Gas gangrene with Clostridium septicum in a neutropenic patient.

Gas gangrene is a rare presentation of a necrotizing fasciitis, caused by Clostridium perfringens, C. septicum and other clostridial species. With its rapid progression it is a potentially life-threatening infection, that poses as a challenge in the clinical management requiring an interdisciplinary approach.Here we present a 62-year-old woman, who developed neutropenic fever while undergoing chemotherapy for triple negative breast cancer. She presented with a high fever, reporting little pain in her left thigh accompanied by redness and induration locally. Subsequently the patient developed pain and redness of the back of the left hand. The initial findings suggested cellulitis and immediate empiric treatment with intravenous meropenem was started. Despite the antibiotic treatment the patient rapidly developed septic shock along with progression of the local infection. Emergency surgical debridement revealed extensive necrosis of the soft tissues including extensive myonecrosis of the thigh. On the left hand an extensive debridement was performed, the left lower limb could not be preserved and exarticulation of the left hip was required. Microbiologically C. septicum was isolated in different samples, confirming gas gangrene. As there was no local entry portal on the skin, hematogenous seeding from intestinal translocation in this neutropenic patient was suspected. The empiric antibiotic treatment was tailored to intravenous penicillin and complemented with clindamycin for toxin inhibition. Following radical debridement and antibiotic treatment, the patient could be stabilized. After repetitive debridement wound closure was achieved and the patient was discharged for rehabilitation. Antibiotic treatment was continued for four weeks.This rare case of gas gangrene in a neutropenic patient shows the complexity in the diagnostic and therapeutic management of necrotizing soft tissue infections in immunocompromised patients. It particularly highlights the importance of an interdisciplinary management with fast recognition of the disease and rapid, if needed radical, surgical debridement as well as tailored antibiotic treatment for a successful outcome.

New-onset autoantibodies to selenoprotein P following severe burn injury.

The liver-derived selenium (Se) transporter selenoprotein P (SELENOP) declines in critical illness as a negative acute phase reactant and has recently been identified as an autoantigen. Hepatic selenoprotein biosynthesis and cotranslational selenocysteine insertion are sensitive to inflammation, therapeutic drugs, Se deficiency, and other modifiers. As severe burn injury induces a heavy inflammatory burden with concomitant Se depletion, we hypothesized an impairment of selenoprotein biosynthesis in the acute post-burn phase, potentially triggering the development of autoantibodies to SELENOP (SELENOP-aAb). To test this hypothesis, longitudinal serum samples from severely burned patients were analyzed over a period of six months. Newly occurring SELENOP-aAb were detected in 8.4% (7/83) of the burn patients, with onset not earlier than two weeks after injury. Prevalence of SELENOP-aAb was associated with injury severity, as aAb-positive patients have suffered more severe burns than their aAb-negative counterparts (median [IQR] ABSI: 11 [7-12] vs. 7 [5.8-8], p = 0.023). Autoimmunity to SELENOP was not associated with differences in total serum Se or SELENOP concentrations. A positive correlation of kidney-derived glutathione peroxidase (GPx3) with serum SELENOP was not present in the patients with SELENOP-aAb, who showed delayed normalization of GPx3 activity post-burn. Overall, the data suggest that SELENOP-aAb emerge after severe injury in a subset of patients and have antagonistic effects on Se transport. The nature of burn injury as a sudden event allowed a time-resolved analysis of a direct trigger for new-onset SELENOP-aAb, which may be relevant for severely affected patients requiring intensified acute and long-term care.

Serum Selenium-Binding Protein 1 (SELENBP1) in Burn Injury: A Potential Biomarker of Disease Severity and Clinical Course.

