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Cytokine: more than a new word, a new concept proposed by Stanley Cohen thirty years ago
www.elsevier.com/locate/issn/10434666 Cytokine 28 (2004) 242e247
Cytokine: more than a new word, a new concept proposed by Stanley Cohen thirty years ago In 1974, Cohen et al. (Cohen S, Bigazzi PE, Yoshida T. Similarities of T cell function in cell-mediated immunity and antibody production. Cell Immunol 1974;12:150e9) introduced for the first time the word ‘‘cytokine’’. To commemorate this 30-year anniversary, we interviewed him. Could you remind us the main steps of your career? I attended Columbia University (BA and MD) and trained in Pathology at Harvard, followed by a fellowship with Drs. Baruj Benacerraf and Robert McCluskey at NYU. My initial research was done at the University of Buffalo and the University of Connecticut. Currently I am Chair of the Department of Pathology at the University of Medicine and Dentistry-New Jersey Medical School. I Co-chaired five of the International Lymphokine Workshops, and served on NIH and DOD Study Sections, as well as the editorial boards of several journals. I won the ParkeeDavis Award in 1977 for work on the in vivo role of lymphokines, and for the work that led to the identification of cytokines as a new category of biologic mediator. In 1986, I received a seven-year ‘‘Outstanding Investigator Award’’ from the NIH for research in these areas. I am currently the SecretaryeTreasurer of the American Society of Investigative Pathology. What was your main field of investigation in 1974? My initial interest, just prior to my work on cytokines, was on cell-mediated immunity and delayed hypersensitivity, mostly with respect to the cells that populated these reactions. Just prior to that, both John David and Barry Bloom had described MIF, and a number of other lymphocyte-derived factors (collectively called ‘‘lymphokines’’ by Dudley Dumonde). At that point, there was substantial controversy as to whether such factors were merely in vitro artifacts or whether they had biological significance, and much of doi:10.1016/j.cyto.2004.10.006
my work therefore focused on demonstrating in vivo roles for them. Takeshi Yoshida and I were able to demonstrate that lymphokines were produced in vivo, that they could be identified at reaction sites of cellular immunity, and that the injection of these factors into intact animals induced physiologic or pathologic responses, depending upon experimental conditions. Mine was also one of two laboratories (the other being that of Carolyn Geczy and Alain deWeck) to simultaneously first describe the successful production of anti-lymphokine antibodies, and my group was able to use these to confirm in vivo presence and activity of lymphokines.
Do you remember how you went to create this neologism? Was it long before it appeared in your 1974 paper? In other words could you tell how occurred the birth of the word ‘‘cytokine’’? In the course of the above work, we discovered that B cells, as well as T cells, could make MIF, and that mediators from either source had similar functional, physicochemical, and antigenic properties. This, plus Joe Oppenheim’s work on monocyte-derived LAF suggested the possibility that many cells other than lymphocytes could make such mediators. In this view, what was special about lymphoid cells was not the capacity to make mediators, but rather the fact that antigen could stimulate them to do so. Several reports described MIF-like activity in fibroblast cultures, but in those days, MIF was quantitated by a ratio of migration of macrophages in the presence of activated to control preparations, and there was no control other than medium for those experiments. Because of analogies with interferon, we thought that the use of virus-infected nonlymphoid cells would be a good test of this, since uninfected cultures would be the appropriate control. We found that virus infection of number of cell lines, including kidney cells, led to the production of MIFlike, chemotactic, and mitogenic activities indistinguishable from activities of known lymphokines. We also
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were able to show antigenic similarity between virusinduced MIF and lymphoid-derived MIF using the antilymphokine antibody described above. Based on this work, I postulated that that lymphokines produced by the immune system were merely a subset of a more general family of hormone-like mediators produced by many different kinds of cells. In analogy with Dumonde’s use of the word lymphokine, I suggested the name ‘‘cytokine’’ for these various functional macromolecules. Did the people of your lab acquired rapidly the use of this word? At that time, we had regular joint meetings of the immunologists at my institution (U.Conn.) and at Yale, and the word took hold first both in my lab and among several of the labs in Connecticut. Have you been proposing this word during conferences or meetings before it appeared in your review? Actually, I did little to ‘‘propose’’ the word, other than to use it in talks at various meetings, including the Lymphokine Workshops. Certainly there was no attempt to ‘‘market’’ the word itself, as I thought the concept was more important than the label. Basically, I began to think of the immune system as a specialized part of a total body (paracrine and autocrine) hormonal network, with the cytokines functionally analogous to hormones on the exocrine systems. Do you think that immunologists were (or still are?) too self-absorbed, considering the immune system working independently of the other systems, failing to integrate it within the functioning of the whole organism? The field of neuroimmunology comes to mind as an attempt to relate two integrative systems, and it is now appreciated that cytokines play roles in various organ systems, including the reproductive system. However, I think that there is too much emphasis on cytokines as mediators of host response and too little as mediators of normal growth, development, and homeostasis. Did you meet reluctant people about the use of this new word? Although the very first paper describing virus-induced lymphokine-like mediators was not well received by the immunologic journals (other than ‘‘Infection and Immunity’’, which published it), the other papers appeared in a number of other journals as well, and in
