RESUMO
Rationale: 17ß-estradiol (E2) can directly promote the growth of ERα-negative cancer cells through activation of endothelial ERα in the tumor microenvironment, thereby increasing a normalized tumor angiogenesis. ERα acts as a transcription factor through its nuclear transcriptional AF-1 and AF-2 transactivation functions, but membrane ERα plays also an important role in endothelium. The present study aims to decipher the respective roles of these two pathways in ERα-negative tumor growth. Moreover, we delineate the actions of tamoxifen, a Selective Estrogen Receptor Modulator (SERM) in ERα-negative tumors growth and angiogenesis, since we recently demonstrated that tamoxifen impacts vasculature functions through complex modulation of ERα activity. Methods: ERα-negative B16K1 cancer cells were grafted into immunocompetent mice mutated for ERα-subfunctions and tumor growths were analyzed in these different models in response to E2 and/or tamoxifen treatment. Furthermore, RNA sequencings were analyzed in endothelial cells in response to these different treatments and validated by RT-qPCR and western blot. Results: We demonstrate that both nuclear and membrane ERα actions are required for the pro-tumoral effects of E2, while tamoxifen totally abrogates the E2-induced in vivo tumor growth, through inhibition of angiogenesis but promotion of vessel normalization. RNA sequencing indicates that tamoxifen inhibits the E2-induced genes, but also initiates a specific transcriptional program that especially regulates angiogenic genes and differentially regulates glycolysis, oxidative phosphorylation and inflammatory responses in endothelial cells. Conclusion: These findings provide evidence that tamoxifen specifically inhibits angiogenesis through a reprogramming of endothelial gene expression via regulation of some transcription factors, that could open new promising strategies to manage cancer therapies affecting the tumor microenvironment of ERα-negative tumors.
Assuntos
Neoplasias , Tamoxifeno , Camundongos , Animais , Tamoxifeno/farmacologia , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Células Endoteliais/metabolismo , Angiogênese , Expressão Gênica , Endotélio/metabolismo , Linhagem Celular Tumoral , Microambiente Tumoral/genéticaRESUMO
OBJECTIVE: ERα (estrogen receptor alpha) exerts nuclear genomic actions and also rapid membrane-initiated steroid signaling. The mutation of the cysteine 451 into alanine in vivo has recently revealed the key role of this ERα palmitoylation site on some vasculoprotective actions of 17ß-estradiol (E2) and fertility. Here, we studied the in vivo role of the arginine 260 of ERα which has also been described to be involved in its E2-induced rapid signaling with PI-3K (phosphoinositide 3-kinase) as well as G protein in cultured cell lines. Approach and Results: We generated a mouse model harboring a point mutation of the murine counterpart of this arginine into alanine (R264A-ERα). In contrast to the C451A-ERα, the R264A-ERα females are fertile with standard hormonal serum levels and normal control of hypothalamus-pituitary ovarian axis. Although R264A-ERα protein abundance was normal, the well-described membrane ERα-dependent actions of estradiol, such as the rapid dilation of mesenteric arteries and the acceleration of endothelial repair of carotid, were abrogated in R264A-ERα mice. In striking contrast, E2-regulated gene expression was highly preserved in the uterus and the aorta, revealing intact nuclear/genomic actions in response to E2. Consistently, 2 recognized nuclear ERα-dependent actions of E2, namely atheroma prevention and flow-mediated arterial remodeling were totally preserved. CONCLUSIONS: These data underline the exquisite role of arginine 264 of ERα for endothelial membrane-initiated steroid signaling effects of E2 but not for nuclear/genomic actions. This provides the first model of fertile mouse with no overt endocrine abnormalities with specific loss-of-function of rapid ERα signaling in vascular functions.
