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Sarcomas are defined as a group of mesenchymal malignancies with over 100 heterogeneous subtypes. As a rare and difficult to diagnose entity, micrometastasis is already present at the time of diagnosis in many cases. Current treatment practice of sarcomas consists mainly of surgery, (neo)adjuvant chemo- and/or radiotherapy. Although the past decade has shown that particular genetic abnormalities can promote the development of sarcomas, such as translocations, gain-of-function mutations, amplifications or tumor suppressor gene losses, these insights have not led to established alternative treatment strategies so far. Novel therapeutic concepts with immunotherapy at its forefront have experienced some remarkable success in different solid tumors while their impact in sarcoma remains limited. In this review, the most common immunotherapy strategies in sarcomas, such as immune checkpoint inhibitors, targeted therapy and cytokine therapy are concisely discussed. The programmed cell death (PD)-1/PD-1L axis and apoptosis-inducing cytokines, such as TNF-related apoptosis-inducing ligand (TRAIL), have not yielded the same success like in other solid tumors. However, in certain sarcoma subtypes, e.g. liposarcoma or undifferentiated pleomorphic sarcoma, encouraging results in some cases when employing immune checkpoint inhibitors in combination with other treatment options were found. Moreover, newer strategies such as the targeted therapy against the ancient cytokine macrophage migration inhibitory factor (MIF) may represent an interesting approach worth investigation in the future.

Background Lipedema is a common adipose tissue disorder affecting women, characterized by a symmetric subcutaneous adipose tissue deposition, particularly of the lower extremities. Lipedema is usually underdiagnosed, thus remaining an undertreated disease. Importantly, no histopathologic or molecular hallmarks exist to clearly diagnose the disease, which is often misinterpreted as obesity or lymphedema. Materials and methods The aim of the present study is to characterize in detail morphologic and molecular alterations in the adipose tissue composition of lipedema patients compared with healthy controls. Detailed histopathologic and molecular characterization was performed using lipid and cytokine quantification as well as gene expression arrays. The analysis was conducted on anatomically matched skin and fat tissue biopsies as well as fasting serum probes obtained from 10 lipedema and 11 gender and body mass index–matched control patients. Results Histologic evaluation of the adipose tissue showed increased intercellular fibrosis and adipocyte hypertrophy. Serum analysis showed an aberrant lipid metabolism without changes in the circulating adipokines. In an adipogenesis gene array, a distinct gene expression profile associated with macrophages was observed. Histologic assessment of the immune cell infiltrate confirmed the increased presence of macrophages, without changes in the T-cell compartment. Conclusions Lipedema presents a distinguishable disease with typical tissue architecture and aberrant lipid metabolism, different to obesity or lymphedema. The differentially expressed genes and immune cell infiltration profile in lipedema patients further support these findings.

BACKGROUND Chylous leak is an uncommon complication after head and neck surgery and typically results from a lesion of the thoracic duct (TD). Beside conservative treatment, different minimally invasive and surgical procedures exist, of which almost all lead to a total closure of the TD. METHODS We report on a rare case of microsurgical lymphovenous anastomosis to treat a TD lesion. An additional systematic review on surgical procedures to treat TD lesions with special attention to lymphovenous anastomoses was performed according to the PRISMA guidelines. RESULTS A 52-year-old patient with a chylous fistula after modified radical neck dissection was successfully treated by a lymphovenous anastomosis of the TD and external jugular vein with additional coverage by sternocleidomastoid muscle flap. The patient showed a complete resolution of chylous leak with an uneventful postoperative course.The systematic search of literature yielded 684 articles with 4 case reports on lymphovenous anastomosis in chylous leak with a high success rate. Other surgical techniques include transcervical, thoracoscopic, or video-assisted thoracoscopic TD ligation, either alone or combined with a local muscle flap. CONCLUSIONS Lymphovenous anastomosis of the TD is a feasible and safe technique allowing for treatment of cervical TD lesions, especially if minimally invasive procedures fail. Compared with other techniques, lymphatic circulation can successfully be maintained.

After skin tissue injury or pathological removal, vascularization timing is paramount in graft survival. As full thickness skin grafts often fail to become perfused over larger surfaces, split-thickness grafts are preferred and can be used together with biomaterials, which themselves are non-angiogenic. One way of promoting vascular ingrowth is to “pre-vascularize” an engineered substitute by introducing endothelial cells (ECs). Since it has been previously demonstrated that surface structured biomaterials have an effect on wound healing, skin regeneration, and fibrosis reduction, we proposed that a microvascular-rich lipoconstruct with anisotropic topographical cues could be a clinically translatable vascularization approach. Murine lipofragments were formed with three polydimethylsiloxane molds (flat, 5 µm, and 50 µm parallel gratings) and implanted into the dorsal skinfold chamber of male C57BL/6 mice. Vascular ingrowth was observed through intravital microscopy over 21 days and further assessed by histology and protein identification. Our investigation revealed that topographical feature size influenced the commencement of neovascular ingrowth, with 5 µm gratings exhibiting early construct perfusion at 3 days post-operation, and 50 µm being delayed until day 5. We therefore postulate that surface structured lipoconstructs may serve as an easily obtained and produced construct suitable for providing soft tissue and ECs to tissue defects. Statement of Significance Skin graft failures due to inadequate or uneven perfusion frequently occur and can be even more complicated in deep, difficult to heal wounds, or bone coverage. In complex injuries, biomaterials are often used to cover bone structures with a standard split thickness graft; however, perfusion can take up to 3 weeks. Thus, any means to promote faster and uniform vascularization could significantly reduce healing time, as well as lower patient down-time. As pre-vascularized constructs have reported success in research, we created a cost-efficient, translatable method with no additional laboratory time as adipose tissue can be harvested and used immediately. We further used surface topography as an aspect to modulate construct perfusion, which has been reported for the first time here.

BACKGROUND/AIM: Among the most important prognostic factors in melanoma is the sentinel lymph node (SLN) status. MATERIALS AND METHODS: Using our electronic database we identified 109 of 890 SLN-negative patients with progressive disease (PD). These patients were characterized for melanoma type, molecular type, sequence and extent of metastatic spread. RESULTS: A total of 61 of 109 SLN-negative patients had PD in the SLN-basin indicating false-negative SLN (group-1). Forty eight of 109 patients had PD at distant sites and were therefore impossible to be identified using SLN biopsy (group-2). Despite distant spread these patients had significantly more single organ metastasis (p

Objective Lymphedema is a chronic, progressive and burdensome disease that is known to have a substantial impact on quality of life (QOL). Hence, the assessment of QOL is an important aspect of any study which seeks to evaluate outcomes after lymph reconstructive surgery. We therefore aimed to analyze currently available patient-reported outcome measurements (PROMS) for patients with lower limb lymphedema (LLL) with regard to their psychometric properties. Furthermore, we intended to evaluate the change in QOL in patient undergoing lymphovenous anastomosis (LVA) or vascularized lymph node transfer (VLNT). Methods A literature research of four databases on studies that included PROMS for LLL was conducted. All selected studies were assessed for validity according Consensus-based Standards for the selection of Health Measurement Instruments. Studies that used the Lymph Quality of Life Measure for Limb Lymphedema (LYMQOL) after LVA and/or VLNT were included for quantitative analysis. Results In total, 988 studies were screened, of which 30 studies were included in this review. We identified six validated PROMS to assess LLL. Of those, the LYMQOL was the most commonly used questionnaire. Based on the LYMQOL, a significant improvement of QOL in LLL was noted in all studies after reconstructive lymph surgery (LVA/VLNT) regardless of lymphedema etiology, stage, or time since diagnosis. On the basis of the Consensus-based Standards for the selection of Health Measurement Instruments criteria, the Lymphoedema Functioning, Disability and Health Questionnaire for Lower Limb Lymphoedema seems to be superior in terms of its psychometric properties. Conclusions A significant improvement in the QOL in patients with LLL after reconstructive lymph surgery can be observed. Future studies on reconstructive lymph surgery need to include both objective and congruent volume measurements, as well as data on QOL based a well-validated PROM such as the Lymphoedema Functioning, Disability and Health Questionnaire for Lower Limb Lymphoedema.