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addition, I was speculating about physiologic roles beyond cellular immunity for these agents (embryogenesis, growth and development, T cell helper function, etc.), so there was fertile soil. The word caught on almost immediately. Are there other papers or concept you had difficulties to publish, before it became well recognized? Only one other time. A little over a decade ago, I had begun to be interested in cytokine-mediated cell lymphoid proliferation and this rapidly evolved into an interest in the initiation of DNA synthesis in human cells. We adapted a system described by Manju Das, in which isolated quiescent nuclei were used as targets for potential initiating factors, and found that a protease or protease-like factor present in activated lymphocytes (ADR) was necessary for them to initiate synthesis (J Cell Biochem, 51:157,1993; Exp Cell Res 205:302,1993). To study the mechanism in detail we had to go to a more defined model, and we utilized plasmid constructs containing putative replication origin sequences from the human rRNA gene (Exp Cell Res, 209:123,1993). Very few human origins of replication had been characterized at that time, and we had difficulty in convincing reviewers that we had both bona fide replication origins and DNA synthesis (as opposed to repair). However, several other laboratories subsequently also reported origins in the regions we were utilized, and we were able to show actual increase in DNA following initiation (as compared to more indirect evidence of DNA synthesis). Since then, the field has blossomed enormously, and more sophisticated approaches than I am capable of have been brought to bear on the topic. However, the exact role of a protease in replication remains both unclear and relatively unstudied, and I suspect that I will return to exploring it. My (somewhat naı¨ ve) working hypothesis is that specific matrix associated docking sites are critical for initiation of DNA replication, and that the protease may uncover or expose these sites at critical moments. When you wrote this new word for the first time in your 1974 review, were you thinking/expecting that it will become so widely used? As I indicated above, I was much more interested in the extrapolation of the concept to the whole organism from the viral-induction studies. I was hoping that giving it a name would help the concept become more concrete. In fact, during the period in which I was CoChair of the Nomenclature Committee of the International Union of Immunological Societies, I did not propose it as standard nomenclature. I probably should have, but it clearly turned out to be unnecessary.
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You were in the Nomenclature Committee when in 1979 the word ‘‘interleukin’’ was created. Don’t you think, that the word was far to limiting (i.e. to leukocytes)? It is a common misconception that the term interleukin was accepted for adoption by the committee. When it was proposed, both Byron Waksman and I, as Co-chairs, felt it would turn out to be too restrictive (even then) and there was general agreement by the committee that it would be premature to refer to mediators as interleukins. This view was conveyed to the proposers. However, shortly thereafter, an editorial appeared in a journal (I forget which one) stating that the term ‘‘interleukin’’ had been discussed and considered by the committee (true) and that henceforth the term would be used in scientific communications by the scientists writing the editorial. This was a trueetrueunrelated kind of position to take, but it caught on in the general scientific community. Do you remember when you read or heard the word ‘‘cytokine’’ for the first time mentioned by another scientist? I believe people started using it in earnest at the first and second international lymphokine workshops (1976 and 1979) that I co-organized and co-chaired. The word ‘‘cytokine’’ appears more than 14 000 fold in the title of scientific papers or reviews. It seems that you rarely used it in the titles of your own papers! I tended to use it more in reviews and presentations than in primary articles. Again, I was more interested in the phenomena than the nomenclature. Also, my interests began to move first in the direction of cytokine-mediated cell proliferation, and then to a role for protease activity in the initiation of DNA replication, and then to origins of replication in human preparations. In retrospect, it would have been more ‘‘political’’ to publish more cytokine-focused papers with the word ‘‘cytokine’’.
This study showed me the possibility of analyzing complex systems where purified components are not available (as was the case for most of the early studies of lymphokines and cytokines by ourselves and others). (2) Sonozaki H, Cohen S. The macrophage disappearance reaction: mediation by a soluble lymphocyte-derived factor. Cellular Immunol. 2:341, 1971. I believe this was the first study to show an unequivocal role for a lymphokine in an in vivo system. (3) Flanagan, T. D., Yoshida, T. and Cohen, S. Production of macrophage migration inhibition factors by virus-infected cell cultures. Infect. & Immun. 8:145, 1973. The first report of induced MIF production in cells not of the immune system (although we had previously described chemotactic factors in similar settings). (4a) Cohen, S., Fisher, B., Yoshida, T. and Bettigole, R. Serum MIF activity in patients with lymphoproliferative diseases. New Eng. J. Med. 290:82, 1974. (4b) Torisu, M., Yoshida, T. and Cohen, S. Serum migration inhibitory factor in port-transplantation hepatic dysfunction. Clin. Immunol. & Immunopath. 3:369, 1975. Demonstration of the presence of lymphokines in specific disease states. This was the first in vivo demonstration of the presence of a given cytokine, if one except the in vivo description of interferon by A. Isaacs and G. Hitchcock and by J. Vilcek, in the early 1960s. (5a) Yoshida, T. and Cohen, S. The production of antiguinea pig lymphokine antibody. J. Immunol. 114:688, 1975. (5b) Yoshida, T., Bigazzi, P.E. and Cohen, S. Biologic and antigenic similarity of virus-derived MIF. Proc. Nat. Acad. Sci. USA 72:1641, 1975.
(1) Cohen S. The requirement for the association of two adjacent rabbit gamma G antibody molecules in the fixation of complement by immune complexes. J Immunol 100:407, 1968.
