A comparison between amylase levels from peritonsillar, dental and neck abscesses.

OBJECTIVES: Pus of peritonsillar abscess (PTA) contains very high amylase levels in some patients. The objective of this study was to further test this finding and to check whether high amylase levels in peritonsillar abscess originate from contamination by saliva during aspiration. STUDY DESIGN: Prospective study. SETTING: Tertiary care university hospital. PARTICIPANTS: The study includes 64 patients with PTA, 8 patients with a neck abscess and 12 patients with a dental abscess. MAIN OUTCOME MEASURE: Amylase levels of pus and serum were compared between the groups. Clinical data regarding hospitalisation length, recurrence rate and previous antibiotic treatment were also collected. RESULTS: Mean amylase levels in the pus of the PTA group were 3045 U/L (median 59 U/L), 13 U/L in the neck abscess group (P = 0.001) and 22 U/L in the dental abscess group (P = 0.001). Mean serum amylase was higher in the PTA group; PTA - 50 U/L, neck abscess - 37 U/L (P = 0.002) and dental abscess - 26 U/L (P < 0.002). All of the patients with amylase levels above 65 U/L had a first episode of PTA. In contrast, 40% of patients with amylase lower than 65 U/L had recurrent PTA (P = 0.003). CONCLUSION: A clear association is seen between minor salivary glands and peritonsillar abscess. The high amylase level in peritonsillar pus is not from contamination with saliva.

Streptococcus anginosus l-cysteine desulfhydrase gene expression is associated with abscess formation in BALB/c mice.

Streptococcus anginosus, an anginosus group bacterium, is frequently isolated from odontogenic abscesses, and is the oral bacterium that is primarily responsible for producing hydrogen sulfide from l-cysteine through the action of its l-cysteine desulfhydrase (ßC-S lyase) enzyme. However, the relationship between its production of hydrogen sulfide and abscess formation has not been investigated. To elucidate the etiological role of hydrogen sulfide in abscess formation, we initially measured, using specific primers, expression of the lcd gene, which encodes ßC-S lyase, in the pus of abscesses that formed in BALB/c mice following subcutaneous injection of S. anginosus into the dorsa. Expression of lcd was >15-fold higher when l-cysteine was present than when it was absent. A mouse virulence assay revealed that the mean diameter of abscesses caused by S. anginosus FW73 plus l-cysteine was greater than that of abscesses caused by S. anginosus FW73 in the absence of l-cysteine. These findings demonstrate that the lcd gene of S. anginosus is upregulated in mouse abscesses and that hydrogen sulfide, the product of a reaction catalyzed by ßC-S lyase, plays an etiological role in odontogenic abscess formation.

Pyk2 is required for neutrophil degranulation and host defense responses to bacterial infection.

The appropriate regulation of neutrophil activation is critical for maintaining host defense and limiting inflammation. Polymorphonuclear neutrophils (PMNs) express a number of cytoplasmic tyrosine kinases that regulate signaling pathways leading to activation. One of the most highly expressed, but least studied, kinases in PMNs is proline rich kinase 2 (Pyk2). By analogy to the related focal adhesion kinase, Pyk2 has been implicated in regulating PMN adhesion and migration; however, its physiologic function has yet to be described. Using pyk2(-/-) mice, we found that this kinase was required for integrin-mediated degranulation responses, but was not involved in adhesion-induced cell spreading or activation of superoxide production. Pyk2-deficient PMNs also manifested reduced migration on fibrinogen-coated surfaces. The absence of Pyk2 resulted in a severe reduction in paxillin and Vav phosphorylation following integrin ligation, which likely accounts for the poor degranulation and cell migration. Pyk2(-/-) mice were unable to efficiently clear infection with Staphylococcus aureus in a skin abscess model, owing in part to the poor release of granule contents at the site of infection. However, Pyk2-deficient PMNs responded normally to soluble agonists, demonstrating that this kinase functions mainly in the integrin pathway. These data demonstrate the unrealized physiologic role of this kinase in regulating the adhesion-mediated release of PMN granule contents.

Contribution of coagulases towards Staphylococcus aureus disease and protective immunity.