Oxidative stress, systemic inflammation, and metabolic derangements are hallmarks of burn pathophysiology. Severely burned patients are highly susceptible to infectious complications. Selenium-binding protein 1 (SELENBP1) modulates intracellular redox homeostasis, and elevated serum concentrations have been associated with adverse clinical outcomes in trauma patients. We hypothesized that serum SELENBP1 at hospital admission and during hospitalization may constitute a meaningful biomarker of disease severity and the clinical course in burn injury, with pulmonary infection as primary endpoint. To this end, we conducted a prospective cohort study that included 90 adult patients admitted to the Burn Center of the University Hospital Zurich, Switzerland. Patients were treated according to the local standard of care, with high-dose selenium supplementation during the first week. Serum SELENBP1 was determined at nine time-points up to six months postburn and the data were correlated to clinical parameters. SELENBP1 was initially elevated and rapidly declined within the first day. Baseline SELENBP1 levels correlated positively with the Abbreviated Burn Severity Index (ABSI) (R = 0.408; p < 0.0001). In multiple logistic regression, a higher ABSI was significantly associated with increased pulmonary infection risk (OR, 14.4; 95% CI, 3.2-88.8; p = 0.001). Similarly, baseline SELENBP1 levels constituted a novel but less accurate predictor of pulmonary infection risk (OR, 2.5; 95% CI, 0.7-8.9; p = 0.164). Further studies are needed to explore the additional value of serum SELENBP1 when stratifying patients with respect to the clinical course following major burns and, potentially, for monitoring therapeutic measures aimed at reducing tissue damage and oxidative stress.

Risk Factors for Nonunion After Distal Phalangeal Fractures of the Hand.

PURPOSE: This study aimed to evaluate the risk factors for distal phalanx fracture nonunion. METHODS: We retrospectively reviewed all adult patients treated for distal phalanx fractures at our institution between January 2015 and December 2019 with a minimum one-year follow-up period for potential risk factors. The absence of consolidation signs on follow-up radiographs at least 12 months after trauma was defined as nonunion. RESULTS: This study included 124 patients with 143 fractures available for follow-up. Nonunion was diagnosed in 19 patients, 18 of whom initially presented with an open fracture. On the day of the injury, 17 patients with open fractures presented to the hospital. In 16 nonunion cases, the traumatic mechanism was a crush injury. All nonunions occurred in tuft fractures, and none required revision surgery at the follow-up visit. CONCLUSIONS: Our findings suggest that tuft involvement in open fractures is the main risk factor for nonunion of distal phalangeal fractures. However, after a minimum of 1 year of follow-up, none of the tuft nonunions required revision surgery. TYPE OF STUDY/LEVEL OF EVIDENCE: Therapeutic IV.

Changes in coagulation and temperature management in burn patients - A survey of burn centers in Switzerland, Austria and Germany.

BACKGROUND: Severely burned patients suffer from both coagulopathy and hypothermia, with a lack of international consensus and appropriate treatment guidelines. This study examines recent developments and trends in coagulation and temperature management in European burn centers. METHODS: A survey was sent to burn centers in Switzerland, Austria and Germany in 2016 and again in 2021. The analysis was performed using descriptive statistics, with categorical data reported in absolute numbers (n) and percentages (%) and numerical data reported as mean and standard deviation. RESULTS: The rate of completed questionnaires was 84 % (16 of 19 questionnaires) in 2016 and 91 % (21 of 22 questionnaires) in 2021. The number of global coagulation tests performed has decreased over the observation period in favor of single factor determination and bed-side point-of-care coagulation tests. This has also led to increased administration of single factor concentrates in therapy. Although many centers had a defined treatment protocol for hypothermia in 2016, coverage increased such that in 2021 all centers surveyed had such a protocol. The body temperature was measured more consistently in 2021; thus, hypothermia was more actively sought, detected and treated. CONCLUSION: A point-of-care guided, factor-based coagulation management and the maintenance of normothermia have gained importance in the care of burn patients in recent years.

Burn Scar Ectropion Correction: Surgical Technique for Functional Outcomes.

