Ablating Myocardium Using Nanosecond Pulsed Electric Fields: Preclinical Assessment of Feasibility, Safety, and Durability.

BACKGROUND: Unlike conventional microsecond pulsed electrical fields that primarily target the cell membranes, nanosecond pulses are thought to primarily electroporate intracellular organelles. We conducted a comprehensive preclinical assessment of catheter-based endocardial nanosecond pulsed field ablation in swine. METHODS: A novel endocardial nanosecond pulsed field ablation system was evaluated in a total of 25 swine. Using either a low-dose (5-second duration) or high-dose (15-second duration) strategy, thoracic veins and discrete atrial and ventricular sites were ablated. Swine survived for <1 (n=1), ≈2 (n=7), ≈7 (n=6), 14 (n=2), or ≈28 (n=9) days, and venous isolation was assessed before euthanize. Safety assessments included evaluation of esophageal effects, phrenic nerve function, and changes in venous caliber. All tissues were subject to careful gross pathological and histopathologic examination. RESULTS: All (100%) veins (13 low-dose, 34 high-dose) were acutely isolated, and all reassessed veins (6 low-dose, 15 high-dose) were durably isolated. All examined vein lesions (10 low-dose, 22 high-dose) were transmural. Vein diameters (n=15) were not significantly changed. Of the animals assessed for phrenic palsy (n=9), 3 (33%) demonstrated only transient palsy. There were no differences between dosing strategies. Thirteen mitral isthmus lesions were analyzed, and all 13 (100%) were transmural (depth, 6.4±0.4 mm). Ventricular lesions were 14.7±4.5 mm wide and 7.1±1.3 mm deep, with high-dose lesions deeper than low-dose (7.9±1.2 versus 6.2±0.8 mm; P=0.007). The esophagus revealed nontransmural adventitial surface lesions in 5 of 5 (100%) animals euthanized early (2 days) post-ablation. In the 10 animals euthanized later (14-28 days), all animals demonstrated significant esophageal healing-8 with complete resolution, and 2 with only trace fibrosis. CONCLUSIONS: A novel, endocardial nanosecond pulsed field ablation system provides acute and durable venous isolation and linear lesions. Transient phrenic injury and nontransmural esophageal lesions can occur with worst-case assessments suggesting limits to pulsed field ablation tissue selectivity and the need for dedicated assessments during clinical studies.

Hypertrophic Granulation Wounds Treated With Silver Nitrate Sticks or With Topical Steroid: Rate of Wound Closure.

Hypertrophic granulation (HG) is abnormal granulation tissue raised above the level of surrounding intact skin and is thought to delay wound healing. Effective treatment to eliminate HG could speed healing, but this is not well studied. Two common treatments are chemical cautery with silver nitrate, and the use of topical steroids. In a Midwestern burn and wound clinic, both of these treatments are employed. A quality improvement project compared wound size reduction for HG wounds undergoing the two treatment modalities. Retrospective chart review identified HG wounds treated during a 1-year period, and compared wound size prior to and after 1 month of treatment (length, width, and surface area). Results were presented to the clinic staff, and 1 year later the quality assurance project was repeated. The initial audit found data on 18 patients treated with silver nitrate cautery, and 16 patients treated with 1% hydrocortisone cream. Median length and width were decreased by 5 mm with silver nitrate cautery treatment, and by 14 mm with 1% hydrocortisone treatment (P < .05). The repeat audit 1 year later found 10 patients treated with silver nitrate, and 38 treated with hydrocortisone. Median length and width decreased by 0 and 2 mm respectively for silver nitrate cautery treatment, and 15 and 10 mm for 1% hydrocortisone treatment (P < .05). In conclusion, this uncontrolled review suggests faster healing in HG wounds treated with 1% hydrocortisone compared with those treated with silver nitrate cautery. Confirmation with controlled and randomized studies is warranted.

Evaluating Real-World National and Regional Trends in Definitive Closure in U.S. Burn Care: A Survey of U.S. Burn Centers.

