May 18th, 2013 by Michael Ricciardi
[UPDATED – May 24, 2013; see note at bottom] A team of reproductive biologists led by Shoukhrat Mitalipov at the Oregon Health and Science University (Beaverton, OR) have successfully created ‘patient-specific’ embryonic stem cells (ESCs) through cloning.
The improved cloning technique used by the team is much more efficient than previous approaches that have achieved only partial successes. The breakthrough will surely reignite ethical debate over the controversial technique that has long-promised cures for diseases such as diabetes and Parkinson’s disease, to name just two.
The purpose of cloning ESCs is to create perfectly matched (thus ‘ patient-specific’) cell lines (cultures). ESCs are ‘pluripotent’ and capable of becoming any other cell type (except placenta cells) — given the right molecular prompts. But the technique requires the destruction of embryos in order to derive the ESCs that can then be made to ‘differentiate’ into any other type of cell (e.g., heart cells, liver cells, neurons, etc.).
This required destruction of the source embryo is what has sparked the ethical debate over its use (and the banning of stem cell research of this kind in many states). Additionally, there have been recent cases of exposed fraud* and, newer, alternative techniques (i.e., cell reprogramming via ‘induced pluripotent stem cells’ [iPSCs] or transdifferentiation) have been developed that would seem to obviate the need for cloning.
The cloning technique used here is known as ‘therapeutic cloning’, or somatic cell nuclear transfer (SCNT) and is essentially the same technique — with a few important ‘tweaks’ — used to create the first cloned sheep (‘Dolly’). In SCNT cells, a donor cell is fused with an egg (also a single cell) from which the nucleus (containing the egg’s DNA) has been removed. The egg cell then reprograms the transferred DNA from the donor cell into an embryonic state which then begins dividing until a multi-cell stage (i.e., a blastocyst) is achieved. The newly divided cells are then harvested to create a cell line (lineage) that is genetically matched to the donor and capable of differentiating into any cell type desired.
* Woo Suk Hwang of Seoul National University in S. Korea, published studies in 2004 and 2005 which fraudulently claimed to have successfully cloned ESCs.
The New and Improved Experiments
Mitalipov et al perfected their technique using monkey eggs before working with human eggs. In their technique, the researchers used an inactivated Sendai virus to induce the egg and donor cell fusion stage and then applied an electric jolt to trigger embryonic development (i.e., cell division). Following initial failures to generate stable cell lines, caffeine was added to protect the egg from premature development. These techniques are not new, but when used in the right combination, produce stable, growing ESC colonies.
The experiments, begun in September, 2012, took only a few months to achieve success. The techniques were successfully applied to cloning monkey cell lines back in 2007 and observers have wondered why it took six years to to do the same with human ESCs. Mitalipov asserts that much of that time was spent dealing with US stem cell regulations on ESC research (imposed under George W. Bush).
Some of the cloned cell lines were created using fetal skin cells while others were derived from an 8-month old patient with a rare metabolic syndrome. These latter cells were used to prove that ESCs could be derived from matured donor cells. Also, the technique used here did not require destruction of a large number of eggs; it took 15 eggs to generate the latter cell line and just 5 eggs to generate the former.
The team conducted several test to verify that these were viable SCNT cells capable of differentiating into other cell types (including self-contracting heart cells, or cardiocytes).
The Future of Therapeutic Cloning
This improved cloning technique to derive patient-specific ESCs comes along at a time when many are questioning the need for therapeutic cloning. These critics point to fairly new iPSC technology — first developed in 2006 (Takahashi K, Yamanaka S) — and an improved version of this technique (RNA-induced Pluripotent Stem cells, or RiPS cells; Rossi et al), which seem to be just as successful at generating patient-specific cell lines — with no embryonic destruction required.
As long as the technique requires destroying even a single embryo, no funding from the US National Institutes of Health (which are funded by taxpayers) can be used to derive or experiment with SCNT cells — putting a brake on most new medical research in this field.
Further, the procedure is expensive in that eggs are acquired through paid donors (who are paid between $USD 3000 – 7000) and bio-ethicists have argued that this exploits the poor and could promote an “organ trade”.
Their are even fears that the breakthrough could trigger a new round of “cloning hysteria” over the possibility of cloning a human being. But fellow researcher (and co-author of the paper) Masahito Tachibana asserts that human cloning is not possible using their (SCNT) technique,and that why this is so will be explained in a forth-coming paper (note: earlier experiments with monkey embryos by Mitalipov failed to produce an actual cloned monkey).
In the meantime, researchers look forward to clinical comparison between the two techniques (i.e., iPS and SCNT cells). Some have argued that iPS cells are not truly “reprogrammed”, due to defects caused by inserted DNA (but see the linked article above, ‘RiPS cells’), and, that SCNT cells are more similar to ESCs derived from in vitro fertilization techniques. To that end, Mitalipov and Tachibana have begun a new study comparing the two, derived-cell techniques using an identical donor cell for the fused-cell source.
Medical biologists around the world are eagerly awaiting the results — and anticipating a renewed ethical debate.
UPDATE (May 24, 2013): Questions surrounding the duplicate use of ES images and mislabeling of images (cloned verses control) have emerged regarding the recently published paper (in the journal Cell). Read the Science magazine news report.
Some source material for this post came from the Sci Am article: ‘Patient-Specific Human Embryonic Stem Cells Created by Cloning’ by David Cyranoski and Nature magazine (published May 15, 2013)
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