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Background: The phenomenon of hormesis is characterized by a biphasic dose-response, exhibiting opposite effects in the low- and high-dose zones. In this study, we explored the possibility that the hormesis concept may describe the interactions between two tumors implanted in a single mouse, such that the resulting tumors are of different sizes. Materials and methods: We used two murine tumors of spontaneous origin and undetectable immunogenicity growing in BALB/c mice. A measure of cell proliferation was obtained by immunostaining for Ki-67 protein and by using the [3H] thymidine uptake assay. For serum fractionation, we utilized dialysis and chromatography on Sephadex G-15. Results: The larger primary tumor induced inhibitory or stimulatory effects on the growth of the smaller secondary one, depending on the ratio between the mass of the larger tumor relative to that of the smaller one, with high ratios rendering inhibition and low ratios inducing stimulation of the secondary tumor. Conclusion: Since metastases can be considered as natural secondary tumor implants in a tumor-bearing host and that they constitute the main problem in cancer pathology, the use of the concept of hormesis to describe those biphasic effects might have significant clinical implications. In effect, if the tumor-bearing host were placed in the inhibitory window, tumor extirpation could enhance the growth of distant metastases and, reciprocally, if placed in the stimulatory window, tumor extirpation would result not only in a reduction or elimination of primary tumor load but also in a slower growth or inhibition of metastases. © 2010 Springer-Verlag. Fil: Bruzzo Iraola, Juan. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina Fil: Chiarella, Paula. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina Fil: Meissl, Roberto Jose. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina Fil: Ruggiero, Raul Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; Argentina

We have previously observed that acellular extracts from necrotic areas (NE) of the non-metastatic murine mammary adenocarcinoma M3, enhance in vitro cell detachment and spontaneous lung metastases. In the present study, using different proteinase inhibitors along with NE, only the calcium chelator EDTA could significantly abrogate the enhanced cell detachment from M3 produced by NE. The typical cleavage products of type IV collagenase were detected inside the tumor necrotic area, mainly in association with necrobiotic cells, as evaluated by Western blot analysis and immunohistochemical assays. Zymography revealed the presence of 72- and 92-kDa gelatinase/type IV collagenase in NE. Moreover, NE increased the in vitro invasive ability of cultured M3 cells. The use of specific antibodies against both 72- and 92-kDa type IV collagenases in the invasion assay showed that only the latter was able to revert the enhanced invasiveness to the baseline. It can be concluded that tumor necrosis is an important source of gelatinase/type IV collagenase, mainly in its 92 kDa form, and plays a major role in tumor invasion.

MPA induces mammary tumors in virgin BALB/c mice with an average latency of 52 weeks. In order to determine whether the simultaneous administration of a chemical carcinogen, N-methyl-N-nitrosourea (MNU), shortened the latency of MPA-induced tumors, a total of 60 virgin female BALB/c mice were treated with either MNU + MPA or MNU or MPA. The experiment lasted 7 months. The incidence and latency of mammary tumors were significantly different between the 3 groups: 15/19 (79%) in MNU + MPA-treated mice with a latency of 154 +/- 19 days; 3/20 (15%) in MNU-treated mice with a latency of 179 +/- 7 days; 0/20 (tumors only start appearing after 10 months) in MPA-treated mice. Histologically, MNU + MPA-induced tumors were similar to the few tumors observed in MNU-treated mice: most of them were type B adenocarcinomas with a high degree of necrosis and calcification. Only one of the MNU + MPA-induced tumors expressed high levels of ER and PR and proved to be MPA-responsive in further passages. All the other tumors showed low or non-detectable levels of ER and PR together with an independent pattern of tumor growth. In MNU-treated mice the only tumor that was transplanted proved to be hormone independent and had low levels of PR and ER. In both MNU and MNU + MPA treated mice lung adenocarcinomas were detected. Cystic uterine glandular hyperplasias were observed in all animals. It can be concluded that MPA and MNU potentiate their carcinogenic effect in mammary gland.

