Translocation of particles and inflammatory responses after exposure to fine particles and nanoparticles in an epithelial airway model

ABSTRACT: BACKGROUND: Experimental studies provide evidence that inhaled nanoparticles may translocate over the airspace epithelium and cause increased cellular inflammation. Little is known, however, about the dependence of particle size or material on translocation characteristics, inflammatory response and intracellular localization. RESULTS: Using a triple cell co-culture model of the human airway wall composed of epithelial cells, macrophages and dendritic cells we quantified the entering of fine (1 mum) and nano-sized (0.078 mum) polystyrene particles by laser scanning microscopy. The number distribution of particles within the cell types was significantly different between fine and nano-sized particles suggesting different translocation characteristics. Analysis of the intracellular localization of gold (0.025 mum) and titanium dioxide (0.02-0.03 mum) nanoparticles by energy filtering transmission electron microscopy showed differences in intracellular localization depending on particle composition. Titanium dioxide nanoparticles were detected as single particles without membranes as well as in membrane-bound agglomerations. Gold nanoparticles were found inside the cells as free particles only. The potential of the different particle types (different sizes and different materials) to induce a cellular response was determined by measurements of the tumour necrosis factor-alpha in the supernatants. We measured a 2-3 fold increase of tumour necrosis factor-alpha in the supernatants after applying 1 mum polystyrene particles, gold nanoparticles, but not with polystyrene and titanium dioxide nanoparticles. CONCLUSION: Quantitative laser scanning microscopy provided evidence that the translocation and entering characteristics of particles are size-dependent. Energy filtering transmission electron microscopy showed that the intracellular localization of nanoparticles depends on the particle material. Both particle size and material affect the cellular responses to particle exposure as measured by the generation of tumour necrosis factor-alpha.

Matrix metalloproteinase-19 is a predictive marker for tumor invasiveness in patients with oropharyngeal squamous cell carcinoma

AIMS: To evaluate the expression of matrix metalloproteinase-19 (MMP-19) in oropharyngeal squamous cell carcinoma along with its association with structural features of invasiveness. To investigate whether MMP-19 expression correlates with lymphatic or systemic metastasis and prognosis in patients who have received definitive radiotherapy. METHODS AND RESULTS: The histological evaluation of the invasive front was based on Bryne's malignancy grading system. We correlated the immunohistochemical expression pattern with morphological parameters which characterize tumor invasiveness such as keratinization, nuclear polymorphism, invasion pattern, and the host inflammatory response. Local immunoreactivity for MMP-19 was positively correlated with tumor invasiveness as reflected in its structural characteristics and the degree of nuclear polymorphism, and negatively correlated with the inflammatory response of the host. No correlation existed between MMP-19 expression and clinicopathological features (TNM stage, grade of differentiation) or a patient''s outcome and prognosis. CONCLUSIONS: This latter finding probably reflects the unique change for MMPs from high immunoreactivity within healthy tissue areas and non-invasive tumor parts, through absence in the least invasive neoplastic regions, to strong re-expression at a highly invasive front of the same tumor. Our findings indicate that MMP-19 can be used as a marker for tumor invasiveness in patients with oropharyngeal squamous cell carcinoma.

Heme oxygenase-1 is a critical regulator of nitric oxide production in enterohemorrhagic Escherichia coli-infected human enterocytes

Enterohemorrhagic Escherichia coli (EHEC) are the causative agent of hemolytic-uremic syndrome. In the first stage of the infection, EHEC interact with human enterocytes to modulate the innate immune response. Inducible NO synthase (iNOS)-derived NO is a critical mediator of the inflammatory response of the infected intestinal mucosa. We therefore aimed to analyze the role of EHEC on iNOS induction in human epithelial cell lines. In this regard, we show that EHEC down-regulate IFN-gamma-induced iNOS mRNA expression and NO production in Hct-8, Caco-2, and T84 cells. This inhibitory effect occurs through the decrease of STAT-1 activation. In parallel, we demonstrate that EHEC stimulate the rapid inducible expression of the gene hmox-1 that encodes for the enzyme heme oxygenase-1 (HO-1). Knock-down of hmox-1 gene expression by small interfering RNA or the blockade of HO-1 activity by zinc protoporphyrin IX abrogated the EHEC-dependent inhibition of STAT-1 activation and iNOS mRNA expression in activated human enterocytes. These results highlight a new strategy elaborated by EHEC to control the host innate immune response.

