Michael Fossel Michael is President of Telocyte

June 20, 2018

Aging and Disease: 2.7 Cell Senescence, Changes In Molecular Turnover, Mitochondria

Filed under: mitochondria,senescent cells,Telomeres — Tags: , , — webmaster @ 9:28 am

Many people assume that mitochondria (and free radicals) lie at the heart of aging or that they are “the cause” of aging. Despite having a central role in aging, the assumption that mitochondria cause aging is both simplistic and – not to put too fine a point on it – is totally at odds with both logic and evidence.

Mitochondria do not cause aging.

This is not to say that mitochondria don’t play a central role in aging cells, but playing a central role is not the same as being the cause nor the same as an overarching explanation for aging, let alone an explanation for age-related disease. By analogy, fevers and elevated white blood cell counts (leukocytoses) play a central role in the clinical presentation of many viral infections, but they neither cause nor explain any viral infection. To describe the presentation of a viral infection, you should include the clinical symptoms and signs, including fevers and high white counts, but while they are an integral part of the process of infection, they don’t cause infection. With that in mind, let’s consider mitochondria, starting with a high-level perspective and then looking at the way mitochondria change in aging cells. In so doing, we will see how mitochondrial aging plays a role in cell aging, but we will also see that mitochondrial aging is secondary to changes within the nucleus, and specifically to changes in gene expression as modulated by telomere shortening.

Previously, we pointed out that every cell in your body is – biologically speaking – several billion years old, yet those cells only show evidence of aging during the several decades of a typical human lifespan. We each inherit a line of cells that didn’t age for billions of years, and yet begin aging during the several decades after birth. The same is true for mitochondria. Every mitochondria in the human body is at least 1.5 billion years old, the time at which the first mitochondria and eukaryotic cell joined forces. If you suggest that all aging is due to aging mitochondria, then you are immediately brought up short by the biological provenance of mitochondria. You end up forced to admit that mitochondria didn’t age for 1.5 billion years and yet (as with your cells themselves) begin aging during the decades after birth.

You inherit mitochondria that have never aged and that then age rapidly during your lifespan. If mitochondria cause aging, then why didn’t they age for the past 1.5 billion years? Why suddenly now? In short, if you want to explain aging by pointing to mitochondria, then you are going to have to back up a step and explain mitochondrial aging itself.

To put it bluntly: if mitochondrial aging causes human aging, then what causes mitochondrial aging?

As we’re just seen, you can’t very well blame “wear and tear”, since your mitochondria are all 1.5 billion years old and they did perfectly well until you came along. If mitochondria play a key role (and they do), you still have to ask why mitochondria sometimes do age and sometimes don’t, or why mitochondria didn’t age for billions of years, but then suddenly age after conception. We can’t simply ignore the question, wave our hands, and blame “free radical damage”.

Yet it is clear that mitochondria show changes during cell aging. There are two clear changes in metabolism: they become less efficient at creating energy (ATP) and they become more likely to produce free radicals (ROS). Putting it graphically, we see that as telomeres shorten, as gene expression changes, and as cells progress toward senescence, we find that the mitochondria show a decrease in ATP production and an increase in ROS production. Putting it differently, the ratio of ATP/ROS goes down with age. As our cells age, they get sloppier, taking more effort to produce usable cellular energy, while simultaneously creating more damaging ROS as they do so. To be specific, young cells typically produce about 1-4% ROS for every mole of ATP, but this percentage increases as our cells senesce. Nor, as we will see, is ROS production the only problem.

