RESUMO
Airway epithelial cells mount a tolerogenic microenvironment that reduces the proinflammatory potential of respiratory dendritic cells (DCs). We recently demonstrated that tracheal epithelial cells continuously secrete soluble mediators that affect the reactivity of local innate immune cells. Using transcriptional profiling, we now observed that conditioning of DCs by tracheal epithelial cells regulated 98 genes under homeostatic conditions. Among the most upregulated genes were Ms4a8a and Ym1, marker genes of alternatively activated myeloid cells. Ex vivo analysis of respiratory DCs from nonchallenged mice confirmed a phenotype of alternative activation. Bioinformatic analysis showed an overrepresentation of hormone-nuclear receptors within the regulated genes, among which was the glucocorticoid receptor. In line with a role for glucocorticoids, pharmacological blockade as well as genetic manipulation of the glucocorticoid receptor within DCs inhibited Ms4a8a and Ym1 expression as well as MHC class II and CD86 regulation upon epithelial cell conditioning. Within epithelial cell-conditioned medium, low amounts of glucocorticoids were present. Further analysis showed that airway epithelial cells did not produce glucocorticoids de novo, yet were able to reactivate inactive dehydrocorticosterone enzymatically. The results show that airway epithelial cells regulate local immune responses, and this modulation involves local production of glucocorticoids and induction of an alternative activation phenotype in DCs.
Assuntos
Microambiente Celular/imunologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Receptores de Glucocorticoides/genética , Mucosa Respiratória/imunologia , Mucosa Respiratória/metabolismo , Transdução de Sinais/imunologia , Animais , Brônquios/citologia , Brônquios/imunologia , Brônquios/metabolismo , Microambiente Celular/genética , Citocinas/metabolismo , Citocinas/fisiologia , Células Dendríticas/citologia , Feminino , Imunofenotipagem , Inflamação/genética , Inflamação/imunologia , Inflamação/prevenção & controle , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Cultura Primária de Células , Distribuição Aleatória , Receptor Cross-Talk/imunologia , Receptores de Glucocorticoides/biossíntese , Receptores de Glucocorticoides/deficiência , Mucosa Respiratória/citologia , Transdução de Sinais/genética , Linfopoietina do Estroma do TimoRESUMO
Intestinal epithelial cells (IECs) act as a physical barrier separating the commensal-containing intestinal tract from the sterile interior. These cells have found a complex balance allowing them to be prepared for pathogen attacks while still tolerating the presence of bacterial or viral stimuli present in the lumen of the gut. Using primary human IECs, we probed the mechanisms that allow for such a tolerance. We discovered that viral infections emanating from the basolateral side of IECs elicit a stronger intrinsic immune response in comparison to lumenal apical infections. We determined that this asymmetric immune response is driven by the clathrin-sorting adaptor AP-1B, which mediates the polarized sorting of Toll-like receptor 3 (TLR3) towards the basolateral side of IECs. Mice and human IECs lacking AP-1B showed an exacerbated immune response following apical stimulation. Together, these results suggest a model where the cellular polarity program plays an integral role in the ability of IECs to partially tolerate apical commensals while remaining fully responsive to invasive basolateral pathogens.
Assuntos
Polaridade Celular/imunologia , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Receptor 3 Toll-Like/metabolismo , Complexo 1 de Proteínas Adaptadoras/genética , Complexo 1 de Proteínas Adaptadoras/metabolismo , Animais , Células Cultivadas , Técnicas de Silenciamento de Genes , Humanos , Interferons/metabolismo , Interleucina-6/metabolismo , Mucosa Intestinal/metabolismo , Mucosa Intestinal/virologia , Camundongos , Receptor 3 Toll-Like/agonistas , Vírus/imunologiaRESUMO
Intestinal epithelial cells (IECs) are exposed to the low-oxygen environment present in the lumen of the gut. These hypoxic conditions on one hand are fundamental for the survival of the commensal microbiota and, on the other hand, favor the formation of a selective semipermeable barrier, allowing IECs to transport essential nutrients/water while keeping the sterile internal compartments separated from the lumen containing commensals. The hypoxia-inducible factor (HIF) complex, which allows cells to respond and adapt to fluctuations in oxygen levels, has been described as a key regulator in maintaining IEC barrier function by regulating their tight junction integrity. In this study, we sought to better evaluate the mechanisms by which low oxygen conditions impact the barrier function of human IECs. By profiling miRNA expression in IECs under hypoxia, we identified microRNA 320a (miRNA-320a) as a novel barrier formation regulator. Using pharmacological inhibitors and short hairpin RNA-mediated silencing, we could demonstrate that expression of this microRNA (miRNA) was HIF dependent. Importantly, using overexpression and knockdown approaches of miRNA-320a, we could confirm its direct role in the regulation of barrier function in human IECs. These results reveal an important link between miRNA expression and barrier integrity, providing a novel insight into mechanisms of hypoxia-driven epithelial homeostasis.