ANIMAL MODEL OF HUMAN DISEASE
Carcinoma of the Urinarv Bladder
Animal Model: Carcinoma of the Urinarv Bladder, Induced in Fischer Rats by N-[4-(5-Nitro-2furvD)-2-thiazolvl]formamide Contributed by: Samuel M. Cohen, MD, PhD, and Gilbert H. Friedell, MD, Departments of Pathology, St. Vincent Hospital and University of Massachusetts Medical School, Worcester, MA 01604.
Biologi Features
N-[4-(5-Nitro-2-furvl)-2-thiazolvl]formamide (FAN FT) reproducibly induces a 100% incidence of tumors of the urinary bladder in rats,"2 mice,3 hamsters,4 and dogs 5 without sex predominance. Only bladder tumors occur when FANFT is fed at a dose of 0.2% of the diet to weanling male inbred Fischer 344 rats 2; if FANFT is administered for the lifetime of the rats or for 23 or 36 weeks followed by control diet, all of the rats die from bladder cancer before the age of 20 months. The lesions of the urinary bladder are usually multiple, begin in the fundus, and progress through successive histologic stages from mild hyperplasia to invasive cancer. Hyperplasia is present in all rats within 2 weeks after FANFT administration is begun. It increases in severity through 8 weeks, after which nodular and papillary lesions appear together with the hvperplasia. These lesions, particularly the nodular ones, graduallv enlarge, protrude into the lumen, and may grow downward into the underlying stroma in a "broad front" pattern. Although occasionally present earlier, bv 25 to 30 weeks cellular and nuclear atvpia are evident within the lesions and invasion through the basement membrane can be demonstrated. Increased anaplasia and continued growth of the tumors occurs; after 40 to 30 weeks, invasion of the muscle laver is present. Invasion through the muscular wall into surrounding tissues occurs if the rat survives long enough, and metastases to inguinal and para-aortic lymph nodes, lungs, and other tissues are occasionallv found at necropsv. Publication sponsored by the Registrs of Comparative Pathology of the Armed Forces Institute of and supported by Public Health Service Grant RR 00301 from the Division of Research Resources. US Department of Health, Education and Welfare. under the auspices of Universities Associated for Research and Education in Pathologp- Inc. Research supported by Grant CA413945 from the National Cancer Institute through the National Bladder Cancer Project 0002-9440/79/0607-0849$01.00 849 © 1979 American Association of Pathologists
Pathology
850
COHEN/FRIEDELL
American Journal of Pathology
Obstruction of the ureters and subsequent hydronephrosis rarely occur in Fischer rats fed FANFT, nor do tumors develop in the ureters or renal pelvis as they do in Sprague-Dawley rats late in the course of the disease.' Microscopic hematuria is detectable after 10 weeks of FANFT feeding, becomes grossly visible by 25 to 40 weeks, and is severe late in the course of the disease, when the rat loses weight, develops severe anemia and an elevated blood urea nitrogen, and then dies 50 to 80 weeks after FANFT feeding was started. Nearly all the tumors are transitional cell carcinomas (Figure 1), although focal squamous metaplasia is common. Glandular metaplasia is seen less frequently. Undifferentiated carcinoma or spindle cell variants are occasionally induced; a sarcoma is rarely observed. Stromal infiltration of mononuclear cells, including lymphocytes, histiocytes, and plasma cells, appears with the onset of hyperplasia and increases with the progression to neoplasia. Lymphoid follicles are seldom seen in rats, unlike their presence in mice I with FANTF-induced bladder tumors. Mast cells are also present beneath the involved epithelium and increase with the development of the tumor 2 (Figure 2). The proliferation of submucosal blood vessels is increasingly evident during the development of epithelial hyperplasia; the pattern of proliferation changes at 10 weeks, with the appearance of nodular and papillary lesions.6 Calculi are not seen in the urine. Calcification is present infrequently in the tumors and then only in areas of necrosis. Its presence is unrelated to size or extent of invasion of the lesion. Comparison With Human Disease
FANFT-induced bladder carcinomas resemble the human disease in several ways. Most of the lesions are of transitional cell type and papillary, although most advanced tumors are polypoid masses protruding into the lumen (Figure 1) (a rather uncommon form of human bladder cancer). As with the human disease, the FANFT-induced lesions progress from epithelial hyperplasia to noninvasive carcinoma, invasion, and eventually distant metastases. The progression of the disease in the rat can be followed by urinary cytology,7 and hematuria is a frequent early sign of thq disease, as it is in the human. There would appear to be a particularly close parallel between human disease and rat model in the pathogenesis of early lesions, and histopathologic and cytologic studies have been done, including transmission and scanning electron microscopy. Pleomorphic microvilli represent a marker of irreversibility in the rat bladder epithelium proliferation in response to the carcinogen,8 and comparable changes are seen in human tumors. Pleomorphic microvilli are also seen on exfoliated bladder epithelial cells from human or animal subjects.
