Adult stem cells in MS for reversal

In Multiple Sclerosis (MS), the immune system of a patient turns on his or her nervous system, destroying the myelin sheaths that serve as insulation around nerves, disrupting the transmission of nerve signals. The myelin damage often occurs in a patchy manner, at first. See the Medline Plus page from the National Institutes for Health for more information, including a patient tutorial in English and in Spanish. The Journal of the American Medical Association has a similar patient education page in pdf.

There have been trials using adult stem cell treatments in the form of bone marrow transplants and injection of adult stem cells in different manners for several years. (See ClinicalTrial.gov for some of these - if my search lapses, search "stem cell multiple sclerosis.)

The March issue of Lancet Neurobiology reports (Free access to the abstract) success from research at Northwestern University using the patients' own bone marrow stem cells, after harvesting those cells, preserving them, and then using chemotherapy to destroy the immune system before replacing the patient's stem cells. A review of the study is at the Science Daily and at this blog, Science Codex.

Here's the information on this trial from Clinical Trials.gov.

Any sort of bone marrow transplant is dangerous due to the lost red blood cells, platelets (to make blood clots), and the white blood cells that function as the immune system. This trial was set up to preserve all but the immune system. While it's still not a procedure to take lightly, it appears that the researchers at Northwestern have made it safer.

"Tea-bag" Adult Stem Cell Treatment for Stroke

British researchers report an amazing recovery for a 49 year old man who suffered a hemorrhagic stroke on October 15, 2008. The researchers at the company, "Biocompatibles," used adult stem cells from a healthy donor. The cells had been engineered to cause them to produce a protein that helps prevent "programmed" cell death (even after the bleeding stops and the pressure is removed) and embedded in tiny beads that had been sewn up in a cloth "tea-bag."

From the press release, published on the Medical News Today Neurology and Neuroscience website:

Stroke is one of the leading causes of death in the elderly population in the developed world. The incidence rate has been reported as 145 per 100,000. Hemorrhagic stroke is responsible for ~15 to 20% of all stroke and it is the least treatable form of stroke. It is associated with the highest morbidity and mortality rate of all stroke with only 44% of affected patients surviving the first 30 days. Only 20% of these survivors regain functional independence. The cascade of events starts with the sudden rupture of a blood vessel in the brain, causing haemorrhage and pressure inside the skull. Surgery may be used to relieve the pressure; but the haemorrhage causes a longer-term process of programmed cell death, or apoptosis, and it is this that causes the lasting neurological damage.

The CellBeads™ are delivered directly to the injury site during the surgery. They are programmed to deliver CM1, a proprietary version of a naturally occurring protein, GLP-1, which has been shown to have powerful anti-apoptotic effects. The delivery mechanism is a cluster of human adult mesenchymal stem cells obtained from a healthy donor and encapsulated in alginate beads. The cells are genetically engineered to produce the protein, which is delivered continuously, directly to the injury site. The alginate beads protect the stem cells from the body's immune system, which would otherwise destroy the foreign cells. CellBeads™ are transplanted within a retrievable mesh device and are removed completely after a treatment period of 14 days. Retrieval of the implant prevents possible long-term side effects from the transplanted cells.

The research is a "Phase I/II" trial, which means that the doctors and scientists are actually testing the safety of the treatment, and not the actual effectiveness of the treatment, itself. In other words, "does the treatment do more harm than good."

The CEO of Biocompatibles, Crispin Simon (that name is as British as tea bags), spoke to a Reuters reporter for a story published at Forbes online, stressing that the patient is young and other wise healthy, and had the standard of care for hemorrhagic strokes, surgery to relieve the pressure from the blood on the cells around the stroke. 10% to 20% of patients have similar recovery, without the Biocompatible beads.

Still, the report is a welcome source of hope for anyone who has watched and waited helplessly after a patient or a loved one had a hemorrhagic stroke.

No Patent on Human Embryonic Stem Cells: EU

A Thanksgiving present from the European Union!


The Enlarged Board of Appeal of the European Patent Office (EPO) did not take the day off. Early on November 27, the EPO announced that they would not allow the development of human embryonic stem cells to be patented as filed by WARF in 1996, since that technique depended on the destruction of human embryos.

For background on WARF, see this post on the patents, here.

