A leishmaniose é uma doença causada por parasitos do gênero Leishmania e transmitida por insetos flebotomíneos. O objetivo deste trabalho foi comparar o perfil da infecção por L. amazonenses em BALB/c, BALB/c nude e C57BL/6, e avaliar a expressão das proteínas CP, LACK, LRR17, metacaspase, PDI e STI, em amastigotas isolados de BALB/c e BALB/c nude. Observamos em 13 semanas o aumento mais precoce da espessura da pata em BALB/c do que em BALB/c nude. A carga parasitária nos animais nude foi superior à dos demais. Por outro lado, animais C57BL/6 controlaram a infecção a partir da sexta semana. A análise da expressão gênica nas lesões mostrou um padrão misto Th1/Th2 em todos os animais, e maior expressão de IFN-g, IL-4, IL1b e iNOS em BALB/c. Em baço e linfonodo observamos baixa expressão de CD3e e elevada expressão de iNOS em BALB/c nude. A análise por FACs mostrou em baço e linfonodo a esperada baixa porcentagem de células TCD4 e T CD8 em BALB/c nude, no entanto com capacidade de produzir IFN-g e IL-4. A LACK e a metacaspase estão aumentadas nos parasitas isolados de BALB/c nude. A metacaspase apresentou maior atividade também em nude. Não observamos diferença na expressão da LACK e metacaspase em amastigotas isolados de macrófagos infectados na presença de IFN-g e IL-4. / Leishmaniasis is caused by parasites of Leishmania genus, transmitted by phlebotomine sandflies. The aim of this study was to compare the profile of the infection by L. amazonenses in BALB/c, BALB/c nude and C57BL/6 and to analyze the expression of CP, LACK, LRR17, metacaspase, PDI and STI proteins, in amastigotes isolated from BALB/c and BALB/c nude lesions. We observed during 13 weeks an early increase in footpad thickness in BALB/c compared to BALB/c nude. Parasite loads in nude mice were higher than the others. C57BL/6 mice controlled infection since the sixth week. Gene expression analysis of the lesions indicated a mixed Th1/Th2 pattern in all mice, and higher expression of IFN-g, IL-4, IL1b and iNOS in BALB/c. In spleens and lymph nodes we observed lower expression of CD3e and high expression of iNOS in BALB/c nude. FACs analysis of spleens and lymph nodes showed low percentages of TCD4 and T CD8 cells in BALB/c nude, but producing IFN-g and IL-4. LACK and metacaspase are increased in parasites isolated from BALB/c nude, and metacaspase had higher enzymatic activity in amastigotes from nude. No differences in LACK or metacaspase in amastigotes isolated from macrophages infected in the presence of IFN-g and IL-4 were observed.

A Baccharis coridifolia é uma planta tóxica da família Compositae, gênero Baccharis L., conhecida popularmente como \"mio-mio\", a qual é encontrada no sul do Brasil e também em algumas regiões do Estado de São Paulo. Em bovinos, a intoxicação provoca lesões necróticas no trato gastrintestinal e nos tecidos linfóides sólidos em geral, com exceção do timo. Já em camundongos, a B. coridifolia promove, além das lesões em órgãos linfóides, lesões necróticas também no timo. Os princípios ativos da B. coridifolia foram isolados e identificados como sendo os tricotecenos macrocíclicos: roridinas A, D e E; verrucarinas A e J e miotoxina A. Em relação ao mecanismo de ação, trabalhos recentes in vitro demonstraram que as roridinas e as verrucarinas, promovem apoptose em tecido linfóide. Deste modo, considerando a toxicidade desta planta aos tecidos linfóides, o objetivo do presente estudo foi investigar se linfócitos tumorais (linfoma) seriam mais sensíveis aos efeitos tóxicos da B. coridifolia que linfócitos normais. Para tal, fez-se um experimento in vitro com culturas de linfócitos provenientes do baço e timo de camundongos hígidos, bem como células do linfoma murino A20 e carcinoma mamário de Ehrlich tratadas com o Resíduo Hexânico (RH) obtido do extrato etanólico de B. coridifolia por 24 horas. A viabilidade celular foi verificada pela técnica MTT, na qual se evidenciou diminuição da viabilidade das duas linhagens tumorais tratadas, linfoma A20 e carcinoma de Ehrlich, sem alteração na viabilidade dos linfócitos normais. Para melhor caracterizar este efeito tóxico do RH em linfócitos tumorais, camundongos nude (NMRI-nu/nu) foram inoculados com células A20, sendo posteriormente tratados por via oral, durante sete dias, com RH, associado ou não a uma aplicação intraperitoneal do quimioterápico ciclofosfamida no primeiro dia de tratamento. Resultados mostraram que os animais tratados com RH foram menos afetados com a ocorrência de metástase do linfoma primário para outros órgãos e que as involuções tumorais foram mais expressivas no grupo submetido à associação deste tratamento com uma aplicação de ciclofosfamida. Estes resultados permitem sugerir que linfócitos tumorais são mais suscetíveis aos efeitos tóxicos da B. coridifolia. / Baccharis coridifolia is a toxic plant from Compositae family, genus Baccharis