Comparison of two non-bronchoscopic methods for evaluating inflammation in patients with acute hypoxaemic respiratory failure

INTRODUCTION: The simple bedside method for sampling undiluted distal pulmonary edema fluid through a normal suction catheter (s-Cath) has been experimentally and clinically validated. However, there are no data comparing non-bronchoscopic bronchoalveolar lavage (mini-BAL) and s-Cath for assessing lung inflammation in acute hypoxaemic respiratory failure. We designed a prospective study in two groups of patients, those with acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) and those with acute cardiogenic lung edema (ACLE), designed to investigate the clinical feasibility of these techniques and to evaluate inflammation in both groups using undiluted sampling obtained by s-Cath. To test the interchangeability of the two methods in the same patient for studying the inflammation response, we further compared mini-BAL and s-Cath for agreement of protein concentration and percentage of polymorphonuclear cells (PMNs). METHODS: Mini-BAL and s-Cath sampling was assessed in 30 mechanically ventilated patients, 21 with ALI/ARDS and 9 with ACLE. To analyse agreement between the two sampling techniques, we considered only simultaneously collected mini-BAL and s-Cath paired samples. The protein concentration and polymorphonuclear cell (PMN) count comparisons were performed using undiluted sampling. Bland-Altman plots were used for assessing the mean bias and the limits of agreement between the two sampling techniques; comparison between groups was performed by using the non-parametric Mann-Whitney-U test; continuous variables were compared by using the Student t-test, Wilcoxon signed rank test, analysis of variance or Student-Newman-Keuls test; and categorical variables were compared by using chi-square analysis or Fisher exact test. RESULTS: Using protein content and PMN percentage as parameters, we identified substantial variations between the two sampling techniques. When the protein concentration in the lung was high, the s-Cath was a more sensitive method; by contrast, as inflammation increased, both methods provided similar estimates of neutrophil percentages in the lung. The patients with ACLE showed an increased PMN count, suggesting that hydrostatic lung edema can be associated with a concomitant inflammatory process. CONCLUSIONS: There are significant differences between the s-Cath and mini-BAL sampling techniques, indicating that these procedures cannot be used interchangeably for studying the lung inflammatory response in patients with acute hypoxaemic lung injury.

Coordinated changes in mRNA turnover, translation, and RNA processing bodies in bronchial epithelial cells following inflammatory stimulation.

Bronchial epithelial cells play a pivotal role in airway inflammation, but little is known about posttranscriptional regulation of mediator gene expression during the inflammatory response in these cells. Here, we show that activation of human bronchial epithelial BEAS-2B cells by proinflammatory cytokines interleukin-4 (IL-4) and tumor necrosis factor alpha (TNF-alpha) leads to an increase in the mRNA stability of the key chemokines monocyte chemotactic protein 1 and IL-8, an elevation of the global translation rate, an increase in the levels of several proteins critical for translation, and a reduction of microRNA-mediated translational repression. Moreover, using the BEAS-2B cell system and a mouse model, we found that RNA processing bodies (P bodies), cytoplasmic domains linked to storage and/or degradation of translationally silenced mRNAs, are significantly reduced in activated bronchial epithelial cells, suggesting a physiological role for P bodies in airway inflammation. Our study reveals an orchestrated change among posttranscriptional mechanisms, which help sustain high levels of inflammatory mediator production in bronchial epithelium during the pathogenesis of inflammatory airway diseases.

The causative pathogen determines the inflammatory profile in cerebrospinal fluid and outcome in patients with bacterial meningitis

BACKGROUND The brain's inflammatory response to the infecting pathogen determines the outcome of bacterial meningitis (BM), for example, the associated mortality and the extent of brain injury. The inflammatory cascade is initiated by the presence of bacteria in the cerebrospinal fluid (CSF) activating resident immune cells and leading to the influx of blood derived leukocytes. To elucidate the pathomechanisms behind the observed difference in outcome between different pathogens, we compared the inflammatory profile in the CSF of patients with BM caused by Streptococcus pneumonia (n = 14), Neisseria meningitidis (n = 22), and Haemophilus influenza (n = 9). METHODS CSF inflammatory parameters, including cytokines and chemokines, MMP-9, and nitric oxide synthase activity, were assessed in a cohort of patients with BM from Burkina Faso. RESULTS Pneumococcal meningitis was associated with significantly higher CSF concentrations of IFN-γ , MCP-1, and the matrix-metalloproteinase (MMP-) 9. In patients with a fatal outcome, levels of TNF-α, IL-1 β, IL-1RA, IL-6, and TGF-α were significantly higher. CONCLUSION The signature of pro- and anti-inflammatory mediators and the intensity of inflammatory processes in CSF are determined by the bacterial pathogen causing bacterial meningitis with pneumococcal meningitis being associated with a higher case fatality rate than meningitis caused by N. meningitidis or H. influenzae.

