Eradication of multidrug-resistant A. baumannii in burn wounds by antiseptic pulsed electric field.
ABSTRACT: Emerging bacterial resistance to multiple drugs is an increasing problem in burn wound management. New non-pharmacologic interventions are needed for burn wound disinfection. Here we report on a novel physical method for disinfection: antiseptic pulsed electric field (PEF) applied externally to the infected burns. In a mice model, we show that PEF can reduce the load of multidrug resistant Acinetobacter baumannii present in a full thickness burn wound by more than four orders of magnitude, as detected by bioluminescence imaging. Furthermore, using a finite element numerical model, we demonstrate that PEF provides non-thermal, homogeneous, full thickness treatment for the burn wound, thus, overcoming the limitation of treatment depth for many topical antimicrobials. These modeling tools and our in vivo results will be extremely useful for further translation of the PEF technology to the clinical setting, as they provide the essential elements for planning of electrode design and treatment protocol.
Project description:Introduction:The purpose of this study was to evaluate whether cryopreserved (frozen) adipose-derived regenerative cells (ADRCs) have a therapeutic effect on burn wound healing as well as freshly isolated (fresh) ADRCs. Methods:Full thickness burns were created on dorsum of nude mice and burn wound was excised. The wound was covered by artificial dermis with; (i) fresh ADRCs, (ii) frozen ADRCs, and (iii) PBS (control). The assessment for wound healing was performed by morphological, histopathological and immunohistochemical analyses. Results:In vivo analyses exhibited the significant therapeutic effect of frozen ADRCs on burn wound healing up to the similar or higher level of fresh ADRCs. There were significant differences of wound closure, epithelized tissue thickness, and neovascularization between the treatment groups and control group. Although there was no significant difference of therapeutic efficacy between fresh ADRC group and frozen ADRC group, frozen ADRCs improved burn wound healing process in dermal regeneration with increased great type I collagen synthesis compared with fresh ADRCs. Conclusions:These findings indicate that frozen ADRCs allow us to apply not only quickly but also for multiple times, and the cryopreserved ADRCs could therefore be useful for the treatment of burn wounds in clinical settings.
Project description:Healing of burn wounds is often associated with scar formation due to excessive inflammation and delayed wound closure. To date, no effective treatment is available to prevent the fibrotic process. The Renin Angiotensin System (RAS) was shown to be involved in fibrosis in various organs. Statins (e.g. Atorvastatin), Angiotensin receptor antagonists (e.g. Losartan) and the combination of these drugs are able to reduce the local RAS activation, and reduced fibrosis in other organs. We investigated whether inhibition of the RAS could improve healing of burn wounds by treatment with Atorvastatin, Losartan or the combination of both drugs. Therefore, full and partial thickness burn wounds were inflicted on both flanks of Yorkshire pigs. Oral administration of Atorvastatin, Losartan or the combination was started at post-burn day 1 and continued for 28 days. Full thickness wounds were excised and transplanted with an autologous meshed split-thickness skin graft at post-burn day 14. Partial thickness wounds received conservative treatment. Atorvastatin treatment resulted in enhanced graft take and wound closure of the full thickness wounds, faster resolution of neutrophils compared to all treatments and reduced alpha-smooth muscle actin positive cells compared to control treatment. Treatment with Losartan and to a lesser extent the combination therapy resulted in diminished graft take, increased wound contraction and poorer scar outcome. In contrast, Losartan treatment in partial thickness wounds decreased the alpha-smooth muscle actin+ fibroblasts and contraction. In conclusion, we showed differential effects of Losartan and Atorvastatin in full and partial thickness wounds. The extensive graft loss seen in Losartan treated wounds is most likely responsible for the poor clinical outcome of these full thickness burn wounds. Therefore, Losartan treatment should not be started before transplantation in order to prevent graft loss. Atorvastatin seems to accelerate the healing process in full thickness wounds possibly by dampening the pro-inflammatory response.
Project description:We used a modified Walker-Mason scald burn rat model to demonstrate that Pseudomonas aeruginosa, a common opportunistic pathogen in the burn ward and notable biofilm former, establishes biofilms within deep partial-thickness burn wounds in rats.Deep partial-thickness burn wounds, ~10% of the TBSA, were created in anesthetized male Sprague-Dawley rats (350-450 g; n = 84). Immediately post-burn, 100 µl of P. aeruginosa in phosphate-buffered saline at 1 × 103, 1 × 104, or 1 × 105 cells/wound was spread over the burn surface . At 1, 3, 7, and 11 days post-burn, animals were euthanized and blood and tissue were collected for complete blood counts, colony-forming unit (CFU) counts, biofilm gene expression, histology, scanning electron microscopy (SEM), and myeloperoxidase activity in the burn eschar.P. aeruginosa developed robust biofilm wound infections, plateauing at ~1 × 109 CFU/g burn tissue within 7 days regardless of inoculum size. Expression of Pseudomonas alginate genes and other virulence factors in the infected wound indicated formation of mature P. aeruginosa biofilm within the burn eschar. Compared to un-inoculated wounds, P. aeruginosa infection caused both local and systemic immune responses demonstrated by changes in systemic neutrophil counts, histology, and myeloperoxidase activity within the burn wound. Additionally, SEM showed P. aeruginosa enmeshed within an extracellular matrix on the burn surface as well as penetrating 500-600 µm deep into the eschar.P. aeruginosa establishes biofilms within deep partial-thickness burn wounds and invades deep into the burned tissue. This new in vivo biofilm infection model is valuable for testing novel anti-biofilm agents to advance burn care.
