Dates
Location
8:00 AM - 8:15 AM
Kenneth W. Liechty, MD; Daria A. Narmoneva, PhD; Carlos Zgheib, PhD, MS
8:15 AM - 9:15AM
Gregory S. Schultz, PhD
Details
9:15 AM - 9:30 AM
9:30 AM - 10:30 AM
Sandeep Gopalakrishnan MS, PhD, MAPWCA; Timothy J. Koh, PhD
Details
10:30 AM - 11:30 AM
Peter M. Abadir, MD; Meghan Brennan, MD
Details
11:30 AM - 11:45 AM
11:45 AM - 12:45 PM
Jun Araki, MD, PhD; Takahiro Umehara, PhD
Details
Jun Araki, MD, PhD
Takahiro Umehara, PhD
12:45 PM - 2:00 PM
Vickie Driver, DPM; Mitchell Sanders, PhD, Daria A. Narmoneva, PhD; Carlos Zgheib, PhD, MS; Praveen Arany, DDS, PhD; Jignesh Patel; Neil Muchin
Details
2:00 PM - 3:00 PM
Subhadip Ghatak, PhD; Steven M. Jay, PhD; Shannon L. Stott, PhD
Details
3:00 PM - 4:00 PM
Eva Nozik, PhD; Arjun Deb, PhD
Details
4:00 PM - 4:15 PM
4:15 PM - 5:15 PM
Jordan Yaron, PhD
Details
6:30 PM-8:30 PM
Registered WHS members are invited to attend. National Harbor Capital Canopy (Walking Distance from Gaylord)
7:00 AM -9:00 AM
9:00 AM - 9:15 AM
9:15 AM - 10:30 AM
10:30 AM - 10:45 A.M
10:45 AM - 11:45 AM
Carlos Zgheib, PhD, MS
Details
11:45 AM - 12:00 PM
12:00 PM - 1:30 PM
1:30 PM - 1:45 PM
1:45 PM - 4:00 PM
Kenneth W. Liechty, MD; Piul Rabbani, PhD; Willeke Daamen, PhD
Details
4:00 PM - 4:15 PM
4:15 PM - 5:15 PM
Kenneth W. Liechty, MD; Piul Rabbani, PhD; Willeke Daamen, PhD
Details
5:15 PM - 7:45 PM
7:00 AM -9:00 AM
9:00 AM - 9:15 AM
9:15 AM - 10:15 AM
Details
10:15 AM - 10:30 AM
10:30 AM - 11:30 AM
Details
11:30 AM - 2:00 PM
12:15 PM - 2:00 PM
Sashwati Roy, PhD; Marjana Tomic-Canic, PhD; Kenneth W. Liechty, MD; Bijan Najafi, PhD
Details
2:00 PM - 2:15 PM
2:15 PM - 3:15 PM
Julie Hakim, MD; Kaitlyn Sadtler, PhD
Details
3:15 PM - 3:30 PM
3:30 PM - 4:30 PM
Alisha Oropallo, PhD; Liping Tang, PhD; Jun Xia, MD
Details
3:15 PM - 3:30 PM
3:30 PM - 4:30 PM
Alisha Oropallo, PhD; Liping Tang, PhD; Jun Xia, MD
Details
4:30 PM - 4:45 PM
4:45 PM - 5:45 PM
Geoffrey C. Gurtner, MD; Rodica Pop-Busui MD, PhD; Chandan K. Sen, PhD; Brian M. Schmidt, DPM, Kellen Chen, PhD, Sashwati Roy, PhD
Details
5:45 PM - 6:00 PM
6:00 PM - 6:30 PM
Timothy Koh, MD; Luisa A. DiPietro, PhD
Details
6:30 PM - 7:00 PM
7:00 PM - 7:25 PM
7:00 PM - 8:30 PM
9:15 AM - 10:15 AM
K. Dev Verma, MD; Rosa Sherafat-Kazemzadeh, MD; Julie Morabito, MD
Details
10:15 AM - 10:30 AM
10:30 A.M. - 11:30 A.M.
Details
11:30 AM
11:30 AM - 2:00 PM
P1.
Skin wound healing requires both cell proliferation and migration. However, the molecular mechanisms governing these activities are not fully understood. The catecholamine epinephrine is a hormone released systemically as part of the stress response and is rapidly degraded within several minutes. On the other hand, levels of catecholamines may be modulated locally within the wound environment, independent of serum levels. We previously reported that 1) epinephrine can inhibit keratinocyte migration and delay wound healing, and 2) the PNMT enzyme that converts norepinephrine into epinephrine is upregulated in wounded epidermis. Here we probed the wound microenvironment for the local generation and metabolism of catecholamines, hypothesizing that these changes could correlate with healing parameters. Using a mouse acute wound model, we assessed tissue catecholamine levels using UHPLC-ED and analyzed tissue gene expressions with quantitative real-time PCR. We find that the tissue and serum concentrations of epinephrine were elevated on days 2 -4 post wounding, relative to unwounded controls (day 0). Likewise, we noted higher mRNA expression of the enzymatic pathways for catecholamine metabolism— dopa decarboxylase (Ddc), monoamine oxidase B (Moab), catechol-O-methyltransferase (Comt), and solute carrier family 6 member 2 (Slc6a2)—on days 1-3 compared to day 0. These enzymes play pivotal roles in the metabolism and catabolism of catecholamines; thus, their elevation can result in increase or decrease in local catecholamine concentrations. Additionally, the expression of the beta-2-adrenergic-receptor (β2AR) receptor for epinephrine was upregulated on days 1-3 compared to day 0. Histological analysis revealed that on day 3 post wounding, there was a decrease in re-epithelization compared to the earlier day 2, and the subsequent days 4-7, suggesting that elevated epinephrine and its signaling through elevated levels of its receptor could be negatively impacting on healing at that time. On the other hand, the increased expression of the enzymatic pathways resulting in degradation of catecholamines might be supporting the increase in re-epithelization observed on days 5-7. Together, these results suggests that the lag in early wound closure might be due to local wound environment modulation of levels of epinephrine and its β2AR receptor, while the increase in wound re-epithelization observed later in the wound healing trajectory may be abetted by upregulation of catecholamine metabolism and its degradation. This avenue of research provides insights into the role of the catecholamine signaling pathway in wound healing program.
P2.
Hydrogels have been the material of choice for regenerative medicine applications due to their biocompatibility that can facilitate cellular attachment, proliferation and provide structural integrity to tissues by serving as scaffolds. The present study aimed at constructing a porous hydrogel composite for the repair of chronic skin wounds. The chitosan-based hydrogel is serving as a natural extracellular matrix (ECM) to enhance cell proliferation and control release of zinc oxide nanoparticles i.e. ZnO-NPs, as the antimicrobial agent. The composite hydrogel contained a blends of chitosan, sodium alginate and elastin were manufactured by the freeze gelation method. The antimicrobial properties against Escherichia coli (E.coli) and Staphylococcus aureus (S. aureus) were tested by incorporating varying concentrations of ZnO-NPs (0, 0.001, 0.01, 0.1 and 1% w/w) into the scaffolds. ZnO-NPs were characterized by UV visible spectroscopy, Scanning electron microscopy (SEM) analysis, Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction (XRD) analysis. The fabricated gels were characterized by SEM analysis, FTIR, XRD, swelling ratio, degradation behavior and release kinetics of loaded ZnO-NPs. In vitro cytocompatibility of the fabricated composite was investigated using human adipose stem cells (ADSCs) by MTT and lactate dehydrogenase (LDH) assay, further assessed by SEM analysis and PKH26 staining. The SEM and XRD analysis confirmed the successful loading of ZnO-NPs into these scaffolds. Fluorescence PKH26 stained images and SEM analysis of ADSCs seeded scaffolds revealed biocompatible nature of the scaffold. ZnO-NPs and all loaded nanocomposite scaffolds exhibited good antibacterial activities. The findings suggested that the developed composite gels have potential clinically for tissue engineering and chronic wound treatment.