The EPO has upheld its previous ruling from earlier this year:

The EPC does not allow patenting inventions whose commercial exploitation would be contrary to public order ("ordre public") or morality. Furthermore, the Convention prohibits patenting on uses of human embryos for industrial or commercial purposes.

Needless to say, Geron is not happy.

The (manufactured) Stem Cell Debate at Dartmouth

I don't believe I've ever seen a report on a presentation that allowed half the space for "debate," after the fact.

The Stem Cell Debate at Dartmouth

Sunday, November 16, 2008

Father Tadeusz Pacholczyk, Ph.D. was recently invited to give a lecture entitled “Stem Cells and Cloning: Understanding the Scientific Issues and the Moral Objections” at Aquinas House, in observance of the Feast of St. Luke, the patron saint of medical professionals. Pacholczyk, or Father Tad as he encourages his audience members to call him, is the Director of Education for the National Catholic Bioethics Center. He arrived at this position after receiving degrees in philosophy, biochemistry, molecular cell biology, and chemistry, a Ph.D. in Neuroscience from Yale University, and years of research in molecular biology, bioethics, and dogmatic theology. In a free public lecture lasting more than two hours, Pacholczyk outlined both the scientific and ethical considerations of human embryonic stem cell research and to a lesser extent cloning, giving justifications for the Catholic Church’s positions on these technologies.

After giving an in depth layman’s version of the science involved in stem cell research and a history of both scientific milestones and relevant policy decisions, Pacholczyk corrected what he believed were some of the most pervasive myths about stem cell research. He believes that individuals and organizations within the media and others who engage in expensive advertising campaigns have deliberately misled the American people in an effort to reframe the debate over the use of human embryos for research.

**************
The Dartmouth Review understands that this is an issue on which reasonable moral people can disagree, and so Michael S. Gazzaniga ‘61, Ph.D., Director of the Sage Center for the Study of Mind at the University of California, Santa Barbara was asked to explain some of the ethical justifications. He indicated that, “The handling of human tissue has always commanded the respect of the biomedical community and always will.” However, Gazzaniga does not consider an embryo to be in possession of the same moral status as an adult human, while acknowledging that the issue has “deep meaning to millions of people.”

The Review has raised several ethics questions regarding the virtual debate they created by interviewing Dr. Gazzaniga after Dr. Pacholczyk's talk.

Will they seek out opposing views in the future or is it only Catholic priests who require such answers? Will they now give Dr. Pacholczyk an opportunity to respond?

In addition, Dr. Gazzaniga finds the determination as to when a human being becomes a human being fairly simplistic:

Asked the basic question underlying this debate and that about abortion, when a human embryo becomes a human being, Gazzaniga called it a “social decision, not unlike the kind a society makes about when to call someone legally blind.”

Does Dr. Gazzaniga's emphasis on contrasting "adult" human beings with embryonic human beings indicate that he finds differing moral values in the lives of infants, children, and "adults," does he extend these differences to the state of function of the brain, and can he justify these variations at least as well as we can our culture's definition of "legally blind?"

Induced Pluripotent stem cells without viruses

"The adenovirus will infect the cells but then will clear themselves from the cells. After a few cell divisions there are no traces of the virus in the cell," Hochedlinger said. "You can't tell the virus was ever there."

Science Magazine has published a report on induced Pluripotent (iPS) stem cells from liver cells (hepatacytes) that do not show any trace of the viruses initially used to cause the regression from adult cells to embryonic-like stem cells. The report, by Hochedlinger's group at Massachusetts Gener al Hospital and Harvard, is behind a pay-wall, but there is a review here at the Washington Post.

The simple explanation is that viruses are used to carry copies of genes that turn on proteins which cause the cells to divide and produce embryonic-like stem cells. Prior research used viruses that might become permanently inserted into the DNA of the cells. These viruses did not cause an infection in the culture because the viruses used were not good at causing themselves to be reproduced or inserting themselves into neighboring cells. However, because they inserted themselves into the DNA, there was a risk that the cells could become mutated or even cause tumors due to the abnormal DNA that resulted. In the case of Hochedlinger's cells, the adenovirus used does not insert into the DNA nearly as often, and when it does, the cell is able to repair the DNA in subsequent copies of the DNA as it reproduces the nucleus of the cell in order to divide to become two cells.

