The Role Of Pattern-recognition Receptors In The Pathogenesis Of Behcet’s Disease And Vogt-koyanagi-harada Syndrome

BackgroundUveitis is one of leading causes of blindness. Behcet’s disease andVogt-Koyanagi-Harada disease are the two most common uveitis entities inChina. The pathogenesis of these two diseases is still unknow. Butinfections and immune system are involved in the pathogenesis of thesetwo diseases. Pattern-recognition receptors(PRRs) is the important part ofinnate immune system and play an important role in the defence ofmicroorganism. The dysfunction of PRRs could induce the disorder ofimmune system and lead to the happening of diseases. The present studywas designsed to investigate the questions as follows:1. The espression ofPRRs in the MDMs from Behcet’s disease and Vogt-Koyanagi-Haradadisease.2. The role of PRRs in the pathogenesis of these two diseases.Collectively this project will provide a new insight into the pathogenesisof uveitis and be hopefully expected to offer a new strategy in the study ofthe prevention and treatment of different uveitis entities. PartⅠ The role of pattern-recognition receptors in thepathogenesis of Behcet’s diseasePurposeBehcet’s disease (BD) is a chronic systemic inflammatory disorder ofunknown etiology. Pattern-recognition receptors(PRRs) are critical in theinnate immune response to microbial invaders. In this study weinvestigated the role of PRRs in the pathogenesis of BD.MethodsTLR2/4expression, IL-1β and reactive oxygen species (ROS) productionwere studied in monocyte-derived macrophages (MDMs) obtained fromBD patients, acute anterior uveitis (AAU) patients and healthy controlsusing real-time PCR, flow cytometry and ELISA. The NLRP3inflammasome of MDMs was downregulated by RNA interference. Thelevels of phosphorylated P38, Erk1/2and JNK MAPK were evaluatedusing flow cytometry.ResultsTLR2/4expression was significantly increased in MDMs from activeBD patients. IL-1β and ROS production of peptidoglycan (PGN)/lipopolysaccharide (LPS)-induced MDMs from active BD patients was significantly increased compared with inactive BD patients, AAU patientsand healthy controls. ROS activator and inhibitor significantly increasedand decreased the production of IL-1β respectively. The production ofIL-1β was significantly decreased after the NLRP3inflammasome wasdownregulated. The phosphorylation levels of p38and ERK1/2in MDMsfrom BD patients and controls were increased following stimulation witheither PGN or LPS. Both SB203580(p38inhibitor) and PD98059(ERK1/2inhibitor) significantly decreased the production of IL-1β.ConclusionsThe results suggest that TLR2/4expression in MDMs from active BDpatients is significantly increased. Interaction of TLR2/4with their ligandsPGN/LPS is involved in BD pathogenesis possibly by the induction ofIL-1β through a ROS-NLRP3dependent pathway.PartⅡ The role of pattern-recognition receptors in thepathogenesis of Vogt-Koyanagi-Harada diseasePurposeActivation of pattern-recognition receptors (PRRs) signaling pathwaysby microbial products can drive host-defense pathways, inflammatoryresponses and adaptive immunity. In the present study, we investigated the role of PRRs in the pathogenesis of Vogt-Koyanagi-Harada disease, atypical autoimmune disease.MethodsMonocyte-derived macrophages (MDMs) from VKH patients, AAUpatients and healthy controls were subjected to analysis of TLR2,3and4expression. IL-1β and reactive oxygen species (ROS) production wasevaluated using real-time PCR, flow cytometry and ELISA. ALentivirus-mediated inhibitor of Nod-like receptor pyrindomain-containing protein3(NLRP3) was transfected to MDMs toexamine its effect on IL-1β production. Flow cytometry was used toevaluate the levels of phosphorylated P38, Erk1/2and JNK by MDMs.ResultsThe expression of TLR3and TLR4, but not TLR2, was significantlyincreased in MDMs from VKH patients with active uveitis compared withinactive VKH, AAU and healthy controls. VKH patients with active uveitisshowed an elevated level of IL-1β and ROS in MDMs with or withoutpolyinosine-polycytidylic acid (poly I:C) and lipopolysaccharide (LPS)stimulation. IL-1β production could be significantly upregulated anddownregulated by a ROS activator or inhibitor respectively.Downregulation of the NLRP3inflammasome significantly inhibited the production of IL-1β. The phosphorylation levels of p38and ERK1/2weresignificantly higher in MDMs stimulated by poly I:C and LPS from activeVKH patients compared to inactive VKH patients, AAU patients andhealthy controls. Inhibition of p38or ERK1/2significantly decreased IL-1βexpression.ConclusionsThe present results showed that the increased expression of TLR3/4inMDMs was involved in the pathogenesis of VKH disease by the inductionof IL-1β through a ROS-NLRP3dependent pathway.