To understand why mitochondria lose efficiency, and why it is the result of telomere shortening, we need to understand a bit about mitochondria and their dependence upon the nucleus. It’s true that the mitochondria have their own 37 genes and that these code for 13 proteins that are critical to aerobic metabolism, but it is equally true that most mitochondrial enzymes are imported from the nucleus. Likewise, while the rate of mitochondrial DNA damage is 3 to 20-fold higher than that of nuclear DNA damage (due to the high concentration of mitochondrial ROS, the lack of protective histones, the lack of introns, and less effective mitochondrial DNA repair), mitochondrial DNA repair, replication, and transcription, as well as mitochondrial maintenance generally (including membrane maintenance) are entirely dependent upon the nuclear DNA. Moreover, most of the aerobic enzymes (including those required for the citric acid cycle) have components that are coded for in nuclear genes. In other words, mitochondrial function is almost entirely dependent upon gene expression in the nucleus, which is modulated by telomere shortening. The mitochondria and mitochondrial energy metabolism (including both ATP and ROS production) are completely at the mercy of nuclear gene expression, and are not independent. Aging occurs because changes in nuclear gene expression result in mitochondrial dysfunction, not the other way around.

Looking at this visually, almost all the enzymes required for normal mitochondrial function are based in the nucleus, not in the mitochondria. The mitochondria, far from being an island unto itself, is entirely dependent upon nuclear DNA, the rates of nuclear gene expression, and telomere modulation of that gene expression.

We have already pointed out that, as cells age, the mitochondria become less adept at producing energy, as well as increasingly prone to producing ROS. By implication, these two changes in energy metabolism are both the result of changes in gene expression within the nucleus, but these are far from the only changes that occur with cell aging. A young, fully functional cell has four processes which address the dangers of ROS. These four processes are:

 

  1. Production:           ROS are produced within the mitochondria
  2. Sequestration:     Membranes restrict ROS within the mitochondria
  3. Trapping:             Scavenger molecules “capture” ROS outside the mitochondria
  4. Repair:                 DNA repair and molecular turnover deal with ROS damage

 

The bulk (perhaps 93%) of ROS are produced within the mitochondria. In young cells, the rate of production is relatively low and even those ROS that are produced are generally kept within the mitochondrial membranes. As cells age (and as telomeres shorten and gene expression alters), the enzymes required for efficient energy production are turned over less rapidly, are less available, and are more likely to be denatured: the outcome is that the rate of ROS production climbs. Moreover, a similar problem is occurring within the membranes that wall off the mitochondria: the enzymes responsible for cell membrane molecules – mostly lipid molecules – are also down regulated, with the result that membrane molecular turnover slows, the percent of damaged (oxidized) lipids increases, and the membranes are increasingly porous, allowing ROS leakage to increase. Adding to the membrane problem is that membrane leakage also rises in the nuclear membranes, so the rate of DNA damage rises even as the rate of DNA repair begins to fall. In short, as telomeres shorten and gene expression changes, the mitochondria not only produce more ROS, but are more likely to leak ROS into the cytoplasm and the nucleus.

In the young cell, such ROS are rapidly trapped by multiple mechanisms, including SOD (superoxide dismutase), catalase, and other enzymes coded for in the nucleus (as well as trapping due to dietary scavenger agents, such as urate, tocopherols, etc.). As cells age, however, these scavenger enzymes are recycled more slowly and at any given time the older cell has an increasing percentage of dysfunctional ROS scavenger molecules. Worse yet, the rate of DNA repair declines, as does the rate of recycling of damaged molecules generally.

The result of these four processes is the “perfect storm”, a multiplicative set of four effects causing an exponential increase in overall cell damage.

  1. Mitochondrial ROS production increases,
  2. Mitochondrial and nuclear membrane leakage increases,
  3. ROS damage increases as scavenger efficiency declines, and
  4. Cellular responses to damage (molecular recycling and DNA repair) decline.

It is little wonder that antioxidants have little practical effect upon cell aging or age-related diseases. An increase in dietary anti-oxidants shows almost no effect upon the third of these four effects (scavenging) and no effect whatsoever upon the other three effects (production, leakage, or recycling/repair of damage.

As cells age, there are critical changes within the mitochondria as well as in the cell’s ability to respond to the increasingly dysfunctional mitochondria. However, all of these changes – in production, sequestration, scavenging, and repair – are the result of changes occurring within the nucleus. Much of what happens in cell aging can be seen in the increasing dysfunction of the mitochondria, but all of these changes are secondary to changes within the nucleus.