The bacterial pathogen Staphylococcus aureus seeds abscesses in host tissues to replicate at the center of these lesions, protected from host immune cells via a pseudocapsule. Using histochemical staining, we identified prothrombin and fibrin within abscesses and pseudocapsules. S. aureus secretes two clotting factors, coagulase (Coa) and von Willebrand factor binding protein (vWbp). We report here that Coa and vWbp together are required for the formation of abscesses. Coa and vWbp promote the non-proteolytic activation of prothrombin and cleavage of fibrinogen, reactions that are inhibited with specific antibody against each of these molecules. Coa and vWbp specific antibodies confer protection against abscess formation and S. aureus lethal bacteremia, suggesting that coagulases function as protective antigens for a staphylococcal vaccine.

Contribution of the prostaglandin E2/E-prostanoid 2 receptor signaling pathway in abscess formation in rat zymosan-induced pleurisy.

Abscess formation is a classic host response to infection by many pathogenic microorganisms. Here, we studied the role of prostaglandins (PGs) and their signal transduction in abscess formation. Zymosan was injected into the pleural cavity of rats. Expression of enzymes involved in PG synthesis, their receptors, and cytokines in exudate leukocytes and abscesses were analyzed by polymerase chain reaction, Western blotting, and immunohistochemistry. Treatment with ketorolac, a cyclooxygenase (COX)-1 inhibitor, or N-[2-cyclohexyloxy-4-nitrophenyl] methanesulfonamide (NS-398), a COX-2 inhibitor, reduced the size of abscesses and the number of cells recovered from the abscess. COX-2 was detected in leukocytes of the exudate and a marginal area of abscesses. Among detected terminal PG synthases, the major one was cytosolic PGE synthase. Membrane-bound PGE synthase (mPGES)-1 was detected in cells that were similar to the COX-2-expressing cells in morphology and localization. A high level of the E-prostanoid (EP)(2) receptor and a low level of the EP(4) receptor were detected. The expression pattern of the EP(2) receptor paralleled that of COX-2 and mPGES-1. 11,15-O-Dimethyl PGE(2) (ONO-AE1-259), an EP(2) receptor agonist, and rolipram, a phosphodiesterase type-4 inhibitor, reversed the effects of COX inhibitors on abscess formation. In contrast, 16-(3-methoxymethyl) phenyl-omega-tetranor-3,7-dithia PGE(1) (ONO-AE1-329), an EP(4) receptor agonist, did not reverse the effects of NS-398. Moreover, NS-398 reduced the mRNA levels in exudate leukocytes of some proinflammatory and fibrogenic cytokines, which was reversed by ONO-AE1-259. These results suggest that PGE(2) generated via COX-1 and COX-2 may interact with the EP(2) receptor and may up-regulate in cAMP-dependent fashion the production of cytokines that promote abscess formation.

Deficiency of iNOS contributes to Porphyromonas gingivalis-induced tissue damage.

Periodontitis is a chronic inflammatory disease that results in extensive soft and hard tissue destruction of the periodontium. Porphyromonas gingivalis possesses an array of virulence factors and has been shown to induce expression of inducible nitric oxide synthase (iNOS) in inflammatory cells. The aim of this study was to investigate the effect of eliminating iNOS in a murine model of P. gingivalis infection. This was achieved by utilizing a P. gingivalis-induced skin abscess model, and an alveolar bone loss model employing an oral infection of P. gingivalis in iNOS knockout mice. The results indicated that iNOS knockout mice exhibit more extensive soft tissue damage and alveolar bone loss in response to P. gingivalis infection compared to wild-type mice. The local immune response to P. gingivalis in iNOS knockout mice was characterized by increased numbers of polymorphonuclear monocytes, while the systemic immune response was characterized by high levels of interleukin-12. The iNOS is required for an appropriate response to P. gingivalis infection.

Induction of type 3 deiodinase activity in inflammatory cells of mice with chronic local inflammation.