Assuntos
Lesões das Artérias Carótidas/tratamento farmacológico , Endotélio Vascular/efeitos dos fármacos , Estradiol/farmacologia , Receptor alfa de Estrogênio/agonistas , Terapia de Reposição de Estrogênios , Estrogênios/farmacologia , Fertilidade/efeitos dos fármacos , Artérias Mesentéricas/efeitos dos fármacos , Mutação Puntual , Animais , Aterosclerose/metabolismo , Aterosclerose/patologia , Aterosclerose/prevenção & controle , Lesões das Artérias Carótidas/metabolismo , Lesões das Artérias Carótidas/patologia , Lesões das Artérias Carótidas/fisiopatologia , Proliferação de Células/efeitos dos fármacos , Endotélio Vascular/lesões , Endotélio Vascular/metabolismo , Endotélio Vascular/fisiopatologia , Ativação Enzimática , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Ciclo Estral/efeitos dos fármacos , Feminino , Masculino , Artérias Mesentéricas/metabolismo , Artérias Mesentéricas/fisiopatologia , Camundongos Endogâmicos C57BL , Óxido Nítrico Sintase Tipo III/metabolismo , Ovariectomia , Reepitelização/efeitos dos fármacos , Transdução de Sinais , Fatores de Tempo , Útero/efeitos dos fármacos , Útero/metabolismo , Remodelação Vascular/efeitos dos fármacos , Vasodilatação/efeitos dos fármacosRESUMO
17ß-Estradiol induces the postnatal development of mammary gland and influences breast carcinogenesis by binding to the estrogen receptor ERα. ERα acts as a transcription factor but also elicits rapid signaling through a fraction of ERα expressed at the membrane. Here, we have used the C451A-ERα mouse model mutated for the palmitoylation site to understand how ERα membrane signaling affects mammary gland development. Although the overall structure of physiological mammary gland development is slightly affected, both epithelial fragments and basal cells isolated from C451A-ERα mammary glands failed to grow when engrafted into cleared wild-type fat pads, even in pregnant hosts. Similarly, basal cells purified from hormone-stimulated ovariectomized C451A-ERα mice did not produce normal outgrowths. Ex vivo, C451A-ERα basal cells displayed reduced matrix degradation capacities, suggesting altered migration properties. More importantly, C451A-ERα basal cells recovered in vivo repopulating ability when co-transplanted with wild-type luminal cells and specifically with ERα-positive luminal cells. Transcriptional profiling identified crucial paracrine luminal-to-basal signals. Altogether, our findings uncover an important role for membrane ERα expression in promoting intercellular communications that are essential for mammary gland development.
Assuntos
Epitélio/metabolismo , Receptor alfa de Estrogênio/biossíntese , Glândulas Mamárias Animais/embriologia , Comunicação Parácrina/fisiologia , Animais , Células Epiteliais/metabolismo , Células Epiteliais/transplante , Estradiol/metabolismo , Receptor alfa de Estrogênio/genética , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Lipoilação/fisiologia , Glândulas Mamárias Animais/metabolismo , Glândulas Mamárias Animais/transplante , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Transdução de SinaisRESUMO
OBJECTIVE: Estrogens exert beneficial effect on the blood vascular system. However, their role on the lymphatic system has been poorly investigated. We studied the protective effect of the 17ß estradiol-the most potent endogenous estrogen-in lymphedema-a lymphatic dysfunction, which results in a massive fluid and fat accumulation in the limb. APPROACH AND RESULTS: Screening of DNA motifs able to mobilize ERs (estrogen receptors) and quantitative real-time polymerase chain reaction analysis revealed that estradiol promotes transcriptional activation of lymphangiogenesis-related gene expression including VEGF (vascular endothelial growth factor)-D, VEGFR (VEGF receptor)-3, lyve-1, and HASs (hyaluronan synthases). Using an original model of secondary lymphedema, we observed a protective effect of estradiol on lymphedema by reducing dermal backflow-a representative feature of the pathology. Blocking ERα by tamoxifen-the selective estrogen modulator-led to a remodeling of the lymphatic network associated with a strong lymphatic leakage. Moreover, the protection of lymphedema by estradiol treatment was abrogated by the endothelial deletion of the receptor ERα in Tie2-Cre; ERαlox/lox mice, which exhibit dilated lymphatic vessels. This remodeling correlated with a decrease in lymphangiogenic gene expression. In vitro, blocking ERα by tamoxifen in lymphatic endothelial cells decreased cell-cell junctions, inhibited migration and sprouting, and resulted in an inhibition of Erk but not of Akt phosphorylation. CONCLUSIONS: Estradiol protection from developing lymphedema is mediated by an activation of its receptor ERα and is antagonized by tamoxifen. These findings reveal a new facet of the estrogen influence in the management of the lymphatic system and provide more evidence that secondary lymphedema is worsened by hormone therapy.
