Cell Spreading and Mitochondrial DNA Deletions

Researchers here argue for decreased cell spreading in old skin to be a cause of higher levels of mitochondrial DNA deletions in longer-lived skin cells. Their methodology leaves open the possibility of other possible causes for the data they have gathered, however. I don't believe that they have convincingly demonstrated causality at this point. Nonetheless worth reading, I think.

Why is this interesting? Because mitochondrial DNA damage is strongly implicated as a contributing cause of degenerative aging, but there is considerable debate over how and why this damage occurs and accumulates with age. The SENS rejuvenation research viewpoint is to skip the debate over causes and just repair the damage and measure the benefits that result, but this is not a popular viewpoint in the scientific community, where most participants are aiming for complete understanding at some indefinite future date rather than the production of useful therapies as soon as possible. So we are going to see much more research in the future exploring this aspect of biochemistry.

Mitochondria are the power plants of the cell, each cell containing a swarm of hundreds of these descendants of symbiotic bacteria, each of which contains at least one copy of the remnant DNA left over from that of their ancestors. Evolution has moved much of this DNA to the cell nucleus, or it has atrophied, leaving just a small number of genes that are passed from mother to child. Mitochondrial populations are very dynamic, constantly dividing and fusing, passing chunks of protein machinery between one another, and culled by cell quality control mechanisms when damaged. Damage occurs to cellular machinery all the time, and near all of it is repaired. Mitochondrial DNA (mtDNA) deletions can be a real problem, however: DNA encodes for the proteins needed for correct function, and there is a way in which a mitochondrion with just the right type of damage can fall into a malfunctioning state that provides it an advantage in replication and resistance to quality control. When that happens the whole cell is quickly taken over by the descendants of that dysfunctional mitochondrion. The cell itself becomes broken, exporting harmful reactive molecules into surrounding tissues. A small but influential population of cells are in this state by the time old age rolls around, and they cause significant harm.

Why does this DNA damage happen? Some researchers believe it is due to the proximity of mitochondrial DNA to the energetic processes by which mitochondria produce chemical energy stores, coupled with comparatively poor DNA repair processes available in the mitochondria. Other researchers consider that the damage happens during mitochondrial replication, and other changes taking place in cells over the course of aging might explain a rising level of errors that occur during this replication. There are other theories - in biochemistry there are always other theories - and the one described in the following open access paper is one such.

Age-associated reduction of cell spreading induces mitochondrial DNA common deletion by oxidative stress in human skin dermal fibroblasts: implication for human skin connective tissue aging

In human skin, dermal fibroblasts are responsible for collagen homeostasis. Consequently, impaired dermal fibroblast function is a major contributing factor in human skin connective tissue aging. We previously reported that a prominent characteristic of dermal fibroblasts in aged skin is reduced spreading and contact with collagen fibrils, causing cells to lose their typical elongated spindle-like morphology and become shorter with a rounded and collapsed morphology. In young healthy skin, dermal fibroblasts attach to intact collagen fibrils and achieve normal cell spreading and shape. However, in aged dermis the collagen fibrils are fragmented, which impairs fibroblast-collagen interactions. These alterations impair fibroblast spreading and function. While cell shape is known to regulate many cellular functions, the molecular basis of their impact on dermal fibroblast function and skin connective tissue aging are not well understood.

Although dermal fibroblasts are the major cell type responsible for the maintenance of dermal connective tissue homeostasis, little is known about the role of mtDNA common deletion in aging dermal fibroblasts. Dermal fibroblasts have a very low proliferative rate which would allow for an accumulation of mtDNA deletion. Additionally, the relationship between age-related reduced cell spreading, which is a prominent feature of aged dermal fibroblasts, and mtDNA common deletion has been virtually unexplored. Based on this information, we explored the possible connection between age-related reduced cell spreading and mtDNA common deletion in the dermis of human skin. We found that mtDNA common deletion is significantly increased in both naturally aged and photoaged human skin dermis in vivo, and that reduced fibroblast spreading induces the increase in mtDNA common deletion through increased endogenous reactive oxygen species (ROS).

