Functional Characterization of Cultured Keratinocytes after Acute Cutaneous Burn Injury

In addition to forming the epithelial barrier against the outside environment keratinocytes are immunologically active cells. In the treatment of severely burned skin, cryoconserved keratinocyte allografts gain in importance. It has been proposed that these allografts accelerate wound healing also due to the expression of a favourable--keratinocyte-derived--cytokine and growth factor milieu. In this study the morphology and cytokine expression profile of keratinocytes from skin after acute burn injury was compared to non-burned skin. Skin samples were obtained from patients after severe burn injury and healthy controls. Cells were cultured and secretion of selected inflammatory mediators was quantified using Bioplex Immunoassays. Immunohistochemistry was performed to analyse further functional and morphologic parameters. Histology revealed increased terminal differentiation of keratinocytes (CK10, CK11) in allografts from non-burned skin compared to a higher portion of proliferative cells (CK5, vimentin) in acute burn injury. Increased levels of IL-1α, IL-2, IL-4, IL-10, IFN-γ and TNFα could be detected in culture media of burn injury skin cultures. Both culture groups contained large amounts of IL-1RA. IL-6 and GM-CSF were increased during the first 15 days of culture of burned skin compared to control skin. Levels of VEGF, FGF-basic, TGF-ß und G-CSF were high in both but not significantly different. Cryoconservation led to a diminished mediator synthesis except for higher levels of intracellular IL-1α and IL-1ß. Skin allografts from non-burned skin show a different secretion pattern of keratinocyte-derived cytokines and inflammatory mediators compared to keratinocytes after burn injury. As these secreted molecules exert auto- and paracrine effects and subsequently contribute to healing and barrier restoration after acute burn injury therapies affecting this specific cytokine/growth factor micromilieu could be beneficial in burned patients

Evaluation of LHP® (1% hydrogen peroxide) cream versus petrolatum and untreated controls in open wounds in healthy horses: a randomized, blinded control study

<p>Abstract</p> <p>Background</p> <p>Treatment and protection of wounds in horses can be challenging; protecting bandages may be difficult to apply on the proximal extremities and the body. Unprotected wounds carry an increased risk of bacterial contamination and subsequent infection which can lead to delayed wound healing. Topical treatment with antimicrobials is one possibility to prevent bacterial colonization or infection, but the frequent use of antimicrobials ultimately leads to development of bacterial resistance which is an increasing concern in both human and veterinary medicine.</p> <p>Methods</p> <p>Standardized wounds were created in 10 Standardbred mares. Three wounds were made in each horse. Two wounds were randomly treated with LHP^®or petrolatum and the third wound served as untreated control. All wounds were assessed daily until complete epithelization. Protocol data were recorded on day 2, 6, 11, 16, 21 and 28. Data included clinical scores for inflammation and healing, photoplanimetry for calculating wound areas and swab cytology to assess bacterial colonization and inflammation. Bacterial cultures were obtained on day 2, 6 and 16.</p> <p>Results</p> <p>Mean time to complete healing for LHP^®treated wounds was 32 days (95%CI = 26.9-37.7). Mean time to complete healing for petrolatum and untreated control wounds were 41.6 days (95%CI = 36.2-47.0) and 44.0 days (95%CI = 38.6-49.4) respectively. Wound healing occurred significantly faster in LHP^®wounds compared to both petrolatum (p = 0.0004) and untreated controls (p < 0.0001). There was no significant difference in time for healing between petrolatum and untreated controls. Total scores for bacteria and neutrophils were significantly (p < 0.0001) lower for LHP^®treated wounds compared to petrolatum from day 16 and onwards. <it>Staphylococcus aureus </it>and <it>Streptococcus zooepidemicus </it>were only found in cultures from petrolatum treated wounds and untreated controls.</p> <p>Conclusions</p> <p>Treatment with LHP^®reduced bacterial colonization and was associated with earlier complete wound healing. LHP^®cream appears to be safe and effective for topical wound treatment or wound protection.</p

