Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
[CANCER RESEARCH 31, 463-467, April 1971) Nucleic Acid Metabolism in Proliferating and Differentiating Colonie Cells of Man and in Neoplastic Lesions of the Colon1 Frank Troncale, Ralph Hertz, and Martin Lipkin Departments of Medicine, The New York Hospital, Memorial Hospital for Cancer and Allied Diseases, and the Cornell University Medical College ¡F.T.,M. L.¡,and the Department of Surgery, Memorial Hospital for Cancer and Allied Diseases, [R. H.J, New York, New York 10021 SUMMARY Enzymes involved in the metabolism of nucleic acid precursors were assayed in proliferating and maturing cells in the colon of man and in cells removed from polypoid lesions of the colon. Cells were separated from superficial and deeper layers of colonie mucosa by a recently developed tissue-planing instrument. Gradients of thymidine kinase, thymidine phosphorylase, and adenine and hypoxanthine phosphoribosyltransferase activities were found to characterize different stages of cell differentiation in normal colon. Thymidine kinase and phosphorylase were highest in young, proliferating cells and decreased during differentiation and migration of the cells to the mucosal surface. Phosphoribosyltransferase activities were lowest in young, proliferating cells and increased during cell differentiation. In the polypoid lesions including carcinomas patterns of enzyme activity characterizing young, proliferative cells were found. INTRODUCTION The mucosal lining of the gastrointestinal tract of man is continuously replaced by epithelial cells that migrate from the deep portion of the mucosal crypts to the surface. Most of these cells have a life-span of 2 to 4 days and are rapidly extruded from the surface of the mucosa into the lumen of the intestine (4, 23, 30). During migration, the epithelial cells undergo rapid morphological and biochemical changes as they differentiate into cells that carry out mature functional activities in each region of the intestine. These include the cessation of DNA synthesis and proliferative activity. How ever, in areas of colonie mucosa near hyperplasias and polypoid lesions and in the lesions themselves, epithelial cells are present that continue to synthesize DNA and proliferate throughout their entire life-span, as they migrate to the surface of the mucosa (10). In mammalian cells, factors that are believed to have a role in DNA synthesis and proliferative activity include protein and histone synthesis (5, 11, 31), ribosomal content (19), membrane potential (3), and cell mass (20). However, metabolic regulatory controls that lead to the onset of DNA synthesis and its cessation during the differentiation of normal intestinal cells have not been well defined. In small intestinal cells of rodents, it has recently been shown that, as DNA synthesis stops, marked changes develop in the activities of enzymes that have a role in the synthesis of nucleotide precursors of DNA and RNA (13, 16). Differences in the stability and turnover of these enzymes and their templates are also present during the differentiation and migration of rodent small intestinal cells, and it has been suggested that these factors may have a role in the differentiation of the cells (17). These characteristics of differentiating intestinal cells have not been studied in man, either in normal or diseased states. In this study, we have begun to explore these properties of intestinal cells in man, and we have measured the activities of enzymes involved in nucleic acid metabolism in proliferating and differentiating colonie epithelial cells, as they migrate to different levels of the colonie crypts. Several experiments were carried out: (a) the location in the colonie crypts of proliferating and nonproliferating epithelial cells was studied after pulse injection of TdR-3H,2 (mucosal biopsies and microautoradiography); (b) enzyme activities were studied in proliferative and nonproliferative cells removed separately from different layers of normal colonie mucosa