L., known as \"mio-mio\". It is found in the south of Brazil, as well as in some regions of São Paulo state. In bovines, its intoxication leads to necrotic injuries to gastrointestinal tract and linfoid tissues. In mice, B. coridifolia leads, apart from linfoid solids in general to necrotic injuries in thymus. B. coridifolia\'s active principles were isolated and identified as being macrocyclic tricotecenes: roridines A, D and E; verrucarines A and J and miotoxine A. Related to action pathway, recent research (in vitro) has shown that roridines and verrucarines, B. coridifolia\'s active principles, lead to apoptosis in linfoid tissues. Therefore, considering this plant\'s toxicity to linfoid tissues, this work aimed the study of whether tumoral lymphocytes (lymphoma) would be more sensitive to B. coridifolia\'s effects than regular lymphocytes. In this sense, an experiment was carried out (in vitro) using healthy mice\'s lymphocytes from spleens and thymus, as well as cells from murine A20 lymphoma and Ehrlich breast carcinoma treated with a residue (RH, hexane obtained) from a B. coridifolia\'s ethanol effusion (24 hours). The cell viability was tested using MTT technique, where a decrease in viability was found for treated tumoral cells lineages, A 20 lymphoma and Ehrlich carcinoma, with no changes in normal lymphocytes viability. To find a better characterization to RH\'s toxic effect in tumoral lymphocytes, nude mice (NMRI-nu/nu) were inoculated with A 20 cells and then were orally treated, during 7 days, with RH, and, in some cases, with intraperitonial injection of cyclophosphamide, during the first day of treatment. Results showed that RH treated animals were less affected to primary lymphoma metastasis in other organs and tumoral involutions were more prominent in the group treated with cyclophosphamide concomitantly. These results suggest that tumoral lymphocytes are more susceptible to B. coridifolia\'s toxic effects.

Dietary factors may affect development and progression of prostate cancer. Experimental and epidemiological studies have suggested an effect of phytoestrogens on prostate cancer. Lignans are the predominant phytoestrogen in a Western diet. The effects of a diet rich in phytoestrogens and in particular lignans, as compared to a control diet, were assessed in several prostate cancer models. In paper I, 70 athymic nude mice with transplanted subcutaneous LNCaP tumours, an androgen sensitive human prostate cancer cell line, were fed one out of six phytoestrogen rich diets or a control diet after tumour injection. The rye diet, with high lignan content, decreased tumour take and growth, decreased secretion of prostate specific antigen and increased apoptosis. Addition of fat to the rye diet decreased the beneficial effects. In paper II, transgenic mice designed to develop prostate cancer (TRAMP) were fed rye bran or a control diet from the age of four weeks. Rye bran decreased prostate epithelial cell volume by 20%, and increased cell apoptosis by 31% as compared to the control diet. In paper III, we examined the effects of 7-hydroxymatairesinol (HMR), a purified lignan, in nude mice with subcutaneous LNCaP tumours in two different concentrations as compared to a control diet. Mice on the HMR diets had a reduced tumour take rate, lower total tumour volume, increased proportion of non-growing tumours, and increased apoptosis as compared to the control diet. Paper IV was a three week intervention study exploring the effects of rye bran bread vs. a control diet in men with prostate cancer. The men in the rye group had increased levels of plasma enterolactone and in biopsies from the prostate after the intervention an increase in apoptosis was observed in comparison with biopsies obtained before the intervention. In paper V, we examined the association between plasma levels of enterolactone, and risk of prostate cancer in a nested case control study. In the Northern Sweden Health and Disease Cohort, enterolactone concentrations were measured in plasma obtained at a mean time of 5 years before diagnosis from 265 cases of prostate cancer, and from 525 matched controls. We found no significant association between plasma enterolactone and risk of prostate cancer. Men with very low enterolactone levels (bottom decile) however, had significantly higher risk of prostate cancer. Phytoestrogen rich diet including soy, rye bran, substances purified from rye, and a purified lignan (HMR) all inhibited prostate tumour growth. However, it cannot be concluded that the effects observed were due solely to lignans as other components in rye grain such as tannins, phytic acid, ferulic acid, vitamins and minerals may have contributed to the beneficial effects. Thus, additional studies are needed to further elucidate the effects of phytoestrogens on prostate cancer development and progression.

