Genotyping of circulating cell-free DNA enables noninvasive tumor detection in myxoid liposarcomas.

Soft tissue sarcomas (STS) are rare tumors of mesenchymal origin. About 50% of patients with STS experience relapse and more than 30% will die within 10 years after diagnosis. In this study we investigated circulating free DNA (cfDNA) and tumor-specific genetic alterations therein (circulating tumor DNA, ctDNA) as diagnostic biomarkers. Plasma concentrations and fragmentation of cfDNA was analyzed with quantitative PCR. Patients with STS (n = 64) had significantly higher plasma concentrations and increased fragmentation of cfDNA when compared to patients in complete remission (n = 19) and healthy controls (n = 41) (p < 0.01 and p < 0.001). Due to overlapping values between patients with STS and controls, the sensitivity and specificity of these assays is limited. Sensitive assays to detect genomic alterations in cfDNA of synovial sarcomas (t(X;18)), myxoid liposarcomas (t(12;16) and TERT C228T promoter mutation) and well-differentiated/de-differentiated liposarcomas (MDM2 amplifications) were established. ctDNA was quantified in nine liposarcoma patients during the course of their treatment. Levels of breakpoint t(12;16) and TERT C228T ctDNA correlated with the clinical course and tumor burden in patients with myxoid liposarcomas (n = 4). ctDNA could detect minimal residual disease and tumor recurrence. In contrast, detection of MDM2 amplifications was not sensitive enough to detect tumors in patients with well-differentiated/de-differentiated liposarcomas (n = 5). Genotyping of cfDNA for tumor specific genetic alterations is a feasible and promising approach for monitoring tumor activity in patients with myxoid liposarcomas. Detection of ctDNA during follow-up examinations despite negative standard imaging studies might warrant more sensitive imaging (e.g. PET-CT) or closer follow-up intervals to timely localize and treat recurrences.

Evaluation of the Implantable Doppler Probe for Free Flap Monitoring in Lower Limb Reconstruction.

BACKGROUND: Timely reexploration and reanastomoses can salvage failing free flaps. The use of the implantable Doppler probe provides direct evidence of vascular impairment of the microvascular anastomoses and allows for postoperative NPWT. The aim of this retrospective study was to compare the Doppler probe to conventional monitoring techniques for free flap monitoring in lower limb reconstruction and to identify risk factors for perfusion disturbance and reexploration. METHODS: All patients receiving free muscle flap reconstruction for lower limb soft tissue defects at our department from 2000 to 2013 were included, and all adverse events, timely detection of perfusion problems, and outcome of revision surgery were assessed by chart analysis. RESULTS: For lower limb reconstruction, 110 free muscle transfers were performed of which 41 muscle flaps were conventionally monitored and 69 flaps were monitored using the implantable Doppler probe. In 18 cases, the free muscle flaps needed revision because of perfusion disturbances. The salvage rate was 80% with monitoring by the implantable Doppler probe compared with 62.5% using conventional monitoring methods resulting in success rates of 95.7 and 92.7%, respectively. CONCLUSION: The use of the implantable Cook-Swartz Doppler probe represents a safe monitoring method for lower limb reconstruction, which allows for the additional use of NPWT. Higher salvage and revision success rates can be attributed to an earlier detection of perfusion impairment. However, a larger patient cohort is necessary to verify superiority over conventional postoperative monitoring.

Aging Changes of the Male Lips-A Lesser Evil Than in Females?

BACKGROUND: Although aging changes of the female perioral region and aesthetic procedures for perioral rejuvenation are described in literature, specific data for the male population are lacking. This study aims to evaluate perioral aging process in the male population to compare quality and quantity of the observed changes to the female population. METHODS: Magnetic resonance imaging (MRI) scans of 22 male subjects, were recruited into 2 age groups: young (20-35 years) and old (65-80 years) and the following parameters were measured: length of visible upper lip, thickness of upper lip at the level of vermilion border, thickness of upper lip at the level of the mid lip, and cross-section area of upper lip. Cadaver transverse sections of the upper lip of 10 individuals in 2 age groups, young (<40 years, n = 5) and old (>80 years, n = 5) underwent histological analysis of the relevant anatomical structures. RESULTS: The MRI analyses does not show a statistically significant difference in upper lip lengths of old and young subjects (P = 0.213). Upper lip thickness decreases statistically significant (P < 0.001), the midline sagittal upper lip area does not show a significant change (P = 0.57). Histomorphometry shows a statistically significant decrease of cutis thickness, increase of subcutaneous fat, thinning of the orbicularis muscle, and a broadening of the orbicularis oris angle. CONCLUSIONS: The aging changes of the perioral region in men and women are similar in regard to the general processes, yet the extent of the changes is less striking in the men.