Dominance of chemokine ligand 2 and matrix metalloproteinase-2 and -9 and suppression of pro-inflammatory cytokines in the epidural compartment after intervertebral disc extrusion in a canine model

BACKGROUND CONTEXT In canine intervertebral disc (IVD) disease, a useful animal model, only little is known about the inflammatory response in the epidural space. PURPOSE To determine messenger RNA (mRNA) expressions of selected cytokines, chemokines, and matrix metalloproteinases (MMPs) qualitatively and semiquantitatively over the course of the disease and to correlate results to neurologic status and outcome. STUDY DESIGN/SETTING Prospective study using extruded IVD material of dogs with thoracolumbar IVD extrusion. PATIENT SAMPLE Seventy affected and 13 control (24 samples) dogs. OUTCOME MEASURES Duration of neurologic signs, pretreatment, neurologic grade, severity of pain, and outcome were recorded. After diagnostic imaging, decompressive surgery was performed. METHODS Messenger RNA expressions of interleukin (IL)-1β, IL-2, IL-4, IL-6, IL-8, IL-10, tumor necrosis factor (TNF), interferon (IFN)γ, MMP-2, MMP-9, chemokine ligand (CCL)2, CCL3, and three housekeeping genes was determined in the collected epidural material by Panomics 2.0 QuantiGene Plex technology. Relative mRNA expression and fold changes were calculated. Relative mRNA expression was correlated statistically to clinical parameters. RESULTS Fold changes of TNF, IL-1β, IL-2, IL-4, IL-6, IL-10, IFNγ, and CCL3 were clearly downregulated in all stages of the disease. MMP-9 was downregulated in the acute stage and upregulated in the subacute and chronic phase. Interleukin-8 was upregulated in acute cases. MMP-2 showed mild and CCL2 strong upregulation over the whole course of the disease. In dogs with severe pain, CCL3 and IFNγ were significantly higher compared with dogs without pain (p=.017/.020). Dogs pretreated with nonsteroidal anti-inflammatory drugs revealed significantly lower mRNA expression of IL-8 (p=.017). CONCLUSIONS The high CCL2 levels and upregulated MMPs combined with downregulated T-cell cytokines and suppressed pro-inflammatory genes in extruded canine disc material indicate that the epidural reaction is dominated by infiltrating monocytes differentiating into macrophages with tissue remodeling functions. These results will help to understand the pathogenic processes representing the basis for novel therapeutic approaches. The canine IVD disease model will be rewarding in this process.

Myositides : What is the current situation?

This article gives a review of the classification, diagnostic procedures and treatment of idiopathic inflammatory myopathies from a neurological point of view. The myositis syndromes can be subdivided into four groups, polymyositis (PM), dermatomyositis (DM), inclusion body myositis (IBM) and necrotizing myopathy (NM), which substantially differ clinically and pathophysiologically. Myositis may also occur in association with cancer or autoimmune systemic diseases (overlap syndrome). Diagnosis of inflammatory myopathies is based on clinical symptoms, determination of creatine phosphokinase and acute phase parameters in blood (e.g. C-reactive protein and erythrocyte sedimentation rate), electromyography results and findings of magnetic resonance imaging (MRI) in muscle. A muscle biopsy is mandatory to confirm the diagnosis. High quality randomized controlled trials of treatment regimens for inflammatory myopathies are sparse; however, empirical experience indicates a clear effectiveness of immunosuppressive treatment of PM, DM and NM.

Salivary Pellicle vs Whole Saliva: The Response of Oral Fibroblasts Based on a Genome-Wide Microarray.