Sex steroids are inevitable in women. However, long-term exposure to sex steroids increases the risk of breast cancer. A complete understanding of sex steroid control of the breast and how it relates to breast cancer risk is still lacking. Angiogenesis and proteolytic enzyme activity are crucial for the process by which tumors evolve into a vascularized, invasive phenotype. Matrix metalloproteinases are potent matrixdegrading enzymes that affect several steps in tumor progression including angiogenesis. In the female reproductive organs, sex steroids regulate angiogenesis and MMP activity, yet little is known how sex steroids affect these crucial events in normal and malignant breast tissue. This thesis elucidates a link between sex steroids, MMP activity, and angiogenesis. It is shown that estradiol down-regulates while tamoxifen up-regulates the protein expression and activity of MMP-2 and MMP-9 in human breast cancer cells in vitro and in human breast cancer xenografts in vivo. The results further suggest that a biological consequence of this regulation may be modulation of tumor angiogenesis. The net effect of adding tamoxifen to estradiol treatment was an increase in extracellular levels of the endogenous angiogenesis inhibitor endostatin and decreased levels of the tumor promoter TGF-β1 compared to estradiol treatment only. This was accompanied by reduced vasculature and decreased tumor growth. Similarly, a regulatory effect of estradiol and tamoxifen on endostatin generation was observed in normal human breast tissue by whole-tissue culture and microdialysis in human breast tissue in situ. In conclusion, the results presented in this thesis suggest previously unknown mechanisms of action of estradiol and tamoxifen in breast cancer and in normal human breast tissue, and novel means by which estradiol may tip the scale to favor angiogenesis. This knowledge may be important for the understanding of sex steroid dependent breast carcinogenesis and in the future development of tissue-specific preventive as well as therapeutic strategies against breast cancer.

A Baccharis coridifolia é uma planta tóxica da família Compositae, gênero Baccharis L., conhecida popularmente como \"mio-mio\", a qual é encontrada no sul do Brasil e também em algumas regiões do Estado de São Paulo. Em bovinos, a intoxicação provoca lesões necróticas no trato gastrintestinal e nos tecidos linfóides sólidos em geral, com exceção do timo. Já em camundongos, a B. coridifolia promove, além das lesões em órgãos linfóides, lesões necróticas também no timo. Os princípios ativos da B. coridifolia foram isolados e identificados como sendo os tricotecenos macrocíclicos: roridinas A, D e E; verrucarinas A e J e miotoxina A. Em relação ao mecanismo de ação, trabalhos recentes in vitro demonstraram que as roridinas e as verrucarinas, promovem apoptose em tecido linfóide. Deste modo, considerando a toxicidade desta planta aos tecidos linfóides, o objetivo do presente estudo foi investigar se linfócitos tumorais (linfoma) seriam mais sensíveis aos efeitos tóxicos da B. coridifolia que linfócitos normais. Para tal, fez-se um experimento in vitro com culturas de linfócitos provenientes do baço e timo de camundongos hígidos, bem como células do linfoma murino A20 e carcinoma mamário de Ehrlich tratadas com o Resíduo Hexânico (RH) obtido do extrato etanólico de B. coridifolia por 24 horas. A viabilidade celular foi verificada pela técnica MTT, na qual se evidenciou diminuição da viabilidade das duas linhagens tumorais tratadas, linfoma A20 e carcinoma de Ehrlich, sem alteração na viabilidade dos linfócitos normais. Para melhor caracterizar este efeito tóxico do RH em linfócitos tumorais, camundongos nude (NMRI-nu/nu) foram inoculados com células A20, sendo posteriormente tratados por via oral, durante sete dias, com RH, associado ou não a uma aplicação intraperitoneal do quimioterápico ciclofosfamida no primeiro dia de tratamento. Resultados mostraram que os animais tratados com RH foram menos afetados com a ocorrência de metástase do linfoma primário para outros órgãos e que as involuções tumorais foram mais expressivas no grupo submetido à associação deste tratamento com uma aplicação de ciclofosfamida. Estes resultados permitem sugerir que linfócitos tumorais são mais suscetíveis aos efeitos tóxicos da B. coridifolia. / Baccharis coridifolia is a toxic plant from Compositae family, genus Baccharis L., known as \"mio-mio\". It is found in the south of