Specifically, it is telomere shortening and its effect upon gene expression within the nucleus that results in mitochondrial aging. Mitochondrial changes are a key part of the tragedy that plays out on the stage of cellular aging, but that tragedy is directed by the nuclear chromosomes – and by our telomeres.

Next Time: 2.8 Cell Senescence, Changes In Molecular Turnover, Extra-Cellular Molecules

June 13, 2018

Aging and Disease: 2.6 Cell Senescence, Changes In Molecular Turnover, DNA Repair

Why are we more likely to get cancer as we age?

Not only does the incidence of cancer go up with age, but it goes up exponentially. Why? Moreover, the exponential rise is seen in most species, regardless of their lifespan. It’s not the years, it’s the aging process, regardless of time. Why? The key to these questions lies with the rise of DNA damage as we age. But just as with other kinds of cell damage – free radicals in the mitochondria, for example – the issue is not the rate of damage, but the rate of maintenance. In the case of DNA, however, the key feature to maintenance isn’t the rate molecular turnover, but the rate of DNA repair. DNA is the only molecule that is repaired rather than simply replaced. We replace (i.e., recycle) all other molecules in our cells (and even outside of our cells), but we never replace DNA. Instead, we repair it with great effort and in exquisite detail. DNA carries priceless information in its structure, so rather that just recycling the molecule (breaking it down and building a new molecule), our cells go to enormous lengths (and enormous metabolic cost) to find and repair every single error. Without delving into detail, let’s look at an overview of DNA damage, DNA repair, and the clinical implications for aging cells – and aging people.

DNA damage is continual, as is repair. DNA damage occurs continually due to radiation, oxidation, toxins, viruses, and even spontaneous thermal disruption (even at normal body temperatures) with an incidence estimated at up to 106 hits per cell per day. If unrepaired, the result will not only be a dysfunctional individual cell, but a cell that divides without control, thereby harming (and even killing) the entire organism. Ultimately, uncontrolled cell division is expressed clinically as cancer. Left unrepaired, DNA damage becomes fatal. Clearly DNA repair is critical, and must be both constant and all-but-flawless for any organism to survive.

 

DNA repair is, like most biological concepts, remarkably (almost indescribably) complex. No matter how we discuss it, there will be exceptions, qualifications, and additional intricacies which remain unaddressed in our discussion. We will therefore and of necessity, present a simplified summary of DNA repair, one which presents only a high-level, conceptual view of the cell’s response to a single type of DNA damage (single-base errors), while ignoring other types of DNA damage (e.g., double-strand breaks). With this caveat in mind, we will characterize DNA repair as being handled by four basic families of DNA repair enzymes which have these functions:

  1. Identification: find the damaged DNA base and flag it for removal
  2. Excision: remove the damaged DNA base from the strand
  3. Replacement: insert the correct DNA base into place in the strand
  4. Ligation: link the new DNA base to neighboring bases in the strand

In the aging cell, and correlated with telomere shortening, the expression of all four of these types of DNA repair enzymes are down-regulated. This down regulation is typical of cell senescence and is modulated by the telomere. As the telomere shortens, all four repair processes are down-regulated. DNA repair continues, but at a slower pace. Young cells repair DNA almost instantly, older cells repair DNA but at a more lackadaisical pace. The result is that, at any given moment, older cells are more likely to have unrepaired DNA.

The result is that slower DNA repair – and the rising percentage of (as yet) unrepaired DNA damage – means a higher likelihood that such damage will affect the cell’s ability to control cell division. For example, if the damage occurs to the DNA repair genes themselves or to the genes that are central to the cell cycle braking system (which would otherwise prevent cells with DNA damage from dividing), then the cell may replicate and carry the DNA damage into the daughter cells. The result is a cascade of increasing cell damage and a decreasing ability to control cell division. In short, the stage is set for malignancy, clinical cancer, and death.