obtained at operation; (c) enzyme activities were studied in cells lining the surfaces of polypoid lesions of the colon, some of which are believed to have an increased susceptibility to development of carcinoma (22, 27). MATERIALS AND METHODS Microautoradiographic Location of Proliferating and Nonproliferating Cells. Two patients each were given injections of 10 mCi of TdR-3H. Both patients had inoperable carcinomas of the colon with metastic lesions. Biopsies of colonie mucosa were taken from a colostomy opening in Patient 1 and from the rectal mucosa of Patient 2, 1 and 2 hr, respectively, after injection of TdR-3H. Microautoradiographs were prepared, and the location of cells incorporating TdR-3 H in the colonie crypts was determined microscopically (25). Preparation of Specimens of Colonie Mucosa for Enzyme Assay. Other specimens of histologically normal colonie 1This project was supported by NIH Grants and Awards 5 F03 AM 44662, CA-08921, and K 3-AM4468 from the USPHS. Received September 8, 1970; accepted December 14, 1970. 2The abbreviation used is: TdR-3H, thymidine-methyl-3 H. APRIL 1971 Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1971 American Association for Cancer Research. 463 Frank Troncale, Ralph Hertz, and Martin Lipkin mucosa were removed at operation from colons that contained a variety of different colon lesions: carcinomas, diverticular disease, a lipoma, and adenomatous polyps. Strips of normal mucosa 1 inch wide and 2 to 3 inches long were taken. The strips were always located 2 to 3 inches away from the lesion. Specimens were quickly placed on ice and transported to a cold room maintained at 32°F. The mucosa was separated from underlying tissues by careful dissection with scissors, laid out flat on a dissection board, cut into strips 1 cm wide and 6 to 10 cm long, and then stretched and clamped tight on the platform of a recently developed tissue-planing apparatus (16). This instrument contains a razor mounted on a movable overhead housing. The blade quickly moves across the fresh mucosa and removes progressively deeper layers of mucosa. A micrometer attachment regulated the depth of each plane that was cut. By this technique, the mucosa was separated into 3 approximately equal layers that were histologically identified as upper, middle, and lower thirds of the colonie crypts. Specimens of colonie mucosa were also obtained from patients at proctoscopy by gently scraping the surface of colon with a modified surgical spoon curet. Mucosal trauma was minimal, and histological examination of the scrapings revealed sheets of surface mucosal cells, in some instances attached to the upper portion of the crypts. The surface epithelium of adenomatous polyps, villous adenomas, and carcinomas was also gently scraped off with a No. 15 Bard-Parker knife from specimens obtained either at surgery or through the proctoscope. Hyperplastic polyps around 5 mm in size were homogenized completely. The tissue removed was homogenized in ice-cold Tris-HCl (pH 7.4) and adjusted to a volume of 0.5 ml. Homogenates were centrifuged in the cold at 15,000 X g for 15 min; the supernatant was used for enzyme assay as reported previously (16), and precipitates were assayed for DNA. In the specimens of mucosa removed by the razor planing apparatus, the total amount of DNA present averaged 0.19 mg; and in the specimens removed by surface scraping, the amount averaged 0.22 mg. Enzyme activities increased linearly during the time of incubation. Thymidine Rimise Assay (ATP:Thymidine S'-Phospho- The reaction mixture contained 150 mamóles of adenine-8-14C (specific activity, 200 cpm/mpmole), 50 ni/mióles of 5-phospho-D-ribosylpyrophosphate, 1 jmiole of MgCl2, 10 /¿molesof Tris-HCl buffer (pH 8.0), and 0.1 ml of supernatant containing enzyme, in a total volume of 0.2 ml. The reaction mixture was incubated at 37°for 30 min, and the reaction was stopped by immersion in boiling water. After cooling, the mixture was centrifuged at low speed for 5 min, and an aliquot of the supernatant was spotted