Association of Metabolic Profile and Microbiome in Chronic Pressure Ulcer Wounds
ABSTRACT: Chronic, non-healing wounds contribute significantly to the suffering of patients with co-morbidities in the clinical population with mild to severely compromised immune systems. Normal wound healing proceeds through a well-described process. However, in chronic wounds this process seems to become dysregulated at the transition between resolution of inflammation and re-epithelialization. Bioburden in the form of colonizing bacteria is a major contributor to the delayed headlining in chronic wounds such as pressure ulcers. However how the microbiome influences the wound metabolic landscape is unknown. Here, we have used a Systems Biology approach to determine the association between the taxonomic and metabolomic profile of wounds colonized by bacteria. Pressure ulcer biopsies were harvested from primary chronic wounds and bisected into top and bottom sections prior to analysis of microbiome by pyrosequencing and analysis of metabolome using 1H nuclear magnetic resonance (NMR) spectroscopy. Bacterial taxonomy revealed that wounds were colonized predominantly by three main phyla, but differed significantly at the genus level. While taxonomic profiles demonstrated significant variability between wounds, metabolic profiles shared significant similarity based on the depth of the wound biopsy. Association between taxonomy and metabolic landscape indicated significant wound-to-wound similarity in metabolite enrichment sets and metabolic pathway impacts, especially with regard to amino acid metabolism. To our knowledge, this is the first demonstration of a statistically robust correlation between bacterial colonization and metabolic landscape within the chronic wound environment.
Project description:Chronic, non-healing wounds contribute significantly to the suffering of patients with co-morbidities in the clinical population with mild to severely compromised immune systems. Normal wound healing proceeds through a well-described process. However, in chronic wounds this process seems to become dysregulated at the transition between resolution of inflammation and re-epithelialization. Bioburden in the form of colonizing bacteria is a major contributor to the delayed headlining in chronic wounds such as pressure ulcers. However how the microbiome influences the wound metabolic landscape is unknown. Here, we have used a Systems Biology approach to determine the biochemical associations between the taxonomic and metabolomic profiles of wounds colonized by bacteria. Pressure ulcer biopsies were harvested from primary chronic wounds and bisected into top and bottom sections prior to analysis of microbiome by pyrosequencing and analysis of metabolome using 1H nuclear magnetic resonance (NMR) spectroscopy. Bacterial taxonomy revealed that wounds were colonized predominantly by three main phyla, but differed significantly at the genus level. While taxonomic profiles demonstrated significant variability between wounds, metabolic profiles shared significant similarity based on the depth of the wound biopsy. Biochemical association between taxonomy and metabolic landscape indicated significant wound-to-wound similarity in metabolite enrichment sets and metabolic pathway impacts, especially with regard to amino acid metabolism. To our knowledge, this is the first demonstration of a statistically robust correlation between bacterial colonization and metabolic landscape within the chronic wound environment.
Project description:UNLABELLED:Chronic nonhealing wounds have been heralded as a silent epidemic, causing significant morbidity and mortality especially in elderly, diabetic, and obese populations. Polymicrobial biofilms in the wound bed are hypothesized to disrupt the highly coordinated and sequential events of cutaneous healing. Both culture-dependent and -independent studies of the chronic-wound microbiome have almost exclusively focused on bacteria, omitting what we hypothesize are important fungal contributions to impaired healing and the development of complications. Here we show for the first time that fungal communities (the mycobiome) in chronic wounds are predictive of healing time, associated with poor outcomes, and form mixed fungal-bacterial biofilms. We longitudinally profiled 100, nonhealing diabetic-foot ulcers with high-throughput sequencing of the pan-fungal internal transcribed spacer 1 (ITS1) locus, estimating that up to 80% of wounds contain fungi, whereas cultures performed in parallel captured only 5% of colonized wounds. The "mycobiome" was highly heterogeneous over time and between subjects. Fungal diversity increased with antibiotic administration and onset of a clinical complication. The proportions of the phylum Ascomycota were significantly greater (P = 0.015) at the beginning of the study in wounds that took >8 weeks to heal. Wound necrosis was distinctly associated with pathogenic fungal species, while taxa identified as allergenic filamentous fungi were associated with low levels of systemic inflammation. Directed culturing of wounds stably colonized by pathogens revealed that interkingdom biofilms formed between yeasts and coisolated bacteria. Combined, our analyses provide enhanced resolution of the mycobiome during impaired wound healing, its role in chronic disease, and impact on clinical outcomes. IMPORTANCE:Wounds are an underappreciated but serious complication for a diverse spectrum of diseases. High-risk groups, such as persons with diabetes, have a 25% lifetime risk of developing a wound that can become chronic. The majority of microbiome research related to chronic wounds is focused on bacteria, but the association of fungi with clinical outcomes remains to be elucidated. Here we describe the dynamic fungal communities in 100 diabetic patients with foot ulcers. We found that communities are unstable over time, but at the first clinical presentation, the relative proportions of different phyla predict healing times. Pathogenic fungi not identified by culture reside in necrotic wounds and are associated with a poor prognosis. In wounds stably colonized by fungi, we identified yeasts capable of forming biofilms in concert with bacteria. Our findings illuminate the associations of the fungal mycobiome with wound prognosis and healing.
