Henrik Calum

Thermal injury induces impaired function in polymorphonuclear neutrophil granulocytes and reduced control of burn wound infection

Severe thermal injury induces immunosuppression, involving all parts of the immune system, especially when large fractions of the total body surface area are affected. An animal model was established to characterize the burn‐induced immunosuppression. In our novel mouse model a 6% third‐degree burn injury was induced in mice with a hot‐air blower. The third‐degree burn was confirmed histologically. The mice were allocated into five groups: control, shave, burn, infection and burn infection group. At 48 h, a decline in the concentration of peripheral blood leucocytes was observed in the group of mice with burn wound. The reduction was ascribed to the decline in concentration of polymorphonuclear neutrophil leucocytes and monocytes. When infecting the skin with Pseudomonas aeruginosa, a dissemination of bacteria was observed only in the burn wound group. Histological characterization of the skin showed a more polymorphonuclear neutrophil granulocytes (PMNs)‐dominated inflammation in the group of mice with infected burn wound compared with the with burn wound group. In contrast, a higher degree of inflammation was observed in the burn wound group compared with the group of mice with infected burn wound. Furthermore, the oxidative burst and the phagocytic capacity of the PMNs were reduced in the group of mice with burn wound. Using this novel mouse model of thermal injury a decline of peripheral leucocytes was observed, whereas the increased local inflammatory response at the site of infection showed reduced capacity to contain and eliminate the infection.

Pseudomonas aeruginosa biofilm aggravates skin inflammatory response in BALB/c mice in a novel chronic wound model

Chronic wounds are presumed to persist in the inflammatory state, preventing healing. Emerging evidence indicates a clinical impact of bacterial biofilms in soft tissues, including Pseudomonas aeruginosa (PA) biofilms. To further investigate this, we developed a chronic PA biofilm wound infection model in C3H/HeN and BALB/c mice. The chronic wound was established by an injection of seaweed alginate‐embedded P. aeruginosa PAO1 beneath a third‐degree thermal lesion providing full thickness skin necrosis, as in human chronic wounds. Cultures revealed growth of PA, and both alginate with or without PAO1 generated a polymorphonuclear‐dominated inflammation early after infection. However, both at days 4 and 7, there were a more acute polymorphonuclear‐dominated and higher degree of inflammation in the PAO1 containing group (p < 0.05). Furthermore, PNA‐FISH and supplemented DAPI staining showed bacteria organized in clusters, resembling biofilms, and inflammation located adjacent to the PA. The chronic wound infection showed a higher number of PAO1 in the BALB/c mice at day 4 after infection as compared to C3H/HeN mice (p < 0.006). In addition, a higher concentration of interleukin‐1beta in the chronic wounds of BALB/c mice was observed at day 7 (p < 0.02), despite a similar number of bacteria in the two mouse strains. The present study succeeded in establishing a chronic PA biofilm infection in mice. The results showed an aggravating impact of local inflammation induced by PA biofilms. In conclusion, our findings indicate that improved infection control of chronic wounds reduces the inflammatory response and may improve healing.

Novel experimental Pseudomonas aeruginosa lung infection model mimicking long-term host–pathogen interactions in cystic fibrosis

The dominant cause of premature death in patients suffering from cystic fibrosis (CF) is chronic lung infection with Pseudomonas aeruginosa. The chronic lung infection often lasts for decades with just one clone. However, as a result of inflammation, antibiotic treatment and different niches in the lungs, the clone undergoes significant genetic changes, resulting in diversifying geno‐ and phenotypes. Such an adaptation may generate different host responses. To experimentally reflect the year‐long chronic lung infection in CF, groups of BALB/c mice were infected with clonal isolates from different periods (1980, 1988, 1997, 1999 and 2003) of the chronic lung infection of one CF patient using the seaweed alginate embedment model. The results showed that the non‐mucoid clones reduced their virulence over time, resulting in faster clearing of the bacteria from the lungs, improved pathology and reduced pulmonary production of macrophage inflammatory protein‐2 (MIP‐2) and granulocyte colony‐stimulating factor (G‐CSF). In contrast, the mucoid clones were more virulent and virulence increased with time, resulting in impaired pulmonary clearing of the latest clone, severe inflammation and increased pulmonary MIP‐2 and G‐CSF production. In conclusion, adaptation of P. aeruginosa in CF is reflected by changed ability to establish lung infection and results in distinct host responses to mucoid and non‐mucoid phenotypes.

Augmented effect of early antibiotic treatment in mice with experimental lung infections due to sequentially adapted mucoid strains of Pseudomonas aeruginosa

BACKGROUND Effects of treatment with tobramycin initiated 1 or 24 h post-infection were investigated in a new version of a pulmonary infection model in mice. The model reflects the differentiated behaviour of Pseudomonas aeruginosa mucoid strains isolated from the lungs of one chronically infected cystic fibrosis (CF) patient at different time periods during chronic lung infection. METHODS BALB/c mice were challenged with alginate-embedded mucoid clinical isolates isolated in 1988, 1997 or 2003. Mice were euthanized on day 1, 2 or 3 post-infection for estimation of quantitative bacteriology, histopathology, and measurement of granulocyte colony-stimulating factor (G-CSF) and macrophage inflammatory protein 2 (MIP-2). RESULTS There was a significant reduction of bacteria when comparing treatment initiated 1 h post-infection with treatment initiated after 24 h for isolates 1997 and 2003. Treatment initiated 1 h post-infection also resulted in a reduction of the pulmonary cytokines G-CSF, for all three isolates, and MIP-2, for isolates 1997 and 2003. Histological evaluation showed a shift from the acute-type inflammatory immune response to a chronic-type in mice infected with isolate 2003. CONCLUSIONS A significant reduction in the number of bacteria was observed when initiating treatment 1 h post-infection compared with initiating treatment after 24 h, although the latest isolate avoided complete clearance. Early antibiotic treatment directed at the mucoid phenotype in mice also reduced the inflammation and, thereby, the lung tissue damage.

