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O P E N A C C E S S S O U R C E : Trends in Molecular Medicine @ Cell
Highlights
- Conversion of somatic cells to induced pluripotent stem cells and reset of the germline age with each generation represent two examples of rejuvenation.
- We propose a model of ‘ground zero,’ the mid‐embryonic state characterized by the lowest biological age at which both organismal life and aging begin.
- We propose that the zygote–ground zero period is associated with rejuvenation, wherein the biological age is decreased, telomeres are extended, and molecular damage is cleared.
- Ground zero of aging and organismal life may be related to the phylotypic period in the evolutionary hourglass model.
Cells may naturally proceed or be forced to transition to a state with a radically lower biological age, that is, be rejuvenated. Examples are the conversion of somatic cells to induced pluripotent stem cells and rejuvenation of the germline with each generation. We posit that these processes converge to the same ‘ground zero’, the mid-embryonic state characterized by the lowest biological age where both organismal life and aging begin. It may also be related to the phylotypic state. The ground zero model clarifies the relationship between aging, development, rejuvenation, and de-differentiation, which are distinct throughout life. By extending the rejuvenation phase during early embryogenesis and editing the genome, it may be possible to achieve the biological age at the ground zero lower than that achieved naturally.
The Advent of Rejuvenation
It has recently been realized that cells can be rejuvenated; that is, they can naturally proceed to or be experimentally induced to transition to the states characterized by lower biological ages than their original states [1,2]. This understanding has the potential to transform what we know about the aging process and life itself. It is known that aging is malleable because its long-term course may be adjusted by numerous genetic and environmental interventions, which can decelerate or accelerate the aging process. In addition, various studies established the regulation of organismal biological age by metabolic manipulation, senescent cell ablation, immune interventions, and other approaches (i.e., by applying the strategies which target systems that globally affect the organismal state) [1] (Figure 1). Yet, all of these approaches slow down aging or affect the biological age of the tested organ systems (e.g., as assessed by aging hallmarks [3] or biomarkers [4]) only marginally, and their effects on other organ systems are typically not assessed. Moreover, for most of these approaches, the long-term effects on healthspan (see Glossary) and lifespan remain unknown. Thus, although these approaches are extremely important and may somewhat decrease the biological age, they may only represent fragmentary or partial rejuvenation strategies.
Figure 1. Partial and Complete Rejuvenation.
At advanced ages, aging is associated with several hallmarks (shown by seven circles), which may be targeted by certain candidate interventions (shown next to each circle), thereby slightly reducing the biological age of an organism. However, because these approaches affect only some pathways, cells, or organ systems without affecting others, they may lead to only fragmentary or partial rejuvenation. By contrast, complete rejuvenation (shown by a large circle in the middle of the figure) should reset the age of all cells and systems, such that the organism becomes essentially indistinguishable from the same organism in the much younger state. Abbreviation: mTOR, mammalian target of rapamycin.
On the other hand, complete rejuvenation may be defined by an exhaustive reset of every age-related feature of a cell or organism, so that they become essentially indistinguishable from those of younger cells or organisms. This age reset represents the transition that is conceptually (but not necessarily mechanistically) opposite to the aging process and is not limited to the reversal of a single or a few parameters (e.g., protein activity, metabolite level, or gene expression) or aging hallmarks (e.g., DNA damage, epigenetic alterations, telomere attrition, protein aggregation, or accumulation of aberrant mitochondria). If a parameter, molecule, or cell state changes with age, this change may lead to a multiplicity of possible effects: (i) damaging when the change results in a by-product of metabolism, mutation, or a post-translational modification that alters protein function; (ii) protective when the change leads to an increased expression of stress response system components, such as damage repair; or (iii) neutral when the change bears no functional consequences [5]. Therefore, it is difficult to infer the functional impacts of age-related changes merely from the fact that they occur. On the other hand, a complete rejuvenation is a biological process of turning back time that involves the reset of all age-related changes. Rejuvenation is also the process that distinguishes living organisms from inanimate objects; for example, mechanical systems such as cars and gadgets irreversibly age, whereas life continually renews itself with each generation.
