A study on the ability of quaternary ammonium groups attached to a polyurethane foam wound dressing to inhibit bacterial attachment and biofilm formation

Bacterial infection of acute and chronic wounds impedes wound healing significantly. Part of this impediment is the ability of bacterial pathogens to grow in wound dressings. In this study, we examined the effectiveness of a polyurethane (PU) foam wound dressings coated with poly diallyl‐dimethylammonium chloride (pDADMAC‐PU) to inhibit the growth and biofilm development by three main wound pathogens, Staphylococcus aureus, Pseudomonas aeruginosa, and Acinetobacter baumannii, within the wound dressing. pDADMAC‐PU inhibited the growth of all three pathogens. Time‐kill curves were conducted both with and without serum to determine the killing kinetic of pDADMAC‐PU. pDADMAC‐PU killed S. aureus, A. baumannii, and P. aeruginosa. The effect of pDADMAC‐PU on biofilm development was analyzed quantitatively and qualitatively. Quantitative analysis, colony‐forming unit assay, revealed that pDADMAC‐PU dressing produced more than eight log reduction in biofilm formation by each pathogen. Visualization of the biofilms by either confocal laser scanning microscopy or scanning electron microscopy confirmed these findings. In addition, it was found that the pDADMAC‐PU‐treated foam totally inhibited migration of bacteria through the foam for all three bacterial strains. These results suggest that pDADMAC‐PU is an effective wound dressing that inhibits the growth of wound pathogens both within the wound and in the wound dressing.     

Time course study of delayed wound healing in a biofilm‐challenged diabetic mouse model

Bacterial biofilm has been shown to play a role in delaying wound healing of chronic wounds, a major medical problem that results in significant health care burden. A reproducible animal model could be very valuable for studying the mechanism and management of chronic wounds. Our previous work showed that Pseudomonas aeruginosa (PAO1) biofilm challenge on wounds in diabetic (db/db) mice significantly delayed wound healing. In this wound time course study, we further characterize the bacterial burden, delayed wound healing, and certain aspects of the host inflammatory response in the PAO1 biofilm‐challenged db/db mouse model. PAO1 biofilms were transferred onto 2‐day‐old wounds created on the dorsal surface of db/db mice. Control wounds without biofilm challenge healed by 4 weeks, consistent with previous studies; none of the biofilm‐challenged wounds healed by 4 weeks. Of the biofilm‐challenged wounds, 64% healed by 6 weeks, and all of the biofilm‐challenged wounds healed by 8 weeks. During the wound‐healing process, P. aeruginosa was gradually cleared from the wounds while the presence of Staphylococcus aureus (part of the normal mouse skin flora) increased. Scabs from all unhealed wounds contained 10⁷ P. aeruginosa, which was 100‐fold higher than the counts isolated from wound beds (i.e., 99% of the P. aeruginosa was in the scab). Histology and genetic analysis showed proliferative epidermis, deficient vascularization, and increased inflammatory cytokines. Hypoxia inducible factor expression increased threefold in 4‐week wounds. In summary, our study shows that biofilm‐challenged wounds typically heal in approximately 6 weeks, at least 2 weeks longer than nonbiofilm‐challenged normal wounds. These data suggest that this delayed wound healing model enables the in vivo study of bacterial biofilm responses to host defenses and the effects of biofilms on host wound healing pathways. It may also be used to test antibiofilm strategies for treating chronic wounds.     

Cadexomer iodine provides superior efficacy against bacterial wound biofilms in vitro and in vivo