Brazil, as well as in some regions of São Paulo state. In bovines, its intoxication leads to necrotic injuries to gastrointestinal tract and linfoid tissues. In mice, B. coridifolia leads, apart from linfoid solids in general to necrotic injuries in thymus. B. coridifolia\'s active principles were isolated and identified as being macrocyclic tricotecenes: roridines A, D and E; verrucarines A and J and miotoxine A. Related to action pathway, recent research (in vitro) has shown that roridines and verrucarines, B. coridifolia\'s active principles, lead to apoptosis in linfoid tissues. Therefore, considering this plant\'s toxicity to linfoid tissues, this work aimed the study of whether tumoral lymphocytes (lymphoma) would be more sensitive to B. coridifolia\'s effects than regular lymphocytes. In this sense, an experiment was carried out (in vitro) using healthy mice\'s lymphocytes from spleens and thymus, as well as cells from murine A20 lymphoma and Ehrlich breast carcinoma treated with a residue (RH, hexane obtained) from a B. coridifolia\'s ethanol effusion (24 hours). The cell viability was tested using MTT technique, where a decrease in viability was found for treated tumoral cells lineages, A 20 lymphoma and Ehrlich carcinoma, with no changes in normal lymphocytes viability. To find a better characterization to RH\'s toxic effect in tumoral lymphocytes, nude mice (NMRI-nu/nu) were inoculated with A 20 cells and then were orally treated, during 7 days, with RH, and, in some cases, with intraperitonial injection of cyclophosphamide, during the first day of treatment. Results showed that RH treated animals were less affected to primary lymphoma metastasis in other organs and tumoral involutions were more prominent in the group treated with cyclophosphamide concomitantly. These results suggest that tumoral lymphocytes are more susceptible to B. coridifolia\'s toxic effects.

Thesis (MMedSc )--University of Stellenbosch, 2011. / ENGLISH ABSTRACT: The present study investigated in particularly the specific nature of the supporting stromal layer located between the implanted human cyst and host murine tissue, which has yet to be reported. During an initial phase of this study, the particular light microscopic properties of the existing hematoxylin and eosin (H&E) stained experimental cyst was investigated, with regards to the presence or absence of specific morphological features, namely spongiosis, exocytosis, epithelial keratinization, epithelial thickness and hyperplasia, and the vascularity and fibrosis present in the stroma of these experimental sections. Subsequent analysis reported significant spongiosis, in addition to increased exocytosis of immune cells and epithelial keratinization in a number of cysts. Additionally, increased epithelial thickness and hyperplasia was reported in only 2 / 10 experimental tissues, whereas increased vascularity was observed in the stroma following analysis of H&E and Special staining, such as Verhoeff-von Gieson and Masson trichrome results. During the second phase of the study, immunohistochemical analysis with a particularly wide array of antibodies raised against specific human and mouse antigens had been applied. This involved automated immunohistochemical staining with mouse anti-human primary antibodies, in addition to manual staining with rabbit anti-mouse primary antibodies. Subsequent visualization was achieved by means of linking to biotinylated secondary antibodies, and Streptavidin-HRP incubation for standard visualization, followed by counterstaining with Hematoxylin. Maintained positive expression of cytokeratins 5, 13, and 14 was demonstrated in both control human vaginal mucosa and experimental cysts, whereas similar findings were not reported for cytokeratin 1, given the vast keratinization which was observed. Human collagen type IV and laminin of the basement membrane reported positive expression in 9 / 10 and 6 / 10 control human vaginal mucosa tissues respectively. In comparison, negative mouse collagen type IV and laminin was reported in most experimental cysts compared to positive staining in positive control mouse tissues. Immunohistochemical staining for human elastin, fibronectin, von Willebrand factor, and fibroblasts revealed maintained positive staining in all control human vaginal mucosa and experimental cysts. However, maintained expression of CD34 (endothelial marker), CD1a (langerhans cells), and human VEGFR-3 in experimental cysts was not