An in vitro collagen perfusion wound biofilm model; with applications for antimicrobial studies and microbial metabolomics

BackgroundThe majority of in vitro studies of medically relevant biofilms involve the development of biofilm on an inanimate solid surface. However, infection in vivo consists of biofilm growth on, or suspended within, the semi-solid matrix of the tissue, whereby current models do not effectively simulate the nature of the in vivo environment. This paper describes development of an in vitro method for culturing wound associated microorganisms in a system that combines a semi-solid collagen gel matrix with continuous flow of simulated wound fluid. This enables culture of wound associated reproducible steady state biofilms under conditions that more closely simulate the dynamic wound environment. To demonstrate the use of this model the antimicrobial kinetics of ceftazidime, against both mature and developing Pseudomonas aeruginosa biofilms, was assessed. In addition, we have shown the potential application of this model system for investigating microbial metabolomics by employing selected ion flow tube mass spectrometry (SIFT-MS) to monitor ammonia and hydrogen cyanide production by Pseudomonas aeruginosa biofilms in real-time. ResultsThe collagen wound biofilm model facilitates growth of steady-state reproducible Pseudomonas aeruginosa biofilms under wound like conditions. A maximum biofilm density of 1010 cfu slide-1 was achieved by 30 hours of continuous culture and maintained throughout the remainder of the experiment. Treatment with ceftazidime at a clinically relevant dose resulted in a 1.2 – 1.6 log reduction in biofilm density at 72 hours compared to untreated controls. Treatment resulted in loss of complex biofilm architecture and morphological changes to bacterial cells, visualised using confocal microscopy. When monitoring the biofilms using SIFT-MS, ammonia and hydrogen cyanide levels peaked at 12 hours at 2273 ppb (±826.4) and 138 ppb (±49.1) respectively and were detectable throughout experimentation. ConclusionsThe collagen wound biofilm model has been developed to facilitate growth of reproducible biofilms under wound-like conditions. We have successfully used this method to: (1) evaluate antimicrobial efficacy and kinetics, clearly demonstrating the development of antimicrobial tolerance in biofilm cultures; (2) characterise volatile metabolite production by P. aeruginosa biofilms, demonstrating the potential use of this method in metabolomics studies

Burn Injury Reduces Neutrophil Directional Migration Speed in Microfluidic Devices

Thermal injury triggers a fulminant inflammatory cascade that heralds shock, end-organ failure, and ultimately sepsis and death. Emerging evidence points to a critical role for the innate immune system, and several studies had documented concurrent impairment in neutrophil chemotaxis with these post-burn inflammatory changes. While a few studies suggest that a link between neutrophil motility and patient mortality might exist, so far, cumbersome assays have prohibited exploration of the prognostic and diagnostic significance of chemotaxis after burn injury. To address this need, we developed a microfluidic device that is simple to operate and allows for precise and robust measurements of chemotaxis speed and persistence characteristics at single-cell resolution. Using this assay, we established a reference set of migration speed values for neutrophils from healthy subjects. Comparisons with samples from burn patients revealed impaired directional migration speed starting as early as 24 hours after burn injury, reaching a minimum at 72–120 hours, correlated to the size of the burn injury and potentially serving as an early indicator for concurrent infections. Further characterization of neutrophil chemotaxis using this new assay may have important diagnostic implications not only for burn patients but also for patients afflicted by other diseases that compromise neutrophil functions

A new model for preclinical testing of dermal substitutes for human skin reconstruction