The cells are infected, they change because of the infection, but their granddaughters are able to get rid of the infection, while continuing to act like embryonic stem cells instead of grown up liver cells. The new "adeno-iPS" cells pass all the tests for "stemness."

Just think, no need for egg cells, no cloning, no destruction of embryos, and we're one step closer to healing within the body - to learning how to heal without transplant rejection or tumors due to the treatment. One day, we may be able to regenerate organs and tissues in place, as needed.

"iPS cells would have never been discovered without human embryonic stem cells"

In fact, Mr. Siegal, without the objection to cloning and embryonic stem cell research, iPS cells researchers might have taken a little longer to develop therapy that holds promise for regeneration and healing in the body, without transplants, intermediate cells and without costing the life of another human being.

Bernie Siegal, that lawyer who sued the Raelians for custody of their supposed cloned children and who lauded Hu Wu Suk for his own cloned children, is hosting one of his "World Stem Cell Summits" in Madison, Wisconsin next week. He is joined by Robin Alta Charo in promoting the Summit and reminding all of us that the push is not over for more clone-and-kill embryos and federal tax funded research depending on embryo creation and destruction. No matter how successful induced Pleuripotent Stem (iPS) cells are turning out to be.

Siegal founded the Genetics Policy Institute, with the help of the National Heritage Foundation Inc:

The GPI is funded by individuals, foundations, academic institutions and scientific societies, including the American Society For Cell Biology, the American Society for Biochemistry and Molecular Biology, the Michael J. Fox Foundation for Parkinson's Research and the Huffington Foundation, according to Siegel.

R. Alta Charo (for some reason, she drops the "Robin") is a lawyer/bioethicist ( "for Hire")will deliver a keynote address at the Summit. She lobbies against physicians with consciences and in favor of abortion, cloning and embryo stem cell research. She's especially fond of the latter two, because (as she says in this audio copy of her lecture at the American Society of Bioethics and Humanities convention in 2006)she believes they will support her Progressive politics and belief that the research will support her personal belief that there is no Creator, so humans aren't so special, after all.

For a brief review of the history of "ethics for sale," look at this set of my posts.)

Human-pig embryo approved in UK

The "cybrid" or hybrid human-animal embryos are created in the laboratory by Somatic Cell Nuclear Transplantation, using emptied eggs from animals and the nuclear and cellular DNA from humans.. We know that there are currently experiments on-going with the human embryos made using emptied cow eggs (more on the "ease" of making these embryos, here), and now the British have authorized the development of pig-human embryos.

The experimenters admit that the problem will be achieving embryos and embryonic stem cells that do not contain DNA left from the egg. Proving the purity and "human-ness" of the stem cells will be a complication that I do not believe they will be able to overcome, at least for transplantation into humans, except possibly in the case of severe, last-hope disease and trauma.

The ethical debates about xeno-transplants and treatments using living organs, cells and tissues from animals carry the risks of transmitting animal diseases that humans have no immunity for and the development of new strains of disease that cross species lines. Ethicists have predicted that at least the early patients will have to live their lives in isolation at the worst, and have life-long surveillance at the best. (more on the debate, here and here.)

However, the researchers will probably be able to develop other uses, such as the early warning chemical weapon detection systems that are being developed by our own military, using human embryonic stem cells.

Rather than humanitarian and medical hope, I believe that time will show us that the research is the result of pure greed, with each lab hoping to come up with a product that can be patented and sold. I'm disappointed that the courts and "ethics" bodies in the US and UK have allowed these patents of human organisms. The drive to "create" new human cells and artifacts using human DNA is the logical outcome.

Human-DNA-in-cow-egg embryo created in UK

Scientists in the UK report that they have created an embryo using the transfer of human nuclear DNA from an embryonic human cell into the oocyte of a cow that has had the nucleus removed. These embryos are the "hybrids" or "cybrids" that we've been discussing for the last few years.

From the Guardian:

Apparently these researchers have achieved some success - but by using the nucleus from a very early embryonic cell, which might be easier to reprogramme than an adult cell. At the moment it is impossible to assess the significance of this report until we know more details of what has been achieved ... the results have been repeated and, importantly, they have been reviewed by independent researchers in the usual way."

Josephine Quintavalle, of the pressure group Comment on Reproductive Ethics, said the research should not worry those opposed to hybrid embryos because the Newcastle work did not seem convincing. "The embryos didn't survive, they were created from embryonic stem cells rather than adult tissue, and there's a lot of question marks over the research."

