November 15, 2002
Therapeutic Cloning Provides Younger Stem Cells

There is an important report out in New Scientist about therapeutic cloning and the relative vigor of different stem cell types. But it is difficult to puzzle out exactly what this report is saying. If anyone with relevant knowledge can go read the full article in New Scientist I'd appreciate your feedback in the comments for this post: (or email me)

Now New Scientist has uncovered a patent application that claims cloned stem cells have a big advantage over other stem cells. A team at Advanced Cell Technology (ACT) in Massachusetts, working with Malcolm Moore of the Memorial Sloan-Kettering Cancer Center in New York, cloned skin cells from two cows. They then injected blood-forming stem cells (which also give rise to immune cells) from the cloned fetuses back into the cows. One cow had its immune systems suppressed with drugs.

The cloned cells seemed to have an amazing ability to take over from adult ones, replacing up to 50 per cent of the cows' blood stem cells after just one infusion, even in the cow whose immune system was untouched.

UPDATE: Upon reflection, my interpretation: First, they took skin cells from the cows. Then they cloned them by putting their nucleuses into unfertilized eggs (from the same or different cows?). Then they grew cow fetuses to the 100 day point. Then they harvested blood stem cells from the fetuses. Then they injected those blood stem cells back into the adult cows that had been cloned. These cloned stem cells outcompeted the existing blood stem cells. Okay, then how did they know that the cloned cells outcompeted the existing stem cells that were already in the adults? Did they inject each cow's clone-derived stem cells into the other cow in the pair? They would need some genetic marker that would let them tell the cloned cells apart from the native cells of the cow. Possibly they used mitochondrial DNA for that purpose by using a different cow as the egg donor for the cloning.

What This Result Says About The Prospects For Adult Stem Cells

While this article doesn't convey the details of these experiments with sufficient clarity the article does seem to be claiming that the cloned stem cells were in some sense younger than the more adult stem cells. If this is what the ACT researchers found then this is very important. Many scientists are trying to coax adult stem cells into becoming various differentiated cell types and some successes are even being reported. However, the cells in the adult stem cell reservoirs in the body age along the rest of the body. This latest result underscores this point. As scientists find ways to coax adult stem cells into becoming various assorted differentiated cell types will they find that the adult stem cells will be too old and tired to become enough of each needed cell type to be useful? Or will they find that the differentiated cells so produced are too old to function properly?

Old People Most Need Replacement Cells. The Old people are the group who have the most amount of illness and who are most in need of having their stem cells coaxed into making various kinds of differentiated replacement cells. Yet the adult stem cells of old people are also going to be old and therefore less vigorous than those of younger adults.

Youthful Adult Stem Cells Will Help Make Us Young Again

Youthful adult stem cells are not just useful for treating illnesses. If the adult stem cell reservoirs could be replenished with youthful cells then we could become at least partially youthful again. Just as embryonic stem cells (whether created by classical fertilization or by cloning) can be converted into fully differentiated cells it ought to be possible to find ways to coax them into becoming each of the specialized adult stem cell types. This is desireable because adult stem cells are creating differentiated cells throughout our bodies every day. They are making new skin cells, blood cells, neurons, and assorted other cell types.

However, coaxing embryonic stem cells to become adult stem cell types may not turn out to be any easier than coaxing a given adult stem cell type into becoming other different adult stem cell types. We need to understand in far more detail what makes each adult stem cell type be that type and not some other adult stem cell type or the embryonic stem cell type. It is a lot easier to find indications that one has created a differentiated cell type because differentiated cell types have unique proteins that are well known and used for doing what they do and they make various chemicals that can be tested for. But to stop differentiation at an intermediate step (ie at the adult stem cell step) is harder.

We Need To Find Out Why Adult Stem Cells Are Less Vigorous

While this latest report may demonstrate an advantage of stem cells created by cloning it doesn't answer the question of why those cells are more vigorous. If the report really did find that adult skin fibroblast cells, when cloned, became stem cells that were more vigorous than existing adult stem cells in the same animal then why? One potential explanation is that the cloning process presented the differentiated adult fibroblast's genome with chemical signals that induced it to grow its telomeres. However, there are other possibilities. One big one is that the cloning procedure may just have selected for less damaged DNA. If the scientists had to clone many nucleuses in order to get one that worked then the scientists may have just been selecting for a cell that had the least amount of accumuation of aging-related damage to its genome.