BACKGROUND Lymphoedema is a progressive and potentially disabling disease. A growing number of studies show promising clinical results after microsurgical reconstruction. However, this treatment is currently not supported by level 1 evidence and insurance coverage is variable. METHODS Electronic records of 55 patients with limb lymphoedema, who were eligible for lymphovenous bypass surgery and/or lymphatic tissue transfer in our department from 2017 to 2020, were reviewed. Correspondence between our department and health insurers was analysed. A web-based search and individual telephone interviews were conducted to identify health insurer policies. RESULTS We included 42 patients undergoing 46 operations and evaluated the correspondence between our department and nine different health insurers. Overall, reimbursement of costs was approved in 67% (n = 31) of all surgeries and was refused in 33% (n = 15). The mean number of applications for reconsideration sent to insurers was 1.3 ± 0.7. The time between confirmation of the indication and the final decision ranged from 6 to 300 days (mean 50 days). Reimbursement of cost coverage ranged from 0% to 100% depending on the individual insurance company. No insurance company had policies publicly available online and all stated that they determine coverage only when provided with specific patient details on a case-by-case basis. CONCLUSION Insurance companies in Switzerland do not have a uniform policy regarding cost coverage for lymphatic surgery procedures. Moreover, the decision process appeared to be rather uniform within the respective insurance company and independent of the individual case. Standardised evaluation criteria including patient reported outcome measures should be developed to underscore the beneficial effects of lymphatic surgery and facilitate insurance coverage.

ZusammenfassungIm Frühstadium von Nervenläsionen stellt die klinische Differenzierung zwischen Neurapraxie, Axonotmesis und Neurotmesis oftmals eine große Herausforderung dar. Vor allem im Frühstadium von Nervenläsionen ist eine korrekte Klassifizierung des Schädigungstyps jedoch von essenzieller Bedeutung, da hiermit das therapeutische Konzept, insbesondere das operative Vorgehen und damit die Prognose, maßgebend bestimmt werden. Neben einer ausführlichen klinischen Statuierung und Anamneseerhebung bedarf es zur Präzisierung der Diagnose den Einsatz ergänzender elektrophysiologischer (funktioneller) und/oder bildgebender Untersuchungen. Eine elektrophysiologische Diagnostik kann hierbei Auskunft über Lokalisation, Schweregrad, Verlauf, Schädigungstyp und einer beginnenden oder stattgehabten Reinnervation geben. Präoperativ sollten hinsichtlich der funktionellen Diagnostik eine Neurografie, Nadel-Elektromyografie (EMG) und ggf. evozierte Potentiale (EP) und bildgebend eine Nervensonografie oder Magnetresonanztomografie (MRT) durchgeführt werden. Ergänzend kann eine EMG auch intraoperativ erfolgen.

Background:. Numerous approaches have been employed to treat chronic lymphocele and cutaneous lymphatic fistulas (LFs) with little success. Given a high incidence and substantial consequences for patients, there is an ongoing demand for effective therapeutic and preventive strategies. The aim of this study was to evaluate the results after microscopic lymphatic ligation (MLL) and lymphovenous anastomosis (LVA) as a therapeutic and preventive approach in this context. Methods:. Demographic data, surgical characteristics, complications, and the overall outcome of all patients undergoing surgery for postoperative LF from 2014 to 2019 were collected retrospectively. Patients were categorized in accordance with predefined inclusion/exclusion criteria and with their treatment. Statistical analysis was conducted using descriptive, summary statistics to identify a central tendency. Results:. Thirty-four patients underwent indocyanine-green-lymphangiography guided revision surgery for LF. Two patients were lost to follow-up at 6 months. LF was successfully treated in all patients (n = 32) with a multimodal approach. Only MLL was performed in 22 patients and MLL/LVA in 10 patients. LF resolved in 78% of all patients with MLL only or MLL/LVA. In the remaining 22%, LF resolved after additional sclerotherapy within 3 months. Conclusions:. Treatment of LF should follow a standardized staged surgical approach to optimize outcome. LF was treated successfully in all our patients. We therefore propose a multimodal interdisciplinary approach to this common clinical problem that includes adjunctive sclerotherapy.

OBJECTIVE: This study aimed to determine the benefits of sclerotherapy with OK-432 for the treatment of postoperative chronic lymphocele. BACKGROUND: Postoperative chronic lymphocele formation is common and accounts for a high postoperative morbidity. Nonsurgical strategies comprise repetitive percutaneous fluid aspiration or percutaneous sclerotherapy. OK-432 has been used to treat congenital lymphatic malformations with several reports of promising results. We hypothesized that it is more beneficial than repetitive percutaneous fluid aspiration for the treatment of symptomatic lymphocele. METHODS: Two cohorts of melanoma patients who developed recurrent lymphocele after lymph node dissection from January 2013 to August 2017 were compared. The first cohort was treated with repetitive percutaneous fluid aspiration (n = 20). The second cohort received OK-432 sclerotherapy (n = 20). Primary end points were overall treatment duration, number of treatment sessions, and the clinical success in both cohorts. Secondary end points were surgical site infection rate, need for additional antibiotic treatment, wound healing disorders, and the need for revision surgery. RESULTS: Mean overall duration of treatment with sclerotherapy was significantly shorter than with repetitive aspiration (9.4 ± 7.2 vs 47.5 ± 31.9 days, P < 0.01). Mean number of sclerotherapy treatment sessions were 2.5 ± 1.2. Clinical success with OK-432 was 19 of 20, and that with repeated aspiration was 7 of 20 (χ = 15.82, P < 0.001). No surgical site infection occurred in the sclerotherapy cohort, which was significantly lower than those treated with repetitive aspiration (P < 0.03). Surgical revision was mandatory in 12 of 20 patients who were treated with repetitive aspiration, and only 1 of 20 patients in the sclerotherapy cohort. CONCLUSION: Sclerotherapy with OK-432 for the treatment of postoperative lymphocele is highly beneficial with a significant reduction of morbidity and the overall treatment time compared with repetitive aspiration.

Zusammenfassung. Die körperdysmorphe Störung (KDS) zählt zu den somatoformen Störungen und ist durch ein übermäs­siges Beschäftigen mit einem oder mehreren wahrgenommenen «Schönheitsmakeln» gekennzeichnet. Typischerweise liegt der Fokus auf dem Gesichtsbereich, beispielsweise der Hautbeschaffenheit oder der Form von Gesichtsteilen. Neben charakteristischen Verhaltensweisen, wie einem ständigen Kontrollieren des Defektes im Spiegel, erzeugt die Erkrankung einen hohen Leidensdruck bei den Betroffenen, welcher nicht selten mit einem sozialen Rückzug einhergeht. Im Patienten- klientel des Plastischen Chirurgen hat das Erkrankungsbild mit einer Prävalenz von bis zu 20 % eine hohe Bedeutung, zumal Operationen mit einer Zunahme der Beschwerden einhergehen können. Wir empfehlen daher, Patienten, welche einen plastisch-ästhetischen Eingriff wünschen, ­während der Anamnese im Hinblick auf die typischen Symptome und Verhaltensweisen zu beobachten und einzelne Fragen aus den bekannten Screening-Fragebögen in das Gespräch einfliessen zu lassen, um die die Erkrankung rechtzeitig zu erkennen.

