Effects of Porphyromonas gingivalis LipopolysaccharideTolerized Monocytes on Inflammatory Responses in Neutrophils

Periodontitis is a chronic inflammatory disease induced by bacteria. Exposure of the host to periodontal pathogens and their virulence factors induces a state of hyporesponsiveness to subsequent stimulations, which is termed endotoxin tolerance. The role and mechanism of lipopolysaccharide (LPS)–tolerized monocytes in inflammatory responses in neutrophils are currently unclear. Here, conditioned supernatants were collected from THP-1 cells treated with or without repeated 1 μg/ml Porphyromonas gingivalis (P.gingivalis) LPS. The chemotactic response of freshly isolated neutrophils recruited by supernatants was determined by a transwell migration assay, which demonstrated a reduced migration of neutrophils stimulated with supernatants from tolerized THP-1 cells in comparison to non-tolerized THP-1 cells. In addition, there was a marked increase in reactive oxygen species (ROS) generation and a significant decrease in Caspase 3 activities in neutrophils treated with supernatants from THP-1 cells that were treated repeatedly with P.gingivalis LPS in comparison to single treatment. A cytokine antibody array was then used to assess cytokine expression patterns in THP-1 cells. In tolerized THP-1 cells, 43 cytokine (43/170) expression levels were decreased, including chemokine ligand 23 (CCL23) and IFN-γ, while 11 cytokine (11/170) expression levels were increased, such as death receptor 6 (DR6). Furthermore, there was decreased production of IFN-γ and epithelial neutrophil activating peptide-78 (ENA-78) in THP-1 cells after stimulation with repeated P. gingivalis LPS in comparison to single challenge, which was confirmed by ELISA. Therefore, P.gingivalis LPS- tolerized THP-1 cells were able to depress neutrophil chemotaxis and apoptosis, and contribute to respiratory burst, which might be related to the changes in cytokine expression patterns in THP-1 cells.     

Porphyromonas gingivalis Gingipain-Dependently Enhances IL-33 Production in Human Gingival Epithelial Cells

The cytokine IL-33 is constitutively expressed in epithelial cells and it augments Th2 cytokine-mediated inflammatory responses by regulating innate immune cells. We aimed to determine the role of the periodontal pathogen, Porphyromonas gingivalis, in the enhanced expression of IL-33 in human gingival epithelial cells. We detected IL-33 in inflamed gingival epithelium from patients with chronic periodontitis, and found that P. gingivalis increased IL-33 expression in the cytoplasm of human gingival epithelial cells in vitro. In contrast, lipopolysaccharide, lipopeptide, and fimbriae derived from P. gingivalis did not increase IL-33 expression. Specific inhibitors of P. gingivalis proteases (gingipains) suppressed IL-33 mRNA induction by P. gingivalis and the P. gingivalis gingipain-null mutant KDP136 did not induce IL-33 expression. A small interfering RNA for protease-activated receptor-2 (PAR-2) as well as inhibitors of phospholipase C, p38 and NF-κB inhibited the expression of IL-33 induced by P. gingivalis. These results indicate that the PAR-2/IL-33 axis is promoted by P. gingivalis infection in human gingival epithelial cells through a gingipain-dependent mechanism.     

Altered T-Cell Responses by the Periodontal Pathogen Porphyromonas gingivalis

Several studies support an association between the chronic inflammatory diseases periodontitis and atherosclerosis with a crucial role for the periodontal pathogen Porphyromonas gingivalis. However, the interplay between this pathogen and the adaptive immune system, including T-cells, is sparsely investigated. Here we used Jurkat T-cells to determine the effects of P. gingivalis on T-cell-mediated adaptive immune responses. We show that viable P. gingivalis targets IL-2 expression at the protein level. Initial cellular events, including ROS production and [Ca²⁺]_i, were elevated in response to P. gingivalis, but AP-1 and NF-κB activity dropped below basal levels and T-cells were unable to sustain stable IL-2 accumulation. IL-2 was partially restored by Leupeptin, but not by Cathepsin B Inhibitor, indicating an involvement of Rgp proteinases in the suppression of IL-2 accumulation. This was further confirmed by purified Rgp that caused a dose-dependent decrease in IL-2 levels. These results provide new insights of how this periodontal pathogen evades the host adaptive immune system by inhibiting IL-2 accumulation and thus attenuating T-cell proliferation and cellular communication.     