To better understand trends in burn treatment patterns related to definitive closure, this study sought to benchmark real-world survey data with national data contained within the National Burn Repository version 8.0 (NBR v8.0) across key burn center practice patterns, resource utilization, and clinical outcomes. A survey, administered to a representative sample of U.S. burn surgeons, collected information across several domains: burn center characteristics, patient characteristics including number of patients and burn size and depth, aggregate number of procedures, resource use such as autograft procedure time and dressing changes, and costs. Survey findings were aggregated by key outcomes (number of procedures, costs) nationally and regionally. Aggregated burn center data were also compared to the NBR to identify trends relative to current treatment patterns. Benchmarking survey results against the NBR v8.0 demonstrated shifts in burn center patient mix, with more severe cases being seen in the inpatient setting and less severe burns moving to the outpatient setting. An overall reduction in the number of autograft procedures was observed compared to NBR v8.0, and time efficiencies improved as the intervention time per TBSA decreases as TBSA increases. Both nationally and regionally, an increase in costs was observed. The results suggest resource use estimates from NBR v8.0 may be higher than current practices, thus highlighting the importance of improved and timely NBR reporting and further research on burn center standard of care practices. This study demonstrates significant variations in burn center characteristics, practice patterns, and resource utilization, thus increasing our understanding of burn center operations and behavior.

A phase 3, open-label, controlled, randomized, multicenter trial evaluating the efficacy and safety of StrataGraft® construct in patients with deep partial-thickness thermal burns.

OBJECTIVE: This phase 3 study evaluated StrataGraft construct as a donor-site sparing alternative to autograft in patients with deep partial-thickness (DPT) burns. METHODS: Patients aged ≥18 years with 3-49% total body surface area (TBSA) thermal burns were enrolled. In each patient, 2 DPT areas (≤2000cm2 total) of comparable depth after excision were randomized to either cryopreserved StrataGraft or autograft. Coprimary endpoints were: the difference in percent area of StrataGraft treatment site and autograft treatment site autografted at Month 3 (M3), and the proportion of patients achieving durable wound closure of the StrataGraft site without autograft at M3. Safety assessments were performed in all patients. Efficacy and safety follow-up continued to 1 year. RESULTS: Seventy-one patients were enrolled. By M3, there was a 96% reduction in mean percent area of StrataGraft treatment sites that required autografting, compared with autograft treatment sites (4.3% vs 102.1%, respectively; P<.0001). StrataGraft treatment resulted in durable wound closure at M3 without autografting in 92% (95% CI: 85.6, 98.8; n/n 59/64) of patients for whom data were available. The most common StrataGraft-related adverse event was pruritus (15%). CONCLUSIONS: Both coprimary endpoints were achieved. StrataGraft may offer a new treatment for DPT burns to reduce the need for autografting. CLINICAL TRIAL IDENTIFIER: NCT03005106.

Complex Multidisciplinary Cranial and Scalp Reconstruction for Patient Salvage.

OBJECTIVE/BACKGROUND: Patients can develop scalp and cranial defects as a result of neoplasm, trauma, or infection. Reconstruction of these defects can be difficult in some patients and may require a multidisciplinary approach using creative solutions usually used for disease processes in other areas of the body, such as severe burns. METHODS: A series of 9 patients were treated using multidisciplinary techniques for reconstruction of complex cranial and scalp defects. Data on patient characteristics, initial treatment, and preparatory and definitive reconstructive treatment were retrospectively collected. Outcomes were measured as full solution, partial solution, or failure. RESULTS: Three patients had a full solution/wound closure, 5 had a partial solution, and 1 failed reconstructive attempt. Full solution patients tended to be younger, received reconstruction treatment modalities for longer periods of time, and had more definitive surgeries. Initial and preparatory surgeries did not tend to promote a full solution, though having fewer preparatory surgeries that were not related to wound vacuum-assisted closure use tended to be associated with a better outcome. Infection of the scalp or cranium did not tend to change the result. CONCLUSIONS: Reconstructive salvage of complex cranial and scalp defects takes time, so patience and earlier recognition of need for atypical reconstruction is beneficial. Patient characteristics may influence outcomes, but judicious choice of materials and techniques is more critical to patient success. Use of a multidisciplinary approach to complex cranial and scalp reconstruction is a beneficial option for many patients for whom standard reconstruction methods are not viable.

