Theodoros Adamis

Immunomodulatory clarithromycin treatment of experimental sepsis and acute pyelonephritis caused by multidrug-resistant Pseudomonas aeruginosa.

ABSTRACT Clarithromycin was administered intravenously to 55 rabbits to evaluate its effect on experimental sepsis caused by multidrug-resistant Pseudomonas aeruginosa. Acute pyelonephritis was induced after ligation of the right ureter and injection of 108 CFU of the test isolate per kg of body weight into the renal pelvis. The animals were divided into six groups: group A, controls; group B, rabbits that received one intravenous dose of 80 mg of clarithromycin per kg concomitantly with bacterial challenge; group C, rabbits that received two doses of clarithromycin, the second one of which was given 2 h after the first one; group D, rabbits that received 15 mg of amikacin per kg; group E, rabbits that received one dose of clarithromycin and amikacin; and group F, rabbits that received two doses of clarithromycin and amikacin. Serum endotoxin levels were estimated by the QCL-1000 Limulus amoebocyte lysate assay, tumor necrosis factor alpha (TNF-α) levels were measured by a bioassay, and malondialdehyde (MDA) levels were measured by the thiobarbiturate assay. Viable bacterial counts in various tissue samples were also assessed. The mean survival times of the animals in groups A, B, C, D, E, and F were 4.50, 7.69, 4.07, 4.55, 11.55, and 11.60 days, respectively (P = 0.033 for group D versus group F, P = 0.006 for group D versus group E, P = not significant for group B versus group E, P = 0.042 for group C versus group F). Serum endotoxin levels were similar between groups at all sampling times; TNF-α and MDA levels in groups B, C, E, and F decreased significantly over follow-up. The numbers of viable bacterial cells in the infected kidney were similar among the groups; those in the liver, spleen, lungs, and mesenteral lymph nodes were significantly decreased in groups B, E, and F compared to those in groups A and D. It is concluded that a prolongation of survival in animals with experimental sepsis caused by multidrug-resistant P. aeruginosa was achieved after coadministration of clarithromycin and amikacin and that the increased survival was probably attributable to the immunomodulatory properties of clarithromycin.

Efficacy and pharmacodynamics of linezolid, alone and in combination with rifampicin, in an experimental model of methicillin-resistant Staphylococcus aureus endocarditis

OBJECTIVES To evaluate the efficacy of oral linezolid, with or without rifampicin, on valve vegetations and secondary foci of infection compared with vancomycin, in the absence or presence of rifampicin, in experimental endocarditis caused by methicillin-resistant Staphylococcus aureus. METHODS Treatment groups were controls (n = 16), linezolid (n = 15), vancomycin (n = 15), linezolid and rifampicin (n = 15), vancomycin and rifampicin (n = 13), linezolid relapse (n = 11) and vancomycin relapse (n = 9). Therapy lasted 5 days in all groups, with survival of animals in the linezolid relapse and vancomycin relapse groups being recorded for an additional 5 days. Blood was drawn to determine the linezolid concentration, and valve vegetations, and kidney, liver, lung and spleen segments were collected for culture. RESULTS Survival in each individual group was higher than that in the control group; bacterial load in valve vegetations was reduced by all treatment regimens, with linezolid exhibiting bactericidal effects. Bactericidal activity of linezolid was noted in all secondary foci of infection except the lung, where only the combination of rifampicin with linezolid was bactericidal. CONCLUSIONS Orally administered linezolid is effective in limiting bacterial growth in the secondary foci of endocarditis. Co-administration of rifampicin favoured the suppression of bacterial growth in the lung.

Clarithromycin is an effective immunomodulator when administered late in experimental pyelonephritis by multidrug-resistant Pseudomonas aeruginosa

BackgroundTo apply clarithromycin as an immunomodulatory treatment in experimental urosepsis by multidrug-resistant Pseudomonas aeruginosa.MethodsAcute pyelonephritis was induced in 40 rabbits after inoculation of the test isolate in the renal pelvis. Therapy was administered upon signs of sepsis in four groups: A, controls; B, intravenous clarithromycin; C, amikacin; and D, both agents. Survival and vital signs were recorded; blood was sampled for culture and estimation of pro-inflammatory mediators; monocytes were isolated for determination of apoptotic rate and ex vivo TNFα secretion. Quantitative cultures and biopsies of organs were performed after death.ResultsIncreased rectal temperature and oxygen saturation were found in groups B and D compared to A and C. Mean survival of groups A, B, C and D was 2.65, 7.15, 4.25 and 8.70 days respectively. No differences were noted between groups concerning bacterial load in blood and tissues and serum endotoxins. Serum MDA and total caspase-3 activity of monocytes of group D decreased following treatment compared to other groups. Negative correlation was detected between cytoplasmic caspase-3 and ex vivo secretion of TNFα of blood monocytes of group A; similar correlation was not found for any other group. Pathology scores of liver and lung of group B were lower than group A.ConclusionClarithromycin administered late in experimental urosepsis by multidrug-resistant P. aeruginosa prolonged survival and ameliorated clinical findings. Its effect is probably attributed to immunomodulatory intervention on blood monocytes.