The Daiokanzoto (TJ-84) Kampo Formulation Reduces Virulence Factor Gene Expression in Porphyromonas gingivalis and Possesses Anti-Inflammatory and Anti-Protease Activities

Kampo formulations used in Japan to treat a wide variety of diseases and to promote health are composed of mixtures of crude extracts from the roots, bark, leaves, and rhizomes of a number of herbs. The present study was aimed at identifying the beneficial biological properties of Daiokanzoto (TJ-84), a Kampo formulation composed of crude extracts of Rhubarb rhizomes and Glycyrrhiza roots, with a view to using it as a potential treatment for periodontal disease. Daiokanzoto dose-dependently inhibited the expression of major Porphyromonas gingivalis virulence factors involved in host colonization and tissue destruction. More specifically, Daiokanzoto reduced the expression of the fimA, hagA, rgpA, and rgpB genes, as determined by quantitative real-time PCR. The U937-3xκB-LUC monocyte cell line transfected with a luciferase reporter gene was used to evaluate the anti-inflammatory properties of Daiokanzoto. Daiokanzoto attenuated the P. gingivalis-mediated activation of the NF-κB signaling pathway. It also reduced the secretion of pro-inflammatory cytokines (IL-6 and CXCL8) by lipopolysaccharide-stimulated oral epithelial cells and gingival fibroblasts. Lastly, Daiokanzoto, dose-dependently inhibited the catalytic activity of matrix metalloproteinases (-1 and -9). In conclusion, the present study provided evidence that Daiokanzoto shows potential for treating and/or preventing periodontal disease. The ability of this Kampo formulation to act on both bacterial pathogens and the host inflammatory response, the two etiological components of periodontal disease, is of high therapeutic interest.     

Porphyromonas gingivalis Mediates Inflammasome Repression in Polymicrobial Cultures through a Novel Mechanism Involving Reduced Endocytosis

The interleukin (IL)-1β-processing inflammasome has recently been identified as a target for pathogenic evasion of the inflammatory response by a number of bacteria and viruses. We postulated that the periodontal pathogen, Porphyromonas gingivalis may suppress the inflammasome as a mechanism for its low immunogenicity and pathogenic synergy with other, more highly immunogenic periodontal bacteria. Our results show that P. gingivalis lacks signaling capability for the activation of the inflammasome in mouse macrophages. Furthermore, P. gingivalis can suppress inflammasome activation by another periodontal bacterium, Fusobacterium nucleatum. This repression affects IL-1β processing, as well as other inflammasome-mediated processes, including IL-18 processing and cell death, in both human and mouse macrophages. F. nucleatum activates IL-1β processing through the Nlrp3 inflammasome; however, P. gingivalis repression is not mediated through reduced levels of inflammasome components. P. gingivalis can repress Nlrp3 inflammasome activation by Escherichia coli, and by danger-associated molecular patterns and pattern-associated molecular patterns that mediate activation through endocytosis. However, P. gingivalis does not suppress Nlrp3 inflammasome activation by ATP or nigericin. This suggests that P. gingivalis may preferentially suppress endocytic pathways toward inflammasome activation. To directly test whether P. gingivalis infection affects endocytosis, we assessed the uptake of fluorescent particles in the presence or absence of P. gingivalis. Our results show that P. gingivalis limits both the number of cells taking up beads and the number of beads taken up for bead-positive cells. These results provide a novel mechanism of pathogen-mediated inflammasome inhibition through the suppression of endocytosis.     

