Nuclear DNA from one individual is compatible with mitochondrial DNA from another, and the procedure can be successfully performed with eggs that have been previously frozen. But even very low levels of contamination with donor mitochondrial DNA could cause the resulting stem cells to ultimately revert to having the mitochondrial type the transplant procedure is supposed to prevent, suggesting a need for procedures that can eliminate mitochondrial contamination during nuclear transplant.
A team from Columbia University published those conclusions in the May 19, 2016, online issue of Cell Stem Cell.
Mitochondria are the only organelles in the body that come with their own DNA, probably reflecting an evolutionary history of being independent organisms that took symbiosis as far as it can go.
Like nuclear DNA, mitochondrial DNA can contain disease-causing mutations. According to the Thomson Reuters Incidence & Prevalence Database, the incidence of mitochondrial disease in the working-age population is about six per 100,000 individuals, putting them on a par with disorders like Huntington's disease and amyotrophic lateral sclerosis.
As one might expect from the fact that mitochondria have 37 genes, and also use proteins produced from nuclear DNA, mitochondrial disease is an umbrella category. But because mitochondria are the main cellular energy producers, mitochondrial diseases tend to affect organs with a high energy demand, notably the nervous system and the heart.
Mitochondrial DNA is inherited exclusively from the mother, because sperm contribute only DNA and no cytoplasm during fertilization. One consequence is that it is possible to combine the nuclear DNA of a woman with mitochondrial disease with the cytoplasm of an egg from a healthy donor, which could cure her children of the disease.
The procedure is sometimes called three-parent IVF, though for now, at least in the U.S., that moniker is theoretical, as its clinical use is banned by the FDA. (See BioWorld Today, Feb. 27, 2014.)
In the U.K., Parliament has in principle approved a related procedure, pronuclear transfer, with strict controls, though there are no reports of any children having been born as a result.
In 2012, researchers reported success in combining the cytoplasm of one human egg with the nuclear DNA from another, fertilizing the egg, and growing the resulting embryos to the blastocyst stage that allowed the derivation of stem cells, via a method they called spindle transfer. (See BioWorld Today, Oct. 25, 2012.)
The spindles themselves contain mitochondria, but in the work published in 2012, the authors found that those mitochondria ultimately disappeared after transplantation, leading to cells with nuclear DNA from one donor and mitochondrial from another.
In the current study, the authors derived stem cell lines after such nuclear transfer to take a more comprehensive look at the fate of both sets of mitochondrial DNA after mitochondrial replacement.
There have been some concerns that mitochondrial DNA that is completely unrelated to the nuclear DNA it needs to cooperate with could itself prove detrimental to mitochondrial function, and so the team also wanted to see how easy, or difficult, it is for nuclear DNA from one individual and mitochondrial DNA from another to essentially play nice.
The team also looked at whether the eggs could be frozen before the procedure.
"Mitochondrial replacement can be performed with cryopreserved oocytes . . . which obviates the need for synchronization" of the donors' and recipients' menstrual cycles, making the procedure that much easier, Dieter Egli told BioWorld Today.
The results of compatibility testing were likewise encouraging. When the team tested disparate mitochondrial genotypes, with one set of African and one set of Caucasian mitochondrial DNA in some cases, Egli said that "we found no advantage" of having well-matched mitochondrial and nuclear DNA during the procedure.
But in terms of actually getting rid of the offending mitochondrial DNA, the results were not a slam dunk.
"In most cases, this [transfer] is very effective and the mitochondrial exchange is complete," Egli said. But "in some cases, the mitochondrial genotype that we are trying to eliminate comes back."
Typically, the mitochondrial DNA from the nucleus donor declined over time. But in some cases, the proportion of mitochondrial DNA from the nucleus donor increased over time. Such an increase "could potentially wipe out the therapeutic benefit" of the treatment, Michio Hirano told BioWorld Today.
Hirano, a professor of neurology at Columbia University Medical Center, and Egli, assistant professor of molecular genetics at Columbia University, are co-corresponding authors of the paper.
Depending on both the nature of the mutation and the exact proportion of the DNA type, partial mitochondrial replacement "could be beneficial," Hirano said. But there is still a risk.
A female child conceived via the procedure could also pass on the condition to her children if her nuclear mother's mitochondrial DNA were to gain the upper hand in her germline cells.
The team is planning to test whether other methods of combining nucleus and cytoplasm could eliminate any mitochondria being transferred along with the nucleus.
Other groups have reported that polar body transfer, another method for SCNT, can avoid mitochondrial contamination more often than spindle transfer.
The work now published in Cell Stem Cell also used only eggs with normal mitochondrial DNA, and the team wants to test whether using cells with mitochondrial DNA mutations will change the results.
Finally, they want to test whether there are benefits to using eggs that are genetically matched on certain characteristics. Their current study showed that the procedure could lead to viable cells without such matching. But there may be subtle benefits to matching that would become apparent only with closer study.