The first assumption to be examined is that only the stem cells are the target cells for the carcinogenic process. In other words, those cells which are near or terminally-differentiated will not give rise to cancer. As a corollary, it assumed that the stem cell pool in different organs varies with not only age, but the organ itself due to a variety of biological reasons. In order to illustrate this point, it will be further assumed that there are three major types of organ systems: “closed,” “open” and “mixed.” “Closed organ” systems refer to organs, such as the liver or the kidney, where the stem cells are tightly controlled by the differentiated daughter cells in order to maintain a constant volume. On the other hand, in “open organ” systems, such as the skin and lining of the G.I.tract, the stem cells are constantly dividing because the differentiated cells are constantly being lost. In organs, such as the lung or testis, while the volume size is maintained, stem cells are constantly dividing to replace those being lost. This classification system might have relevance to those risk factors interacting with the carcinogen, such as radiation, which might affect cell proliferation. They should affect “closed organ” systems more than “open organ” systems, since without a source of mitogenic stimulation (either endogenous growth factors or hormones or exogenous chemicals or cell death and removal), the initiated stem cell would be held in check by the negative growth regulators of the differentiated cells.
Going back to the assumption that the stem cell pool might vary in certain tissues, it would be predicted that the stem cell pool might decrease in “closed organ” systems with age/developmental/pregnancy status. The number of stem cells in “open organ” systems might be expected to remain relatively constant. On the other hand, the stem cell pool might be expected to decrease with age in closed systems. This might explain, in large part, “age at the time of exposure” effects, as has been noted with the relationship between breast cancer and pregnancy status. Pregnancy would be expected to deplete the stem cell pool to produce differentiated milk-producing cells. Therefore, there would be fewer target cells for the initiation of the carcinogenic process with earlier and more frequent pregnancies. Women, who accrue initiated stem cells in their breast tissues with age but do not get pregnant, have the chance of having these initiated stem cells promoted every month by the hormones associated with the menstrual cycle.
Biological observations related to the role of stem cells in carcinogenesis include the fact that dividing cells seem to be more susceptible to the cytotoxic and mutagenic action of carcinogen initiators (Rossman and Klein, 1988). In addition, cell division seems to be imperative for the promoting phase of carcinogenesis (Bout well, 1974). While recently much attention has been given to the role of mitogenesis in carcinogenesis (Ames and Gold, 1990a), it should be pointed out that in the cases where apparent mitogenesis of cells in a given organ (or the lack thereof) has not been correlated with carcinogenesis, it must be shown that the initiated stem cell behaves identically to the normal stem cell. If initiated stem cells respond to a mitogen differently than the normal stem cell, one cannot simply use mitogenesis of tissues as a “biomarker” (Ledda-Columbano et al., 1989). Evidence for this might be from the observation that Phenobarbital treatment of initiated rat livers does not lead to general enlargement or overall proliferation of hepatocytes, but does lead to the clonal expansion of the initiated cell into a large focus of cells. In other words, Phenobarbital appears to stimulate, selectively, the initiated cell to proliferate (Kaufman et al., 1988).
An additional assumption related to the carcinogenic process is that the process involves multi-steps and multi-mechanisms. In brief, the initiation, promotion, progression concept, while recently criticized (Ames and Gold, 1990a; Ames, 1991), is applicable to most carcinogenic processes. The one exception might be the formation of teratomas. Even here, this cancer could be explained by a non-initiation/promotion/progression hypothesis without having to resort to the rejection of this major concept. First, one has to realize that “initiation,” and “progression” is operational concepts, derived from whole animal studies, not from experiments involving cells in culture or molecular events in the test tube. No implied mechanisms are intrinsic to these terms. While the observations leading to these concepts were derived from experimental rodent studies and while, in real life, neither rodents or human beings are subjected to those conditions that allow one to clearly observe distinet events taking place in the experimental conditions, it is further assumed the concept is applicable to the real life human situation, In the experimental condition, it appears to be a fact that initiation is the result of an irreversible event. Since, by definition, mutagenesis is an irreversible event (while some types of mutations can be back mutated, for all practical purposes, in this situation it would be highly unlikely). In addition, true mutagens, such as UV light, appear to be good initiators (Fry et al., 1982). On the other hand, if a critical gene is stably altered by an epigenetic event (e.g., a gene is stably turned on or off by, let’s say, an ethylating or de-ethylating event), then it would be possible for an initiating event not to involve a mutagenic event. In the case of teratomas, if a critical oncogene is de-repressed in the developing embryo, the cell might grow as a tumor with the microenvironment providing the promoting stimulus. When the microenvironment changes due to the growth, some of the cell could terminally differentiate giving rise to many of the disorganized tissues found in the teratomas.
From whole animal studies, it appears that the promotion phase is an interrupt able process (Bout well, 1974; Pitot et al., 1981). While the underlying mechanism is still unknown, it is assumed that if initiation involves the irreversible alterations of a single stem cell, then promotion is that process which allows the colnal expansion of that cell. Several additional assumptions have to be made. First, the gene (s) altered in the initiated stem cell must prevent terminal differentiation of that stem cell. If the initiated cells are stem cells which cannot terminally differentiate, then any mitogenic stimuli (i.e., promotion) would allow these cells to accumulate as “non-terminally” differentiated cells in a tissue, whereas the normal stem cells, after proliferation, would terminally differentiate. The more these undifferentiating cells are forced to proliferate, the larger this focus of cells becomes in the tissue. In the skin, as normal cells differentiate, they eventually die and are removed. Only the undifferentiated initiated cells would remain and accumulate as a papilloma. In the closed organ, such as the liver, the assumption is that the stem cell, perhaps the oval cell (Fausto, 1990; Sell, 1990; Sirica et al., 1990), will selectively proliferate after a mitogenic or promoting stimuli accumulating as a focus of incompletely terminally differentiated hepatocytes. A great deal of experimental evidence, which led to the “oncogeny as partially blocked ontogeny” hypothesis (Potter, 1981b) and in vitrol in vivo studies (Miller et al., 1987; Yuspa and Morgan, 1981; Yuspa et al., 1985), seems to be consistent with these assumptions. While it is generally assumed that promotion involves mitogenesis, some have argued that not all mitogens are promoters and not all promoters are mitogens. However, one has to keep this clearly in mind and to be critical of the so-called exceptions. In the first place, if the tumor, consisting of billions of clonally derived cells results from some “promoting” treatment, then it should be obvious that, while all cells might not have divided during the promoting treatment, the initiated cells did. For how else could a single initiated cell become a tumor of billions of cells? In the second place. It has to be shown that a “non-promoting” mitogen actually can sustain mitogenesis for a long period of time and can actuallyu promote the initiated cell. For if it can only trigger mitogenesis for a short period of time or that it preferentially acts as a mitogen only for the normal stem cell and not the initiated cell, then these superficial observations do not constitute evidence against promoters as mitogens.