Clonal Origin of Skin and Bone Tumors Produced by Repeated Beta-irradiation in Mosaic Cell Mice

Clonal origin of skin and bone tomors produced by repeated beta-irradiation was determined by using mice with cellular mosaicism created by random X-chromosome inactivation, on the basis of phosphoglycerate kinase-1 (PGK). The backs of female C3H/He (Pgk-1^a/Pgk-1^b) mice were exposed to beta rays from ⁹⁰Sr-⁹⁰Y at a dose of 3 Gy per exposure 3 times weekly until tumors appeared. The cumulative tumor incidence reached 100% 500 days after the beginning of irradiation, as determined by the Kaplan-Meier method. All 8 tumors examined were of a single PGK phenotype: 5 squamous cell carcinomas and 2 osteosarcomas of A-type, and 1 squamous cell carcinoma of B-type. The absence of double PGK phenotype (AB-type) tumors indicated the monoclonal origin of the tumors produced by repeated irradiation.     

Long-term Exposure to the Anti-inflammatory Agent Phenylbutazone Induces Kidney Tumors in Rats and Liver Tumors in Mice

Long-term toxicity and carcinogenicity of phenylbutazone, a nonsteroidal anti-inflammatory drug, were evaluated in F344/N rats and B6C3F1 mice. In 2-year studies, phenylbutazone was given in corn oil by gavage 5 days per week to groups of 50 rats of each sex at doses of 0, 50, or 100 mg/kg body weight, and to groups of 50 mice at doses of 0, 150, or 300 mg/kg body weight. Body weights and survival were similar among groups. Major target organs are kidneys in rats, and liver in mice. Kidney: inflammation, papillary necrosis, and mineralization in both sexes of rats, and hyperplasia and dilatation of the pelvis epithelium, and cysts in female rats. Uncommon tubular cell tumors of the kidney were found in 13 exposed rats: 5 in the 50 mg group and 4 in the 100 mg group of males; 4 in dosed female rats; none in controls. In female rats, dose-related increases in hyperplasia of the pelvis transitional epithelium, and 2 carcinomas were discovered. Urinary bladder: papillomas of the transitional epithelium were seen in 2 low-dose male and in 1 low-dose female rats. Forestomach: ulcers in rats, with acanthosis, hyperkeratosis, and basal cell hyperplasia in female rats; however, no neoplasms were associated with these lesions. Liver: primarily in male mice exposed to. phenylbutazone, hemorrhage, centrilobular cytomegaly and karyomegaly, fatty metamorphosis, cellular degeneration, and coagulative necrosis were seen; clear cell foci were observed in male mice. In summary, under the conditions of these 2-year oral intubation studies, phenylbutazone is associated with renal carcinogenicity in rats, as evidenced by increases in tubular cell neoplasms in both sexes. Evidence of carcinogenicity for male mice was shown by increased incidences and multiplicity of liver tumors. No carcinogenic activity was found for female mice.     

Induction of Transplantable Tumors by Repeated Subcutaneous Injections of Natural and Synthetic Vitamin E in Mice and Rats

Natural vitamin E and synthetic vitamin E (dl/-α-tocopheryl acetate) were tested for their tnmorigenicity in rodents. Transplantable tumors, at the site of injection, were induced by repeated injections of these compounds in two strains of mice, NFS/N and C57BL/6N × C3H/He Fl, and in a strain of rats, Fischer 344. Natural vitamin E was tumorigenic in both strains of female mice only when injected with soya oil. In contrast, dl-α-tocopheryl acetate alone was capable of inducing tumors in Fischer 344 rats. Only one out of 5 male NFS/N mice given dl-α-tocopheryl acetate developed a tumor. Therefore, Fischer 344 rats were more susceptible to tumor formation by dl-α-tocopheryl acetate than NFS/N mice. dl-α-Tocopheryl acetate with soya oil or with palm oil also resulted in the formation of transplantable tumors in NFS/N mice and Fischer 344 rats. There was no difference in the tumor incidence between mice treated with dl-α-a-tocopheryl acetate alone and dl-α-tocopheryl acetate plus soya oil or palm oil. However, in rats, the incidence was lower for a group treated with dl-α-tocopheryl acetate plus palm oil than for those with dl-αa-tocopheryl acetate alone and with dl-α-tocopheryl acetate plus soya oil.     