The administration of MPA to virgin female BALB/c mice led to the development of mammary adenocarcinomas, which in further in vivo transplants gave rise to both MPA-dependent and MPA-independent lines. In this paper we chose one of the MPA-dependent lines with high contents of estrogen (ER) and progesterone (PR) receptors, and were able to demonstrate that a) the growth of these tumors could be manipulated by the administration or the withdrawal of the hormonal supply; b) PR were down-regulated in MPA-treated mice; c) progesterone had the same stimulatory effect as MPA on tumor growth; d) tumors did not grow in estrogen-treated mice; e) tumor growth was much lower in males than in females; f) the presence of the ovaries had a positive influence on tumor growth, even in the presence of MPA; g) the withdrawal of progestin pellets in ovariectomized mice usually led to complete remissions followed by regrowth of the tumors after several weeks; and h) the regrowing tumors maintained their steroid receptor pattern and (in 3 out of 4 cases) their hormone-dependent behavior in further passages.

Introduction: It has been demonstrated that leukemia inhibitory factor (LIF) induces epithelium apoptosis through Stat3 activation during mouse mammary gland involution. In contrast, it has been shown that this transcription factor is commonly activated in breast cancer cells, although what causes this effect remains unknown. Here we have tested the hypothesis that locally produced LIF can be responsible for Stat3 activation in mouse mammary tumors. Methods: The studies were performed in different tumorigenic and non-tumorigenic mammary cells. The expression of LIF and LIF receptor was tested by RT-PCR analysis. In tumors, LIF and Stat3 proteins were analyzed by immunohistochemistry, whereas Stat3 and extracellular signal-regulated kinase (ERK)1/2 expression and phosphorylation were studied by Western blot analysis. A LIF-specific blocking antibody was used to determine whether this cytokine was responsible for Stat3 phosphorylation induced by conditioned medium. Specific pharmacological inhibitors (PD98059 and Stat3ip) that affect ERK1/2 and Stat3 activation were used to study their involvement in LIF-induced effects. To analyze cell survival, assays with crystal violet were performed. Results: High levels of LIF expression and activated Stat3 were found in mammary tumors growing in vivo and in their primary cultures. We found a single mouse mammary tumor cell line, LM3, that showed low levels of activated Stat3. Incidentally, these cells also showed very little expression of LIF receptor. This suggested that autocrine/paracrine LIF would be responsible for Stat3 activation in mouse mammary tumors. This hypothesis was confirmed by the ability of conditioned medium of mammary tumor primary cultures to induce Stat3 phosphorylation, activity that was prevented by pretreatment with LIF-blocking antibody. Besides, we found that LIF increased tumor cell viability. Interestingly, blocking Stat3 activation enhanced this effect in mammary tumor cells. Conclusion: LIF is overexpressed in mouse mammary tumors, where it acts as the main Stat3 activator. Interestingly, the positive LIF effect on tumor cell viability is not dependent on Stat3 activation, which inhibits tumor cell survival as it does in normal mammary epithelium. © 2007 Quaglino et al.; licensee BioMed Central Ltd. Fil:Quaglino, A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Schere-Levy, C. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Romorini, L. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Kordon, E.C. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.

Background Chronic injury deregulates cellular homeostasis and induces a number of alterations leading to disruption of cellular processes such as cell cycle checkpoints and apoptosis, driving to carcinogenesis. The stress protein heme oxygenase-1 (HO-1) catalyzes heme degradation producing biliverdin, iron and CO. Induction of HO-1 has been suggested to be essential for a controlled cell growth. The aim of this work was to analyze the in vivo homeostatic response (HR) triggered by the withdrawal of a potent carcinogen, p-dimethylaminoazobenzene (DAB), after preneoplastic lesions were observed. We analyzed HO-1 cellular localization and the expression of HO-1, Bcl-2 and cell cycle related proteins under these conditions comparing them to hepatocellular carcinoma (HC). Methods The intoxication protocol was designed based on previous studies demonstrating that preneoplastic lesions were evident after 89 days of chemical carcinogen administration. Male CF1 mice (n = 18) were used. HR group received DAB (0.5 % w/w) in the diet for 78 days followed by 11 days of carcinogen deprivation. The HC group received the carcinogen and control animals the standard diet during 89 days. The expression of cell cycle related proteins, of Bcl-2 and of HO-1 were analyzed by western blot. The cellular localization and expression of HO-1 were detected by immnunohistochemistry. Results Increased expression of cyclin E/CDK2 was observed in HR, thus implicating cyclin E/CDK2 in the liver regenerative process. p21cip1/waf1 and Bcl-2 induction in HC was restituted to basal levels in HR. A similar response profile was found for HO-1 expression levels, showing a lower oxidative status in the carcinogen-deprived liver. The immunohistochemical studies revealed the presence of macrophages surrounding foci of necrosis and nodular lesions in HR indicative of an inflammatory response. Furthermore, regenerative cells displayed changes in type, size and intensity of HO-1 immunostaining. Conclusion These results demonstrate that the regenerative capacity of the liver is still observed in the pre-neoplastic tissue after carcinogen withdrawal suggesting that reversible mechanism/s to compensate necrosis and to restitute homeostasis are involved.

