Injection Of 3-methylcholanthrene Research Articles

Inhibition of chemical carcinogenesis by berberine in rats and mice.

Berberine, an alkaloid isolated from the plant Berberis aristata, has been found to inhibit significantly the carcinogenesis induced by 20-methylcholanthrene (200 microg/0.1 mL/mouse) or N-nitrosodiethylamine (NDEA; 0.02% NDEA in distilled water, 2.5 mL/animal by gavage, five days a week for 20 weeks) in a dose-dependent manner in small animals. Administration of berberine (0.5, 2.5 or 5.0 mg kg(-1)) could reduce significantly the incidence of tumour in animals after an injection of 20-methylcholanthrene and increased their life span compared with the control. When berberine (10, 25 or 50 mg kg(-1)) was administered simultaneously with NDEA, the markers of liver injury (liver weight, gamma-glutamyl transpeptidase activity and glutathione S-transferase level) were reduced significantly compared with animals treated with NDEA only, which resulted in all the values being elevated. A similar decrease was noted in the serum levels of lipid peroxide, bilirubin and glutamate pyruvate transaminase. Morphology of liver tissue and levels of marker enzymes indicated that berberine offered protection against chemical carcinogenesis.

Evaluation of Elephantopus scaber on the inhibition of chemical carcinogenesis and tumor development in mice

The effect of the active fraction of Elephantopus scaber L. (Asteraceae) (ES) on skin papillomas induced by 7,12-dimethylbenz(a)anthracene (DMBA) as an initiator and croton oil as promoter was studied in mice. The active fraction of E. scaber (100 mg/kg) on topical application delayed the onset of papilloma formation and reduced the mean number of papillomas and the mean weight of papillomas per mouse. The intraperitoneal administration of the active fraction of E. scaber also had a significant effect on subcutaneous injection of 20-methylcholanthrene (20-MCA)-induced soft tissue sarcomas in mice. It inhibited the incidence of sarcomas and reduced the tumor diameter compared to MCA-treated control animals. The subcutaneous administration of the active fraction of E. scaber significantly inhibited the growth of subcutaneously transplanted DLA and EAC solid tumors, delayed the onset of tumor formation, and increased the life span of tumor bearing mice. The present study thus indicates the tumor inhibitory activity of the active fraction of E. scaber against chemically induced tumors and its ability to inhibit the development of solid tumors.

Pathology of soft‐tissue tumors: Daily diagnosis, molecular cytogenetics and experimental approach

This article reviews problems in diagnostic pathology and molecular cytogenetics of soft-tissue tumors. Also discussed are the origin of soft-tissue sarcomas and the molecular basis of effective target therapy for sarcomas. Molecular cytogenetic analysis of tumor-specific chromosomal translocations and associated fusion gene transcripts offers a useful adjunct to the diagnosis of soft-tissue tumors, but recent studies have indicated a growing number of fusion gene variations in each tumor type. In pleomorphic sarcoma/malignant fibrous histiocytoma, the alternative lengthening of telomeres (ALT) mechanism may result in formation of anaphase bridges and marked nuclear pleomorphism. The histogenesis of soft-tissue sarcomas has been a matter of controversy. In the present experimental model using s.c. injection of 3-methylcholanthrene in C57BL/6 mice pretreated with bone marrow-transplantation from green fluorescent protein (GFP)-positive green mice, the bone marrow-derived mesenchymal stem cells as well as the tissue-resident mesenchymal cells in the peripheral soft tissues are possible originators of sarcomagenesis. Little is known about a molecular basis of target therapy for sarcomas. Platelet-derived growth factor-BB (PDGF-BB) enhances the invasive activity of malignant peripheral nerve sheath tumor (MPNST) cells through platelet-derived growth factor receptor (PDGFR) phosphorylation, whereas imatinib mesylate inhibited such activity, suggesting that targeting PDGFR-beta may result in the establishment of novel treatment for MPNST. In addition, emmprin is a transmembrane glycoprotein on tumor cells that stimulates peritumoral fibroblasts to produce matrix metalloproteinases (MMP), playing a crucial role in tumor progression, invasion and metastasis. The MMP upregulation mechanism mediated by tumor-associated emmprin may be a potentially useful target in anti-tumor invasion therapy for sarcomas.

Effect of cytochrome P450 1A induction on oxidative damage in rat brain.