Dissociation of pentameric to monomeric C-reactive protein localizes and aggravates inflammation: in vivo proof of a powerful proinflammatory mechanism and a new anti-inflammatory strategy.

BACKGROUND: The relevance of the dissociation of circulating pentameric C-reactive protein (pCRP) to its monomeric subunits (mCRP) is poorly understood. We investigated the role of conformational C-reactive protein changes in vivo. METHODS AND RESULTS: We identified mCRP in inflamed human striated muscle, human atherosclerotic plaque, and infarcted myocardium (rat and human) and its colocalization with inflammatory cells, which suggests a general causal role of mCRP in inflammation. This was confirmed in rat intravital microscopy of lipopolysaccharide-induced cremasteric muscle inflammation. Intravenous pCRP administration significantly enhanced leukocyte rolling, adhesion, and transmigration via localized dissociation to mCRP in inflamed but not noninflamed cremaster muscle. This was confirmed in a rat model of myocardial infarction. Mechanistically, this process was dependent on exposure of lysophosphatidylcholine on activated cell membranes, which is generated after phospholipase A2 activation. These membrane changes could be visualized intravitally on endothelial cells, as could the colocalized mCRP generation. Blocking of phospholipase A2 abrogated C-reactive protein dissociation and thereby blunted the proinflammatory effects of C-reactive protein. Identifying the dissociation process as a therapeutic target, we stabilized pCRP using 1,6-bis(phosphocholine)-hexane, which prevented dissociation in vitro and in vivo and consequently inhibited the generation and proinflammatory activity of mCRP; notably, it also inhibited mCRP deposition and inflammation in rat myocardial infarction. CONCLUSIONS: These results provide in vivo evidence for a novel mechanism that localizes and aggravates inflammation via phospholipase A2-dependent dissociation of circulating pCRP to mCRP. mCRP is proposed as a pathogenic factor in atherosclerosis and myocardial infarction. Most importantly, the inhibition of pCRP dissociation represents a promising, novel anti-inflammatory therapeutic strategy.

Comparison between endothelial progenitor cells and human umbilical vein endothelial cells on neovascularization in an adipogenesis mouse model.

Volume stability and growth of tissue engineered adipose tissue equivalents using adipose-derived stem cells (ASCs) rely strongly on angiogenesis and neovascularization to support the maintenance of cells. An attractive cellular approach is based on coimplantation of endothelial cells to create a vascular network. Endothelial progenitor cells (EPCs) are a promising cell population, since they can be easily isolated from autologous human peripheral blood and thus represent a clinically feasible option. We have previously shown in in vitro and semi-in vivo studies that ASCs exert beneficial effects on EPCs in terms of enhanced tube formation and formation of blood vessels, respectively. In this study, we investigated the in vivo effects of coimplantation on endothelial cell-mediated neovascularization and ASC-mediated adipose tissue formation. For this purpose, human ASCs and human EPCs (or HUVECs as direct comparison to EPCs) were suspended alone or in coculture in fibrin and subcutaneously injected into the back of athymic nude mice and explanted after 1, 3 or 6months. Our results show that monocultures of EPCs or HUVECs were not able to perform vasculogenesis and constructs exhibited complete resorption already after 1month. However, a remarkable difference between EPCs and HUVECs was detected when coimplanted with ASCs. While coimplanted HUVECs gave rise to a stable neovasculature which was characterized by perfusion with erythrocytes, coimplanted EPCs showed no ability to form vascular structures. In the case of HUVEC-derived neovasculature, coimplanted ASCs displayed perivascular properties by stabilizing these neovessels. However, formation of human adipose tissue was independent of coimplanted endothelial cells. Our results indicate that HUVECs are superior to EPCs in terms of promoting in vivo neovascularization and recruiting perivascular cells for vessel stabilization when coimplanted with ASCs.