demonstrated, compared to positive expression in control human vaginal mucosa. Subsequent analysis of murine antigens illustrated uniformly negative staining for mouse fibronectin, langerhans cells (CD207), and fibroblasts, in addition to negative staining in positive control mouse tissue sections. Furthermore, negative staining for mouse VEGFR-2 was reported in all experimental cysts; however strong positive staining of this marker in mouse kidney tissue had been reported. The findings of this study suggested that the exact nature of the stromal layer is of both human and murine origin. Furthermore, the tissue region located beneath the human vaginal epithelium is suggested to be of human nature, whereas the second distinct region located at the periphery of experimental cyst tissues, is suggested to be murine origin; however the findings of immunohistochemical analysis could not illustrate definitively the exact nature of the intermediate stromal layer, but could in fact demonstrate a mixture of human and murine tissue. / AFRIKAANSE OPSOMMING: Die huidige studie het die spesifieke molekulêre en histologiese eienskappe van die stromale laag geleë tussen menslike sist- en muis velweefsel bestudeer, wat tans nog nie bekend is nie. Gedurende die eerste fase van hierdie studie is die besondere lig-mikroskopiese eienskappe van die bestaande hematoksilien en eosien (H&E) eksperimentele siste bestudeer, met betrekking tot die aan- of afwesigheid van spesifieke morfologiese eienskappe, naamlik spongiose, eksositose van immuunselle, epiteel keratinisasie, epiteel dikte en hiperplasie, en laastens die stromale vaskulariteit en fibrose. Gevolglike analise het daarop gedui dat beduidende spongiose, eksositose en epiteel keratinisasie gevind word in die eksperimentele siste in vergelyking met kontrole vaginal weefsel. Hierteenoor is die verdikking van die epiteel en hiperplasie in slegs 2 / 10 eksperimentele siste gevind, terwyl vermeerderde vaskulariteit aangedui is na gevolglike H&E en spesiale (soos byvoorbeeld Verhoeff-von Gieson en Masson trichrome) kleuringsresultate. Die tweede fase van die studie het die immunokleuring met verskeie mens- en muis spesifieke antiliggame behels, waarby die uitdrukking van verskeie mens antigene vergelyk is met dié van muis. As sulks is ge-automatiseerde immunohistochemie toegepas met muis primêre antiliggame, tesame met fisiese kleuring met konyn primêre antiliggame toegepas. Gevolglike visualisasie is aangedui deur middel van binding met sekondêre antiliggaam en Streptavidin- HRP, gevolg deur teenkleuring met Hematoksilien. Algehele behoud van positiewe uitdrukking van sitokeratien 5, 13, en 14 is bevind, terwyl sitokeratien 1 uitdrukking nie daarwerklik vergelykbaar is met dié van kontrole mens vaginale weefsel nie. Die uitdrukking van mens kollageen IV en laminien van die basaal membraan is verder bestudeer, en het egter positiewe kleuring in 9 / 10 en 6 / 10 van kontrole mens vaginale mukosa aangedui. In vergelykking hiermee kon die huidige bevindings egter net positiewe kleuring in 4 / 10 en 3 / 10 eksperimentele siste vir kollageen IV en laminien onderskeidelik, illustreer. Immunohistochemiese analise van menslike elastien, fibronektien, von Willebrand (vW) faktor en fibroblaste het op deurgaans positiewe uitdrukking van hierdie merkers aangedui in beide eksperimentele en kontrole menslike weefsel. In teenstelling hiermee is volgehoue uitdrukking van CD34 (endoteel merker), CD1a (Langerhans sel merker) en mens VEGFR-3 in ekperimentele siste egter nie illustreerbaar nie, in vergelykking met deurgaans positiewe uitdrukking van hierdie antigene in kontrole mens vaginale mukosa. In opvolging is deurgaans negatiewe uitdrukking van muis fibronektien, langerhans sel (CD207) en fibroblaste bevestig, terwyl negatiewe kleuring ook deurgaans in positiwe kontrole muis weefsel, bekom deur die disseksie van ‘n naakte muis, gevind is. Verder is ook negatiewe kleuring vir VEGFR-2 in alle eksperimentele siste gevind, terwyl egter sterk positiewe kleuring in muis nierweefsel as positiewe weefsel gevind is. Die resultate van die huidige studie het daarop gedui dat die stromale laag onderliggend tot mens vaginale epiteel van menslike oorsprong is, terwyl die periferale stroma onderliggend tot muis velweefsel, ongetwyfeld van muis oorsprong is. Laastens kon die spesifieke oorsprong van die tussenliggende stroma nie aangedui word nie, maar dat dit moontlik uit beide menslike- en muisweefsel bestaan.