P3.
Background: Fibroblast-populated collagen matrices are useful and appropriate experimental models to study cell-dependent structural remodeling and tension generation in soft tissue. Throughout the last 50 years these models have been measured in two dimensions, or characterized based on their weight change, as a response to a tested variable. We recently demonstrated how optical coherence tomography (OCT) can be used to measure the cross-sectional morphology of anchored collagen matrices; here, we demonstrate that a three-dimensional reconstruction based on a complete set of 2D scans of developing matrices can be performed, allowing detailed visualization and characterization of 3D surface wrinkles or curvatures of these tissues undergoing compaction. Methods: A 20 microliter mixture of human fibroblasts and type I collagen in the presence or absence of chitosan was plated centrally in a well of a prewarmed 12-well plate, allowed to polymerize then submerged in media. Matrices were immediately released or allowed to generate tension for 3 days, then released from anchorage. 3D scans were taken prior to release, 15 minutes after release, and 24 hours after release. Scans were then imported into the open-source program 3D Slicer for reconstruction and registration. Results: Control collagen matrices demonstrated patterns of wrinkles along the upper surface and a sharp outside contour, while matrices with chitosan demonstrated smoother edges and a thicker profile. Conclusion: The combination of OCT and 3D rendering software will allow more intricate complex data collection and analysis in tissue models.
P4.
INTRODUCTION
Common methods to limit in-vivo inflammatory signaling consist of either pharmacological therapies or highly invasive implant-based approaches. More recently, however, data have been supportive of ultrasound splenic stimulation, which has been shown to be an effective non-invasive modulator of the cholinergic anti-inflammatory pathway (CAP) in rodent models[DD1] . Here, we test the capacity of splenic ultrasound stimulation to modulate LPS-induced TNF-α production in a non-rodent burn model, to identify whether splenic ultrasound might be a promising therapy to reduce the burn-induced inflammatory response.
METHODS
New Zealand white rabbits underwent three 1-cm contact thermal skin-burn injuries to each ear, with burns controlled so that only the ventral skin is injured down to the perichondrium. One set of rabbits was allocated to undergo ultrasound stimulation (test group), while a control set of rabbits was not stimulated. Beginning one day after burn induction, test rabbits underwent daily 10-minute splenic ultrasound stimulation under isoflurane sedation for a total of eight stimulations[DD2], with control rabbits also administered isoflurane. Blood samples were taken from both test and control groups at baseline (before burn and stimulation), after burn induction but before stimulation, and at the conclusion of ultrasound stimulation. Blood samples were stimulated with LPS for 4 hours, plasma collected, and TNF-α levels determined by ELISA.
RESULTS
Measurement of plasma TNF-α in LPS-stimulated blood samples revealed a highly significant decrease in the test group of rabbits compared to the control rabbits Plasma TNF-α in blood samples unstimulated with LPS remained below the detection limit.
CONCLUSION/DISCUSSION
Ultrasound splenic stimulation shows activation of CAP in a non-rodent burn model, modulating the sensitivity of blood cells to activation by an inflammatory stimulus. These data not only support previous findings, but also suggest that this technology may have beneficial implications in human clinical settings.
P5.
Purpose: Partial thickness burns have traditionally been treated with a combination of topical antimicrobial application, debridement, and serial dressing changes. This methodology is painful, extends hospital length of stay, and may result in higher expense to both the patient and burn facility. Extensive work has been done on developing dressing and skin substitute strategies to mitigate these concerns. Transcyte® is a bioengineered skin substitute comprised of an extracellular matrix of allogeneic human dermal fibroblasts and is indicated in the treatment of partial thickness burns. This work will describe the experience and outcomes in a series of patients where this bioengineered skin substitute was used in the treatment of partial thickness wounds at a single American Burn Association verified burn center.
Methods: This is a case series presentation of patients who were grafted with Transcyte® for their partial thickness burn injuries. It will review challenges in the application and management, outcomes, and outpatient follow up.
Results: Eleven patients underwent placement of the bioengineered skin substitute Transcyte®. Burn injuries were most commonly caused by flame and scald injuries. Total body surface area (TBSA) burned ranged from 1% to 54%. Comorbidities most commonly included type 2 diabetes mellitus and hypertension, but also included end stage renal disease, neuropathy, and psoriasis. Surgical staff training was required to appropriately prepare and time the application of this skin substitute intraoperatively. Only one patient required autografting to an area of skin previously grafted with Transcyte®. Four patients were discharged immediately following surgery and managed on an outpatient basis with an average inpatient length of stay (LOS) of three days. Areas grafted with Transcyte® were mainly the extremities. Grafted areas were covered in petroleum impregnated non-stick dressings, followed by layers of dry gauze. Dressing removal was performed with a multidisciplinary team to facilitate education, training, wound assessment, and discharge planning if a patient remained inpatient. Average time to wound healing was 23.5 days in patients who healed. The average number of clinical follow up visits before determining a healed wound was 1.75 visits.
Conclusion: Partial thickness burn coverage with bioengineered skin substitutes like Transcyte® may offer alternative treatment strategies compared to local wound care. In some cases, it may offer shorter hospital stays, mitigate daily dressing changes, and allow outpatient care and follow up.
P6.
Introduction:
Suprathel® is an absorbable synthetic wound dressing used in partial-thickness wounds of varying etiologies. Research showing acceptable healing as well as improved patient comfort in partial-thickness burns as well as split-thickness skin graft donor sites is well-established. Toxic epidermal necrolysis (TENS) is an immune-mediated disorder affecting cutaneous and mucosal surfaces, often as a reaction to medication, that involves at least 30% of body surface area. Patients diagnosed with the condition are best treated in burn centers given the complexity of the wound care requirements and similarities to large partial-thickness thermal burns. We present our experience treating 6 patients with biopsy-confirmed toxic epidermal necrolysis and document time-to-healing as well as pain associated with treatment with usage of the product at our community-based burn and wound center.
Methods:
A retrospective chart review, evaluating patient characteristics including patient age, patient sex, total body surface area involved and tbsa grafted, time-to healing (>95% epithelialized), as well as pain scores during hospitalization for the 6 patients diagnosed and treated between December 2021 and November 2022.
Results:
All patients experienced complete (>95%) wound healing and epithelialization within __ days post-placement. In addition, average pain scores as documented by nursing were reviewed pre- and post-placement.
Discussion:
Suprathel® poly-lactic acid absorbable polymer has been shown to increase patient pain and lead to healing in various partial-thickness wounds often cared for in the bur arena. Likewise, TENS is an immune-mediated desquamating skin condition best treated in burn centers given the complexity and skin surface size involved. Numerous wound care strategies have been utilized and reviewed in the literature. Given the complexity of the wounds as well as the discomfort experienced by most patients, strategies and wound care regimens that can accelerate healing as well as increase patient comfort and reduce pharmacological pain treatments would be beneficial. Our experience demonstrates acceptable wound healing time compared to the literature, as well as increased patient comfort as documented pre-and post placement.
P6.
Introduction:
Suprathel® is an absorbable synthetic wound dressing used in partial-thickness wounds of varying etiologies. Research showing acceptable healing as well as improved patient comfort in partial-thickness burns as well as split-thickness skin graft donor sites is well-established. Toxic epidermal necrolysis (TENS) is an immune-mediated disorder affecting cutaneous and mucosal surfaces, often as a reaction to medication, that involves at least 30% of body surface area. Patients diagnosed with the condition are best treated in burn centers given the complexity of the wound care requirements and similarities to large partial-thickness thermal burns. We present our experience treating 6 patients with biopsy-confirmed toxic epidermal necrolysis and document time-to-healing as well as pain associated with treatment with usage of the product at our community-based burn and wound center.