In this opinion piece, we propose the ground zero model of aging. We first build the model, which is based mainly on in vitro data as applied to organismal aging and rejuvenation, and then, on this backdrop, we discuss the relationships between development, aging, and rejuvenation. We further consider the origin of this ground zero and its relationship to other embryonic models and finally discuss rejuvenation strategies to reset or lower the ground zero state.
Aging and Rejuvenation of Germline and Somatic Cells
It is often discussed that, because the germline is immortal, it does not age [6,7]; this notion dates to the 19th century, when August Weismann proposed the separation of ageless germline and aging soma. However, at the time of conception, the contributing human germline has typically been maintained in a metabolically active state for two or more decades and must have accumulated damage, such as metabolic by-products, epimutations, and modified irreplaceable proteins. In other words, it has become biologically older than its earlier, embryonic state. Although the germline biological age at the time of conception is expected to be much younger than that of somatic tissues of the same organism, and although some of the accumulated damage may be removed by designated molecular systems, rejuvenation in the prezygotic state could only be partial because, in the absence of cell division (as in the oocyte), there are always more damage forms than the means of protecting against them [8]. Also, although some germ cells may accumulate more damage than others and therefore may lead to early mortality and abnormalities in the offspring (this damage will also increase with the age of the host), all germ cells unavoidably accumulate some damage. Thus, for the new life to begin in the same young state as in the previous generation, the zygote must somehow remove this damage and decrease its biological age to the level of the germline age in the previous generation. In other words, it appears that the germline ages during development and adult life, and then it is rejuvenated in the offspring after conception.
The complete rejuvenation conceptualized above (i.e. the transition from a state characterized by a higher biological age to the state with lower biological age) is not limited to early embryogenesis. Takahashi and Yamanaka’s groundbreaking discovery of induced pluripotent stem cells (iPSCs) [9] has made it clear that somatic cells may also be rejuvenated. The conversion of somatic cells to iPSCs, which corresponds to the state of embryonic stem (ES) cells, is accompanied by incremental cell heterogeneity, with many cells in the population acquiring different cell states and some becoming rejuvenated [6]. Mechanistic details of somatic rejuvenation are not fully clear; for example, it is not known how damage is removed during this process, if it is a gradual and coordinated process, and if different damage forms are cleared according to their own temporal trajectories.
A Model of the Ground Zero of Organismal Life
By convention, the age of a person is counted from the day he/she is born. But when does his/her aging begin? To consider this question, one needs to ask another question: When does organismal life begin? There are several common answers: conception, first neural activity, first heartbeat, first breath, or simply birth. However, the beginning of organismal (as opposed to cellular) life at conception contradicts the observations that an early embryo (i) may be naturally or experimentally split, generating two (i.e., twins) or more organisms [10]; (ii) can be combined with other embryos of the same species [11] and even with ES cells/iPSCs of other species [12] generating chimeric organisms; (iii) initially relies on maternal gene products rather than its own [13]; (iv) gradually extends telomeres (from early cleavage to blastocyst through a recombination-based mechanism and subsequently using telomerase) [14]; (v) gradually removes epigenetic marks [15]; (vi) decreases structural entropy [16]; (vii) gradually inactivates chromosome X and develops biased paternal/material monoallelic gene expression [17]; (viii) is unable to distinguish self- from non-self (its acquired immune system is formed later) [18]; and (ix) may become another individual by swapping its genetic material via somatic cell nuclear transfer. Together, this suggests gradual rejuvenation during early embryogenesis as opposed to the aging of the newly formed organism starting from zygote or early cleavage. However, this rejuvenation process is reversed at some point during development, wherein telomeres again begin to shorten, self-recognition is established, biological age starts to increase, and so forth [13., 14., 15., 16., 17., 18.]. It is unclear if all these changes during early embryogenesis and their subsequent reversal closer to mid-embryogenesis occur simultaneously, gradually, or in waves or whether each follows its own temporal trajectory, because highly resolved measurements are currently lacking for many of them. However, the direction of changes is already clear from the current data.
All this leads to a model wherein early embryos are gradually rejuvenated, for example, by extending their telomeres, erasing epigenetic marks and clearing up and diluting molecular damage, and this continues up to a particular time during early development. Conception represents a starting point for this process, culminating in the state of the lowest biological age, the ground zero of organismal life and aging (Figure 2). In effect, the period from conception to this stage may be viewed as a preparatory stage, which is associated with damage clearance and rejuvenation, for subsequent development of the organism. This suggests that organismal life begins after this preparatory stage and is associated with the formation of the body plan, immune system, neural activity, and so forth. With regard to the beginning of life, this argument clearly distinguishes conception (beginning of cellular life) from ground zero (beginning of organismal life).