Fixing Telomere Length: If telomere length is the problem a technique may be found to take adult stem cells out of the body, bath them with in genes or drugs that induce telomerase to make long telomeres, and then to transfer the reinvigorated stem cells back into the body. There is a caution here. Even if such a procedure worked and the stem cells became more vigorous the cells might still have a dangerous amount of accumulated DNA damage in them as well. The risk of cancer and metabolic disorders might be increased.

Fixing Accumulated DNA Damage: However, if adult stem cells become old due to accumulation of DNA damage then that becomes a much harder problem to fix. The genome is 3 billion base pairs lone. It will some day be possible to locate and identify all the mutations in an extracted line of adult stem cells. But then all the mutations (or at least all the important ones) would need to be fixed. Gene therapy to fix that amount of damage is far in advance of where gene therapy is today.

Select Cells With Less Damaged DNA: Another possible approach would be to take out large numbers of adult stem, let each cell divide, and take one of each pair and sequence its DNA. Look for cells that have the least amount of DNA damage. Then fix their telomeres and/or do gene therapy to fix the damage that the chosen cells do have. This approach depends on DNA sequencing technology that is under development but not yet available. It also depends on gene therapy technology that is not yet available.

The Relevance Of This Result To Bioethics Debates

Some people have ethical objections to using embryonic stem cells in to develop medical therapies. Embryonic stem cells can be created by fertilizing an egg with a sperm to make an embryo and then letting the embryo divide. However, cloning has now been demonstrated (at least in other species besides humans) as a way to create a viable embryo. This is done by taking an unfertilized egg, taking out its nucleus and putting an adult nucleus into it. Do that enough times and some small percentage of the resulting cells will be capable of developing into a fetus and then baby and then adult. But since cloning can create viable embryos (at least some small percentage of the time) many of the same people who object to using fertilized eggs to create stem cells also object to the use of cloning to make stem cells as well. In their view the fact that a cell has the potential to become a full human adult should imbue that cell with special rights and legal protections.

Note that it is not the objective of this essay to argue for or against the various positions taken in the ethics debate about various forms cloning and stem cell research. My goal here is to show why this latest report about ACT's research is pertinent to those who are making supporting arguments.

Many who argue against embryonic stem cell research claim that ways will be found to induce adult stem cells to do anything that embryonic stem cells can do. One objection to that line of argument is that it may take longer to find ways to tell adult stem cells to become less differentiated and then to differentiate down a different path. For instance, find a way to make blood stem cells to become not just locked into being blood stem cells so that they can then be told to become muscle stem cells. But this latest report strengthens another argument against adult stem cells as competitors to embryonic stem cells: adult stem cells are older and therefore generally less able to do any job they can be coaxed into doing. Therefore adult stem cells may be less able to replace embryonic stem cells for many therapeutic purposes.

ACT Sounds Financially Weak

Will ACT be able to do anything with the results that were the occasion for this discussion? There is reason to doubt on that score. ACT is not doing too well:

Advanced Cell, based in Worcester, Mass., temporarily suspended Cibelli's human cloning efforts for lack of money, and also sold its cattle-cloning subsidiary, Cyagra LLC, to raise cash.

Geron, the Menlo Park, Calif.-based industry leader, laid off a third of its work force and cut research spending to bolster its lagging stock price.