PURPOSE Whole saliva comprises components of the salivary pellicle that spontaneously forms on surfaces of implants and teeth. However, there are no studies that functionally link the salivary pellicle with a possible change in gene expression. MATERIALS AND METHODS This study examined the genetic response of oral fibroblasts exposed to the salivary pellicle and whole saliva. Oral fibroblasts were seeded onto a salivary pellicle and the respective untreated surface. Oral fibroblasts were also exposed to freshly harvested sterile-filtered whole saliva. A genome-wide microarray of oral fibroblasts was performed, followed by gene ontology screening with DAVID functional annotation clustering, KEGG pathway analysis, and the STRING functional protein association network. RESULTS Exposure of oral fibroblasts to saliva caused 61 genes to be differentially expressed (P < .05). Gene ontology screening assigned the respective genes into 262 biologic processes, 3 cellular components, 13 molecular functions, and 7 pathways. Most remarkable was the enrichment in the inflammatory response. None of the genes regulated by whole saliva was significantly changed when cells were placed onto a salivary pellicle. CONCLUSION The salivary pellicle per se does not provoke a significant inflammatory response of oral fibroblasts in vitro, whereas sterile-filtered whole saliva does produce a strong inflammatory response.

Organ inflammation in porcine Escherichia coli sepsis is markedly attenuated by combined inhibition of C5 and CD14.

Sepsis is an infection-induced systemic inflammatory syndrome, potentially causing organ failure. We previously showed attenuating effects on inflammation, thrombogenicity and haemodynamics by inhibiting the Toll-like receptor co-factor CD14 and complement factor C5 in a porcine Escherichia coli-induced sepsis model. The present study explored the effect on organ inflammation in these pigs. Tissue samples were examined from the combined treatment group (n = 8), the positive (n = 8) and negative (n = 6) control groups after 4h of sepsis. Inflammatory biomarkers were measured using ELISA, multiplex and qPCR analysis. Combined inhibition of C5 and CD14 markedly attenuated IL-1β by 31-66% (P < 0.05) and IL-6 by 54-96% (P < 0.01) in liver, kidney, lung and spleen; IL-8 by 65-100% in kidney, lung, spleen, and heart (P < 0.05) and MCP-1 by 46-69% in liver, kidney, spleen and heart (P < 0.05). Combined inhibition significantly attenuated tissue factor mRNA upregulation in spleen (P < 0.05) and IP-10 mRNA upregulation in four out of five organs. Finally, C5aR mRNA downregulation was prevented in heart and kidney (P < 0.05). Combined inhibition of C5 and CD14 thus markedly attenuated inflammatory responses in all organs examined. The anti-inflammatory effects observed in lung and heart may explain the delayed haemodynamic disturbances observed in septic pigs receiving combined inhibition of C5 and CD14.

Effects of Estrogen on Muscle Damage in Response to an Acute Resistance Exercise Protocol