Induction of Renal Cell Tumors in Rats and Mice, and Enhancement of Hepatocellular Tumor Development in Mice after Long-term Hydroquinone Treatment

Hydroquinone (HQ) was administered to F344 rats and B6C3F₁ mice of both sexes at a level of 0.8% in the diet for two years. This treatment induced renal tubular hyperplasia as well as adenomas, predominantly in males of both species, and was associated with chronic nephropathy in rats. In addition, the occurrence of epithelial hyperplasia of the renal papilla was increased in male rats. Foci of cellular alteration of the liver were significantly reduced in number by HQ in rats, but in contrast, were increased in mice, where development of hepatocellular adenoma was also enhanced in males. The incidence of squamous cell hyperplasia of the forestomach epithelium was significantly higher in mice of both sexes given HQ than in the controls, but no corresponding increase in tumor development was observed. The present study strongly indicates potential renal carcinogenicity of HQ in male rats and hepatocarcinogenicity in male mice. Thus, it is possible that HQ, which is present in the human environment, may play a role in cancer development in man.     

Rare Activation of the Human c-Ha-ras Transgene of Mice in Hemangioendothelial Sarcomas and Liver Tumors Induced by Glu-P-1

A transgenic mouse (Tg), having the human c-Ha- ras proto-oncogene, has been demonstrated to develop hemangioendothelial sarcomas (HESs) which are associated with the transgene mutation, but not to develop liver tumors. In this study, we examined the effects of 2-amino-6-methyldipyrido [1,2- a :3',2'- d ] imidazole (Glu-P-1), a food-borne carcinogen, which has been demonstrated to induce HESs and liver tumors in CDF₁ mice, on Tg mice. Chronic administration of 0.05% Glu-P-1 in the diet induced HESs in Tg (7/17), but not in 18 non-transgenic mice (N-Tg). With basal diet, two out of 17 Tg but none of 22 N-Tg, developed HESs. In contrast, Glu-P-1 administration induced liver tumors, both in Tg and in N-Tg; 16/17 in Tg and 13/18 in N-Tg. The incidence of hepatocellular carcinomas in Tg was higher than that in N-Tg (8/17 versus 3/18). With basal diet, only one out of 17 Tg and none of 22 N-Tg developed liver tumors. The Ha- ras mutation in tumors developed by the groups administered Glu-P-1, was examined. No mutations were detected in the transgene and mouse c-Ha- ras genes in all three HESs examined. In contrast, when 29 liver tumors taken from Tg were examined, two mutations of the transgene were detected in two HCCs. No mouse c-Ha- ras gene mutations were detected in any of the 47 liver tumors examined, which had developed in Tg and N-Tg mice. These results suggest that the transgene plays a role in the development of HESs induced by Glu-P-1, but not as a result of its mutation. Futher, the transgene plays no significant role in the development of liver tumors induced by Glu-P-1, but does play a role in the malignant conversion of some liver tumors, as a result of its mutation.     

Effect of an Inhibitor of Tumor Promotion, α-Difluoromethylornithine, on Tumor Induction by Repeated Beta Irradiation in Mice

α-Difluoromethylornithine (DFMO) is an inhibitor of putrescine biosynthesis and of tumor promotion. Studies were made of its effect on induction of skin and bone tumors by repeated beta irradiation of the back of female ICR mice 3 times weekly at a dose of 3 Gy per exposure. When given at a concentration of 10 mg/ml in the drinking water, DFMO significantly delayed the time of tumor emergence and decreased the yield of skin tumors, although the cumulative tumor incidence computed by the Kaplan-Meier method finally reached 100%. These results indicate that tumor-promoting activity is involved in repeated beta irradiation, besides its tumor-initiating activity.     

Site-specific Mutation of the Human c-Ha-ras Transgene Induced by Dimethylbenzanthracene Causes Tissue-specific Tumors in Mice

Forestoraach squamous cell carcinomas, lung adenocarcinomas and spleen angiosarcomas were induced by dimethylbenzanthracene (DMBA) in the rasH2 transgenic mouse line carrying human c-Ha-ras genes with their own promoter, encoding the prototype p21 gene product. Fifteen out of 21 mice (71%) developed forestomach squamous cell carcinomas, while 15 out of 21 (71%) had lung adenocarcinomas and 3 out of 21 (14%) showed spleen angiosarcomas within 8 weeks after a single administration of 50 mg/kg DMBA intraperitoneally. Somatic mutation at the 61st codon of the transgenes, from CAG(Gln) to CTG(Len), was detected in all these newly developed tumors. However, non-transgenic littermates demonstrated no tumors at all. These findings provide strong evidence that the somatic mutational activation of human c-Ha- ras genes is a critical event in tumorigenesis and a close relationship is therefore strongly suggested between the tissue-specific development of tumors and the somatic mutation of human c-Ha-ras genes in these rasH2 transgenic mice.     