Clarithromycin: immunomodulatory therapy of experimental sepsis and acute pyelonephritis by Escherichia coli.

The potency of clarithromycin as immunomodulator was assessed in an experimental model of sepsis based on acute pyelonephritis by susceptible Escherichia coli. 55 rabbits were utilized; 5 for preliminary pharmacokinetic study and 50 for treatment. The latter were divided into 5 groups of treatment, A: controls; B: clarithromycin pretreatment; C: amikacin pretreatment; D: clarithromycin treatment on presentation of pulmonary oedema; and E; amikacin treatment on presentation of pulmonary oedema. Survival was recorded; tumour necrosis factor-alpha (TNFα), and malondialdehyde (MDA) were estimated in serum; activities of caspase-3 in monocyte cytosolic extracts were studied; and bacterial counts made in various organs. Median survival of animals of groups A, B, C, D and E was 1.0, 21.0, 12.5, 2.0 and 5.0 d, respectively. TNFα and MDA and monocyte caspase-3 activity of group A increased over time; no increases were detected in groups B and C. Concentrations of MDA and activities of monocytic caspase-3 were decreased after administration of clarithromycin in group D, an effect not occurring in group E. Bacterial load was decreased in renal tissue of group D compared to group A. It is concluded that intravenous clarithromycin might constitute a promising immunomodulator in sepsis even in the advent of pulmonary oedema.

n-6 Polyunsaturated Fatty Acids Enhance the Activities of Ceftazidime and Amikacin in Experimental Sepsis Caused by Multidrug-Resistant Pseudomonas aeruginosa

ABSTRACT Recent in vitro and ex vivo studies disclosed an enhancement of the activity of antimicrobials on multidrug-resistant Pseudomonas aeruginosa by n-6 polyunsaturated fatty acids (PUFAS); therefore their effect was evaluated in experimental sepsis in 60 rabbits. Solutions of gamma-linolenic acid (GLA) and arachidonic acid (AA) were administered intravenously with ceftazidime and amikacin in rabbits with sepsis caused by one multidrug-resistant isolate. Therapy was started after bacterial challenge in five groups comprising 12 animals in each group: A, normal saline; B, antimicrobials; C, 99% ethanol and antimicrobials; D, GLA and antimicrobials; and E, AA and antimicrobials. Blood was sampled for the estimation of levels of endotoxins in serum (lipopolysaccharide), leukocytes, tumor necrosis factor alpha (TNF-α) and antimicrobials. Animals were sacrificed 210 min after bacterial challenge for tissue cultures. All animals had considerable endotoxemia and evolved leukopenia. The number of viable cells in blood, lung, and mesenteric lymph nodes was significantly reduced in groups D and E compared to that in other groups. Levels of antimicrobials in serum were inadequate to achieve bacterial killing due to the level of resistance. n-6 PUFAs did not influence TNF-α. It is concluded that intravenous coadministration of n-6 PUFAs and antimicrobials enhanced antimicrobial bacterial killing in experimental sepsis caused by multidrug-resistant P. aeruginosa.

Early apoptosis of blood monocytes is a determinant of survival in experimental sepsis by multi-drug-resistant Pseudomonas aeruginosa.

Apoptosis of blood monocytes was studied in experimental sepsis by multi‐drug‐resistant Pseudomonas aeruginosa. Thirty‐six rabbits were used, divided into the following groups: A (n = 6), sham; B (n = 6), administered anaesthetics; and C (n = 24), acute pyelonephritis induced after inoculation of the test isolate in the renal pelvis. Blood was sampled at standard time intervals for estimation of tumour necrosis factor (TNF)‐α and isolation of monocytes. Half the monocytes were incubated and the other half was lysed for estimation of the cytoplasmic activity of caspase‐3 by a kinetic chromogenic assay. No animal in groups A and B died; those in group C were divided into two subgroups, CI (n = 8) with present activity of caspase‐3 of blood monocytes at 3·5 h and CII (n = 16) with absent activity. Their median survival was 2·0 and 3·5 days, respectively (P = 0·0089). Ex vivo secretion of TNF‐α from monocytes was higher by monocytes of subgroup CII than subgroup CI at 3·5 h (P = 0·039) and of group A than CII at 48 h (P = 0·010). Median change of caspase‐3 activity between 3·5 and 24 h of sampling was 56·1 and −5·8 pmol/min per 104 cells for subgroups CI and CII (P = 0·040), respectively. Respective changes between 3·5 and 48 h were 28 981·0 and 0 pmol/min per 104 cells (P = 0·036). Early induction of apoptosis in blood monocytes is of prime importance for the survival of the septic host and might be connected to changes of monocyte potential for the secretion of TNF‐α.