Polycyclic and halogenated aromatic hydrocarbons (PAHs and HAHs) can enhance the generation of reactive oxygen species (ROS) by inducing cytochrome P450 1A (CYP 1A) in vivo and in vitro. While the brain is vulnerable to oxidative injury, whether or not CYP 1A induction in the brain can produce measurable levels of oxidative damage has not been reported. The objective of this study was to investigate the effect of this induction on oxidative damage to the CNS. Time course changes in rat brain CYP 1A activity were determined by measurement of ethoxyresorufin O-deethylase (EROD) activity in whole brain homogenates. Three days after exposure of rats to five daily injections of 3-methylcholanthrene (3-MC) an approximately sevenfold increase in EROD activity was observed. Hepatic levels were increased 60-100 fold. This increase in CYP 1A activity was not accompanied by increased protein or lipid oxidation as measured by tryptophan fluorescence and TBAR formation, or decreased glutamine synthetase (GS) activity. These findings indicate that if increased CYP 1A activity in the brain following 3-MC treatment leads to increased ROS generation, the increase is insufficient to overwhelm the endogenous antioxidant defense system, produce detectable oxidative damage, and alter glutamate homeostasis.

Regression and prevention of autochthonous tumors induced by 3-methylcholanthrene after injection of a T-cell receptor α/β positive and CD4/CD8 double negative T-cells

Both the therapeutic and preventative effects of a murine T-cell line, tMK-2, with T-cell receptor (TCR) α/β positive and CD4 −/8 − double negative (DN) phenotype against autochthonously tumors induced by subcutaneous (s.c.) injection of 3-methylcholanthrene (MC) were examined. Complete regression of the tumor was observed when administration of tMK-2 cells was begun on tumors 5 mm in diameter. The tumor mass in five out of five mice was reduced in size after the administration of tMK-2 cells regardless of the routes of administration: s.c. injection of tMK-2 cells (5×10 7 cells) once a week around tumors, intraperitoneal (i.p.) injection (5×10 7 cells), or intravenous (i.v.) injection (1×10 7 cells). The tumors regressed to the status of a scar within 1 month of initial injection, and this status was maintained throughout the remainder of the 3 months period of tMK-2 cell injection. One month after discontinuation of tMK-2 cell administration, the diameter of the tumors had not increased regardless of the route of injection. The control groups consisted of either untreated mice, mice with i.v. injection of 1 μg of recombinant murine interleukin (IL)-12 once a week, or mice with s.c. injection of autologous splenocytes (5×10 7) from BALB/c mice once a week. Continuous growth of tumors was observed in each group and all control mice died due to bleeding ulcerations of the tumors. Tumor development was effectively prevented when tMK-2 cells were administrated 1 week after the s.c. injection of MC. In the groups receiving s.c., i.p., and i.v. injection of tMK-2 cells, no MC-induced tumors developed, whereas four out of five of the control mice developed autochthonous tumors. The tMK-2 cells also exerted in vitro NK-like cytotoxic activity, and their killing activity was strongly increased in the presence of both IL-2 and IL-12. These results suggest that the injected T-cells with TCR α/β positive and CD4 −/8 − DN phenotype and NK-like activity are important in the therapy as well as the prevention of tumor development.

Aryl hydrocarbon hydroxylase activity levels in the hepatic microsomal fraction from MC- or TCDD-treated mice: a comparison between aromatic hydrocarbon responsive and non-responsive strains

The effects of in vivo administration of polycyclic aromatic hydrocarbons on the levels of aryl hydrocarbon hydroxylase (AHH) activity in aromatic hydrocarbon (Ah) responsive and non-responsive strains of mice were studied using the hepatic microsomal fraction. Injection of 3-methylcholanthrene (MC; 42 mg kg body wt.) or 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD; 120 μg kg body wt.) into both strains produced marked enhancement of AHH activity except for MC treatment of Ah non-responsive strains. Addition of 7,8-benzoflavone (BNF) to the microsomal AHH assay mixture prepared from mice previously injected with vehicle (olive oil) alone caused an increase in activity when the mice were responsive, while BNF lowered the activity in non-responsive strains. With regard to MC-injected mice, BNF and 3-methyl-sulphonyl-4,5,3',4'-tetrachlorobiphenyl (3-MSF-TCB) decreased microsomal AHH activity in Ah-responsive mice, whereas these drugs enhanced the activity in Ah-non-responsive strains. 3-MSF-TCB also had inhibitory potency on AHH activity, but the mechanism of inhibition seems to be somewhat different from that of BNF. It may also suggest that cytochrome P-450 isozymes inhibited by BNF are different from those inhibited by 3-MSF-TCB.

Modulation of lung tumor development in mice with the soybean-derived Bowman-Birk protease inhibitor.