Macrophage Subset Sensitivity to Endotoxin Tolerisation by Porphyromonas gingivalis

Macrophages (MΦs) determine oral mucosal responses; mediating tolerance to commensal microbes and food whilst maintaining the capacity to activate immune defences to pathogens. MΦ responses are determined by both differentiation and activation stimuli, giving rise to two distinct subsets; pro-inflammatory M1- and anti-inflammatory/regulatory M2- MΦs. M2-like subsets predominate tolerance induction whereas M1 MΦs predominate in inflammatory pathologies, mediating destructive inflammatory mechanisms, such as those in chronic P.gingivalis (PG) periodontal infection. MΦ responses can be suppressed to benefit either the host or the pathogen. Chronic stimulation by bacterial pathogen associated molecular patterns (PAMPs), such as LPS, is well established to induce tolerance. The aim of this study was to investigate the susceptibility of MΦ subsets to suppression by P. gingivalis. CD14^hi and CD14^lo M1- and M2-like MΦs were generated in vitro from the THP-1 monocyte cell line by differentiation with PMA and vitamin D₃, respectively. MΦ subsets were pre-treated with heat-killed PG (HKPG) and PG-LPS prior to stimulation by bacterial PAMPs. Modulation of inflammation was measured by TNFα, IL-1β, IL-6, IL-10 ELISA and NFκB activation by reporter gene assay. HKPG and PG-LPS differentially suppress PAMP-induced TNFα, IL-6 and IL-10 but fail to suppress IL-1β expression in M1 and M2 MΦs. In addition, P.gingivalis suppressed NFκB activation in CD14^lo and CD14^hi M2 regulatory MΦs and CD14^lo M1 MΦs whereas CD14^hi M1 pro-inflammatory MΦs were refractory to suppression. In conclusion, P.gingivalis selectively tolerises regulatory M2 MΦs with little effect on pro-inflammatory CD14^hi M1 MΦs; differential suppression facilitating immunopathology at the expense of immunity.     

Extract from Rumex acetosa L. for Prophylaxis of Periodontitis: Inhibition of Bacterial In Vitro Adhesion and of Gingipains of Porphyromonas gingivalis by Epicatechin-3-O-(4β→8)-Epicatechin-3-O-Gallate (Procyanidin-B2-Di-Gallate)

Background

The aerial parts of Rumex acetosa L. have been used in traditional European medicine for inflammatory diseases of the mouth epithelial tissue. The following study aimed to investigate the influence of a proanthocyanidin-enriched extract from R. acetosa extract against the adhesion of Porphyromonas gingivalis (P. gingivalis), a pathogen strongly involved in chronic and aggressive periodontitis. A further goal was to define the bioactive lead structures responsible for a potential antiadhesive activity and to characterize the underlying molecular mechanisms of the antiadhesive effects.

Methodology

An extract of R. acetosa (RA1) with a defined mixture of flavan-3-ols, oligomeric proanthocyanidins and flavonoids, was used. Its impact on P. gingivalis adhesion to KB cells was studied by flow cytometry, confocal laser scanning microscopy and in situ adhesion assay using murine buccal tissue. RA1 and its compounds 1 to 15 were further investigated for additional effects on gingipain activity, hemagglutination and gene expression by RT-PCR.