on diethylaminoethyl paper and treated according to the method of Breitman (6). Hypoxanthine Phosphoribosyltransferase Assay (IMP: Pyrophosphate Phosphoribosyltransferase, EC 2.4.2.8). The reaction mixture and assay method were the same as above except for the use of 150 m/imoles of hypoxanthine-8-'4C (specific activity, 200 cpm/m/L/mole) in place of labeled adenine. Data on enzyme activities in upper, middle, and lower regions of colonie crypts obtained from normal strips of mucosa were subjected to the following statistical analysis. The numerical values for enzyme activity in each region of the crypt were correlated with the average distance of the cells in each region from the bottom of the crypt. The average cell distance in each of the 3 regions was taken from data on histological slides. Data on enzyme activities from polyps, villous adenomas, carcinomas, and surface mucosa were subjected to an analysis of variance to bring out differences among the various groups. RESULTS Microautoradiographic Measurements. The fraction of cells incorporating TdR-3 H into DNA at each cell position in the colonie crypts is shown in Chart 1. Most cells that were synthesizing DNA were located in the lower third of the transferase, EC 2.7.1.21). Thymidine kinase was assayed with the reaction mixture of Behki and Morgan (2) and the diethylaminoethyl paper method of Breitman (6). The 0.5 M Tris-HCl buffer (pH 8.0 at 37°)contained NaF, 1 mg/ml. Thymidine Phosphorylase Assay (Thymidine: Orthophosphate Deoxyribosyltransferase, EC 2.4.2.4). The reaction mixture contained 1 pinole of thymidine-2-14C (specific activity, 10s cpm/|/mole), 20 Amóles of phosphate buffer (pH 7.5), and 0.1 ml of supernatant containing enzyme in a total volume of 0.5 ml. Incubation was carried out for 30 min at 37°.The reaction was stopped by immersion in boiling water, and 50 jul of supernatant were spotted on Whatman No. 3MM Chromatographie paper strips. The end product of the reaction, thymine, was separated from thymidine in a decending Chromatographie system with the use of ethyl ace ta te :H2 O :formic acid (12:7:1) (upper layer used as solvent) (12). Adenine Phosphoribosyl transferase Assay (AMP:Pyrophosphate Phosphoribosyltransferase, EC 2.4.2.7). 464 Bottom 0 0.2 0.4 0.6 0 Fraction of cells labeledwith TdR- 0.2 0.4 Chart 1. Changes in the fraction of cells labeled at each cell position in microautoradiographs of the colonie crypts. Patient 1 (left), 1 hr after injection of TdR-3H, and Patient 2 (right), 2 hr after injection of TdR-3 H. — at cell positions 30 and 60, boundaries between the upper, middle, and lower thirds of the crypts. Very few cells in the upper third are synthesizing DNA. CANCER RESEARCH VOL. 31 Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1971 American Association for Cancer Research. Nucleic Acid Metabolism in Human Colon crypts, below cell position 30. The number of DNA-synthesizing cells gradually decreased in the midregion of the crypts from about cell position 30 to cell position 60. Few cells synthesized DNA as they reached the upper third of the crypts, and none were synthesizing by the time they reached the surface. Similar spatial distributions of cells in DNA synthesis in colonie crypts have been found in other studies (23, 24). These cells migrate rapidly from the deeper crypt regions to the surface, from where they are extruded into the intestinal lumen. Enzyme Activities in Regions of Normal Colonie Crypt. The comparative amounts of enzyme activity in the upper, middle, and lower thirds of the colonie crypts in normal mucosa are shown in Chart 2. There is a 4-fold decrease in the activity of thymidine kinase (12 colons) in the upper compared to the lower third of the crypts, indicating that the level of thymidine kinase falls off very rapidly during the migration of the cells through the middle third of the crypts. Since these cells migrate at a velocity of 1 to 2 cell positions/hour (26) the half-life of thymidine kinase is in the order of hours. Some enzyme activity is present