BACKGROUND: Eyelid scarring after severe burn injury of the face is a significant complication endangering vision in addition to the burn scar sequelae. Scar contraction leads to asymmetry and malposition of the eyelid axis, resulting in corneal exposure, eyelid retraction, and incomplete eyelid closure. In consequence, dryness and irritation of the cornea can lead to keratitis, corneal opacity, and vision impairment. In this study, we present our surgical technique for lateral canthopexy in combination with full-thickness skin grafting (FTSGing) in patients with eyelid axis distortion after scar contraction of the periorbital region after severe burn injuries of the face. METHODS: In this retrospective, single-center case study, we present 5 consecutive patients who experienced severe burn injuries to the face between 2014 and 2019. Patients were suffering from ectropion and malposition of the eyelid axis. In all cases, we performed lateral transosseous canthopexy and FTSGing. RESULTS: Improved symmetry and complete eyelid closure were restored in all 5 patients. The following ophthalmological examinations showed resolved corneal erosions, as well as reduction of chemosis and epiphora. Further vision impairment was successfully prohibited. Surgical revision with FTSGing was required in 2 patients because of recurrence of unilateral lower eyelid retraction. CONCLUSIONS: Lateral transosseous canthopexy represents a suitable surgical method to durably correct eyelid malposition, ectropion, and incomplete lid closure in patients with severe scarring of the periorbital region after burns of the face. Early detection of patients at risk and timing of surgical intervention are of great importance.

Evolution of a concept with enzymatic debridement and autologous in situ cell and platelet-rich fibrin therapy (BroKerF).

BACKGROUND: Deep partial-thickness burns are traditionally treated by tangential excision and split thickness skin graft (STSG) coverage. STSGs create donor site morbidity and increase the wound surface in burn patients. Herein, we present a novel concept consisting of enzymatic debridement of deep partial-thickness burns followed by co-delivery of autologous keratinocyte suspension and plated-rich fibrin (PRF) or fibrin glue. MATERIAL AND METHODS: In a retrospective case study, patients with deep partial-thickness burns treated with enzymatic debridement and autologous cell therapy combined with PRF or fibrin glue (BroKerF) between 2017 and 2018 were analysed. BroKerF was applied to up to 15% total body surface area (TBSA); larger injuries were combined with surgical excision and skin grafting. Exclusion criteria were age <18 or >70 years, I°, IIa°-only, III° burns and loss of follow-up. RESULTS: A total of 20 patients with burn injuries of 16.8% ± 10.3% TBSA and mean Abbreviated Burn Severity Score 5.45 ± 1.8 were identified. Of the patients, 65% (n = 13) were treated with PRF, while 35% (n = 7) were treated with fibrin glue. The mean area treated with BroKerF was 7.5% ± 0.05% TBSA, mean time to full epithelialization was 21.06 ± 9.2 days and mean hospitalization time was 24.7 ± 14.4 days. Of the patients, 35% (n = 7) needed additional STSG, 43% (n = 3) of whom had biopsy-proven wound infections. CONCLUSION: BroKerF is an innovative treatment strategy, which, in our opinion, will show its efficacy when higher standardization is achieved. The combination of selective debridement and autologous skin cells in a fibrin matrix combines regenerative measures for burn treatment. LAY SUMMARY: Patients suffering from large burn wounds often require the use of large skin grafts to bring burned areas to heal. Before the application of skin grafts, the burned skin must be removed either by surgery or using enzymatic agents. In this article, we describe a method where small areas of skin are taken and skin cells are extracted and sprayed on wound areas that were treated with an enzymatic agent. The cells are held in place by a substance extracted from patients' blood (PRF) that is sprayed on the wound together with the skin cells. We believe this technique can be helpful to reduce the need of skin grafts in burned patients and improve the healing process.

Effect of a factor-based coagulation management on blood product use after major burn injury: A retrospective cohort study.

BACKGROUND: Transfusion of allogenic blood products was shown to be associated with more adverse events and a higher mortality in severely burned patients. This study investigated the impact of a goal-directed and factor-based coagulation algorithm on blood product use and clinical outcomes in severely burned patients. METHODS: This retrospective cohort study included adult patients admitted to the burn center of the University Hospital Zurich with major burn injuries compromising 20-80% of total body surface area. We compared two 3-year periods, one before the introduction of a goal-directed coagulation and transfusion algorithm (period 1: 2009-2011) and one after (period 2: 2016-2018). We applied linear and logistic regression models adjusted for confounders. RESULTS: We analyzed 36 patients (27.8% female) versus 42 patients (14.3% female) in period 1 and 2, respectively. Comorbidities and burn types were comparable between both collectives. Treatment according to the coagulation algorithm resulted in an overall reduction of 33 units of red blood cells (95% CI -52.8 to -12.9, p = 0.002), 9 units fresh frozen plasma (95% CI -14.7 to -2.6, p = 0.006) and 1.4g fibrinogen (95% CI -2.2 to -0.5, p = 0.001) per patient. We observed less infections (61.8% vs. 41.5%, p = 0.11) and a reduced mortality (38.9% vs. 26.8%, p = 0.33) during the algorithm treated period, although not significant. CONCLUSION: Treatment of severely burned patients with a goal-directed coagulation algorithm reduced blood product use and resulted in target-oriented administration of coagulation factors to improve outcomes.