BACKGROUND: Currently, acellular dermal substitutes used for skin reconstruction are usually covered with split-thickness skin grafts. The goal of this study was to develop an animal model in which such dermal substitutes can be tested under standardized conditions using a bioengineered dermo-epidermal skin graft for coverage. METHODS: Bioengineered grafts consisting of collagen type I hydrogels with incorporated human fibroblasts and human keratinocytes seeded on these gels were produced. Two different dermal substitutes, namely Matriderm(®), and an acellular collagen type I hydrogel, were applied onto full-thickness skin wounds created on the back of immuno-incompetent rats. As control, no dermal substitute was used. As coverage for the dermal substitutes either the bioengineered grafts were used, or, as controls, human split-thickness skin or neonatal rat epidermis were used. Grafts were excised 21 days post-transplantation. Histology and immunofluorescence was performed to investigate survival, epidermis formation, and vascularization of the grafts. RESULTS: The bioengineered grafts survived on all tested dermal substitutes. Epidermis formation and vascularization were comparable to the controls. CONCLUSION: We could successfully use human bioengineered grafts to test different dermal substitutes. This novel model can be used to investigate newly designed dermal substitutes in detail and in a standardized way

Analyse der Lebensqualität nach chirurgischer Spaltung des ersten Strecksehnenfachs bei Tendovaginitis stenosans de Quervain

Background: Evaluation of the quality of life following surgical release of the first extensor compartment in adult patients with de Quervain disease. Patients and methods: This retrospective study included hospital chart review and patient-reported outcome assessment using the German version of the Disabilities of the Arm, Shoulder and Hand (DASH) Outcome Measure and the validated German Michigan Hand Outcomes Questionnaire (MHQ). Results: A total of 46 patients underwent a surgical release of the first extensor compartment for de Quervain disease. Postoperative German MHQ was 78±4. Postoperative DASH score was 19±5. Satisfaction with surgery was rated positive in 85%, and 89% would undergo the procedure again. Patients with a transversal incision line had a DASH result of 11±6 and a German MHQ score of 83±7, patients with a longitudinal incision had a DASH score of 22±9 and a German MHQ score of 80±6. The patients pain level after transversal incision was markedly lower (resting pain=0.4±0.3; stress pain=1.0±0.6) than in patients operated with a longitudinal incision (resting pain=2.4±0.9; stress pain=3.1±1.1). Conclusion: The surgical approach for the treatment of de Quervain disease is associated with a high postoperative quality of life, a low postoperative morbidity and a low level of postoperative pain. A transversal incision is associated with better postoperative results than the longitudinal approach.Hintergrund: Die Zufriedenheit von Patienten nach chirurgischer Spaltung des ersten Strecksehnenfaches bei De Quervain Tendinopathie ist unbekannt. Aufgrund der weiten Verbreitung dieser ambulanten Operation stellten wir uns die Frage, wie sich die Lebensqualität nach offen chirurgischer Spaltung des ersten Strecksehnenfaches bei erwachsenen Patienten mit einer Tendovaginitis stenosans de Quervain verhält. Patienten und Methoden: Retrospektiv wurden Patientendaten sowie die Patienten-orientierten Fragebögen Michigan Hand Outcomes Questionnaire (MHQ) und Disabilities of the Arm, Shoulder and Hand (DASH) Outcome Measure ausgewertet und analysiert. Primäre Endpunkte: DASH-Ergebnis (0=keine Einschränkung, 100=maximale Einschränkung) und MHQ-Ergebnis (0=maximale Einschränkung, 100=keine Einschränkung), sekundäre Endpunkte: Schmerzen (Numerische Rating Skala (NRS) 0-10), Zufriedenheit, Narkoseform, Schnittführung, Komplikationen. Ergebnisse: Wir führten bei insgesamt 46 Patienten eine offen chirurgische Spaltung des ersten Strecksehnenfachs durch. Der postoperative MHQ-Score war 78±4 Punkte und der postoperative DASH-Score 19±5 Punkte. Die Zufriedenheit der Patienten mit dem chirurgischen Vorgehen lag bei 85%, wobei sich 89% dieser Prozedur erneut unterziehen würden. Patienten mit einer quer zum Handgelenk verlaufenden Hautschnittführung hatten einen MHQ-Score von 83±7 Punkten und einen DASH-Score von 11±6 Punkten, während Patienten mit einer Längsinzision einen MHQ-Score von 80±6 Punkten und einen DASH-Score von 22±9 Punkten aufwiesen (n.s.). Das Schmerzniveau nach querer Schnittführung (Ruheschmerz=0,4±0,3; Belastungsschmerz= 1,0±0,6) war deutlich geringer als bei Patienten mit longitudinaler Inzision (Ruheschmerz=2,4±0,9; Belastungsschmerz=3,1±1,1, n.s.). Schlussfolgerung: Die offen chirurgische Spaltung des ersten Strecksehnenfaches bei Tendovaginitis de Quervain ist mit einer hohen postoperativen Lebensqualität, einer geringen Morbidität sowie einem niedrigen Grad an postoperativen Schmerzen verbunden. Die quere Inzision zeigte zudem bessere postoperative Ergebnisse als die longitudinale Schnittführung