Methods:
A retrospective chart review, evaluating patient characteristics including patient age, patient sex, total body surface area involved and tbsa grafted, time-to healing (>95% epithelialized), as well as pain scores during hospitalization for the 6 patients diagnosed and treated between December 2021 and November 2022.
Results:
All patients experienced complete (>95%) wound healing and epithelialization within __ days post-placement. In addition, average pain scores as documented by nursing were reviewed pre- and post-placement.
Discussion:
Suprathel® poly-lactic acid absorbable polymer has been shown to increase patient pain and lead to healing in various partial-thickness wounds often cared for in the bur arena. Likewise, TENS is an immune-mediated desquamating skin condition best treated in burn centers given the complexity and skin surface size involved. Numerous wound care strategies have been utilized and reviewed in the literature. Given the complexity of the wounds as well as the discomfort experienced by most patients, strategies and wound care regimens that can accelerate healing as well as increase patient comfort and reduce pharmacological pain treatments would be beneficial. Our experience demonstrates acceptable wound healing time compared to the literature, as well as increased patient comfort as documented pre-and post placement.
P7.
Canonical Wnt signaling pathway is quiescent in many mammalian organs and gets activated in response to injury. Wnt signaling promotes fibrotic wound healing following acute cutaneous injury by epithelial migration, differentiation and myofibroblast activation. Topical application of Wnt signaling inhibitor promotes regenerative cutaneous repair. To study Wnt signaling modulation in chronic wound repair, we created wounds in STZ-induced type I diabetes C57Bl/6J mouse model. Full thickness excisional wounds activated Wnt signaling in both dermal and epidermal layers identified by beta-catenin immunohistological staining and axin2 transcript levels. Topical application of a small molecule Wnt signaling inhibitor significantly promoted regenerative wound healing in full thickness excisional wound injury models. We provided strong positive implications of Wnt pathway inhibition during chronic cutaneous wound repair in animal models. However, there is a large gap in our understanding of Wnt signaling activation in chronic non-healing human wounds. To understand chronic wound pathologies in humans, we obtained de-identified human wound biopsies from Vanderbilt University Medical Center. Human chronic wound pathologies which include diabetic ulcer, keloids, hypertrophic scars, and melanoma re-excision wounds to screen for fibrosis, collagen deposition and beta-catenin (Wnt signaling) activation. We aim to identify Wnt dependent or independent chronic wound pathologies. Our preliminary results identified that not all chronic wounds are driven by Wnt activation. While hypertrophic scars and decubitus ulcers showed significant Wnt activation, beta catenin activity was not identified in wounds such as keloids. This understanding is crucial to establish a correlation between Wnt signaling activation and chronic non-healing wounds. Our future studies are focused on the identification of the cell types regulated by Wnt activation in vivo and in vitro. Our studies will pave the way to use Wnt signaling inhibitors for selective personalized therapy for chronic wounds.
P8.
Diabetic wound healing impairment is a rapidly rising clinical condition with considerable morbidity. In gene therapy, microRNAs (miR) produce highly specific bio-responses and can treat severe and various diseases. Different types of nanoparticles have been used as carriers to deliver miR, improving therapeutic efficacy by increasing biostability and enhancing cellular uptake and site targeting. In the present study, we compare specific bio-active characters/properties of three candidate nano-carriers: gold (bioactive, metal), silica (bioinert, oxide), and cerium oxide (bioactive, oxide) following modification with miR-146a in the presence of ROS which levels are significantly high in diabetic wounds. Results from the electrochemical studies, enzyme-mimetic activities, and structure-property measurements showed inactivity towards ROS and degradation over cycling for miR-146a conjugated gold and silica nanoparticles, respectively. In contrast, cerium oxide nanoparticle (CNP) conjugates exhibited unique ROS scavenging behavior, conjugation-conferred electrochemical stability, and protected miR-146a from oxidation. Overall, our results support CNPs’ ability to stabilize miR while retaining its antioxidant properties and its impact in nanomedicine.
P9.
Processing of placental membranes results in varying impact on native tissue attributes including extracellular matrix (ECM) composition, growth factors and cytokines, and endogenous cells. Harsher processing methodologies, including dehydration and sterilization, can result in partial loss of protein structure and functionality through denaturation as well as permanent damage to the structural properties of the tissue. A proprietary hypothermic storage processing technique, Allofreshä, has been developed to preserve and maintain the native characteristics of amnion (HSAM†) and chorion (HSCM°) membranes. In this study, the effect of hypothermic storage on placental membranes was evaluated and compared to native fresh tissue by evaluating tissue structure, composition, and in vitro degradation characteristics.
Donor-matched fresh and hypothermically stored placental membranes were evaluated using immunohistochemistry and scanning electron microscopy (SEM). H&E-stained images were used to assess changes in tissue thickness and structure. To evaluate degradation characteristics in vitro, donor-matched membranes were placed in simulated wound fluid (SWF) and cultured at 37°C over 17 days. Tissue loss and architecture were assessed using dry weight and SEM, respectively.
Both HSAM and HSCM maintained a tissue structure characteristic of native amnion and chorion, respectively. HSAM retained intact epithelial, stromal, and spongy layers, while HSCM retained the reticular, basement membrane, and trophoblastic layers. When comparing thickness, no observed changes were seen between fresh amnion and HSAM; however, HSCM was thinner than fresh samples, attributed to a debridement step during processing. In vitro degradation studies found that fresh amnion and chorion and hypothermically stored tissues degraded at similar rates, highlighting the impact of hypothermic storage on retaining fresh tissue characteristics. Furthermore, SEM images of hypothermically stored degraded samples presented with similar tissue architecture when compared to native tissues.
These results demonstrate that hypothermically stored amnion and chorion membranes maintain the native characteristics of fresh tissue.
† Affinity, Organogenesis, Canton, MA
° Novachor, Organogenesis, Canton, MA
P10.
Background: The reconstruction of soft tissue defect on the trochanteric area is challenging. Due to significant complications of regional flap, free tissue transfer is an appropriate option. However, this area has poor recipient vessels. Then we present perforators as the recipient vessels to facilitate the use of free flap coverage for successful reconstruction of defects in the trochanteric area.
Methods: From 2013 to 2017, 10 patients underwent free flap reconstruction for soft tissue defects of the trochanteric area. After preoperative CT or MRI images confirmed the enhanced perforating artery, the site on the skin was identified by Doppler. If the vessel was confirmed reliable, the operation was performed in the same manner as other free flaps.
Results: All flaps survive, and the perforators selected for surgery include four superficial circumflex iliac artery perforators, four tensor fasciae latae artery perforators, and two inferior gluteal artery perforators. The average diameter of the recipient artery was 0.97 mm and the vein was 0.94 mm. One case exhibited arterial insufficiency caused by compression of hematoma; however, the flap fully survived through revised surgery.
Conclusion: The reconstruction of soft tissue defects in the trochanteric area are limited in recipient vessels. However, using a perforator vessel as a recipient facilitates the reconstruction via free flap coverage. This method provides acceptable flap survival and sufficient padding, with less morbidity and collateral injury.
P11.
Background: The present study investigates whether modifiable behavioral factors including current cigarette smoking, heavy alcohol consumption, and regular exercise are associated with the risk of lower extremity amputation (LEA) in diabetic patients.
Methods: 2,644,440 diabetic patients (aged≥20 years) were analyzed by using the database of the Korean National Health Insurance Service. (Table 1) Cox proportional hazard regression was used to assess adjusted hazard ratios for the behavioral factors with the risk of LEA under adjustment for potential confounders.
