Pre-treatment with probiotics prolongs survival after experimental infection by multidrug-resistant Pseudomonas aeruginosa in rodents: an effect on sepsis-induced immunosuppression.

Based on several randomised clinical studies indicating benefit from oral probiotic intake for the prevention of hospital-acquired infections in critically ill patients, this study aimed to explain the mechanism of action of probiotics for the prevention of lethal experimental infection by multidrug-resistant (MDR) Pseudomonas aeruginosa. Experiments using an Escherichia coli strain susceptible to all antimicrobials were also conducted. C57BL/6 mice were pre-treated intraperitoneally with sterile water for injection or Lactobacillus plantarum. Survival was recorded and mice were sacrificed for measurement of apoptosis and tissue bacterial overgrowth and for isolation and culture of splenocytes for cytokine production. Experiments were repeated after pre-treatment with a commercial preparation of four probiotics (L. plantarum, Lactobacillus acidophilus, Saccharomyces boulardii and Bifidobacterium lactis; LactoLevure(®)). Peripheral blood mononuclear cells (PBMCs) of healthy volunteers were stimulated by heat-killed P. aeruginosa following pre-treatment with medium or probiotics. Pre-treatment with L. plantarum significantly prolonged survival after challenge by either MDR P. aeruginosa (66.7% vs. 31.3%; P=0.026) or E. coli (56.0% vs. 12.0%, P=0.003). Survival benefit was even more pronounced when mice were pre-treated with LactoLevure(®). Tissue bacterial outgrowth and apoptosis of white blood cells and splenocytes were not altered. TNFα and IL-10 production by splenocytes of mice pre-treated with probiotic was increased and IFNγ production was decreased. Pre-treatment with LactoLevure(®) restored production of IL-17. Stimulation of human PBMCs after probiotic pre-treatment was accompanied by reduced gene expression of SOCS3. The results suggest that the protective effect of probiotics is mediated through prevention of sepsis-induced immunosuppression.

Immune responses in relation to the type and time of thermal injury: an experimental study.

BACKGROUND: Thermal injuries are followed by a complex immune response, but the relationship between the severity of burn injury and the time exposure to the thermal injury on the extent of the immune response is still not known. OBJECTIVE: This study focuses on characterising the effect of temperature and time exposure on the post-burn immune response. METHODS: We used 120 C57BL/6 male mice divided equally in 5 burn groups and one sham operated group (groups A-E and sham). Ten mice per group were sacrificed at 24 and 48 h after burn injury and whole blood was collected; specimens of liver, lung, spleen, kidney and bowel were excised. Apoptosis and TREM-1 expression on circulating blood cells were measured. Splenocytes were isolated and stimulated for cytokine production; the rate of apoptosis of splenocytes was also measured. RESULTS: Production of IL-17 from splenocytes of mice group D was enhanced. Considerable effects were shown on the apoptosis of circulating lymphocytes and of spleen cells. The apoptotic rates varied between groups and also evolved after 24 and 48 h. To examine the origin of this differential response, quantitative bacterial cultures of liver, lung and kidney were made but no differences were observed compared with sham-operated animals. LIMITATIONS: This study was based on an experimental murine model. CONCLUSION: There is a unique response for each type of injury depending on the temperature of the thermal source and the exposure time.

Immunomodulatory intervention with interferon-γ in Escherichia coli pyelonephritis.

PURPOSE: We investigated the efficacy of recombinant human interferon-γ in experimental pyelonephritis due to Escherichia coli. MATERIALS AND METHODS: Pyelonephritis was induced by intrapelvic inoculation of bacteria after ureteral ligation in 38 rabbits assigned to 1 of 3 groups, including group 1-16 controls, group 2-14 rabbits treated with intravenous recombinant human interferon-γ and group 3-8 rabbits treated with intravenous recombinant human interferon-γ plus amikacin. Bacterial counts, cytokines and malondialdehyde were measured in blood. Peripheral blood mononuclear cells were isolated to measure TNFα transcripts, cytokine stimulation and apoptosis. Survival was recorded, and the tissue bacterial load and myeloperoxidase activity were measured after sacrifice. RESULTS: The mortality rate in groups 1, 2 and 3 was 66.7%, 25% and 12.5%, respectively. The circulating bacterial count and tissue bacterial load were less in group 2 than in group 1. Circulating malondialdehyde negatively correlated with the bacterial load of the spleen. Although the number of TNFα transcripts in circulating peripheral blood mononuclear cells did not differ, peripheral blood mononuclear cells isolated from group 2 at 48 hours produced much greater concentrations of tumor necrosis factor-α after stimulation with Pam3Cys. In parallel, the apoptosis rate of circulating monocytes was increased in group 2 at 48 hours. Lung myeloperoxidase activity at 24 hours, serving as indirect evidence of neutrophil infiltration, was decreased in group 2. CONCLUSIONS: Recombinant human interferon-γ administration prolonged survival in rabbits with experimental E. coli urosepsis. Its action was probably related to increased bacterial phagocytosis after modulation of oxidant status and reversal of monocyte immunoparalysis.