Figure 2. The Ground Zero Model of Organismal Life and Aging.
(A) Molecular features associated with aging are U-shaped. During early embryogenesis, telomeres are extended and entropy is decreased, suggesting a transition from an older to a younger state, but after the inflection point, defined as the ground zero state (or period), their trajectories are reversed. The actual changes of features associated with aging during development may be more complex (e.g., may be asynchronized or in waves). (B) Ground state as the beginning of organismal life and aging. The biological age of an organism decreases in early embryogenesis and increases starting at mid-embryogenesis. Ground zero corresponds to the lowest biological age of an organism. In this model, the zygote defines the beginning of cellular life, and ground zero represents the beginning of organismal life. © The beginning of aging at mid-embryogenesis suggests approaches to achieve a lower biological age than that naturally possible. In the ground zero model, the period from the zygote to ground zero represents rejuvenation, and aging begins at ground zero. Extending or enhancing the rejuvenation period (as well as reducing the load of damaging mutations) may lead to a lower biological age (green) than naturally achieved (purple) at ground zero and therefore a reduced biological age throughout life, leading to an extended lifespan and healthspan.
One obvious exception to this principle (rejuvenation during early embryogenesis until a certain point during development) is the organismal genome. In contrast to the gradual acquisition of younger features during early embryogenesis, the genome is formed at conception and cannot be rejuvenated (i.e., mutations are irreversible). Instead, the genome is ‘rejuvenated’ at the level of species; that is, the germline acquires mutations during the life of an organism, but after conception, the most deleterious genotypes are eliminated due to early-life mortality and decreased fitness during adult life [19]. In other words, purifying selection supports mutation–selection balance and may be viewed as rejuvenation of the genome at the level of species. Advances in genome editing [20] provide additional opportunities for rejuvenation of the genome by removing damaging mutations, which may be used when these tools further improve with regard to off-target effects [21].
Starting from the ground zero state, to begin a new life cycle, some cells of the organism need to (i) erase age-related (and also cell type–specific) somatic epigenetic patterns through a cell reprogramming process, generating primordial germ cells; (ii) establish sex-specific epigenetic patterns in these cells, which subsequently enable meiotic maturation and fertilization; (iii) remove these epigenetic patterns after fertilization, jumpstarting the developmental program [22]; and (iv) establish ground zero epigenetic patterns and beginning of a new life cycle. The specific solutions for these innovations differ among animals, whereas mid-embryogenesis, the timing that generally corresponds to the ground zero state is considered the most conserved state that characterizes metazoans that age and even other eukaryotes [23]. It is known as the phylotypic state (Box 1).
Box 1.
Relationship between Ground Zero, Phylotypic State, Entropic Minimum, and the Ancestral State of Animals
The notion of ground zero reverberates with other embryonic models. At the level of gametes, zygotes, and early embryos, various vertebrates look very different, but during development, they acquire a common state (phylotypic state) wherein they can be virtually indistinguishable from one another, and then they diverge again. In this hourglass model, the phylotypic state is defined by the expression of evolutionarily old genes and constrained variance in gene expression [41,42]. It is possible that the phylotypic state matches the ground zero state. Likewise, some metrics of entropy, such as structural entropy, may be lowest during mid-embryogenesis, and this might correspond to the lowest biological age. The period approximately related to the phylotypic state may also be associated with the loss of regeneration and the embryo–fetal transition [43]. Interestingly, mortality is also highest during early embryogenesis, wherein the species genome is rejuvenated by eliminating embryos inviable due to a combination of damaging alleles, which would be consistent with the use of genes with higher selection coefficients during this stage. Overall, all these milestones in embryonic life may converge to the same state, corresponding to the beginning of organismal life. This state may correspond to the ancestral state, perhaps the ancestor of all animals, with innovations supported by new genes and functions extending to both before and after this state.
Relationship between Aging, Rejuvenation, and Development
The ground zero model has several implications, and the one that should be addressed first is how aging, rejuvenation, and development are related to one another.