Results: The risk of LEA was significantly increased by current cigarette smoking and heavy alcohol consumption (HR 1.436; 95% CI, 1.367-1.508; HR 1.082; 95% CI, 1.011-1.158) but significantly decreased by regular exercise (HR 0.745; 95% CI, 0.706-0.786) after adjusting for age, sex, smoke, alcohol drink, exercise, low income, hypertension, dyslipidemia, body mass index, using insulin, oral antidiabetic drugs, and diabetic duration. (Table 2 & Figure 1) The synergistically increased risk of LEA was observed with the more overlap of each inappropriate behavior. The hazard ratio of LEA by current cigarette smoking, heavy alcohol consumption, and regular exercise increased with age, short diabetic duration, and 3 or less oral diabetic drugs. (Table 3)
Conclusion: The modification of behaviors including current smoking, heavy alcohol intake, and regular exercise prevents LEA then improve physical, emotional, and social quality of life in diabetic patients.
P12.
The best outcomes for patients with venous leg ulcers require an accurate diagnosis. Neglecting lymphedema results in higher recidivism. To maximize a national effort to improve outcomes, the existing gaps between Vein and Wound centers must be bridged to enhance educational endeavors and integrate Societal guidelines. Two potential databases are the US Wound Registry and the American Vein & Lymphatic Society (AVLS) Patient Reported Outcome (PRO) Venous Registry data bases.
Data base integration requires a common shared clinical definition of VLUs, and accurate consistent identification and documentation of veins treated, excluding the presence of tendon or bone in the wound bed, atypical ulcers and organ dysfunction. A true VLU is typically epidermal and dermal loss only. Venous reflux ultrasound identifies axial and/or deep venous reflux. Recent published AVLS PRO data demonstrated 85% of leg wounds are venous in origin and 97% possess surgically correctable disease in 1,794 unique subjects (1,878 limbs). Significant gaps in AVLS PRO data are documentation of mixed ulcers, diagnosis of diabetes, and accurate identification of atypical ulcers. These gaps are potentially bridged by the Wound registry data.
The lymphedema of VLUs remains vastly underrecognized and undertreated. The US Wound Registry identified 29,298 patients with venous stasis or venous insufficiency as of October 2015, with 4,029 simultaneously diagnosed with lymphedema. The integrated data of the US Wound Registry and AVLS PRO databases could vastly improve the appropriate diagnosis lymphedema as a critical component of “phlebolymphedema?€?, creating new measures of treatment documentation and outcomes determination. Lymphedema education, therapeutic feedback and documented potential patient quality of life outcomes with respect to VLU therapies could have vast health care economic implications to strengthen reimbursement coverage. Successful lymphedema treatment may benefit from a Certified Lymphedema Therapist, a persistent relative novelty for many vein and wound centers. Additional research should include micronutrients for endothelial function, Micronized purified flavanoid fractions, endothelial glycocalyx restorative interventions, single nucleotide polymorphisms.
Conclusions;
Vein and wound center patient treatment goals are aligned; improve patient outcomes and quality of life. Significant gaps are present in accurate clinical diagnos, recognition and lymphedema treatment with integrated CLT management. The US Wound Registry and AVLS PRO databases are both nationally recognized resources aligned with Societal guidelines. If we are to bridge the gap to new levels of holistic patient care metrics, we must bridge the datasets. The opportunity to improve patient cares for those suffering with VLUs and lymphedema is at hand with venous, lymphatic, wound and CLT leadership.
P13.
Introduction: Traditional management of Pyoderma gangrenosum consists of pharmaceutical and local wound care practices. In this case series we explore a treatment algorithm of pharmaceutical regimen preoperatively (standard of care and micronutrients), followed by full pyoderma wound excisional surgical debridement, pH controlled hypochlorous acid wound cleansing using negative pressure wound therapy and the sequential use of an ovine forestomach matrix graft. We hypothesize that by addressing wound pH imbalance, lack of structural extracellular matrix and management of periwound dermal lymphatic stasis with negative pressure wound therapy, wound resolution can occur without resulting pathergy.
Methods: 3 patients were evaluated by dermatology and standard of care therapeutics initiated 4-12 weeks prior to surgical debridement. Pyoderma wounds underwent surgical excisional sharp and ultrasonic debridement. Negative pressure wound therapy with hypochlorous acid instillation was applied, changed every 3-4 days. With adequate granulation tissue, ovine graft was placed, a 2-layer compression wrap applied, changed weekly. When the graft was fully incorporated, split thickness skin grafting, application of ovine graft to the donor site or ovine graft was utilized to continue epithelialization. The primary endpoint was worsening of wound size after surgical debridement. The secondary endpoints included time to 100% granulation of the graft and time to complete epithelialization.
Results: 3 patients underwent this treatment algorithm; there was no worsening or increase in the size of the wound. The mean time to 100% granulation was 2 weeks. One patient had a STSG at week 2 and had 50% graft take. Repeat wound biopsies in 2 patients demonstrated no neutrophilic infiltration. One patient healed by secondary intention by week 19 and the third patient continues in wound care with progressive epithelialization.
Discussion: The addition of excisional surgical debridement/resection, pH controlled hypochlorous acid, negative pressure wound therapy and ovine foregut matrix to the management of pyoderma wounds appears to be a safe and effective method. This treatment algorithm is counterintuitive to the current standard surgical practice of pyoderma due to relative concerns of pathergy development. Although this is a small sample size the results are promising and warrant a larger sample size with additional performance of interval biopsies and biochemical data to determine cellular response.
P14.
Purpose
Several critical factors are thought to contribute to the chronic wound milieu, including biofilm, bacterial colonization, and elevated wound pH. Chronic wounds have a pH ranging from 7-9, as compared to the normal skin “acid mantle?€? pH of 5.5. Targeting elevated wound pH, bacterial colonization, and biofilm while maintaining a moist wound environment and avoiding cytotoxic cleansing agents are key components of an effective wound management strategy. To maintain a moist wound environment, dressings typically must be changed twice per day. Pure Hypochlorous Acid has been previously demonstrated to have excellent antimicrobial preservative properties, a pH of 5.5, low cytotoxic potential, and effective biofilm management properties and cost effectiveness.
Methods/Results
This retrospective case series evaluated the efficacy of moist-to-moist, twice daily pure hypochlorous acid (pHA) dressings for six patients with complex chronic postsurgical wounds. The wound etiologies included 3 patients with exposed orthopedic hardware in the leg/ankle, 1 patient on a cardiac transplant list with a wound at a Left Ventricular Assist Device driveline entry site, 1 patient with dehisced abdominal incisions from a combined liver-kidney transplant, and 1 patient with a dehisced cervical spine wound from malignancy resection involving a vertebral body.
In this series of 6 patients, moist-to-moist pHA soaked gauze dressings were initiated at initial consult and continued through last follow up (3 patients) and wound closure (3 patients). Both in-person and virtual visits were utilized for follow up. All patients were advised to use adjunctive micronutrients and were provided with protein intake recommendations. For the 3 patients with leg/ankle wounds with exposed orthopedic hardware, limb edema/lymphedema was managed with dynamic dermal and static compression. Wound size was documented, and debridement was performed as indicated. No adjunctive advanced dressings were utilized.
Three patients had closed wounds within 3-6 visits, over the course of 10-25 weeks. Three patients had 2-4 visits with significant progression to closure; of these, one had improved 94% in 4 weeks, another improved 50% in 11 weeks, and the third improved 9% in 4 weeks. For the patient with just 9% improvement since treatment initiation, her wound bed appearance markedly improved, becoming 100% granular. Follow up is ongoing. No patient experienced adverse events.
