Abstract
BackgroundAlthough pneumococcal pneumonia is one of the most common causes of death due to infectious diseases, little is known about pneumococci-lung cell interaction. Herein we tested the hypothesis that pneumococci activated pulmonary epithelial cell cytokine release by c-Jun-NH2-terminal kinase (JNK)MethodsHuman bronchial epithelial cells (BEAS-2B) or epithelial HEK293 cells were infected with S. pneumoniae R6x and cytokine induction was measured by RT-PCR, ELISA and Bioplex assay. JNK-phosphorylation was detected by Western blot and nuclear signaling was assessed by electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP). JNK was modulated by the small molecule inhibitor SP600125 and AP1 by transfection of a dominant negative mutant.ResultsS. pneumoniae induced the release of distinct CC and CXC, as well as Th1 and Th2 cytokines and growth factors by human lung epithelial cell line BEAS-2B. Furthermore, pneumococci infection resulted in JNK phosphorylation in BEAS-2B cells. Inhibition of JNK by small molecule inhibitor SP600125 reduced pneumococci-induced IL-8 mRNA expression and release of IL-8 and IL-6. One regulator of the il8 promoter is JNK-phosphorylated activator protein 1 (AP-1). We showed that S. pneumoniae time-dependently induced DNA binding of AP-1 and its phosphorylated subunit c-Jun with a maximum at 3 to 5 h after infection. Recruitment of Ser63/73-phosphorylated c-Jun and RNA polymerase II to the endogenous il8 promoter was found 2 h after S. pneumoniae infection by chromatin immunoprecipitation. AP-1 repressor A-Fos reduced IL-8 release by TLR2-overexpressing HEK293 cells induced by pneumococci but not by TNFα. Antisense-constructs targeting the AP-1 subunits Fra1 and Fra2 had no inhibitory effect on pneumococci-induced IL-8 release.ConclusionS. pneumoniae-induced IL-8 expression by human epithelial BEAS-2B cells depended on activation of JNK and recruitment of phosphorylated c-Jun to the il8 promoter.
Highlights
Pneumococcal pneumonia is one of the most common causes of death due to infectious diseases, little is known about pneumococci-lung cell interaction
Antisense-constructs targeting the activator protein 1 (AP-1) subunits Fra1 and Fra2 had no inhibitory effect on pneumococci-induced IL-8 release
S. pneumoniae induced cytokine release in human lung epithelial BEAS-2B cells To characterize the inflammatory activation of human lung epithelial cells by S. pneumoniae, we infected BEAS2B cell with pneumococci strain R6x with an infection dose of 106 cfu/ml
Summary
Pneumococcal pneumonia is one of the most common causes of death due to infectious diseases, little is known about pneumococci-lung cell interaction. Pneumonia is the most common cause of death due to infectious diseases in industrialized countries [1]. Despite the availability of vaccines and antibiotic treatments, mortality rates remain high [2,4]. The number of antibiotic resistant strains is increasing and even vancomycin-tolerant strains have been observed [5]. Cytokine liberation and subsequent recruitment and activation of leucocytes are a hallmark in pneumococci pneumonia usually leading to elimination of the pathogens. Immune cells like alveolar macrophages significantly contribute to the activation of the host immune system, evidence has been presented that lung epithelium considerably participates in the recognition of invading pathogens and initiation of the host response [6]. Since the pulmonary epithelium constitutes a large surface, which is in direct contact with invading pathogens, analysis of the interaction between pathogens and pulmonary epithelial cells is of considerable interest
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