Creatine Kinase (CK) is used as a measure of exercise-induced muscle membrane damage. During acute eccentric (muscle lengthening) exercise, muscle sarcolemma, sarcoplasmic reticulum, and Z-lines are damaged, thus causing muscle proteins and enzymes to leak into the interstitial fluid. Strenuous eccentric exercise produces an elevation of oxygen free radicals, which further increases muscle damage. Muscle soreness and fatigue can be attributed to this membrane damage. Estradiol, however, may preserve membrane stability post-exercise (Brancaccio, Maffulli, & Limongelli, 2007; Carter, Dobridge, & Hackney, 2001; Tiidus, 2001). Because estradiol has a similar structure to Vitamin E, which is known to have antioxidant properties, and both are known to affect membrane structure, researchers have proposed that estrogen acts as an antioxidant to provide a protective effect on the post-exercise muscle of women (Sandoval & Matt, 2002). As a result, it has been postulated that muscles in women incur less damage in response to an acute strenuous exercise as compared to men. PURPOSE: To determine if circulating estrogen concentrations are related to muscle damage, as measured by creatine kinase activity and to determine gender differences in creatine kinase as a marker of muscle damage in response to an acute heavy resistance exercise protocol. METHODS: 7 healthy, resistance-trained, eumenhorrheic women (23±3 y, 169±9.1 cm, 66.4±10.5 kg) and 8 healthy, resistance-trained men (25±5 y, 178±6.7 cm, 82.3±9.33 kg) volunteered to participate in the study. Subjects performed an Acute Resistance Exercise Test (ARET) consisting of 6 sets of 5 repetitions Smith machine squats at 90% of their previously determined 1-RM. Blood samples were taken pre-, mid-, post-, 1 hour post-, 6 hours post-, and 24 hours post-exercise. Samples were stored at -80ºC until analyzed. Serum creatine kinase was measured using an assay kit from Genzyme (Framingham, MA). Serum estradiol was measured by an ELISA from GenWay (San Diego, CA). Estradiol b-receptor presence on granulocytes was measured via flow cytometry using primary antibodies from Abcam (Cambridge, MA) and PeCy7 antibodies (secondary) from Santa Cruz (Santa Cruz, CA). RESULTS: No significant correlations between estrogen and CK response were found after an acute resistant exercise protocol. Moreover, no significant change in estradiol receptors were expressed on granulocytes after exercise. Creatine Kinase response, however, differed significantly between genders. Men had higher resting CK concentrations throughout all time points. Creatine Kinase response increased significantly after exercise in both men and women (p=0.008, F=9.798). Men had a significantly higher CK response at 24 hours post exercise than women. A significant condition/sex/time interaction was exhibited in CK response (p=0.02, F=4.547). Perceived general soreness presented a significant condition, sex interaction (p=0.01, F=9.532). DISCUSSION: Although no estradiol and CK response correlations were found in response to exercise, a significant difference in creatine kinase activity was present between men and women. This discrepancy of our results and findings in the literature may be due to the high variability between subjects in creatine kinase activity as well as estrogen concentrations. The lack of significance in change of estradiol receptor expression on granulocytes in response to exercise may be due to intracellular estradiol receptor staining and non-specific gating for granulocytes rather than additional staining for neutrophil markers. Because neutrophils are the initial cells present in the inflammatory response after strenuous exercise, staining for estrogen receptors on this cell type may allow for a better understanding of the effect of estrogen and its hypothesized protective effect against muscle damage. Furthermore, the mechanism of action may include estradiol receptor expression on the muscle fiber itself may play a role in the protective effects of estradiol rather than or in addition to expression on neutrophils. We have shown here that gender differences occur in CK activity as a marker of muscle damage in response to strenuous eccentric exercise, but may not be the result of estradiol concentration or estradiol receptor expression on granulocytes. Other variables should be examined in order to determine the mechanism involved in the difference in creatine kinase as a marker of muscle damage between men and women after heavy resistance exercise.

Differential stress response and susceptibility to oxidative injury in alveolar macrophages from C57BL/6J and C3H/HeJ mice following oxidant exposure

Studies have demonstrated a variable response to ozone among individuals and animal species and strains. For instance, C57BL/6J mice have a greater inflammatory response to ozone exposure than C3H/HeJ mice. In these studies, I utilized these strain differences in an effort to derive a mechanistic explanation to the variable strain sensitivity to ozone exposure. Therefore, alveolar macrophages (AM) from C57BL/6J and C3H/HeJ mice were exposed in vitro to hydrogen peroxide ($\rm H\sb2O\sb2$), heat and acetyl ceramide or in vivo to ozone. Necrosis and DNA fragmentation in macrophages from the two murine strains were determined to assess cytotoxicity following these treatments. In addition, synthesis and expression of the stress proteins, stress protein 72 (SP72) and heme oxygenase (HO-1), were examined following treatments. The in vitro experiments were conducted to eliminate the possibility of in vivo confounders (i.e., differences in breathing rates in the two strains) and thus directly implicate some inherent difference between cells from the two murine strains. $\rm H\sb2O\sb2$ and heat caused greater cytotoxicity in AM from C57BL/6J than C3H/HeJ mice and DNA fragmentation was a particularly sensitive indicator of cell injury. Similarly, AM from C57BL/6J mice were more sensitive to ozone exposure than cells from C3H/HeJ mice. Exposure to either 1 or 0.4 ppm ozone caused greater cytotoxicity in macrophages from C57BL/6J mice compared to macrophages from C3H/HeJ mice. The increased sensitivity of AM to injury was associated with decreased synthesis and expression of stress proteins. AM from C57BL/6J mice synthesized and expressed significantly less stress proteins in response to heat and ozone than AM from C3H/HeJ mice. Heat treatment resulted in greater synthesis and expression of SP72. In addition, macrophages from C57BL/6J mice expressed lower amounts of HO-1 than macrophages from C3H/HeJ mice following 0.4 ppm ozone exposure. Therefore, AM from C57BL/6J mice are more susceptible to oxidative injury than AM from C3H/HeJ mice which might be due to differential expression of stress proteins in these cells. ^

Myelin basic protein kinase activity in tomato leaves is induced systemically by wounding and increases in response to systemin and oligosaccharide elicitors