Conclusions
This case series demonstrates that moist-to-moist pure Hypochlorous Acid cleanser soaked dressing changes can be effective in a variety of chronic complex postsurgical wounds, including those with exposed orthopedic hardware, bone, LVAD driveline sites, and post-operative sites in immunosuppressed patients. This dressing change regimen is effective, simplified, and cost-efficient.
P15.
P16.
Background
A diabetic foot ulcer is complicated and difficult to treat. The vascular supply is an important factor in wound healing and many diabetic patients have advanced atherosclerosis in the foot. To assess arterial steno-occlusion, computed tomography angiography and conventional angiography have been used and these modalities are expensive and the results may be delayed by the need for a trained clinician reader. If the patient has an ulcer on the foot, it is easy for the patient to agree to proceed with these diagnostic examinations. However, if there is no ulcer on the foot of the patient, the patient may hesitate to proceed with these procedures. Therefore, we assessed the measurement of foot skin temperature as a screening tool for evaluating lower extremity circulation to do more detailed investigations.
Methods
This retrospective study reviewed the medical records of forty-eight diabetic patients who did no ulcers on their feet between December 2018 and January 2021. They were following up at the department of endocrinology and referred me because of neurologic symptoms such as tingling sensation, pain, numbness, and so on. The foot skin temperature was measured with a non-contact infrared thermometer on the dorsum and planta of both feet and the lowest temperature was recorded for statistical analysis. Receiver operating characteristic (ROC) curve analysis was used to evaluate this measurement method as a diagnostic tool.
Results
A total of 76 feet were analyzed. Through the backtracking process, we could found that the optimal threshold value for the prediction of arterial steno-occlusion in the ipsilateral lower extremity was 35.5 Celcius degree. The area under the ROC curve was 0.76. With sensitivity set at 73.1%, the specificity was 75.0%.
Conclusions
Measuring foot skin temperature is a predictive and cost-effective method for assessment of the vascular supply and need for more study and can be used not only to determine whether percutaneous transluminal angioplasty (PTA) is indicated but also to monitor the result of PTA for early detection of steno-occlusion, even in the absence of a wound. Moreover, the technique is easily performed by non-professionals and can be useful in patient education.
P17.
Advancements in pathology have given rise to software applications intending to minimize human error and improve efficacy of image analysis. Still, subjectivity and nonsignificant error rate limitations of the most ubiquitous tissue stain analysis software reveal the need for a highly accurate, automated nuclear quantification software specific to immunohistochemistry with improved precision and efficiency. A new method for the quantification of immunohistochemical biomarkers in keratinocyte nuclei is proposed to overcome these limitations, contributing sensitive shape-focused segmentation, accurate nuclear detection, and automated device-independent color assessment.
P18.
Background:
Patients with stage 4 pelvic pressure injuries that have large, undermined cavities are at high risk for treatment failure and often fall into the category of palliative care. The essential deep lesion has been described as a subdermal muscle infarction, as the deep muscle surface has a rich vascular supply to the dermis that is quite vulnerable to pressure damage. The overlying skin is not as vulnerable and can appear normal. There are few studies that discuss treatment of the broad area of undermining that may occur with extended injury.This case assessed jet lavage irrigation in the outpatient setting using an irrigator accessory with a long narrow tube-tip that allows irrigation of the undermined cavity.
Methods:
An investigational review board study (IRB #6066, Sterling IRB, Atlanta, GA) was established to assess closed pulsatile irrigation in the long-term care setting. From an intention to treat consecutive series of 30 stage 4 pelvic pressure injuries, five patients were selected where the undermining could account for at least 50% of the wound dimension. These patients were all high risk and were considered palliative care by the primary care physicians and nursing home administration. Treatments were done at the bedside and three liters of saline were irrigated with a frequency of three to five days per week. Average age was 78, (range: 69 to 96) with periods of treatment ranging from 10 to 238 weeks.The basic protocol is irrigation of the wound with a pulsatile irrigator that offers a low pressure at 8 to 12 psi and typically uses 3 liters of normal saline. The undermined area and tunnels were carefully irrigated using the tunnel tip until fluid was clear with no remaining debris.
Results: Significant reduction of the undermined cavity was seen in all cases within the first three weeks. No patient developed wound sepsis, but bacterial contamination could not be determined without use of auto-fluorescense digital imaging (MolecuLight). Undermining resolution occurred in four patients and one patient with improving wound died of Covid.
Conclusion: This series highlights the ability of appropriate mechanical debridement and irrigation to quickly resolve the undermining and tunnelling in severe chronic wounds. Positive and notably progressive healing was seen in all five cases. The femoral canal irrigation tip was the critical tool and other standard surgical devices such as the Yankar sucker tip are probably as effective. With the potential for underlying structures to be involved such as bone, tendons, nerves, and vessels, there is no place for topicals and dressings as the primary treatment consideration. Modern surgical literature strongly supports this approach as the standard of care using irrigation and debridement for all undermined, contaminated chronic stage 4 pelvic pressure injuries.
P19.
Background: Antiseptics are an essential component of wound care for controlling the biofilm forming bacteria that cause infection. Assessment of efficacy of wound surface treatment has improved dramatically with the introduction of autofluorescence imaging (AFI). We are now able to measure with a high level of sensitivity, the bacterial contamination levels relevant to our practice. This pre-hypothesis report presents several experiments done in the clinical setting to assess antiseptic use.
From an investigational review board study (IRB# 6066, Sterling IRB, Atlanta, GA) a series of 30 patients were assessed using several FDA approved methods and products for early wound bed preparation in chronic wound care. These treatments included daily low-pressure jet lavage (8-12 PSI) with 3 liters of normal saline, the use of antiseptics on the wounds, and combination of antiseptics irrigated with the jet lavage irrigators. One patient was remarkable with a difficult wound that had been recalcitrant to treatments. There were three stage 4 pelvic pressure injuries including greater trochanter, ischium and sacrum. Bacteria on multiple PCR tests included Psuedomonas, Acinetobacter, and Group D Streptococcus. A 3-day delay yielded a new overgrowth of bacteria estimated at >log 4 colony forming units per gram of tissue assessed with autofluorescence imaging (MolecuLight Inc. Toronto, CA). A binary treatment of 50% AFI improvement was considered the threshold for a positive result.
Results: Experiment 1: Treatment with jet lavage using saline as the irrigant after the wound had 3 days of treatment with only daily saline-soaked gauze dressings, Friday to Monday, revealed greater than 80% bacterial load removal in all tests (n=13). Experiment 2: jet lavage of 3 liters saline followed by 250 ccs modified sodium hypochlorite through the irrigator revealed >90% clearance of bacteria (n-23). Experiment 3: jet lavage irrigation followed by 250 ccs modified sodium hypochlorite, betaine/polyhexanide, 0.018% sodium hypochlorite, and hypochlorous acid showing >90% clearance of bacteria (n-12) with no detectable difference between antiseptics. Experiment 4: Spray bottle 250 ccs irrigation with 2-minute dwelling of soaked 4×4 sponges with each noted antiseptic followed by jet lavage irrigation 3 liters demonstrating minimal removal (0-25% effect of antiseptic irrigator/dwelling) versus >90% effect of the jet lavage irrigation that followed. Regression of each test is P=.001.
Conclusion: Jet lavage irrigation is optimized with irrigation of antiseptics with the irrigator. This boosts bacterial debridement to an acceptable level (>90%). Use of simple squeeze bottle irrigation (measured 1-2 PSI) yielded minimal (<25%) AFI bacterial clearance. These findings were compelling but further controlled clinical trials are warranted. We found no safety issues with these methods.
P20.
Background: Treatment of keloids is challenging. The aim is to present our experience using punch-assisted proliferating core excision followed by postoperative 11 Gy(single fraction) or 8Gy (two fractions, daily) radiotherapy for keloids within 24 hours after surgery.