Examination of clinical samples indicates bacterial biofilms are present in the majority of chronic wounds, and substantial evidence suggests biofilms contribute significantly to delayed healing. Bacteria in biofilms are highly tolerant of antimicrobials, and little data exist to guide the choice of anti‐biofilm wound therapy. Cadexomer iodine (CI) was recently reported to have superior efficacy compared to diverse wound dressings against Pseudomonas aeruginosa biofilms in an ex vivo model. In the current study, the strong performance of CI vs. P. aeruginosa biofilm was confirmed using colony and colony drip‐flow in vitro wound biofilm models. Similar in vitro efficacy of CI was also demonstrated against mature Staphylococcus aureus biofilms using the same models. Additionally, the rapid kill of mature S. aureus and P. aeruginosa colony biofilms was visualized by confocal microscopy using Live/Dead fluorescent stains. Superior in vitro efficacy of CI vs. staphylococcal biofilms was further demonstrated against methicillin‐resistant S. aureus (MRSA) using multiple biofilm models with log reduction, Live/Dead, and metabolic endpoints. Comparator antimicrobial dressings, including silver‐based dressings used throughout and other active agents used in individual models, elucidated only limited effects against the mature biofilms. Given the promising in vitro activity, CI was tested in an established mouse model of MRSA wound biofilm. CI had significantly greater impact on MRSA biofilm in mouse wounds than silver dressings or mupirocin based on Gram‐stained histology sections and quantitative microbiology from biopsy samples (>4 log reduction in CFU/g vs. 0.7–1.6, p < 0.0001). The superior efficacy for CI in these in vitro and in vivo models suggests CI topical products may represent a better choice to address established bacterial biofilm in chronic wounds.     

Preclinical evaluation of a novel silver gelling fiber dressing on Pseudomonas aeruginosa in a porcine wound infection model

The wound environment is a fertile ground for biofilm forming pathogens. Once biofilms form within the wound, they can be very challenging to eradicate. The purpose of this study was to examine the effect of a gelling fiber dressing with silver using a well‐established porcine wound biofilm model. Deep partial thickness wounds were inoculated with Pseudomonas aeruginosa ATCC 27312 and covered with a polyurethane film dressing to promote biofilm formation. Wounds were then divided into treatment groups: gelling fiber dressing with silver, gelling fiber dressing without silver, hydrofiber dressing with silver, benzethonium chloride and ethylenediaminetetraacetic acid and compared to untreated control. Microbiological, biofilm, and histological wound assessments were performed on days 3, 5, and 7 postinfection. Treatment with gelling fiber dressing with silver resulted in significant reduction of P. aeruginosa biofilm when compared to all other treatment groups on every assessment time point. In addition, gelling fiber dressing with silver treatment resulted in detachment of biofilm from the wound, while wounds treated with gelling fiber dressing with and without silver showed more granulation tissue formation on day 3. Our data show that a new gelling fiber dressing with silver was effective in reducing biofilm associated P. aeruginosa in vivo. This study may have important clinical implications especially for wounds heavily colonized with gram‐negative biofilm‐forming bacteria.     

Noncontact,low‐frequency ultrasound as an effective therapy against Pseudomonas aeruginosa–infected biofilm wounds

Bacterial biofilms, a critical chronic wound mediator, remain difficult to treat. Energy‐based devices may potentially improve healing, but with no evidence of efficacy against biofilms. This study evaluates noncontact, low‐frequency ultrasound (NLFU) in the treatment of biofilm‐infected wounds. Six‐millimeter dermal punch wounds in rabbit ears were inoculated with 10⁷ colony‐forming units of Pseudomonas aeruginosa or left as sterile controls. A biofilm was established in vivo using our published model. NLFU treatment was carried out every other day or every day, with contralateral ear wounds acting as internal, untreated controls. Wounds were harvested for several quantitative endpoints and scanning electron microscopy to evaluate the biofilm structure. The P. aeruginosa biofilm consistently impaired wound epithelialization and granulation. NLFU, both every other day and every day, improved healing and reduced bacterial counts relative to untreated controls (p < 0.05). Scanning electron microscopy confirmed a qualitative decrease in bacteria after both treatments. NLFU also reduced inflammatory cytokine expression (p < 0.05). Our study suggests that NLFU is an effective therapy against P. aeruginosa wound biofilm. This represents the first in vivo evidence of energy‐based modalities' impact on wound biofilm, setting the foundation for future mechanistic studies. Continued wound care technology research is essential to improving our understanding, and treatment, of biofilm‐infected chronic wounds.     

