**4. The adaptation**

probabilistic, depending on the distance between genes (Russell, 1998), and natural selection is directional since it follows a direction which favors the survival of the fittest organism.

changes in some organisms subjected to special physiological requirements.

The picture shows the estimated geological age of the last common ancestor of each pair of specified animals. Each time estimate is based on comparisons of amino acid sequences of orthologous proteins; more time had a pair of ani‐ mals to evolve independently, smaller is the percentage of amino acids remaining identical. The final estimates and the time scale has been calibrated to match the fossil evidence that the last common ancestor of mammals and birds lived 310 million years ago. Numbers in the top bar gives data on sequence divergence for a particular protein chosen arbitrarily, i.e the α-chain of hemoglobin. The clear irregularities in growing divergence with increasing time reflect the randomness of the evolutionary process and, probably, the action of natural selection, which drives particularly rapid

Both mutation and recombination define the *allelic variability*, which determines a difference between the genotypes of individuals; this variability will be further shaped by the environ‐ ment, that ultimately determine the presence of different *phenotypes*, based on instructions dictated by the *genotype* (Russell, 1998). Mutations, as source of variation, are quite limited since their rates are very low. On the other side, recombination is a primary source of variation; most of the attention of evolutionary geneticists has focused on the extensive *genetic recombi‐ nation* that takes place during meiosis and, in particular, during pairing of sisters chromatides and its significance as a generator of genetic diversity in organisms with sexual reproduction

In organisms with asexual reproduction, such as bacteria, recombination is very limited since mutations are the only source of gene combinations, thus, asexual organisms may evolve more slowly, under natural selection, than sexual ones (Griffiths et al., 1999). Although the mutations play a more limited role as a source of variability in multicellular organisms with sexual reproduction, together with the effects of gene recombination, they can still be transmitted to

In multicellular eukaryotes these cells are the connection between the different generations since they are the only ones that undergo *meiosis*, as well as *mitosis*, in contrast to *somatic cells*,

(Charlesworth, 1988; Edwards, 2000).

**Figure 5.** Phylogenetic proximity

652 Regenerative Medicine and Tissue Engineering

offspring only by the *germ cells*.

It is impossible to speak of life in a biological sense without postulating a *certain degree of independence* from the environment where life itself takes place. Whatever its level of com‐ plexity, a living organism is a *biological system* delimited by a kind of boundary to face the *external environment* and capable to maintain a stable, constant condition by regulating himself and the *internal environment*.

The demarcation of self with a *membrane* and the establishment of an *internal environment*, opposed to the external one, is a formal requirement to any living organism and the condition of *relative equilibrium* called *homeostasis* (Cannon, 1929) provides the biological system an appropriate level of independence from the environment that allows to acquire other essential properties such as response to stimuli, development, growth and reproduction.

Any change, eitherdecremental orincremental, in the internal/external environment ofthe cell/ organism such that it requires an active response from the cell/organism can be termed de‐ mand (Ciulla et al., 2011). By using the simplified approach *demand-response*, it is possible to classify the demand as a function of its *type*, *intensity* and *duration*. Since the possible responses of the cell/organism are limited and fall within the *adaptive processes*, the demands can be further classifiedincurrent or extra according to the efficiencywith whichthey canbehandled.Current demands can be adequately dealt with in a physiological context, even though some degree of cell/organism injury can still occur; extra demands cannot be sufficiently buffered, and lead to functional impairment and, eventually, to disease or, as the last resort, to death. Anyhow the final outcome of the changes imposed on living organisms depends on the *ratio demandresponse*; anexcess indemandand/or adefectinthe response forthe lackordepletionof adaptive resources, can represent a serious problem for the survival of the organism. In such view, *disease* is the result of an *unfavorable interaction* between the cell/organism and the environment or, by simplifying, between the genetic resources and the environment. The succession of cyclical and stochastic environmental changes suggests that, during the life cycle, the current demands represent the bulk of all demands while extra demands are more often occasional. The relation between current and extra demand can be depicted by using the iceberg metaphor (Figure 6). Thus, the *possibility of damages* and *disease* caused by *unfavorable interactions* with the environ‐ ment are foreseen and their occurrence is managed by coping biological resources involved in tissue maintenance and repair of damages.