Identification of ALP+/CD73+ defining markers for enhanced osteogenic potential in human adipose-derived mesenchymal stromal cells by mass cytometry.

BACKGROUND: The impressive progress in the field of stem cell research in the past decades has provided the ground for the development of cell-based therapy. Mesenchymal stromal cells obtained from adipose tissue (AD-MSCs) represent a viable source for the development of cell-based therapies. However, the heterogeneity and variable differentiation ability of AD-MSCs depend on the cellular composition and represent a strong limitation for their use in therapeutic applications. In order to fully understand the cellular composition of MSC preparations, it would be essential to analyze AD-MSCs at single-cell level. METHOD: Recent advances in single-cell technologies have opened the way for high-dimensional, high-throughput, and high-resolution measurements of biological systems. We made use of the cytometry by time-of-flight (CyTOF) technology to explore the cellular composition of 17 human AD-MSCs, interrogating 31 markers at single-cell level. Subcellular composition of the AD-MSCs was investigated in their naïve state as well as during osteogenic commitment, via unsupervised dimensionality reduction as well as supervised representation learning approaches. RESULT: This study showed a high heterogeneity and variability in the subcellular composition of AD-MSCs upon isolation and prolonged culture. Algorithm-guided identification of emerging subpopulations during osteogenic differentiation of AD-MSCs allowed the identification of an ALP+/CD73+ subpopulation of cells with enhanced osteogenic differentiation potential. We could demonstrate in vitro that the sorted ALP+/CD73+ subpopulation exhibited enhanced osteogenic potential and is moreover fundamental for osteogenic lineage commitment. We finally showed that this subpopulation was present in freshly isolated human adipose-derived stromal vascular fractions (SVFs) and that could ultimately be used for cell therapies. CONCLUSION: The data obtained reveal, at single-cell level, the heterogeneity of AD-MSCs from several donors and highlight how cellular composition impacts the osteogenic differentiation capacity. The marker combination (ALP/CD73) can not only be used to assess the differentiation potential of undifferentiated AD-MSC preparations, but also could be employed to prospectively enrich AD-MSCs from the stromal vascular fraction of human adipose tissue for therapeutic applications.

[Implementation of treatment by enzymatic debridement in burns: results of an interprofessional German-speaking expert workshop]. / Implementierung der Behandlung durch enzymatisches Debridement bei Verbrennungen ­ Ergebnisse eines interprofessionellen, deutschsprachigen Expertenworkshops.

INTRODUCTION: Since its introduction in 2013 Bromelain-based Enzymatic Debridement (ED) is increasingly used in burn centers. Published evidence shows its efficiency in eschar removal as well as a superiority in blood loss and necessity of further surgical procedures compared to standard-of-care. While the procedure is safe and shows reliable results in experienced hands, some practical and logistical issues must be challenged that are not described sufficiently in available literature. METHOD: A multi-professional panel, consisting of experienced users of ED from German-speaking burn units has been invited to an expert workshop. Topics concerning indication, definition of treatment pathways, practical issues, post-treatment and handling of complications have been coordinated in advance to allow discussion during the workshop. RESULTS: To each topic practical recommendations were developed and consented. Summarizing key messages have been additionally highlighted. They aim on helping to achieve optimal results after establishing the technique by new users as well as optimizing results by experienced users. Amongst others, the resulting recommendations deal with indications for ED beyond the classic domain, different treatment pathways depending on burn depth and primary result after ED with adapted post-treatment, management of treatment failure and implementation of infrastructural conditions. DISCUSSION: While efficiency of ED as well as superiority in some aspects of treatment of burn wounds could be shown in available literature, user-oriented recommendations for practical implementation are scarce. Although the recommendations and experts opinions published here are only partly evidenced based, they are still based on the pooled experienced of the panelists that easily outnumbers the cases published in literature so far and allow valuable support for a successful implementation of the technique.