Production of anti-lympokine antibody and its use to provide further evidence of similarity between lymphoid and nonlymphoid generated mediator. As you can imagine it was technically quite difficult to make a polyclonal antibody (in the pre-monoclonal era) to a single small component of a complex mixture that proved impossible to purify to homogeneity (it took molecular cloning to obtain pure mediators, and that came much later).
I was able to show the two-for-one requirement by a mathematical analysis rather than using purified immunoglobulins or purified complement components.
(6a) Gutowski, J.K., Mukherji, B. and Cohen, S. The role of cytoplasmic intermediates in IL-2 induced T cell growth. J. Immunol. 133:3068, 1984.
Among your papers, what are your favorite ones?
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(6b) Wong, R., Gutowski, J.K., Katz, M., Goldfarb, R. and Cohen, S. Induction of DNA synthesis in isolated nuclei by cytoplasmic factors: inhibition by protease inhibitors. Proc. Nat. Acad. Sci. 84:241, 1987. (6c) Fresa, K.L., Autieri, M., Coffman, F.D., Georgoff, I., and Cohen, S. A cytosolic aprotinin-binding activator of DNA replication is tyrosine phosphorylated in its activeform. Exp. Cell. Res., 205: 302, 1993. (6d) Coffman, F.D., Georgoff, I., Fresa, K.L., Sylvester, J., Gonzales, I., and Cohen, S. In vitro replication of plasmids containing human ribosomal gene sequences: origin localization and dependence on an aprotinin-binding cytosolic protein., Exp. Cell Res., 209: 123, 1993. Demonstration of the role of a protease-like cytoplasmic intermediate involved in the regulation/initiation of cell proliferation in lymphoid cells. (7) Fernandes, H., Cohen, S., and Bishayee, S. Glycosylation-induced conformational modification positively regulates receptor-receptor association: a study with an aberrant EGF receptor expressed in cancer cells. J. Biol. Chem. 276:5375,2001. This is among my favorite papers simply because it resulted from Dr. Bishayee allowing me to participate in, and learn from his laboratory about the biochemistry of ligandereceptor interactions.
What is your favorite cytokine(s) and why? MIF. Although described at about the same time as LAF (IL-1) and IFN gamma, MIF focused attention on a potential role of non-antibody mediators in delayed hypersensitivity. Also, it is interesting that the name may be a misnomer because its effect appears to be on cell adhesion rather than movement per se. Its exact physiologic and host defense roles remain mysterious, although now that it has been cloned it is being found in interesting settings, and it is a molecule that I would like to return to studying.
When MIF was ‘‘rediscovered’’ as a product of the pituitary gland (Bernhagen et al. Nature 1993, 365, 756), I supposed it reinforced your concept? I was very excited to read that paper. Since that time, there have been a number of reports of MIF production by various non-immune cell types, and MIF transgenics have provided interesting results as well.
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In your ParkeeDavis Award Lecture (Am. J. Pathol. 1977, 88, 502-528) you mentioned that MAF and MIF were physicochemically similar; but MAF was later identified as gamma-interferon? Yes it was, and it was at that time that it was fully appreciated that MIF was not MAF in another guise. Several of us had initially speculated that MIF, MAF, and MCF might be a single pleiotropic molecule, but that was clearly not the case. My assertion that MIF and MAF were physicochemically similar was merely based on the fact that those activities appeared to co-purify by all of the purification schemes available at the time, and were therefore not separable by those physicochemical procedures. Modern approaches have obviously changed all that.
Nowadays, MIF is mainly considered as a pro-inflammatory cytokine. However, at homeostasis it exists as a circulating cytokine. How do you conciliate these two parameters? What about a natural ‘‘MIF binding protein’’ (as it does for IL-18) to neutralize its pro-inflammatory properties? As you know MIF is constitutively expressed or produced in various tissues, which strongly suggests a homeostatic role. There are several mechanisms whereby circulating MIF might not exert a pro-inflammatory function. These include (1) as you suggest, a binding protein, (2) its presence as a precursor form that is completely or partially functionally inactive, or (3) the concomitant presence of MIF inhibiting factors.
Indeed, you described an MIFIF (MIF inhibitory factor) (Cohen S, Yoshida T. Suppression of B cell MIF production by T cells and soluble T cell-derived factors. J Immunol. 1977;119:719e21). What happened to this factor? We decided to hold off pursuing it until MIF itself was better characterized in molecular terms, and that has happened only relatively recently. I am grateful that MIF has been ‘‘rediscovered’’ in modern molecular terms, but I don’t think we will get back to explore the range of activity of it and related molecules.
What has been for you the most surprising discovery in the area of cytokines? That they may have significant therapeutic potential.
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As a scientist, were not you surprised that despite the complexity of the cytokine network, one could address one individual cytokine as a therapeutic tool or target? If the cytokine network were not so complex, we probably could have cured cancer by now utilizing cytokine-based therapy. Nevertheless, I think that modified cytokines are potential tools for therapy (longer lasting, more powerful, expanded modes of action, targeted cytokines, etc. even as single entities. I would suspect, however, that cytokine cocktails would be more effective.