Pseudomonas aeruginosa uses T3SS to inhibit diabetic wound healing

Diabetic foot ulcers are responsible for more hospitalizations than any other complication of diabetes. Bacterial infection is recognized as an important factor associated with impaired healing in diabetic ulcers. Pseudomonas aeruginosa is the most frequently detected Gram‐negative pathogen in diabetic ulcers. P. aeruginosa infection has been shown to impair healing in diabetic wounds in a manner that correlates with its ability to form biofilm. While the majority of infections in diabetic ulcers are biofilm associated, 33% of infections are nonbiofilm in nature. P. aeruginosa is the most prevalent Gram‐negative pathogen in all diabetic wound types, which suggests that the deleterious impact of P. aeruginosa on healing in diabetic wounds goes beyond its ability to form biofilm and likely involves other factors. The Type III Secretion System (T3SS) virulence structure is required for the pathogenesis of all P. aeruginosa clinical isolates, suggesting that it may also play a role in the inhibition of wound repair in diabetic skin ulcers. We evaluated the role of T3SS in mediating P. aeruginosa–induced tissue damage in the wounds of diabetic mice. Our data demonstrate that P. aeruginosa establishes a robust and persistent infection in diabetic wounds independent of its ability to form biofilm and causes severe wound damage in a manner that primarily depends on its T3SS.     

Development of a novel ex vivo porcine skin explant model for the assessment of mature bacterial biofilms

Bacterial biofilms have been proposed to be a major factor contributing to the failure of chronic wounds to heal because of their increased tolerance to antimicrobial agents and the prolonged inflammation they cause. Phenotypic characteristics of bacterial biofilms vary depending on the substratum to which they attach, the nutritional environment, and the microorganisms within the biofilm community. To develop an ex vivo biofilm model that more closely mimics biofilms in chronic skin wounds, we developed an optimal procedure to grow mature biofilms on a central partial‐thickness wound in 12‐mm porcine skin explants. Chlorine gas produced optimal sterilization of explants while preserving histological properties of the epidermis and dermis. Pseudomonas aeruginosa and Staphylococcus aureus developed mature biofilms after 3 days that had dramatically increased tolerance to gentamicin and oxacillin (～100× and 8,000× minimal inhibitory concentration, respectively) and to sodium hypochlorite (0.6% active chlorine). Scanning electron microscopy and confocal microscopy verified extensive exopolymeric biofilm structures on the explants. Despite a significant delay, a ΔlasI quorum‐sensing mutant of P. aeruginosa developed biofilm as antibiotic‐tolerant as wild‐type after 3 days. This ex vivo model simulates growth of biofilms on skin wounds and provides an accurate model to assess effects of antimicrobial agents on mature biofilms.     

Impact of a novel,antimicrobial dressing on in vivo,Pseudomonas aeruginosa wound biofilm: Quantitative comparative analysis using a rabbit ear model

The importance of bacterial biofilms to chronic wound pathogenesis is well established. Different treatment modalities, including topical dressings, have yet to show consistent efficacy against wound biofilm. This study evaluates the impact of a novel, antimicrobial Test Dressing on Pseudomonas aeruginosa biofilm‐infected wounds. Six‐mm dermal punch wounds in rabbit ears were inoculated with 10⁶ colony‐forming units of P. aeruginosa. Biofilm was established in vivo using our published model. Dressing changes were performed every other day with either Active Control or Test Dressings. Treated and untreated wounds were harvested for several quantitative endpoints. Confirmatory studies were performed to measure treatment impact on in vitro P. aeruginosa and in vivo polybacterial wounds containing P. aeruginosa and Staphylococcus aureus. The Test Dressing consistently decreased P. aeruginosa bacterial counts, and improved wound healing relative to Inactive Vehicle and Active Control wounds (p < 0.05). In vitro bacterial counts were also significantly reduced following Test Dressing therapy (p < 0.05). Similarly, improvements in bacterial burden and wound healing were also achieved in polybacterial wounds (p < 0.05). This study represents the first quantifiable and consistent in vivo evidence of a topical antimicrobial dressing's impact against established wound biofilm. The development of clinically applicable therapies against biofilm such as this is critical to improving chronic wound care.     