Les cellules Natural Killer (NK), actrices majeures de la vigilance anti-tumorale, sont modulées par des facteurs nutritionnels et métaboliques. L'inhibition de leur activité favorise le développement tumoral. Un régime alimentaire hypercalorique induisant l'obésité est un facteur de risque de développer un cancer du sein. Au niveau du micro-environnement tumoral mammaire, la biodisponibilité en certaines molécules contrôle non seulement les cellules néoplasiques mais, également les cellules immunes infiltrées. Ainsi, la leptine, sécrétée à forte concentration par les adipocytes mammaires, pourrait favoriser la croissance tumorale et altérer les cellules NK. L'arginine fortement consommée par les cellules tumorales et les cellules suppresseurs dérivées des myéloïdes pourrait faire défaut aux cellules NK. L'objectif de cette thèse est de caractériser les activités fonctionnelles et métaboliques des cellules NK en situation de stress nutritionnel. Dans un premier temps, nous avons exploré, in vivo, l'impact d'un régime hypercalorique sur l'activité des cellules NK et sur le développement tumoral mammaire. Ensuite, nous avons cherché à identifier les potentielles altérations fonctionnelles des cellules NK en mimant, in vitro, les conditions retrouvées au niveau du micro-environnement tumoral telles que la présence de concentration élevée en leptine et la déplétion en arginine. Des souris Balb-c "nude" femelles ont été soumises à un régime hypercalorique (HC) versus une diète normo-calorique (NC) pendant 6 mois. Au bout de 5 mois, des cellules tumorales mammaires (MCF-7 ; groupes NCT et HCT) ou le véhicule (groupes NC et HC) ont été implantés au niveau de la quatrième paire de glandes mammaires. Sous régime HC, le développement tumoral s'accompagne d'une perte de masse grasse, de masse maigre et de poids corporel avec un volume et un poids de tumeur augmentés. Cette diète induit au niveau tumoral une sur-expression des ARNm d'enzymes impliquées dans la glycolyse et une sous-expression des acteurs du cycle de Krebs. Sous régime HC, l'expression de la caspase 3 clivée et des récepteurs des oestrogènes β et de la progestérone est réduite alors que celle du Ki67 est accrue. Les cellules NK des souris HC ont une cytotoxicité diminuée. Bien que la présence de tumeur stimule l'activité lytique des cellules NK, la cytotoxicité de ces cellules reste inférieure dans le groupe HCT comparativement à celle du groupe NCT. La leptine stimule, in vitro, de façon dose-dépendante l'activité métabolique des cellules NK. A fortes concentrations, elle active leur cytotoxicité vis-à-vis des cellules cibles MDA-MB-231. Cet effet passe par une stimulation de l'expression de TRAIL et de l'IFN-γ par les cellules NK. En revanche, vis-à-vis des cellules cibles MCF7, les cellules NK présentent une activité lytique réduite en présence de fortes concentrations de leptine, probablement en lien avec une réduction de l'expression de la perforine. En réponse à une déplétion en arginine dans le milieu de culture, la prolifération et la cytotoxicité des cellules NK sont abaissées. L'altération de la reconnaissance des cellules cibles par les récepteurs NKp46 et NKp30, la moindre transmission du signal activateur par la chaine ζ et la faible production d'IFN-γ peuvent expliquer l'inhibition de la cytotoxicité des cellules NK. Ainsi, un apport énergétique élevé favorise le développement tumoral mammaire notamment eninhibant la cytotoxicité des cellules NK. De plus, la leptine à fortes concentrations stimule ou réduit, in vitro, la cytotoxicité des cellules NK selon la nature des cellules cancéreuses mammaires cibles. Une déplétion en arginine, in vitro, quant à elle, inhibe la prolifération et la cytotoxicité des cellules NK. Ces travaux contribuent à mieux comprendre l'impact du micro-environnement sur la réponse antitumorale des cellules NK. / Natural killer (NK) cells are critical mediators of anti-tumor immunity. A high-calorie diet inducing obesity is associated with breast cancer development. NK cells are modulated by dietary and metabolic factors and a decrease in their lytic activity promotes mammary tumor development. In the breast microenvironment, high concentration of leptin can be secreted by mammary adipocytes and thereby could stimulate tumor growth and control immune cells. Arginine, strongly consumed by tumor and myeloid-derived suppressor cells, could be lacking to NK cells. The aim of this work is to characterize the functional and metabolic activities of NK cells in response to nutritional stress. Initially, we explored in vivo the impact of a high-calorie diet on NK cells activity and mammary tumor development. Then, we identified potential functional alterations in NK cells by mimicking the conditions found in the tumor microenvironment such as the presence of high leptin concentration and arginine depletion. Female Balb-c nude mice were fed a high-caloric diet (HC) versus a standard caloric diet (SC) for 6 months. After five months, mammary tumor cells (MCF-7, SCT, HCT) or MatrigelTM (SC, HC) were implanted into the fourth mammary fat pads. The tumor development in HC diet-fed mice was associated with a decrease in body weight, body fat and lean mass and an increase in volume and weight of tumors. This diet induced tumor over-expression, at the transcriptional level, of enzymes involved in glycolysis and a down-expression of citrate cycle actors. Protein tumor levels of cleaved caspase 3, estrogen β and progesterone receptors were reduced while Ki67 was increased in the HC diet-fed mice. NK cell cytotoxicity of HC diet-fed mice was reduced. Although the presence of tumor stimulated NK cell lytic activity, this later was lower in the HCT group compared to the one of SCT mice. In vitro, leptin stimulated, in dose-dependent manner, the metabolic activity of NK cells. High leptin concentrations enhanced NK cell cytotoxicity against the MDA-MB-231 target cells. This phenomenon involved the increase of expression of TRAIL and IFN-γ in NK cells. However, against the MCF-7 target cells, NK cell lytic activity was reduced in the presence of high concentrations of leptin, probably in link to the decreased perforin expression. NK cell proliferation and cytotoxicity were impaired in response to arginine depletion. This inhibition of NK cell cytotoxicity could be linked to a low target cells recognition by NKp46 and NKp30, a reduced activating signal transmission by ζ chain and a low production of IFN-γ. Thus, high energy intake promotes mammary tumor development in particular by inhibiting NK cell cytotoxicity. In vitro, high leptin concentrations stimulate or reduce NK cell cytotoxicity according to the breast cancer cell targets. Furthermore, arginine depletion inhibits NK cell proliferation and cytotoxicity in vitro. These findings provide insight into the microenvironment impacts on NK cell antitumor response in tumor development.