Assuntos
Linfedema Relacionado a Câncer de Mama/prevenção & controle , Estradiol/administração & dosagem , Receptor alfa de Estrogênio/agonistas , Terapia de Reposição Hormonal , Linfangiogênese/efeitos dos fármacos , Vasos Linfáticos/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Linfedema Relacionado a Câncer de Mama/metabolismo , Linfedema Relacionado a Câncer de Mama/patologia , Linfedema Relacionado a Câncer de Mama/fisiopatologia , Modelos Animais de Doenças , Implantes de Medicamento , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Vasos Linfáticos/metabolismo , Vasos Linfáticos/patologia , Vasos Linfáticos/fisiopatologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ovariectomia , Fosforilação , Moduladores Seletivos de Receptor Estrogênico/toxicidade , Tamoxifeno/toxicidadeRESUMO
Women now live more than a third of their lives after the onset of menopause. The decline in endogenous estrogen production during this period is accompanied by functional disorders that affect quality of life. These symptoms may be relieved by menopausal hormone therapy (MHT) initially based on the administration of equine conjugated estrogens (mainly in the United States, oral route) or the natural estrogen, 17ß-estradiol (in Europe, transdermal route). Estrogen receptor α (ERα), but not ERß, mediates most of the physiological effects of estrogens. ERα belongs to the superfamily of nuclear receptors and regulates the transcription of genes via its activation functions AF1 and AF2. In addition to these classical genomic actions, estrogens can activate a subpopulation of ERα present at the cell membrane and thereby induce rapid signals. In this review, we will summarize the evolution of MHTs in last decades, as well as treatments that use various selective estrogen receptor modulators (SERMs). Next, we will describe recent advances in the understanding of the mechanisms of estrogen action, in particular the respective roles of nuclear and membrane ERα as well as the potential implications for future therapies.
Assuntos
Terapia de Reposição de Estrogênios/tendências , Menopausa/efeitos dos fármacos , Moduladores Seletivos de Receptor Estrogênico/uso terapêutico , Animais , Calibragem , Receptor alfa de Estrogênio/metabolismo , Terapia de Reposição de Estrogênios/métodos , Terapia de Reposição de Estrogênios/normas , Feminino , Humanos , Menopausa/fisiologia , Qualidade de Vida , Moduladores Seletivos de Receptor Estrogênico/administração & dosagemRESUMO
OBJECTIVE: Chronic nonhealing wounds are a substantial medical concern and are associated with morbidity and mortality; thus, new treatment strategies are required. The first step toward personalized/precision medicine in this field is probably in taking sex differences into account. Impaired wound healing is augmented by ischemia, and we previously demonstrated that 17ß-estradiol exerts a major preventive effect against ischemia-induced skin flap necrosis in female mice. However, the equivalent effects of testosterone in male mice have not yet been reported. We then investigated the role of steroid hormones in male mice using a skin flap ischemia model. APPROACH AND RESULTS: Castrated male mice developed skin necrosis after ischemia, whereas intact or castrated males treated with testosterone were equally protected. Testosterone can (1) activate the estrogen receptor after its aromatization into 17ß-estradiol or (2) be reduced into dihydrotestosterone, a nonaromatizable androgen that activates the androgen receptor. We found that dihydrotestosterone protected castrated wild-type mice by promoting skin revascularization, probably through a direct action on resistance arteries, as evidenced using a complementary model of flow-mediated outward remodeling. 17ß-estradiol treatment of castrated male mice also strongly protected them from ischemic necrosis through the activation of estrogen receptor-α by increasing skin revascularization and skin survival. Remarkably, 17ß-estradiol improved skin survival with a greater efficiency than dihydrotestosterone. CONCLUSIONS: Testosterone provides males with a strong protection against cutaneous necrosis and acts through both its estrogenic and androgenic derivatives, which have complementary effects on skin survival and revascularization.