But she added: "What it has done is wake up the public to this reality, that while parliament is getting in a tizz about this, while the whole country is up in arms discussing it, the HFEA is already issuing licences."

Supposedly, if the technique is perfected to allow the embryos to survive longer, these embryos will allow the study of the early embryo and production of embryonic stem cells in order to learn more about and find cures for diseases like diabetes and Parkinson's.

However, even if the embryos are disorganized and fail early, or if they are destroyed at day 5 or 6 or whenever, the ethical determination as to whether they are "human" or "bovine" has not been cleared up. We won't know what they are until several labs and several trials successfully create these embryos.

If the embryos appear to divide in an organized manner, producing human proteins and the differentiation necessary to create human embryonic stem cells, then they are essentially human embryos. This is a case of the old if it walks like a duck, quacks like a duck, etc., logic.

Since the stated intention is to destroy the embryo, and we don't know whether they are human or not, those of us who find the killing of humans, even at the earliest stages will also hold that it is inherently unethical to even begin the process.

A discussion about the discussions about the announcement can be read at one of Nature.com's blogs, "The Great Beyond."

From the thread, "UK hybrid embryo: in perspective - April 02, 2008,"

New Scientist has attacked the group for announcing the achievement through the media rather than through a scientific publication. The Independent focuses on the ethical debate. Not many organisations outside the UK gave it any coverage at all, and those that did may have been under the impression that it was a world first, not mentioning previous achievements in the field (eg. Life Scientist, Australia).

Nature Reviews Stem Cell Heart Treatments

The journal, Nature, has published a review article, "Stem-cell therapy for cardiac disease,"
about treatment of heart disease with stem cells, focusing on the many types of cells that are being used in research, including bone marrow derived stem cells and progenitors and "resident" cardiomyocyte stem cells. The latter are actually found in the heart and can be harvested from the patient who needs them and used to repair damaged heart disease.

The abstract promises more than I ever thought I'd read in a "First tier" journal.

Heart failure is the leading cause of death worldwide, and current therapies only delay progression of the disease. Laboratory experiments and recent clinical trials suggest that cell-based therapies can improve cardiac function, and the implications of this for cardiac regeneration are causing great excitement. Bone-marrow-derived progenitor cells and other progenitor cells can differentiate into vascular cell types, restoring blood flow. More recently, resident cardiac stem cells have been shown to differentiate into multiple cell types present in the heart, including cardiac muscle cells, indicating that the heart is not terminally differentiated. These new findings have stimulated optimism that the progression of heart failure can be prevented or even reversed with cell-based therapy.

New York Times article on cloned humans

The New York Times (free one time registration required) has a news piece on the Stemagen cloned human embryos, with reference to "making copies of people" and implantation of cloned embryos for reproduction.

One of the men who donated the fibroblast skin cells is also the owner of Stemagen.

The NYT has more on the story behind the cloned human embryos:

The Stemagen scientists, led by Andrew French, an animal cloner recruited from Australia, used skin cells from Dr. Wood and another Stemagen employee as the DNA source. They used 29 eggs donated by young women at the fertility clinic that Dr. Wood manages.

Five blastocysts were developed. One was shown to be a clone by genetic testing, the scientists reported, and two others also showed good evidence of being clones.

Dr. Wood said the key to success might have been choosing egg donors who were known to be fertile and healthy because they had previously been successful donors at his fertility clinic.

The women were also donating at the same time to couples wanting babies. Some eggs went to the couples and the others to the research, with the consent of both the donors and the couples. The donors were paid for the eggs that went to the in vitro fertilization but not to the research, Dr. Wood said.

Therapeutic cloning has been hampered by lack of access to healthy eggs, in part because it is often considered unethical to pay women for such donations. Dr. Daley of Harvard said Stemagen’s “egg sharing” approach appeared to be a reasonable way to obtain eggs.

The media will have fun with this story.

Human embryos cloned in California

Scientists at Stemgen, a La Jolla, California laboratory have published a report on the successful cloning of human embryos in the journal, Stem Cells. (The article is available free, due to the open access policy of the journal.)

The authors are very clear: these are human embryos produced by somatic cell nuclear transfer or cloning. The embryos were clones of the men who donated the fibroblast skin cells.