in the upper third of the crypts, an area where virtually all cells have stopped making DNA. Chart 20 shows a 4-fold increase in adenine phosphoribosyltransferase activity as the cells migrated from the lower to upper third of the crypts. The activity of this enzyme increases rapidly as cells stop proliferating in the midregion of the crypts. Hypoxanthine phosphoribosyltransferase activity also increased during migration of the cells to the surface area of mucosa as seen in Chart 1c. However, this increase was not as marked as seen with adenine phosphoribosyltransferase. A decrease in the activity of thymidine phosphorylase was observed as cells migrated from the middle to the upper third of the colonie crypts (Chart 2d). The decrease in thymidine kinase and increase in adenine phosphoribosyltransferase activities were significantly correlated with cell distance from the bottoni of the crypt (0.05 > p > 0.01), and for hypoxan thine phosphoribosyltransferase the increase in activity was borderline significant (0.1 > p > 0.05). A linear increase or decrease in thymidine phosphorylase activity was not observed. Enzyme Activities in Polypoid Lesions. In Chart 3, comparative enzyme activities expressed per mg of DNA are shown in cells removed in vivo from the surface of the colonie mucosa and from the surface of the lesions. Cells were removed from the mature nonproliferative zone of normal mucosa; small hyperplastic polyps less than 1 cm in diameter; large polyps greater than 1 cm in diameter, most of which were adenomatous; villous adenomas; and carcinomas. Data on enzyme activities in lower-third proliferative cells shown in Chart 2 are replotted as¿ for comparison. As shown in Chart 3a, thymidine kinase activity was significantly greater in villous adenoma and carcinoma cells than in the other specimens (p < 0.005) and approximated levels of activity found in proliferative cells of normal colonie tissue. Adenine phosphoribosyltransferase activity (Chart 3b) was significantly greater in the mature surface cells of normal E S = 'S 3 3 £ _o 2: 1121 C E 2 2 lili Chart 2. Change in activity (mean ±S.E.) of 4 enzymes of purine and pyrimidine biosynthesis in human colonie crypts measured when the crypts were separated into lower, middle, and upper third by a razor planning instrument. Note differing magnitude of the first ordinales. The number of colon specimens studied is in parentheses, a, thymidine kinase; b, adenine phosphoribosyltransferase; c, hypoxanthine phos phoribosyltransferase; d, thymidine phosphorylase. colon than in the other tissues (0.025 >p> 0.01). Activity of this enzyme progressively decreased in the cells of polyps of increasing size. In villous adenomas and carcinomas, levels of activity reached those found in the immature proliferative cells of normal colonie tissue. The activity of hypoxanthine phosphoribosyltransferase was significantly higher in mature colon cells and small hyperplastic polyps (0.05 > p > 0.025) than the other lesions and decreased to reach the low levels found in immature proliferative cells (Chart 3c). However, a similar activity gradient was not found with thymidine phosphorylase; the amount of activity in hyperplastic tissue appeared to be closer to those found in proliferative than in mature cells (Chart 3d). DISCUSSION Previous work has shown that the mean generation time of actively proliferating colonie epithelial cells in man is about 2 days. In the proliferative region, approximately 30% of epithelial cells are making DNA, and others are moving through one of the phases of the proliferative cell cycle. Under normal conditions, most migrating epithelial cells in the colon of man stop making DNA 12 or more hr before they reach the luminal surface of the intestine, a situation analogous to small intestine when cells reach the upper region of the crypts and begin to move onto the villi (28). As cells migrate through the midregion of the crypts of both small intestine and colon, although progressively fewer cells replicate, they are still capable of being recalled into the proliferative cell cycle and of making new DNA, if new cells are needed, as for example after radiation damage (8); these cells are in a "transitional" stage between the proliferative and mature phases of their life cycle (28). However, once they have differentiated sufficiently to reach the colonie crypt surfaces or migrate onto the villi, they normally are no longer able to proliferate. This cessation of DNA synthesis and proliferative activity is APRIL 1971 Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1971 American Association for Cancer Research. 