Sex Hormone Dependency of Diethylnitrosamine-induced Liver Tumors in Mice and Chemoprevention by Leuprorelin

The prevalence of liver tumors throughout the world makes it imperative to seek chemopreventive agents. This tumor appears to be hormone-responsive and hormonal manipulations may therefore be beneficial. On this basis, both sexes of 12-day-old B6C3FJ mice were injected i.p. with diethylnitrosamine (DEN) at the dose of 2.5 μg/g body weight and observed for 32 weeks (males) or 36 weeks (females). In 100% of male mice, liver tumors were observed with an average diameter of 2.72 mm and multiplicity of 60.8. Orchidectomy at 6 weeks of age in these mice inhibited the incidence, multiplicity and size to 63%, 5.6 and 1.54 mm, respectively. By further implantation with an E₂ pellet at monthly intervals, these parameters were reduced to 26%, 0.6 and 0.61 mm, respectively. Administration of a gonadotropin-blocking chemical, leuprorelin, to DEN-treated male mice significantly reduced the multiplicity and size of tumors to 18.3 and 2.54 mm ( P <0.01 compared to those of DEN only). In female mice, the incidence of liver tumor was significantly smaller than that of males. However, ovariectomy and/or testosterone supplement significantly increased the occurrence of liver tumor. An anti-estrogen, toremifene, caused a marked further decrease of liver tumors. Mitotic indices with bromodeoxyuridine in tumor tissues paralleled the occurrence of liver tumors. Serum testosterone levels were significantly reduced by orchidectomy or by leuprorelin administration. These results further confirm that liver tumor is testosterone-responsive and hormonal manipulation by surgical orchidectomy or by chemical orchidectomy i.e. by leuprorelin, could substantially prevent the appearance of liver tumors.     

Rapid Growth and Spontaneous Metastasis of Human Germinal Tumors Ectopically Transplanted into Scid (Severe Combined Immunodeficiency) and Scid-nudestreaker Mice

Xenograft acceptance, growth and spontaneous metastasis of ectopically transplanted human germinal tumors were compared among scid mice, athymic nude mice and F₂ hybrids constructed from scid and nude mice, in relation to the impairments of T and B cell functions in these mice. In scid mice which are deficient in T and B cell functions, human yolk sac tumor (YST-2) that originated from the ovary grew to enormous sizes in 100% of the animals after both subcutaneous and intraperitoneal transplantation, while only half (59.1% and 51.9%) of the subcutaneous and none of the intraperitoneal transplants were accepted in usual athymic nude mice (BALB/c- nu/nu and CDl- nu/nu ). The YST-2 grew rapidly in scid mice, developing 3 to 10 times larger tumors compared to nude-streaker (AKR/J- nu^str/nu^str ) and usual nude mice, respectively. Furthermore, ectopically transplanted tumors spontaneously metastasized to distant organs (mostly to the lung) in scid mice (but less frequently in leaky scid mice), while metastases have never been found in nude mice. Although a xenograft of human classic (typical or pure) seminoma of the testis has never been established in nude mice, it grows slowly in one-third (36.4%) of scid mice and very rapidly in all of scid-nu ( scid/scid; nu^str/nu^str ) double mutant mice. Spontaneous metastases of xenografted seminomas were also observed in distant organs (lymph node, lung, liver, spleen, and kidney). The metastastic distribution of the two human germinal tumors in scid and scid- nu^str mice mimics that found in human. These results (xenograft acceptance, growth of transplanted tumors and degree of metastatic spread) were compatible with the level of T and B cell impairments indicated by FACS analysis, as well as mitogen responses, serum IgG and morphological features of the thymus.     