Principal Findings

RA1 (5 to 15 μg/mL) reduced P. gingivalis adhesion in a dose-dependent manner to about 90%. Galloylated proanthocyanidins were confirmed to be responsible for this antiadhesive effect with epicatechin-3-O-gallate-(4β,8)-epicatechin-3’-O-gallate (syn. procyanidin B2-di-gallate) being the lead compound. Ungalloylated flavan-3-ols and oligomeric proanthocyanidins were inactive. RA1 and the galloylated proanthocyanidins strongly interact with the bacterial virulence factor Arg-gingipain, while the corresponding Lys-gingipain was hardly influenced. RA1 inhibited also hemagglutination. In silico docking studies indicated that epicatechin-3-O-gallate-(4β,8)-epicatechin-3’-O-gallate interacts with the active side of Arg-gingipain and hemaglutinin from P. gingivalis; the galloylation of the molecule seems to be responsible for fixation of the ligand to the protein. In conclusion, the proanthocyanidin-enriched extract RA1 and its main active constituent procyanidin B2-di-gallate protect cells from P. gingivalis infection by inhibiting bacterial adhesion to the host cell. RA1 and procyanidin B2-di-gallate appear to be promising candidates for future cytoprotective preparations for oral mouth care products.     

SGK1 negatively regulates inflammatory immune responses and protects against alveolar bone loss through modulation of TRAF3 activity

Serum- and glucocorticoid-regulated kinase 1 (SGK1) is a serine/threonine kinase that plays important roles in the cellular stress response. While SGK1 has been reported to restrain inflammatory immune responses, the molecular mechanisms involved remain elusive, especially in oral bacteria-induced inflammatory milieu. Here, we found that SGK1 curtails Porphyromonas gingivalis–induced inflammatory responses through maintaining levels of tumor necrosis factor receptor-associated factor (TRAF) 3, thereby suppressing NF-κB signaling. Specifically, SGK1 inhibition significantly enhances production of proinflammatory cytokines, including tumor necrosis factor α, interleukin (IL)-6, IL-1β, and IL-8 in P. gingivalis–stimulated innate immune cells. The results were confirmed with siRNA and LysM-Cre–mediated SGK1 KO mice. Moreover, SGK1 deletion robustly increased NF-κB activity and c-Jun expression but failed to alter the activation of mitogen-activated protein kinase signaling pathways. Further mechanistic data revealed that SGK1 deletion elevates TRAF2 phosphorylation, leading to TRAF3 degradation in a proteasome-dependent manner. Importantly, siRNA-mediated traf3 silencing or c-Jun overexpression mimics the effect of SGK1 inhibition on P. gingivalis–induced inflammatory cytokines and NF-κB activation. In addition, using a P. gingivalis infection–induced periodontal bone loss model, we found that SGK1 inhibition modulates TRAF3 and c-Jun expression, aggravates inflammatory responses in gingival tissues, and exacerbates alveolar bone loss. Altogether, we demonstrated for the first time that SGK1 acts as a rheostat to limit P. gingivalis–induced inflammatory immune responses and mapped out a novel SGK1–TRAF2/3–c-Jun–NF-κB signaling axis. These findings provide novel insights into the anti-inflammatory molecular mechanisms of SGK1 and suggest novel interventional targets to inflammatory diseases relevant beyond the oral cavity.     

Porphyromonas gingivalis oral infection exacerbates the development and severity of collagen-induced arthritis

Introduction

Clinical studies suggest a direct influence of periodontal disease (PD) on serum inflammatory markers and disease assessment of patients with established rheumatoid arthritis (RA). However, the influence of PD on arthritis development remains unclear. This investigation was undertaken to determine the contribution of chronic PD to immune activation and development of joint inflammation using the collagen-induced arthritis (CIA) model.

Methods

DBA1/J mice orally infected with Porphyromonas gingivalis were administered with collagen II (CII) emulsified in complete Freund’s adjuvant (CFA) or incomplete Freund’s adjuvant (IFA) to induce arthritis. Arthritis development was assessed by visual scoring of paw swelling, caliper measurement of the paws, mRNA expression, paw micro-computed tomography (micro-CT) analysis, histology, and tartrate resistant acid phosphatase for osteoclast detection (TRAP)-positive immunohistochemistry. Serum and reactivated splenocytes were evaluated for cytokine expression.