Lipedema is a chronic adipose tissue disorder characterized by the disproportional subcutaneous deposition of fat and is commonly misdiagnosed as lymphedema or obesity. The molecular determinants of the lipedema remain largely unknown and only speculations exist regarding the lymphatic system involvement. The aim of the present study is to characterize the lymphatic vascular involvement in established lipedema. The histological and molecular characterization was conducted on anatomically-matched skin and fat biopsies as well as serum samples from eleven lipedema and ten BMI-matched healthy patients. Increased systemic levels of vascular endothelial growth factor (VEGF)-C (P = 0.02) were identified in the serum of lipedema patients. Surprisingly, despite the increased VEGF-C levels no morphological changes of the lymphatic vessels were observed. Importantly, expression analysis of lymphatic and blood vessel-related genes revealed a marked downregulation of Tie2 (P P = 0.02) consistent with an increased macrophage infiltration (P = 0.009), without changes in the expression of other lymphatic markers. Interestingly, a distinct local cytokine milieu, with decreased VEGF-A (P = 0.04) and VEGF-D (P = 0.02) expression was identified. No apparent lymphatic anomaly underlies lipedema, providing evidence for the different disease nature in comparison to lymphedema. The changes in the lymphatic-related cytokine milieu might be related to a modified vascular permeability developed secondarily to lipedema progression.

Background: The deep inferior epigastric perforator flap (DIEP) has gained widespread popularity in autologous breast reconstruction due to its natural aesthetic results and muscle-sparing design. However, donor site results regarding aesthetic outcome are often less favorable. Despite several studies on surgical technique and improvements of flap harvest, only a small number of studies addressing donor site aesthetic exist. Methods: A systematic review was performed on donor site aesthetic after DIEP flap harvest according to the PRISMA guidelines. Secondary interests were abdominal wall morbidity and complication rates in general. The following electronic databases were included to identify relevant studies: Medline, Embase, PubMed, Cochrane, and Web of Science. Results: The search yielded 95 articles for full-text analysis, with only four key studies found on donor site aesthetic specifically. Regarding complication rates, the overall rate of abdominal hernia ranges from 0 to 7.1%. Abdominal bulging occurs in 2.3% to 33%. Besides abdominal wall integrity, wound dehiscence (12–39%), seroma (1–48%), hematoma (1–15%), infections (1–12%), fat necrosis (0–11%) and umbilical necrosis (2–3%) significantly impair donor site aesthetic. Conclusions: This systematic review reveals that the topic of donor site aesthetic and potential improvement of results has largely been neglected in the past decade. This study provides a thorough evaluation of potential problems that plastic surgeons may encounter and includes recommendations to improve results.

Objective The aim of our systematic meta-analysis was to find out if lipofilling to the breast alters follow-up imaging procedures and leads to an increased number of biopsies because of suspicious findings. Methods We conducted a systematic meta-analysis of the literature including all prospective and retrospective studies focusing on imaging outcomes in patients with a history of breast cancer who have received one or more lipofilling procedures after oncologic surgery to the breast. Results Twelve studies met the inclusion criteria, comprising 1711 patients and at least 2261 lipofilling procedures. 564 patients (33%) were followed up only with ultrasound, 735 patients (43%) only received mammography, 273 (16%) had a combination of ultrasound, mammography and MRI, and 37 patients (2.1%) were followed up via ultrasound and mammography. A collective of 102 patients making up a matched-cohort study received ultrasound, mammography, MRI, and PET/CT, while only 51 of them made up the investigation group who had autologous fat grafting (3%). Biopsy rates were 1%-24% with a medium of 6.5% over all groups. Medium follow-up was 18.8 months (range 6-50 months). The rate of local oncologic events among the patients with lipofilling procedures detected during the study periods was 0.7%. Conclusion Lipofilling to the breast after oncologic operations appears to be a safe procedure with overall low biopsy and local recurrence rate. Suspicious imaging occurs in most cases out of physiologic remodeling and inflammation processes at the operation site and needs to be distinguished from malignant focusses. The amount of required biopsies stands in relation to the used imaging method and the time to follow-up.

Sentinel lymph node dissection (SLND) and radical lymph node dissection (RLND) represent standard surgical procedures in skin cancer patients. Complications may arise, with seroma formation being the most prevalent. Although both conservative and surgical approaches have been undertaken in effort to reduce the occurrence of postoperative seroma, none have been able to show a beneficial effect. Therefore, the aim of our study was to examine the rate of complications in RLNDs and SLNDs in respect to the influence of different preventative techniques on potential seroma formation. We performed an evaluation of all SLNDs and RLNDs within a 2-year period at our hospital. The primary focus was seroma formation, with secondary points being infection, wound healing problems (WHP), and lymphedema, as well as the total length of hospital stay and revision surgery. Three hundred eighty-nine patients were recorded, of which 268 patients (69 %) underwent an SLND while 121 patients (31 %) underwent an RLND. Seroma in RLNDs occurred significantly more frequently with 46.3 % compared to SLNDs with only 11.6 %. Pressure and vacuum dressings were associated with a considerably higher chance of seroma formation, namely 3.5 and 2.1 times more than with a normal dressing. Other methods such as ligation, clipping, fibrin glue, or a sartorius flap showed no effects. RLNDs are associated with a higher rate of seroma regardless of surgical or postoperative approach, whereas SLNDs only induce a few side effects. Further infections, wound healing problems, and lymphedema were significantly associated with seroma formation. Level of Evidence: Level IV, risk / prognostic study

Vascularized lymph node (VLN) transfer is an emerging strategy to re-establish lymphatic drainage in chronic lymphedema. However, the biological processes underlying lymph node integration remain elusive. This study introduces an experimental approach facilitating the analysis of short-term molecular and cellular effects of ischemia/reperfusion on VLN flaps. Lymph node flaps were dissected pedicled on the lateral thoracic vessels in 44 Lewis rats. VLN flaps were exposed to 45 or 120 minutes ischemia by in situ clamping of the vascular pedicle with subsequent reperfusion for 24 hours. Flaps not exposed to ischemia/reperfusion served as controls. Lymph nodes and the perinodal adipose tissue were separately analyzed by Western blot for the expression of lymphangiogenic and angiogenic growth factors. Moreover, morphology, microvessel density, proliferation, apoptosis and immune cell infiltration of VLN flaps were further assessed by histology and immunohistochemistry. Ischemia for 120 minutes was associated with a markedly reduced cellularity of lymph nodes but not of the perinodal adipose tissue. In line with this, ischemic lymph nodes exhibited a significantly lower microvessel density and an increased expression of VEGF-D and VEGF-A. However, VEGF-C expression was not upregulated. In contrast, analyses of the perinodal adipose tissue revealed a more subtle decrease of microvessel density, while only the expression of VEGF-D was increased. Moreover, after 120 minutes ischemia, lymph nodes but not the perinodal adipose tissue exhibited significantly higher numbers of proliferating and apoptotic cells as well as infiltrated macrophages and neutrophilic granulocytes compared with non-ischemic flaps. Taken together, lymph nodes of VLN flaps are highly susceptible to ischemia/reperfusion injury. In contrast, the perinodal adipose tissue is less prone to ischemia/reperfusion injury.

Background Chronic secondary lymphedema is a well-known complication in oncologic surgery. Autologous lymph node transplantation, lymphovenous anastomosis, and other lymphatic surgeries have been developed in the last decades with rising clinical application. Animal models to explore the pathophysiology of lymphedema and microsurgical interventions have reached great popularity, although the induction of stable lymphedema in animals is still challenging. The aim of this review was to systematically assess lymphedema animal models and their potential use to study surgical interventions. Materials and methods A systematic review according to the PRISMA guidelines was performed without time or language restriction. Studies describing new or partially new models were included in chronological order. Models for primary and secondary lymphedema were assessed, and their potential for surgical procedures was evaluated. Results The systematic search yielded 8590 discrete articles. Of 180 articles included on basis of title, 84 were excluded after abstract review. Ninety-six were included in the final analysis with 24 key articles. Conclusions No animal model is perfect, and many models show spontaneous lymphedema resolution. The rodent limb appears to be the most eligible animal model for experimental reconstruction of the lymphatic function as it is well accessible for vascularized tissue transfer. There is a need for standardized parameters in experimental lymphedema quantification. Also, more permanent models to study the effect of free vascularized lymph node transfer are needed.