As an MD, were not you disappointed by the relative low clinical applications of the tremendous amount of data accumulated in the field? In general, there has been a lack of solid translational research in every area of biomedicine, as compared to basic research and direct clinical investigation. This has been probably due, at least in the US, to a failure of study sections, in the past, to consider translational research within the mantra of ‘‘hypothesis testing research’’. This mindset seems to have disappeared, and I suspect more clinical applications will appear at an exponential pace.
What have been for you the main achievements in the field of cytokines during these last 30 years? The application of molecular biological techniques to the field. Classical protein purification studies proved incapable of precise isolation and identification. Molecular cloning has led to the ability to obtain almost any cytokine of interest in pure form for study. Also, molecular analysis has allowed us to further understand mechanism of action at the level of ligandereceptor interaction.
What are the main questions you have about cytokines for which you are still waiting for the answer? Cytokines tend to participate in biologic reactions as a cytokine cascade, with multiple factors interacting, and with multiple regulatory loops. This plus the pleiotropic nature of cytokine activities makes it exceedingly difficult to understand how they actually produce a specific defense function in a specific set of circumstances, become part of an etiopathogenic pathway, or, for that matter regulate physiologic processes. Thus, understanding of cytokine action at the ‘‘systems’’ level awaits major new techniques and insights.
What are the main cytokines that have been published in the time of purified factors that never got confirmation once cloning became available? What were the sources of artifacts? I am not suredpeople tend to publish successes more than failures. However, an eosinophilic chemotactic factor that Peter Ward and I described in J. Exp. Med. has never been cloned and there is no evidence for it as an independent and unique entity. On the other hand, I am sure that there are many cytokines, defined in terms of biologic activity, that have not been cloned yet but which are real and interesting entities. A major potential source of artifact relates to both of the effects mentioned in item 14. I am sure that some so-called unique cytokines have been or will be proved to be due to interaction of several already well-defined cytokines. Also, there is a tendency to associate a newly discovered activity with a new cytokine, when it may merely be another manifestation of a cytokine already known.
In your 1974 Cell. Immunol. paper you were referring to ‘‘antigen specific factor’’. Those have vanished after the cloning period, isn’t it? This is the same for some others, like the ‘‘suppressive factor of allergy’’ for example. It is hard to believe that there were not some real phenomena underpinning some very well-performed science dealing with those issues. I wonder whether these have truly vanished or whether no one is interested in studying them with modern methodology.
Five journals have included the word ‘‘cytokine’’ in their title; hundreds of books in many languages have been published with the word you created. Any pride about it? Actually, it is interesting that a concept that I initially had trouble getting published took on such an astounding life of its own. I like to think that over and above the research findings that demonstrated the existence of this category of biologic mediator, which were very novel at the time, the association of the concept with a label helped to get people interested in studying it. I take pride in the fact that a cadre of young scientists, including John David, Barry Bloom, Joe Oppenheim, Jan Vilcek, Edgar Pick, Alain deWeck, and their scientific descendants and many others besides our own laboratory helped convert the field from a study of antibodyeantigen reactions and T cell-B cell interactions to a study of one of the major hormonal integrative pathways of the body as a whole (with the continuing encouragement and advice of Byron Waksman). Also, we got along well and had a great deal of fun.
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What would you tell to a young scientist to motivate him/ her to join the field of cytokines?
Have you had the chance to explain the world of cytokines to your children or grandchildren or to the general public?
This is the era of multi-factorial investigation (gene arrays, proteomics, informatics, etc.) allowing us to focus on ‘‘systems biology’’ which is a new way of saying physiology, but does imply a quantitative and predictive approach. Cytokine production and action, to the extent that these involve multiple, complex, interacting loops and cascades, represent ideal systems to study from this point of view.
I have been active in our university’s Mini-Med program, which was spearheaded by Dr. Jacob Lindenthal (a social epidemiologist in the Department of Psychiatry). In this program, we introduce interested lay audiences to the concepts of disease and disease processes, and I deal with immunology, with obvious attention to my favorites, the cytokines. Also, my wife, Dr. Marion Cohen, has been charged by the American Society of Investigative Pathology to develop a program for high school teachers to help them introduce ‘‘abnormal biology’’ and host defense into their biology curricula, and I talk about cytokines in the context of that program (which made its first appearance at the latest FASEB meeting). My children don’t need me to explain cytokines; all are research physicians (a pediatric endocrinologist, a medical endocrinologist, and an oncologist). My grandchildren range from age two to ten, and are still on T cells and B cells. Cytokines will come later.
Any regret(s)? Yes. As the field grew exponentially, my interest drifted towards less-explored research pathways. I regret that I did not maintain an active interest in cytokine research per se. I also regret that I took on some administrative career activities that seem to interfere with science.
As an MD, did you miss the clinic? Actually, I was always interested in research that could have the possibility of clinical application, rather than the treatment of patients with therapies already at hand, which is why I became an experimental pathologist rather than an internist.
What are your non-scientific interests? Photography, music, pen and custom knife collecting, trying to understand both modern physics and my grandchildren.
Don’t you think that effort should be made to teach the general public that may know the word ‘‘hormone’’ but not yet the word ‘‘cytokine’’? Many others, besides us, are working towards introducing medicine-related topics into very early stages of general education, in part because medically aware patients can interact more effectively with their physicians, and in part because the topics covered really are an essential part of modern biology/biomedicine. I think that it might be easier at first, if only for historical reasons, to introduce the concept of cytokines as regulatory components of host defense systems than to deal with it as a homeostatic and developmental mediator.