Results

Mice induced for PD and/or arthritis developed periodontal disease, shown by decreased alveolar bone and alteration of mRNA expression in gingival tissues and submandibular lymph nodes compared to vehicle. P. gingivalis oral infection increased paw swelling and osteoclast numbers in mice immunized with CFA/CII. Arthritis incidence and severity were increased by P. gingivalis in mice that received IFA/CII immunizations. Increased synovitis, bone erosions, and osteoclast numbers in the paws were observed following IFA/CII immunizations in mice infected with P gingivalis. Furthermore, cytokine analysis showed a trend toward increased serum Th17/Th1 ratios when P. gingivalis infection was present in mice receiving either CFA/CII or IFA/CII immunizations. Significant cytokine increases induced by P. gingivalis oral infection were mostly associated to Th17-related cytokines of reactivated splenic cells, including IL-1β, IL-6, and IL-22 in the CFA/CII group and IL-1β, tumor necrosis factor-α, transforming growth factor-β, IL-6 and IL-23 in the IFA/CII group.

Conclusions

Chronic P. gingivalis oral infection prior to arthritis induction increases the immune system activation favoring Th17 cell responses, and ultimately accelerating arthritis development. These results suggest that chronic oral infection may influence RA development mainly through activation of Th17-related pathways.     

Polymicrobial Oral Infection with Four Periodontal Bacteria Orchestrates a Distinct Inflammatory Response and Atherosclerosis in ApoEnull Mice

Periodontal disease (PD) develops from a synergy of complex subgingival oral microbiome, and is linked to systemic inflammatory atherosclerotic vascular disease (ASVD). To investigate how a polybacterial microbiome infection influences atherosclerotic plaque progression, we infected the oral cavity of ApoE^null mice with a polybacterial consortium of 4 well-characterized periodontal pathogens, Porphyromonas gingivalis, Treponema denticola, Tannerealla forsythia and Fusobacterium nucleatum, that have been identified in human atherosclerotic plaque by DNA screening. We assessed periodontal disease characteristics, hematogenous dissemination of bacteria, peripheral T cell response, serum inflammatory cytokines, atherosclerosis risk factors, atherosclerotic plaque development, and alteration of aortic gene expression. Polybacterial infections have established gingival colonization in ApoE^null hyperlipidemic mice and displayed invasive characteristics with hematogenous dissemination into cardiovascular tissues such as the heart and aorta. Polybacterial infection induced significantly higher levels of serum risk factors oxidized LDL (p < 0.05), nitric oxide (p < 0.01), altered lipid profiles (cholesterol, triglycerides, Chylomicrons, VLDL) (p < 0.05) as well as accelerated aortic plaque formation in ApoE^null mice (p < 0.05). Periodontal microbiome infection is associated with significant decreases in Apoa1, Apob, Birc3, Fga, FgB genes that are associated with atherosclerosis. Periodontal infection for 12 weeks had modified levels of inflammatory molecules, with decreased Fas ligand, IL-13, SDF-1 and increased chemokine RANTES. In contrast, 24 weeks of infection induced new changes in other inflammatory molecules with reduced KC, MCSF, enhancing GM-CSF, IFNγ, IL-1β, IL-13, IL-4, IL-13, lymphotactin, RANTES, and also an increase in select inflammatory molecules. This study demonstrates unique differences in the host immune response to a polybacterial periodontal infection with atherosclerotic lesion progression in a mouse model.     