In Situ Deactivation of Interleukin-6 Enhances Early Peripheral Nerve Regeneration in a Murine Injury Model.

BACKGROUND: Systemic alteration of interleukin-6 (IL-6) influences peripheral nerve regeneration. We investigated the potential influences of in situ (at the coaptation site) IL-6 modulation in a peripheral-nerve-transection/sciatic-nerve-graft in vivo model. METHODS: We quantified the elongation of regenerating axons, the number of arborizing axons, and the number of branches per arborizing axon 7 days after the injury in mice expressing axonal fluorescent proteins (thy-1-YFP mice). Sciatic nerves from nonexpressing mice (C57Bl6 or IL-6(-/-) mice) were grafted into those expressing yellow fluorescent protein. We altered the in situ IL-6 concentration by loading a topical gelatin sponge with an inhibiting IL-6 receptor antibody or IL-6 combined with a soluble IL-6 receptor. Sciatic nerves from IL-6(-/-) mice were grafted into an additional group. The contralateral sham-operated side served as control in all the groups. RESULTS: Axonal elongation increased significantly with the in situ application of the IL-6 receptor antibody, while topical IL-6 significantly reduced the regeneration distance. The number of arborizing axons increased significantly in nerves grafted from IL-6(-/-) mice, whereas branches per arborizing axons remained stable. CONCLUSION: In situ IL-6 receptor inhibition and IL-6(-/-) nerve grafting enhance early peripheral nerve regeneration in an acute murine injury model.

Comparison of Schwann cells and olfactory ensheathing cells for peripheral nerve gap bridging.

INTRODUCTION: Previously, we introduced the biogenic conduit (BC) as a novel autologous nerve conduit for bridging peripheral nerve defects and tested its regenerative capacity in a short- and long-term setting. The aim of the present study was to clarify whether intraluminal application of regeneration-promoting glial cells, including Schwann cells (SC) and olfactory ensheathing cells (OEC), displayed differential effects after sciatic nerve gap bridging. MATERIAL AND METHODS: BCs were generated as previously described. The conduits filled with fibrin/SC (n = 8) and fibrin/OEC (n = 8) were compared to autologous nerve transplants (NT; n = 8) in the 15-mm sciatic nerve gap lesion model of the rat. The sciatic functional index was evaluated every 4 weeks. After 16 weeks, histological evaluation followed regarding nerve area, axon number, myelination index and N ratio. RESULTS: Common to all groups was a continual improvement in motor function during the observation period. Recovery was significantly better after SC transplantation compared to OEC (p < 0.01). Both cell transplantation groups showed significantly worse function than the NT group (p < 0.01). Whereas nerve area and axon number were correlated to function, being significantly lowest in the OEC group (p < 0.001), both cell groups showed lowered myelination (p < 0.001) and lower N ratio compared to the NT group. DISCUSSION: SC-filled BCs led to improved regeneration compared to OEC-filled BCs in a 15-mm-long nerve gap model of the rat.

The long-term functional recovery of repair of sciatic nerve transection with biogenic conduits.

INTRODUCTION: The aim of this study was to evaluate long-term regenerative capacity over a 15-mm nerve gap of an autologous nerve conduit, the biogenic conduit (BC), 16 weeks after sciatic nerve transection in the rat. METHODS: A 19-mm long polyvinyl chloride (PVC) tube was implanted parallely to the sciatic nerve. After implantation, a connective tissue cover developed around the PVC-tube, the so-called BC. After removal of the PVC-tube the BCs filled with fibrin (n = 8) were compared to autologous nerve grafts (n = 8). Sciatic functional index (SFI) was evaluated every 4 weeks, histological evaluation was performed at 16 weeks postimplantation. Regenerating axons were visualized by retrograde labelling. RESULTS: SFI revealed no significant differences. Nerve area and axon number in the BC group were significantly lower than in the autologous nerve group (P < 0.05; P < 0.01). Analysis of myelin formation showed no significant difference in both groups. Analysis of N-ratio revealed lower values in the BC group (P < 0.001). CONCLUSION: This study reveals the suitability of BC for nerve gap bridging over a period of 16 weeks with functional recovery to comparable extent as the autologous nerve graft despite impaired histomorphometric parameters.

An in vivo engineered nerve conduit--fabrication and experimental study in rats.