Representation of the iceberg metaphor, illustrating the boundary between health as a result of individual adaptabili‐ *ty* and disease. The tip of the iceberg corresponds to overt disease; the huge part below the water line is where indi‐ vidual adaptability successfully buffers environmental demands; just below the surface is the grey zone of subclinical disease. Demands are defined as current or extra according to how efficiently they can be handled by the single or‐ ganism.

Modified from an Open Access source: http://www.intechopen.com/books/advances-in-regenerative-medicine/ inflammation-angiogenesis-cross-talk-and-endothelial-progenitor-cells-a-crucial-axis-in-regenerating.

**Figure 6.** The iceberg metaphor to understand diseases

Therefore, as changes in the internal/external environment occur continuously, it is evident that *allocating biological resources* in order to buffer changes is an essential requirement for life and living organisms have, indeed, evolved specific *adaptive processes* to meet the demands imposed by changes in the environment. In this context, it is therefore not superfluous to remind that the *availability of resources* is only possible if there is a corresponding *genetic resource*. The *genetic program* that governs *genetic resources* is a *finite sequence of instructions* whose possible combinations are somehow limited by the same program. Furthermore, especially in complex organisms that require *stages of development* to reach a final *adult form* and a *full functionality*, genetic resources are not only quantitatively limited but also *temporally regulat‐ ed* by the constraints of the life cycle. Thus in the same manner will also be limited the *adaptive processes*, including *organism adaptation* or *plasticity* and *adaptability* (Kutschera, 2009). The *plasticity* is the ability to express a broad variety of phenotypes in response to environmental changes and, in complex organisms with sexual reproduction, is at a maximum during *embryogenesis* and *early extrauterine life*. The *adaptability* could instead be applied to the more limited process of adaptation which occurs in adult life and varies between organs and species (Figura 7).

of the cell/organism are limited and fall within the *adaptive processes*, the demands can be further classifiedincurrent or extra accordingto the efficiencywith whichtheycanbehandled.Current demands can be adequately dealt with in a physiological context, even though some degree of cell/organism injury can still occur; extra demands cannot be sufficiently buffered, and lead to functional impairment and, eventually, to disease or, as the last resort, to death. Anyhow the final outcome of the changes imposed on living organisms depends on the *ratio demandresponse*; anexcess indemandand/or adefectinthe response forthe lackordepletionof adaptive resources, can represent a serious problem for the survival of the organism. In such view, *disease* is the result of an *unfavorable interaction* between the cell/organism and the environment or, by simplifying, between the genetic resources and the environment. The succession of cyclical and stochastic environmental changes suggests that, during the life cycle, the current demands represent the bulk of all demands while extra demands are more often occasional. The relation between current and extra demand can be depicted by using the iceberg metaphor (Figure 6). Thus, the *possibility of damages* and *disease* caused by *unfavorable interactions* with the environ‐ ment are foreseen and their occurrence is managed by coping biological resources involved in

Representation of the iceberg metaphor, illustrating the boundary between health as a result of individual adaptabili‐ *ty* and disease. The tip of the iceberg corresponds to overt disease; the huge part below the water line is where indi‐ vidual adaptability successfully buffers environmental demands; just below the surface is the grey zone of subclinical disease. Demands are defined as current or extra according to how efficiently they can be handled by the single or‐

Modified from an Open Access source: http://www.intechopen.com/books/advances-in-regenerative-medicine/

Therefore, as changes in the internal/external environment occur continuously, it is evident that *allocating biological resources* in order to buffer changes is an essential requirement for life and living organisms have, indeed, evolved specific *adaptive processes* to meet the demands imposed by changes in the environment. In this context, it is therefore not superfluous to

inflammation-angiogenesis-cross-talk-and-endothelial-progenitor-cells-a-crucial-axis-in-regenerating.

tissue maintenance and repair of damages.

654 Regenerative Medicine and Tissue Engineering

**Figure 6.** The iceberg metaphor to understand diseases

ganism.

The lower panel shows the environmental fluctuations as result of temperature changes (street light colors) during the history of Earth. The upper panel shows how genetic resources respond accordingly to these fluctuations by tun‐ ing adaptability and plasticity. The bars depict which resource is mainly involved in each time points.