Vol. 95, No. 3 June 1979
CARCINOMA OF THE URINARY BLADDER
Figure 1-Noninvasive papillary transitional cell
carcinoma occurring as a polypoid mass protruding into the lumen of the bladder. A mononuclear cell infiltrate, including mast cells, is present in the subepithelial stroma. (H&E. x 100) Figure 2-Transitional cell carcinoma with numerous darkly stained mast cells in the delicate fibrovascular core of the tumor. (H&E, x 400)
851
852
COHEN/FRIEDELL
American Journal of Pathology
Usefulness of the Model
The use of an inbred strain makes possible studies on tumor-host interaction, including various immunologic studies,9 during different stages of the disease. The model is also useful for studies of other aspects of bladder cancer, including biochemical changes in the host during carcinogenesis, and various therapeutic investigations. A two-stage process in bladder carcinogenesis has recently been demonstrated using FANFT as an initiating agent,10 and the model may be a useful way to investigate possible promoting agents and possible initiating agents. Availability
FANFT-induced tumors in Fischer rats are readily transplantable; we have established tumor lines both by transplantation and in tissue culture. These are available for various studies.'1 Fischer rats are available from commercial sources. References 1. Ertiirk E, Price JM, Morris JE, Cohen S, Leith RS, von Esch AM, Crovetti AJ: The production of carcinoma of the urinary bladder in rats by feeding N-[4-(5-nitro-2furyl)-2-thiazolyl]formamide. Cancer Res 27:1998-2002, 1967 2. Tiltman AJ, Friedell GH: The histogenesis of experimental bladder cancer. Invest Urol 9:218-226, 1971 3. Erturk E, Cohen SM, Bryan GT: Urinary bladder carcinogenicity of N-[4-(5-nitro2-furyl)-2-thiazolyl]formamide in female Swiss mice. Cancer Res 30:1309-1311, 1970 4. Croft WA, Bryan GT: Production of urinary bladder carcinomas in male ham-
5.
6. 7. 8.
9. 10. 11.
sters by N-[4-(5-nitro-2-furyl-)2-thiazolyl]formamide, N-[4-(5-nitro-2-furyl)-2-thiazolyllacetamide, or formic acid 2-[4-(5-nitro-2-furyl)-2-thiazolyl]hydrazide. J Natl Cancer Inst 51:941-949, 1973 Ertfirk E, Atassi SA, Yoshida 0, Cohen SM, Price JM, Bryan GT: Comparative urinary and gallbladder carcinogenicity of N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide and N-[4-(5-nitro-2-furyl)-2-thiazolyl]acetamide in the dog. J Natl Cancer Inst 45:535-542, 1970 Cohen SM, Tatematsu M, Arai H, Fukushima S, Ogiso T, Tsuda H, Ito N: Neovascularization during urinary bladder carcinogenesis induced by N-[4-(5-nitro-2furyl)-2-thiazolyl]formamide (FANFT). Proc Jpn Cancer Assoc 36:241, 1977 Jacobs JB, Arai M, Cohen SM, Friedell GH: Light and scanning electron microsopy of exfoliated bladder epithelial cells in rats fed N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide. J Natl Cancer Inst 57:63-66, 1976 Jacobs JB, Arai M, Cohen SM, Friedell GH: Early lesions in experimental bladder cancer: Scanning electron microscopy of cell surface markers. Cancer Res 36:25122517, 1976 Johansson S, Cohen SM, Yang JPS, Arai M, Friedell GH: The influence of N-[4-(5nitro-2-furyl)-2-thiazolyl]formamide and phenacetin on the immune status in male Fischer rats. Invest Urol 15:308-311, 1978 Cohen SM, Arai M, Friedell GH: Promoting effect of DL-tryptophan and saccharin in urinary bladder carcinogenesis in the rat. Proc Am Assoc Cancer Res 19:4, 1978 Cohen SM, Arai J, Jacobs J, Friedell GH, Ito N: Transplantation and tissue culture of rat urinary bladder transitional cell carcinoma. Proc Jpn Cancer Assoc 35:209, 1976
Carcinoma of the Urinarv Bladder
Animal Model: Carcinoma of the Urinarv Bladder, Induced in Fischer Rats by N-[4-(5-Nitro-2furvD)-2-thiazolvl]formamide Contributed by: Samuel M. Cohen, MD, PhD, and Gilbert H. Friedell, MD, Departments of Pathology, St. Vincent Hospital and University of Massachusetts Medical School, Worcester, MA 01604.