Project description:Diabetics chronic wounds are characterized by high levels of oxidative stress (OS) and are often colonized by biofilm-forming bacteria that severely compromise healing and can result in amputation. However, little is known about the role of skin microbiota in wound healing and chronic wound development. We hypothesized that high OS levels lead to chronic wound development by promoting the colonization of biofilm-forming bacteria over commensal/beneficial bacteria. To test this hypothesis, we used our db/db -/- mouse model for chronic wounds where pathogenic biofilms develop naturally after induction of high OS immediately after wounding. We sequenced the bacterial rRNA internal transcribed spacer (ITS) gene of the wound microbiota from wound initiation to fully developed chronic wounds. Indicator species analysis, which considers a species' fidelity and specificity, was used to determine which bacterial species were strongly associated with healing wounds or chronic wounds. We found that healing wounds were colonized by a diverse and dynamic bacterial microbiome that never developed biofilms even though biofilm-forming bacteria were present. Several clinically relevant species that are present in human chronic wounds, such as Cutibacterium acnes, Achromobacter sp., Delftia sp., and Escherichia coli, were highly associated with healing wounds. These bacteria may serve as bioindicators of healing and may actively participate in the processes of wound healing and preventing pathogenic bacteria from colonizing the wound. In contrast, chronic wounds, which had high levels of OS, had low bacterial diversity and were colonized by several clinically relevant, biofilm-forming bacteria such as Pseudomonas aeruginosa, Enterobacter cloacae, Corynebacterium frankenforstense, and Acinetobacter sp. We observed unique population trends: for example, P. aeruginosa associated with aggressive biofilm development, whereas Staphylococcus xylosus was only present early after injury. These findings show that high levels of OS in the wound significantly altered the bacterial wound microbiome, decreasing diversity and promoting the colonization of bacteria from the skin microbiota to form biofilm. In conclusion, bacteria associated with non-chronic or chronic wounds could function as bioindicators of healing or non-healing (chronicity), respectively. Moreover, a better understanding of bacterial interactions between pathogenic and beneficial bacteria within an evolving chronic wound microbiota may lead to better solutions for chronic wound management.
Project description:Diabetic foot ulcers (DFUs) lead to nearly 100,000 lower limb amputations annually in the United States. DFUs are colonized by complex microbial communities, and infection is one of the most common reasons for diabetes-related hospitalizations and amputations. In this study, we examined how DFU microbiomes respond to initial sharp debridement and offloading and how the initial composition associates with 4 week healing outcomes. We employed 16S rRNA next generation sequencing to perform microbial profiling on 50 samples collected from 10 patients with vascularized neuropathic DFUs. Debrided wound samples were obtained at initial visit and after one week from two DFU locations, wound bed and wound edge. Samples of the foot skin outside of the wounds were also collected for comparison. We showed that DFU wound beds are colonized by a greater number of distinct bacterial phylotypes compared to the wound edge or skin outside the wound. However, no significant microbiome diversity changes occurred at the wound sites after one week of standard care. Finally, increased initial abundance of Gram-positive anaerobic cocci (GPAC), especially Peptoniphilus (p < 0.05; n = 5 subjects), was associated with impaired healing; thus, GPAC's abundance could be a predictor of the wound-healing outcome.