Male strain A mice were treated with a single i.p. injection of 3-methylcholanthrene (MCA). Four months later, the number of lung tumors was counted. In mice treated three times a week, for 8 weeks, with crude soybean extract containing the Bowman-Birk protease inhibitor (BBI), the number of lung tumors was significantly lower than in control animals receiving carcinogen treatment only (40-70% of controls). On the other hand, treatment initiated 8 weeks after MCA only had no effect on tumor development. A reduction in the number of lung tumors was also found in animals treated i.p. or orally with purified BBI three times a week for 8 weeks following MCA administration. It is concluded that BBI is capable of partially blocking the development of lung tumors in mice.

Effect of H2-Receptor Antagonists on Acetaminophen-lnduced Hepatic Injury

AbstractThe effects of H2-antagonists on acetaminophen-induced hepatic injury were examined. Rats were administered acetaminophen at the dose of 800 mg/kg body, 60 hr after injection of 3-methylcholanthrene. As an H2-antagonist. Cimetidine (200 mg/kg). ranitidine (50 mg or 100 mg/kg). or famotidine (20 mg or 40 mg/kg) was administered before and after acetaminophen injection. The group administered only acetaminophen had been severely damaged as evaluated by changes in serum transaminase, P-450 content, aminopynne demethylation. glutathione content and histological study, but administration of 200 mg Cimetidine together with acetaminophen significantly reduced the hepatic injury to nearly the control level. Ranitidine had no protective effect against hepatic injury at the dose of 50 mg. which appears to have the same antacid effect as 200 mg cimeitdine. whereas it had a slight but significant protective effect as evaluated by the transaminase level, glutathione content and histological study at the dose of 100 mg. Famotidine had no effect against acetaminophen induced hepatic injury. Because famotidine had no effect, the protection by H2-antagonist against acetaminophen-induced hepatic injury cannot be explained by the decrease in hepatic blood flow alone. Therefore, inhibition of P-450 activity seems to be more important for reducing the generation of the reactive metabolites of acetaminophen than hepatic blood flow decrease.

Suppressive effects of 3-methylcholanthrene on the in vitro antitumor activity of naturally cytotoxic cells.

Transient suppression of splenic natural killer (NK), natural cytotoxic (NC), and peritoneal macrophage cytotoxicity was observed following a single injection of 3-methylcholanthrene (3-MC) into C3H/HeN mice. Natural killer cell activity was depressed by 30-60% 4-6 d after injection of 1.0 mg 3-MC. Levels of NK reactivity returned to normal 8 d post 3-MC injection, and no suppression of natural killing was seen when tested 6 wk after 3-MC treatment. 3-MC did not affect Propionibacterium acnes augmentation of NK cell activity when tested both 6 d and 6 wk after carcinogen injection. The results indicate that the observed suppression of naturally cytotoxic cells may not be important in allowing 3-MC-induced tumors to grow, since suppression is not long-lasting. Therefore, any effect on tumor growth mediated by a suppression of naturally cytotoxic cells would have to be exerted at the earliest stages of tumor development.

Effect of Tuftsin on In Vivo Development of 3-Methylcholanthrene-Induced Primary Fibrosarcoma and Lewis Lung Carcinoma in Mice<xref ref-type="fn" rid="FN2">2</xref>

The effect of tuftsin therapy on tumor development was examined in a murine primary fibrosarcoma and the Lewis lung carcinoma systems. Following im injection of 3-methylcholanthrene (CAS: 56-49-5) on day 0, C57BL/10ScSn mice were treated weekly with 3 ip tuftsin injections beginning on day 1 or day 60. Similar patterns of tumor development were observed regardless of whether tuftsin therapy was immediate or delayed. Only modest differences in experimental and control tumor incidences were found upon termination of studies; however, treated animals developed significantly fewer tumors than controls early during the observation periods. Thus mean tumor latent periods varied significantly when therapy began on day 1 (103.6 days in controls vs. 119.1 in treated mice; P = .02) or 2 months later (104.6 days in controls vs. 115.3 in treated mice; P = .01). One day subsequent to intra-footpad implantation of 10(5) Lewis lung carcinoma cells, C57BL/6 mice received at least 10 iv injections of tuftsin and were compared with controls for variations in survival or lung tumor development. The mean survival time in treated mice, 41.2 days, differed sharply from that (30.1 days) in controls (P = .00001). Similar groups of mice varied significantly in mean metastatic lung colony counts when examined on day 30; there were 15.1 colonies in controls and 8.0 in experimental animals (P = .03).

Systemic effects of methylcholanthrene in mice. II. Immune responses to sheep red cells.