This study demonstrates, for the first time, that SCNT can be utilized to generate cloned human blastocysts using differentiated adult donor
nuclei remodeled and reprogrammed by human oocytes. Evidence of successful SCNT was shown with DNA fingerprinting analyses of three SCNT cloned blastocysts where embryo genomic DNA was that of the donor fibroblast cell line and were not fragmented oocytes or of parthenogenetic origin.

. . .DNA fingerprints from three SCNT blastocysts were consistent with those of the somatic cell donor employed with no evidence of contamination from the egg donors, indicating that embryonic development was being controlled by the donor cell genome.

The cloned human embryos were produced using donated oocytes less than 2 hours old and the DNA from the skin cells of men. (The eggs were donated by women for the use of other couples, see below.) The use of male donor DNA allows for easier distinction from any possible parthenogenetically produced embryos, which would be female. Any embryos that are male serve to prove the success of the experiment.

In this case, the cloned embryos were actually compared to parthenogenetically produced embryos created by stimulating oocytes to become embryos. These embryos only contain the DNA of the women who donated the eggs. Parthenotes are not clones, because of the rearrangement of genes that happens when the eggs are produced with half of the normal chromosomes which would be matched by the haploid sperm if fertilization took place.

It appears that the group had a very high success rate, with approximately 2/3 or 16 of 25 of the enucleated oocytes producing very early embryonic organisms, which (who) demonstrated cell development and division similar to embryos produced by in vitro fertilization. 10 of the embryos developed to day 3 and 5 of those went to day 5, with the formation of blastocysts. Blastocysts are embryos that have developed enough cells to form a layer of cells around a hollow center, and eventually the inner cell mass, the differentiated grouping of embryonic stem cells at one spot within the sphere. All 5 of the blastocysts formed inner cell masses. The authors do not report any stem cell lines from these embryonic stem cells, but note that they are trying to do so - either from these embryos or from additional cloning.

The Discussion includes speculation that the success rate was so high because the oocyte donors were young women who were able to produce so many eggs through stimulation of their ovaries that there were more than enough for the use by the parents (couples?) to whom they were donating for the production of embryos for implantation and pregnancy. Although the article states that all 3 of the parents were able to get pregnant from the eggs that went to them, that could not have been known at the time the eggs were taken to the experimental lab. Some went to the in vitro lab and some went to the experimenters within less than 2 hours. It takes at least a few hours after in vitro fertilization to determine whether any embryos were formed.

If embryonic stem cell lines are developed from this technique, perhaps some group will compare them to embryonic-like stem cells developed by reprogramming.

Myths on Myths about stem cells

There's a new Public Broadcasting System (your tax dollars at work) television show on "stem cells," "Mapping Stem Cell Research: Terra Incognita."

You don't have to go any farther than the top of the home page, with its picture of a girl in a wheelchair and this quote,

"Some people consider stem cell biology to be the Holy Grail of Regenerative Medicine, while others view embryonic stem cell use as morally wrong."

to see that it's propaganda for embryonic stem cell research and cloning for embryonic stem cells. The authors immediately begin the pattern of using the term "stem cells" for both of the two basic kinds of stem cells: those that require the destruction of a human life and those that don't.

Here are the first three points from the "Myths and Realities" page, with my comments in Bold after each.

MYTH
Stem cell research uses aborted fetuses.
REALITY
Stem cells can be totipotent (a fertilized egg with the “total potential” to give rise to all different types of cells in the body), multipotent (stem cells that can give rise to a small number of different cell types), or pluripotent (stem cells that can give rise to any type of cells in the body except those that are needed to develop a fetus). While pluripotent stem cells could be developed from fetal tissue or even adults, they are best derived from early-stage embryos, a mass of cells that is only a few days old—not aborted fetuses.

The authors skip over the significance of the fact that embryronic stem cells come from destroyed human embryos in the lab, it is true that most stem cell research does not use tissues obtained from abortions. Nowadays, however, the term "fetus" is too often used by the media (and even researchers who ought to know better) for all pre-born human beings. The proper definition of human embryo is the organism from fertilization or the beginning of the first cell division to 7-8 weeks of age. The term "fetus" in humans is properly used from 8 weeks until birth.

More on the claims about what is the "best" source of stem cells and about "embryonic-like stem cells," below.