Porphyromonas gingivalis GroEL Induces Osteoclastogenesis of Periodontal Ligament Cells and Enhances Alveolar Bone Resorption in Rats

Porphyromonas gingivalis is a major periodontal pathogen that contains a variety of virulence factors. The antibody titer to P. gingivalis GroEL, a homologue of HSP60, is significantly higher in periodontitis patients than in healthy control subjects, suggesting that P. gingivalis GroEL is a potential stimulator of periodontal disease. However, the specific role of GroEL in periodontal disease remains unclear. Here, we investigated the effect of P. gingivalis GroEL on human periodontal ligament (PDL) cells in vitro, as well as its effect on alveolar bone resorption in rats in vivo. First, we found that stimulation of PDL cells with recombinant GroEL increased the secretion of the bone resorption-associated cytokines interleukin (IL)-6 and IL-8, potentially via NF-κB activation. Furthermore, GroEL could effectively stimulate PDL cell migration, possibly through activation of integrin α1 and α2 mRNA expression as well as cytoskeletal reorganization. Additionally, GroEL may be involved in osteoclastogenesis via receptor activator of nuclear factor κ-B ligand (RANKL) activation and alkaline phosphatase (ALP) mRNA inhibition in PDL cells. Finally, we inoculated GroEL into rat gingiva, and the results of microcomputed tomography (micro-CT) and histomorphometric assays indicated that the administration of GroEL significantly increased inflammation and bone loss. In conclusion, P. gingivalis GroEL may act as a potent virulence factor, contributing to osteoclastogenesis of PDL cells and resulting in periodontal disease with alveolar bone resorption.     

Hyperlipidemia Impaired Innate Immune Response to Periodontal Pathogen Porphyromonas gingivalis in Apolipoprotein E Knockout Mice

A finely-tuned innate immune response plays a pivotal role in protecting host against bacterial invasion during periodontal disease progression. Hyperlipidemia has been suggested to exacerbate periodontal health condition. However, the underlying mechanism has not been addressed. In the present study, we investigated the effect of hyperlipidemia on innate immune responses to periodontal pathogen Porphyromonas gingivalis infection. Apolipoprotein E-deficient and wild-type mice at the age of 20 weeks were used for the study. Peritoneal macrophages were isolated and subsequently used for the study of viable P. gingivalis infection. ApoE^−/− mice demonstrated inhibited iNOS production and impaired clearance of P. gingivalis in vitro and in vivo; furthermore, ApoE^−/− mice displayed disrupted cytokine production pattern in response to P. gingivalis, with a decreased production of tumor necrosis factor-α, interleukin-6 (IL-6), IL-1β and monocyte chemotactic protein-1. Microarray data demonstrated that Toll-like receptor (TLR) and NOD-like receptor (NLR) pathway were altered in ApoE^−/− mice macrophages; further analysis of pattern recognition receptors (PRRs) demonstrated that expression of triggering receptors on myeloid cells-1 (TREM-1), an amplifier of the TLR and NLR pathway, was decreased in ApoE^−/− mice macrophages, leading to decreased recruitment of NF-κB onto the promoters of the TNF-α and IL-6. Our data suggest that in ApoE^−/− mice hyperlipidemia disrupts the expression of PRRs, and cripples the host’s capability to generate sufficient innate immune response to P. gingivalis, which may facilitate immune evasion, subgingival colonization and establishment of P. gingivalis in the periodontal niche.     

Brucella Cyclic β-1,2-Glucan Plays a Critical Role in the Induction of Splenomegaly in Mice

Brucella, the etiological agent of animal and human brucellosis, is a bacterium with the capacity to modulate the inflammatory response. Cyclic β-1,2-glucan (CβG) is a virulence factor key for the pathogenesis of Brucella as it is involved in the intracellular life cycle of the bacteria. Using comparative studies with different CβG mutants of Brucella, cgs (CβG synthase), cgt (CβG transporter) and cgm (CβG modifier), we have identified different roles for this polysaccharide in Brucella. While anionic CβG is required for bacterial growth in low osmolarity conditions, the sole requirement for a successful Brucella interaction with mammalian host is its transport to periplasmic space. Our results uncover a new role for CβG in promoting splenomegaly in mice. We showed that CβG-dependent spleen inflammation is the consequence of massive cell recruitment (monocytes, dendritics cells and neutrophils) due to the induction of pro-inflammatory cytokines such as IL-12 and TNF-α and also that the reduced splenomegaly response observed with the cgs mutant is not the consequence of changes in expression levels of the characterized Brucella PAMPs LPS, flagellin or OMP16/19. Complementation of cgs mutant with purified CβG increased significantly spleen inflammation response suggesting a direct role for this polysaccharide.     