Methods: 20 patients with keloids were treated with this protocol. The mean follow-up period was 6 months. The outcome was reported with a recurrence-free rate. We also reported side effects.
Results and Conclusion: Our preliminary results show positive outcomes of the current protocol. Our present study needs further long-term validation with more diverse patients.
P21.
Introduction:
For patients with full thickness complex wounds, autologous split-thickness skin grafts (STSG) are generally regarded as the mainstay of treatment. However, STSG’s are often ineffective by themselves in the treatment or closure of complex wounds when deeper anatomic structures such as bone or tendon are involved. In those situations, placement of “dermal analogues?€? to form a well-vascularized tissue layer on which to place a subsequent STSG are used. The time to incorporation of those analogues is often variable but measured in weeks if not months. In this study, we present the outcomes of Matriderm® graft usage on healing complex full-thickness wounds in a community-based burn and wound center in several patients, as well as observations on usage of the product to optimize success in the in- and outpatient arena to accelerate the time-to-healing of these complex wounds.
Methods:
A retrospective chart review of all patients who received the Matriderm® product in their wound healing algorithm, with photographic documentation was performed. We present pre-operative wound evaluation and measurements, complicating factors and comorbidities, as well as final closure and time-to-healing (>95% epithelialized) as data points.
Results:
Ongoing patient enrollment. Further information forthcoming as patient enrollment continues.
Discussion:
Matriderm® collagen-elastin dermal matrix is already proving to be a useful tool in the algorithm for complex full-thickness wound healing. Our experience with its usage has shown accelerated time to healing and reliable, sturdy wound closure. In our community-based burn and wound healing center it is considered extremely reliable, easy-to use and handle, and shortens time-to-healing in these complex wounds. Further study comparing it to previously treated patients with other dermal analogues will be forthcoming.
P22.
Background: Dupuytren’s contracture (DC) is a fibrotic disease which presents with thickening of the palmar fascia causing irreversible flexion of digits in its most aggressive form. Presence of the highly contractile alpha smooth muscle actin (aSMA) protein in myofibroblasts, combined with cell proliferation, is thought to contribute to this disease. Three observations suggest that DD fibroblasts could have short telomeres: the disease affects mostly older individuals; the tissue is highly proliferative; and recurring disease requires more proliferation. Cells with short telomeres in vitro are growth arrested and demonstrate phenotypes that resemble aging in vivo. Therefore our goal was to develop an in vitro model of replicative senescent DC cells to study myofibroblast behavior. Methods: To examine expression of aSMA, transforming growth factor beta (TGF- beta) was used to differentiate DC fibroblasts into myofibroblasts. A group of early or low population doubling (PD) (‘young’: yDC) and late or high PD cells (‘aged’: aDC) derived from 3 patient cultures were cultured using standard conditions in the presence or absence of TGF- beta. Western blotting was used to measure aSMA expression. Proliferating myofibroblasts were visualized using aSMA immunocytochemistry combined with click-staining of ethynyl deoxyuridine (EdU)-labeled nuclei counterstained with DAPI. ImageJ was used to analyze the western blot for relative density, and a two-way t test was performed. Results: aSMA expression was found to be higher in aDC cells treated with TGF- beta (n = 4, p = 1.56E-04), which was further supported by the fluorescent microscopy images. Collagen matrices populated with aDC cells contracted less than yDC; the difference was negated when contraction was plotted against cell number. Conclusions: in vitro aDC cells may not proliferate as well as yDC cells but can effectively function as myofibroblasts, correlating published data showing that certain aggressive DC is not correlated with high cellularity.
P23.
P24.
Background:
Four active young adults with Inflammatory Bowel Diseases (IBD) developed excruciatingly painful slough-filled lower leg pyoderma gangrenosum (PG) wounds. Two of the young people were adherent to aggressive treatment plans for their inflammatory illness, including high-dose steroids and antibiotics. The other two refused, believing these treatments would cause gut microbiome imbalances and make their underlying condition worse.
All four patients had to limit their usual productive activities (work or school) due to pain, medication side effects, and wound-related disability. When it became apparently that their health care providers were unfamiliar with effective wound management methods for PG, the patients and/or their families searched online for alternative treatments.
Pyoderma gangrenosum is a notoriously painful inflammatory wound often related to autoimmune disorders. Traditional debridement, including both manual and sharp, is contraindicated because any manipulation of the ulcers causes them to increase in size (pathergy). Polymeric membrane dressings* (PMDs) are an ideal choice for PG wounds for many reasons. PMDs address the underlying cause of the ulcers (inflammation) directly by subduing the nociceptor (pain-sensing nerves) response. PMDs’ components work synergistically with the body to gently continuously debride wounds, avoiding pathergy while eliminating barriers to healing. PMDs also promote brisk wound healing by concentrating nutrients in the wound bed and fostering an ideal moisture balance across the entire wound surface. PMDs do not adhere to the wound bed, avoiding this potential source of trauma. Finally, PMDs are recognized as a “pain relieving?€? dressing because of their effect upon the nociceptor system.
Methods:
Each of the patients decided (with their local physician’s consent) to try PMDs with email or smartphone guidance from the author. Patients/families performed all dressing changes themselves. No wound cleansing or manual debriding was ever required. In collaboration with their local physicians, all antibiotic and prescription pain medications were gradually eliminated.
Results:
All of the wounds were already markedly cleaner and most were measurably smaller by day four. Pain medication use rapidly decreased and activity rapidly increased. Three of the four wounds fully closed in 3½ months using PMDs as the only topical wound treatment. The fourth patient became unable to obtain PMDs; her wound remained open.
Conclusion:
All of the patients’ goals were met or exceeded when PMDs were used as directed. Persistent use of PMDs resulted in decreased inflammation, excellent pain relief, brisk atraumatic slough removal, increased quality of life, and steady healing to complete closure of three very challenging PG wounds managed with PMDs. When PMDs were halted prematurely (the fourth patient), the PG wound stopped closing.
P25.
Impaired wound healing, often the result of chronic metabolic diseases or infection, is characterized by the dysregulation of immune responses in tissue repair. In the wound bed, EVs have been reported to promote paracrine signaling, immune cell crosstalk and re-epithelialization, but the mechanisms by which they accomplish these essential functions has been obscured by the heterogeneity of EVs produced and their diverse payloads, as well as a lack of clarity on their biologic activity in vivo. Therefore, we have focused on three major areas of unmet need in the study of EVs in wound healing. First, we have addressed the challenge of EV heterogeneity by deploying single vesicle flow cytometry (vFC) for the quantitative analysis of individual EVs, which helps understand subpopulations of EVs at various stages of wound healing. This work led to the discovery that CD44 was down-regulated on the surface of EVs from diabetic obese mice. Second, we used -omic technologies such as mass spectrometry and RNA sequencing to identify specific Serpin family serine proteases and regulatory miRNA payloads that were dysregulated in diabetic obese mice. Third, we assessed the biological activity of EVs based on their ability accelerate tissue repair in animal models of impaired wound healing. This work has led to the development of engineered EVs that deliver specific tagged payloads to monitor uptake and promote wound closure in diabetic mice.
P26.
A novel peel and place negative pressure wound therapy (NPWT)* dressing† has been created that effectively promotes granulation tissue formation and re-epithelialization while substantially decreasing tissue ingrowth when the dressing is worn for 35 days with weekly dressing changes. The peel and place dressing was compared to reticulated open cell foam (ROCF) ^plus an interface layer (IFL).