Effects of Endotoxin in Mice Bearing Solid Metastasizing Tumors and Treated with Lysozyme Hydrochloride

Summary

The effects of the i.v. administration of endotoxin (6.25–50 μg/mouse on day 13 after tumor implantation) in mice treated orally with lysozyme hydrochloride (100 mg/kg on days 5–12 from tumor implantation) were examined using Lewis lung carcinoma in the C57B1 mouse and MCa mammary carcinoma of CBA mice. On primary tumor growth, endotoxin alone causes a dose-dependent and statistically significant reduction with a nadir on day + 2 from endotoxin treatment. Combined with lysozyme, endotoxin causes an effect independent of the dose used, corresponding to the effect caused by endotoxin alone at the dose of 25 μg/mouse. No tumor regression was recorded in any of the treated groups. Endotoxin is virtually devoid of effects at the metastatic level. In the same conditions, lysozyme causes a reduction of primary tumor growth and a more pronounced inhibition of lung metastasis formation as expected from its already reported effects. The antitumor activity of endotoxin, unlike lysozyme, can be ascribed to tumor hemorrhagic necrosis due to tumor necrosis factor (TNF) production, as determined in tumor homogenates. Endotoxin does not increase the antitumor effects in mice treated with lysozyme, as expected from the data obtained with the more immunogenic SAI sarcoma, although lysozyme increased the mitogenic response to ConA of ex vivo isolated splenocytes, in vitro cultured in the presence of IL-2.     

苏乐康胶囊抑制理化致癌因子诱发小鼠的肿瘤发生

本文用苏乐康胶囊,剂量为39mg,每天1次灌胃,连续7天,抑制~(60)钴γ线全身照射0.5Gy和二乙基亚硝胺(DENA)皮下注射,剂量为0.1mg,每周注射1次,共8次,诱发雄性LACA小鼠肺肿瘤的发生。该胶囊不仅明显地抑制和阻断肺肿瘤的发生,其发生率由80.0％降到26.1％,P<0.01,而且肺癌的发生率也明显降低,由26.7％降到0.0%,P<0.01。该胶囊阻断γ线和DENA的联合致癌作用机制与其中人参、黄芪、维生素E等具有还原和抗氧化作用有关,能捕获自由基,减轻或消除其对靶细胞的损伤。     

Promotion of Skin Carcinogenesis by Dimethylarsinic Acid in Keratin (K6)/ODC Transgenic Mice

Dimethylarsinic acid (DMA) is a major metabolite of inorganic arsenicals in mammals, and arsenic exposure is associated with tumor development in a wide variety of human tissues, particularly the skin. Transgenic mice with ornithine decarboxylase (ODC) targeted to hair follicle keratinocytes are much more sensitive than littermate controls to carcinogens. In this study we investigated the promoting effect of DMA on skin carcinogenesis in such K6/ODC transgenic mice. The back skin of female C57BL/6J K6/ODC transgenic mice, 10 to 14 weeks old, was initiated with topical application of 7,12-dimethylbenz[α]anthracene (DMBA) at a dose of 50 μg or acetone alone on day 1 of the experiment, followed by treatment with 3.6 mg of DMA, 5 μg of 12-O-tetradecanoylphorbol-13-acetate (TPA) or neutral vehicle (control) twice a week for 18 weeks. Mice were killed 1 week after the end of the treatment. Induction of skin tumors was significantly accelerated in the DMA-treated group, as well as in the TPA-treated group, indicating that DMA has a promoting effect on skin tumorigenesis in K6/ODC transgenic mice.     

枸杞子多糖与放化疗合用对脑荷G422瘤小鼠的治疗作用

本实验观察了枸杞子多糖与~(60)Co头部照射及卡氮介(BCNU)合用对脑荷G422瘤小鼠生存期和脾脏T淋巴细胞增殖的影响,结果显示:三者合用不仅能产生明显的协同抑瘤作用,而且可明显改善荷瘤鼠的细胞免疫功能.     

Inhibitory Effects of Low-Dose Aloe-Emodin on the Development of Colorectal Tumors in Min Mice