New Microbicidal Functions of Tracheal Glands: Defective Anti-Infectious Response to Pseudomonas aeruginosa in Cystic Fibrosis

Tracheal glands (TG) may play a specific role in the pathogenesis of cystic fibrosis (CF), a disease due to mutations in the cftr gene and characterized by airway inflammation and Pseudomonas aeruginosa infection. We compared the gene expression of wild-type TG cells and TG cells with the cftr ΔF508 mutation (CF-TG cells) using microarrays covering the whole human genome. In the absence of infection, CF-TG cells constitutively exhibited an inflammatory signature, including genes that encode molecules such as IL-1α, IL-β, IL-32, TNFSF14, LIF, CXCL1 and PLAU. In response to P. aeruginosa, genes associated with IFN-γ response to infection (CXCL10, IL-24, IFNγR2) and other mediators of anti-infectious responses (CSF2, MMP1, MMP3, TLR2, S100 calcium-binding proteins A) were markedly up-regulated in wild-type TG cells. This microbicidal signature was silent in CF-TG cells. The deficiency of genes associated with IFN-γ response was accompanied by the defective membrane expression of IFNγR2 and altered response of CF-TG cells to exogenous IFN-γ. In addition, CF-TG cells were unable to secrete CXCL10, IL-24 and S100A8/S100A9 in response to P. aeruginosa. The differences between wild-type TG and CF-TG cells were due to the cftr mutation since gene expression was similar in wild-type TG cells and CF-TG cells transfected with a plasmid containing a functional cftr gene. Finally, we reported an altered sphingolipid metabolism in CF-TG cells, which may account for their inflammatory signature. This first comprehensive analysis of gene expression in TG cells proposes a protective role of wild-type TG against airborne pathogens and reveals an original program in which anti-infectious response was deficient in TG cells with a cftr mutation. This defective response may explain why host response does not contribute to protection against P. aeruginosa in CF.     

Role of Staphylococcus aureus Virulence Factors in Inducing Inflammation and Vascular Permeability in a Mouse Model of Bacterial Endophthalmitis

Staphylococcus (S.) aureus is a common causative agent of bacterial endophthalmitis, a vision threatening complication of eye surgeries. The relative contribution of S. aureus virulence factors in the pathogenesis of endophthalmitis remains unclear. Here, we comprehensively analyzed the development of intraocular inflammation, vascular permeability, and the loss of retinal function in C57BL/6 mouse eyes, challenged with live S. aureus, heat-killed S. aureus (HKSA), peptidoglycan (PGN), lipoteichoic acid (LTA), staphylococcal protein A (SPA), α-toxin, and Toxic-shock syndrome toxin 1 (TSST1). Our data showed a dose-dependent (range 0.01 μg/eye to 1.0 μg/eye) increase in the levels of inflammatory mediators by all virulence factors. The cell wall components, particularly PGN and LTA, seem to induce higher levels of TNF-α, IL-6, KC, and MIP2, whereas the toxins induced IL-1β. Similarly, among the virulence factors, PGN induced higher PMN infiltration. The vascular permeability assay revealed significant leakage in eyes challenged with live SA (12-fold) and HKSA (7.3-fold), in comparison to other virulence factors (~2-fold) and controls. These changes coincided with retinal tissue damage, as evidenced by histological analysis. The electroretinogram (ERG) analysis revealed a significant decline in retinal function in eyes inoculated with live SA, followed by HKSA, SPA, and α-toxin. Together, these findings demonstrate the differential innate responses of the retina to S. aureus virulence factors, which contribute to intraocular inflammation and retinal function loss in endophthalmitis.