All animal work was approved by the relevant Institutional Animal Care and Use Committee and complied with all applicable national and local regulations. The study design required that an IFL be used with ROCF treatments to limit tissue ingrowth allowing for 7 day dressing changes. Peel and place and ROCF+IFL dressings were applied to deep, excisional wounds (extending 2 cm into the muscle) created paraspinally in 2 swine. Peel and place treated wounds received continuous NPWT at -125 mmHg for 35 days. ROCF +IFL treated wounds received NPWT for 28 days followed by IFL alone until study term. Dressing changes were performed every 7 days. 2D and 3D wound images were acquired at each observation day. 2D color thresholding analysis enabled quantification of fibrinous material observed on wound bed surfaces. 3D imaging was used to record wound area and volume. Additionally, wound sections were collected at term for histologic and morphometric analysis including granulation tissue thickness, percent re-epithelialization, and prevalence of fibrinous material on the wound bed.
Fibrinous material was observed on both peel and place and ROCF+IFL treated wound beds at early observation days but showed considerable reduction, without intentional debridement, by day 35. Thresholding analysis of 2D color images showed a 62.3% average fibrinous material coverage in the peel and place treatments at day 7 that decreased to 10.7% at day 35. Morphometric analysis confirmed limited serocellular debris coverage at day35 with a mean area of 3.9%.
The fibrinous serocellular material did not seem to negatively impact wound healing. Image analysis showed a rapid reduction of peel and place treated wound area from 22 cm2 on Day 0 to 0.1 cm2 by day 35. 3D image analysis for the peel and place dressing showed a wound volume fill of 87.6% after 7 days, and 98.9% fill by day 35.
Histomorphometry assessments showed 100% re-epithelialization. A veterinary pathologist reported granulation tissue to be of good quality as evidenced by collagen maturity and vascularization.
Results of this study illustrate that the novel extended wear, peel and place, NPWT dressing, applied to deep excision wounds, may be worn for 35 days without negatively impacting wound healing. Furthermore, the peel and place dressing can promote quality granulation tissue formation, re-epithelialization, and wound closure.
*3M™ V.A.C.® Therapy; † Novel Peel and Place Dressing (3M, San Antonio, TX); ^3M™ V.A.C.® Granufoam™ Dressing
P27.
Background: Pressure ulcers (PU) are chronic wounds associated with significant morbidity. Limited therapeutics are available for prevention and treatment of PU. Silk fibroin is a biocompatible polymer that can be fabricated into nanostructures, termed nanosilk. Nanosilk is characterized by a high strength-to-density ratio and strengthens the biomechanical properties of skin. We have previously shown that application of an 8% nanosilk spray reduces the incidence of PU development. Here, we hypothesize that a single application of a 4% nanosilk cream would have improved delivery and therefore efficacy at preventing PU progression by protecting the skin against extracellular matrix degradation and decreasing inflammation.
Methods: 12-13 week C57BL/6 mice underwent implantation of a 6.35mm diameter steel disc beneath the external oblique muscle latero-caudally (n=41). After approximately two weeks to allow for complete healing, a disc magnet (5 mm diameter, 2 mm thick) is applied to the skin superficial to the implanted disk for 10, three-hour cycles consisting of two hours with the magnet applied and one hour of rest with the magnet off. Photos were taken at the conclusion of each cycle for staging. PU development was evaluated using the established National Pressure Ulcer Staging System with grades from 0 to 4. At the conclusion of 10 cycles, wound tissue was collected and processed for histologic analysis with tissue sections stained for CD45, a pan-leukocyte marker, and trichrome, a stain for collagen. 10 random high-powered fields (HPF, 20x) were imaged and analyzed using NIS Elements – Advanced Research imaging software to quantify the number of CD45+ cells and the area of collagen deposition. Four study groups were included: a non-treated control group, one application (1x) of control pluronic gel (PLG), 1x Cavilon® spray, and 1x 4% nanosilk cream (pluronic gel base). Each treatment was applied prior to cycle one. Statistical significance was determined by one-way ANOVA (p<0.05 significant).
Results: A single application of 4% nanosilk cream significantly decreased the PU stage starting at cycle six compared to the untreated control (p=0.02), PLG control (p=0.04), and 1x Cavilon® spray (p=0.01). Additionally, 4% nanosilk significantly lowered inflammatory cell infiltrate (p=0.03) and increased collagen deposition (p=0.04) in PU by the end of 10 cycles.
Conclusion: A 4% nanosilk cream was more effective at preventing high stage PU, decreasing inflammation, and increasing collagen deposition than both a control gel and Cavilon® spray. Topical nanosilk cream shows significant promise as a potential therapeutic for prevention of PU.
P28.
Background: Normal wound healing is a highly complex biological process sensitive to several molecular regulatory sequences and cell processes. Dysregulation of associated gene expression can lead to compromised wound healing. In recent research, non-coding microRNA(miR)-based therapy has been shown to produce remarkable therapeutic efficacy. In our previous studies, it was demonstrated that expression of miR146a is significantly downregulated in diabetic wounds. We have also shown that delivery of exogeneous miR-146a by conjugation to octahedral cerium oxide nanoparticles (CNPs) significantly improved healing and cell health through regulation of proinflammatory genes expression and scavenging of ROS. In this study, we propose that CNP morphology plays an important role in effective miR146a delivery to cells and inflammation reduction via shape-dependent material properties and interaction with cell structures.
Methods: A short organic linker was used to conjugate miR146a to CNPs. CNPs were chosen due to their unique ROS-scavenging activity in addition to their ability to carry and deliver miRs. CNPs with different morphologies (octahedral, rod, and cube), synthesized using a single step method, were used to study potential shape-dependent miR146a delivery performance. As Ce3+/Ce4+ is a key factor for antioxidant property, differences in redox state ratio between the various shape CNPs were analyzed using X-ray photoelectron spectroscopy (XPS). The corresponding change in physicochemical properties and antioxidant activities were studied by several materials characterization techniques and SOD enzyme mimetic activity assay. Cytotoxicity and miR146a gene target expression were characterized by cell viability assay and PCR, respectively, for each particle morphology as measures of potential wound healing efficacy.
Results: SOD activities of octahedral, rod, cube CNP were measured to be 98.2, 79.2 and 12.9%. These results appear to partially correlate with the trend observed for Ce3+/Ce4+ ratio from XPS analysis (octahedral (26.8%), cube (16.1%), rod (66.6%)). No cytotoxicity was observed for conjugated samples analyzed using a macrophage cell line (RAW 264.7). Additionally, it was seen that rod shape CNPs were the most effective in miR delivery over other tested shapes (cube and octahedra) by a substantial margin, in a per particles basis. Measurement of IL-6 gene expression was determined to decrease significantly for octahedra CNP-miR146a treated cells.
Conclusion: Our results show that the shape of redox-active nanovector cerium oxide plays a major role in delivery efficacy for particle conjugated miR146a and in their ability to reduce inflammation, with octahedral shape being the most effective.
P29.
Background: Autologous nerve grafting has been applied as the best method of treating peripheral nerve defects but it has problems such as donor site morbidity. If the defect is more than 2 cm, direct suture is difficult and the nerve conduit is used for nerve regeneration. In addition, nerve repair by suture can damage nerve tissue. Therefore, we would like to repair nerve using hydrogel of mussel adhesive protein. We use mussel adhesive protein to induce nerve regeneration without suture of nerve injury site and compare with conventional nerve repair by suture.
Methods: We made a 10mm long defect in the sciatic nerve of 46 rats and divided to subgroups. 1) Normal (positive control), 2)Sham (negative control), 3)Direct anastomosis –suture, 4)Direct anastomosis by fibrin glue, 5)Direct anastomosis by Mussel adhesive protein(MAP), 6)Direct anastomosis by MAP with Substance P, 7)15 mm gap model with silicon tube by suture, 8)15 mm gap model with silicon tube by fibrin glue, 9)15 mm gap model with silicon tube by MAP 10) 15 mm gap model with silicon tube by MAP with substance P. After 6 weeks, nerve conduction velocity(NCV) and histological observations were made.