465 Frank Troncale, Ralph Hertz, and Martin Lipkin «e 0 (231 (U) (8l 12) (71 (21 (6l (111 5Z AOrS 3oE'S 2ESi 1st= 161"|,9liÃ-(201 1101 'EThymine formed, limóles /mg 30-—-©fin(19) DNA/ M (91 (91 121 SP LP VA 6l CA181i(71 L Chart 3. Change in enzyme activity (mean ±S.E.) in cells removed from normal colon and from neoplastic lesions. M, cells scraped by spoon curet from the surface of normal mucosa; SP, small polyps less than 1 cm; LP, surface cells scraped from polyps 1 cm or larger in diameter, VA, from villous adenomas; CA, from carcinomas. L, lowerthird proliferated cells of normal mucosa from Chart 2, shown here for comparison. Numbers in parentheses refer to the number of speci mens studied, a, b, c, and d refer to same enzymes as in Chart 2. accompanied by the rapid development of morphological and biochemical features that prepare the cells to function as elements lining the surface of the gastrointestinal tract. In this study, thymidine kinase activity was high in proliferative crypt cells and declined as the cells approached the surface of the mucosa. Recently, it was shown in small intestine of the rat (13, 16) that the levels of several of the enzymes involved in the synthesis and degradation of nucleic acid precursors change very rapidly during the normal differentiation of the 466 epithelial cells. Thymidine kinase, while present in young, proliferating cells located in the crypts, was not present in villous cells. This enzyme is widespread in proliferating cells and is absent from only a few cell types (9). Previous studies have shown that the half-life of thymidine kinase is characteristically in the order of minutes to a few hours (9), and in small intestine and liver of rat it is 2.6 hr (7, 16). Enzyme activity is increased in rapidly growing tumors (15) and in proliferating tissue culture cells (21). In the present study, thymidine phosphorylase activity did not increase during migration and differentiation of colonie cells of man. This is in contrast to the findings in small intestine, where a marked increase was observed during cell migration (16). This increase, together with the decrease in enzyme activity observed in leukemic cells (14, 29), had suggested that this enzyme might be involved in the regulatory control of DNA synthesis by limiting the availability of thymidine (16). In this instance, it is not known whether the failure to detect an increase in thymidine phosphorylase activity during the migration of colonie cells might be connected with their susceptibility to continued DNA synthesis and mitosis. Thymidine synthetase activity, not measured in this study, could reflect DNA synthesis more directly than either thymidine kinase or thymidine phosphorylase. In the colon, as in small intestine, adenine and hypoxanthine phosphoribosyltransferase activities increased with normal differentiation and migration of the cells. However, in the colon, the magnitude of the adenine phosphoribosyltransferase increase was greater than hypoxanthine phosphoribosyltransferase, in contrast to the findings in small intestine (16). In contributing to the formation of AMP's and IMP's, these enzymes may make it possible for salvage pathways to reclaim nucleic acid precursor materials from the intestine. The question of why levels of various enzymes change during differentiation of normal cells and the possible significance of these changes has received attention in recent years. In