Experimental autoimmune orchitis (EAO) induced in Swiss mice could be reduced by means of the utilization of micronized frustules of fossil diatoms (DS) containing 54% of SiO2. Experimental mice were sensitized with testicular Antigen (Ag) in Complete Freund’s Adjuvant (CFA) inoculated twice, on day 0 and day 21. 100 μg of DS suspension was inoculated into sensitized mice 10 times, once every 4 days, subcutaneously, starting on day 7 after the first Ag inoculation. Mice receiving the DS treatment showed a diminution of the delayed hypersensitivity reaction, lower antibody titer and decreased incidence of testicular injury as well as reduced grade and extension of the lesions. Possible explanation of these results would suggest alteration of monocyte and/or macrophage normal behaviour as well as alteration of antibody synthesis by different mechanisms.

Counter-irritation (CI) triggered by different non-specific irritant stimuli delayed the growth of a murine tumour of non-detected immunogenicity. The syngeneic LB tumour transplant by itself also induced CI and decreased the number of leukocytes migrating to a secondary s.c. irritant stimulus, e.g. sponge or carrageenan. On the other hand, partial inhibition of cell migration by treatment with either 0.5 mg kg-1 indomethacin or 0.3 mg kg-1 piroxicam retarded LB tumour growth, presumably by a mechanism unrelated to inhibition of immune responses by PGE2. It is suggested that CI may play a role in the early stages of concomitant resistance. Images Figure 5

The non-metastatic murine mammary adenocarcinoma M3 and its metastatic variant MM3 were used to evaluate the role of intratumoral necrosis in cell detachment and metastasis. Accelular extracts from necrotic areas of both tumors increased in vitro cellular detachment from M3 but not from MM3 fragments. Furthermore, the in vivo inoculation of the necrotic extracts within non-metastatic M3 tumors gave rise to pulmonary metastases. Histological studies revealed in M3 a central necrosis limited by an uninterrupted peripheral ring of well preserved cells, while in MM3 necrotic and non-necrotic areas alternated. It is concluded that the distribution of necrosis within the primary tumor by facilitating cell detachment is, at least in part, responsible for the development of metastases.

Various immunization assays were used to demonstrate the lack of immunogenicity of three BALB/c tumours of spontaneous origin and of a fourth one resulting from foreign body tumorigenesis. All four tumours inhibited the growth of a second implant of the same tumour into the contralateral flank. In our tumour models "concomitant immunity" (1) was not mediated by macrophage or T-cell dependent immune reactions: both thymectomized BALB/c and nude mice (treated or untreated with silica) gave the same results as intact mice; (2) showed some degree of non-specificity, inhibiting the growth of a different tumour in 3/4 cases; though, the existence of a specific component could not be discarded; (3) was proportional to the volume of the primary tumour at the time of the second challenge; (4) was dependent on actively growing primary tumour, not being obtained with progressively increasing daily inocula of irradiated tumour cells; (5) was detectable in an actively growing secondary tumour; recurrent growth after partial surgical excision was inhibited and (6) involved cytostasis of the secondary tumour: a syngeneic graft of the overlying skin led to tumour growth while histological studies revealed the presence of viable tumour cells. It is postulated that "concomitant immunity" or resistance can be generated without the active participation of the immune system and that tumour-related factors are, in certain cases, responsible for blocking the growth of secondary tumours. Images Figure 5