Murine glia express the immunosuppressive cytokine, interleukin-10, following exposure to Borrelia burgdorferi or Neisseria meningitidis.

There is growing appreciation that resident glial cells can initiate and/or regulate inflammation following trauma or infection in the central nervous system (CNS). We have previously demonstrated the ability of microglia and astrocytes, resident glial cells of the CNS, to respond to bacterial pathogens by rapid production of inflammatory mediators. However, inflammation within the brain parenchyma is notably absent during some chronic bacterial infections in humans and nonhuman primates. In the present study, we demonstrate the ability of the immunosuppressive cytokine, interleukin-10 (IL-10), to inhibit inflammatory immune responses of primary microglia and astrocytes to B. burgdorferi and N. meningitidis, two disparate gram negative bacterial species that can cross the blood-brain barrier in humans. Importantly, we demonstrate that these organisms induce the delayed production of significant quantities of IL-10 by both microglia and astrocytes. Furthermore, we demonstrate that such production occurs independent of the actions of bacterial lipopolysaccharide and is secondary to the autocrine or paracrine actions of other glia-derived soluble mediators. The late onset of IL-10 production by resident glia following activation, the previously documented expression of specific receptors for this cytokine on microglia and astrocytes, and the ability of IL-10 to inhibit bacterially induced immune responses by these cells, suggest a mechanism by which resident glial cells can limit potentially damaging inflammation within the CNS in response to invading pathogens, and could explain the suppression of inflammation seen within the brain parenchyma during chronic bacterial infections.

Induction of Nod1 and Nod2 intracellular pattern recognition receptors in murine osteoblasts following bacterial challenge.

Osteoblasts produce an array of immune molecules following bacterial challenge that could recruit leukocytes to sites of infection and promote inflammation during bone diseases, such as osteomyelitis. Recent studies from our laboratory have shed light on the mechanisms by which this cell type can perceive and respond to bacteria by demonstrating the functional expression of members of the Toll-like family of cell surface pattern recognition receptors by osteoblasts. However, we have shown that bacterial components fail to elicit immune responses comparable with those seen following challenge with the intracellular pathogens salmonellae and Staphylococcus aureus. In the present study, we show that UV-killed bacteria and invasion-defective bacterial strains elicit significantly less inflammatory cytokine production than their viable wild-type counterparts. Importantly, we demonstrate that murine osteoblasts express the novel intracellular pattern recognition receptors Nod1 and Nod2. Levels of mRNA encoding Nod molecules and protein expression are significantly and differentially increased from low basal levels following exposure to these disparate bacterial pathogens. In addition, we have shown that osteoblasts express Rip2 kinase, a critical downstream effector molecule for Nod signaling. Furthermore, to begin to establish the functional nature of Nod expression, we show that a specific ligand for Nod proteins can significantly augment immune molecule production by osteoblasts exposed to either UV-inactivated bacteria or bacterial lipopolysaccharide. As such, the presence of Nod proteins in osteoblasts could represent an important mechanism by which this cell type responds to intracellular bacterial pathogens of bone.

Osteoblasts express the inflammatory cytokine interleukin-6 in a murine model of Staphylococcus aureus osteomyelitis and infected human bone tissue.

Staphylococcus aureus is the single most common cause of osteomyelitis in humans. Incidences of osteomyelitis caused by S. aureus have increased dramatically in recent years, in part due to the appearance of community-acquired antibiotic resistant strains. Therefore, understanding the pathogenesis of this organism has become imperative. Recently, we have described the surprising ability of bone-forming osteoblasts to secrete a number of important immune mediators when exposed to S. aureus in vitro. In the present study, we provide the first evidence for the in vivo production of such molecules by osteoblasts during bacterial infection of bone. These studies demonstrate the expression of the key inflammatory cytokine interleukin-6 by osteoblasts in organ cultures of neonatal mouse calvaria, and in vivo using a mouse model that closely resembles the pathology of trauma-induced staphylococcal osteomyelitis, as determined by confocal microscopic analysis. Importantly, we have established the clinical relevancy of these findings in infected human bone tissue from patients with S. aureus-associated osteomyelitis. As such, these studies demonstrate that bacterial challenge of osteoblasts during bone diseases, such as osteomyelitis, induces cells to produce inflammatory molecules that can direct appropriate host responses or contribute to progressive inflammatory damage.

Signaling via Toll-like receptor 5 can initiate inflammatory mediator production by murine osteoblasts.

Murine osteoblasts express Toll-like receptor 5 (TLR5), and this expression is upregulated following exposure to bacteria or to the TLR5 agonist, flagellin. Importantly, flagellin activates transcriptional regulators and elicits proinflammatory cytokine production, suggesting TLR5 functionality. TLR5 may represent an important mechanism underlying the recognition of bacterial pathogens by osteoblasts during bone infections.

Cultured astrocytes express toll-like receptors for bacterial products.

It has become apparent that astrocytes may be important contributors to inflammatory immune responses within the brain in response to microbial challenges. To date, the mechanisms that underlie activation of this major glial cell type by such challenges have not been investigated. In the present study, we present evidence for members of a recently discovered family of receptors for highly conserved microbial components, the Toll-like receptors (TLRs), in isolated cultures of primary murine astrocytes. We describe the low-level constitutive expression of messenger RNA-encoding TLR2, TLR4, TLR5, and TLR9 in resting cultures of these cells. Importantly, the level of expression of messenger RNA for each of these receptors is markedly elevated following exposure to specific bacteria-derived ligands for these receptors. The functional expression of these receptor proteins is further supported by the ability of known ligands for each TLR to induce both message expression and protein secretion of the proinflammatory cytokine, interleukin-6. In addition, the recent availability of antibodies to TLR2 and TLR4 has enabled us to demonstrate directly the presence of these receptors on astrocytes by Western blot and immunofluorescence analysis, respectively. Furthermore, we have confirmed the sensitivity of such receptor expression to ligand stimulation. The present demonstration of Toll-like microbial pattern-recognition receptors on primary astrocytes provides a mechanistic link between bacterial challenge and inflammatory immune responses that may be an important component of the pathologies of bacterially induced inflammatory CNS disorders.