The ultimate understanding of how a multicellular organism, such as the buman being, maintains its ability to regulate cell growth, proper differentiation, adaptive responses and wound-healing will necessitate adhering to the principle that an organism is “greater than the sum of its parts” (Eagle, 1965). This holistic concept, together with the hierarchical (Brody, 1973) and cybernetic (Potter, 1974) concepts, has been utilized to explain how the regulation of ions and molecules is achieved at the molecular/biochemical levels, at the cellular level and at the organ and system levels. The idea that there exists a cybernetic-like control system in multicellular organisms to regulate cell functions within and between tissues was created by Claude Bernard (1878). W.B. Cannon coined the term, homeostasis, to describe this regulatory process (1929). In addition, Weiss and Kavanan (1957) postulated the existence of growth regulators of cell proliferation, while Mazia (1961) suggested that “removal of a block” was necessary for cell division, Later, Potter (1983) integrated both the positive and negative factors into a scheme whereby, locally and systematically, they could regulate growth.
The explanation of how a constant “milieu interieur” is maintained was provided by the conceptualization of positive and negative factors, produced by cells of one type/tissue which affects distal cells of another type/tissue. It predated the discovery of most of the actual molecular entities involved in homeostasis. Eventually, this concept of homeostasis by a cybernetic process was fleshed out with the discovery of various hormones, biologically active peptides, lymphokines and neurotransmitters. Note, that oncogenes code for these kinds of molecules (and their receptors). The bilolgical process by which cells influence control over each other in the multicellular organism is referred to as intercellular communication. In general there are three forms of intercellular communication; extracellular communication would refer to how cells communicate with distal cells via the secretion of a molecular signals (e.g., hormone, peptide, growth regulator, neurotransmitter); intracellular communication would refer to those transmembrane-signalling elements triggered by the extracellular signals; and Intercellular communication would refer to the transfer of ions and small regulatory molecules from contiguous cells through a protein channel on the cell membrane, called the gap junction (Loewenstein, 1990).
This interconnected communication network allows a multicellular organism to control its fundamental cellular, tissue, organ and system functions. For example, a hormone secreted by the hypothalamus (extracellular) could trigger intracellular biochemical changes in distal cells of another organ, which in turn, could either increase or decrease intercellular communication in that tissue which has a receptor for that hormone. By increasing intercellular communication, these hormone triggered cells might now differentiate or if already differentiated, produce a substance unique to that hormone signal. Alternatively, if the biochemical changes occurring in the targeted cells cause a decrease in gap junction-mediated intercellular communication, then the cells have an opportunity to divide.
The key to this homeostatic process is an understanding of the fundamental role the gap junction plays in multicellular organisms. This channel-like structure, made of proteins which are highly conserved through evolution, is found in almost all cells of the solid tissues of metazoans (Revel, 1988). These gap junction channels convey ions and small molecular weight molecules from one cell to all others which are connected by these channels. In effect, all coupled cells are in equilibrium for these ions and small molecules. If any one cell has a momentary increase or decrease in these substances, these will soon equilibrate via these channels. These channels allow a single cell to be either a “sink” or “point source” to the other coupled cells for these ions and small molecules (Sheridan, 1987). Therefore, if critical a level of a regulatory molecule or ion is needed before a cell can proliferate of differentiate modulation of the gap junction function or number might be needed. For example, if a series of cells are coupled and only some of them receive a growth factor molecule, the intracellular changes induced by the extracellular signal would be shared by all the coupled cells, such that no one cell would have enough to start the cell division process if the gap junctions remained open (Lowenstein, 1967). On the other hand, it has been postulated that if the gap junction function is down regulated by the growth factor, these critical intracellular signals would reach a critical mass and trigger the cell division process. This appears to be the case, since it has been shown that various growth factors and hormones can down regulate gap junction function (Aylsworth et al., 1989; Maldonado et al., 1988).
Gap junction communication has also been implicated in the regulation of normal development and differentiation (Lo, 1985; Schultz, 1985). The fertilized egg and early blastula cells do not have gap junctions (Lo and Gilula, 1974). As the early embryo starts to develop and differentiate, gap junctional communication starts to appear (Lo and Gilula, 1974). Even stem or progenitor cells in various human tissues (e.g., breast, kidney) do not appear to have gap junctions (Chang et al., 1987; Chang et al., 1990). If developing embryos are treated with antibodies or anti-sense RNA directed to gap junctions. Abnormal development ensues (Bevilacqua et al., 1989; Warner et al., 1984). Several chemicals, such as dilation, retinoid and alcohol, known to be teretogens but not mutagens, have been shown to down regulate gap junctions (Trosko and Chang, 1988a).