Cytokine and surface receptor diversity of NK cells in resistant C3H/HeN and susceptible BALB/c mice with chronic Pseudomonas aeruginosa lung infection

The purpose of the present study was to investigate whether NK cells from resistant C3H/HeN mice and susceptible BALB/c mice showed different release of cytokines and expression of surface molecules during chronic P. aeruginosa lung infection using alginate‐embedded P. aeruginosa mimicking the infection in cystic fibrosis. Lung cell suspensions were depleted of lymphocytes by magnetic cell sorting. The concentrations of IFN‐γ, IL‐1β and GM‐CSF were estimated by ELISA at day 1 and 2 after infection. Non‐infected mice were used as controls. Flow cytometry was used to estimate the surface expression of the LFA‐1 and Fc receptors on NK cells. At day 2, IFN‐γ levels increased in C3H/HeN mice but decreased in BALB/c mice. The GM‐CSF levels increased only in the C3H/HeN mice at day 1 and 2. Surface expression of LFA‐1 on the NK cells was higher in C3H/HeN mice at day 1 and 2. In contrast, the expression of Fc receptors was significantly lower on NK cells in C3H/HeN mice at day 1 and 2. In conclusion, the present results show phenotypic differences in NK cells in the two mice strains in chronic P. aeruginosa lung infection, indicating different modulating effects in the Th1/Th2 balance.

Cryptococcal rib osteomyelitis as primary and only symptom of idiopathic CD4 penia.

A 59-year old man with idiopathic CD4 lymphopenia presented with extensive disseminated Cryptococcus neoformans infection including a large rib cryptoccocoma, vertebral spondylitis and pleural empyema. Complete resection of the affected part of the rib was necessary after failure of initial antifungal treatment. The vertebral spondylitis has been successfully managed at 3 years of follow-up by continuous itraconazole treatment and regular MRI combined with leucocyte scintigraphy assessment.

Animal models of chronic wound care: the application of biofilms in clinical research

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Burn Mouse Models

Severe thermal injury induces immunosuppression, involving all parts of the immune system, especially when large fractions of the total body surface area are affected. An animal model was established to characterize the burn-induced immunosuppression. In our novel mouse model a 6 % third-degree burn injury was induced with a hot-air blower. The third-degree burn was confirmed histologically. At 48 h, a decline in the concentration of peripheral blood leucocytes was observed in the group of mice with burn wound. The reduction was ascribed to the decline in concentration of polymorphonuclear neutrophil leucocytes and monocytes. When infecting the skin with Pseudomonas aeruginosa, a dissemination of bacteria was observed only in the burn wound group. Histological characterization of the skin showed an increased polymorphonuclear neutrophil granulocytes dominated inflammation in the group of mice with infected burn wound compared with the burn wound only group. The burn mouse model resembles the clinical situation and provides an opportunity to examine or develop new strategies like new antibiotics and immune therapy, in handling burn wound victims much.

Dietzia papillomatosis bacteremia

ABSTRACT The clinical significance of Dietzia papillomatosis is for the moment limited to the rare skin disease confluent and reticulated papillomatosis. We present a case of infection with D. papillomatosis in a 2-year-old boy with known syringomyelia. The microbiological diagnosis was done using 16S rRNA gene sequencing. This is the first report of bacteremia with D. papillomatosis.

Noninvasive measurement of reepithelialization and microvascularity of suction-blister wounds with benchmarking to histology: Suction-blister wound healing model in humans

We explored use of the suction‐blister wound model in the assessment of not only epidermal regeneration but also pain, the microvascular response and bacteriology. The effects of topical zinc sulfate were studied to articulate the methodologies in this double‐blind trial. One epidermal suction blister (10 mm) was induced on each buttock in 30 healthy volunteers (15 females:15 males) and deroofed on day 0. The wounds were randomized to daily treatment with 1.4% zinc sulfate shower gel (n = 20), placebo (n = 20) or control (n = 20). Digital photography coupled with planimetry, transepidermal water loss (TEWL) measurement and optical coherence tomography (OCT) was benchmarked to the gold standard of histology of 60 full‐thickness wound biopsies on day 4. Pain increased after application of the shower gels. Microvessel density, determined from OCT images, increased from day 0 to day 2 in the three groups but increased more with the placebo than with the zinc shower gel (p = 0.003) or the control treatment (p = 0.002) and correlated (rS = 0.313, p = 0.015) with the inflammatory response on day 4, as determined by histology. Coagulase‐negative staphylococci were more common in wounds compared with skin (p = 0.002) and was reduced (p = 0.030) with zinc sulfate treatment. Planimetric analysis of digital wound images was not biased (p = 0.234) compared with histology, and TEWL measurements showed no correlation (rS = 0.052, p = 0.691) with epithelialization. Neoepidermal formation, determined by histology, did not differ (p = 0.290) among the groups. Zinc sulfate reduced (p = 0.031) the release of lactate dehydrogenase from cultured gel‐treated keratinocytes isolated from the blister roofs. Therefore, combination of the standardized suction‐blister wound model with noninvasive planimetry and OCT is a useful tool for assessing wound therapies. Zinc sulfate transiently dampened inflammation and reduced bacterial growth.