Biologi Features
N-[4-(5-Nitro-2-furvl)-2-thiazolvl]formamide (FAN FT) reproducibly induces a 100% incidence of tumors of the urinary bladder in rats,"2 mice,3 hamsters,4 and dogs 5 without sex predominance. Only bladder tumors occur when FANFT is fed at a dose of 0.2% of the diet to weanling male inbred Fischer 344 rats 2; if FANFT is administered for the lifetime of the rats or for 23 or 36 weeks followed by control diet, all of the rats die from bladder cancer before the age of 20 months. The lesions of the urinary bladder are usually multiple, begin in the fundus, and progress through successive histologic stages from mild hyperplasia to invasive cancer. Hyperplasia is present in all rats within 2 weeks after FANFT administration is begun. It increases in severity through 8 weeks, after which nodular and papillary lesions appear together with the hvperplasia. These lesions, particularly the nodular ones, graduallv enlarge, protrude into the lumen, and may grow downward into the underlying stroma in a "broad front" pattern. Although occasionally present earlier, bv 25 to 30 weeks cellular and nuclear atvpia are evident within the lesions and invasion through the basement membrane can be demonstrated. Increased anaplasia and continued growth of the tumors occurs; after 40 to 30 weeks, invasion of the muscle laver is present. Invasion through the muscular wall into surrounding tissues occurs if the rat survives long enough, and metastases to inguinal and para-aortic lymph nodes, lungs, and other tissues are occasionallv found at necropsv. Publication sponsored by the Registrs of Comparative Pathology of the Armed Forces Institute of and supported by Public Health Service Grant RR 00301 from the Division of Research Resources. US Department of Health, Education and Welfare. under the auspices of Universities Associated for Research and Education in Pathologp- Inc. Research supported by Grant CA413945 from the National Cancer Institute through the National Bladder Cancer Project 0002-9440/79/0607-0849$01.00 849 © 1979 American Association of Pathologists
Pathology
850
COHEN/FRIEDELL
American Journal of Pathology
Obstruction of the ureters and subsequent hydronephrosis rarely occur in Fischer rats fed FANFT, nor do tumors develop in the ureters or renal pelvis as they do in Sprague-Dawley rats late in the course of the disease.' Microscopic hematuria is detectable after 10 weeks of FANFT feeding, becomes grossly visible by 25 to 40 weeks, and is severe late in the course of the disease, when the rat loses weight, develops severe anemia and an elevated blood urea nitrogen, and then dies 50 to 80 weeks after FANFT feeding was started. Nearly all the tumors are transitional cell carcinomas (Figure 1), although focal squamous metaplasia is common. Glandular metaplasia is seen less frequently. Undifferentiated carcinoma or spindle cell variants are occasionally induced; a sarcoma is rarely observed. Stromal infiltration of mononuclear cells, including lymphocytes, histiocytes, and plasma cells, appears with the onset of hyperplasia and increases with the progression to neoplasia. Lymphoid follicles are seldom seen in rats, unlike their presence in mice I with FANTF-induced bladder tumors. Mast cells are also present beneath the involved epithelium and increase with the development of the tumor 2 (Figure 2). The proliferation of submucosal blood vessels is increasingly evident during the development of epithelial hyperplasia; the pattern of proliferation changes at 10 weeks, with the appearance of nodular and papillary lesions.6 Calculi are not seen in the urine. Calcification is present infrequently in the tumors and then only in areas of necrosis. Its presence is unrelated to size or extent of invasion of the lesion. Comparison With Human Disease
FANFT-induced bladder carcinomas resemble the human disease in several ways. Most of the lesions are of transitional cell type and papillary, although most advanced tumors are polypoid masses protruding into the lumen (Figure 1) (a rather uncommon form of human bladder cancer). As with the human disease, the FANFT-induced lesions progress from epithelial hyperplasia to noninvasive carcinoma, invasion, and eventually distant metastases. The progression of the disease in the rat can be followed by urinary cytology,7 and hematuria is a frequent early sign of thq disease, as it is in the human. There would appear to be a particularly close parallel between human disease and rat model in the pathogenesis of early lesions, and histopathologic and cytologic studies have been done, including transmission and scanning electron microscopy. Pleomorphic microvilli represent a marker of irreversibility in the rat bladder epithelium proliferation in response to the carcinogen,8 and comparable changes are seen in human tumors. Pleomorphic microvilli are also seen on exfoliated bladder epithelial cells from human or animal subjects.