Risk Factors for Mortality and Prolonged Hospitalization in Electric Burn Injuries.

Electrical injuries are rare, but very destructive with high morbidity and mortality, prolonged hospital length of stay and need for repeated procedures. The aim of study was to investigate characteristics and management of electrical injuries and predisposing factors for mortality and prolonged length of stay. Patient charts were reviewed retrospectively to identify patients admitted with electrical injuries at the Zurich Burns Center (2005-2019). Patient characteristics, management, and outcome were analyzed and risk factors for mortality and prolonged hospitalization were assessed. Eighty-nine patients were included, mostly males (86.5%), between 21 and 40 years (50.6%), with high-voltage (74.2%) occupational injuries (66.3%). Median intensive care unit and hospital stays were 6 (first and third IQR: 2.0; 30.0) and 18 (9.0; 48.0) days. Low-voltage patients had a median of 2 (1.5; 3.0) procedures, compared to 4 (2.0; 10.8) in high-voltage. The amputation rate was 13.5%, and a total of 46 flaps were required. Fifty-four patients had at least one serious complication. Mortality was 18% in high-voltage patients, mostly after multiple organ failure (35%). High total body surface area (TBSA), renal failure and cardiovascular complications were risk factors for mortality (P < .001) in multivariate regression models. Determinants for prolonged hospital stay were TBSA and sepsis (P < .01), and additionally abdominal complications and limb loss for intensive care unit stay (P < .05). Electrical injuries are still cause of significant morbidity and mortality, mostly involve young men in their earning period. Several risk factors for in-hospital mortality and prolonged stay were identified and can support physicians in the management and decision making in these patients.

Response of routine inflammatory biomarkers and novel Pancreatic Stone Protein to inhalation injury and its interference with sepsis detection in severely burned patients.

BACKGROUND: Inhalation of thermal and chemical products of combustion evokes an immune response measurable at a systemic level. Inhalation injury related kinetics of currently available inflammatory biomarkers and novel Pancreatic Stone Protein (PSP) as well as their interference with septic events has not been addressed to literature yet. METHODS: Analysis of the influence of inhalation injury and ARDS on biomarker kinetics (PSP, procalcitonin (PCT), C-reactive Protein (CRP), white blood cells (WBC)) in 90 patients admitted to Zurich Burn Center between May 2015 and October 2018 with burns ≥15% total body surface area (TBSA) over 14 days. RESULTS: Twenty-five (27%) of 90 included patients presented with inhalation injury (median age 52 years [IQR 27], median TBSA 31.5% [IQR 21], mean ABSI-Score 7±3). At admission, only WBC demonstrated significantly higher values in the inhalation injury group (p=0.011). Acute respiratory distress syndrome (ARDS) was present in 32% without association to the severity of inhalation injury (p=0.11). WBC, CRP and PCT failed to delineate inhalation injury related inflammation from septic progression at most time points. PSP was the strongest marker to identify septic patients both by its higher values and steeper increase over time (p<0.001). CONCLUSION: Inhalation injury leads to an inflammatory response at a systemic level with alterations of biomarkers. While routine inflammatory markers demonstrated strong interferences between inhalation injury with its associated ARDS and evolving sepsis, PSP reliably identified septic patients in a setting of inflammatory turbulences secondary to inhalation injury.

Safety of enzymatic debridement in extensive burns larger than 15% total body surface area.