Assuntos
Di-Hidrotestosterona/farmacologia , Estradiol/farmacologia , Terapia de Reposição Hormonal , Isquemia/prevenção & controle , Neovascularização Fisiológica/efeitos dos fármacos , Pele/irrigação sanguínea , Pele/efeitos dos fármacos , Retalhos Cirúrgicos/irrigação sanguínea , Cicatrização/efeitos dos fármacos , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Receptor alfa de Estrogênio/agonistas , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Isquemia/metabolismo , Isquemia/patologia , Isquemia/fisiopatologia , Masculino , Artérias Mesentéricas/efeitos dos fármacos , Camundongos Pelados , Camundongos Endogâmicos C57BL , Camundongos Knockout , Necrose , Orquiectomia , Ratos Wistar , Pele/metabolismo , Pele/patologia , Retalhos Cirúrgicos/patologia , Fatores de Tempo , Sobrevivência de Tecidos/efeitos dos fármacosRESUMO
BACKGROUND: To date, all studies conducted on breast cancer diagnosis have focused on the expression of the full-length 66-kDa estrogen receptor alpha (ERα66). However, much less attention has been paid to a shorter 46-kDa isoform (ERα46), devoid of the N-terminal region containing the transactivation function AF-1. Here, we investigated the expression levels of ERα46 in breast tumors in relation to tumor grade and size, and examined the mechanism of its generation and its specificities of coregulatory binding and its functional activities. METHODS: Using approaches combining immunohistochemistry, Western blotting, and proteomics, antibodies allowing ERα46 detection were identified and the expression levels of ERα46 were quantified in 116 ERα-positive human breast tumors. ERα46 expression upon cellular stress was studied, and coregulator bindings, transcriptional, and proliferative response were determined to both ERα isoforms. RESULTS: ERα46 was expressed in over 70% of breast tumors at variable levels which sometimes were more abundant than ERα66, especially in differentiated, lower-grade, and smaller-sized tumors. We also found that ERα46 can be generated via internal ribosome entry site-mediated translation in the context of endoplasmic reticulum stress. The binding affinities of both unliganded and fully-activated receptors towards co-regulator peptides revealed that the respective potencies of ERα46 and ERα66 differ significantly, contributing to the differential transcriptional activity of target genes to 17ß estradiol (E2). Finally, increasing amounts of ERα46 decrease the proliferation rate of MCF7 tumor cells in response to E2. CONCLUSIONS: We found that, besides the full-length ERα66, the overlooked ERα46 isoform is also expressed in a majority of breast tumors. This finding highlights the importance of the choice of antibodies used for the diagnosis of breast cancer, which are able or not to detect the ERα46 isoform. In addition, since the function of both ERα isoforms differs, this work underlines the need to develop new technologies in order to discriminate ERα66 and ERα46 expression in breast cancer diagnosis which could have potential clinical relevance.
Assuntos
Neoplasias da Mama/genética , Receptor alfa de Estrogênio/genética , Regulação Neoplásica da Expressão Gênica , Processamento Alternativo , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Análise por Conglomerados , Estresse do Retículo Endoplasmático , Estradiol/farmacologia , Receptor alfa de Estrogênio/metabolismo , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Imuno-Histoquímica , Ligação Proteica , Biossíntese de Proteínas , Isoformas de Proteínas , Proteoma , Proteômica/métodos , Estudos RetrospectivosRESUMO
Estrogen receptors (ER) are used as therapeutic targets, either for contraception or for the hormonal replacement therapy in menopausal women, but also in physiopathology for breast cancer treatment. It is therefore important to understand the tissue-specificity of the actions of ERα to optimize the benefits/risks ratio in each tissue. Besides the conventional nuclear ERα acting as a transcription factor, many studies have demonstrated that ERα is also able to mediate extra nuclear signaling, enabling rapid actions of estrogen. Recently, new transgenic mouse models were used to study these effects, and allowed to genetically segregate membrane versus nuclear actions of a steroid hormone receptor, demonstrating their in vivo tissue-specific roles.