Delayed wound healing in diabetic (db/db) mice with Pseudomonas aeruginosa biofilm challenge: a model for the study of chronic wounds

Chronic wounds are a major clinical problem that lead to considerable morbidity and mortality. We hypothesized that an important factor in the failure of chronic wounds to heal was the presence of microbial biofilm resistant to antibiotics and protected from host defenses. A major difficulty in studying chronic wounds is the absence of suitable animal models. The goal of this study was to create a reproducible chronic wound model in diabetic mice by the application of bacterial biofilm. Six‐millimeter punch biopsy wounds were created on the dorsal surface of diabetic (db/db) mice, subsequently challenged with Pseudomonas aeruginosa (PAO1) biofilms 2 days postwounding, and covered with semiocclusive dressings for 2 weeks. Most of the control wounds were epithelialized by 28 days postwounding. In contrast, none of biofilm‐challenged wounds were closed. Histological analysis showed extensive inflammatory cell infiltration, tissue necrosis, and epidermal hyperplasia adjacent to challenged wounds—all indicators of an inflammatory nonhealing wound. Quantitative cultures and transmission electron microscopy demonstrated that the majority of bacteria were in the scab above the wound bed rather than in the wound tissue. The model was reproducible, allowed localized cutaneous wound infections without high mortality, and demonstrated delayed wound healing following a biofilm challenge. This model may provide an approach to study the role of microbial biofilms in chronic wounds as well as the effect of specific biofilm therapy on wound healing.     

Pseudomonas aeruginosa biofilm hampers murine central wound healing by suppression of vascular epithelial growth factor

Biofilm‐infected wounds are clinically challenging. Vascular endothelial growth factor and host defence S100A8/A9 are crucial for wound healing but may be suppressed by biofilms. The natural course of Pseudomonas aeruginosa biofilm infection was compared in central and peripheral zones of burn‐wounded, infection‐susceptible BALB/c mice, which display delayed wound closure compared to C3H/HeN mice. Wounds were evaluated histopathologically 4, 7 or 10 days post‐infection. Photoplanimetry evaluated necrotic areas. P. aeruginosa biofilm suppressed vascular endothelial growth factor levels centrally in BALB/c wounds but increased peripheral levels 4–7 days post‐infection. Central zones of the burn wound displayed lower levels of central vascular endothelial growth factor as observed 4 and 7 days post‐infection in BALB/c mice compared to their C3H/HeN counterparts. Biofilm suppressed early, centrally located S100A8/A9 in BALB/c and centrally and peripherally later on in C3H/HeN wounds as compared to uninfected mice. Peripheral polymorphonuclear‐dominated inflammation and larger necrosis were observed in BALB/c wounds. In conclusion, P. aeruginosa biofilm modulates wounds by suppressing central, but inducing peripheral, vascular endothelial growth factor levels and reducing host response in wounds of BALB/c mice. This suppression is detrimental to the resolution of biofilm‐infected necrosis.     

Efficacy of a surfactant‐based wound dressing on biofilm control

The aim of this study was to evaluate the efficacy of both a nonantimicrobial and antimicrobial (1% silver sulfadiazine—SSD) surfactant‐based wound dressing in the control of Pseudomonas aeruginosa, Enterococcus sp, Staphylococcus epidermidis, Staphylococcus aureus, and methicillin‐resistant S. aureus (MRSA) biofilms. Anti‐biofilm efficacy was evaluated in numerous adapted American Standards for Testing and Materials (ASTM) standard biofilm models and other bespoke biofilm models. The ASTM standard models employed included the Minimum biofilm eradication concentration (MBEC) biofilm model (ASTM E2799) and the Centers for Disease Control (CDC) biofilm reactor model (ASTM 2871). Such bespoke biofilm models included the filter biofilm model and the chamberslide biofilm model. Results showed complete kill of microorganisms within a biofilm using the antimicrobial surfactant‐based wound dressing. Interestingly, the nonantimicrobial surfactant‐based dressing could disrupt existing biofilms by causing biofilm detachment. Prior to biofilm detachment, we demonstrated, using confocal laser scanning microscopy (CLSM), the dispersive effect of the nonantimicrobial surfactant‐based wound dressing on the biofilm within 10 minutes of treatment. Furthermore, the non‐antimicrobial surfactant‐based wound dressing caused an increase in microbial flocculation/aggregation, important for microbial concentration. In conclusion, this nonantimicrobial surfactant‐based wound dressing leads to the effective detachment and dispersion of in vitro biofilms. The use of surfactant‐based wound dressings in a clinical setting may help to disrupt existing biofilm from wound tissue and may increase the action of antimicrobial treatment.     

