A Demonstration in Mice of Whole Mitochondria Delivered as a Therapy
Mitochondria, the swarming power plants of the cell, become damaged and dysfunctional with age. Can this be addressed by delivering complete, whole, new mitochondria as a therapy? There have been signs in past years that cells can ingest and incorporate mitochondria from the surrounding environment, but few useful demonstrations to show whether or not this is common in living tissues. In the research here, researchers achieve that result, delivering mitochondria into tissues as a therapy, and using this approach to treat an animal model of Parkinson's disease. This neurodegenerative condition is associated with degraded mitochondrial function, especially in the dopamine-generating neurons in the brain; depletion of that cell population produces the visible symptoms of the disease.
Unfortunately it isn't clear as to whether usefulness in addressing mitochondrial dysfunction in Parkinson's will translate to usefulness in addressing the type of mitochondrial dysfunction thought to cause aging in general. The contribution to aging is based on damage to mitochondrial DNA resulting in mutant mitochondria that are both malfunctioning and capable of outcompeting the normal mitochondria present in a cell quite quickly. Delivering new, fully functional mitochondria might not do much in this situation; they would simply be outcompeted again. It still seems worth the attempt if it turns out to be comparatively easy to replicate this demonstration in mice, on the grounds that you never know in certainty until you try, but I'm not optimistic based on the current understanding of the situation. On the other hand, one potentially interesting application of mitochondrial uptake might be to provide people an upgrade from a comparatively poor mitochondrial haplotype to a comparatively better mitochondrial haplotype, as different mitochondrial genomes have different performance characteristics.
Mitochondrial dysfunction is associated with a large number of human diseases, including neurological and muscular degeneration, cardiovascular disorders, obesity, diabetes, aging and rare mitochondrial diseases. Replacement of dysfunctional mitochondria with functional exogenous mitochondria is proposed as a general principle to treat these diseases. Here we found that mitochondria isolated from human hepatoma cell could naturally enter human neuroblastoma cell line, and when the mitochondria were intravenously injected into mice, all of the mice were survived and no obvious abnormality appeared. The results of in vivo distribution suggested that the exogenous mitochondria distributed in various tissues including brain, liver, kidney, muscle and heart, which would benefit for multi-systemically mitochondrial diseases.
In normal mice, mitochondrial supplement improved their endurance by increase of energy production in forced swimming test; and in experimental Parkinson's disease (PD) model mice induced by respiratory chain inhibitor MPTP, mitochondrial replacement prevented experimental PD progress through increasing the activity of electron transport chain, decreasing reactive oxygen species level, and preventing cell apoptosis and necrosis. Since effective drugs remain elusive to date for mitochondrial diseases, the strategy of mitochondrial replacement would provide an essential and innovative approach as mitochondrial therapy.
I wouldn't be writing this off in a haste. I've read enough papers in the last couple of years showing that this method rescues cells in fibrotic tissue as well as neurons in models of PD and AD.
I'm wondering if delivering healthy mitochondria could be combined with this approach
https://www.ncbi.nlm.nih.gov/pubmedhealth/behindtheheadlines/news/2016-07-13-pomegranate-compound-could-combat-complications-of-ageing-/
They claim to be able to kill off damaged mitochondria in rodents. Even had it published in Nature. Results of clinical trials are expected this year.
"Delivering new, fully functional mitochondria might not do much in this situation; they would simply be outcompeted again. It still seems worth the attempt if it turns out to be comparatively easy to replicate this demonstration in mice, on the grounds that you never know in certainty until you try, but I'm not optimistic based on the current understanding of the situation."
If that is the case, why did mitochondrial supplementation produce benefits in this experiment?
Hi Jim,
These weren't aging mice: the authors actually don't state the age of their animals, but the costs and/or hassle of raising mice while they age or acquiring pre-aged likely means that their silence on the matter means they were in the defaultage range, somewhere between weanlings and early full adulthood. So the observed benefits can't really be attributed to anything related to the age-related accumulation of deletions, since the baseline level was likely very low. Also, the benefits were observed over the course of 2 hours, which is much more rapid-onset than you'd expect if transfused mitochondria were somehow "drowning out" mutant mt (whose effects are largely downstream, likely via effects on other cells).
The acute mitochondrial "supplementation" in the otherwise-untreated mice likely just gave them a quick burst of extra ATP, at no biosynthetic cost. Animals administered the mitochondrial toxin MPP+/MPTP, which causes reduced energy production and excess ROS production, would have benefited from extra ATP and cytosolic ROS scavenging, rescuing some dopaminergic neurons, rather than any effect on deletion-bearing mitochondria.
Thanks for the comprehensive reply Michael.