OBJECTIVES: Bromelain-based enzymatic debridement has emerged as an alternative to surgical eschar removal. Indications include partial thickness, mixed pattern, and full-thickness burns. Enzymatic debridement has been approved by the European Medicines Agency for treating burn wounds affecting <15% total body surface area (TBSA). Data and evidence for the treatment of areas >15% TBSA in one session is scarce. The aim of this retrospective study was to retrospectively analyze off-label use of enzymatic debridement in a single burn center for large TBSA burns. METHODS: Between 01/2017 and 12/2018, 59 patients with partial- to full-thickness burns underwent enzymatic debridement in a single center study. Patients were categorized into two groups: the regular use group with a treated area less than 15% TBSA and the off-label group (OG) with larger TBSA debrided in one session. Treatment was evaluated for systemic inflammatory reaction, bleeding, hemodynamic instability and electrolyte shifts. RESULTS: In total, 49 patients were treated in the regular use group with a median application area of 6% (IQR 2.5-9.5) and 10 patients were treated in the off-label group with a median application area of 18% (IQR 15-19) TBSA. We found no significant differences regarding blood pressure, body temperature or hemodynamic stability during and after enzymatic debridement. No treatment-related serious adverse events were observed in either group. Catecholamine use was similar in both groups. No differences in leukocyte counts, CRP, PCT and lactate prior to application and during the following three days were observed. Sodium, potassium, chloride and phosphate levels did not differ. We found no evidence of an electrolyte shift. Survival was 49 of 49 patients (100%) in the RG and 7 of 10 patients (70%) in the OG (p = 0.004). CONCLUSION: Enzymatic debridement did not result in any expected or unexpected side effects in the patient groups investigated. These preliminary results indicate the potential safety of bromelain-based enzymatic debridementin the treatment of burns greater than 15% TBSA.

Characterisation of clinical manifestations and treatment strategies for invasive beta-haemolytic streptococcal infections in a Swiss tertiary hospital.

AIMS OF THE STUDY: Invasive streptococcal infections affect more than half a million patients worldwide every year and have a high lethality. Little is known about the epidemiology and microbiological characteristics of streptococcal infections in Switzerland. This case series study aims to describe the demographics, known risk factors for streptococcal skin and soft tissue infections, clinical presentations, treatment and outcomes of patients admitted to the University Hospital Zurich between 2000 and 2014 with invasive streptococcal infections caused by Streptococcus pyogenes (group A Streptococcus), Streptococcus dysgalactiae ssp. equisimilis or the Streptococcus anginosus group, as well as the microbiological characteristics of the clinical isolates. METHODS: Data collected retrospectively from patients hospitalised between 2000 and 2014 with invasive streptococcal infections were analysed. M protein gene (emm) typing of the bacterial clinical isolates was carried out according to the Centers for Disease Control and Prevention guidelines. RESULTS: A total of 86 patients with invasive beta-haemolytic streptococcal infections were included in this study, of which 49% presented with necrotising fasciitis (NF). The median age was 44 years and half were female. The most common risk factor was acute skin lesions. C-reactive protein levels were significantly higher in patients with NF, as were acute renal failure and distributive shock. Beta-lactam antibiotics were given to most patients, and intravenous immunoglobulins were given to 18% of patients within the first 24 hours. All patients suffering from NF underwent surgery. The overall case fatality rate was 8.1% at 30 days post admission. All Group A Streptococcus strains were susceptible to penicillin and clindamycin, and we found resistance to tetracycline in 11.9% of strains. The most common emm-type isolated was emm1 (44.4%). CONCLUSIONS: Invasive beta-haemolytic streptococcal infections, the most severe presentation of which is NF, remain a serious clinical issue and require rapid diagnosis and treatment. This is the first representative analysis monitoring clinical and microbiological characteristics of patients with a severe invasive beta-haemolytic streptococcal infection and treated in Zurich, Switzerland. In addition to the detailed reporting of various clinical and microbiological characteristics, we show that C-reactive protein levels, acute renal failure and distributive shock were higher in the patients with NF. We also found a low case fatality rate compared to other reports. The detailed clinical data and microbiological characteristics depicted in this study will lead to a better understanding of regional differences in severe invasive streptococcal infections.

Pharmacokinetics and Biodistribution of Tacrolimus after Topical Administration: Implications for Vascularized Composite Allotransplantation.

