RESUMO
Endotoxin tolerance assures proper regulation of the TLR4 signaling pathway and avoids uncontrolled inflammation, limiting tissue damage and endotoxin shock development. Though underlying molecular mechanisms are still undefined, evidence indicates the involvement of microRNAs, which represent a new layer of regulation of inflammatory pathways. Here, we report that LPS and other inflammatory stimuli repress miR-511-5p expression in human monocytes, while anti-inflammatory stimuli, such as TGF-ß and glucocorticoids, have the opposite effect. MiR-511-5p levels selectively influenced cell activation when endotoxin was used, while biological activity of other TLR agonists was unaffected. Consistent with this, TLR4 was validated as the miR-511-5p direct target responsible for glucocorticoids- and TGF-ß-mediated inhibition of pro-inflammatory cytokines production observed in endotoxin tolerant monocytes. MiR-511-5p thus acts as an intracellular mediator of glucocorticoids and TGF-ß for the induction of endotoxin tolerance in human monocytes.
Assuntos
Regulação para Baixo/efeitos dos fármacos , Glucocorticoides/farmacologia , MicroRNAs/genética , Transdução de Sinais/efeitos dos fármacos , Receptor 4 Toll-Like/genética , Linhagem Celular Tumoral , Células Cultivadas , Dexametasona/farmacologia , Citometria de Fluxo , Regulação da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Subunidade p40 da Interleucina-12/genética , Subunidade p40 da Interleucina-12/metabolismo , Lipopolissacarídeos/farmacologia , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Receptor 4 Toll-Like/metabolismo , Fator de Crescimento Transformador beta/farmacologiaRESUMO
The presence of a growing tumor establishes a chronic state of inflammation that acts locally and systemically. Bone marrow responds to stress signals by expanding myeloid cells endowed with immunosuppressive functions, further fostering tumor growth and dissemination. How early in transformation the cross-talk with the bone marrow begins and becomes detectable in blood is unknown. Here, gene expression profiling of the bone marrow along disease progression in a spontaneous model of mammary carcinogenesis demonstrates that transcriptional modifications in the hematopoietic compartment occurred as early as preinvasive disease stages. The transcriptional profile showed downregulation of adaptive immunity and induction of programs related to innate immunity and response to danger signals triggered by activating transcription factor 3. Transcriptional reprogramming was paralleled by the expansion of myeloid populations at the expense of erythroid and B lymphoid fractions. Hematopoietic changes were associated with modifications of the bone marrow stromal architecture through relocalization and increased density in the interstitial area of Nestin+ mesenchymal cells expressing CXCL12 and myeloid cells expressing CXCL12 receptor CXCR4. These early events were concomitant with deregulation of circulating miRNAs, which were predicted regulators of transcripts downregulated in the bone marrow and involved in lymphoid differentiation and activation. These data provide a link between sensing of peripheral cancer initiation by the bone marrow and hematopoietic adaptation to distant noxia through transcriptional rewiring toward innate/inflammatory response programs. SIGNIFICANCE: The bone marrow senses distant tissue transformation at premalignant/preinvasive stages, suggesting that circulating messengers, intercepted in the blood, could serve as early diagnostic markers.
Assuntos
Adaptação Fisiológica/genética , Biomarcadores Tumorais/genética , Medula Óssea/patologia , Neoplasias da Mama/patologia , MicroRNA Circulante/genética , Células Estromais/patologia , Transcriptoma , Animais , Apoptose , Biomarcadores Tumorais/sangue , Medula Óssea/metabolismo , Neoplasias da Mama/sangue , Neoplasias da Mama/genética , Proliferação de Células , MicroRNA Circulante/sangue , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Células Estromais/metabolismo , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
An appropriate immune response requires a tight balance between pro- and anti-inflammatory mechanisms. IL-10 is induced at late time-points during acute inflammatory conditions triggered by TLR-dependent recognition of infectious agents and is involved in setting this balance, operating as a negative regulator of the TLR-dependent signaling pathway. We identified miR-125a~99b~let-7e as an evolutionary conserved microRNA cluster late-induced in human monocytes exposed to the TLR4 agonist LPS as an effect of this IL-10-dependent regulatory loop. We demonstrated that microRNAs generated by this cluster perform a pervasive regulation of the TLR signaling pathway by direct targeting receptors (TLR4, CD14), signaling molecules (IRAK1), and effector cytokines (TNFα, IL-6, CCL3, CCL7, CXCL8). Modulation of miR-125a~99b~let-7e cluster influenced the production of proinflammatory cytokines in response to LPS and the IL-10-mediated tolerance to LPS, thus identifying this gene as a previously unrecognized major regulatory element of the inflammatory response and endotoxin tolerance.
Assuntos
Regulação da Expressão Gênica , MicroRNAs/genética , Família Multigênica , Transdução de Sinais , Receptor 4 Toll-Like/metabolismo , Linhagem Celular , Biologia Computacional/métodos , Citocinas/metabolismo , Perfilação da Expressão Gênica , Genes Reporter , Humanos , Tolerância Imunológica , Imunofenotipagem , Mediadores da Inflamação/metabolismo , Lipopolissacarídeos/imunologia , Monócitos/imunologia , Monócitos/metabolismo , Interferência de RNA , Receptor 4 Toll-Like/genéticaRESUMO
Beyond the physiology of reproduction, estrogen controls the homeostasis of several tissues. Although macrophages play a key role in tissue remodeling, the interplay with estrogen is still ill defined. Using a transcriptomic approach we first obtained a comprehensive list of genes that are differentially expressed in peritoneal macrophages in response to physiological levels of 17ß-estradiol (E2) injected in intact female mice. Our data also showed the dynamic nature of the macrophage response to E2 and pointed to specific biological programs induced by the hormone, with cell proliferation, immune response and wound healing being the most prominent functional categories. Indeed, the exogenous administration of E2 and, more importantly, the endogenous hormonal surge proved to support macrophage proliferation in vivo, as shown by cell cycle gene expression, BrdU incorporation and cell number. Furthermore, E2 promoted an anti-inflammatory and pro-resolving macrophage phenotype, which converged on the induction of genes related to macrophage alternative activation and on IL-10 expression in vivo. Hormone action was maintained in an experimental model of peritoneal inflammation based on zymosan injection. These findings highlight a direct effect of estrogen on macrophage expansion and phenotypic adaptation in homeostatic conditions and suggest a role for this interplay in inflammatory pathologies.