BACKGROUND: Several types of nerve conduits have been used for peripheral nerve gap bridging. This study investigated the in vivo engineering of a biological nerve conduit and its suitability for nerve gap bridging. MATERIAL AND METHODS: A 19-mm long polyvinyl chloride (PVC) tube was implanted parallely to the sciatic nerve. After implantation, a connective tissue cover developed around the PVC-tube, the so-called biogenic conduit. Histological cross-sections were performed after 1, 2, 3, and 4 weeks. Wall thicknesses were measured and all vessels per cross-section were counted. The biogenic conduit filled with fibrin was used to bridge a 15-mm long nerve gap in the sciatic lesion model of the rat (n = 8). The results of nerve repair with the conduit were compared to the autologous nerve graft (n = 8). Sciatic functional index (SFI), nerve area, axon count, myelination index, and ratio of total myelinated fiber area/nerve area (N-ratio) were analyzed after 4 weeks. RESULTS: The wall thickness of biogenic conduits increased over the 4 weeks implantation time. Biogenic conduits revealed highest number of vessels per cross-section after 4 weeks. The results of SFI analysis did not show significant difference between the repairs with biogenic conduit and autologous nerve graft. Nerve area and axon count in the biogenic conduit group were significantly lower than in the autologous nerve group (P < 0.001). The biogenic conduit group showed significant higher myelination values, but lower N-ratio when compared to the nerve graft group (P < 0.001). CONCLUSIONS: The in vivo engineered conduits allow nerve gap bridging of 15 mm. However, quality of regeneration after 4 weeks observation time is not comparable to autologous nerve grafts. Whether biogenic conduits might be a suitable alternative to artificial and biological conduits for gap bridging will have to be evaluated in further studies.

Ten years stable internal fixation of metacarpal and phalangeal hand fractures-risk factor and outcome analysis show no increase of complications in the treatment of open compared with closed fractures.

BACKGROUND: : Stable internal screw/plating systems for hand fractures have evolved during the last 20 years. The improved versatility leads to the increased use of these materials in open fractures, with the benefit of early mobilization. The aim of this retrospective study is to discern whether the broadening of the indications for these implants is accompanied by increased complication rates. METHODS: : Data from 365 patients treated during the last 10 years at our department for metacarpal or phalangeal fractures with stable internal fixation by screw or plate were gathered and analyzed. RESULTS: : Uneventful bony consolidation was observed in 91.2% (n = 333). The functional results were excellent to acceptable in 85.2%, whereas in 14.8% (n = 54), the result was unsatisfactory, the latter group presenting with concominant soft tissue injury. There was no statistically significant difference in infection and nonunion rates when comparing open and closed fractures. CONCLUSION: : These results confirm that most patients with open metacarpal and phalangeal fractures can be treated by stable internal fixation.

A new evaluation tool for monitoring devices and its application to evaluate the implantable Doppler probe.

Evaluation parameters for free flap monitoring devices are used inconsistently, leading to considerable confusion about the quality and applicability of these devices. A comparison of different systems and different clinical series is almost impossible. The ultimate questions that need to be answered are those regarding the efficiency and the effectiveness of the system. A new tool consisting of two simple parameters that sufficiently and comparably describe the quality of monitoring devices is developed. The flap failure reduction rate describes the percentage of saved flaps (effectiveness). The revision success rate describes the efficiency. Literature reevaluation shows inconsistent results, although all authors describe a positive experience. This shows the limited value of the classical parameters. Larger studies have flap failure reduction rates of 5 to 12% (5 to 12% of monitored flaps are saved). Revision success rates of 75 to 90% prove that the system is efficient enough in daily use. Reevaluation of the smaller reported series result in lower parameters, which shows that there is a significant learning curve for this device. The new parameters alleviate the confusion surrounding evaluation of monitoring systems by giving specific information about effectiveness and efficiency. The benefits of the implanted Doppler probe can now be clearly described. However, in several studies the benefit of the system is overrated.

The use of the vacuum-assisted closure in microsurgical reconstruction revisited: application in the reconstruction of the posttraumatic lower extremity.