A Different Way To Get Younger Stem Cells

At least in some cases there may be a way to get relatively younger stem cells without either using embryos or cloning. One way to do it is to take stem cells from the umbilical cord of a newborn bady. The umbilical cord is going to be thrown out anyway. Using it for this purposee does not cost any real or potential life. Most kinds of ethical objections that are raised about cloned and embryo stem cells don't really apply to umbilical stem cells because umbilical stem cells are more differentiated than embryo stem cells (ie they are not at the point in development where a cell is at when it is ready to become a full fetus). The problem with umbilical stem cells up to now has been that they are few in number and haven't been easy to grow. However, here's a recent report of a better way to get stem cells from umbilical cords:

One obstacle to using cord blood more routinely as a source of stem cells in transplantation patients is the amount of blood required. Clinical trials have established that higher numbers of blood cells per kilogram of body weight of the recipient are associated with improved transplantation outcome. However, the amount of blood cells collected from cords is often not sufficient for an adult recipient. Scientists have therefore attempted to culture and expand cord blood-derived cells before transplanting them into patients. As they report in the October 21 issue of the Journal of Clinical Investigation, Irwin Bernstein and colleagues (Fred Hutchinson Cancer Center, Seattle, and University of Washington, Seattle), have been successful in doing so. Exposing human cord blood to a particular molecule called Delta-1 under defined culture conditions resulted in an over 100-fold increase in the number of the most immature stem cells. Other progenitors that maintained the potential to differentiate into multiple different blood cell types were also expanded.

On one hand the stem cells in a new born's umbilical cord should be very young and vigorous. On the other hand, these umbilical cord cells are already partially differentiated stem cells that are good at making blood cell types. Therefore it may be much more difficult to coax them to become all the other cell types that embryonic stem cells can become.

An Interesting Result With Fetal Neural Stem Cells

Here's another short report that again is not sufficiently detailed. Some sort of stem cell is induced to become a more differentiated neuron.

In November 11 advanced online Nature Neuroscience, Ping Wu and colleagues at the University of Texas Medical Branch, Galveston, Texas, USA, show that a novel priming procedure for fetal human neural stem cells (hNSCs) can transform them into cholinergic neurons in adult rat CNS (Nature Neuroscience, DOI:10.1038/nn974, November 11, 2002).

Did they take a fairly advanced rat fetus and extract already partially differentiated stem cells from the hippocampus to use to perform this procedure? Or did they take stem cells out of a fetus that was at a much earlier stage in development? Were the extracted stem cells more like embryonic stem cells or where they as differentiated as the sorts of stem cells that are found in adult brains?

Regardless of the type of stem cell these researchers started out with there is one note of caution to bear in mind when reports are made that stem cells have been converted into differentiated cells: It is not a certainty that the conversion was done perfectly accurately. Since we do not know exactly what pattern of genetic regulatory state characterises each cell type we don't know whether some attempt to make, say, a cholinergic neuron really made exactly that and nothing more or nothing less. Each cell type needs large sets of genes turned on and off in a pattern unique to that cell type. Its possible that some of these experiments are yielding cells that have a few extra genes turned on or a few genes turned off that shouldn't be turned on or off.

Update: In The Long Run Embryonic Stem Cells Won't Be Necessary

Scientists are gradually identifying and elucidating the function of the genes that control embryonic development and cellular differentiation:

Philadelphia, PA In the search to understand the nature of stem cells, researchers at the University of Pennsylvania School of Medicine have identified a regulatory gene that is crucial in maintaining a stem cell's ability to self-renew. According to their findings, the Foxd3 gene is a required factor for pluripotency the ability of stem cells to turn into different types of tissue in the mammalian embryo. Their research is presented in the October 15th issue of the journal Genes and Development.

"Stem cells represent a unique tissue type with great potential for disease therapy, but if we are to use stem cells then we ought to know the basis of their abilities," said Patricia Labosky, PhD, an Assistant Professor in the Department of Cell and Developmental Biology. "Among the stem cell regulatory genes, it appears that Foxd3 gene expression keeps stem cells from quickly differentiating that is, developing into different types of tissue holding back the process so that an embryo will have enough stem cells to continue developing normally."

This latest report about Foxd3 adds to an existing list of genes that control embryonic development:

"Our findings implicate Foxd3 as one of the few genes serving as a 'master switch' of the developing embryo," said Labosky. "These genes determine the fate of cells by turning on or off other genes in response to signals in the embryo."

Foxd3 joins previously identified genes, such as Oct4, Fgf4, and Sox2, which control the pluripotency of embryonic stem cells in the early stages of embryogenesis. In their experiments, Labosky and her colleagues found that these genes are still expressed despite the lack of Foxd3. This suggests Foxd3 functions either downstream of Oct4, Fgf4 and Sox2, or along a parallel pathway.