The impact of multidrug resistance on the pathogenicity of Escherichia coli: an experimental study.

Based on the controversial findings of clinical studies regarding the influence of multidrug resistance on mortality, 10 susceptible and 10 multidrug-resistant (MDR) and extended-spectrum beta-lactamase-producing isolates of Escherichia coli were applied to stimulate monocytes isolated from healthy donors. Immune mediators were estimated in supernatants. Four susceptible isolates (Group A) and four MDR isolates (Group B) were used to initiate acute pyelonephritis in 48 rabbits following inoculation of the pathogen into the right renal pelvis. Survival was recorded and blood monocytes were isolated and incubated to estimate the ex vivo release of tumour necrosis factor-alpha (TNFalpha). Release of TNFalpha, interleukin (IL)-6 and IL-8 was higher after 2 h and 4 h of stimulation by MDR isolates compared with susceptible isolates. The opposite occurred for the release of IL-12. Death occurred in 22 rabbits in Group A (91.7%) compared with 12 in Group B (50.0%) (P=0.003). Monocytes isolated at 24 h from Group A rabbits released significantly higher TNFalpha than monocytes from Group B. Tissue bacterial load after animal death was significantly higher in the kidneys of Group A rabbits. It is concluded that susceptible and MDR E. coli stimulate monocytes resulting in a different pattern of release of pro-inflammatory cytokines, which is accompanied by prolonged survival following experimental sepsis by MDR isolates.

Immunomodulatory Effect of Three-Day Continuous Administration of Clarithromycin for Experimental Sepsis Due to Multidrug-Resistant Pseudomonas aeruginosa

Abstract Based on former animal studies showing the effect of clarithromycin in experimental sepsis by multidrug-resistant Pseudomonas aeruginosa following administration of single doses, the significance of its administration for three consecutive days was evaluated. Acute pyelonephritis was induced in 20 rabbits after inoculation of the test isolate in the renal pelvis. Therapy was administered upon signs of sepsis in group B; A served as control. Survival was recorded; monocytes were isolated for determination of ex vivo TNFα secretion. Quantitative cultures of organs were performed after death. Mean survival of groups A and B was 2.65 and 7.95 days respectively. At 24 hours, serum malondialdehyde of group B, which is an index of the oxidant status in serum, was lower than A. ex vivo release of TNFα by the isolated monocytes of group B was lower than A at 3.5 and 48 hours. Tissue bacterial load was similar in two groups after animal death. It is concluded that clarithromycin possessed considerable immunomodulatory effects restraining release of TNFα from blood monocytes.

Early Cutaneous Alterations in Experimental Sepsis by Pseudomonas aeruginosa

Background: To evaluate whether histopathologic findings of skin in sepsis by Pseudomonas aeruginosa correlate with the clinical course. Methods: Histological alterations after bacterial challenge by one susceptible (A) and two multidrug-resistant isolates (B and C) of P. aeruginosa were studied in 18 rabbits. Sepsis was induced by the intravenous infusion of 1 × 108 CFU by a catheter in the right jugular vein; blood was sampled for the estimation of tumor necrosis factor α (TNF-α) and malondialdehyde (MDA). Skin biopsies were collected along with a subcutaneous fat specimen for culture. Results: The mean survival was 0.85, 1.75 and 11.00 days after challenge by isolates A, B and C, respectively. The main histologic findings of skin were: inflammation and swelling of the dermis; thickening of the endothelium and infiltration of vessel wall and lumen by polymorphonuclear leukocytes; extravasation of red blood cells, and necrobiotic changes of the hair follicles. Serum TNF-α was elevated in animals challenged by isolate A compared to challenge by isolates B and C. Concentrations of MDA were similar for all isolates. Mean log10 of viable cells isolated from subcutaneous fat were 5.74, 2.74 and 1.40 after challenge by isolates A, B and C, respectively. Conclusions: Prolongation of survival was accompanied by lower serum TNF-α, decreased viable cells from subcutaneous fat and intensified inflammatory response in the dermis and subcutaneous tissue. These findings might be of importance for immunomodulatory intervention.