The ability of quaternary ammonium groups attached to a urethane bandage to inhibit bacterial attachment and biofilm formation in a mouse wound model

For proper wound healing, control of bacteria or bacterial infections is of major importance. While caring for a wound, dressing material plays a key role as bacteria can live in the bandage and keep re‐infecting the wound. They do this by forming biofilms in the bandage, which slough off planktonic bacteria and overwhelm the host defense. It is thus necessary to develop a wound dressing that will inhibit bacterial growth. This study examines the effectiveness of a polyurethane foam wound dressing bound with polydiallyl‐dimethylammonium chloride (pDADMAC) to inhibit the growth of bacteria in a wound on the back of a mouse. This technology does not allow pDADMAC to leach away from the dressing into the wound, thereby preventing cytotoxic effects. Staphylococcus aureus, Pseudomonas aeruginosa and Acinetobacter baumannii were chosen for the study to infect the wounds. S. aureus and P. aeruginosa are important pathogens in wound infections, while A. baumannii was selected because of its ability to acquire or upregulate antibiotic drug resistance determinants. In addition, two different isolates of methicillin‐resistant S. aureus (MRSA) were tested. All the bacteria were measured in the wound dressing and in the wound tissue under the dressing. Using colony‐forming unit (CFU) assays, over six logs of inhibition (100%) were found for all the bacterial strains using pDADMAC‐treated wound dressing when compared with control‐untreated dressing. The CFU assay results obtained with the tissues were significant as there were 4–5 logs of reduction (100%) of the test organism in the tissue of the pDADMAC‐covered wound versus that of the control dressing‐covered wound. As the pDADMAC cannot leave the dressing (like other antimicrobials), this would imply that the dressing acts as a reservoir for free bacteria from a biofilm and plays a significant role in the development of a wound infection.     

The role of allogenic keratin‐derived dressing in wound healing in a mouse model

Keratin is an interesting protein needed for wound healing and tissue recovery. We have recently proposed a new, simple and inexpensive method to obtain fur and hair keratin‐derived biomaterials suitable for medical application. The aim of the study was to evaluate the role of the fur keratin‐derived protein (FKDP) dressing in the allogenic full‐thickness surgical skin wound model. The data obtained using scanning electron microscopy showed that employed processed biomaterial had higher surface porosity compared with control raw material. From the MTS test, it was found keratin biomaterial is not only toxic to the NIH/3T3 cell line (p < 0.05), but also enhances cell proliferation compared with the control. In vivo studies have shown keratin dressings are tissue biocompatible, accelerate wound closure and epithelialization to the statistically significant differences on day 5 (p < 0.05) in comparison to control wounds. Histological examination revealed, that in FKDP‐treated wounds the inflammatory response contained predominantly macrophages whilst their morphological untreated variants showed mixed cell infiltrates rich in neutrophils. Predominant macrophages based response creates more favorable environment for healing. In FKDP‐dressed wounds the number of microhemorrhages was also significantly decreased (p < 0.05) as compared with undressed wounds. Applied keratin dressing favors reconstruction of a more regular skin structure and assures better cosmetic effect in terms of scar formation and appearance. In conclusion, fur keratin‐derived protein dressings significantly accelerated wound healing in the mouse model. Further studies are needed to determine the molecular mechanisms involved in the multilayer wound healing process and to assess the possible use of these dressings for medical purposes.     