Aloe-emodin (AE), a natural anthraquinone compound, has been reported to exhibit anticancer activityin various cancer cell lines and anti-inflammatory effects in murine macrophages. In the present study, weinvestigated the cancer chemopreventive effects of AE in an Apc-deficient Min mouse model. In the firstexperiment, male Min mice were fed a basal diet or diets containing 5 ppm AE and 10 ppm AE for 12 weeks.The dietary administration of 5 ppm AE significantly reduced the number of colorectal tumors. In a secondexperiment, we investigated the effects of AE on colitis-related colon carcinogenesis in Min mouse treated withdextran sodium sulfate (DSS). Female Min mice were administered 1% DSS in their drinking water for 7 days.AE was given to mice in their diet at a dose of 5 or 50 ppm for 5 weeks. Feeding with AE significantly reducedthe number of colorectal tumors. When proliferation of cells in normal-appearing colonic mucosa was assessedby monoclonal anti-rat Ki-67 antibody (MIB-5) immunohistochemistry in experiments 1 and 2, the AE treatmentsignificantly decreased the mean MIB-5-labeling index. These results suggest that the dietary administration oflow-dose AE may have chemopreventive effects against development of colorectal tumors in Min mice, possiblyin part by reducing cell proliferation in colorectal mucosa.     

Characterization of High-grade Neuroendocrine Tumors of the Lung in Relation to menin Mutations

It has been suggested that mutations in the menin gene play a role in the development of multiple endocrine neoplasia type 1 (MEN1)-associated and of sporadic forms of low- and intermediategrade neuroendocrine tumors of the lung. In the present study, eight tumor specimens of large cell neuroendocrine carcinoma (LCNEC) and 13 of small cell lung cancer (SCLC), which represent a high-grade category of neuroendocrine tumors, were examined for the potential involvement of menin alterations as well as for the expression of various neuroendocrine markers and p53 and Rb abnormalities. All specimens expressed multiple neuroendocrine markers as expected and almost invariably carried p53 and Rb alterations. Unexpectedly, however, mutations in the menin gene were not detected in any of the high-grade neuroendocrine tumors examined. We thus conclude that menin mutations do not play a crucial role in the pathogenesis of high-grade subsets, in contrast to their suggested significant role in the development of low- and intermediate-grade subsets. Interestingly, loss of heterozygosity (LOH) in the menin gene appeared to be more prevalent in LCNEC (50%) than in SCLC (22%), suggesting a possible distinction between SCLC and LCNEC.     

Similarity in the Effect of Caffeine on DNA Synthesis after UV Irradiation between Xeroderma Pigmentosum Variant Cells and Mouse Cells

The effect of UV irradiation on the rate of DNA synthesis was compared among normal human, xeroderma pigmentosum (XP, group A and variant) and mouse cells with and without caffeine in the culture medium after UV irradiation. At the same levels of survival, approximately 37%, all cells showed reduction in the rate of synthesis 0–3 h after UV irradiation followed by a recovery to normal or near-normal level 12 h later. In the presence of caffeine, no change in the recovery patterns was observed in normal human and XP A cells. XP variant cells and mouse cells showed little or no recovery in the presence of caffeine even after 12 h, when full recovery was obtained without caffeine. XP variant and mouse cells appear to have a common response in that post-irradiation treatment with caffeine inhibits reinitiation of UV-reduced DNA replication. Enhancement by caffeine of UV-killing in XP variant and mouse cells may he due to the retarded resumption of DNA replication.     

Changes in Cell Proliferative Parameters of SCCVII and EMT6 Murine Tumors after Single-dose Irradiation

To understand better the repopulation kinetics of tumor cells after radiotherapy, we Investigated changes in cell proliferative parameters after single-dose irradiation of SCCVII tumors in C3H/He mice and EMT6 tumors in Balb/c mice. The following parameters were determined 0–15 days after single irradiation at 20 or 30 Gy; dividing fraction (DP), potential doubling time (Tpot), number of clonogenic cells per tumor (Ncln), and volume doubling time (Tvol). DF and Tpot were determined by in vivo - in vitro cytokinesis-block assay with cytochalasin B, Ncln was measured by in vivo - in vitro colony-forming assay, and Tvol was determined by growth delay assay. In both tumors, longer Tpot and lower DF and Ncln were obtained for 3–4 days after irradiation, but in SCCVII tumors these values returned to the pretreatment levels 9 days after irradiation. In EMT6 tumors, Tpot, DF, and Ncln did not return to the pretreatment levels even 12 days after irradiation. In the regrowth phase of both tumors following irradiation at 20 Gy, Tvol was longer than the pretreatment level, although Tpot was similar in SCCVII and only slightly longer in EMT6. Therefore, the cell loss factor in the regrowth phase was considered to be higher than the pretreatment level in both tumors. From the results, recruitment of previously quiescent cells into the proliferative pool in these tumors was suggested to contribute to repopulation after radiation.