Introdução. O câncer é responsável por uma das principais causas de morte entre mulheres e nota-se um aumento na sua incidência. A linhaça tem sido preconizada como capaz de reduzir o risco do câncer de mama. Objetivo. Verificar o efeito de dietas ricas em linhaça marrom e dourada na evolução do câncer de mama. Métodos. Células cancerosas mamárias humanas (tipo MCF-7) foram inseridas no tecido subcutâneo de camundongos fêmeas nude. Quando os tumores já puderam ser claramente identificados, após 7 semanas, os animais constituíram 3 grupos, um com dieta controle e os outros dois com as variedades de linhaça marrom e dourada. As sementes de linhaça foram moídas para produção de farinhas que foram utilizadas na suplementação das dietas oferecidas aos animais. O crescimento dos tumores implantados nos animais foi acompanhado por oito semanas. Resultados. As linhaças das variedades marrom e dourada mostraram-se eficazes na diminuição da taxa de crescimento dos tumores MCF-7, não sendo verificada nenhuma diferença entre elas. Conclusão. Ambas variedades de linhaça reduziram o crescimento dos tumores. Assim, o consumo de linhaça deve ser estimulado a fim de diminuir a incidência do câncer de mama. / Introduction. Breast cancer is one of the main causes of death among women and an increase in its incidence has been noticed. There is evidence that flaxseed intake could reduce breast cancer risk. Objective. To verify the effect of brown and golden flaxseed rich diets on breast cancer. Methods. Cancer cells of the MCF-7 line were injected subcutaneously into athymic mice. After seven weeks, when tumors were already established, the animals were randomly divided in three groups: (1) control, fed on a basal diet; (2) and (3) mice fed on the basal diet supplemented with 10% brown and with 10% golden flaxseed, respectively. Flaxseed was freshly ground for production of the diets offered to the animals. Tumor growth was monitored weekly for eight weeks. Results. The two varieties of flaxseed inhibited tumor growth, without any difference between them. Conclusion. Both flaxseed varieties were able to reduce tumor growth. Therefore, their intake should be stimulated with the aim to reduce breast cancer incidence.