Project description:Diabetic foot ulcers (DFUs) are a debilitating complication of diabetes in which bacterial presence, including the frequent colonizer Staphylococcus aureus, contributes to inhibition of healing. MicroRNAs (miRs) play a role in healing and host response to bacterial pathogens. However, the mechanisms by which miR response to cutaneous S. aureus contributes to DFU pathophysiology are unknown. Here, we show that S. aureus inhibits wound closure and induces miR-15b-5p in acute human and porcine wound models and in chronic DFUs. Transcriptome analyses of DFU tissue showed induction of miR-15b-5p to be critical, regulating many cellular processes, including DNA repair and inflammatory response, by suppressing downstream targets IKBKB, WEE1, FGF2, RAD50, MSH2, and KIT. Using a human wound model, we confirmed that S. aureus-triggered miR-15b-5p induction results in suppression of the inflammatory- and DNA repair-related genes IKBKB and WEE1. Inhibition of DNA repair and accumulation of DNA breaks was functionally confirmed by the presence of the pH2AX within colonized DFUs. We conclude that S. aureus induces miR-15b-5p, subsequently repressing DNA repair and inflammatory response, showing a mechanism of inhibition of healing in DFUs previously unreported, to our knowledge. This underscores a previously unknown role of DNA damage repair in the pathophysiology of DFUs colonized with S. aureus.
Project description:BACKGROUND:The aim of this study was to assess the possible association between different factors such as age, sex, antibiotic consumption duration, angiogenesis and pain and "acceleration of wound healing" in pilonidal sinus patients after treating with platelet-rich plasma (PRP). METHODS:In this clinical trial, 110 patients were randomly divided into treatment arm and control group. After surgery, control group underwent classic wound dressing and the treatment arm experienced PRP gel therapy. Before achieving complete healing, wound incisional biopsy was performed in order to evaluate angiogenesis. During the study, other data such as pain and antibiotic consumption duration were also collected. Wound healing time of pilonidal sinus disease was analyzed using Extended and Stratify Cox model. Data were analyzed using R and STATA software. p<0.05 were considered statistically significant. RESULTS:The average wound volume was calculated 41.9 ± 8.01 cc in the controls and 42.35 ± 10.81 in the treatment arm group. The mean of healing time was 8.7 ± 1.18, 4.8 ± 0.87 weeks for control and treatment arm, respectively. There was a significant and strong negative association between healing time and wound volume (p<0.01). Moreover, a significant negative association was found between pain duration and angiogenesis (p<0.001), a strong positive significant association was found between healing time of the treatment arms (p<0.01), and the rate of wound healing for participants treated with PRP gel was 37.2 times more than that of controls. CONCLUSION:Authors hope for these finding to help the future researches to more thoroughly focus on the mentioned factors in order to find a suitable strategy for wound healing using PRP.
Project description:Chronic wounds-including diabetic foot ulcers, venous leg ulcers, and pressure ulcers-represent a major health problem that demands an urgent solution and new therapies. Despite major burden to patients, health care professionals, and health care systems worldwide, there are no efficacious therapies approved for treatment of chronic wounds. One of the major obstacles in achieving wound closure in patients is the lack of epithelial migration. Here, we used multiple pre-clinical wound models to show that Caveolin-1 (Cav1) impedes healing and that targeting Cav1 accelerates wound closure. We found that Cav1 expression is significantly upregulated in wound edge biopsies of patients with non-healing wounds, confirming its healing-inhibitory role. Conversely, Cav1 was absent from the migrating epithelium and is downregulated in acutely healing wounds. Specifically, Cav1 interacted with membranous glucocorticoid receptor (mbGR) and epidermal growth factor receptor (EGFR) in a glucocorticoid-dependent manner to inhibit cutaneous healing. However, pharmacological disruption of caveolae by M?CD or CRISPR/Cas9-mediated Cav1 knockdown resulted in disruption of Cav1-mbGR and Cav1-EGFR complexes and promoted epithelialization and wound healing. Our data reveal a novel mechanism of inhibition of epithelialization and wound closure, providing a rationale for pharmacological targeting of Cav1 as potential therapy for patients with non-healing chronic wounds.