But why are these processes limited in adult life? In this regard, it should be remembered that in complex organism the evolution towards multicellularity is a form of adaptation to the environment that involves a high cost in terms of biological resources; in other words,

**Figure 7.** Relationship between plasticity and adaptability in a changing environment

*differentiation, specialization*, *establishment of functional hierarchies* and development of a *fully developed complex organisms* implies a considerable expense of biological resources, thus reducing the availability of resources for other forms of adaptation in adult life such as the *regeneration* process.

Therefore in the presence of an *extra demand* causing a damage the response of complex organisms in adult life mainly consists in *functional* and *structural adaptation* since *re-genera‐ tion* processes after *injury* are limited. Indeed in such organisms, *generation* and *re-generation* are capabilities shared by the same deputy cells named *stem cells*, the only ones capable of differentiating into all cell types that make up a multicellular organism (Paragraph 2). The number of stem cells is limited and, as we saw earlier, their *potency* is maximal during the prenatal life in the *embryonic stem cells*, and progressively decreases in post-natal life remaining confined within the *adult stem cells*.

These cells are therefore to be understood as a kind of *functional reserve*, restricted to a *specific tissue*, that could be recruited to support adaptation processes that allow the organism to better fit in with the changed environment and, thus, attain a new equilibrium. At this regard the *functional hierarchy* of complex organisms allocates *functional reserve* where it is needed and establishes the *distribution of the available resources*. In complex organisms such as mammals, many adult tissues contain populations of adult stem cells that have the capacity for *renewal after disease* or *aging*; these tissues include brain, bone marrow, peripheral blood, blood vessels, skeletal muscle, skin, teeth, heart, gut, liver, ovarian epithelium, and testis. Thus, the primary role of adult stem cells in a living organism is to *maintain* and *repair* the tissue in which they are found. Unfortunately there is a very small number of adult stem cells in each tissue, with large numerical differences between a tissue and another, and, therefore, the *re-generative potential* is unevenly distributed and, in any case, is very limited. The reason why some *highly specialized tissues* have limited regenerative capacity is not yet known, but we can not exclude that the extreme structural and functional specialization reached by some tissues is an inherent limit to regeneration (Table 1).

Recently a number of experiments have reported that certain adult stem cell types can differentiate into cell types seen in organs or tissues other than those expected from the cells' predicted origin or *lineage*; for example, brain stem cells that differentiate into blood cells or blood-forming cells that differentiate into cardiac muscle cells, suggesting the idea that cells might have an *alternative fate* after maturation according to their specific identity. The two known processes by which cells are able to turn into other cell types are: *trans-differentiation*, consisting in the direct conversion from one cell type to another, and *de-differentiation*, or the reversion to a less-differentiated cell type and the subsequent maturation to a different lineage. Thus the *cell identity*, which had so far been considered a rigid and durable characteristic involving a one-way process from precursor to mature cell, was shown to exhibit not only intrinsic plasticity (Scadden, 2007) but also a certain degree of adaptation depending on the interplay between genome and microenvironment. In fact it was demonstrated that mature cells are able to switch not only their functional phenotype but also their gene expression profile into that of stem cells, thereby acquiring pluripotent plasticity (Ciulla et al., 2011). The

\* Hayflick demonstrated in 1965 a limited number of cell division (Hayflick, 1965)

*differentiation, specialization*, *establishment of functional hierarchies* and development of a *fully developed complex organisms* implies a considerable expense of biological resources, thus reducing the availability of resources for other forms of adaptation in adult life such as the *re-*

Therefore in the presence of an *extra demand* causing a damage the response of complex organisms in adult life mainly consists in *functional* and *structural adaptation* since *re-genera‐ tion* processes after *injury* are limited. Indeed in such organisms, *generation* and *re-generation* are capabilities shared by the same deputy cells named *stem cells*, the only ones capable of differentiating into all cell types that make up a multicellular organism (Paragraph 2). The number of stem cells is limited and, as we saw earlier, their *potency* is maximal during the prenatal life in the *embryonic stem cells*, and progressively decreases in post-natal life remaining