Project description:Using Sprague-Dawley rats (350-450 g; n = 61) and the recently updated Walker-Mason rat scald burn model, we demonstrated that Pseudomonas aeruginosa readily formed biofilms within full-thickness burn wounds. Following the burn, wounds were surface-inoculated with P. aeruginosa in phosphate-buffered saline (PBS), while sterile PBS was used for controls. On post-burn days 1, 3, 7, and 11, animals were euthanized and samples collected for quantitative bacteriology, bacterial gene expression, complete blood cell counts, histology, and myeloperoxidase activity. Robust biofilm infections developed in the full-thickness burn wounds inoculated with 1 × 104 CFU of P. aeruginosa. Both histology and scanning electron microscopy showed the pathogen throughout the histologic cross-sections of burned skin. Quantigene analysis revealed significant upregulation of alginate and pellicle biofilm matrix genes of P. aeruginosa within the burn eschar. Additionally, expression of P. aeruginosa proteases and siderophores increased significantly in the burn wound environment. Interestingly, the host's neutrophil response to the pathogen was not elevated in either the eschar or circulating blood when compared to the control burn. This new full-thickness burn biofilm infection model will be used to test new anti-biofilm therapies that may be deployed with soldiers in combat for immediate use at the site of burn injury on the battlefield.
Project description:BACKGROUND:Clinically, severe burns remain one of the most challenging issues, but an ideal treatment is yet absent. Our purpose is to compare the efficacy of stem cell therapy in a preclinical model of burn wound healing. METHODS:Research reports on mesenchymal stem cells (MSCs) for burn wound healing were retrieved from 5 databases: PubMed, Embase, MEDLINE, Web of Science, and the Cochrane Library. The primary outcomes reported in this article include the un-healing rate of the wound area, the closure rate, and the wound area. Secondary outcomes included CD-31, vascular density, interleukin (IL)-10, thickness of eschar tissue, vascular endothelial growth factor (VEGF), and white blood cell count. Finally, a subgroup analysis was conducted to explore heterogeneity that potentially impacted the primary outcomes. A fixed-effects model with a 95% confidence interval (CI) was performed when no significant heterogeneity existed. Otherwise, a random-effects model was used. All data analysis was conducted by using Engauge Digitizer 10.8 and R software. RESULTS:Twenty eligible articles were finally included in the analysis. Stem cell therapy greatly improved the closure rate (2.00, 95% CI 0.52 to 3.48, p?=?0.008) and compromised the wound area (-?2.36; 95% CI -?4.90 to 0.18; p?=?0.069) rather than the un-healing rate of the wound area (-?11.10, 95% CI -?32.97 to 10.78, p?=?0.320). Though p was 0.069, there was a trend toward shrinkage of the burn wound area after stem cell therapy. Vascular density (4.69; 95% CI 0.06 to 9.31; p?=?0.047) and thickness of eschar tissue (6.56, 95% CI 1.15 to 11.98, p?=?0.017) were also discovered to be significantly improved in the burn site of stem cell-treated animals. Moreover, we observed that animals in the stem cell group had an increased white blood cell count (0.84, 95% CI 0.01 to 1.66, p?=?0.047) 5?days post treatment. Other indicators, such as VEGF (p?=?0.381), CD-31 (p?=?0.335) and IL-10 (p?=?0.567), were not significantly impacted. CONCLUSIONS:Despite limited data from preclinical trials, this meta-analysis suggests that stem cell therapy is curative in decreasing the burn wound area and provides some insights into future clinical studies of stem cell therapy for burns.
Project description:Background: An experimental study was undertaken to determine the efficacy of the epidermal growth factor (EGF) with tocotrienol-rich fraction (TRF) cream in the wound-healing process on skin with deep partial-thickness burn in rats. Methods: A total of 180 Sprague-Dawley rats were randomly divided into six groups of six each and were: untreated control, treated with Silverdin® cream, base cream, base cream with c% EGF, base cream with 3% TRF or base cream with c% EGF and 3% TRF, respectively. Creams were applied once daily for 21 consecutive days. Six animals from each group were sacrificed using anaesthetic overdose on the third, seventh, 11th, 14th and 21st day post-burn. Skin tissues with the wound to be examined were excised for macroscopic and microscopic evaluation and biochemical analyses. Results: EGF + TRF formulation decreased the number of neutrophils, lymphocytes and myofibroblasts post-burn. However, no effects on the number of adipose cells in the healing process were recorded. In addition, lipid peroxidation and nitrite production were found to be reduced post-burn, reducing oxidative stress. Conclusions: Results of the present study indicate that the addition of EGF with TRF have ameliorating effects on deep-partial thickness burn healing parameters.