Background Congenital hypoplasia of facial bones has traditionally been treated by orthognathic surgery. However, the inherent invasiveness of orthognathic surgery often leads to a high complication rate. Facial fat grafting could be a less invasive method to correct facial deformities. Objectives The aim of this study was to evaluate the results of microfat grafting for facial contouring. Methods This retrospective chart review evaluated 166 patients who were treated with microfat grafting for maxillary and/or mandibular hypoplasia. Pretreatment and posttreatment photographs were compared regarding improvement of facial contour, and complications were recorded. Results The follow-up period ranged from 4 months to 10 years (mean, 2 years 7 months). Thirty-eight percent of the patients had a refill procedure 6 or more months after the first procedure. A majority of the evaluated patients stated that they benefited from the microfat grafting, with ratings of excellent (50%), sufficient (48%), and poor (2%). Complications included visible fat lobules under the lower eyelid skin (7%), which was seen during the first 4 years and was resolved by changing the injection cannulae and technique, and fat resorption, which was seen in all patients, with a clinical range from ±15% in the immobile malar area and chin region to ±50% in the mobile lip area. Conclusions Facial microfat grafting is a valuable alternative to more complicated advancement osetotomies being performed in patients solely for aesthetic reasons. The low morbidity and rapid recovery make facial microfat grafting a welcome tool in the armamentarium of the modern facial aesthetic surgeon. Level of Evidence 4![Graphic][1] Therapeutic [10.1093/asj/sjv084][2] [1]: /embed/inline-graphic-1.gif [2]: /lookup/doi/10.1093/asj/sjv084

Treatment of fracture dislocations of the PIP joint represents a hand surgical challenge. In hemi-hamate arthroplasty, the palmar joint surface is reconstructed using an osteochondral graft from the hamate and the immediate stability permits early movement.We performed hemi-hamate reconstructions in 10 patients (mean age 34.9 years), who sustained fractures of the base of the middle phalanx of ≥50% surface and dorsal PIP dislocations. Outcomes were assessed by clinical exam and X-ray. Evaluation criteria were range of motion of PIP and DIP joints, grip strength, joint alignment, complications and donor site morbidity. Mean follow-up was 8.6 months (range 3-14). Fractures mostly involved the 4th and 5th fingers, all patients were male. Indications were subluxation and comminuition in acute cases of6 weeks (5 patients) and chronic pain and morning stiffness in chronic cases (5 patients).Operative treatment was performed in average after 93 days (range 0-371 days) after injury. Average PIP motion was 71° (range 0-90); DIP motion was 54° (range 10-90) with a mean PIP flexion contracture of 6.5° (range 0-20). Grip strength averaged 95% of the opposite hand. 4 patients had revision surgery (2× arthrolysis PIP joint, 2× screw shortening, 1× neurolysis R. dorsalis N. ulnaris).Hemi-hamate autograft arthroplasty represents an effective procedure to address severe PIP joint fracture dislocations. It restores the comminuted articular surface in chronic injuries and in the acute injury it is a challenging but valuable alternative to extension block splinting. However, donor site morbidity and revision surgery have to be taken into account.

Background In the early postoperative period after pancreas-kidney transplantation, pancreatic venous thrombosis is a major complication that leads to allograft dysfunction and graft loss. Beside ischemia and reperfusion injury, immunosuppressive drugs have been accused of supporting thrombogenicity. The aim of this study was to evaluate the effect of commonly applied immunosuppressants on microvascular thrombus formation in normal and postischemic tissue in vivo. Methods In the skin fold chambers of tacrolimus-, cyclosporine A-, antithymocyte globulin-, rapamycine-, or saline-treated mice, light/dye-induced microvascular thrombus formation was studied. Additional mice underwent ischemia and reperfusion of the skin fold chamber tissue and received tacrolimus, antithymocyte globulin, or saline before reperfusion. Additionally, the effect of prednisolone was tested in animals with ischemia and reperfusion. Concentrations of sP-selectin, soluble vascular cell adhesion molecule-1, and asymmetric dimethylarginine were assessed by enzyme-linked immunosorbent assay. Immunohistochemistry of the skin fold chamber tissue served for analysis of vascular endothelial nitric oxide synthase and inducible nitric oxide synthase expression. Results In normal tissue, tacrolimus, cyclosporine A, antithymocyte globulin, and rapamycine accelerated microvascular thrombus formation significantly when compared with saline. Whereas ischemia and reperfusion in saline-treated mice enhanced thrombus formation, thrombogenicity was not further increased by ischemia and reperfusion in tacrolimus - or antithymocyte globulin-treated animals. Application of prednisolone reversed the tacrolimus- and antithymocyte globulin-induced prothrombotic effect. Antithymocyte globulin increased sP-selectin and soluble vascular cell adhesion molecule-1, whereas tacrolimus induced asymmetric dimethylarginine production significantly. While tacrolimus and antithymocyte globulin additionally induced endothelial nitric oxide synthase and inducible nitric oxide synthase expression, cyclosporine A influenced only endothelial inducible nitric oxide synthase expression. Conclusion Immunosuppressants enhance thrombus formation in vivo. Although antithymocyte globulin activates the microvascular endothelium, we show for the first time that tacrolimus increases asymmetric dimethylarginine plasma levels. Thus, impaired nitric oxide availability might be the underlying mechanism for the tacrolimus-associated increased thrombogenicity. The efficacy of prednisolone to reverse the tacrolimus-associated and antithymocyte globulin-associated acceleration of thrombus formation underlines the application of this anti-inflammatory drug prior to reperfusion in immunosuppressive regimens.

Secondary lymphedema is a common complication of cancer treatment characterized by chronic limb swelling with interstitial inflammation. The rodent hindlimb is a widely used model for the evaluation of novel lymphedema treatments. However, the assessment of limb volume in small animals is challenging. Recently, high-resolution three-dimensional (3D) imaging modalities have been introduced for rodent limb volumetry. In the present study we evaluated the validity of microcomputed tomography (μCT), magnetic resonance imaging (MRI) and ultrasound in comparison to conventional measuring techniques. For this purpose, acute lymphedema was induced in the mouse hindlimb by a modified popliteal lymphadenectomy. The 4-week course of this type of lymphedema was first assessed in 6 animals. In additional 12 animals, limb volumes were analyzed by μCT, 9.4 T MRI and 30 MHz ultrasound as well as by planimetry, circumferential length and paw thickness measurements. Interobserver correlation was high for all modalities, in particular for μCT analysis (r = 0.975, p, Scientific Reports, 6, ISSN:2045-2322

Cutaneous melanoma is the most deadly cutaneous neoplasm. In order to guide treatment decisions and follow-up of melanoma patients, guidelines for the management of melanoma in Switzerland were inaugurated in 2001 and revised in 2006 and 2016. Recent data on surgical and medical treatments from randomised trials necessitated modification of the treatment and follow-up recommendations.

Cigarette smoking is linked to thromboembolic events; however, a relationship between nicotine exposition and thrombosis has not been established. Thus, we intended to study the effect of acute and chronic nicotine application in an in vivo mouse model.In microvessels of the dorsal skin fold chamber, light-dye-induced thrombus formation was analyzed using intravital fluorescence microscopy. Male and female C57BL/6J mice received nicotine chronically via the drinking water (100 microg/ml) for 8 weeks. An additional series of experiments was performed with acute iv nicotine treatment (3 mg/kg body weight).No significant differences in microvascular thrombus formation were detected after chronic nicotine application in male and female animals when compared with controls. Accordingly, flow cytometric analysis did not show significant effects on platelet activity. Chronic nicotine treatment resulted in a significantly reduced endothelial activation in male, but not in female mice. In contrast, acute iv application of nicotine revealed significantly shorter thrombosis times in arterioles of female mice and a significantly increased endothelial P-selectin expression in mice of both genders.Chronic nicotine application does not promote microvascular thrombus formation in mice of either gender, whereas acute high-dose iv administration caused a significant increase of arteriolar thrombosis in female animals probably via a synergistic effect of increased endothelial P-selectin expression and female hormone levels. A gender-dependency of acute nicotine action can be presumed.