Untersuchung der Lebensqualität nach chirurgischer Versorgung der angeborenen und erworbenen Ringbandstenose

Background: Evaluation of quality of life following surgical release of the first annular flexor tendon pulley in children suffering from pediatric trigger thumb in comparison to adult patients with trigger finger.Material and methods: All patients who underwent an A1 tendon pulley release between 2006 and 2010 at a hand surgery center were reviewed retrospectively. Chart review included operation type, length of hospital stay, and type of anesthesia. Patient satisfaction was assessed using two validated hand questionnaires, the Disabilities of the Arm, Shoulder and Hand (DASH) Outcome Measure and the German Michigan Hand Outcomes Questionnaire (MHQ).Results: A total of 13 children and 147 adults were included in the study. No postoperative complications or recurrence were observed in the pediatric patient population. Postoperative DASH score in children was 0±0 and in adults 14±2. Postoperative German MHQ score was 97±3 in children and 84±2 in adults. Patient satisfaction was rated positive in all children and in 87% of adults; all children and 93% of the adults would undergo this procedure again. Conclusions: The surgical release of the A1 pulley in children and adults is a safe and low-risk procedure. Postoperative quality of life, especially in children, is high; patients and parents were highly satisfied. Therefore, the surgical approach should be recommended as a first-line treatment of the trigger finger in all age groups.Hintergrund: Evaluation der Lebensqualität nach chirurgischer Ringbandspaltung bei angeborener Ringbandstenose des Daumens bei Kindern (Pollex flexus congenitus) sowie erworbener Ringbandstenose bei Erwachsenen. Material und Methoden: Retrospektiv wurden allgemeine Patientendaten nach offener Ringbandspaltung zwischen 2006 und 2010 sowie die Patientenzufriedenheit mit den Patienten-orientierten Fragebögen Michigan Hand Outcomes Questionnaire (MHQ) und Disabilities of the Arm, Shoulder and Hand (DASH) Outcome Measure ausgewertet und analysiert. Ergebnisse: 13 Kinder und 147 erwachsene Patienten wurden in die Studie aufgenommen. Postoperativ kam es bei den Kindern zu keinen Komplikationen oder dem Wiederauftreten eines Schnappphänomens des Daumens. Der postoperative DASH-Score der Kinder war 0±0 Punkte und bei den Erwachsenen 14±2. Der postoperative MHQ-Score lag bei den Kindern bei 97±3 Punkten, der der Erwachsenen bei 84±2. Die postoperative Zufriedenheit wurde bei allen Kindern und bei 87% der erwachsenen Patienten als positiv gewertet. Alle Kinder sowie 93% der Erwachsenen würden sich erneut diesem operativen Eingriff unterziehen. Schlussfolgerung: Die chirurgische Intervention zur Behandlung des Pollex flexus congenitus bei Kindern sowie der Ringbandstenose bei Erwachsenen ist ein sicherer und risikoarmer Eingriff. Die postoperative Lebensqualität, speziell die der Kinder, ist hoch. Daher kann das chirurgische Vorgehen als First-Line-Therapie beim schnellenden Finger in allen Altersgruppen empfohlen werden