We modulated the shape of dermal fibroblasts by disrupting the actin cytoskeleton with latrunculin-A (Lat-A), which rapidly blocks actin polymerization. As expected, disruption of the actin cytoskeleton impaired fibroblast spreading and resulted in a rounded shape. Reduced cell spreading was associated with a significant elevation of mtDNA common deletion. As mitochondrial morphology is crucial for normal mitochondrial function, we assessed mitochondrial morphology. These data indicated that the gross shape of mitochondria was similar between Lat-A treated cells and control cells. It has been reported that cellular damage from reactive oxygen species (ROS) likely plays an important role in mtDNA deletions as well as in the aging process. We therefore examined the relative oxidant levels in fibroblasts using redox-sensitive fluorescent dye. Normal well-spreading fibroblasts displayed a very low level of oxidant-generated fluorescence. In contrast, reduced-spreading fibroblasts displayed intense oxidant-generated fluorescence.

We next investigated whether boosting cellular antioxidant capacity could protect against mtDNA common deletion associated with reduced cell spreading. We chose N-acetyl-cysteine (NAC), which is an antioxidant and metabolic precursor of glutathione. Reduced cell spreading increased mtDNA common deletion in a time-dependent manner, and that the increase was significantly prevented by NAC treatment. These results indicate that the deleterious effects of endogenous oxidative exposure are responsible, at least in part, for reduced-cell-spreading-associated mtDNA common deletion.

I have to think that the conclusion to be drawn here is that messing with the cell cytoskeleton is a bad thing, not that lack of cell spreading is a bad thing (though it probably is, just not demonstrated to be via this methodology). An item that immediately springs to mind is that progeria involves disruption of cytoskeletal structure in cells, and I'm sure people with more experience than I could come up with other off the cuff examples of cytoskeleton dysfunction producing cellular dysfunction. So here I'd want to see a replication of the mitochondrial DNA deletion data using another completely distinct methodology of preventing cell spreading before giving this too much consideration. It is easy to break things in biochemistry and produce results that look somewhat like aging, since breakage causes damage, and aging is an accumulation of damage. It is, however, hard to prove that any given artificial breakage is relevant to normal aging, and most are not.

To finish up for today I'll again make the point that the research community could skip this painstaking investigative work in order to focus on producing methods of repairing mitochondrial DNA damage, or delivering the proteins via another method, such as the allotopic expression technology funded by the SENS Research Foundation and presently under active development by Gensight. Fix the damage and see what happens, and if the repair is good enough and frequent enough then it doesn't matter how the problem occurs. Then, with the luxury of time, back to the labs to figure out every last detail of what happens if you don't take the treatments. The present status quo seems back to front, given that we're all aging to death.


@Lou - I can't get read all of that first paper due to the paywall, but it seems they have created a mouse mutant with a mitochondrial gene disabled in ALL cells. The so this is not a test of the mitochondrial reductive hotpot theory put forward by Aubrey de Grey.

You could perhaps test this by generating stem cells and then fibroblasts with this mitochondrial gene disabled, then inject a handful into a mouse or monkey and see what happens. It would probably just be simpler to allotopically express the gene with some chemical switch to turn it on, let the animals age, and then turn on the allotopic expression of the gene and see what happens.

Posted by: Jim at August 4th, 2015 8:13 AM

Perhaps, there exists chimeras which are cellular mosaics of two closely related species with significantly different longevities. I believe that I read about some experiments like this years ago, but cannot find references. This could be another approach?

Posted by: Lou Pagnucco at August 4th, 2015 12:52 PM

Regarding the PNAS.org study, in my view, the causes for this study’s findings were not "age" itself, but;

-Sod2 deficiency;
-Its influence on varying keratinocyte and epidermal stem cell levels; and
-Its influence on other items shown in the supplementary material, such as varying “mRNA levels of wound healing-related growth factors.”

However, the researchers said they could “identify a previously unidentified age-dependent role for mitochondria in quality and wound closure,” and repeated the age-dependent phrase in the study title.

I guess the “age was the cause” meme is hard to stop repeating.

Posted by: PRice at August 16th, 2015 2:06 AM

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