Results: In vivo results revealed that the NCV was improved more in the experimental groups after 6 weeks from nerve repairing by MAP than in the control group by suture. Histologic(H&E, Toluidine blue staining) and immunohistochemistry(Nestin, MAP−2, GFAP) staining showed more regenerated nerve findings.
Conclusions: In this study, we propose nerve repair by MAP because of shortening operation time and additional nerve damage by suture, Additionally Mussel adhesive protein with substance P propose to be a good method of promoting nerve regeneration.
P30.
Stem cells-based treatment for burn wounds require frozen cells as an off-the-shelf therapy; however, cryopreservation-induced oxidative stress resulted in post-thaw cell death or loss of cell functions, thus arrested their clinical practicality. Although antioxidant priming to stem cells increase their resistant to oxidative stress, but this strategy is still unexplored on cryopreserved cells. Herein, we investigated whether curcumin priming before cryopreservation could preserve the therapeutic potency of thawed stem cells. For this, unprimed and curcumin-primed adipose-derived stem cells (ASCs) were cryopreserved for one month. Post-thawing, cells were assessed for viability by trypan blue assay; metabolic activity by MTT assay; senescence by senescence associated (SA) β galactosidase activity staining assay; migration by scratch healing assay and; mRNA expression by real-time PCR. Subsequently, the healing potential was examined by injecting cells around the wound periphery of acidic burn in rats. Post-healing, skin architecture was histologically examined by hematoxylin & eosin, masson-trichrome and picrosirius red staining’s. Results demonstated that curcumin-primed frozen cells (Cryo/Cur-ASCs) showed better post-thaw viability, metabolic activity and migration ability comparative to unprimed frozen cells (Cryo/ASCs). Curcumin priming alleviated the oxidative damage by activating the ROS-reducing cellular antioxidant system as shown by the evident increase in GSH levels and upregulated mRNA expression of glutathione peroxidase (GPx), superoxide dismutases (SOD1, SOD2), and catalase (CAT). Further, in vivo findings revealed that Cryo/Cur-ASCs-treated wounds exhibited earlier wound closure with an improved architecture (thick epidermis, mature granulation, dense and well-arranged collagen fibers) comparative to Cryo/ASCs and depicted healing capacity almost similar to Fresh/ASCs. In conclusion, our findings suggested that curcumin priming could be effective to alleviate the cryo-induced oxidative stress in post-thawed cells that bring forth the sustained potential of cryopreserved stem cells for burn wounds management.
P31.
Adipose-derived stem cells (ASCs) are a new hope in stem cell-based therapeutics to improve impaired wound healing in diabetic patients, owing to their in situ differentiation, paracrine-release functions, and suitability for autologous use. However, factors like hypoxic conditions in the diabetic wound hamper the ASCs’ therapeutic functions. In this study, we hypothesized that caffeic acid priming before transplantation could promote ASCs’ endurance in a diabetic wound microenvironment. To test our hypothesis, caffeic acid-primed ASCs were incubated in cobalt chloride (CoCl2)-induced hypoxic conditions and assessed in vitro for viability using MTT, senescence using β-galactosidase staining, apoptosis using annexin V staining, and migration using scratch healing assays. Following that, caffeic acid-primed ASCs suspended in platelet rich plasma (PRP) were injected intradermally around a circular excisional wound on the dorsum of a diabetic rat in order to test the healing effectiveness. On completion of wound healing, granulated skin was collected for histopathological analysis. In vitro results revealed that caffeic acid-primed ASCs preserved their cellular shape, proliferation capacity, metabolic state, and migration potential much better than unprimed ASCs in CoCl2-induced hypoxic culture conditions. When co-injected with PRP in an excisionally wounded diabetic rat model, in vivo data showed that caffeic acid-primed ASCs led to faster wound closure, greater neovascularization, and a more compact extracellular matrix compared to unprimed ASCs. In conclusion, this study proposed combinatorial therapy (caffeic acid-primed ASCs + PRP) as an effective stem cell-based modality for the repair of diabetic wounds.
P32.
BACKGROUND: Wound healing in obesity is still unresolved for its indistinct mechanism. Epidermal stem cells (EpiSCs) and adipocytes are considerable elements in skin structure, metabolism and healing, and compulsive tissue hypoxia has been demonstrated in chronic wounds. As HIF-1alpha is a critical transcriptional factor effecting in hypoxia-mediated metabolic changes, adipogenesis and differentiation, we hypothesize that EpiSCs may interact with adipocytes through HIF-1alpha in obesity wound.
METHODS: A model of high fat diet-induced obesity using Sprague-Dawley rats was established, including groups of sham-injury (Sham), injury without cell transplantation (Injury), and injury plus EpiSCs transplantation. A 6-mm diameter full-thickness excision was developed on the dorsal skin of rats. 1×10^5 epidermal basal stem cells isolated from neonatal mice skin and suspended in 30 μl PBS were injected subcutaneously into each wound in EpiSCs group, as equivalent PBS was injected similarly in Sham and Injury groups. Wound samples were harvested and subjected to histological investigations.
RESULTS: Improved histological healing and recruited PerilipinA(+) adipocytes were found in EpiSCs group at 1, 3, 7 and 14 days post injury. Subcutaneous adipose HIF-1alpha expression determined by immunohistochemistry in EpiSCs group was increased at 1 day (P<0.05, vs Sham or Injury), and it remained no significant difference at 3, 7 and 14 days (all P>0.05, vs Sham). Subcutaneous adipose HIF-1alpha expression in Injury group was increased at 7 day (P<0.05, vs Sham or EpiSCs) and 14 days (P<0.05, vs Sham).
CONCLUSIONS: EpiSCs may resume HIF-1alpha-delayed subcutaneous adipocyte differentiation to accelerate healing in obesity.
P33.
P34.
Macrophages are critically involved in regeneration and consecutive phases of wound healing. The dynamic plasticity of macrophages mediates complex function that leads to both tissue-destructive and -reparative outcomes during the inflammatory phase of wound healing. While the emergence of endogenous electric fields (EFs) after wounding has been reported for decades, the role of EFs in macrophage migration and polarization during wound recovery remains unknown. Here, we report the directional guidance effect of EFs on macrophage migration in vitro and an efficient electrical bandage for promoting macrophage polarization and accelerating skin wound healing in vivo. Mouse bone marrow-derived macrophages were cultured in vitro and exposed to EFs of physiological strength. M0 macrophages responded quickly to the applied EF and migrated directionally to the anode. A wearable printed circuit board (PCB)-based electrical bandage with program-controlled delivery was then designed for applying EF to the full-thickness skin wounds in wildtype mice. After a daily, 4-hour EF stimulation at 100mV/mm for 3 days, the wound tissue was fixed and sectioned for Hematoxylin and Eosin staining to evaluate wound closure. Wound re-epithelialization rates were compared to demonstrate a trend of accelerated wound healing from the control wounds at 21.6% (n=22) to the EF treated wounds at 26.8% (n=12, p=0.09) at the early stage of healing. Mouse macrophages were further stained with specific markers to evaluate the impact of long-term EF stimulation on macrophage subpopulations. Our result reveals a shift in macrophage subpopulation in wounds following EFs treatment. The ratio of M1/M2 cells decreased after 3 days of treatment by 30.60%, which indicates that the EF may enhance wound healing by shortening the inflammatory phase after injury. In summary, we describe an electrical method for the manipulation of macrophage migration, phenotype, and functions. A better understanding of the plasticity of wound macrophages in response to EFs would shed light on the therapeutic potential of manipulating macrophage function for optimal wound healing.