Introduction of vacuum-assisted closure (VAC) system into clinical practice has revolutionized wound care. Despite its multiple advantages, however, the VAC is only rarely used in the setting of microsurgical reconstruction. Concerns have been the inability to clinically monitor the flap as well the possibility of flap compression by the device. The authors put their postoperative treatment concept of applying the VAC to free flaps to the test by reviewing their experience with this concept in patients undergoing microsurgical reconstruction of posttraumatic lower-extremity soft tissue defects. Twenty-six patients (22 male, 4 female) were included in this study. Use of the implantable Doppler probe allowed for postoperative flap monitoring. Two flap failures were observed, both in patients with peripheral vascular disease. In conclusion, using the VAC device in the setting of microsurgical reconstruction is safe and allows for increased patient comfort.

Evidence-based plastic surgery: controlled trials in three plastic surgical journals (1990-2005).

Although a myriad of original articles is published annually in plastic surgical journals, assessment of the level of evidence-based medicine has rarely been conducted. A hand search was conducted identifying randomized controlled trials (RCTs) and controlled clinical trials in 3 plastic surgical journals from 1990-2005. The quality of reporting was assessed and additional parameters investigated including report of statistical significance, type of institution, and country affiliation of the first author. Of the 9428 original articles that were analyzed, 172 and 139 articles met the inclusion criteria for RCTs and controlled clinical trials, respectively. Fifty-nine RCTs reported on successful double-blinding with only 20 RCTs reporting the allocation concealment appropriately. Description of participant drop-outs was detected in 64 RCTs and a statistically significant result was reported in 118 RCTs. The annual publication of controlled trials has increased over the last 16 years, with the majority of controlled trials being from North-America and Europe. Execution and publication of controlled trials has increased in the plastic surgical literature. However, the quality of reporting deserves improvement.

A self-made, low-cost infrared system for evaluating the sciatic functional index in mice.

The sciatic functional index (SFI) is a popular parameter for peripheral nerve evaluation that relies on footprints obtained with ink and paper. Drawbacks include smearing artefacts and a lack of dynamic information during measurement. Modern applications use digitized systems that can deliver results with less analytical effort and fewer mice. However, the systems are expensive (€40,000). This study aimed to evaluate the applicability and precision of a self-made, low-cost infrared system for evaluating SFI in mice. Mice were subjected to unilateral sciatic nerve crush injury (crush group; n = 7) and sham operation (sham group; n = 4). They were evaluated on the day before surgery, the 2(nd), 4(th) and 6(th) days after injury, and then every day up to the 23(rd) day after injury. We compared two SFI evaluation methods, i.e., conventional ink-and-paper SFI (C-SFI) and our infrared system (I-SFI). Our apparatus visualized footprints with totally internally reflected infrared light (950 nm) and a camera that can only detect this wavelength. Additionally we performed an analysis with the ladder beam walking test (LBWT) as a reference test. I-SFI assessment reduced the standard deviation by about 33 percent, from 11.6 to 7.8, and cut the variance around the baseline to 21 percent. The system thus requires fewer measurement repetitions and fewer animals, and cuts the cost of keeping the animals. The apparatus cost €321 to build. Our results show that the process of obtaining the SFI can be made more precise via digitization with a self-made, low-cost infrared system.

Lymphangiosarcoma: Is Stewart-Treves Syndrome a Preventable Condition?

BACKGROUND: Stewart-Treves syndrome is a rare complication of breast cancer treatment, representing a lymphangiosarcoma commonly associated with lymphedema and severely impacting patient's outcome. The tumor typically develops in the atrophic, pachydermatous, hyperkeratotic skin of limbs affected by long-standing lymphedema. Clinical data associated with Stewart-Treves syndrome and lymphedema management have rarely been published. METHODS AND RESULTS: In the period between 1980 and 2009, ten patients with Stewart-Treves syndrome were diagnosed and treated at the Foeldiklinik, Hinterzarten, Germany. Nine of the ten patients were female. Five patients had previously suffered from breast cancer (and were treated with mastectomy); two from other malignancies; two patients had primary lymphedema, and one had undergone lower extremity lymphadenectomy. All cancer patients had undergone radiation treatment. In all cases, the sarcoma developed in non-irradiated areas 6-48 years (average 16.3 years) after the onset of lymphedema. None of the patients had received complex decongestive physical therapy (CDT). Two patients had above-elbow amputation, one had shoulder exarticulation, two patients had wide excision and skin grafting, two patients had above-knee amputation procedure, two patients had a below-knee amputation procedure, and one patient had no surgical treatment at all. The time to recurrence after surgery, time to metastasis, patient survival and CDT were recorded. CONCLUSIONS: Patients with lymphedema should be closely examined starting 5 years from the time of lymphedema onset, paying special attention to those with associated malignancies. Only early diagnosis and treatment by radical ablative surgery confers a reasonable prognosis with this rare but aggressive disease. A potential effect of CDT on lymphangiosarcoma has to be studied in a greater patient cohort.