These cells are therefore to be understood as a kind of *functional reserve*, restricted to a *specific tissue*, that could be recruited to support adaptation processes that allow the organism to better fit in with the changed environment and, thus, attain a new equilibrium. At this regard the *functional hierarchy* of complex organisms allocates *functional reserve* where it is needed and establishes the *distribution of the available resources*. In complex organisms such as mammals, many adult tissues contain populations of adult stem cells that have the capacity for *renewal after disease* or *aging*; these tissues include brain, bone marrow, peripheral blood, blood vessels, skeletal muscle, skin, teeth, heart, gut, liver, ovarian epithelium, and testis. Thus, the primary role of adult stem cells in a living organism is to *maintain* and *repair* the tissue in which they are found. Unfortunately there is a very small number of adult stem cells in each tissue, with large numerical differences between a tissue and another, and, therefore, the *re-generative potential* is unevenly distributed and, in any case, is very limited. The reason why some *highly specialized tissues* have limited regenerative capacity is not yet known, but we can not exclude that the extreme structural and functional specialization reached by some tissues is an inherent

Recently a number of experiments have reported that certain adult stem cell types can differentiate into cell types seen in organs or tissues other than those expected from the cells' predicted origin or *lineage*; for example, brain stem cells that differentiate into blood cells or blood-forming cells that differentiate into cardiac muscle cells, suggesting the idea that cells might have an *alternative fate* after maturation according to their specific identity. The two known processes by which cells are able to turn into other cell types are: *trans-differentiation*, consisting in the direct conversion from one cell type to another, and *de-differentiation*, or the reversion to a less-differentiated cell type and the subsequent maturation to a different lineage. Thus the *cell identity*, which had so far been considered a rigid and durable characteristic involving a one-way process from precursor to mature cell, was shown to exhibit not only intrinsic plasticity (Scadden, 2007) but also a certain degree of adaptation depending on the interplay between genome and microenvironment. In fact it was demonstrated that mature cells are able to switch not only their functional phenotype but also their gene expression profile into that of stem cells, thereby acquiring pluripotent plasticity (Ciulla et al., 2011). The

*generation* process.

656 Regenerative Medicine and Tissue Engineering

confined within the *adult stem cells*.

limit to regeneration (Table 1).

*alternative fate* has indeed opened a new window of opportunity revealing the significant opportunities that may arise from *engineering of adaptive processes* (Figure 8).

Representation of biological resources allocation by genetic program in order to buffer changes in the internal/exter‐ nal environment. All living organisms have evolved specific *adaptive processes* to meet the demands imposed by envi‐ ronment.

**Figure 8.** Allocation of biological resources for adaptation processes

In the last decade the topic of *adult stem cell repair* of the infarcted myocardium was among the most popular in the scientific community and has gained growing popularity among top scientific journals. The objective of these studies is, therefore, to optimize the adaptive processes by probing the possibility of *manipulating the cellular identity*. By suggesting possible *alternative fates* for the cells, this research model clashes with the *dogma of the cell cycle* and is faced with the problem of how to ascertain the identity of the cells in a context in which the identity itself is no longer a certainty but, rather, a dynamic concept (Ciulla et al., 2011).

Thus the issue of cell identity or phenotype led to develop alternative techniques to direct visualization of histological structures; among them the refined, however complex, techniques of *fluorescence microscopy* based on the confocal representation of *fluorochromes* visible at different wavelengths. The employment of these laboratory techniques in the belief that *methodological complexity* equates to biological soundness has produced a paradox culminated in a scientific nonsense: the same experiment, carried out by different investigators, produced discrepant findings, as illustrated by an article on Nature (Orlic et al., 2001). After this setback the isolated instances of *trans-differentiation* observed in some vertebrate species following transplantation of adult stem cells have been debated by the scientific community and the observations so far made have been explained alternatively as a result of the fusion of a donor cell with a recipient one; in addition, even when trans-differentiation has been detected, only a small percentage of cells undergo to this process. This episode, seen as a drama by the scientific community, points out that as the complexity of a study increases its informative content paradoxically deteriorates. The scientific basis for this type of reasoning can easily be found in the field of mathematics, according to Gödel's Incompleteness Theorem: *a great complexity is a source of incompleteness because it increases the likelihood that true sentences cannot* *be proved* (Calude and Jurgensen, 2005). In this regard it should be emphasized that the direct visualization of histological structures across a large number of fields coupled with immuno‐ histochemistry assay on contiguous slices in clear guarantees, in any case, a high spatial resolution (Kwok et al., 2010; Ciulla et al., 2013).