Assuntos
Receptor alfa de Estrogênio/fisiologia , Animais , Humanos , Especificidade de Órgãos/fisiologiaRESUMO
Estrogen receptor alpha (ERα) activation functions AF-1 and AF-2 classically mediate gene transcription in response to estradiol (E2). A fraction of ERα is targeted to plasma membrane and elicits membrane-initiated steroid signaling (MISS), but the physiological roles of MISS in vivo are poorly understood. We therefore generated a mouse with a point mutation of the palmitoylation site of ERα (C451A-ERα) to obtain membrane-specific loss of function of ERα. The abrogation of membrane localization of ERα in vivo was confirmed in primary hepatocytes, and it resulted in female infertility with abnormal ovaries lacking corpora lutea and increase in luteinizing hormone levels. In contrast, E2 action in the uterus was preserved in C451A-ERα mice and endometrial epithelial proliferation was similar to wild type. However, E2 vascular actions such as rapid dilatation, acceleration of endothelial repair, and endothelial NO synthase phosphorylation were abrogated in C451A-ERα mice. A complementary mutant mouse lacking the transactivation function AF-2 of ERα (ERα-AF2(0)) provided selective loss of function of nuclear ERα actions. In ERα-AF2(0), the acceleration of endothelial repair in response to estrogen-dendrimer conjugate, which is a membrane-selective ER ligand, was unaltered, demonstrating integrity of MISS actions. In genome-wide analysis of uterine gene expression, the vast majority of E2-dependent gene regulation was abrogated in ERα-AF2(0), whereas in C451A-ERα it was nearly fully preserved, indicating that membrane-to-nuclear receptor cross-talk in vivo is modest in the uterus. Thus, this work genetically segregated membrane versus nuclear actions of a steroid hormone receptor and demonstrated their in vivo tissue-specific roles.
Assuntos
Membrana Celular/metabolismo , Núcleo Celular/metabolismo , Receptor alfa de Estrogênio/genética , Ovário/fisiologia , Útero/metabolismo , Análise de Variância , Animais , Western Blotting , Movimento Celular , Biologia Computacional , Células Endoteliais , Receptor alfa de Estrogênio/metabolismo , Feminino , Processamento de Imagem Assistida por Computador , Imuno-Histoquímica , Lipoilação/genética , Camundongos , Camundongos Transgênicos , Análise em Microsséries , Ovário/metabolismo , Mutação Puntual/genética , Receptor Cross-Talk/fisiologiaRESUMO
Estrogens directly promote the growth of breast cancers that express the estrogen receptor α (ERα). However, the contribution of stromal expression of ERα in the tumor microenvironment to the protumoral effects of estrogen has never been explored. In this study, we evaluated the molecular and cellular mechanisms by which 17ß-estradiol (E2) impacts the microenvironment and modulates tumor development of ERα-negative tumors. Using different mouse models of ER-negative cancer cells grafted subcutaneously into syngeneic ovariectomized immunocompetent mice, we found that E2 potentiates tumor growth, increases intratumoral vessel density, and modifies tumor vasculature into a more regularly organized structure, thereby improving vessel stabilization to prevent tumor hypoxia and necrosis. These E2-induced effects were completely abrogated in ERα-deficient mice, showing a critical role of host ERα. Notably, E2 did not accelerate tumor growth when ERα was deficient in Tie2-positive cells, even in mice grafted with wild-type bone marrow. These results were extended by clinical evidence of ERα-positive stromal cell labeling in the microenvironment of human breast cancers. Together, our findings therefore show that E2 promotes the growth of ERα-negative cancer cells through the activation of stromal ERα (extra-hematopoietic Tie-2 positive cells), which normalizes tumor angiogenesis and allows an adaptation of blood supply to tumors, thereby preventing hypoxia and necrosis. These findings significantly deepen mechanistic insights into the impact of E2 on tumor development with potential consequences for cancer treatment.
Assuntos
Estradiol/metabolismo , Receptor alfa de Estrogênio/deficiência , Melanoma/irrigação sanguínea , Melanoma/patologia , Neovascularização Patológica/patologia , Animais , Linhagem Celular Tumoral , Proliferação de Células , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Melanoma/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células Estromais , Microambiente TumoralRESUMO
The C-type lectin dendritic cell-specific intercellular adhesion molecule-3 grabbing nonintegrin (DC-SIGN) mediates the innate immune recognition of microbial carbohydrates. We investigated the function of this molecule in the host response to pathogens in vivo, by generating mouse lines lacking the DC-SIGN homologues SIGNR1, SIGNR3, and SIGNR5. Resistance to Mycobacterium tuberculosis was impaired only in SIGNR3-deficient animals. SIGNR3 was expressed in lung phagocytes during infection, and interacted with M. tuberculosis bacilli and mycobacterial surface glycoconjugates to induce secretion of critical host defense inflammatory cytokines, including tumor necrosis factor (TNF). SIGNR3 signaling was dependent on an intracellular tyrosine-based motif and the tyrosine kinase Syk. Thus, the mouse DC-SIGN homologue SIGNR3 makes a unique contribution to protection of the host against a pulmonary bacterial pathogen.