During illness, changes in thyroid hormone metabolism occur, so-called nonthyroidal illness (NTI). NTI has been characterized by a fall of serum T(3) due to decreased extrathyroidal conversion of T(4) into T(3) by liver type 1 deiodinase (D1), without an increase in serum TSH. Type 3 deiodinase (D3) was thought not to play an important role during NTI, but recently it has been shown that D3 activity is up-regulated in liver and skeletal muscle of critically ill patients related to hypoxia. We studied D3 gene expression and activity in liver and muscle/subcutis of mice during illness, which was induced by two different stimuli: bacterial endotoxin (lipopolysaccharide) administration, resulting in an acute systemic response, and a turpentine injection in each hindlimb, resulting in a local sc abscess. Lipopolysaccharide induced a rapid decrease in liver D1 and D3 activity but not skeletal muscle of hindlimb. In contrast, local inflammation induced by turpentine did not decrease liver D1 and D3 activity but increased markedly D3 activity in the muscle/subcutis sample containing the abscess, associated with strongly increased IL-1beta and IL-6 mRNA expression. Inflammatory cells, surrounding the abscess showed D3 and T(3)-transporter monocarboxylate transporter-8 immunoreactivity, whereas muscle cells did not show any immunoreactivity. In conclusion, local inflammation strongly induces D3 activity in inflammatory cells, especially in invading polymorphonuclear granulocytes, suggesting enhanced local degradation of T(3).

Increased intracellular activity of matrix metalloproteinases in neutrophils may be associated with delayed healing of infection without neutropenia in myelodysplastic syndromes.

To investigate the influence of the intracellular activity of type II and type IV collagenases [matrix metalloproteinases (MMP)-2, MMP-8, and MMP-9] in neutrophils from patients with myelodysplastic syndromes (MDS), we tried to measure intracellular activity using flow cytometric techniques. We also studied the clinical features of patients showing high activity. The intracellular collagenase activity, expressed as a ratio to the standardized fluorescence intensity, in patients with MDS was significantly higher than normal volunteers (19.5+/-14.8 vs 13.3+/-6.8, p=0.024). The difference among subcategories of MDS according to the French-American-British (FAB) and WHO classifications was not significant. No significant influence of three variables of the International Prognostic Scoring System (IPSS) was seen on activity. Of 8 patients with activity of more than 26.9 (mean+2 standard deviations of normal controls), 5 experienced an episode of delayed healing of infection without neutropenia, while 1 of 43 patients with activity of less than 26.9 experienced such an episode (p=0.0002). The average collagenase activity of six patients with delayed healing of infection without neutropenia (44.7+/-28.9) was significantly higher than that of other MDS patients (16.0+/-7.1, p=0.005) (Fig. 4). It was also significantly higher than the activity of the control group (13.3+/-6.8, p=0.011). Our report suggests that increased collagenase activity in neutrophils may delay healing of infection. In addition, we suggest that increased collagenase activity may be an independent prognostic factor for the susceptibility to severe infection in MDS.

IFN-gamma is effective in reducing infections in the mouse model of chronic granulomatous disease (CGD).

Chronic granulomatous disease (CGD) is a genetic disorder characterized by recurrent bacterial and fungal infections and tissue granuloma formation. CGD phagocytes are unable to generate superoxide because of mutations in any of four proteins of the phagocyte NADPH oxidase. Prophylactic recombinant human interferon-gamma (IFN-gamma) has been shown to reduce the frequency and severity of infections in CGD patients, but its mechanism(s) remains undefined, and its benefit has been questioned. We investigated the prophylactic effect of IFN-gamma in the mouse model of the major autosomal recessive (p47(phox)) form of CGD. In a prospective, randomized, placebo-controlled study, we compared IFN-gamma, 20,000 U administered subcutaneously (s.c.) three times weekly, to placebo in 118 p47(phox-/-) mice. By 6 weeks of study, there were 3 infections in the IFN-gamma group compared with 13 infections in the placebo group (77% reduction in infections, p<0.01). By 18 months of study, there were 7 infections in the IFN-gamma group compared with 18 infections in the placebo group (39% reduction in infections, p<0.01). Two animals receiving IFN-gamma had seizures after 7 months in the study. No other toxicities were observed. Peripheral blood phagocytes from IFN-gamma treated p47(phox-/-) mice produced no superoxide, excluding restoration of the oxidative burst as a mechanism for the IFN-gamma effect. There were no differences in either peritoneal macrophage nitrate production or thioglycollate-induced peritoneal exudate between treatment groups. This animal model demonstrates a prophylactic benefit of IFN-gamma similar to that seen in humans and provides an opportunity to investigate the mechanism(s) of action for IFN-gamma in CGD.

Suppression of polymorphonuclear leucocyte chemotaxis by Pseudomonas aeruginosa elastase in vitro: a study of the mechanisms and the correlation with ring abscess in pseudomonal keratitis.