We begin to see why cancer rises with age. As cells lose telomere length, DNA repair slows, and the risk of cancer rises. Worse yet, however, each of the steps involved in DNA repair are multiplicative, that is, each step will have an impact on all subsequent steps. So if detection slows and the number of DNA errors doubles, then if excision slows, the number of DNA errors goes up another factor of two, i.e., the DNA errors go up four-fold. When you add in replacement and ligation, the effects multiply one another again, with the result that if we down-regulate all of the steps in DNA repair, the increase goes up exponentially.

Most people assume that cancer rates climb with age because of a longer lifetime means a greater cumulative exposure to carcinogens. In fact, the rate of cancer isn’t correlated with years so much as it is with percent of lifespan. For example, mice have an exponential increase in cancer, just as humans do, despite the fact that the average lifespan of a mouse is about 40 time shorter than the average lifespan of a human. It’s not the years, it’s the rate of DNA repair that determines how fast that exponential curve rises. Ultimately, the deciding factor is not cumulative exposure, but the rate of repair. Mice slow DNA repair over a short lifespan and their rate of cancer goes up exponentially in only two years; humans slow DNA repair over a long lifespan and their rate of cancer goes up exponentially over a much longer lifespan. It’s not a matter of having good DNA repair genes, nor is it a matter of chronology. The deciding factor is neither time nor genes, but gene expression and gene expression is controlled by telomere shortening.

If we take the curves for cancer in mice and humans and overlap them to show not years but lifespans, then the curves become identical. It’s not the years, it’s the rate of repair. If we want to prevent or treat cancer, we shouldn’t be focusing as much on exposure to carcinogens, but on cell senescence. Putting it bluntly, if only slightly simplistically, the reason we get more cancer as we age isn’t a matter of what we were exposed to, but the rate at which we repair the damage that is constantly in play over our lifetimes.

We get cancer because of cell senescence.

 

Next Time: 2.7 Cell Senescence, Changes In Molecular Turnover, Mitochondria

June 8, 2018

Aging and Disease: 2.5 Cell Senescence, Changes In Molecular Turnover, Most Molecules

Filed under: Aging diseases,mitochondria,senescent cells — Tags: , — webmaster @ 3:17 pm

As the human body is composed of cells, so are cells composed of molecules. It is true that the cell encompasses a plethora of organelles (membranes, mitochondria, nuclei, Golgi bodies, ribosomes, etc.), but each of these organelles is in turn composed of pools of various molecules. Just as cytoplasm is a “soup” of molecules, organelles are also collections of molecules. The precise types of molecules, their numbers, and their rates of turnover vary from organelle to organelle. The most common molecular types are lipids and proteins, but with admixtures of other molecules, such as carbohydrates (such as sugars), as well as hybrid molecules, such as glycolipids and glycoproteins. As you’d guess, the complexity not only doesn’t stop there, it barely begins there. To talk of a few simple molecular types is only a fuzzy, naïve sketch of that complexity involved in living cells. We’ll look a bit deeper at t he molecular types, then take a simpler view and focus on the only critical feature in aging cells, namely molecular turnover.

Looking at the cell as a whole, the typical human cell is (by numbers) about half lipid molecules and about half protein molecules. Most of the lipids are in membranes (such as the cell membranes, the mitochondrial membranes, and the nuclear membranes); most (but by no means all) of the proteins are in solution in the intracellular fluid. While the membranes have far more lipid molecules than protein molecules, the proteins are heavier (and larger). So while lipids are more numerous (about 50 x as numerous) than proteins in the membranes, the mass of the lipids and the proteins (as well as glycoproteins, etc.) are about equal.

While the lipids determine how the membrane acts in a general sense, it’s proteins that determine the functional (and the active) properties of the membrane. So while there aren’t as many protein molecules, but what they lack in numbers they more than make up for in their importance to cell activity. What about cytoplasm? About 40% of body weight is made of the intracellular fluid, where lipids are heavily outnumbered. In the cytoplasm, it’s the proteins, the electrolytes, and other molecules that determines the activity.