Nas últimas décadas tem aumentado o interesse pelos enxertos de pele humana radioesterilizadas, para aplicação principalmente em queimaduras extensas e profundas. Isto se deve ao fato destes enxertos apresentarem rápida aderência e menor potencial antigênico, em comparação com os demais tratamentos utilizados. A proposta deste estudo foi avaliar a histoarquitetura do enxerto de pele humana irradiada com doses de 25 kGy, 50 kGy e não irradiada, durante o processo de reparação tecidual, em camundongos Nude submetidos a enxertia de pele na região dorsal. Três grupos de animais receberam enxertos de pele humana irradiada (25 kGy e 50 kGy) e não irradiada e foram eutanasiados no 3º, 7º e 21º dia após a realização da cirurgia. Após os procedimentos histológicos de rotina, as amostras de tecido foram coradas com hematoxilina e eosina (HE) para a quantificação de queratinócitos, fibroblastos, células de defesa e vasos sanguíneos e a reação de imunofluorescência (IF) foi realizada para a determinação da expressão de colágeno do tipo I humano e do colágeno dos tipos I e III de camundongo. A quantificação, tanto das células quanto dos tipos de colágeno foi realizada por análise de imagem, utilizando o programa Image-Pro PLus 6.0. Os resultados histológicos demostraram que a pele humana irradiação, quando enxertada, influencia o aumento do número de células no local de cicatrização ao longo do tempo, principalmente na dose de 25 kGy, além de proporcionar uma melhor dispersão destas células. No 21º dia, os três grupos de animais com enxertia de pele humana tiveram parte do enxerto incorporado no processo de cicatrização. O grupo não irradiado apresentou maior incorporação do enxerto (43%), porém menor produção de colágeno do tipo III de camundongo (22%). Já os grupos com enxertia de pele irradiada apresentaram menor incorporação do enxerto (6 e 15%), mas com maior produção de colágeno do tipo III de camundongo (35% e 28%, para 25 kGy e 50 kGy, respectivamente). Com este estudo pôde-se concluir que o grupo irradiado a 25 kGy, apresenta maior proliferação celular e formação de vasos,além de melhor remodelamento da região de cicatrização. / Over the last few years it has increased the interest in the human skin grafts radio sterilized for application mainly in extensive and deep burns. Because these grafts quickly grip and present antigenic lower potential, compared with other treatments used. The purpose of this study was to evaluate the histoarchitecture of human skin grafts irradiated with doses 25 kGy, 50 kGy and non-irradiated during the pepair tissue process in nude mice submitted by skin grafting in the dorsal region. Three groups of animals received irradiated human skin grafts (25 kGy and 50 kGy) and non-irradiated and were euthanized on the 3rd, 7th and 21th day after the surgery. Indeed, routine histologic procedures, tissue samples were stained with hematoxylin and eosin (HE) for quantification of keratinocytes, fibroblasts, immune cells and blood vessels and immunofluorescence (IF) was performed to determine the expression human collagen type I and collagen type I and III mouse. Therefore, quantification of both the cells and the collagen types was performed by image analysis using Image-Pro Plus 6.0 software. Histologic results demonstrated at a dose of 25 kGy that human skin irradiation when grafted influences the increase in the number of cells in wound site over time and it provides better dispersion of these cells. In addition, on the 21st day, three groups of animals with human skin graft were embedded part of the graft in the healing process. On the other hand, the group not irradiated showed greater incorporation of the graft (43 %), but less production of collagen type III mouse (22 %). Since the groups irradiated skin graft showed lower graft incorporation (6 and 15%), but with greater production of collagen type III mice (35 % and 28 % to 25 kGy and 50 kGy, respectively). In conclusion, this study presented that the group irradiated to 25 kGy and it has a higher cell proliferation and vessel formation, and better remodeling of the healing area.

Introdução. O câncer é responsável por uma das principais causas de morte entre mulheres e nota-se um aumento na sua incidência. A linhaça tem sido preconizada como capaz de reduzir o risco do câncer de mama. Objetivo. Verificar o efeito de dietas ricas em linhaça marrom e dourada na evolução do câncer de mama. Métodos. Células cancerosas mamárias humanas (tipo MCF-7) foram inseridas no tecido subcutâneo de camundongos fêmeas nude. Quando os tumores já puderam ser claramente identificados, após 7 semanas, os animais constituíram 3 grupos, um com dieta controle e os outros dois com as variedades de linhaça marrom e dourada. As sementes de linhaça foram moídas para produção de farinhas que foram utilizadas na suplementação das dietas oferecidas aos animais. O crescimento dos tumores implantados nos animais foi acompanhado por oito semanas. Resultados. As linhaças das variedades marrom e dourada mostraram-se eficazes na diminuição da taxa de crescimento dos tumores MCF-7, não sendo verificada nenhuma diferença entre elas. Conclusão. Ambas variedades de linhaça reduziram o crescimento dos tumores. Assim, o consumo de linhaça deve ser estimulado a fim de diminuir a incidência do câncer de mama. / Introduction. Breast cancer is one of the main causes of death among women and an increase in its incidence has been noticed. There is evidence that flaxseed intake could reduce breast cancer risk. Objective. To verify the effect of brown and golden flaxseed rich diets on breast cancer. Methods. Cancer cells of the MCF-7 line were injected subcutaneously into athymic mice. After seven weeks, when tumors were already established, the animals were randomly divided in three groups: (1) control, fed on a basal diet; (2) and (3) mice fed on the basal diet supplemented with 10% brown and with 10% golden flaxseed, respectively. Flaxseed was freshly ground for production of the diets offered to the animals. Tumor growth was monitored weekly for eight weeks. Results. The two varieties of flaxseed inhibited tumor growth, without any difference between them. Conclusion. Both flaxseed varieties were able to reduce tumor growth. Therefore, their intake should be stimulated with the aim to reduce breast cancer incidence.