Bacteria, or the culture supernatants of an elastase non-producing strain of Pseudomonas aeruginosa, elicited a chemotactic response from polymorphonuclear leucocytes (PMN) in vitro. The chemoattractive capacity was diminished under the presence of Boc-Phe-Leu-Phe-Leu-Phe, a receptor antagonist of N-formyl-Met-Leu-Phe (fMLP) which is a bacterial chemotactic peptide to PMN. This indicated that the chemoattractant derived from Pseudomonas aeruginosa was a fMLP-like molecule(s). In contrast, culture supernatants of an elastase producing strain of Pseudomonas aeruginosa produced negligible chemotactic response from PMN. Indeed, an inhibitory effect of the culture supernatants or of purified Pseudomonas aeruginosa elastase (PAE) on PMN chemotaxis was observed when fMLP was used as a chemoattractant. Another fMLP-induced function of PMN, respiratory burst activation, was also diminished by pretreatment of PMN with PAE. PAE hydrolysed fMLP at the Met-Leu bond and diminished the chemoattractant capacity. In addition, a receptor analysis with fML-3H-P demonstrated a decrease in numbers of fMLP receptors on PMN without changing the dissociation constant values after the treatment of the cells with PAE. In the primary structure of the fMLP receptor previously reported, a preferential amino acid sequence for cleavage by PAE was identified in what was believed to be an extracellular portion of the receptor molecule. These results suggested that PAE could diminish PMN infiltration in response to Pseudomonas aeruginosa in vivo by cleavage of the fMLP-like pseudomonal chemotactic ligand and the receptors on PMN.

Detection of intraabdominal abscess by serum lysozyme estimation.

BACKGROUND: The failure of new, innovative, and often expensive tests to show the presence of intraabdominal infection caused us to reexamine the value of an inexpensive and almost forgotten one. Lysozyme is a bacteriolytic enzyme located within the lysosomes of phagocytic cells including leukocytes. METHODS: We measured serum concentrations of lysozyme by a standard turbidimetric method in both a murine model of intraabdominal infection and in trauma patients with intraabdominal abscesses or other acquired infections. RESULTS: In mice with intraabdominal abscess secondary to cecal ligation and puncture (n = 35) serum lysozyme activity increased compared with sham-operated controls (n = 20; p < 0.001). In trauma patients with intraabdominal abscess after injury and surgery (n = 19), there was also an increase in serum lysozyme activity compared with controls (n = 15; p < 0.001) or with patients with lung infection (n = 21; p < 0.001). The increase of serum lysozyme activity occurred before intraabdominal sepsis was clinically apparent. CONCLUSIONS: In this study, serum lysozyme concentration has a high specificity related to the presence of an intraabdominal abscess and is an indirect measure of the sequestration of leukocytes to the site of an established or developing collection of intraabdominal pus. The estimation of serum lysozyme may be an aid to differentiate critically ill patients with a potential occult intraabdominal abscess.

Evaluation of bacterial interference and beta-lactamase production in management of experimental infection with group A beta-hemolytic streptococci.

The in vivo effects of penicillin and cefprozil therapy on the interaction between organisms commonly recovered from inflamed tonsils were studied by using a subcutaneous abscess model in mice. These organisms were group A beta-hemolytic streptococci (GABHS), Streptococcus salivarius (which is capable of interfering with GABHS), and Staphylococcus aureus. In mice infected with GABHS and S. salivarius alone or in combination, penicillin eliminated both organisms and cefprozil eliminated GABHS and S. aureus but not S. salivarius. Penicillin did not, however, reduce the number of GABHS or S. salivarius in the presence of S. aureus. The present study demonstrated the ability of beta-lactamase-producing S. aureus to protect GABHS from penicillin. However, no such protection was present following the administration of cefprozil. Furthermore, the preservation of S. salivarius that interferes with GABHS growth may provide protection from reinfection with GABHS. This study supports and provides an explanation for the increased efficacies of cephalosporins administered orally over that of penicillin when treating patients with acute GABHS pharyngitis or tonsillitis.

Efficacy of ampicillin-sulbactam versus that of cefoxitin for treatment of Escherichia coli infections in a rat intra-abdominal abscess model.