Cytokine: more than a new word, a new concept proposed by Stanley Cohen thirty years ago In 1974, Cohen et al. (Cohen S, Bigazzi PE, Yoshida T. Similarities of T cell function in cell-mediated immunity and antibody production. Cell Immunol 1974;12:150e9) introduced for the first time the word ‘‘cytokine’’. To commemorate this 30-year anniversary, we interviewed him. Could you remind us the main steps of your career? I attended Columbia University (BA and MD) and trained in Pathology at Harvard, followed by a fellowship with Drs. Baruj Benacerraf and Robert McCluskey at NYU. My initial research was done at the University of Buffalo and the University of Connecticut. Currently I am Chair of the Department of Pathology at the University of Medicine and Dentistry-New Jersey Medical School. I Co-chaired five of the International Lymphokine Workshops, and served on NIH and DOD Study Sections, as well as the editorial boards of several journals. I won the ParkeeDavis Award in 1977 for work on the in vivo role of lymphokines, and for the work that led to the identification of cytokines as a new category of biologic mediator. In 1986, I received a seven-year ‘‘Outstanding Investigator Award’’ from the NIH for research in these areas. I am currently the SecretaryeTreasurer of the American Society of Investigative Pathology. What was your main field of investigation in 1974? My initial interest, just prior to my work on cytokines, was on cell-mediated immunity and delayed hypersensitivity, mostly with respect to the cells that populated these reactions. Just prior to that, both John David and Barry Bloom had described MIF, and a number of other lymphocyte-derived factors (collectively called ‘‘lymphokines’’ by Dudley Dumonde). At that point, there was substantial controversy as to whether such factors were merely in vitro artifacts or whether they had biological significance, and much of doi:10.1016/j.cyto.2004.10.006
my work therefore focused on demonstrating in vivo roles for them. Takeshi Yoshida and I were able to demonstrate that lymphokines were produced in vivo, that they could be identified at reaction sites of cellular immunity, and that the injection of these factors into intact animals induced physiologic or pathologic responses, depending upon experimental conditions. Mine was also one of two laboratories (the other being that of Carolyn Geczy and Alain deWeck) to simultaneously first describe the successful production of anti-lymphokine antibodies, and my group was able to use these to confirm in vivo presence and activity of lymphokines.
Do you remember how you went to create this neologism? Was it long before it appeared in your 1974 paper? In other words could you tell how occurred the birth of the word ‘‘cytokine’’? In the course of the above work, we discovered that B cells, as well as T cells, could make MIF, and that mediators from either source had similar functional, physicochemical, and antigenic properties. This, plus Joe Oppenheim’s work on monocyte-derived LAF suggested the possibility that many cells other than lymphocytes could make such mediators. In this view, what was special about lymphoid cells was not the capacity to make mediators, but rather the fact that antigen could stimulate them to do so. Several reports described MIF-like activity in fibroblast cultures, but in those days, MIF was quantitated by a ratio of migration of macrophages in the presence of activated to control preparations, and there was no control other than medium for those experiments. Because of analogies with interferon, we thought that the use of virus-infected nonlymphoid cells would be a good test of this, since uninfected cultures would be the appropriate control. We found that virus infection of number of cell lines, including kidney cells, led to the production of MIFlike, chemotactic, and mitogenic activities indistinguishable from activities of known lymphokines. We also
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were able to show antigenic similarity between virusinduced MIF and lymphoid-derived MIF using the antilymphokine antibody described above. Based on this work, I postulated that that lymphokines produced by the immune system were merely a subset of a more general family of hormone-like mediators produced by many different kinds of cells. In analogy with Dumonde’s use of the word lymphokine, I suggested the name ‘‘cytokine’’ for these various functional macromolecules. Did the people of your lab acquired rapidly the use of this word? At that time, we had regular joint meetings of the immunologists at my institution (U.Conn.) and at Yale, and the word took hold first both in my lab and among several of the labs in Connecticut. Have you been proposing this word during conferences or meetings before it appeared in your review? Actually, I did little to ‘‘propose’’ the word, other than to use it in talks at various meetings, including the Lymphokine Workshops. Certainly there was no attempt to ‘‘market’’ the word itself, as I thought the concept was more important than the label. Basically, I began to think of the immune system as a specialized part of a total body (paracrine and autocrine) hormonal network, with the cytokines functionally analogous to hormones on the exocrine systems. Do you think that immunologists were (or still are?) too self-absorbed, considering the immune system working independently of the other systems, failing to integrate it within the functioning of the whole organism? The field of neuroimmunology comes to mind as an attempt to relate two integrative systems, and it is now appreciated that cytokines play roles in various organ systems, including the reproductive system. However, I think that there is too much emphasis on cytokines as mediators of host response and too little as mediators of normal growth, development, and homeostasis. Did you meet reluctant people about the use of this new word? Although the very first paper describing virus-induced lymphokine-like mediators was not well received by the immunologic journals (other than ‘‘Infection and Immunity’’, which published it), the other papers appeared in a number of other journals as well, and in