American Burn Association Guidelines on the Management of Acute Pain in the Adult Burn Patient: A Review of the Literature, a Compilation of Expert Opinion, and Next Steps.

The ABA pain guidelines were developed 14 years ago and have not been revised despite evolution in the practice of burn care. A sub-committee of the American Burn Association's Committee on the Organization and Delivery of Burn Care was created to revise the adult pain guidelines. A MEDLINE search of English-language publications from 1968 to 2018 was conducted using the keywords "burn pain," "treatment," and "assessment." Selected references were also used from the greater pain literature. Studies were graded by two members of the committee using Oxford Centre for Evidence-based Medicine-Levels of Evidence. We then met as a group to determine expert consensus on a variety of topics related to treating pain in burn patients. Finally, we assessed gaps in the current knowledge and determined research questions that would aid in providing better recommendations for optimal pain management of the burn patient. The literature search produced 189 papers, 95 were found to be relevant to the assessment and treatment of burn pain. From the greater pain literature 151 references were included, totaling 246 papers being analyzed. Following this literature review, a meeting to establish expert consensus was held and 20 guidelines established in the areas of pain assessment, opioid medications, nonopioid medications, regional anesthesia, and nonpharmacologic treatments. There is increasing research on pain management modalities, but available studies are inadequate to create a true standard of care. We call for more burn specific research into modalities for burn pain control as well as research on multimodal pain control.

Guidelines for Thrombolytic Therapy for Frostbite.

The data are insufficient to support standardized treatment of all patients with frostbite with thrombolytic therapy. The following guidelines, however, should be applied to all patients with cyanosis persisting proximal to the distal phalanx (Grade 3 or 4 frostbite injury) and demonstrated loss of perfusion at or proximal to the middle phalanx immediately after rewarming.

Evaluation and Management of the Burn Patient: A Case Study and Review.

Advances in the management of burn patients have contributed to significant improvements in morbidity and mortality over the last century. The physiologic insult from this injury pattern, however, still requires extensive surgical intervention, resuscitation and multidisciplinary care. This paper will review the standard of care of these patients in the context of a recent case study from our institution.

Palliative Coverage of Cranial Defect following Failed Cranial Flap for Advanced Squamous Cell Carcinoma: Case Report.

INTRODUCTION: With advanced-stage head and neck cancers, patients may develop large and/or complex wounds despite multiple reconstruction attempts. Wound coverage may require novel approaches to palliate the patient. METHOD: We present the case of a 56-year-old female with advanced squamous cell carcinoma of the scalp and skull who required multiple surgical interventions. Despite our best reconstructive efforts, the patient subsequently developed scalp infection and sepsis, necessitating further debridement for source control. She then required coverage of the exposed dura and skull to prevent further infection. RESULTS: The calvarial wound was covered with a dermal regeneration template and held in place by a vacuum-assisted closure (VAC) device. This coverage prevented additional infection and morbidity, was relatively easy and comfortable to manage, and demonstrated healing and development of granulation. Unfortunately, the patient succumbed to her systemic cancer before application of a palliative split-thickness skin graft. CONCLUSION: A VAC device and dermal regeneration template constituted an excellent modality for managing the complex calvarial wound encountered by otolaryngology, neurosurgery, and burn/wound services. The technique provided appropriate palliation for a patient with advanced head and neck cancer.

Propylthiouracil-Associated Leukocytoclastic Necrotizing Cutaneous Vasculitis: A Case Report and Review of the Literature.