Assays of splenic plaque-forming cells (PFC) for sheep red cells (SRC), done in parallel in control and methylcholanthrene (MCA)-injected mice of strains C3H/eB and C57BL/6, disclosed immunodepression, the degree of which depended on strain and sex of mice, and time elapsed between injection of MCA and performance of test. The most severe and long-lasting effects were observed in male C3H/eB mice, and the least impairment was seen in female C57BL/6 mice. After surgical splenectomy for assay of PFC, MCA-injected mice were observed for appearance of induced tumors up to 52 weeks after administration of carcinogen. In most groups of C57BL/6, but only rarely in C3H/eB mice, a correlation was found in individual mice between immunodepression and appearance of tumor. Since, however, the incidence of induced tumors was higher in C57BL/6 than in C3H/eB mice, immunodepression cannot be considered a universally obligatory step in carcinogenesis, but its involvement may be limited to particular phenotypes.

12-0-Tetradecanoylphorbol-13-acetate-induced amplification of mesenchymal tumorigenesis in the mouse skin.

Skin tumors were induced in 6-week-old female Swiss albino mice by a single subcutaneous (SC) injection of 20-methylcholanthrene (MCA) in the right scapular region and the animals were then divided into four groups. Mice in group I did not receive further treatment. Six weeks after MCA injection, those in groups II and III received twice weekly applications of 0.1 ml acetone and 1.8 nmol 12-0-tetradecanoylphorbol-13-acetate (TPA) in 0.1 ml acetone, respectively, at the site of MCA injection until tumor development. Group IV animals were divided into four subsets and administered two, four, six, or eight TPA applications commencing 6 weeks after carcinogen injection. The effect of TPA pretreatment on MCA-induced tumorigenesis was studied in animals in group V. In mice treated with MCA alone, the most predominant mesenchymal tumor type is fibrosarcoma with induction of some rhabdomyosarcomas. Mixed mesenchymal tumors consisting of fibrosarcoma, rhabdomyosarcoma, or hibernoma were observed in only 12% of the animals. The number of animals bearing mixed mesenchymal tumors such as fibrosarcoma, rhabdomyosarcoma, hibernoma, and/or liposarcoma increased to 46% in mice receiving MCA + TPA until tumor development. Interestingly, liposarcomas were not found at all in animals treated with MCA alone. The data indicates that TPA application to precancerous mouse skin enhances mesenchymal tumorigenesis.

Natural resistance mechanisms may play a role in protection against chemical carcinogenesis.

Mice deprived of B lymphocytes by the chronic administration of anti-IgM antibodies have been shown to possess a heightened natural resistance (NR) to micro-organisms, to parental bone marrow, and to natural killer (NK)-sensitive tumors in vitro and in vivo. Experiments described in this communication indicate that the latent period of primary tumors induced by the injection of methylcholanthrene (MC) is also prolonged in these mice. This observation suggests that NR mechanisms may provide protection against primary chemically induced tumors.

Effect of solvents on methylcholanthrene-induced carcinogenesis in mice.

We have shown previously, with X-chromosome linked phosphoglycerate kinase (PGK) cell markers, that the fibrosarcomas induced in mice by subcutaneous injection of 0.2-2.0 mg of methylcholanthrene (MCA) dissolved in olive oil had a multicellular origin. In this study we used a constant dose of MCA injected subcutaneously and compared the effects on tumorigenesis of using benzene or olive oil as the solvents. The benzene and olive oil groups differed in tumor incidence (100% versus 50%), location (muscle versus skin), and type (rhabdomyosarcomas versus fibrosarcomas). Each of the nine olive-oil MCA-induced tumors showed both PGK enzyme types, whereas 13 ot 16 tumors induced by MCA in benzene displayed only a single PGK type. This difference in PGK phenotypes suggests that the MCA-benzene neoplasms arose from one or a few cells. We conclude from these results that the nature of the solvent plays an important role in the type and number of cells from which tumors induced by subcutaneous injection of MCA arise.

Vitamin C acts as a cocarcinogen to methylcholanthrene in guinea-pigs

Large doses of vitamin C were found to promote the induction of sarcomas by methylcholanthrene. The tumours appeared earlier in the group of guinea-pigs treated with vitamin C after injection of methylcholanthrene than in the control group injected only with methylcholanthrene. The treatment of guinea-pigs with vitamin C after induction of tumours thus did not inhibit the growth of tumours.

Effect of prolonged immunization of mice with bovine serum albumin on the incidence of chemically induced tumors.

The effect of prolonged immunization of mice with bovine serum albumin (BSA) on tumors elicited by injection of 3-methylcholanthrene was studied. The incidence of tumors and rate of tumor growth were higher in mice immunized with BSA than in control mice injected with syngeneic serum. The possibility of an immunosuppressive effect of the prolonged immunization with BSA is discussed.