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addition, I was speculating about physiologic roles beyond cellular immunity for these agents (embryogenesis, growth and development, T cell helper function, etc.), so there was fertile soil. The word caught on almost immediately. Are there other papers or concept you had difficulties to publish, before it became well recognized? Only one other time. A little over a decade ago, I had begun to be interested in cytokine-mediated cell lymphoid proliferation and this rapidly evolved into an interest in the initiation of DNA synthesis in human cells. We adapted a system described by Manju Das, in which isolated quiescent nuclei were used as targets for potential initiating factors, and found that a protease or protease-like factor present in activated lymphocytes (ADR) was necessary for them to initiate synthesis (J Cell Biochem, 51:157,1993; Exp Cell Res 205:302,1993). To study the mechanism in detail we had to go to a more defined model, and we utilized plasmid constructs containing putative replication origin sequences from the human rRNA gene (Exp Cell Res, 209:123,1993). Very few human origins of replication had been characterized at that time, and we had difficulty in convincing reviewers that we had both bona fide replication origins and DNA synthesis (as opposed to repair). However, several other laboratories subsequently also reported origins in the regions we were utilized, and we were able to show actual increase in DNA following initiation (as compared to more indirect evidence of DNA synthesis). Since then, the field has blossomed enormously, and more sophisticated approaches than I am capable of have been brought to bear on the topic. However, the exact role of a protease in replication remains both unclear and relatively unstudied, and I suspect that I will return to exploring it. My (somewhat naı¨ ve) working hypothesis is that specific matrix associated docking sites are critical for initiation of DNA replication, and that the protease may uncover or expose these sites at critical moments. When you wrote this new word for the first time in your 1974 review, were you thinking/expecting that it will become so widely used? As I indicated above, I was much more interested in the extrapolation of the concept to the whole organism from the viral-induction studies. I was hoping that giving it a name would help the concept become more concrete. In fact, during the period in which I was CoChair of the Nomenclature Committee of the International Union of Immunological Societies, I did not propose it as standard nomenclature. I probably should have, but it clearly turned out to be unnecessary.
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You were in the Nomenclature Committee when in 1979 the word ‘‘interleukin’’ was created. Don’t you think, that the word was far to limiting (i.e. to leukocytes)? It is a common misconception that the term interleukin was accepted for adoption by the committee. When it was proposed, both Byron Waksman and I, as Co-chairs, felt it would turn out to be too restrictive (even then) and there was general agreement by the committee that it would be premature to refer to mediators as interleukins. This view was conveyed to the proposers. However, shortly thereafter, an editorial appeared in a journal (I forget which one) stating that the term ‘‘interleukin’’ had been discussed and considered by the committee (true) and that henceforth the term would be used in scientific communications by the scientists writing the editorial. This was a trueetrueunrelated kind of position to take, but it caught on in the general scientific community. Do you remember when you read or heard the word ‘‘cytokine’’ for the first time mentioned by another scientist? I believe people started using it in earnest at the first and second international lymphokine workshops (1976 and 1979) that I co-organized and co-chaired. The word ‘‘cytokine’’ appears more than 14 000 fold in the title of scientific papers or reviews. It seems that you rarely used it in the titles of your own papers! I tended to use it more in reviews and presentations than in primary articles. Again, I was more interested in the phenomena than the nomenclature. Also, my interests began to move first in the direction of cytokine-mediated cell proliferation, and then to a role for protease activity in the initiation of DNA replication, and then to origins of replication in human preparations. In retrospect, it would have been more ‘‘political’’ to publish more cytokine-focused papers with the word ‘‘cytokine’’.
This study showed me the possibility of analyzing complex systems where purified components are not available (as was the case for most of the early studies of lymphokines and cytokines by ourselves and others). (2) Sonozaki H, Cohen S. The macrophage disappearance reaction: mediation by a soluble lymphocyte-derived factor. Cellular Immunol. 2:341, 1971. I believe this was the first study to show an unequivocal role for a lymphokine in an in vivo system. (3) Flanagan, T. D., Yoshida, T. and Cohen, S. Production of macrophage migration inhibition factors by virus-infected cell cultures. Infect. & Immun. 8:145, 1973. The first report of induced MIF production in cells not of the immune system (although we had previously described chemotactic factors in similar settings). (4a) Cohen, S., Fisher, B., Yoshida, T. and Bettigole, R. Serum MIF activity in patients with lymphoproliferative diseases. New Eng. J. Med. 290:82, 1974. (4b) Torisu, M., Yoshida, T. and Cohen, S. Serum migration inhibitory factor in port-transplantation hepatic dysfunction. Clin. Immunol. & Immunopath. 3:369, 1975. Demonstration of the presence of lymphokines in specific disease states. This was the first in vivo demonstration of the presence of a given cytokine, if one except the in vivo description of interferon by A. Isaacs and G. Hitchcock and by J. Vilcek, in the early 1960s. (5a) Yoshida, T. and Cohen, S. The production of antiguinea pig lymphokine antibody. J. Immunol. 114:688, 1975. (5b) Yoshida, T., Bigazzi, P.E. and Cohen, S. Biologic and antigenic similarity of virus-derived MIF. Proc. Nat. Acad. Sci. USA 72:1641, 1975.
(1) Cohen S. The requirement for the association of two adjacent rabbit gamma G antibody molecules in the fixation of complement by immune complexes. J Immunol 100:407, 1968.