In response to wounding, a 48-kDa myelin basic protein (MBP) kinase is activated within 2 min, both locally and systemically, in leaves of young tomato plants. The activating signal is able to pass through a steam girdle on the stem, indicating that it moves through the xylem and does not require intact phloem tissue. A 48-kDa MBP kinase is also activated by the 18-amino acid polypeptide systemin, a potent wound signal for the synthesis of systemic wound response proteins (swrps). The kinase activation by systemin is strongly inhibited by a systemin analog having a Thr-17 → Ala-17 substitution, which is a powerful antagonist of systemin activation of swrp genes. A 48-kDa MBP kinase activity also increases in response to polygalacturonic acid and chitosan but not in response to jasmonic acid or phytodienoic acid. In def1, a mutant tomato line having a defective octadecanoid pathway, the 48-kDa MBP kinase is activated by wounding and systemin as in the wild-type plants. This indicates that MBP kinase functions between the perception of primary signals and the DEF1 gene product. In response to wounding, the MBP kinase is phosphorylated on phosphotyrosine residues, indicating a relationship to the mitogen-activated protein kinase family of protein kinases.

Repeated, but not acute, stress suppresses inflammatory plasma extravasation

Clinical findings suggest that inflammatory disease symptoms are aggravated by ongoing, repeated stress, but not by acute stress. We hypothesized that, compared with single acute stressors, chronic repeated stress may engage different physiological mechanisms that exert qualitatively different effects on the inflammatory response. Because inhibition of plasma extravasation, a critical component of the inflammatory response, has been associated with increased disease severity in experimental arthritis, we tested for a potential repeated stress-induced inhibition of plasma extravasation. Repeated, but not single, exposures to restraint stress produced a profound inhibition of bradykinin-induced synovial plasma extravasation in the rat. Experiments examining the mechanism of inhibition showed that the effect of repeated stress was blocked by adrenalectomy, but not by adrenal medullae denervation, suggesting that the adrenal cortex mediates this effect. Consistent with known effects of stress and with mediation by the adrenal cortex, restraint stress evoked repeated transient elevations of plasma corticosterone levels. This elevated corticosterone was necessary and sufficient to produce inhibition of plasma extravasation because the stress-induced inhibition was blocked by preventing corticosterone synthesis and, conversely, induction of repeated transient elevations in plasma corticosterone levels mimicked the effects of repeated stress. These data suggest that repetition of a mild stressor can induce changes in the physiological state of the animal that enable a previously innocuous stressor to inhibit the inflammatory response. These findings provide a potential explanation for the clinical association between repeated stress and aggravation of inflammatory disease symptoms and provide a model for study of the biological mechanisms underlying the stress-induced aggravation of chronic inflammatory diseases.

Inhibition of interferon γ induced interleukin 12 production: A potential mechanism for the anti-inflammatory activities of tumor necrosis factor

Inflammation is associated with production of cytokines and chemokines that recruit and activate inflammatory cells. Interleukin (IL) 12 produced by macrophages in response to various stimuli is a potent inducer of interferon (IFN) γ production. IFN-γ, in turn, markedly enhances IL-12 production. Although the immune response is typically self-limiting, the mechanisms involved are unclear. We demonstrate that IFN-γ inhibits production of chemokines (macrophage inflammatory proteins MIP-1α and MIP-1β). Furthermore, pre-exposure to tumor necrosis factor (TNF) inhibited IFN-γ priming for production of high levels of IL-12 by macrophages in vitro. Inhibition of IL-12 by TNF can be mediated by both IL-10-dependent and IL-10-independent mechanisms. To determine whether TNF inhibition of IFN-γ-induced IL-12 production contributed to the resolution of an inflammatory response in vivo, the response of TNF+/+ and TNF−/− mice injected with Corynebacterium parvum were compared. TNF−/− mice developed a delayed, but vigorous, inflammatory response leading to death, whereas TNF+/+ mice exhibited a prompt response that resolved. Serum IL-12 levels were elevated 3-fold in C. parvum-treated TNF−/− mice compared with TNF+/+ mice. Treatment with a neutralizing anti-IL-12 antibody led to resolution of the response to C. parvum in TNF−/− mice. We conclude that the role of TNF in limiting the extent and duration of inflammatory responses in vivo involves its capacity to regulate macrophage IL-12 production. IFN-γ inhibition of chemokine production and inhibition of IFN-γ-induced IL-12 production by TNF provide potential mechanisms by which these cytokines can exert anti-inflammatory/repair function(s).