Project description:Burn injury results in an immediate compromised skin state, which puts the affected patient at an immediate risk for infection, including sepsis. For burn patients that develop infections, it is critical to rapidly identify the etiology so that an appropriate treatment can be administered. Current clinical standards rely heavily on culture-based methods for local and systemic infection testing, which can often take days to complete. While more advanced methods (ie, MALDI or NAAT) have improved turnaround times, they may still suffer from either the need for pure culture or sensitivity and specificity issues. Peptide nucleic acid fluorescent in situ hybridization (PNA-FISH) offers a way to reduce this time from days to hours and provide species-specific identification. While PNA-FISH has had great utility in research, its use in clinical microbiology diagnostics has been minimal (including burn wound diagnostics). This work describes a nonculture-based identification technique using commercial available U.S. FDA-approved PNA-FISH probes for the identification of common clinical pathogens, Pseudomonas aeruginosa and Staphylococcus aureus, present in burn wound infections. Additionally, calcofluor white was included for identification of Candida albicans. All three pathogens were identified from a tri-species infected deep-partial thickness rat burn wound model. These species were clearly identifiable in swab and tissue samples that were collected, with minimal autofluorescence from any species. Although autofluorescence of the tissue was present, it did not interfere or was otherwise minimized through sample preparation and analysis. The methodology developed was done so with patient care and diagnostic laboratories in mind that it might be easily transferred to the clinical setting.
Project description:Burn wound injury affects soldiers and civilians alike, often resulting in a dynamic, but un-orchestrated, host response that can lead to infection, scarring, and potentially death. To mitigate these factors, it is important to have a clinically relevant model of burn wound infection that can be utilized for advancing burn wound treatments. Our previous reports have demonstrated the ability of Pseudomonas aeruginosa to generate a biofilm infection within a modified Walker-Mason rat burn model of deep-partial (DPT) and full-thickness (FT) burn wounds (10% total body surface area) in male Sprague-Dawley rats (350-450 g). Here, we further define this model with respect to the host response when challenged with P. aeruginosa infection between the two burn types. Following burn injury and immediate surface exposure to P. aeruginosa, inflammation at the local and systemic levels were monitored for an 11 days period. Compared to burn-only groups, infection with P. aeruginosa further promoted local inflammation in both DPT and FT burn wounds, which was evident by enhanced cellular influx (including neutrophils and monocytes), increased levels of several pro-inflammatory cytokines (IL-1?, IL-6, GRO/KC, andMIP-1?), and reduced IL-10. Systemically, only minor changes were seen in circulating white blood cells and cytokines; however, increases in high mobility group box-1 (HMGB-1) and hyaluronan, as well as decreases in fibronectin were noted particularly in FT burns. Compared to the burn-only group, P. aeruginosa infection resulted in sustained and/or higher levels of HMGB-1 and hyaluronan. Combined with our previous work that defined the burn depth and development of P. aeruginosa biofilms within the wound, this study further establishes this model by defining the host response to the burn and biofilm-infection. Furthermore, this characterization shows several similarities to what is clinically seen and establishes this model for future use in the development and testing of novel therapeutics for burn wound treatment at home and on the battlefield.
Project description:Burn injuries are a leading cause of morbidity including prolonged hospitalization, disfigurement, and disability. Currently there is no Food and Drug Administration-approved burn therapeutics. A clinical distinction of burn injuries from other acute wounds is the event of the so-called secondary burn wound progression within the first week of the injury, in which a burn expands horizontally and vertically from its initial boundary to a larger area. Therefore, an effective therapeutics for burns should show dual abilities to prevent the burn wound progression and thereafter promote burn wound healing. Herein we report that topically applied F-5 fragment of heat shock protein-90? is a dual functional agent to promote burn wound healing in pigs. First, F-5 prevents burn wound progression by protecting the surrounding cells from undergoing heat-induced caspase 3 activation and apoptosis with increased Akt activation. Accordingly, F-5-treated burn and excision wounds show a marked decline in inflammation. Thereafter, F-5 accelerates burn wound healing by stimulating the keratinocyte migration-led reepithelialization, leading to wound closure. This study addresses a topical agent that is capable of preventing burn wound progression and accelerating burn wound healing.
Project description:An adhesive yet easily removable burn wound dressing represents a breakthrough in second-degree burn wound care. Current second-degree burn wound dressings absorb wound exudate, reduce bacterial infections, and maintain a moist environment for healing, but are surgically or mechanically debrided from the wound, causing additional trauma to the newly formed tissues. We have developed an on-demand dissolvable dendritic thioester hydrogel burn dressing for second-degree burn care. The hydrogel is composed of a lysine-based dendron and a PEG-based crosslinker, which are synthesized in high yields. The hydrogel burn dressing covers the wound and acts as a barrier to bacterial infection in an in vivo second-degree burn wound model. A unique feature of the hydrogel is its capability to be dissolved on-demand, via a thiol-thioester exchange reaction, allowing for a facile burn dressing removal.