AIM: The high doses of oral tacrolimus (TAC) (1,2) necessary to prevent acute rejection (AR) after vascularized composite allotransplantation (VCA) are associated with systemic adverse effects. The skin is the most antigenic tissue in VCA and the primary target of AR. However, the short-term use of topical TAC (Protopic®), as an off-label adjunct to oral TAC, to treat AR episodes pro re nata (PRN), has yielded inconsistent results. There is lack of data on the pharmacokinetics and tissue distribution of topical TAC in VCA, that hampers our understanding of the reasons for unreliable efficacy. Toward this goal, we evaluated the ability of topical TAC to achieve high local tissue concentrations at the site of application with low systemic concentrations. MATERIALS AND METHODS: We assessed the pharmacokinetics and tissue distribution of topical TAC (Protopic®, 0.03%) after single or repeated topical application in comparison to those after systemic delivery in rats. Animals received a single topical application of TAC ointment (Group 1) or an intravenous (IV) injection of TAC (Group 2) at a dose of 0.5 mg/kg. In another experiment, animals received daily topical application of TAC ointment (Group 3), or daily intraperitoneal (IP) injection of TAC (Group 4) at a dose of 0.5 mg/kg for 7 days. TAC concentrations in blood and tissues were analyzed by Liquid Chromatography-Mass Spectrometry (LC/MS-MS). RESULTS: Following single topical administration, TAC was absorbed slowly with a Tmax of 4 h and an absolute bioavailability of 11%. The concentrations of TAC in skin and muscle were several folds higher than whole blood concentrations. Systemic levels remained subtherapeutic (< 3 ng/ml) with repeated once daily applications. CONCLUSION: Topical application of TAC ointment (Protopic®, 0.03%) at a dose of 0.5 mg/kg/day provided high concentrations in the local tissues with low systemic exposure. Repeated topical administration of TAC is well tolerated with no local or systemic adverse effects. This study confirms the feasibility of topical application of TAC for site specific graft immunosuppression and enables future applications in VCA.

Effect of Systemic Adipose-derived Stem Cell Therapy on Functional Nerve Regeneration in a Rodent Model.

Regardless of etiology, peripheral nerve injuries (PNI) result in disruption/loss of neuromuscular junctions, target muscle denervation, and poor sensorimotor outcomes with associated pain and disability. Adipose-derived stem cells (ASCs) have shown promise in neuroregeneration. However, there is a paucity of objective assessments reflective of functional neuroregeneration in experimental PNI. Here, we use a multimodal, static, and dynamic approach to evaluate functional outcomes after ASC therapy in a rodent PNI model. METHODS: Lewis rats were divided into 3 groups: 10 mm sciatic nerve resection ("CUT" group; n = 10), transection and repair ("REP" group; n = 10), transection and repair plus single-dose ASCs ("ASC" group; n = 12). Allogeneic (Brown Norway rat) ASCs (1 × 106) were administered intravenously on postoperative day 1. Functional outcome was assessed by static sciatic index, toe spread factor, and a dynamic swim test on a weekly basis for 6 weeks. Sciatic nerves and gastrocnemius muscles were harvested at endpoint (6 weeks) for histological analysis. RESULTS: The ASC group showed accelerated functional recovery on the swim test at 2 weeks postoperatively, with continued improvement over 4 weeks, culminating in superior overall outcomes at 6 weeks compared with the REP group. The CUT group showed no significant improvement from baseline. Nerve histomorphometry correlated well with the swim test results in the ASC group. Gastrocnemius muscle weights showed no difference between the REP and the ASC groups. CONCLUSION: Our study confirms that early, single dose, systemic administration of ASC after PNI accelerates and enhances overall motor recovery on static and dynamic functional tests as evidenced by improvements in voluntary as well as involuntary motions.

Adipose tissue and the vascularization of biomaterials: Stem cells, microvascular fragments and nanofat-a review.