The KyberSept trial in septic patients showed that antithrombin (AT) reduced 90-day mortality significantly in a subgroup of patients not receiving concomitant heparin for thrombosis prophylaxis. Microvascular thrombosis is a key pathophysiologic mechanism during sepsis, ischemia/reperfusion and disseminated intravascular coagulation (DIC). Therefore, this study investigated the antithrombotic property of AT as potential monotherapy in an experimental endotoxemia model in order to omit concomitant heparin.Using a light/dye injury model in the ear and the cremaster muscle preparation of mice, we quantitatively assessed microvascular thrombus formation in a total of 30 endotoxemic mice by means of intravital fluorescence microscopy. Before thrombus induction animals received a single i.v. bolus of AT (100 or 250 IU/kg), heparin (100 IU/kg) or saline (NaCl).In NaCl-treated endotoxemic animals, light/dye exposure led to complete thrombotic occlusion in arterioles and venules within450 s in the ear model. Heparin delayed thrombotic vessel occlusion by more than 50%. AT significantly prolonged times until thrombotic vessel occlusion in a dose-dependent manner and more effectively than heparin (p0.05 vs. NaCl and heparin). This anti-coagulative effect of AT was especially pronounced in arterioles. Upon light/dye exposure to cremaster muscle preparations in endotoxemic mice AT also caused a 4-fold delay in microvascular thrombus growth with 827+/-77 s until complete thrombotic occlusion.We could characterize for the first time AT-mediated antithrombotic activity during endotoxemia in two models of phototoxicity-induced microvascular thrombosis. Our results clearly demonstrate an additional AT mechanism of action that may be responsible for beneficial effects observed during endotoxemia and DIC. This AT profile may allow future high-dose AT application without giving heparin for thrombosis prophylaxis, an intriguing strategy that is to be tested under clinical conditions.

SummaryA recently published post-hoc analysis of a trial using high-dose antithrombin (AT) in septic patients (KyberSept) revealed significant reduction of lethality when no concomitant heparin was administered, whereas patients with the combination of heparin and AT did not benefit in terms of survival. Therefore, it seems feasible to study the capability of AT in prevention of microvascular thrombus formation to avoid concomitant application of heparin and AT. Using fluorescence microscopy and a light/dyeinjury mouse ear model, the kinetics of thrombus formation were analyzed quantitatively in vivo upon single iv bolus of saline (control), heparin (100 IU/kg), hirudin (1 mg/kg) or AT (25, 50, 100 or 250 IU/kg) (N=7 animals per group each). In controls, light/dye-injury induced complete thrombotic occlusion in all arterioles and venules studied. Heparin and hirudin prevented thrombotic vessel occlusion in 62% and 43% of arterioles and 11% and 28% of venules. AT-250 was found to be more effective than heparin and hirudin, because thrombus formation was completely banned in all arterioles and venules. AT-100 and AT-50 were also capable of significantly blocking thrombus formation in both arterioles and venules. In blood vessels, which finally clogged, the time for development of complete vessel occlusion was delayed after heparin, hirudin and AT-25, but in particular after AT-50 and AT-100. In conclusion, AT-mediated antithrombotic activity has been characterized in a model of phototoxicity-induced microvascular thrombosis formation, demonstrating thatAT delays and prevents thrombus formation in arterioles and venules at least comparably effective as heparin and hirudin.

Cold is supposed to be associated with alterations in blood coagulation and a pronounced risk for thrombosis. We studied the effect of clinically encountered systemic hypothermia on microvascular thrombosis in vivo and in vitro. Ferric chloride-induced microvascular thrombus formation was analyzed in cremaster muscle preparations from hypothermic mice. Additionally, flow cytometry and Western blot analysis was used to evaluate the effect of hypothermia on platelet activation. To test whether preceding hypothermia predisposes for enhanced thrombosis, experiments were repeated after hypothermia and rewarming to 37 degrees C. Control animals revealed complete occlusion of arterioles and venules after 742 +/- 150 and 824 +/- 172 s, respectively. Systemic hypothermia of 34 degrees C accelerated thrombus formation in arterioles and venules (279 +/- 120 and 376 +/- 121 s; P0.05 vs. 37 degrees C). This was further pronounced after cooling to 31 degrees C (163 +/- 57 and 281 +/- 71 s; P0.05 vs. 37 degrees C). Magnitude of thrombin receptor activating peptide (TRAP)-induced platelet activation increased with decreasing temperatures, as shown by 1.8- and 3.0-fold increases in mean fluorescence after PAC-1 binding to glycoprotein (GP)IIb-IIIa and 1.6- and 2.9-fold increases of fibrinogen binding on incubation at 34 degrees C and 31 degrees C. Additionally, tyrosine-specific protein phosphorylation in platelets was increased at hypothermic temperatures. In rewarmed animals, kinetics of thrombus formation were comparable to those in normothermic controls. Concomitantly, spontaneous and TRAP-enhanced GPIIb-IIIa activation did not differ between rewarmed platelets and those maintained continuously at 37 degrees C. Moderate systemic hypothermia accelerates microvascular thrombosis, which might be mediated by increased GPIIb-IIIa activation on platelets but does not cause predisposition with increased risk for microvascular thrombus formation after rewarming.

Objective— By heme degradation, heme oxygenase-1 (HO-1) provides endogenous carbon monoxide and bilirubin, both of which play major roles in vascular biology. The current study aimed to examine whether induction of HO-1 and its byproducts modulate the process of microvascular thrombus formation in vivo. Methods and Results— In individual microvessels of mouse cremaster muscle preparations, ferric chloride-induced thrombus formation was analyzed using intravital fluorescence microscopy. When mice were pretreated with an intraperitoneal injection of hemin, a HO-1 inducer, immunohistochemistry and Western blot protein analysis of cremaster muscle tissue displayed a marked induction of HO-1. In these animals, superfusion with ferric chloride solution induced arteriolar and venular thrombus formation, which, however, was significantly delayed when compared with thrombus formation in animals without HO-1 induction. The delay in thrombus formation in hemin-treated mice was completely blunted by tin protoporphyrin-IX, a HO-1 inhibitor, but not by copper protoporphyrin-IX, which does not inhibit the enzyme. Coadministration of the vitamin E analogue Trolox in HO-1–blocked animals almost completely restored the delay in thrombus formation, implying that, besides CO, the antioxidant HO pathway metabolite bilirubin mainly contributes to the antithrombotic property of HO-1. This was further supported by the fact that bilirubin was found as effective as hemin in delay of ferric chloride-induced thrombus formation. Animals with HO-1 induction revealed reduced P-selectin protein expression in cremaster muscle tissue, which most probably presented the molecular basis for delayed thrombus growth. Conclusion— Local induction of HO-1 activity may be of preventive and therapeutic value for clinical disorders with increased risk of thrombotic events.

Palmar lip injuries of the proximal interphalangeal joint with dorsal fracture-dislocation are difficult to treat and often require major reconstruction. A systematic review was performed and yielded 177 articles. Thirteen articles on hemi-hamate autograft were included in full-text analysis. Results of 71 cases were summarized. Mean follow-up was 36 months and mean proximal interphalangeal joint range of motion was 77°. Overall complication rate was around 35%. Up to 50% of the patients showed radiographic signs of osteoarthritis. However, few of those patients complained about pain or impaired finger motion. Based on this systematic analysis and review, hemi-hamate autograft can be considered reliable for the reconstruction of acute and chronic proximal interphalangeal joint fracture-dislocations with joint involvement >50%, but longer-term follow-up studies are required to evaluate its outcome, especially regarding the rate of osteoarthritis. Level of Evidence: II

A 47-year-old HIV-positive woman presented with fever and a painful swollen right forearm. Clinical presentation and MRI were suggestive for a necrotising fasciitis. Surgical exploration revealed small transparent cystic bodies resembling white caviar, which were identified by their typical morphological features as larval stages (cysticerci) of Taenia crassiceps. Molecular methods, using sequence analysis of the small subunit rRNA gene, definitively confirmed T crassiceps. T crassiceps (Cestodea: Taeniidae) is a tapeworm found in the intestines of red foxes and dogs in the Northern Hemisphere. Human infections are rare and appear to depend on the host's immunocompetence. The eight published cases could not clarify the mode of infection but discuss ingestion of teniid eggs or penetration through a cutaneous wound. The optimal treatment remains unclear. We describe a detailed and successful treatment strategy including extensive surgical interventions, prolonged anthelmintic and antiretroviral treatment.