Project description:BACKGROUND:Chronic wounds have been a great burden to patients and the health care system. The popularity of the internet and smart devices, such as mobile phones and tablets, has made it possible to adopt telemedicine (TM) to improve the management of chronic wounds. However, studies conducted by different researchers have reported contradictory results on the effect of TM on chronic wound management. OBJECTIVE:The aim of this work was to evaluate the efficacy and safety of TM in chronic wound management. METHODS:We systematically searched multiple electronic databases (MEDLINE, EMBASE, and Cochrane Central Register of Controlled Trials [CENTRAL]) to identify eligible studies published from inception to June 12, 2019. Inclusion criteria were randomized controlled trials (RCTs) and interventional cohort studies that investigated the use of TM in chronic wound management. RCT and observational data were analyzed separately. A meta-analysis and qualitative analysis were conducted to estimate endpoints. RESULTS:A total of 6 RCTs and 6 cohort studies including 3913 patients were included. Of these, 4 studies used tablets or mobile phones programmed with apps, such as Skype and specialized interactive systems, whereas the remaining 8 studies used email, telephone, and videoconferencing to facilitate the implementation of TM using a specialized system. Efficacy outcomes in RCTs showed no significant differences in wound healing (hazard ratio [HR] 1.16, 95% CI 0.96-1.39; P=.13), and wound healing around 1 year (risk ratio [RR] 1.05, 95% CI 0.89-1.23; P=.15). Noninferiority criteria of TM were met. A decreased risk of amputation in patients receiving TM was revealed (RR 0.45, 95% CI 0.29-0.71; P=.001). The result of cohort studies showed that TM was more effective than standard care (HR 1.74, 95% CI 1.43-2.12; P<.001), whereas the outcome efficacy RR of wound healing around 1 year (RR 1.21, 95% CI 0.96-1.53; P=.56) and 3 months (RR 1.24, 95% CI 0.47-3.3; P=.67) was not significantly different between TM and standard care. Noninferiority criteria of TM were met for wound healing around 1 year in cohort studies. CONCLUSIONS:Currently available evidence suggests that TM seems to have similar efficacy and safety, and met noninferiority criteria with conventional standard care of chronic wounds. Large-scale, well-designed RCTs are warranted.
Project description:The clinical importance of microbiomes to the chronicity of wounds is widely appreciated, yet little is understood about patient-specific processes shaping wound microbiome composition. Here, a two-cohort microbiome-genome wide association study is presented through which patient genomic loci associated with chronic wound microbiome diversity were identified. Further investigation revealed that alternative TLN2 and ZNF521 genotypes explained significant inter-patient variation in relative abundance of two key pathogens, Pseudomonas aeruginosa and Staphylococcus epidermidis. Wound diversity was lowest in Pseudomonas aeruginosa infected wounds, and decreasing wound diversity had a significant negative linear relationship with healing rate. In addition to microbiome characteristics, age, diabetic status, and genetic ancestry all significantly influenced healing. Using structural equation modeling to identify common variance among SNPs, six loci were sufficient to explain 53% of variation in wound microbiome diversity, which was a 10% increase over traditional multiple regression. Focusing on TLN2, genotype at rs8031916 explained expression differences of alternative transcripts that differ in inclusion of important focal adhesion binding domains. Such differences are hypothesized to relate to wound microbiomes and healing through effects on bacterial exploitation of focal adhesions and/or cellular migration. Related, other associated loci were functionally enriched, often with roles in cytoskeletal dynamics. This study, being the first to identify patient genetic determinants for wound microbiomes and healing, implicates genetic variation determining cellular adhesion phenotypes as important drivers of infection type. The identification of predictive biomarkers for chronic wound microbiomes may serve as risk factors and guide treatment by informing patient-specific tendencies of infection.
Project description:Faster and better wound healing is a longstanding goal. Blood flow, angiogenesis, and tissue oxygenation are important parameters in evaluating the healing process. Optical microangiography (OMAG) allows 3D imaging of tissue vasculature and can provide quantitative blood flow information down to the capillary level of resolution. Dual wavelength laser speckle imaging (DW-LSI) can measure tissue oxygenation status.Cutaneous wound healing of a mouse ear model using a multimodal imaging system that combines OMAG with DWLSI was studied.A complete microvasculature map of the ear in vivo was obtained. The imaging system revealed both hemodynamic and metabolic changes during acute stage wound healing. Blood flow velocity, blood flow direction, as well as changes in concentration of oxygenated hemoglobin (?HbO) and deoxygenated hemoglobin (?Hb) were measured and quantified. In addition, capillary recruitment and angiogenesis were visualized during the chronic stage of repairing.The combination of DW-LSI and OMAG imaging technique may be a powerful tool to visualize and understand microvascular, hemodynamic, and metabolic changes during cutaneous wound healing.