Bacterial translocation induces proinflammatory responses and is associated with early death in experimental severe injury.

OBJECTIVE: An experimental model of severe injury with great lethality was studied to define the impact of bacterial translocation on survival and on inflammatory response. METHODS: Forty-one rabbits were divided into two groups: A, femur myotomy; and B, myotomy and fracture of the femoral bone. Vital signs and survival were recorded. Serum circulating endotoxins (lipopolysaccharides; LPS) were determined and tissue cultures were performed at necropsy. A subgroup of animals was sacrificed at 48 h post injury; LPS was determined in abdominal aorta and portal vein, apoptosis of spleen cells was assessed by flow cytometry, and ex vivo production of tumor necrosis factor alpha by splenocytes was measured. RESULTS: Tissue bacterial burden was increased in animals that died early (i.e., within 48 h after injury) versus rabbits that died later. Portal vein LPS at 48 h was increased in group B compared with group A, whereas circulating LPS did not differ. No difference in apoptosis of either lymphocytes or macrophages of the spleen was found in group B compared with group A. Following stimulation with LPS or phytohemagglutinin, tumor necrosis factor α production by splenocytes of group B was greater than that of group A. CONCLUSIONS: Bacterial translocation primes enhanced proinflammatory responses and it is associated with early death in severe trauma.

Intravenous itraconazole against experimental neutropenic Candida parapsilosis infection: efficacy after suppression of bacterial translocation.

A variety of studies indicate that itraconazole possesses greater intrinsic activity compared to the other azole derivatives against Candida parapsilosis. Efficacy has never been tested in an experimental setting. To this end, C. parapsilosis was used for challenge of 117 rats rendered neutropenic after a course of cyclophosphamide. Rats were assigned to receive intravenous treatment with saline (group A); itraconazole q12h (group B); fluconazole q12h (group C); single dose of ceftriaxone and saline (group D); single dose of ceftriaxone and itraconazole q12h (group E); and single dose of ceftriaxone and fluconazole q12h (group F). Survival was recorded, and yeast outgrowth of liver, spleen, lung, and kidney was measured after sacrifice at serial time intervals. Growth of the test isolate in tissues was significantly lower in group B than in groups A and C after 72 h. However, outgrowth of enterobacteria was found in tissues of groups A, B, and C, implying a phenomenon of bacterial translocation from the gut. When this phenomenon was suppressed with single doses of ceftriaxone, a striking survival benefit of itraconazole-treated animals was found (p = 0.022, group E vs. group F). The present results suggest than in deep infections by C. parapsilosis intravenously administered intraconazole may eradicate the offending agent and provide survival benefit when chemotherapy-induced bacterial translocation from the gut is suppressed. Further clinical evidence is required to support these findings.

Pre-treatment with low-dose endotoxin prolongs survival from experimental lethal endotoxic shock: Benefit for lethal peritonitis by Escherichia coli.

Although LPS tolerance is well-characterized, it remains unknown if it is achieved even with single doses of lipopolysaccharide (LPS) and if it offers protection against lethal bacterial infections. To this end, C57B6 mice were assigned to groups A (sham); B (saline i.p followed after 24h by i.p 30mg/kg LPS); and C (3mg/kg LPS i.p followed after 24h by i.p 30mg/kg LPS). Survival was monitored and animals were sacrificed early after lethal challenge for measurement of tumour necrosis factor-alpha (TNFα) in serum; isolation of splenocytes and cytokine stimulation; and flow-cytometry for apoptosis and TREM-1. Experiments were repeated with mice infected i.p by Escherichia coli after challenging with saline or LPS. Mortality of group B was 72.2% compared with 38.9% of group C (p: 0.020). Serum TNFα of group C was lower than group B. Expression of TREM-1 of group C on monocytes/neutrophils was greater than group B. Release of TNFα, of IFNγ and of IL-17 from splenocytes of group C was lower than group B and the opposite happened for IL-10 showing evidence of cellular reprogramming. In parallel, apoptosis of circulating lymphocytes and of splenocytes of group C was greater compared with group B. Pre-treatment of mice challenged by E. coli with low dose LPS led to 0% mortality compared with 90% of saline pre-treated mice; in these mice, splenocytes improved over-time their capacity for release of IFNγ. It is concluded that single low doses of LPS lead to early reprogramming of the innate immune response and prolong survival after lethal E. coli challenge.