Summary We report a rare case of human Taenia crassiceps infection in a 47-year-old female patient with untreated acquired immunodeficiency syndrome and chronic hepatitis C infection. Little experience exists regarding the appropriate treatment of this infection. Usually, a combination of anthelmintic drugs is applied. Whether surgical measures are indicated have not been clarified. In our patient, initial surgery showed an abscess and fluid collection with numerous transparent cysts localised in the subcutaneous tissue of the cubital fossa. Parasitological and pathological examinations identified these structures as larvae of the cestode T. crassiceps . After treatment with anthelmintic medications, the patient was discharged in good condition. However, the patient presented with the clinical symptoms of an acute fasciitis of the right upper extremity 7 days later. The deteriorating general condition entailing a pre-septical state demanded emergency debridement and fasciectomy of the right arm. After the surgery, the patient recovered fully. Conclusions Surgical treatment appears to be an important measure to reduce the tissue parasite load in patients with severe immunodeficiency. It also has to be questioned whether the bioavailability and the penetration of the drugs commonly administered is sufficiently high to treat such a fulminant infection alone.

Combined pancreas-kidney transplantation is one therapeutic option of Diabetes mellitus. In the early postoperative period pancreatic venous thrombosis is one of the major complications leading to allograft dysfunction and graft loss. One risk factor is the ischemia-reperfusion(I/R)- injury. Additionally, immunosuppressive drugs, such as the calcineurine inhibitor tacrolimus, have been implicated in the development of microvascular thrombosis. Several studies report a beneficial effect of antithrombin III (ATIII)- and N-acetylcysteine-(NAC)-substitution on the ischemia-reperfusion injury. Aim of this study was to analyse the therapeutic effect of ATIII and NAC on microvascular thrombus formation in post-ischemic tissue under immunosuppressive therapy with tacrolimus in vivo.

The aim of this study was to investigate the effect of human recombinant erythropoietin (EPO) on the microcirculation and oxygenation of critically ischemic tissue and to elucidate the role of endothelial NO synthase in EPO-mediated tissue protection. Island flaps were dissected from the back skin of anesthetized male Syrian golden hamsters including a critically ischemic, hypoxic area that was perfused via a collateralized vasculature. Before ischemia, animals received an injection of epoetin beta at a dose of 5,000 U/kg body weight with (n = 7) or without (n = 7) blocking NO synthase by 30 mg/kg body weight L-NAME (Nomega-nitro-L-arginine methyl ester hydrochloride). Saline-treated animals served as control (n = 7). Ischemic tissue damage was characterized by severe hypoperfusion and inflammation, hypoxia, and accumulation of apoptotic cell nuclei after 5 h of collateralization. Erythropoietin pretreatment increased arteriolar and venular blood flow by 33% and 37%, respectively (P < 0.05), and attenuated leukocytic inflammation by approximately 75% (P < 0.05). Furthermore, partial tissue oxygen tension in the ischemic tissue increased from 8.2 to 15.8 mmHg (P < 0.05), which was paralleled by a 21% increased density of patent capillaries (P < 0.05) and a 50% reduced apoptotic cell count (P < 0.05). The improved microcirculation and oxygenation were associated with a 2.2-fold (P < 0.05) increase of endothelial NO synthase protein expression. Of interest, L-NAME completely abolished all the beneficial effects of EPO pretreatment. Our study demonstrates that, in critically ischemic and hypoxic collateralized tissue, EPO pretreatment improves tissue perfusion and oxygenation in vivo. This effect may be attributed to NO-dependent vasodilative effects and anti-inflammatory actions on the altered vascular endothelium.

Thrombotic microangiopathy (TMA) is a fatal complication that occurs after combined pancreaskidney transplantation leading to allograft dysfunction and graft loss. Immunosuppressive drugs, such as calcineurin inhibitors, have been implicated in the development of nonimmune TMA. Tacrolimus has been shown to be associated with decreased NO-production, thus contributing to thrombus formation. ADMA acts as an endogenous NO-synthase inhibitor and is considered to be a marker of endothelial dysfunction. Aim of this study was to analyse the influence of different immunosuppressants on microvascular thrombus formation in vivo and to further examine the underlying endothelial function. Using the skin fold chamber in C57BL/6J mice, microvascular thrombus formation was induced photochemically and was quantitatively analyzed by intravital fluorescence microscopy. Mice were treated with tacrolimus (TAC: 10 mg/kg/d; n = 7), cyclosporine (CYA: 5 mg/kg/d) or sirolimus (RAPA: 1,5 mg/kg/d ip) on 3 consecutive days. Control mice received NaCl 0,9 % (10 ml/kg/d ip; n = 5). Drug plasma levels were examined to ensure therapeutic doses. Additionally, the systemic circulation of sP- and sE-selectin as well as ADMA plasma levels were measured by ELISA. Application of immunosuppressants produced clinically relevant plasma levels (TAC: 8 ± 2 ng/ml, CYA: 214 ± 16 ng/ml; RAPA: 9 ± 1 ng/ml). Microvascular thrombus formation was significantly accelerated in mice receiving TAC compared to control mice (arteriolar and venular occlusion: 251 ± 101s and 179 ± 27 s vs. control 818 ± 221 s and 749 ± 231 s; p < 0,05). Application of CYA significantly increased thrombus formation only in venules (276 ± 49s; p < 0,05 vs. control), whereas RAPA had no significant effect on thrombosis induction. Plasma concentrations of sP-Selectin and sE-selectin were slightly reduced after TAC and CYA application. ADMA-levels however were significantly increased in mice receiving TAC (1,28 ± 0,16 µmol/l vs. control: 0,74 ± 0,13 µmol/l; p < 0,05). The present results show that clinically relevant plasma levels of cyclosporine enhanced thrombus formation only in venules whereas tacrolimus significantly accelerates thrombus formation in arterioles and venules. This effect could be explained by increased ADMA-levels in plasma resulting in impaired NO-bioavailability and increased thrombogenicity, thus presenting a new molecular mechanism in the development of transplant-thrombosis.

Erythropoietin-treatment is generally assumed to be associated with an increased risk for thrombosis due to an elevated hematocrit. However, previous in vitro and in vivo studies of our group revealed a reduction of platelet reactivity and no significant effect on microvascular thrombus formation during treatment with darbepoetin-alpha (DPO), an erythropoietin-derivative with a 3-fold longer half life. Aim of the study was therefore to investigate the effect of DPO application on endothelial function and to further characterize the role of eNOS in this context.