*alternative fate* has indeed opened a new window of opportunity revealing the significant

Representation of biological resources allocation by genetic program in order to buffer changes in the internal/exter‐ nal environment. All living organisms have evolved specific *adaptive processes* to meet the demands imposed by envi‐

In the last decade the topic of *adult stem cell repair* of the infarcted myocardium was among the most popular in the scientific community and has gained growing popularity among top scientific journals. The objective of these studies is, therefore, to optimize the adaptive processes by probing the possibility of *manipulating the cellular identity*. By suggesting possible *alternative fates* for the cells, this research model clashes with the *dogma of the cell cycle* and is faced with the problem of how to ascertain the identity of the cells in a context in which the identity itself is no longer a certainty but, rather, a dynamic concept (Ciulla et al., 2011).

Thus the issue of cell identity or phenotype led to develop alternative techniques to direct visualization of histological structures; among them the refined, however complex, techniques of *fluorescence microscopy* based on the confocal representation of *fluorochromes* visible at different wavelengths. The employment of these laboratory techniques in the belief that *methodological complexity* equates to biological soundness has produced a paradox culminated in a scientific nonsense: the same experiment, carried out by different investigators, produced discrepant findings, as illustrated by an article on Nature (Orlic et al., 2001). After this setback the isolated instances of *trans-differentiation* observed in some vertebrate species following transplantation of adult stem cells have been debated by the scientific community and the observations so far made have been explained alternatively as a result of the fusion of a donor cell with a recipient one; in addition, even when trans-differentiation has been detected, only a small percentage of cells undergo to this process. This episode, seen as a drama by the scientific community, points out that as the complexity of a study increases its informative content paradoxically deteriorates. The scientific basis for this type of reasoning can easily be found in the field of mathematics, according to Gödel's Incompleteness Theorem: *a great complexity is a source of incompleteness because it increases the likelihood that true sentences cannot*

**Figure 8.** Allocation of biological resources for adaptation processes

658 Regenerative Medicine and Tissue Engineering

ronment.

opportunities that may arise from *engineering of adaptive processes* (Figure 8).

Nonetheless research on adult stem cells continues to generate great enthusiasm and has led researchers and clinicians to ask whether adult stem cells could be used for transplants. In the era of biomedicine, transplantations and tissue engineering, an emerging practical issue, however, is what kind of adult stem cells should be used to optimize the adaptive processes after tissue damage. Pioneering studies have focused on the most versatile adult stem cell such as the *hematopoietic* one that, starting from a common precursor, is able to give rise to very different cell lines. It must be remembered that adult hematopoietic stem cells transplantation has been used as a medical procedure in the field of hematology and oncology since the fifties (Rebulla and Giordano, 2004).

Once collected from a donor and administered peripherally to a recipient, the ability of these hematopoietic precursors, identified as *bone marrow mononuclear cells*, to travel through the circulation and to selectively targeting an area of *experimental myocardial damage* produced by means of *cryoinjury* (Ciulla et al., 2004b) has been demonstrated in rats (Ciulla et al., 2003) (Figure 9).

Furthermore, the study of the *mechanisms of homing* of these cells, also showed that this phenomenon is proportional to the extent of the damage (Ciulla et al., 2004a) and, finally, their contribution consists mainly in giving rise to new, actually working, vessels (Ciulla et al., 2006; Ciulla et al., 2007). Another instance is that transplanted cells might have also a *paracrine effect* such as to modulate the response to injury (Ciulla et al., 2008). In the perspective of the autologous infusion, it has been shown that adult stem cells, once removed from the body, have very limited ability to divide, making generation of large quantities of stem cells difficult (Ciulla et al., 2006). Despite these limitations, with a view to improve the healing process in humans, the advantages of using adult stem cells should be remarked as they allow to avoid the ethical and political issues associated with the use of embryonic stem cells.