Inhibition of biofilms by glucose oxidase,lactoperoxidase and guaiacol: the active antibacterial component in an enzyme alginogel

The association of biofilms with wound chronicity has prompted a search for antimicrobial interventions that are effective against biofilms. A patented preparation of glucose oxidase, lactoperoxidase and guaiacol (GLG), which is the antibacterial component of Flaminal®, has been shown to inhibit a wide range of bacteria, but it has not yet been tested on biofilms. This study aims to determine the effect of GLG on biofilms of Staphylococcus aureus, methicillin‐resistant S. aureus and Pseudomonas aeruginosa. Static biofilms were grown in microtitre plates and on coverslips and treated with a range of concentrations of GLG. Effects were monitored by estimating biofilm biomass by staining with crystal violet, biofilm activity by staining with either resazurin or fluorescein diacetate and biofilm viability by staining with LIVE/DEAD® BacLight? Bacterial Viability Kit. GLG was able to prevent the formation of biofilms at concentration ≤0·5% (w/v) and higher concentrations were required to inhibit established biofilms. GLG did not disrupt biofilm biomass. Staphylococci were more susceptible to GLG than P. aeruginosa. These in vitro findings must be verified by in vivo studies.     

Povidone‐iodine ointment demonstrates in vitro efficacy against biofilm formation

Anti‐infectives used to treat chronic exuding wounds are diluted by wound exudates, absorbed into dressings, metabolised by proteases and destroyed by pH. In order to mimic such effects of exudates, the efficacy of six topical wound agents was assessed undiluted and at 10% concentrations, including povidone‐iodine ointment and a silver‐impregnated wound dressing, to remove biofilms of Pseudomonas aeruginosa, multi‐species biofilms of Candida albicans and methicillin‐resistant Staphylococcus aureus (MRSA) in vitro in a Centers for Disease Control and Prevention (CDC) reactor. Povidone‐iodine was also diluted to 3·3% and 33·3% of the commercial concentrations. Viable microorganisms in each preparation were quantified by colony count. No viable P. aeruginosa biofilm material was recovered after 4 and 24 hours of treatment with povidone‐iodine ointment at the 100% and 10% concentrations. No C. albicans/MRSA biofilm material was recovered after 4 and 24 hours of treatment with povidone‐iodine ointment at the 100% concentration. In general, following dilution, povidone‐iodine ointment appeared to exhibit greater biofilm removal than the other agents tested. Further research involving different microorganisms in vitro and in vivo over a longer period of time will help elucidate the full potential of povidone‐iodine ointment and liposomal hydrogel.     

The wound‐healing effects of a next‐generation anti‐biofilm silver Hydrofiber wound dressing on deep partial‐thickness wounds using a porcine model

Topical antimicrobials are widely used to control wound bioburden and facilitate wound healing; however, the fine balance between antimicrobial efficacy and non‐toxicity must be achieved. This study evaluated whether an anti‐biofilm silver‐containing wound dressing interfered with the normal healing process in non‐contaminated deep partial thickness wounds. In an in‐vivo porcine wound model using 2 pigs, 96 wounds were randomly assigned to 1 of 3 dressing groups: anti‐biofilm silver Hydrofiber dressing (test), silver Hydrofiber dressing (control), or polyurethane film dressing (control). Wounds were investigated for 8 days, and wound biopsies (n = 4) were taken from each dressing group, per animal, on days 2, 4, 6, and 8 after wounding and evaluated using light microscopy. No statistically significant differences were observed in the rate of reepithelialisation, white blood cell infiltration, angiogenesis, or granulation tissue formation following application of the anti‐biofilm silver Hydrofiber dressing versus the 2 control dressings. Overall, epithelial thickness was similar between groups. Some differences in infiltration of specific cell types were observed between groups. There were no signs of tissue necrosis, fibrosis, or fatty infiltration in any group. An anti‐biofilm silver Hydrofiber wound dressing did not cause any notable interference with normal healing processes.     