Assuntos
Moléculas de Adesão Celular/fisiologia , Lectinas Tipo C/fisiologia , Receptores de Superfície Celular/fisiologia , Tuberculose/imunologia , Animais , Antígenos CD/fisiologia , MAP Quinases Reguladas por Sinal Extracelular/fisiologia , Feminino , Glicoconjugados/metabolismo , Interleucina-6/biossíntese , Lipopolissacarídeos/metabolismo , Pulmão/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , NF-kappa B/fisiologia , Proteínas Proto-Oncogênicas c-raf/fisiologia , Transdução de Sinais , Receptor 2 Toll-Like/fisiologia , Fator de Necrose Tumoral alfa/biossínteseRESUMO
We recently showed that treatment of macrophages prior to Mycobacterium tuberculosis infection with the pro-inflammatory omega-6 lipid, arachidonic acid (AA) enhanced bacterial killing whereas the anti-inflammatory, omega-3 lipid eicosapentaenoic acid (EPA) stimulated bacterial growth. Here we tested if these effects were depending on when lipids were added to macrophages: before or during Mycobacterium smegmatis or M. tuberculosis infection. Collectively, our data suggested that a high omega-6 diet might be beneficial against mycobacteriosis, while a high omega-3 diet might be detrimental. AA also stimulated TNF-alpha secretion in M. tuberculosis-infected macrophages whereas EPA inhibited this process. AA strongly activated the MAP kinase p38 in uninfected cells but M. tuberculosis infected cells blocked the ability of AA to activate p38; AA-dependent killing is therefore independent of p38. We therefore tested diets enriched in omega-3 and omega-6 lipids on a mouse model of tuberculosis. In contrast to the in vitro results, the omega-6 tended to increase survival of M. tuberculosis in mice, while omega-3- tended to increase pathogen killing. Overall our results together with those previously reported in the literature suggest that it is almost impossible to predict, at the whole organism level, if a diet enriched in omega-3 or -6 will be beneficial or detrimental to intracellular pathogens.
Assuntos
Ácidos Graxos Ômega-3/farmacologia , Ácidos Graxos Ômega-6/farmacologia , Interações Hospedeiro-Patógeno , Macrófagos , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/fisiologia , Animais , Ácido Araquidônico/administração & dosagem , Ácido Araquidônico/farmacologia , Linhagem Celular , Células Cultivadas , Ácido Eicosapentaenoico/administração & dosagem , Ácido Eicosapentaenoico/farmacologia , Ácidos Graxos Ômega-3/administração & dosagem , Ácidos Graxos Ômega-6/administração & dosagem , Feminino , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/microbiologia , Camundongos , Camundongos Endogâmicos BALB C , Mycobacterium smegmatis/efeitos dos fármacos , Mycobacterium smegmatis/crescimento & desenvolvimento , Mycobacterium tuberculosis/crescimento & desenvolvimento , Tuberculose/imunologia , Tuberculose/microbiologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Using signature-tagged transposon mutagenesis, we isolated 23 Mycobacterium tuberculosis mutants, corresponding to 21 genes or genetic regions, attenuated in their ability to parasitize human macrophages. Mutants disrupted in the ABC transporter-encoding genes Rv0986 and Rv0987 were further characterized as being impaired in their ability to bind to host cells.
Assuntos
Elementos de DNA Transponíveis/genética , Genes Bacterianos , Macrófagos/parasitologia , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/patogenicidade , Animais , Proteínas de Bactérias/metabolismo , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , MutagêneseRESUMO
p-Hydroxybenzoic acid derivatives (p-HBADs) are glycoconjugates secreted by all Mycobacterium tuberculosis isolates whose contribution to pathogenicity remains to be determined. The pathogenicity of three transposon mutants of M. tuberculosis deficient in the biosynthesis of some or all forms of p-HBADs was studied. Whilst the mutants grew similarly to the wild-type strain in macrophages and C57BL/6 mice, two of the mutants induced a more severe and diffuse inflammation in the lungs. The lack of production of some or all forms of p-HBADs in these two mutants also correlated with an increased secretion of the pro-inflammatory cytokines tumour-necrosis factor alpha, interleukin 6 and interleukin 12 in vivo. We propose that the loss of production of p-HBADs by tubercle bacilli results in their diminished ability to suppress the pro-inflammatory response to infection and that this ultimately provokes extensive pulmonary lesions in the C57BL/6 model of tuberculosis infection.