MYTH
Somatic cell nuclear transfer using human cells involves the use of fertilized eggs.
REALITY
Somatic cell nuclear transfer, the process in which the nucleus from an adult cell is removed and then transferred to an egg whose nucleus has been removed, is the first step in cloning and can be used to create an embryonic stem cell line. However, an egg cell does not need to be fertilized to be used in this procedure—an unfertilized egg cell can be used.

Here, the authors avoid using "embryo" and throw around the terms "unfertilized egg" and "fertilized egg." An embryo is not a "fertilized egg" - once an egg is fertilized, it becomes an embryo. In Somatic Cell Nuclear Transfer (cloning), the embryo is produced artificially by inserting the DNA of a donor cell and stimulating division and organized development that occurs with natural reproduction. When human DNA is used to produce human embryonic cells in an organized embryo, there can be no doubt that what we are talking about is a human embryo. No matter how he or she is created - or produced - or how severely handicapped by the intentions and actions of the producers, a human embryo is a very young human being.

MYTH
Researchers can use adult stem cells instead of embryonic stem cells. Other treatments using adult stem cells are available to treat conditions such as Parkinson's disease and spinal cord injuries.
REALITY
Adult stem cells lack the versatility and flexibility of embryonic stem cells, making them less likely to lead to medical breakthroughs. Embryonic stem cells have a far greater developmental potential and are more likely to be pluripotent, while adult stem cells are thought to be merely multipotent, or restricted to only certain cell types.

In November 2007, Japanese and American research teams reported new ways to obtain stem cells that behaved like embryonic stem cells from human skin cells—without having to use human embryos. This breakthrough holds great promise in solving the ethical dilemmas of stem cell research, but scientists currently still face technical hurdles and the challenge of finding ways to use these stem cells successfully in medical treatments and therapies.

The biggest lie of all is that embryonic stem cells are more useful in treatments for human beings. Just ask the 20,000 plus in the US alone who have been treated with adult and umbilical cord stem cells or go looking for even one human who has been treated with embryonic stem cells.

While it is true that most ethical, adult stem cells are not "pluripotent," there are many kinds of "multipotent" stem cells and precursor cells in the body. In fact, these are the cells that we probably will use in the future, because they are the cells the body uses to repair itself and because they are less likely to grow out of control or cause tumors.

We are also learning that the desired development of stem cells and precursor cells is influenced by the environment and all sorts of "factors," or chemical and physical signals present in the part of the body where they grow into cells, tissues and organs. The key to future treatment for most disease will probably come from learning to stimulate these conditions and factors.

Besides the ethical dilemma of destroying early human life, embryonic stem cell research has every problem or hurdle that could be cited for adult stem cells: they are difficult to grow, found in small numbers, the cultures may be contaminated with different, undesirable cell lines, and are difficult to control to produce for the exact stem cell line that is needed.

Moreover, no one wants to transplant embryonic stem cells into people. What we want is to produce adult stem cells for treatments.

The last paragraph mentions embryonic-like stem cells. There are several ways to produce stem cells that behave in every way that the unethical stem cells do.

These cells are being used in research to replace the unethical cells produced by destruction of embryos.

The goal of all stem cell research is to have a source of "patient-specific" stem cells from the patient or to find ways to stimulate stem cell production in the body of the patient, when and where they are needed.

The producers of this program are advocating for outdated research methods.While researchers have learned a lot from human embryo research in the past, most of what we use has been developed from research in animal models. The production of new embryonic stem cell lines from human embryos and from cloning is no longer necessary to carry out this research.


(Thanks to Janet, of the Bedford County Citizens Concerned for Human Life, for sending me the link to the website on the show.)

Alabama Citizens for Life Link to LifeEthics

There's a link to LifeEthics.org at the website of Alabama Citizens for Life. I'm flattered that their "Stem Cell Primer" quotes this blog on the division of research into destructive and non-destructive.

The business of adult stem cells

Since Mr. Reed has urged us to follow the money trail, and after my talk with the banker last week, I was pleased to see that others are looking at ethical stem cells as an investment.

The business journal, Bloomberg, reported on several companies that are involved in ethical stem cell research. Evidently, these companies received a boost in their ratings after the news of amniotic stem cells broke over the weekend.

(HT to Rush Limbaugh)