Vol. 95, No. 3 June 1979
CARCINOMA OF THE URINARY BLADDER
Figure 1-Noninvasive papillary transitional cell
carcinoma occurring as a polypoid mass protruding into the lumen of the bladder. A mononuclear cell infiltrate, including mast cells, is present in the subepithelial stroma. (H&E. x 100) Figure 2-Transitional cell carcinoma with numerous darkly stained mast cells in the delicate fibrovascular core of the tumor. (H&E, x 400)
851
852
COHEN/FRIEDELL
American Journal of Pathology
Usefulness of the Model
The use of an inbred strain makes possible studies on tumor-host interaction, including various immunologic studies,9 during different stages of the disease. The model is also useful for studies of other aspects of bladder cancer, including biochemical changes in the host during carcinogenesis, and various therapeutic investigations. A two-stage process in bladder carcinogenesis has recently been demonstrated using FANFT as an initiating agent,10 and the model may be a useful way to investigate possible promoting agents and possible initiating agents. Availability
FANFT-induced tumors in Fischer rats are readily transplantable; we have established tumor lines both by transplantation and in tissue culture. These are available for various studies.'1 Fischer rats are available from commercial sources. References 1. Ertiirk E, Price JM, Morris JE, Cohen S, Leith RS, von Esch AM, Crovetti AJ: The production of carcinoma of the urinary bladder in rats by feeding N-[4-(5-nitro-2furyl)-2-thiazolyl]formamide. Cancer Res 27:1998-2002, 1967 2. Tiltman AJ, Friedell GH: The histogenesis of experimental bladder cancer. Invest Urol 9:218-226, 1971 3. Erturk E, Cohen SM, Bryan GT: Urinary bladder carcinogenicity of N-[4-(5-nitro2-furyl)-2-thiazolyl]formamide in female Swiss mice. Cancer Res 30:1309-1311, 1970 4. Croft WA, Bryan GT: Production of urinary bladder carcinomas in male ham-
5.
6. 7. 8.
9. 10. 11.
sters by N-[4-(5-nitro-2-furyl-)2-thiazolyl]formamide, N-[4-(5-nitro-2-furyl)-2-thiazolyllacetamide, or formic acid 2-[4-(5-nitro-2-furyl)-2-thiazolyl]hydrazide. J Natl Cancer Inst 51:941-949, 1973 Ertfirk E, Atassi SA, Yoshida 0, Cohen SM, Price JM, Bryan GT: Comparative urinary and gallbladder carcinogenicity of N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide and N-[4-(5-nitro-2-furyl)-2-thiazolyl]acetamide in the dog. J Natl Cancer Inst 45:535-542, 1970 Cohen SM, Tatematsu M, Arai H, Fukushima S, Ogiso T, Tsuda H, Ito N: Neovascularization during urinary bladder carcinogenesis induced by N-[4-(5-nitro-2furyl)-2-thiazolyl]formamide (FANFT). Proc Jpn Cancer Assoc 36:241, 1977 Jacobs JB, Arai M, Cohen SM, Friedell GH: Light and scanning electron microsopy of exfoliated bladder epithelial cells in rats fed N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide. J Natl Cancer Inst 57:63-66, 1976 Jacobs JB, Arai M, Cohen SM, Friedell GH: Early lesions in experimental bladder cancer: Scanning electron microscopy of cell surface markers. Cancer Res 36:25122517, 1976 Johansson S, Cohen SM, Yang JPS, Arai M, Friedell GH: The influence of N-[4-(5nitro-2-furyl)-2-thiazolyl]formamide and phenacetin on the immune status in male Fischer rats. Invest Urol 15:308-311, 1978 Cohen SM, Arai M, Friedell GH: Promoting effect of DL-tryptophan and saccharin in urinary bladder carcinogenesis in the rat. Proc Am Assoc Cancer Res 19:4, 1978 Cohen SM, Arai J, Jacobs J, Friedell GH, Ito N: Transplantation and tissue culture of rat urinary bladder transitional cell carcinoma. Proc Jpn Cancer Assoc 35:209, 1976
