RESUMEN
Cantharidin is an active constituent of mylabris, a traditional Chinese medicine, and is a potent and selective inhibitor of protein phosphatase 2A (PP2A) that plays an important role in cell cycle control, apoptosis, and cell-fate determination. In the present study, we found that cantharidin repressed the invasive ability of pancreatic cancer cells and downregulated matrix metalloproteinase 2 (MMP2) expression through multiple pathways, including ERK, JNK, PKC, NF-κB, and ß-catenin. Interestingly, transcriptional activity of the MMP2 promoter increased after treatment with PP2A inhibitors, suggesting the involvement of a posttranscriptional mechanism. By using an mRNA stability assay, we found accelerated degradation of MMP2 mRNA after treatment of cantharidin. Microarray analyses revealed that multiple genes involved in the 3' â 5' decay pathway were upregulated, especially genes participating in cytoplasmic deadenylation. The elevation of these genes were further demonstrated to be executed through ERK, JNK, PKC, NF-κB, and ß-catenin pathways. Knockdown of PARN, RHAU, and CNOT7, three critical members involved in cytoplasmic deadenylation, attenuated the downregulation of MMP2. Hence, we present the mechanism of repressed invasion by cantharidin and other PP2A inhibitors through increased degradation of MMP2 mRNA by elevated cytoplasmic deadenylation.
Asunto(s)
Antineoplásicos/farmacología , Cantaridina/farmacología , Metaloproteinasa 2 de la Matriz/metabolismo , Neoplasias Pancreáticas/tratamiento farmacológico , Estabilidad del ARN/efectos de los fármacos , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Represión Enzimática/efectos de los fármacos , Humanos , Metaloproteinasa 2 de la Matriz/genética , Invasividad Neoplásica , Proteína Fosfatasa 2/antagonistas & inhibidores , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transducción de SeñalRESUMEN
Pertussis toxin (PTX) has been widely used as an adjuvant to induce Th1-mediated organ-specific autoimmune diseases in animal models. However, the cellular and molecular mechanisms remain to be defined. In this study, we showed that dendritic cells (DC) stimulated with PTX (PTX-DC) were able to substitute for PTX to promote experimental autoimmune uveitis (EAU). EAU induced by PTX-DC revealed a typical Th1 response, characterized by high uveitogenic retinal Ag interphotoreceptor retinoid-binding protein (IRBP)-specific IFN-gamma and IL-12 production in the draining lymph nodes, as well as increased levels of anti-IRBP IgG2a and decreased levels of anti-IRBP IgG1 in the serum of IRBP-immunized mice. Furthermore, PTX-DC preferentially induced T cells to produce the Th1 cytokine, IFN-gamma. After being stimulated with PTX, DC exhibited up-regulation of MHC class II, CD80, CD86, CD40, and DEC205. PTX-DC had also increased allostimulatory capacity and IL-12 and TNF-alpha production. Serum IL-12 was increased in naive mice that received PTX-DC i.p. In addition, PTX activated extracellular signal-regulated kinase in DC. Following the inhibition of extracellular signal-regulated kinase signaling, the maturation of PTX-DC was reduced. Subsequently, the ability of PTX-DC to promote IFN-gamma production by T cells in vitro and to induce EAU in vivo was blocked. The results suggest that PTX might exert an adjuvant effect on DC to promote their maturation and the production of proinflammatory cytokines, thereby eliciting a Th1 response.