Would there be any value in carrying out the reverse of this experiment - taking some mitochondria from the cell lines that the SENSRF has that are null for certainly mitochondrial genes and injecting them into mice to observe the effects? Would these mitochondria then rapidly take over any cells they were taken up into?
How many of the 13 genes does a mitochondria need to lose before it rapidly takes over a cell?
"The acute mitochondrial "supplementation" in the otherwise-untreated mice likely just gave them a quick burst of extra ATP, at no biosynthetic cost."
Are we soon going to witness "mitochondrial doping" in the Tour de France? Or not if it is undetectable?
@Michael Having concerns about the experimental process is good but I'd like to remind you, pretty much the same process was used when the first senolytics were discovered and you didn't seem too preoccupied pointing out the shortcomings back then.
I've read an article with a similar technique of mitochondrial transfer used on aged human lung tissue from an year or two ago, and it worked there too. So, again, I wouldn't write this one off so hastily.
@Anonymoose - I don't know if the two situations are that comparable.
With senescent cells, the initial 2011 experimental result involved a mouse model was an accelerated model of aging with a faster rate of production of senescent cells. The possible weakness was that it proved senescent cells were harmful in the levels produced in those mice, but perhaps in regular mice senescent cells never reached a level that had a harmful effect on the animals.
With this experiment the mice aren't really aged in any way. They have damaged (or depleted?) mitochondria and some cell death due to the toxic to mitochondria drug administered. AdG's theory on how mitochondria are directly involved in aging is that mutant mitochondria rapidly take over cells, and by old age this number of cells has reached a significant (if still small) percentage of the whole body. With this experiment replacing damaged mitochondria might help (as would boosting the chemical output of the remaining dopamigenic cells). In an aged mouse the mutant mitochondria would quickly take over the cell again, so there may not be any benefit.
Well, the question is how quickly. If it takes months or years, then it could still be something worth doing even without a means to get rid of the mutant mitochondria.
@Arcanyn: I believe that the process hasn't been observed in mid-occurrence in cells; researchers only see the before and the after. That argues for something fairly rapid.
Posted by: Jim at March 8th, 2017 1:06 AM : Would there be any value in carrying out the reverse of this experiment - taking some mitochondria from the cell lines that the SENSRF has that are null for certainly mitochondrial genes and injecting them into mice to observe the effects? Would these mitochondria then rapidly take over any cells they were taken up into?
That's a clever thought. I should say first that there is significant skepticism within the Foundation's network of scientists that have voiced an opinion that the main result is real, and we'd all certainly like to see it replicated before doing much of anything with it.
Even supposing that the finding is a real phenomenon, however, I'm not sure that injecting mitochondria bearing null mutations would work. First, I'm not confident that such mitochondria would be as effective at surviving in the circulation or gaining access to tissue cells, although we don't know the mechanism of the latter yet: assuming that it requires significant outlay of ATP, such mitochondria might simply not be able to pull it off.
Supposing that they got into postmitotic cells, would they then clonally expand - and would they do so successfully enough to entirely take over the host cell (become homoplasmic)? The mitochondrial mutations that expand clonally in aging cells are not null mutations in specific, individual genes of the OXPHOS machinery (as in our cell lines), but large deletions of the mitochondrial genome that take out the genes encoding mitochondrial transfer RNAs, meaning that they are not just unable to produce the particular proteins for which they lack the specific genes, but are unable to make any of their own proteins, whether they have the genes for them or not.
Whether a clean null mutation for a single mitochondrially-encoded protein in the electron transport chain would clonally expand the same way that a deletion-bearing mitochondrial genome would depends on the exact mechanism whereby clonal expansion occurs - something that is not universally agreed. Dr. de Grey's "Survival of the Slowest" hypothesis to explain the clonal expansion of deletion-bearing mitochondria (for which evidence has mounted substantially in the last decade) is that they selectively escape cellular quality control via selective mitochondrial autophagy ("mitophagy"), because with their OXPHOS completely shut down, they are no longer producing any free radicals, so their membranes don't become damaged and they aren't picked up for selective disposal; meanwhile, their genetically-normal but self-damaged cousins are routinely culled. If that's correct, and if it's the only such mechanism, then you'd expect that a full null mutation for an OXPHOS component that were used under all substrates (such as our ATP8 mutation) would similarly tend to clonally expand.
However, mitochondria do undergo fusion with each other, and although their degree of complementation is limited by the way individual genomes are packaged (most mitochondrial protein-DNA complexes contain only a single mitochondrial genome each and seem to mostly operate with local autonomy), there is some ability for an intact genome to provide a protein that another genome inside the same fused mitochondrion or mitochondrial network lacks. Depending on how strong this effect is, it might sustain OXPHOS and effectively mask a single null mutation in a way that can't happen for a mito with a large deletion.