We examined the efficacy of ampicillin-sulbactam (2:1) and cefoxitin in the treatment of infections caused by Escherichia coli strains exhibiting increasing levels of beta-lactamase-mediated resistance to ampicillin-sulbactam in the rat intra-abdominal abscess model. Cefoxitin was superior to ampicillin-sulbactam in the treatment of infections caused by all strains. Treatment with ampicillin-sulbactam resulted in a statistically significant decrease in CFU per gram of abscess in comparison with treatment with ampicillin alone for both the moderately resistant and the resistant strains, with an inverse correlation between the MIC and the absolute decrease in CFU per gram of abscess.

Impaired neutrophil killing in a patient with defective degranulation of myeloperoxidase.

A case of recurrent, superficial abscesses in an 18 year old girl, is described. Staphylococcus aureus was the pathogen most often implicated and on several occasions the abscesses required surgical drainage. Defects in humoral immunity, neutrophil chemotaxis or opsonophagocytosis were not observed. However, her neutrophil's ability to kill ingested S. aureus in vitro was impaired. This was associated with impaired luminol-dependent chemiluminescence in response to stimulation by either latex beads, or the chemotactic peptide FMLP plus cytochalasin B. Oxygen uptake and superoxide anion production were normal but release of myeloperoxidase by this patient's neutrophils occurred more slowly and to a lower extent than in control cells. These data suggest that the recurrent infections and diminished in vitro neutrophil bactericidal activity observed in this patient are associated with impaired degranulation of myeloperoxidase.

Effect of sepsis on activity of pyruvate dehydrogenase complex in skeletal muscle and liver.

The effect of chronic sepsis on the concentration of active pyruvate dehydrogenase complex has been investigated in liver and skeletal muscle of normal, sterile inflammatory, and chronic septic (small and large abscess) animals. Hyperdynamic sepsis was induced by the intraperitoneal introduction of a rat fecal-agar pellet of known size and bacterial composition (Escherichia coli + Bacteroides fragilis). Total pyruvate dehydrogenase complex activity was not altered in either liver or skeletal muscle in any of the conditions studied. In hepatic tissue, sterile inflammation increased the proportion of active complex 2.5-fold compared with control. The same increase in the concentration of active complex was observed in animals with a small abscess. When the abscess size was increased (large abscess), the concentration of active complex was decreased relative to sterile inflammatory or small abscess septic animals. In contrast to liver, sterile inflammation did not alter the proportion of active complex in skeletal muscle. Sepsis (either small or large septic abscess) resulted in threefold decrease in the concentration of active complex relative to control or sterile inflammatory animals. Changes in the concentration of active complex did not appear to be dependent on the ATP/ADP concentration ratio or tissue pyruvate levels but were consistent with changes in the acetyl-coenzyme A-to-coenzyme A concentration ratio. The mechanism responsible for altered concentration of active complex may be mediated through changes in the activity of the pyruvate dehydrogenase kinase, secondary to alterations in the effector concentration ratios.

Leukocyte elastase alpha 1-proteinase inhibitor complexes may diagnose pancreatic abscesses early.

Complexes of alpha 1-proteinase inhibitor and leukocyte elastase could be demonstrated by crossed immunoelectrophoresis of the peritoneal fluid from four patients who developed a pancreatic abscess during an attack of pancreatitis. No such complexes were seen in 69 patients with acute pancreatitis without an abscess. The complexes were demonstrable 2-3 days before the abscess was clinically evident. They may thus be diagnostically and therapeutically important. The appearance of these complexes denotes the liberation of large amounts of leukocyte elastase. This may help explain the pathophysiology and high mortality of the pancreatic abscess, since leukocyte elastase is known to cause degradation of all components of connective tissue and also degradation and activation of many components within the different cascade systems.

The localization of metallic radionuclides in abscesses in rats.

The accumulation of ten metals in turpentine-induced abscesses of different ages has been studied in rats. The maximum metal concentration ratio between the abscesses and the non-inflamed contralateral muscle was greater than 1 for all the metals studied, the lowest being 2.5 for Zn, and the highest 36 for Th. The ratio remains greater than 1 for more than 28 days for all metals except Ga. There is a marked increase of beta-glucuronidase activity in the inflamed tissue which correlates well with the metal concentration. In contrast the acid phosphatase activity was reduced for the first few days after turpentine injection and never rose above the normal muscle activity. The metals showing the highest uptake in the inflamed lesion appear to be those which are transported in the blood as a transferrin complex.