The purpose of this case report and review of the literature is to provide an exploration of the clinical symptoms, diagnosis, prevention, and management of propylthiouracil (PTU)-associated vasculitis in the intensive care setting. A PubMed search of the available literature was conducted using the MeSH search terms "propylthiouracil" and "vasculitis." The literature search returned 121 articles. Twenty-five were excluded because they were not in English. Fifty-nine case reports or case studies describing PTU-associated vasculitis were included. Data extracted from each case study included patient age, sex, autoimmune markers, laboratory tests, length of time on PTU, treatment for vasculitis, and patient outcomes. The authors reviewed 128 cases of PTU-associated vasculitis. The majority were women (8.8:1 F:M ratio), and the most common presenting symptoms were rash (51.6%), fever (46.9%), and arthralgia (43.8%). In addition to discontinuing PTU, the most common treatment was steroids (71.9%). Eight patients (6.3%) progressed to end-stage renal disease; two (1.6%) required intubation for respiratory failure; and five (3.9%) died of various organ systems failure related to vasculitis development. A high index of suspicion for vasculitis should be maintained, especially when presented with skin manifestations in the presence of PTU therapy. Screening with myeloperoxidase-antinuclear cytoplasmic antibodies is most sensitive. Positive screening should prompt a thorough clinical investigation. In cases of severe skin manifestations, the focus should be on aggressive wound care. Our case report is unique, not only in the size and extent of cutaneous involvement, but also as the first description of mortality secondary to cutaneous manifestations.

Surgeon-Performed Hemodynamic Transesophageal Echocardiography in the Burn Intensive Care Unit.

The use of transesophageal echocardiography (TEE) for resuscitation after burn injury has been reported in small case studies. Conventional TEE is invasive and often requires a subspecialist with a high level of training. The authors report a series of surgeon-performed hemodynamic TEE with an indwelling, less bulky, user-friendly probe. Records of patients treated in a regional burn center who underwent hemodynamic TEE between October 1, 2012 and May 30, 2014 were reviewed. The clinical course of each patient was recorded. All bedside interpretations were retrospectively reviewed for accuracy by a cardiac anesthesiologist. Eleven patients were included in the study. Median age was 68.5 years (interquartile range, 49.5-79.5). Median burn size was 37% TBSA (interquartile range: 16.3-53%). Seven patients were male, and four suffered inhalation injury. The operator's interpretation matched that of the echocardiography technician and cardiac anesthesiologist in all instances. No complications occurred from probe placement. Four patients underwent hemodynamic TEE to determine volume status during resuscitation. Changes in volume status on echocardiography preceded the eventual changes in urine output and vital signs for one patient. Hemodynamic TEE diagnosed cardiogenic shock and was used to titrate inotropes and vasopressors in seven elderly patients. Hemodynamic TEE is a useful adjunct to manage the burn patient who deviates off the expected course, especially if there is a question of cardiac function or volume status. It is less invasive and can be accurately performed by surgical intensivists when transthoracic echo windows are limited. The role of echocardiography in optimizing routine burn resuscitations needs to be further studied.

Failure of the Platelet Function Assay (PFA)-100 to detect antiplatelet agents.

BACKGROUND: Antiplatelet therapy is a complicating factor in patients with traumatic brain injuries (TBI), as well as those with hemorrhagic cerebrovascular accidents (CVAs). Platelet Function Assay (PFA)-100 is a coagulation device that can detect platelet dysfunction caused by aspirin and adenosine diphosphate inhibition. Our retrospective study reviewed the effectiveness of PFA-100 in detecting platelet dysfunction caused by aspirin and clopidogrel and determined its clinical importance. METHODS: All patients with PFA-100 tests from January 2013 to February 2014 were collected. Diagnoses indicative of a TBI or CVA were chosen for analysis. Patients with a normal PFA-100 indicating no platelet dysfunction but with documented aspirin and/or clopidogrel use were selected. An extensive chart review was performed to determine the relevance to their clinical care. RESULTS: A total of 475 patients were evaluated with a PFA-100 from January 2013 to February 2014. PFA-100 detected platelet dysfunction as the result of pre-injury use of antiplatelet agents in TBI and CVA patients with a sensitivity of only 48.6% and a specificity of 74.8%. Had these antiplatelet medications been known during initial workup, these patients would have had a change in management that may have impacted their outcomes. CONCLUSION: Despite its common usage, the PFA-100 is an unreliable tool to assist in the management of TBI and CVA patients. Additional investigation into alternative methods for detecting platelet dysfunction is warranted.