Cigarette smoking increases the risk for myocardial diseases and is associated with a higher incidence of postoperative complications. Previous studies showed an increased leukocyte rolling and adhesion following nicotine application. Next to this the use of estrogen containing oral contraceptives is linked to a higher risk for deep vein thrombosis. However, clinical studies investigating the influence of transdermal nicotine application on cardiovascular risk factors could not show a negative effect of nicotine. Therefore transdermal nicotine replacement is regarded as a safe aid to smoking cessation. It is still not clear whether nicotine or other substances in cigarette smoke are responsible for the prothrombotic risk associated with cigarette smoking. Aim of this study was therefore to investigate the influence of nicotine on microvascular thrombus formation in vivo and to further characterize the underlying mechanisms. In microvessels of dorsal skin fold chambers of mice, light-dye-induced thrombus formation was analyzed using intravital fluorescence microscopy. Male and female C57BL/ 6J mice received nicotine chronically via the drinking water (100 µg/ml) for 8 weeks. Control animals of both gender received normal drinking water. Subsequently plasma levels of cotinine, a stable metabolite of nicotine, were determined. To study the influence of nicotine on endothelial cell function immunohistochemical tissue sections for the vascular localization of P-selectin, PAI-1, PAF-R and ICAM-1 were performed. Furthermore the plasma concentrations of soluble P-selectin, E-selectin, ICAM-1 and VCAM-1 were quantified by ELISA. To characterize the influence of nicotine on platelet activation, flowcytometric analyses of the expression of P-selectin, GPIIb/IIIa and CD107a on murine platelets were carried out. Chronic nicotine exposition caused cotinine plasma levels similar to those seen in chronic smokers (> 100 ng/ml vs. controls: < 10 ng/ml). In male animals no significant differences in microvascular thrombus formation were detected following chronic nicotine application, when compared to controls (complete arteriolar and venular occlusion: 640 ± 96 s and 427 ± 51 s vs. control: 577 ± 78 s and 531 ± 102 s). Likewise in female animals no significant differences were found, following long-term nicotine application (complete arteriolar and venular occlusion: 715 ± 130 s and 493 ± 79 s vs. control: 866 ± 24 s and 481 ± 70 s). In line with this, flow cytometric analyses did not show significant effects of nicotine on P-selectin, GPIIb/IIIa and CD107a expression on resting platelets or upon stimulation with thrombin (20 U/ml). In contrast to this chronic nicotine application caused a marked reduction of vascular expression of P-selectin, PAF-R and PAI-1 in male mice, while in female mice only PAI-1 was found significantly reduced. Additionally chronic nicotine treatment lead to a significant reduction of sP-selectin and sE-selectin circulation in male mice. The present study shows no significant influence of chronic nicotine treatment on microvascular thrombus formation and platelet adhesion molecule expression despite elevated cotinine levels in mice of both gender. However, chronic nicotine treatment resulted in a significant reduction of endothelial adhesion molecule expression in male mice, while there was only a slight effect in female mice.

Diabetic macro- and microangiopathy are associated with a high risk of vascular complications. The diabetic patient exhibits a pathological coagulation state, with an increased synthesis of coagulation factors and plasminogen activator inhibitor 1 (PAI-1) as well as an enhanced aggregation of platelets. Previous studies have shown that C-peptide can reduce leucocyte-endothelial cell interaction and improve microvascular blood flow in patients with type 1 diabetes. In the present study, we examined in vivo whether C-peptide is able to reduce platelet activation and through that microvascular thrombus formation.In the microvessels of cremaster muscle preparations taken from normal and diabetic mice, ferric chloride-induced thrombus formation was analysed using intravital fluorescence microscopy.I.V. administration of C-peptide in high dose (70 nmol/kg), but not in low dose (7 nmol/kg), caused a significant delay in arteriolar and venular thrombus growth in normal and diabetic mice. This effect was repressed by cremaster muscle superfusion with insulin (100 microU/ml) in diabetic animals, but particularly in normal animals. In parallel, immunohistochemistry demonstrated a higher number of PAI-1-expressing vessels in cremaster muscle tissue from control animals and from animals treated with C-peptide and insulin compared with tissue from animals with C-peptide treatment application alone.We conclude that C-peptide possesses antithrombotic actions in vivo. A causal role of PAI-1 in this scenario needs to be further addressed. However, the reversal of C-peptide action by insulin may invalidate the use of this peptide as a treatment option to improve rheology and microcirculation in diabetic patients.

Strategies for myocardial regeneration focus on intravascular administration or mobilisation of stem cells (SC). SDF-1 is discussed as homing factor for SC to injured myocardium. We investigated the role of SDF-1 for mediating SC homing using different in vitro and in vivo approaches in the mouse. In hearts of C57/BL6J mice the LAD coronary artery was ligated (n=4). After 24 h SDF-1 cardiac mRNA expression was measured by RT-PCR. Chemoattractant activity of SDF-1 (100–500 ng/ml) for murine c-kit+ bone marrow SC was studied using Boyden chamber assays (0.2 × 106 SC in upper chamber) (n=3). The microvascular bed of right cremaster muscles was topically exposed to SDF-1 (200 ng/ml) and studied by intravital fluorescence microscopy upon intra-arterial injection of green-fluorescently labelled SC (2 × 106 c-kit+) (n=5). SC rolling was recorded during first pass, while firm adherence of SC to the venular endothelium was assessed after 1 h. Values were compared to those in controls without SDF-1-exposure. SDF-1 mRNA expression was significantly increased 24 h after myocardial infarction. In vitro, SC migrated against an SDF-1 concentration gradient in a dose-dependent fashion (20%–75%). In vivo, the fraction of rolling SC increased significantly upon SDF-1 exposure (14 ± 2% vs. control: 6 ± 1% p

Diabetic macro- and microangiopathy is associated with a high risk of vascular complications and in consequence a high morbidity and mortality rate. The diabetic patient exhibits a pathological coagulation state, which is represented by both an increased synthesis of factor VII, thrombin, tissue factor and plasminogen activator inhibitor 1 (PAI-1) as well as by platelet aggregation enhanced due to elevated GP IIb/IIIa expression and altered calcium homeostasis. Former studies showed improved microvascular blood flow of skin and muscle after C-peptide administration in type 1 diabetic patients. Additionally, positive effects of C-peptide application on leukocyte-endothelial-interaction with increased production of nitric oxide and decreased expression of endothelial P-selectin and ICAM-1 have been reported. The aim of this study was therefore to examine whether the administration of C-peptide has anti-thrombotic effects in vivo. In microvessels of mouse cremaster muscle preparations of diabetic and non-diabetic mice, ferric chloride-induced thrombus formation was analyzed using intravital fluorescence microscopy. C-peptide was applied systemically in low (7 nmol/kg) and high (70 nmol/kg) dose, while control groups received equivalent amounts of heat-inactivated C-peptide. Diabetes was induced by application of streptozotocin (40 mg/kg) on 5 consecutive days. The diabetic metabolic state was confirmed by repeated positive testing for hypergly-cemia and glucosuria. Additionally, flow cytometric analysis of human platelet P-selectin and GP IIb/IIIa expression and immunohistochemistry of endothelial PAI-1 expression within the cremaster muscle tissue were conducted. Administration of C-peptide in high, but not in low dose, caused a significant delay in venular and arteriolar thrombus growth in diabetic mice (complete occlusion: 1171 ± 233 s and 1317 ± 218 s vs. control: 496 ± 50 s and 488 ± 50 s; p

A randomized controlled study (KyberSept-Trial [1]) in patients with severe sepsis showed that anti-thrombin-III significantly reduced lethality in a subgroup of septic patients not receiving concomitant heparin for thrombosis prophylaxis. Therefore, the present study investigated the antithrombotic property of ATIII in an animal study as potential monotherapy in order to omit concomitant heparin therapy. In a light/dye injury model of microvascular thrombosis at the ear of the mouse, we investigated a total of 30 homozygous (SKH1-hr) hairless mice of either sex (10–12 weeks old with a body weight (bw) of 22–40 g) under ip anaesthesia with ketamine/xylazine (90/25 mg/kg bw). Intravital fluorescence microscopy allowed quantitative analysis of the kinetics of thrombus formation in arterioles and venules. Prior to thrombus induction all animals received a single intravenous bolus of hirudin (1 mg/kg bw) or ATIII in dosages of 25, 50, 100 or 250 U/kg bw. Control animals received equivalent volumes of physiologic saline (n=5 animals per group). In saline-treated controls, complete thrombotic occlusion could be induced in 100% of all arterioles (9/9) and venules (40/40) studied. Hirudin prevented thrombotic vessel occlusion in 56% of the arterioles (5/9) and 39% of the venules (9/23). ATIII-250 was found more effective than hirudin, as thrombus formation was banned in all arterioles (13/13) and venules (24/24) (p