intestinal cells, which undergo rapid differentiation and move into new environments within hours, the rapid increase or decrease in the activity of these enzymes appears to be influenced by differential rates of turnover of both the enzymes and their templates. These have been postulated to act as regulatory controls contributing to the normal differentiation of the intestinal cells (17). The rapid appearance or disappearance of metabolic activities and the degradation of some of these enzymes could also involve active synthetic processes, as suggested by experiments in other systems. For example, the degradation of tyrosine aminotransferase (1, 18) and glucose 6-phosphate dehydrogenase (32) may be increased by the synthesis of specific proteins. However, it is not known whether this type of regulatory activity might contribute to changes in enzyme activity in normal differentiating intestinal cells or in the cells of these neoplastic growths. The present study shows that, in the cells of these neoplastic growths, "juvenile" patterns of enzyme activity that are found in proliferative cells persist, and patterns of activity that biochemically identify well-differentiated cells do not develop. CANCER RESEARCH VOL. 31 Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1971 American Association for Cancer Research. Nucleic Acid Metabolism in Human Colon ACKNOWLEDGMENTS We thank Dr. M. E. Balis for helpful suggestions and criticisms and Dr. Melvin Schwartz for aiding the statistical analysis. Microautoradiographs were prepared by Dr. Eleanor Deschner, and technical assistance was provided by Miss Luba Geller. 16. 17. REFERENCES 18. 1. Aurricchio, F., Martin, D., and Tomkins, G. Control of Degradation and Synthesis of Induced Tyrosine Aminotransferase Studied in Hepatoma Cells in Culture. Nature, 224. 806-808, 1969. 2. Behki, R. M., and Morgan, W. S. Studies on the Phosphorylation of Thymidine in Regenerating Rat Liver. Arch. Biochem. Biophys., 107: 427-434, 1964. 3. Ben-Or, S., Eisenberg, S., and Doljanski, F. Electrophoretic Mobilities of Normal and Regenerating Liver Cells. Nature , 188: 1200-1201, 1960. 4. Bertalanffy, F. D. Mitotic Rates and Renewal Time of Digestive Tract Epithelia in the Rat. Acta. Anat., 40: 130-148, 1960. 5. Borun, T. W., Scharff, M. D., and Robbins, E. Rapidly Labeled, Polyribosome-associated RNA Having the Properties of Histone Messenger. Proc. Nati. Acad. Sei. U. S., 58: 1977-1983, 1967. 6. Breitman, T. R. The Feedback Inhibition of Thymidine Kinase. Biochim. Biophys. Acta, 67: 153-155, 1963. 7. Bresnick, E., Williams, S. S., and Mosse, H. Rates of Turnover of Deoxythymidine Kinase and of Its Template RNA in Regenerating and Control Liver. Cancer Res., 27. 469-475, 1967. 8. Cairnie, A. B. Cell Proliferation Studies in the Intestinal Epithelium of the Rat: Response to Continuous Irradiation. Radiation Res., 32: 240-264, 1967. 9. Cleaver, J. E. Thymidine Metabolism: Pathways of Incorporation and Degradation In: A. Neuberger and E. L. Tatum (eds.), Frontiers in Biology, Vol. 6, pp. 43-69. New York: John Wiley and Sons, Inc., 1967. 10. Deschner, E., Lipkin, M., and Solomon, C. Study of Human Rectal Epithelial Cells in Vitro. II. H3-Thymidine Incorporation into Polyps and Adjacent Mucosa. J. Nati. Cancer Inst., 36: 849-857, 1966. 11. Estensen, R. D., and Baserga, R. Puromycin-induced Necrosis of Crypt Cells of the Small Intestine of Mouse. J. Cell Biol., 30: 13-22, 1966. 12. Fink, K., Cline, R. E., Henderson, R. B., and Fink, R. M. Metabolism of Thymine (Methyl-C14 or 2-C14) by Rat Liver in Vitro. J. Biol. Chem., 227: 425-433, 1956. 13. Fortin-Magana, R., Hurwitz, R., Herbst, J. J., and Kretchmer, N. Intestinal Enzymes: Indicators of Proliferation and Differentiation in the Jejunum. Science, 767; 1927-1928, 1970. 14. Gallo, R. C., and Perry, S. The Enzymatic Mechanisms for Deoxythymidine Synthesis in Human Leukocytes. IV. Comparisons between Normal and Leukemic Leukocytes. J. Clin. Invest., 48: 105-116, 1969. 