Pathogenic serum amyloid A 1.1 shows a long oligomer-rich fibrillation lag phase contrary to the highly amyloidogenic non-pathogenic SAA2.2.

Serum amyloid A (SAA) is best known for being the main component of amyloid in the inflammation-related disease amyloid A (AA) amyloidosis. Despite the high sequence identity among different SAA isoforms, not all SAA proteins are pathogenic. In most mouse strains, the AA deposits mostly consist of SAA1.1. Conversely, the CE/J type mouse expresses a single non-pathogenic SAA2.2 protein that is 94% identical to SAA1.1. Here we show that SAA1.1 and SAA2.2 differ in their quaternary structure, fibrillation kinetics, prefibrillar oligomers, and fibril morphology. At 37 °C and inflammation-related SAA concentrations, SAA1.1 exhibits an oligomer-rich fibrillation lag phase of a few days, whereas SAA2.2 shows virtually no lag phase and forms small fibrils within a few hours. Deep UV resonance Raman, far UV-circular dichroism, atomic force microscopy, and fibrillation cross-seeding experiments suggest that SAA1.1 and SAA2.2 fibrils possess different morphology. Both the long-lived oligomers of pathogenic SAA1.1 and the fleeting prefibrillar oligomers of non-pathogenic SAA2.2, but not their respective amyloid fibrils, permeabilized synthetic bilayer membranes in vitro. This study represents the first comprehensive comparison between the biophysical properties of SAA isoforms with distinct pathogenicities, and the results suggest that structural and kinetic differences in the oligomerization-fibrillation of SAA1.1 and SAA2.2, more than their intrinsic amyloidogenicity, may contribute to their diverse pathogenicity.

Photochemical restoration of visual responses in blind mice.

Retinitis pigmentosa (RP) and age-related macular degeneration (AMD) are degenerative blinding diseases caused by the death of rods and cones, leaving the remainder of the visual system intact but largely unable to respond to light. Here, we show that AAQ, a synthetic small molecule photoswitch, can restore light sensitivity to the retina and behavioral responses in vivo in mouse models of RP, without exogenous gene delivery. Brief application of AAQ bestows prolonged light sensitivity on multiple types of retinal neurons, resulting in synaptically amplified responses and center-surround antagonism in arrays of retinal ganglion cells (RGCs). Intraocular injection of AAQ restores the pupillary light reflex and locomotory light avoidance behavior in mice lacking retinal photoreceptors, indicating reconstitution of light signaling to brain circuits. AAQ and related photoswitch molecules present a potential drug strategy for restoring retinal function in degenerative blinding diseases.

Conformational differences between two amyloid ß oligomers of similar size and dissimilar toxicity.

Several protein conformational disorders (Parkinson and prion diseases) are linked to aberrant folding of proteins into prefibrillar oligomers and amyloid fibrils. Although prefibrillar oligomers are more toxic than their fibrillar counterparts, it is difficult to decouple the origin of their dissimilar toxicity because oligomers and fibrils differ both in terms of structure and size. Here we report the characterization of two oligomers of the 42-residue amyloid ß (Aß42) peptide associated with Alzheimer disease that possess similar size and dissimilar toxicity. We find that Aß42 spontaneously forms prefibrillar oligomers at Aß concentrations below 30 µm in the absence of agitation, whereas higher Aß concentrations lead to rapid formation of fibrils. Interestingly, Aß prefibrillar oligomers do not convert into fibrils under quiescent assembly conditions but instead convert into a second type of oligomer with size and morphology similar to those of Aß prefibrillar oligomers. Strikingly, this alternative Aß oligomer is non-toxic to mammalian cells relative to Aß monomer. We find that two hydrophobic peptide segments within Aß (residues 16-22 and 30-42) are more solvent-exposed in the more toxic Aß oligomer. The less toxic oligomer is devoid of ß-sheet structure, insoluble, and non-immunoreactive with oligomer- and fibril-specific antibodies. Moreover, the less toxic oligomer is incapable of disrupting lipid bilayers, in contrast to its more toxic oligomeric counterpart. Our results suggest that the ability of non-fibrillar Aß oligomers to interact with and disrupt cellular membranes is linked to the degree of solvent exposure of their central and C-terminal hydrophobic peptide segments.