Tissue defects in the human body after trauma and injury require precise reconstruction to regain function. Hence, there is a great demand for clinically translatable approaches with materials that are both biocompatible and biodegradable. They should also be able to adequately integrate within the tissue through sufficient vascularization. Adipose tissue is abundant and easily accessible. It is a valuable tissue source in regenerative medicine and tissue engineering, especially with regard to its angiogenic potential. Derivatives of adipose tissue, such as microfat, nanofat, microvascular fragments, stromal vascular fraction and stem cells, are commonly used in research, but also clinically to enhance the vascularization of implants and grafts at defect sites. In plastic surgery, adipose tissue is harvested via liposuction and can be manipulated in three ways (macro-, micro- and nanofat) in the operating room, depending on its ultimate use. Whereas macro- and microfat are used as a filling material for soft tissue injuries, nanofat is an injectable viscous extract that primarily induces tissue remodeling because it is rich in growth factors and stem cells. In contrast to microfat that adds volume to a defect site, nanofat has the potential to be easily combined with scaffold materials due to its liquid and homogenous consistency and is particularly attractive for blood vessel formation. The same is true for microvascular fragments that are easily isolated from adipose tissue through collagenase digestion. In preclinical animal models, it has been convincingly shown that these vascular fragments inosculate with host vessels and subsequently accelerate scaffold perfusion and host tissue integration. Adipose tissue is also an ideal source of stem cells. It yields larger quantities of cells than any other source and is easier to access for both the patient and doctor compared with other sources such as bone marrow. They are often used for tissue regeneration in combination with biomaterials. Adipose-derived stem cells can be applied unmodified or as single cell suspensions. However, certain pretreatments, such as cultivation under hypoxic conditions or three-dimensional spheroids production, may provide substantial benefit with regard to subsequent vascularization in vivo due to induced growth factor production. In this narrative review, derivatives of adipose tissue and the vascularization of biomaterials are addressed in a comprehensive approach, including several sizes of derivatives, such as whole fat flaps for soft tissue engineering, nanofat or stem cells, their secretome and exosomes. Taken together, it can be concluded that adipose tissue and its fractions down to the molecular level promote, enhance and support vascularization of biomaterials. Therefore, there is a high potential of the individual fat component to be used in regenerative medicine.

Evaluation of Porcine Versus Human Mesenchymal Stromal Cells From Three Distinct Donor Locations for Cytotherapy.

Background: Mesenchymal stromal cell (MSC)-based cytotherapies fuel the hope for reduction of chronic systemic immunosuppression in allotransplantation, and our group has previously shown this capability for both swine and human cells. MSCs harvested from distinct anatomical locations may have different behavior and lead to different outcomes in both preclinical research and human trials. To provide an effective reference for cell therapy studies, we compared human and porcine MSCs from omental fat (O-ASC), subcutaneous fat (SC-ASC) and bone marrow (BM-MSC) under rapid culture expansion with endothelial growth medium (EGM). Methods: MSCs isolated from pigs and deceased human organ donors were compared for yield, viability, cell size, population doubling times (PDT), surface marker expression and differentiation potential after rapid expansion with EGM. Immunosuppressant toxicity on MSCs was investigated in vitro for four different standard immunosuppressive drugs. Immunomodulatory function was compared in mixed lymphocyte reaction assays (MLR) with/without immunosuppressive drug influence. Results: Human and porcine omental fat yielded significantly higher cell numbers than subcutaneous fat. Initial PDT was significantly shorter in ASCs than BM-MSCs and similar thereafter. Viability was reduced in BM-MSCs. Porcine MSCs were positive for CD29, CD44, CD90, while human MSCs expressed CD73, CD90 and CD105. All demonstrated confirmed adipogenic differentiation capacity. Cell sizes were comparable between groups and were slightly larger in human cells. Rapamycin revealed slight, mycophenolic acid strong and significant dose-dependent toxicity on viability/proliferation of almost all MSCs at therapeutic concentrations. No relevant toxicity was found for Tacrolimus and Cyclosporin A. Immunomodulatory function was dose-dependent and similar between groups. Immunosuppressants had no significant adverse effect on MSC immunomodulatory function. Discussion: MSCs from different harvest locations and donor species differ in terms of isolation yields, viability, PDT, and size. We did not detect relevant differences in immunomodulatory function with or without the presence of immunosuppressants. Human and pig O-ASC, SC-ASC and BM-MSC share similar immunomodulatory function in vitro and warrant confirmation in large animal studies. These findings should be considered in preclinical and clinical MSC applications.