15. Gordon, H. L., Bardos, T. J., Chmielewicz, Z. F., and Ambrus, J. L. APRIL 1971 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30. 31. 32. Comparative Study of the Thymidine Kinase and Thymidylate Kinase Activities and of the Feedback Inhibition of Thymidine Kinase in Normal and Neoplastic Human Tissue. Cancer Res., 28: 2068-2077, 1968. Imondi, A. R., Balis, M. E., and Lipkin. M. Changes in Enzyme Levels Accompanying Differentiation of Intestinal Epithelial Cells. Exptl. Cell Res., 58: 323-330, 1969. Imondi, A. R., Lipkin, M.. and Balis, M. E. Enzyme and Template Stability as Regulatory Mechanisms in Differentiating Intestinal Epithelial Cells. J. Biol. Chem., 245: 2194-2198, 1970. Kenney, F. T. Turnover of Rat Liver Tyrosine Transaminase: Stabilization after Inhibition of Protein Synthesis. Science, 756: 525-527, 1967. Killander, D., and Zetterberg, A. Quantitative Cytochemical Studies on Interphase Growth. I. Determination of DNA, RNA and Mass Content of Age Determined Mouse Fibroblasts in Vitro and of Interellular Variation in Generation Time. Exptl. Cell Res., 38: 272-284, 1965. Killander, D., and Zetterberg, A. A. Quantitative Cytochemical Investigation of the Relationship between Cell Mass and Initiation of DNA Synthesis in Mouse Fibroblasts in Vitro. Exptl. Cell Res., 40: 12-20, 1965. Kit, S., and Dubbs, D. R. Properties of Deoxythymidine Kinase Partially Purified from Noninfected and Virus-infected Mouse Fibroblast Cells. Virology, 26: 16-27, 1965. Lane, N., and Kaye, G. Pedunculated Adenomatous Polyp of the Colon with Carcinoma, Lymph Node Metastasis, and Suture Line Recurrence. Am. J. Clin. Pathol., 48: 170-182, 1967. Lipkin, M., and Bell, B. Cell Proliferation. In: C. F. Code (ed.), Handbook of Physiology, Alimentary Canal, Vol. 5, pp. 2861-2879. Washington, D. C.: American Physiological Society, 1968. Lipkin, M., Bell, B., and Sherlock, P. Cell Proliferation Kinetics in the Gastrointestinal Tract of Man. I. Cell Renewal in Colon and Rectum. J. Clin. Invest., 42: 767-776, 1963. Lipkin, M., and Quastler, H. Cell Population Kinetics in the Colon of the Mouse. J. Clin. Invest., 41: 141-146, 1966. Lipkin, M., Sherlock, P., and Bell, B. Cell Proliferation Kinetics in the Gastrointestinal Tract of Man. II. Cell Renewal in Stomach, Ileum, Colon, and Rectum. Gastroenterology, 45: 721-729, 1963. Morson, B. C. Factors Influencing the Progress of Early Cancer of the Rectum. Proc. Roy. Soc. Med. Ser. B, 59: 607-608, 1966. Quastler, H., and Sherman, F. G. Cell Population Kinetics in the Intestinal Epithelium of the Mouse. Exptl. Cell Res., / 7: 420-438, 1959. Seitz, J. F., and Luganova, S. The Biochemical Identification of Blood and Bone Marrow Cells of Patients with Acute Leukemia. Cancer Res., 28: 2548-2555, 1968. Shorter, R. G., Moertel, C. G., Titus, J. L., and Reitemeier, R. J. Cell Kinetics in the Jejunum and Rectum of Man. Am. J. Digest. Diseases, 9: 760-763, 1964. Stone, G. E., and Prescott, D. M. Cell Division and DNA Synthesis in Tetrahymena pyriformis Deprived of Essential Amino Acids. J. Cell. Biol., 21: 275-281,1964. Yagil, G., and Feldman, M. The Stability of Some Enzymes in Cultured Cells. Exptl. Cell Res., 54: 29-36, 1969. 467 Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1971 American Association for Cancer Research. Nucleic Acid Metabolism in Proliferating and Differentiating Colonic Cells of Man and in Neoplastic Lesions of the Colon Frank Troncale, Ralph Hertz and Martin Lipkin Cancer Res 1971;31:463-467. Updated version E-mail alerts Reprints and Subscriptions Permissions Access the most recent version of this article at: http://cancerres.aacrjournals.org/content/31/4/463 Sign up to receive free email-alerts related to this article or journal. To order reprints of this article or to subscribe to the journal, contact the AACR Publications Department at [email protected]. To request permission to re-use all or part of this article, contact the AACR Publications Department at [email protected]. Downloaded from cancerres.aacrjournals.org on June 18, 2017. © 1971 American Association for Cancer Research.