Influence of the carboxy terminus of serum amyloid A on protein oligomerization, misfolding, and fibril formation.

The fibrillar deposition of serum amyloid A (SAA) has been linked to the disease amyloid A (AA) amyloidosis. We have used the SAA isoform, SAA2.2, from the CE/J mouse strain, as a model system to explore the inherent structural and biophysical properties of SAA. Despite its nonpathogenic nature in vivo, SAA2.2 spontaneously forms fibrils in vitro, suggesting that SAA proteins are inherently amyloidogenic. However, whereas the importance of the amino terminus of SAA for fibril formation has been well documented, the influence of the proline-rich and presumably disordered carboxy terminus remains poorly understood. To clarify the inherent role of the carboxy terminus in the oligomerization and fibrillation of SAA, we truncated the proline-rich final 13 residues of SAA2.2. We found that unlike full-length SAA2.2, the carboxy-terminal truncated SAA2.2 (SAA2.2ΔC) did not oligomerize to a hexamer or octamer, but formed a high molecular weight soluble aggregate. Moreover, SAA2.2ΔC also exhibited a pronounced decrease in the rate of fibril formation. Intriguingly, when equimolar amounts of denatured SAA2.2 and SAA2.2ΔC were mixed and allowed to refold together, the mixture formed an octamer and exhibited rapid fibrillation kinetics, similar to those for full-length SAA2.2. These results suggest that the carboxy terminus of SAA, which is highly conserved among SAA sequences in all vertebrates, might play important structural roles, including modulating the folding, oligomerization, misfolding, and fibrillation of SAA.

Controlling DNA adsorption and diffusion on lipid bilayers by the formation of lipid domains.

We describe the influence of membrane heterogeneity on the adsorption and diffusion of DNA. Cellular membranes are believed to contain domains (lipid rafts) that influence processes ranging from signal transduction to the diffusion of membrane components. By analogy, we demonstrate that the formation of raft-like domains in supported lipid bilayers provides control over the adsorption and diffusion of DNA. The formation of bilayers from a mixture of the gel phase zwitterionic lipid 1,2-distearoyl-sn-glycero-3-phosphatidylcholine (DSPC) and the fluid phase cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) yielded coexisting DSPC-enriched and DOTAP-enriched phases. We demonstrated the ability to pattern the adsorption of DNA on the heterogeneous bilayers, with the adsorption being restricted to the DOTAP-enriched phase. We further demonstrated that the DSPC-enriched domains acted as obstacles to the lateral diffusion of adsorbed DNA. Fluorescence recovery after photobleaching (FRAP) analysis revealed that the diffusivity of the adsorbed DNA tracked that of the underlying lipid, although the lipid diffusivity changed by an order of magnitude with changes in bilayer composition. Fundamental insight into the adsorption and diffusion of DNA on heterogeneous surfaces may be useful for the design of novel techniques for the size-based separation of DNA.

Enhancing protein stability by adsorption onto raftlike lipid domains.

We demonstrate that the stability of adsorbed proteins can be enhanced by controlling the heterogeneity of the surfaceby creating raftlike domains in a soft liposomal membrane. Recent work has shown that enzymes adsorbed onto highly curved nanoscale supports can be more stable than those adsorbed on flat surfaces with nominally the same chemical structure. This effect has been attributed to a decrease in lateral interenzyme interactions on a curved surface. Exploiting this idea, we asked if adsorbing enzymes onto "patchy" surfaces composed of adsorbing and nonadsorbing regions can be used to reduce lateral interactions even on relatively flat surfaces. We demonstrate that creating domains on which an enzyme can adsorb enhances the stability of that enzyme under denaturing conditions. Furthermore, we demonstrate that the size of these domains has a considerable effect on the degree of stability imparted by adsorption. Such biomimetic raft-inspired systems may find use in applications ranging from biorecognition to the design of novel strategies for the separation of biomolecules and controlling the interaction of multicomponent membrane-bound enzymes.