Aging and Rejuvenation
The relationship between aging and rejuvenation is straightforward: they are essentially opposite to each other; the former makes an organism older, whereas the latter makes it younger. They are naturally linked by ground zero, the timing when early-life rejuvenation ends and aging begins (Figure 2), but, experimentally (e.g., artificially in a test tube), they may be induced at other life stages. Thus, somatic cells may be converted to iPSCs and thus be rejuvenated, whereas unfavorable environmental conditions (high oxygen, lack of nutrients, DNA damage) may lead to aging of the cells, which otherwise would not age. Mechanistically, aging and rejuvenation do not seem to proceed through the same trajectory in the opposite direction. The same mechanistic trajectory of these processes was famously conceived by Fitzgerald to describe the reverse aging of Benjamin Button [24], but this is not observed in real life.
Development and Rejuvenation
Rejuvenation is also different from development. Development is a genetic program that begins at conception and ends roughly at age 20 (although some developmental processes are completed earlier and some later), and its aim is to build a fit organism. Even during the period from conception to ground zero, rejuvenation is different from development because its essence is to remove damage and decrease the biological age rather than build an organism. After ground zero, development is not opposite to rejuvenation, because disassembling what was built is not the same as decreasing the biological age.
Development and Aging
Development and aging differ in timing and purpose (although achieving a specific milestone during development may require a certain biological age of the cells). The former is a genetically programmed process (i.e., there are genes whose purpose is to support development) that begins at conception and ends with a functional adult organism. By contrast, aging begins at ground zero and continues until an organism dies. Also, aging likely has no purpose, and there are no genes that evolved with the sole purpose to cause aging. Aging is a consequence of being alive, a by-product of metabolism that involves accumulation of age-related changes.
Measuring Biological Age
Another issue intimately related to progression through aging and rejuvenation is the concept of biological age. Biological age is an integrative measure of deleterious changes that occur during organismal life [8]. When we look at a person, we can estimate his/her age rather accurately. But some people age slightly faster than the average person, and some age slower. There are now molecular tools available to assess the biological age, most notably epigenetic clocks. The first such clocks were developed for humans [4,25,26] and more recently for mice [27., 28., 29., 30., 31., 32.]. Other clocks are based on gene expression, metabolite patterns, and other features of cells and organisms. The transitions through aging and rejuvenation can be tracked by these biomarkers, and early embryogenesis is not an exception (Figure 3). In fact, it was shown that the epigenetic clock developed for human adults [4] tracks the aging process as early as 45 days after conception [33]. It is important to note that the basis for aging clock applications to study early embryogenesis lies in the method researchers use to build it. By performing regression toward the broad range of chronological ages as well as across tissues, the clock readout tracks the aging process. More specifically, the DNA methylation clock quantifies the levels of errors in the DNA methylome. In reproduction and early embryogenesis, these epigenetic errors, along with other errors, are sufficiently decreased to ensure the normal lifespan of the next generation. Ground zero is the time when an embryo is expected to feature the most ‘perfect’ epigenome naturally accessible, which can be quantified by the clock as the youngest epigenetic age. Therefore, these assays, preferably in combination with other assays, may be used to define the exact timing of ground zero, which is currently unclear. It probably lies somewhere between blastocyst (as its inner cell mass may be divided without aging) and pharyngula (the most conserved state of vertebrate development). It may also be related to gastrulation; as famously stated by Lewis Wolpert, ‘It is not birth, marriage, or death, but gastrulation which is truly the most important time in your life’ [34].
Figure 3. Defining Ground Zero by Following the Biological Age Using DNA Methylation Clocks.
It is also possible that some processes are rejuvenated earlier during development, some later, and some in multiple bouts. This asynchronization can be illustrated by changes in DNA methylation, which is remodeled differently for paternal and maternal DNA during cleavage and is remodeled again later during embryonic development [15]. Likewise, it may be that some processes are rejuvenated prior to fertilization, at the onset of zygotic transcription, in the blastula, or at onset of neurulation. In this sense, the ground zero state may be viewed as the ground zero period. The timing of ground zero will also depend on methods used to assess age reversal of various cellular components. Perhaps some integrative, entropy-like measure is needed to determine the lowest biological age during development.
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