A proof‐of‐concept study of the removal of early and late phase biofilm from skin wound models using a liquid acoustic stream

Chronic wounds fail to progress through the normal stages of healing, with the largest remediable cause of chronicity being presence of a multi‐species biofilm. Removal of biofilm from the wound environment is central to wound care. A device for mechanically removing biofilms from wounds has been devised. The removal is caused by small‐scale liquid currents and shear, generated by acoustically activated microscopic air bubbles. These bubbles and acoustic waves are delivered onto the wound by a gentle liquid stream, allowing cleaning in situ and removal of debris in the run‐off liquid. We have investigated if this liquid acoustic wound stream (LAWS) can remove bacterial biofilm from soft biological wound models and studied the effect of LAWS on the cellular tissues of the substrate. LAWS will efficiently remove early Pseudomonas aeruginosa biofilm from an artificial wound in a pig''s trotter, 24 hours‐mature biofilm of P. aeruginosa from a pre‐wounded human full thickness skin model (EpiDerm FT), and 3‐day mature biofilm of P. aeruginosa or Staphylococcus aureus from a porcine skin explant. Histological examinations of uninfected EpiDerm models that had been treated by LAWS and then stained with Haematoxylin and Eosin, demonstrated no damage to the human tissue, and wound diameter was smaller in the treated skin models compared with untreated samples. Immunofluorescence staining for cytokeratin 14 showed that keratinocytes had migrated further across the wound in the uninfected samples treated by LAWS. We discuss the implications for wound healing and propose further laboratory and clinical studies to demonstrate the removal of biofilm from patients with chronic leg ulcers and the impact on healing.     

Molecular mechanisms of enhanced wound healing by copper oxide‐impregnated dressings

Copper plays a key role in angiogenesis and in the synthesis and stabilization of extracellular matrix skin proteins, which are critical processes of skin formation. We hypothesized that introducing copper into wound dressings would enhance wound repair. Application of wound dressings containing copper oxide to wounds inflicted in genetically engineered diabetic mice (C57BL/KsOlaHsd‐Lepr^db) resulted in increased gene and in situ up‐regulation of proangiogenic factors (e.g., placental growth factor, hypoxia‐inducible factor‐1 alpha, and vascular endothelial growth factor), increased blood vessel formation (p<0.05), and enhanced wound closure (p<0.01) as compared with control dressings (without copper) or commercial wound dressings containing silver. This study proves the capacity of copper oxide‐containing wound dressings to enhance wound healing and sheds light onto the molecular mechanisms by which copper oxide‐impregnated dressings stimulate wound healing.     

Biofilms and delayed healing – an in vitro evaluation of silver‐ and iodine‐containing dressings and their effect on bacterial and human cells

This study investigated whether there are differences in the ability of wound dressings to modulate certain factors known to affect wound healing. A selection of antimicrobial dressings (AQUACEL ^® Ag Extra^™, AQUACEL ^® Ag+ Extra^™, IODOFLEX ^™, ACTICOAT ^™ 7 and PROMOGRAN PRISMA^™ matrix) were tested for their effect on both bacterial bioburden and human dermal fibroblasts. Some dressings underwent further evaluation for activity against Pseudomonas aeruginosa biofilms using a colony‐drip flow reactor model. The ability of in vitro biofilms to produce proteases, and the effect of PROMOGRAN PRISMA matrix on such proteases, was also investigated. All antimicrobial dressings tested reduced vegetative bacterial load; however, only PROMOGRAN PRISMA matrix was able to significantly reduce biofilm populations (P = 0·01). Additionally, PROMOGRAN PRISMA matrix was the only dressing that did not inhibit dermal fibroblast growth. All other dressings were detrimental to cell viability. In vitro biofilms of Pseudomonas aeruginosa were demonstrated as being capable of releasing bacterial proteases into their surroundings, and incubation with PROMOGRAN PRISMA matrix led to a 77% reduction in activity of such proteases (P = 0·002). The unique ability of PROMOGRAN PRISMA matrix to reduce in vitro vegetative bacteria, biofilm bacteria and bacterial proteases while still allowing dermal fibroblast proliferation may help rebalance the wound environment and reduce the occurrence of 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.