Production of anti-lympokine antibody and its use to provide further evidence of similarity between lymphoid and nonlymphoid generated mediator. As you can imagine it was technically quite difficult to make a polyclonal antibody (in the pre-monoclonal era) to a single small component of a complex mixture that proved impossible to purify to homogeneity (it took molecular cloning to obtain pure mediators, and that came much later).
I was able to show the two-for-one requirement by a mathematical analysis rather than using purified immunoglobulins or purified complement components.
(6a) Gutowski, J.K., Mukherji, B. and Cohen, S. The role of cytoplasmic intermediates in IL-2 induced T cell growth. J. Immunol. 133:3068, 1984.
Among your papers, what are your favorite ones?
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(6b) Wong, R., Gutowski, J.K., Katz, M., Goldfarb, R. and Cohen, S. Induction of DNA synthesis in isolated nuclei by cytoplasmic factors: inhibition by protease inhibitors. Proc. Nat. Acad. Sci. 84:241, 1987. (6c) Fresa, K.L., Autieri, M., Coffman, F.D., Georgoff, I., and Cohen, S. A cytosolic aprotinin-binding activator of DNA replication is tyrosine phosphorylated in its activeform. Exp. Cell. Res., 205: 302, 1993. (6d) Coffman, F.D., Georgoff, I., Fresa, K.L., Sylvester, J., Gonzales, I., and Cohen, S. In vitro replication of plasmids containing human ribosomal gene sequences: origin localization and dependence on an aprotinin-binding cytosolic protein., Exp. Cell Res., 209: 123, 1993. Demonstration of the role of a protease-like cytoplasmic intermediate involved in the regulation/initiation of cell proliferation in lymphoid cells. (7) Fernandes, H., Cohen, S., and Bishayee, S. Glycosylation-induced conformational modification positively regulates receptor-receptor association: a study with an aberrant EGF receptor expressed in cancer cells. J. Biol. Chem. 276:5375,2001. This is among my favorite papers simply because it resulted from Dr. Bishayee allowing me to participate in, and learn from his laboratory about the biochemistry of ligandereceptor interactions.
What is your favorite cytokine(s) and why? MIF. Although described at about the same time as LAF (IL-1) and IFN gamma, MIF focused attention on a potential role of non-antibody mediators in delayed hypersensitivity. Also, it is interesting that the name may be a misnomer because its effect appears to be on cell adhesion rather than movement per se. Its exact physiologic and host defense roles remain mysterious, although now that it has been cloned it is being found in interesting settings, and it is a molecule that I would like to return to studying.
When MIF was ‘‘rediscovered’’ as a product of the pituitary gland (Bernhagen et al. Nature 1993, 365, 756), I supposed it reinforced your concept? I was very excited to read that paper. Since that time, there have been a number of reports of MIF production by various non-immune cell types, and MIF transgenics have provided interesting results as well.
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In your ParkeeDavis Award Lecture (Am. J. Pathol. 1977, 88, 502-528) you mentioned that MAF and MIF were physicochemically similar; but MAF was later identified as gamma-interferon? Yes it was, and it was at that time that it was fully appreciated that MIF was not MAF in another guise. Several of us had initially speculated that MIF, MAF, and MCF might be a single pleiotropic molecule, but that was clearly not the case. My assertion that MIF and MAF were physicochemically similar was merely based on the fact that those activities appeared to co-purify by all of the purification schemes available at the time, and were therefore not separable by those physicochemical procedures. Modern approaches have obviously changed all that.
Nowadays, MIF is mainly considered as a pro-inflammatory cytokine. However, at homeostasis it exists as a circulating cytokine. How do you conciliate these two parameters? What about a natural ‘‘MIF binding protein’’ (as it does for IL-18) to neutralize its pro-inflammatory properties? As you know MIF is constitutively expressed or produced in various tissues, which strongly suggests a homeostatic role. There are several mechanisms whereby circulating MIF might not exert a pro-inflammatory function. These include (1) as you suggest, a binding protein, (2) its presence as a precursor form that is completely or partially functionally inactive, or (3) the concomitant presence of MIF inhibiting factors.
Indeed, you described an MIFIF (MIF inhibitory factor) (Cohen S, Yoshida T. Suppression of B cell MIF production by T cells and soluble T cell-derived factors. J Immunol. 1977;119:719e21). What happened to this factor? We decided to hold off pursuing it until MIF itself was better characterized in molecular terms, and that has happened only relatively recently. I am grateful that MIF has been ‘‘rediscovered’’ in modern molecular terms, but I don’t think we will get back to explore the range of activity of it and related molecules.
What has been for you the most surprising discovery in the area of cytokines? That they may have significant therapeutic potential.
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As a scientist, were not you surprised that despite the complexity of the cytokine network, one could address one individual cytokine as a therapeutic tool or target? If the cytokine network were not so complex, we probably could have cured cancer by now utilizing cytokine-based therapy. Nevertheless, I think that modified cytokines are potential tools for therapy (longer lasting, more powerful, expanded modes of action, targeted cytokines, etc. even as single entities. I would suspect, however, that cytokine cocktails would be more effective.
As an MD, were not you disappointed by the relative low clinical applications of the tremendous amount of data accumulated in the field? In general, there has been a lack of solid translational research in every area of biomedicine, as compared to basic research and direct clinical investigation. This has been probably due, at least in the US, to a failure of study sections, in the past, to consider translational research within the mantra of ‘‘hypothesis testing research’’. This mindset seems to have disappeared, and I suspect more clinical applications will appear at an exponential pace.