And of course, SOS could be wrong - or there might be other contributors to clonal expansion, such as "survival of the smallest" (which is strictly speaking compatible with SOS), and it might be that both mechanisms have to be active to effect homoplasmic clonal expansion. In that case, a single null mutation might not have (enough) selective advantage to render the cell homoplasmic.
All that said: aside from sheer curiosity, I'm not sure what there is to be gained from such an experiment. Did you have something in mind, Jim, or were you just throwing it out there?
Posted by: Jim at March 8th, 2017 1:06 AM : How many of the 13 genes does a mitochondria need to lose before it rapidly takes over a cell?
We don't really know: full-on null mutations for a single gene are quite rare. Again, in aging you see overwhelmingly homoplasmic large deletions; other mutations, particularly those that are inherited, tend to be heteroplasmic mixtures of loss-of-function mutations (since the full loss of a protein-coding gene would not let the cell replicate and survive embryogenesis).
Posted by: Jim at March 8th, 2017 1:18 AM:
"The acute mitochondrial "supplementation" in the otherwise-untreated mice likely just gave them a quick burst of extra ATP, at no biosynthetic cost."
Are we soon going to witness "mitochondrial doping" in the Tour de France? Or not if it is undetectable?
Oy vey! That hadn't occurred to me ... maybe that would fuel a sudden rush of investments in mitochondrial genetics, for doping and testing ;) .
Posted by: Anonymoose at March 8th, 2017 8:18 PM: @Michael Having concerns about the experimental process is good but I'd like to remind you, pretty much the same process was used when the first senolytics were discovered and you didn't seem too preoccupied pointing out the shortcomings back then.
At first, I really couldn't figure out what you were talking about; thanks to Jim for providing a jogging correction.To be clear, the first senolytic study had robust results in wild-type mice; the earlier proof-of-concept study on ablation of senescent cells of which you're thinking was not of senolytics, but of activation of a drug-inducible "suicide gene" in p16-expressing cells.
I frequently criticize introduce a totally artificial cause of some pathology, and then fix it by taking away or counterating that artificial cause. This tells us nothing about whether the same intervention can prevent or treat the same pathology when it arises from the "natural" degenerative aging process. Similarly, in the case of the study that is the subject of today's blog post, the infused mitochondria were shown to counteract the acute effects of mitochondrial toxin MPTP/MPP+ - but this really doesn't tell us anything about its ability to rescue cells rendered homoplasmic for deletion-bearing mitochondria, or for cells with other mitochondrial damage or dysfunction driven by degenerative aging.
By contrast, I was only mildly cautious about the original proof-of-concept for senescent cell clearance (for which caution see my blog post at the time) because it was not artificial in this way. While the mutation that causes the excess burden of senescent cells in these mice is only an infrequent cause of senescence in the cells of "normally"-aging animals, the cells themselves are senescent regardless, and the intervention against them didn't rely on interfering with any particular metabolic driver of senescence. Instead, it simply ablated the cells themselves. Remember, one of the key advantages of the "damage-repair" heuristic of SENS is exactly that it attacks the celluar and molecular damage of aging directly, and thus works irrespective of the metabolic drivers of that damage. One could therefore predict with high confidence that similar benefits would accrue when ablating senescent cells in normal, otherwise-healthy but aging mice - and people.
Posted by: Anonymoose at March 8th, 2017 8:18 PM
I've read an article with a similar technique of mitochondrial transfer used on aged human lung tissue from an year or two ago, and it worked there too. So, again, I wouldn't write this one off so hastily.
There are a few studies generally along those lines, but Jim has summarized the reasons why they don't provide us any real evidence in support for a use for this treatment as a therapy against the degenerative aging process. Again, these are models of rescue of acute or ongoing insult from things other than degenerative aging processes, such as ongoing cigarette-smoke-induced lung damage in mice, or mouse models of allergic airway inflammation and acute lung injury, including endotoxin-induced lung injury. None of that supports a benefit for "normally"-aging lung or other tissue, any more than the benefits of chemotherapy in cancer patients is such evidence.
And all the studies I've seen so far involve intercelluar transfer of mitochodnria along with cytoplsmic contents and trophic factors from stem cells to injured cells or tissue in which the donor cells are in direct contact, which is a quite different intervention from intervenous injection of "naked" mitochondria. Can you cite something that is a closer match to the current study?
"Mitochondria were added into the serum and incubation at 37 °C for 0, 15, 30, 60, 120 min, respectively. Afterwards, samples were centrifuged at 3000g for 10 min at 4 °C. The mitochondria were collected and measured by Mitotracker red CMXRos or GFP, and membrane potential assay"
-we know mitochondriain blood are immunogenic, my question is, is 2 hours really long enough to rule out a response,especially one in anin vitro serum assay?