At this point, the evidence, while circumstantial, implicates the up or down regulation of gap junctional communication in triggering quiescent cells to proliferate or proliferating cells to contact-inhibit; undifferentiated cells to differentiate; or differentiated cells to adaptively respond to environmental changes (Trosko and Chang, 1984). This up or down regulation of gap junctional communication can be adaptive or maladaptive. To date, a large number of chemicals, both natural and human-made, have been shown to modulate gap junctional intercellular communication in a variety of mammalian, including human, in vitro systems (Trosko and Chang; 1988a). Some of the chemicals are natural and normal physiological chemicals, such as epidermal growth factor and hormones (Kihara et al., 1990; Larsen, 1983; Madhukar et al., 1989; Maldonado et al., 1988). Others are nutrients, food additives, cigarette tar condensates, drugs, pesticides, pollutants, biological toxins and natural plant products (Trosko and Chang. 1988a). These chemicals have been shown to down regulate gap junctional intercellular communication reversibly, at non-cytotoxic levels, in a dose dependent fashion in both rodent and human in vitro cell systems.
Modulation of the critical cellular function can be at the transcriptional (expression of a gene). Translational (production of a gene-related protein) or at the posttranslational (modification of a synthesized protein) levels. Intracellular changes induced by endogenous or exogenous extra cellular communication molecules can modify levels of calcium, H+ ions, as well as the activation of enzymes which posttranslational modify proteins, increase cyclic AMP and generate free radicals. All of the induced intracellular changes have been associated with the regulation of gap junctions (Saez et al., 1990). In other words, physiological changes can bring about cellular changes, such as cell differentiation, cell division or adaptive cell responses, by triggering intracellular biochemical changes which modify gap junctions.
To bring this concept of intercellular communication directly to the focus of the review, it should be noted that two of the earliest observations characterizing cancer cells were that they lost growth control and they could not normally terminally differentiate. One would predict, assuming that the gap junction played a major role in these two fundamental processes, that cancer cells would have some dysfunction in gap junctional intercellular communication. Lowenstein and his co-workers (Kanno, 1985; Lowenstein, 1966) observed very early in the study of gap junctions that many cancer cells did, indeed, have abnormal gap junction function. Later, assitional evidence showed that tumor metastasis correlated with decreased gap junction function (lijima et al., 1969; Nicolson et al .,1988; Ren et al., 1990). Some of the apparent discrepancies have been explained by the demonstration of selective communication, that is, some cancer cells could communicate with themselves but not with normal cells (Yamasaki et al., 1987).
Linking gap junctions with the initiation / promotion / progression model of carcinogenesis, the demonstration that tumor promoting chemicals and physical conditions could down regulate gap junctions is critical (Murray and Fitzgerald, 1979; Yancey et al., 1979; Yotti et al., 1979). These chemicals are “mitogenic” for the initiated cells. By blocking cell-cell communication, they allow the initiated stem or progenitor cell to escape the suppressing influence of the surrounding communication normal cells. Chemical tumor promoters, such as the phorbol esters, have been shown to decrease the gap junctions on cells in vitro, as well as in vivo (Beer et al., 1988; Janssen-Timmen et al., 1986; Kalimi and Sirsat, 1984; Sugie et al., 1987; Yancey et al., 1982). Even partial keratectomy, a tumor promoting stimulus (Arguis, 1985). Has been associated with the down regulation of gap junctional communication (Dermietzel et al., 1987). On the other hand, anti-tumor promoting chemicals, the retinoid, up regulate gap junctions in phenotypic ally transformed cells to reverse them to a normal phenotype (Mehta et al., 1986; Mehta et al., 1989; Mehta and Lowenstein, 1991). Of some relevance to this analysis is the observation that these tumor promoting chemicals appear to have threshold limits, below which they are unable either to down regulate gap junctions (Trosko and Chang, 1985), and promote the growth of tumors in vivo (Deml and Oesterle, 1987; Goldsworthy et al., 1984; Pitot et al. 1987; Verma and Bout well. 1986).
To link gap junctions to oncogenes and tumor suppressor genes, there have been a series of recent reports showing that cells expressing certain oncogenes (e.g.Ras, Src,Neu,Mos,Raf) had reduced gap junctional intercellular communication (Atkinson et al., 1981; Atkinson et al., 1986; Azarnia and Loewenstein, 1984; Azarnia and Lowenstein, 1987; Azarnia et al., 1988; Chang et al., 1985; Dotto et al., 1989; EI-Doulyy et al., 1988; EI-Fouly et al., 1989a; EI-Fouly et al., 1989b; Kalimi et al., 1990). In addition, non-communicating cancer cells (HeLa), when fused to form a non-tumorigenic hybrid with a normal human fibroblast cell, were shown to have functional gap junctional communication (Kalimi et al., 1990).