Thromboembolism is still a major problem in almost all fields of clinical medicine. The inducible enzyme heme oxygenase 1 (HO-1) provides endogenous iron, vasodilative carbon monoxide (CO) and anti-oxidant biliverdin by decomposition of heme. The aim of this study was to examine whether the expression of HO-1 increases the anti-thrombotic properties of the endothelium and subsequently modulates the process of microvascular thrombus formation in vivo. In micro-vessels of mouse cremaster muscle preparations ferric chloride-induced thrombus formation was analyzed using intravital fluorescence microscopy. When mice were pretreated with an intraperi-toneal injection of hemin, a HO-1 inducer, Western blot protein analysis of cremaster muscle tissue revealed a marked induction of HO-1. Immunohistochemistry displayed a preferential expression of HO-1 in the vascular endothelium of arterioles and venules. In these animals, superfusion with ferric chloride caused arteriolar and venular thrombus growth, which however was significantly delayed (complete occlusion: 670 ± 96 s and 647 ± 129 s) when compared with thrombus formation in control animals (complete occlusion: 143 ± 18 s and 130 ± 25 s). The delay in thrombus formation in hemin-treated mice was abolished by tin protoporphyrin-IX, a HO-1 inhibitor, but not by copper protoporphyrin-IX, which does not inhibit the enzyme. Application of the vitamin E analogue Trolox in HO-1-blocked animals almost completely restored the delay in thrombus formation, implying that - beside CO - the anti-oxidant biliverdin mainly contributes to the anti-thrombotic effect of HO-1. Animals with HO-1 induction exhibited a reduced P-selectin protein expression in cremaster muscle tissue, most probably presenting the molecular basis for delayed thrombus growth. Local induction of HO-1 activity may therefore be of preventive and therapeutic value for clinical disorders with an increased risk of thrombotic events.

As a reaction to reported adverse outcomes after lidocaine infiltration in tumescent liposuction, prilocaine has gained increasing popularity. Previous studies investigating large-volume liposuction procedures found maximum prilocaine levels and methemoglobinemia up to 12 h postoperatively, suggesting that liposuction should be performed as a hospital procedure only. The aim of this study was to determine prilocaine plasma levels and methemoglobinemia in patients after low- to average-volume liposuction for the purpose of defining the required postoperative surveillance period.In 25 patients undergoing liposuction involving less than 2,000 ml prilocaine levels and methemoglobinemia were measured over 4 h postoperatively. Liposuction was conducted after the tumescent technique using a 0.05% hypotonic prilocaine solution with epinephrine.The average prilocaine dose was 6.8 + 0.8 mg/kg, with a maximum dose of 15 mg/kg. The peak prilocaine plasma level of 0.34 mug/ml occurred 3 h after the infiltration. The mean methemoglobinemia at this time point was 0.65%. Only one patient demonstrated a slightly elevated methemoglobin level of 1.4%, but lacked any clinical signs of methemoglobinemia. The prilocaine recovery in the aspirate averaged 36 +/- 4%, indicating that a large amount is removed by suctioning.The patients did not experience high plasma levels of prilocaine or methemoglobinemia undergoing liposuction involving less than 2,000 ml using a 0.05% hypotonic prilocaine solution. The authors therefore conclude that this procedure can be performed safely with a monitoring period of 12 h.

SummaryEbselen, a seleno-organic compound showing glutathione peroxidase-like activity, has potent anti-inflammatory and anti-oxidant effects. Since selenium deficiency is thought to be associated with an increased incidence of vascular thrombosis, we studied the effect of ebselen on blood cell aggregate formation and vessel occlusion in vivo. In individual microvessels of rat cremaster muscle preparations, photochemically induced thrombus formation was analyzed in detail using intravital fluorescence microscopy. In ebselen-pretreated animals (30mg/kg ip), venular thrombus formation was significantly delayed (50% vessel occlusion: 535±34s; initial stasis: 872±82s; complete occlusion: 908±87s) as compared to vehicle-treated controls (416±42; 612±49; 647±51). Moreover, ebselen significantly prolonged the kinetics of arteriolar thrombus formation and even completely prevented blood cell aggregate and thrombus formation in 88.9% of all arterioles studied (p

Ebselen, a seleno-organic compound showing glutathione peroxidase-like activity, has potent anti-inflammatory and anti-oxidant effects. Since selenium deficiency is thought to be associated with an increased incidence of vascular thrombosis, we studied the effect of ebselen on blood cell aggregate formation and vessel occlusion in vivo. In individual microvessels of rat cremaster muscle preparations photochemically induced thrombus formation was analyzed in detail using intravital fluorescence microscopy. In ebselen-pretreated animals (30 mg/kg ip), venular thrombus formation was significantly delayed (50% vessel occlusion: 535 ± 34s; initial stasis: 872 ± 82s; complete occlusion: 908 ± 87s) as compared to vehicle-treated controls (416 ± 42s; 612 ± 49s; 647 ± 51s). Moreover, ebselen significantly prolonged the kinetics of arteriolar thrombus formation and even completely prevented blood cell aggregate and thrombus formation in 88.9% of all arterioles studied (p < 0.05 vs controls: 37.5%). As assessed by flow cytometry, oxidant stress-induced upregulation of CD62P on isolated platelets was found dose-dependently inhibited by increasing concentrations of ebselen (10 – 100 μM), suggesting that the antithrombotic effect of ebselen is achieved by attenuation of CD62P-dependent platelet/leukocyte aggregation. Thus, ebselen represents preventive and therapeutic value for disorders with increased risk for oxidant stress-associated thrombotic events.

Thrombotic microangiopathy (TMA) is a fatal complication that occurs after combined pancreaskidney transplantation leading to allograft dysfunction and graft loss. Immunosuppressive drugs, such as calcineurin inhibitors, have been implicated in the development of nonimmune TMA. Tacrolimus has been shown to be associated with decreased NO-production, thus contributing to thrombus formation. ADMA acts as an endogenous NO-synthase inhibitor and is considered to be a marker of endothelial dysfunction. Aim of this study was to analyse the influence of different immunosuppressants on microvascular thrombus formation in vivo and to further examine the underlying endothelial function. Using the skin fold chamber in C57BL/6J mice, microvascular thrombus formation was induced photochemically and was quantitatively analyzed by intravital fluorescence microscopy. Mice were treated with tacrolimus (TAC: 10 mg/kg/d; n = 7), cyclosporine (CYA: 5 mg/kg/d) or sirolimus (RAPA: 1,5 mg/kg/d ip) on 3 consecutive days. Control mice received NaCl 0,9 % (10 ml/kg/d ip; n = 5). Drug plasma levels were examined to ensure therapeutic doses. Additionally, the systemic circulation of sPand sE-selectin as well as ADMA plasma levels were measured by ELISA. Application of immunosuppressants produced clinically relevant plasma levels (TAC: 8 ± 2 ng/ml, CYA: 214 ± 16 ng/ml; RAPA: 9 ± 1 ng/ml). Microvascular thrombus formation was significantly accelerated in mice receiving TAC compared to control mice (arteriolar and venular occlusion: 251 ± 101s and 179 ± 27 s vs. control 818 ± 221 s and 749 ± 231 s; p < 0,05). Application of CYA significantly increased thrombus formation only in venules (276 ± 49s; p < 0,05 vs. control), whereas RAPA had no significant effect on thrombosis induction. Plasma concentrations of sP-Selectin and sE-selectin were slightly reduced after TAC and CYA application. ADMA-levels however were significantly increased in mice receiving TAC (1,28 ± 0,16 μmol/l vs. control: 0,74 ± 0,13 μmol/l; p < 0,05). The present results show that clinically relevant plasma levels of cyclosporine enhanced thrombus formation only in venules whereas tacrolimus significantly accelerates thrombus formation in arterioles and venules. This effect could be explained by increased ADMA-levels in plasma resulting in impaired NO-bioavailability and increased thrombogenicity, thus presenting a new molecular mechanism in the development of transplant-thrombosis.

Introduction Hypothermia during sepsis significantly impairs patient outcome in clinical practice. Severe sepsis is closely linked to activation of the coagulation system, resulting in microthrombosis and subsequent organ failure. Herein, we studied whether systemic hypothermia accelerates microvascular thrombus formation during lipopolysacharide (LPS)-induced endotoxemia in vivo, and characterized the low temperature-induced endothelial and platelet dysfunctions.