While proteins (as well as glycoproteins, etc.) come in thousands of types, even the lipids are a complex family. The most commons lipid molecules are phospholipids, but there are also cholesterol molecules (sometimes as numerous as phospholipids) and glycolipids, which are sugar-lipid molecules, especially common on the outer cellular. To make things even more complex, some molecules are complex conglomerates of both proteins and lipids.

That’s the introduction to the complexity, but from our standpoint – aging related disease – the important point is not the types of molecules or where they are, but the observation that all of them – every molecule we’ve mentioned – is in continual flux. All of the thousands of different types of molecules are being actively recycled. This is true whether we look at lipids or proteins, organelles or cytoplasm, intracellular molecules or extracellular molecules, molecules of a single type or compound molecules. They all being actively recycled. To be specific, none of them sit around for your lifetime, but all of them are being replaced on a moment-to-moment basis. With the sole exception of DNA, none of the molecules repaired. Instead, they’re simply recycled.

Some of these molecules are recycled slowly, some are recycled quickly. In the case of aerobic enzymes in your mitochondria, where the damage rate is high, these molecules are turned over rapidly; in the case of come cholesterol molecules, where the damage rate is lower, the molecules are turned over more slowly. While you might guess that “damaged” proteins are tagged and turned over more quickly, the reality is that ALL of your proteins – even those that are 100% perfect – are continually recycled. This turnover, proteolysis, is not just a passive “recycling” but is actively regulated and fine-tuned, and is part of the cell cycle and cell division, gene transcription, and general cellular quality control. Where we once viewed proteins a stable molecular pools that were subject only to “wear and tear”, active molecular turnover has been proven by isotopic studies. There is a basal rate of molecular turnover, specific to each protein and each lipid, which occurs regardless of damage: in any given molecular pool, molecules are degraded and replaced whether the molecule is normal or not. However, the rate of degradation can go up or down, depending on the rate of damage. For example, ubiquitin conjugation to globin molecules is markedly enhanced by denaturation of hemoglobin, so although hemoglobin undergoes “recycling” regardless of damage, the rate of that “recycling” goes up in the case of molecular damage.

Not only does this permit fine control of cell functions, but it is the only way to ensure quality control as well: the faster molecules are turned over, the more likely the molecules are to be undamaged and capable of doing their jobs. As we saw in the last blog, the slower the turnover, the higher the percentage of dysfunctional molecules. If we think of this recycling process as cell maintenance, then the slower the maintenance, the less functional the cell as it becomes clogged with molecules that don’t work.

Proteins, lipids and other molecules are turning over continuously and extensively. The turnover of each individual type of molecule is specifically regulated and varies with cell conditions and over time. The regulation of cell processes is not merely controlled at the transcriptional or translational levels, but is finely regulated at the level of protein degradation as well.

How fast do these molecules turnover? Proteins have half-lives varying from a few minutes to several days. The rate of turnover varies depending upon the protein, available nutrients, hormone levels, and especially by cell aging.

But if molecular recycling requires metabolic energy, then why does the cell engage in molecular turnover at all? They answer is to avoid the accumulation of damaged and dysfunctional molecules. It’s much like asking why a home owner spends money on the upkeep of their house. Both the home owner and the cell must spend (money or energy) in order to maintain function. The more they spend, the higher the quality of the house on a day-to-day basis. The less they spend, the more likely the house (or the cell) is to fall apart.

The key observation, from the standpoint of aging and age-related disease, is that almost every molecule we look at shows a deceleration of turnover as cells age. Lipids, proteins, and other molecules sit around longer. The result is leaker membranes, less effective DNA repair, dysfunctional mitochondria, and a host of other gradually increasing failures in the aging cell.

 

Next time: 2.6 Cell Senescence, Changes In Molecular Turnover, DNA Repair

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