Unusual explosive growth of a squamous cell carcinoma of the scalp after electrical burn injury and subsequent coverage by sequential free flap vascular connection--a case report.

BACKGROUND: Squamous cell carcinomos may arise from chronic ulcerating wounds in scars, most commonly postburn scars. Tumour growth usually takes place over months to years. Localization on the scalp is a relatively rare condition. CASE PRESENTATION: This report presents the case of a 63-year-old man with chronic ulceration of a postburn scar of the scalp due to an electrical burn 58 years ago. Sudden tumour growth started within weeks and on presentation already had extended through the skull into frontal cortex. After radical tumour resection, defect was covered with a free radial forearm flap. Local recurrence occurred 6 weeks later. Subsequent wide excision including discard of the flap and preservation of the radial vessels was followed by transfer of a free latissimus dorsi muscle flap, using the radial vessels of the first flap as recipient vessels. The patient received radiotherapy post-operatively. There were no problems with flap survivals or wound healing. Due to rapidly growing recurrence the patient died 2 months later. CONCLUSION: Explosive SCC tumour growth might occur in post-burn scars after more than 50 years. As a treatment option the use of sequential free flap connections might serve in repeated extensive tumour resections, especially in the scalp region, where suitable donor vessels are often located in distance to the defect.

Single-stage dynamic reanimation of the smile in irreversible facial paralysis by free functional muscle transfer.

Unilateral facial paralysis is a common disease that is associated with significant functional, aesthetic and psychological issues. Though idiopathic facial paralysis (Bell's palsy) is the most common diagnosis, patients can also present with a history of physical trauma, infectious disease, tumor, or iatrogenic facial paralysis. Early repair within one year of injury can be achieved by direct nerve repair, cross-face nerve grafting or regional nerve transfer. It is due to muscle atrophy that in long lasting facial paralysis complex reconstructive methods have to be applied. Instead of one single procedure, different surgical approaches have to be considered to alleviate the various components of the paralysis. The reconstruction of a spontaneous dynamic smile with a symmetric resting tone is a crucial factor to overcome the functional deficits and the social handicap that are associated with facial paralysis. Although numerous surgical techniques have been described, a two-stage approach with an initial cross-facial nerve grafting followed by a free functional muscle transfer is most frequently applied. In selected patients however, a single-stage reconstruction using the motor nerve to the masseter as donor nerve is superior to a two-stage repair. The gracilis muscle is most commonly used for reconstruction, as it presents with a constant anatomy, a simple dissection and minimal donor site morbidity. Here we demonstrate the pre-operative work-up, the post-operative management, and precisely describe the surgical procedure of single-stage microsurgical reconstruction of the smile by free functional gracilis muscle transfer in a step by step protocol. We further illustrate common pitfalls and provide useful tips which should enable the reader to truly comprehend the procedure. We further discuss indications and limitations of the technique and demonstrate representative results.

Chorioallantoic membrane angiogenesis model for tissue engineering: a new twist on a classic model.

Tissue-engineering (TE) applications include the isolation, culture, and seeding of cells into a suitable matrix or scaffold before in vivo transplantation. After transplantation, vascularization of the scaffold is a principal limiting factor for cell viability for the first 6-8 days posttransplantation. A model for systematic analysis of this process has been developed. Fertilized White Leghorn eggs were incubated (at 37.8 degrees C in 60% relative humidity) and opened on day 3 of incubation. Preadipocyte-seeded fibrin constructs were implanted in a specially designed plastic cylinder and placed through the opening on the surface of the chorioallantoic membrane (CAM) on day 8 of incubation. Vascularization of the constructs by chorioallantoic blood vessels was assessed for up to 8 days posttransplantation. The survival rate for embryos receiving transplanted constructs was about 90%. Histology confirmed transplant cell viability at day 4 posttransplantation and vascularization of the constructs by avian endothelial cells began at this time. A new in vivo model to study the effect of angiogenesis in TE constructs, including assessments of viability, proliferation, and differentiation of transplanted cells and biomaterial properties, is presented. Advantages include easy access to the vascular network of the CAM, lack of immunocompetence, low costs, and avoidance of animal experiments.

