greatly increase bone deposition during healing. Furthermore, the mice that had MSC injections into the joints did not develop PTA while those that received injections of saline into their joints largely did develop PTA. This study could potentially help produce a therapy that would be used after joint injury and before the patient suffers the initial signs of significant osteoarthritis.

Farshid Guilak, Ph.D., director of orthopedic research at Duke and senior author of the study, summarized the results of this study: “The stem cells were able to prevent post-traumatic arthritis.” The study was published on August 10 in the journal Cell Transplantation.

Another track investigated by Guilak’s lab was so-called “superhealer” strains of laboratory mice that tend to heal from bone fractures much faster than other strains. Guilak and his colleagues examined if MSCs from the MRL/MpJ (MRL) “superhealer” mouse strain would increase bone healing when compared to MSCs from C57BL/6 (B6) mice. This was to determine if the exceptional regenerative abilities of MRL mice was due to their MSCs. Unfortunately, Guilak and colleagues discovered that B6 MSCs did the job just as well, and, in fact, a little better than MSCs from MRL mice. Thus, even though they thought that superhealer mice bred for their super-healing properties would probably fare better than typical mice, in turned out that they were wrong.

“We decided to investigate two therapies for the study, said lead author Brian Diekman, Ph.D., who works as a postdoctoral research fellow in the Guilak lab. “We thought that stem cells from so-called superhealer mice would be superior at providing protection, and instead, we found that they were no better than stem cells from typical mice. We thought that maybe it would take stem cells from superhealers to gain an effect as strong as preventing arthritis after a fracture, but we were surprised – and excited – to learn that regular stem cells work just as well.”

Interestingly, certain people appear to fall into the superhealer category. They bounce back quickly and heal well naturally after a fracture, while other people eventually form cases of arthritis at the fractured joint, said Guilak, who is a professor of orthopedic surgery and biomedical engineering at Duke University.

“The ability of the superhealer mice to have superior healing after a fracture may go beyond the properties of their stem cells and be some beneficial factor, like a growth factor, that we don’t know about yet,” Guilak said.

Diekman said the team looked at markers of inflammation and saw that the stem cells affected the inflammatory environment of the joint after fracture.

“The stem cells changed the levels of certain immune factors, called cytokines, and altered the bone healing response,” said Diekman, who is also with the Duke Department of Biomedical Engineering.

According to Guilak, very few studies have purified stem cells to the degree they were purified for this study. The MSCs used in this study were from bone marrow, and they are not directly involved in the production of blood cells, even though they do play an important support role for blood cell-making stem cells in bone marrow (see C. Shi. Immunology 2012 136(2): 133-8 & also see here).

Diekman said that one of the challenges in the field is devising a precise protocol for identifying, isolating and purifying MSCs from bone marrow, since these stem cells tend to be rather rare in bone marrow.
“We found that by placing the stem cells into low-oxygen conditions, they would grow more rapidly in culture so that we could deliver enough of them to make a difference therapeutically,” Diekman said.