This section is from the book "Symposium Phenomena Of The Tumor Viruses", by U.S. Dept. of Health. Also available from Amazon: Tumor Suppressing Viruses, Genes, and Drugs: Innovative Cancer Therapy Approaches.
These equations are not applicable to experiments which are concerned with the time course of infection after exposure of normal cells to RSV (Prince, Virology, 5: 435, 1958; Temin and Rubin, Virology, 8: 209, 1959). In the latter instance, the initial virus growth is exponential, as it is in any other virus infection. It only levels off to a constant rate after 3 or 4 days. In this regard RSV infection is like influenza virus infection. In fact, the growth curves of the tumor viruses which have been precisely studied are qualitatively indistinguishable from the growth curves for the myxoviruses. They differ only quantitatively, particularly in having a longer latent period and in producing infectious virus at a slower rate.
The comparison of intracellular and extracellular virus in both the established tumor (Rubin, Ann. New York Acad. Sc., 1957) and the freshly infected cells (Temin and Rubin, Virology, 1959) established that RSV is continuously released from cells as soon as the newly synthesized particle gains the property of infectivity, which suggested that, like influenza virus, RSV is completed at the cell membrane.
We have recently been able to show that the initial exponential shape of the RSV growth curve is due, at least in part, to an asynchrony in the time at which individual infected cells start to release virus. The morphologic alteration of chicken fibroblasts into Rous sarcoma cells is associated, at the population level, with the onset of virus production, and may be a direct consequence of virus production. Of particular interest is the change in the cell surface which must accompany the multiplication of any virus that is completed at the cell surface. Cell surface changes are likely to play a critical role in releasing the cell from contact inhibition (Abercrombie et al., Exper. Cell Res. 13: 276, 1957) and thus making the cell a cancer cell.
The evidence in Dr. Beaudreau's paper for exponential cell growth should be supported by experiments in which the increase of cells in any individual passage is measured over at least a tenfold range, rather than the twofold range employed. One cannot establish exponentiality by transferring the contents of 1 dish to 2 dishes to 4 dishes, etc., because one is then only measuring the exponentiality of the manual operation of transfer. From general experience with other cells, it would be surprising to encounter exponential cell growth at cell concentrations from 2.5 to 5 X 10^7 per ml. It is quite rare, even with cell lines well adapted to grow in tissue culture, to get exponential cell growth beyond a concentration of 10^6 cells per ml.
The use of particle counting to determine virus concentration has disadvantages as well as the advantages indicated in Dr. Beaudreau's paper. The chief disadvantage is most strikingly brought out by our recent finding (Rubin, unpublished) that some normal chick embryos are heavily infected with a virus which, in its biologic behavior in vitro, resembles lymphomatosis virus. We detect this virus because it induces resistance in cells to infection by RSV, if it has a head start-hence, its designation RIF for resistance-inducing factor. This agent is physically and immunologically indistinguishable from RSV, but, of course, does not produce the morphologic alteration which characterizes RSV infection. It appears to be ubiquitous in distribution-it is even present in our stocks of RSV. Any investigation of chicken tumor viruses which relies on particle counting will have to take this agent into account. Unless a precise relationship is established between particle count and biologic effect, and this relationship is demonstrated to remain constant throughout the course of an experiment, the particles being counted may be something quite distinct from the biologic problem under investigation.
Dr. Beard: In one principal aspect, the avian myeloblast culture system differs greatly from that of the Rous sarcoma discussed by Dr. Rubin. The myeloblasts established in culture were obtained from chickens with virus-induced myeloblastosis, and, as will be indicated in Dr. Bonar's paper later, it is probable that everyone of these cells carries the virus.
What Dr. Beaudreau has shown is that a system has been devised in which a virus-induced malignant cell will grow at an exponential rate over indefinite periods. Furthermore, the myeloblast is a natural host cell of this virus, both in the chicken and under artificial conditions in vitro. In this respect the myeloblast system is similar to that of the mouse mammary carcinoma studied by Dr. Lasfargues.
As a second principal feature of the myeloblast system, it was shown that the cells liberate the specific virus at a constant rate under the prescribed conditions throughout the period of study.
Dr. Beaudreau's paper has thus showed the recognition of a system of unique value for the study of the relationships between virus and cell in the induction of malignancy and maintenance of malignancy.
Dr. Sanford (National Institutes of Health): In the system described by Dr. Beaudreau, it is important to know the rates of cell necrosis which may distort the data completely. Were these rates measured?
Dr. Alice Moore (Sloan-Kettering Institute): I should like to ask if the infection of normal bone marrow in culture resulted in a change in the morphology of the myeloblasts?
Dr. Beaudreau: In answer to Dr. Rubin's discussion it needs to be said only that the application of the equation described here was simply an attempt to evaluate the rate of product formation (virus formation in this case) with respect to the worker (the cells) in a situation in which the working population was expanding at a rate definable by an exponential relationship. This general type of equation could be used for many purposes, for example, to describe the release of radioactive material from isotope-labeled cells or, for that matter, of automobiles produced by factory workers, as long as the rate of expansion in the number of the workers can be defined and the product counted. The application of this expression to the data of Prince on Rous virus formation was made only for the purpose of pointing out the necessity for consideration of both the virus and the cell together in a discussion of rates of virus formation. From text-figure 7 of the paper, it can be seen that the experimental points during the early part of the infection in Prince's experiments fall below the calculated values which assumed a constant rate of virus formation. This observation may be attributed to either an increasing capacity of the cells to produce virus or, as Dr. Rubin and others have suggested, an increased number of active infected centers. It is of interest that myeloblastosis virus formation shows a reversible effect similar to this under certain nutritionally deficient conditions.
The comparison of tumor virus formation with virus agents that do not induce tumors deserves some comment. Although there may be analogous processes involved in both, the inescapable difference is that the invasion of the cell by tumor virus causes a change in the host cell which makes it durable with respect to the host animal. The nature of this durability is the center of the tumor problem.