An influence of diet on transplantation immunity.

It was discovered by chance that mice raised under otherwise entirely conventional conditions of husbandry but fed upon autoclaved diet (diet A) had stronger cell-mediated immune reactions than those of mice raised under the same conditions but with an unmodified diet (diet B) : skin allografts were rejected more quickly, transplantation tolerance was more difficult to procure and fibrosarcomas induced by the injection of methylcholanthrene (MCA) arose more slowly and less often. Analysis showed that these findings could be explained at least in part by the discovery of Mertin & Hunt (1976, p. 928) that a partial deprivation of polyinsaturated fatty acids led to an intensification of cell-mediated immunity; on the other hand, experiments with dietary mixtures made it seem unlikely that this was the whole explanation and pointed towards some positive immunopotentiation by an ingredient of autoclaved diet. This, it was proposed, might be a compound of unknown composition resulting from the interaction of vitamin A with other dietary constituents. This interpretation was not supported by direct evidence but by confirming that retinol derivatives, especially retinyl acetate, could exercise an immunopotentiation of the kind and degree under investigation: retinyl acetate could counteract the immunosuppressive action of linoleic acid, though retinyl methyl ether was ineffective. Although retinyl derivatives may protect against MCA tumours by impeding its metabolic conversion to an oncogenic form, the effects of an autoclaved diet upon skin allograft survival, the induction of tolerance and the formation of tumours is probably mediated through an immunological mechanism.

Chemical Carcinogenesis in Nude Mice: Comparison Between Nude Mice From Homozygous Matings and Heterozygous Matings and Effect of Age and Carcinogen Dose<xref ref-type="fn" rid="fn2">2</xref>

The incidence and latency periods for local tumor development after sc injection of 3-methylcholanthrene (MCA) into 30-day-old nude mice (nu/nu partially inbred on the CBA/H background) derived from homozygous matings (nu/nu times nu/nu) or heterozygous matings (nu/+ times nu/+) were comparable and did not differ with the immunologically normal controls, even when the carcinogen dosages ranged from 0.01 to 0.10 mg. Similarly, no differences in tumor incidence or latency periods between nude mice from homozygous or heterozygous matings as well as their immunologically normal controls were observed when weight-adjusted doses of MCA equivalent to 0.02 to 0.10 mg in the 30-day-old mice were administered at 120, 210, or 360 days of age. Tumor incidence was lower in nude mice and normal mice when MCA was administered at 210 and 360 days of age, especially in mice given the lower dose of MCA. The lower dosages of MCA (0.01-0.05 mg) had no detectable immunodepressive effects in normal mice. Thus the "normal" tumor incidence in nude mice after MCA administration could not be attributed to: 1) the effect of humoral thymus gland function (in the nude mice derived from heterozygous matings), 2) the immunodepressive effects of the carcinogen (the lower MCA dosages are not immunodepressive), or 3) the age of the mice at administration. These results argue against the thymus dependency of immunologic surveillance.

Inhibition of chemical carcinogenesis by Corynebacterium parvum.

A course of intravenous injections of Corynebacterium parvum starting 4 days before a single intramuscular injection of methylcholanthrene markedly delayed or prevented the development of fibro-sacromas in CBA mice. Similar treatment starting 4 weeks after injection of the carcinogen did not significantly alter the incidence of tumours. In the light of these findings, and previous work on the effect of C. parvum on macrophage activity and the growth of isogeneic tumour transplants, it is postulated that a transformational event occurs soon after injection of methylcholanthrene which results in the emergence of a clone of cells which are selectively liable to destruction by activated macrophages, but which, if not destroyed, multiply so rapidly that subsequent treat­ment fails to prevent, or even perceptibly to delay, the appearance of a tumour.

Accelerated growth of syngeneic tumors in mice treated with methylcholanthrene.

BALB/c and C3H- mice were given a single injection of methylcholanthrene (MCA) and were tested at various times thereafter for immunologic reactivity and resistance to syngeneic tumor challenge. MCA treatment did not affect the rejection of H-2-incompatible tumor allografts, the rejection of H-2-compatible skin allografts, or the induction of delayed hypersensitivity to DNCB as measured by footpad challenge. In contrast, mice treated with MCA exhibited an increase susceptibility to challenge with several syngeneic ultraviolet-induced and MCA-induced fibrosarcomas. These studies demonstrate that in the autochthonous host the measurement of host immune status by in vivo cell-mediated immune responses to exogenous antigens does not necessarily predict the host response to tumor growth.