Infraareolar pectoralis major myocutaneous island flap as treatment of first choice for deep sternal wound infection.

Deep sternal wound infection (DSWI) is a grave complication of median sternotomy, associated with high morbidity, mortality and escalating treatment costs. There is general consensus that optimal treatment comprises radical debridement followed by coverage with a vascularised flap. However, there is ongoing debate regarding the ideal operative procedure. We present our experience with the infraareolar pectoralis major island myocutaneous flap (PEC-MI flap) as treatment of first choice in DSWI. Following a retrospective chart review, data pertaining to patient demographics, type of cardiac surgery performed, prevalence of known DSWI risk factors, identified pathogens, duration of surgery, flap-related complications, duration of hospital stay and antibiotic therapy, as well as mortality were noted. Additionally, we describe the operative technique and review the relevant literature. Twenty-five patients underwent coverage with the PEC-MI flap in our department. The average age was 69.2 years. Nineteen patients underwent coronary artery bypass surgery, 10 valve replacement, two aortic replacement surgery and one pericardiectomy. In six cases, no internal mammary artery was used in cardiac surgery, in 11 cases one and in seven cases both internal mammary arteries were used. The average duration of surgery was 154.2 min and the average hospital stay was 28.4 days. Complications which required revision surgery were haematoma in three cases, one wound dehiscence and one recurrent infection. Two cases required coverage with an additional regional flap. The PEC-MI flap has been used as flap of first choice in our clinic for treatment of DSWI. It is sufficient to raise the flap unilaterally, and it does not require skin grafting. The combination of immunocompetent bulky muscle tissue used to obliterate the sternal cavity and the large skin paddle enabling a low-tension skin closure allows reliable and efficient treatment of this severe complication.

Real-time digital imaging of leukocyte-endothelial interaction in ischemia-reperfusion injury (IRI) of the rat cremaster muscle.

Ischemia-reperfusion injury (IRI) has been implicated in a large array of pathological conditions such as cerebral stroke, myocardial infarction, intestinal ischemia as well as following transplant and cardiovascular surgery. Reperfusion of previously ischemic tissue, while essential for the prevention of irreversible tissue injury, elicits excessive inflammation of the affected tissue. Adjacent to the production of reactive oxygen species, activation of the complement system and increased microvascular permeability, the activation of leukocytes is one of the principle actors in the pathological cascade of inflammatory tissue damage during reperfusion. Leukocyte activation is a multistep process consisting of rolling, firm adhesion and transmigration and is mediated by a complex interaction between adhesion molecules in response to chemoattractants such as complement factors, chemokines, or platelet-activating factor. While leukocyte rolling in postcapillary venules is predominantly mediated by the interaction of selectins with their counter ligands, firm adhesion of leukocytes to the endothelium is selectin-controlled via binding to intercellular adhesion molecules (ICAM) and vascular cellular adhesion molecules (VCAM). Gold standard for the in vivo observation of leukocyte-endothelial interaction is the technique of intravital microscopy, first described in 1968. Though various models of IRI (ischemia-reperfusion injury) have been described for various organs, only few are suitable for direct visualization of leukocyte recruitment in the microvascular bed on a high level of image quality. We here promote the digital intravital epifluorescence microscopy of the postcapillary venule in the cremasteric microcirculation of the rat as a convenient method to qualitatively and quantitatively analyze leukocyte recruitment for IRI-research in striated muscle tissue and provide a detailed manual for accomplishing the technique. We further illustrate common pitfalls and provide useful tips which should enable the reader to truly appreciate, and safely perform the method. In a step by step protocol we depict how to get started with respiration controlled anesthesia under sufficient monitoring to keep the animal firmly anesthetized for longer periods of time. We then describe the cremasteric preparation as a thin flat sheet for outstanding optical resolution and provide a protocol for leukocyte imaging in IRI that has been well established in our laboratories.