What have been for you the main achievements in the field of cytokines during these last 30 years? The application of molecular biological techniques to the field. Classical protein purification studies proved incapable of precise isolation and identification. Molecular cloning has led to the ability to obtain almost any cytokine of interest in pure form for study. Also, molecular analysis has allowed us to further understand mechanism of action at the level of ligandereceptor interaction.
What are the main questions you have about cytokines for which you are still waiting for the answer? Cytokines tend to participate in biologic reactions as a cytokine cascade, with multiple factors interacting, and with multiple regulatory loops. This plus the pleiotropic nature of cytokine activities makes it exceedingly difficult to understand how they actually produce a specific defense function in a specific set of circumstances, become part of an etiopathogenic pathway, or, for that matter regulate physiologic processes. Thus, understanding of cytokine action at the ‘‘systems’’ level awaits major new techniques and insights.
What are the main cytokines that have been published in the time of purified factors that never got confirmation once cloning became available? What were the sources of artifacts? I am not suredpeople tend to publish successes more than failures. However, an eosinophilic chemotactic factor that Peter Ward and I described in J. Exp. Med. has never been cloned and there is no evidence for it as an independent and unique entity. On the other hand, I am sure that there are many cytokines, defined in terms of biologic activity, that have not been cloned yet but which are real and interesting entities. A major potential source of artifact relates to both of the effects mentioned in item 14. I am sure that some so-called unique cytokines have been or will be proved to be due to interaction of several already well-defined cytokines. Also, there is a tendency to associate a newly discovered activity with a new cytokine, when it may merely be another manifestation of a cytokine already known.
In your 1974 Cell. Immunol. paper you were referring to ‘‘antigen specific factor’’. Those have vanished after the cloning period, isn’t it? This is the same for some others, like the ‘‘suppressive factor of allergy’’ for example. It is hard to believe that there were not some real phenomena underpinning some very well-performed science dealing with those issues. I wonder whether these have truly vanished or whether no one is interested in studying them with modern methodology.
Five journals have included the word ‘‘cytokine’’ in their title; hundreds of books in many languages have been published with the word you created. Any pride about it? Actually, it is interesting that a concept that I initially had trouble getting published took on such an astounding life of its own. I like to think that over and above the research findings that demonstrated the existence of this category of biologic mediator, which were very novel at the time, the association of the concept with a label helped to get people interested in studying it. I take pride in the fact that a cadre of young scientists, including John David, Barry Bloom, Joe Oppenheim, Jan Vilcek, Edgar Pick, Alain deWeck, and their scientific descendants and many others besides our own laboratory helped convert the field from a study of antibodyeantigen reactions and T cell-B cell interactions to a study of one of the major hormonal integrative pathways of the body as a whole (with the continuing encouragement and advice of Byron Waksman). Also, we got along well and had a great deal of fun.
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What would you tell to a young scientist to motivate him/ her to join the field of cytokines?
Have you had the chance to explain the world of cytokines to your children or grandchildren or to the general public?
This is the era of multi-factorial investigation (gene arrays, proteomics, informatics, etc.) allowing us to focus on ‘‘systems biology’’ which is a new way of saying physiology, but does imply a quantitative and predictive approach. Cytokine production and action, to the extent that these involve multiple, complex, interacting loops and cascades, represent ideal systems to study from this point of view.
I have been active in our university’s Mini-Med program, which was spearheaded by Dr. Jacob Lindenthal (a social epidemiologist in the Department of Psychiatry). In this program, we introduce interested lay audiences to the concepts of disease and disease processes, and I deal with immunology, with obvious attention to my favorites, the cytokines. Also, my wife, Dr. Marion Cohen, has been charged by the American Society of Investigative Pathology to develop a program for high school teachers to help them introduce ‘‘abnormal biology’’ and host defense into their biology curricula, and I talk about cytokines in the context of that program (which made its first appearance at the latest FASEB meeting). My children don’t need me to explain cytokines; all are research physicians (a pediatric endocrinologist, a medical endocrinologist, and an oncologist). My grandchildren range from age two to ten, and are still on T cells and B cells. Cytokines will come later.
Any regret(s)? Yes. As the field grew exponentially, my interest drifted towards less-explored research pathways. I regret that I did not maintain an active interest in cytokine research per se. I also regret that I took on some administrative career activities that seem to interfere with science.
As an MD, did you miss the clinic? Actually, I was always interested in research that could have the possibility of clinical application, rather than the treatment of patients with therapies already at hand, which is why I became an experimental pathologist rather than an internist.
What are your non-scientific interests? Photography, music, pen and custom knife collecting, trying to understand both modern physics and my grandchildren.
Don’t you think that effort should be made to teach the general public that may know the word ‘‘hormone’’ but not yet the word ‘‘cytokine’’? Many others, besides us, are working towards introducing medicine-related topics into very early stages of general education, in part because medically aware patients can interact more effectively with their physicians, and in part because the topics covered really are an essential part of modern biology/biomedicine. I think that it might be easier at first, if only for historical reasons, to introduce the concept of cytokines as regulatory components of host defense systems than to deal with it as a homeostatic and developmental mediator.