At a molecular level what makes an embryonic cell different than an adult stem cell is probably just a different set of proteins and methyl groups bound at different places on the genome on different genes and regulatory sites. It should eventually become possible to change an adult stem cell (or, for that matter, a fully differentiated cell of any number of cell types) into an embryonic cell by sending in the right kinds and sequences of signals (hormones, drugs that will be discovered, gene therapy) to change the pattern of bound proteins and methyl groups on the DNA. At that point it should be equally possible to change an adult stem cell of one type into an adult stem cell of another type.

The development of a full understanding and the ability to fine tune control of all the genes that govern development could take a long time (10 years? 15 perhaps? my guess is 20 years max). Many scientists who are working with embroynic and adult stem cells are basically hoping that without first taking the time to understand all the details of how genes control cellular growth and differentiation they will be able to find ways to make these cells grow into needed replacement cell types and organs. So the debate about the use of human embryonic stem cells is about what approaches are ethically acceptable as ways to develop new therapies in the short and medium term.

Molecular Switch Positions And Spiritual Definitions Of Life

My guess is that most of the people who hold that it is immoral to use embryonic stem cells in experiments are motivated by a spiritual definition of a human life. In their minds the moment of fertilization is a moment when a spirit enters the fertilized egg. As has already been demonstrated in other species, it is possible to put an adult cell nucleus into an unfertilized egg and the chemicals in the unfertilized egg cytoplasm somehow (as yet not understood) causes changes in the DNA in the nucleus of the adult cell genome that converts it into a state is similar enough to that of a freshly fertilized egg that the cloned cell can develop into an adult of that species. Therefore there is already a method other than fertilization that allows the creation of a new life. These will not remain as the only two methods for creating cells that are capable of developing into mature adults.

Some day we will have the knowledge and techniques to allow us to manipulate adult stem cells into all other types of cells. This would allows us to bypass the use of embryonic stem cells. But it would be a mistake to think this ability would allow us to entirely avoid the need to confront the ethical issues that cause some to oppose the use of embryonic stem cells. The ability to make adult stem cells into increasing numbers of other cell types will eventually extend to include the ability to make adult stem cells into embryonic stem cells. When the details come out about what makes embryonic stem cells different from adult stem cells the bright line that separates them in the minds of many opponents of embryonic stem cell use will become intellectually untenable. If the difference between embryonic and adult stem cells is just a set of molecular switches turned to different states then how can the opponents of embryonic stem cell use define the type of cell that deserves special legal protections? One could start with an adult stem cell and flip just one switch at a time to make it more and more like the switch settings that are characteristic of embryonic stem cells. How many of the switches will have to be flipped into the state that embryonic stem cells have them in before we would have to start treating the cells as embryonic and therefore entitled to special legal protection? Would they all have to be exactly as they are in an embryonic cell in order to warrant legal protection?

Advances in biotechnology promise to obsolesce the historical definitions of how we determine that something has enough of the characteristics of a human to be eligible for some degree of legal protection. The embryonic stem cell and cloning debates are just bush league warm-ups for much larger debates to come. The distance that has historically separated all manner of existing life forms from what we recognize as a human is going to narrow as it becomes possible to create humans in new ways, to give other species some of the qualities of what makes us unique (eg raise the intelligence of other species), and by methods of creating parts and keeping parts alive. We already have the problem of brain-dead humans and humans who are born with brains so defective they lack the thinking abilities that we associated with what it means to be a human. But as it becomes possible to create creatures that possess various subsets of the qualities that define humanity the question of what is a human will grow steadily more difficult to answer. Theoretical philosphical questions will become practical questions needing immediate answers. The debate over such basic questions promises to be highly divisive both within and between different human societies.

Share |      Randall Parker, 2002 November 15 12:06 AM  Aging Reversal

shafir said at March 21, 2003 3:48 PM:

could u please tell me what LLC stands for, as they are used along side MDCK cells during experimetns for Influenza, i need to obtain the meaning of LLC cells for my dessertaion, please help me

from shafir

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