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Specifically tailored amino acid–based formulations were previously shown to have a high potential to avoid stress-mediated degradation of complex molecules such as monoclonal antibodies and viral vectors. By using adenovirus 5 (Ad5) as a model, we studied whether such formulations may also efficiently protect viral vectors in thermal stress experiments and during long-term liquid storage. Algorithm-based amino acid preselection using an excipient database and subsequent application of design of experiments (DoE) in combination with a 37°C challenging model enabled the prediction of long-term storage stability of Ad5. By statistical analysis of the Ad5 infectivity, amino acids with significant influence on Ad5 stability were detected after 2 and 3 weeks of liquid storage at 37°C. Ad5 formulations comprising positively selected amino acids did not reveal any loss of infectivity after 24 months in liquid storage at 5°C. By contrast, a 2 log reduction after 3 months and complete loss of infectivity after 18 months was observed with a standard viral vector formulation. By an optimization round, we designed a simple and well-balanced formulation avoiding MgCl2, previously considered essential in Ad5 formulations. This work demonstrates the efficacy of an algorithm-based development approach in the formulation development for viral vectors.

Highlights • First COVID-19 outpatient study based on risk stratification and early antiviral treatment at the beginning of the disease. • Low-dose hydroxychloroquine combined with zinc and azithromycin was an effective therapeutic approach against COVID-19. • Significantly reduced hospitalisation rates in the treatment group. • Reduced mortality rates in the treatment group., The aim of this study was to describe the outcomes of patients with coronavirus disease 2019 (COVID-19) in the outpatient setting after early treatment with zinc, low-dose hydroxychloroquine and azithromycin (triple therapy) dependent on risk stratification. This was a retrospective case series study in the general practice setting. A total of 141 COVID-19 patients with laboratory-confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in the year 2020 were included. The main outcome measures were risk-stratified treatment decision and rates of hospitalisation and all-cause death. A median of 4 days [interquartile range (IQR) 3–6 days; available for n = 66/141 patients] after the onset of symptoms, 141 patients (median age 58 years, IQR 40–67 years; 73.0% male) received a prescription for triple therapy for 5 days. Independent public reference data from 377 confirmed COVID-19 patients in the same community were used as untreated controls. Of 141 treated patients, 4 (2.8%) were hospitalised, which was significantly fewer (P < 0.001) compared with 58 (15.4%) of 377 untreated patients [odds ratio (OR) = 0.16, 95% confidence interval (CI) 0.06–0.5]. One patient (0.7%) in the treatment group died versus 13 patients (3.4%) in the untreated group (OR = 0.2, 95% CI 0.03–1.5; P = 0.12). No cardiac side effects were observed. Risk stratification-based treatment of COVID-19 outpatients as early as possible after symptom onset using triple therapy, including the combination of zinc with low-dose hydroxychloroquine, was associated with significantly fewer hospitalisations.

Background Extra Corporeal Membrane Oxygenation (ECMO) has become an accepted treatment option for severely ill patients. Due to a limited availability of ECMO support therapy, patients must often be transported to a specialised centre before or after cannulation. According to the ELSO guidelines, an ECMO specialist should be present for such interventions. Here we describe the safety and efficacy of a reduced team approach involving one anaesthesiologist, experienced in specialised intensive care medicine, and a specialised critical care nurse. Methods This study is a 10 years retrospective, single institution analysis of all data collected between January 2007 and December 2016 from the medical records at the University Hospital Bonn, Germany. Results The Bonner mobile ECMO team was deployed in 170 cases for on-site evaluation for ECMO support therapy. 4 (2.4%) patients died prior to arrival or during the implementation of ECMO support. Of the remaining 166 patients, 126 were cannulated at the referring site, 40 were transported without ECMO. Of those, 21 were subsequently cannulated out our centre. 19 patients never received ECMO treatment. The primary indication for ECMO treatment was ARDS (159/166 patients). Veno-venous ECMO was initiated in 137, whilst 10 patients received veno-arterial ECMO treatment. Mean transportation time was 75 ± 36 min, and mean transport distance was 56 ± 57 km. In total, 26 complications were observed, three being directly transport-related. The overall survival was 55%. Conclusions Initiation of extracorporeal membrane oxygenation and subsequent transport can be safely and efficiently performed by a two-man team with good outcome. Electronic supplementary material The online version of this article (10.1186/s12871-019-0687-9) contains supplementary material, which is available to authorized users.

Currently, drug repurposing is an alternative to novel drug development for the treatment of COVID-19 patients. The antimalarial drug chloroquine (CQ) and its metabolite hydroxychloroquine (HCQ) are currently being tested in several clinical studies as potential candidates to limit SARS-CoV-2-mediated morbidity and mortality. CQ and HCQ (CQ/HCQ) inhibit pH-dependent steps of SARS-CoV-2 replication by increasing pH in intracellular vesicles and interfere with virus particle delivery into host cells. Besides direct antiviral effects, CQ/HCQ specifically target extracellular zinc to intracellular lysosomes where it interferes with RNA-dependent RNA polymerase activity and coronavirus replication. As zinc deficiency frequently occurs in elderly patients and in those with cardiovascular disease, chronic pulmonary disease, or diabetes, we hypothesize that CQ/HCQ plus zinc supplementation may be more effective in reducing COVID-19 morbidity and mortality than CQ or HCQ in monotherapy. Therefore, CQ/HCQ in combination with zinc should be considered as additional study arm for COVID-19 clinical trials.

The SARS-CoV-2 virus spreading across the world has led to surges of COVID-19 illness, hospitalizations, and death. The complex and multifaceted pathophysiology of life-threatening COVID-19 illness including viral mediated organ damage, cytokine storm, and thrombosis warrants early interventions to address all components of the devastating illness. In countries where therapeutic nihilism is prevalent, patients endure escalating symptoms and without early treatment can succumb to delayed in-hospital care and death. Prompt early initiation of sequenced multidrug therapy (SMDT) is a widely and currently available solution to stem the tide of hospitalizations and death. A multipronged therapeutic approach includes 1) adjuvant nutraceuticals, 2) combination intracellular anti-infective therapy, 3) inhaled/oral corticosteroids, 4) antiplatelet agents/anticoagulants, 5) supportive care including supplemental oxygen, monitoring, and telemedicine. Randomized trials of individual, novel oral therapies have not delivered tools for physicians to combat the pandemic in practice. No single therapeutic option thus far has been entirely effective and therefore a combination is required at this time. An urgent immediate pivot from single drug to SMDT regimens should be employed as a critical strategy to deal with the large numbers of acute COVID-19 patients with the aim of reducing the intensity and duration of symptoms and avoiding hospitalization and death.

Background Based upon our excellent previous experience with 151 adult patients and 39 children whom had the peritoneal catheter in ventriculoperitoneal shunting placed laparoscopically, we continued following this technique as a first-line-procedure in ventriculoperitoneal shunting. Now we analyzed our experience with additional 405 cases for a better comprehension of the complications, advantages and disadvantages of this procedure on this high number of patients. A strict interdisciplinary setting with the maximum of medical intraoperative competence was our goal and therefore better results. Material and methods N = 405 patients with intraperitoneal shunt insertion from the years 2006–2013 (Follow-up period ranges from 2 to 9 years with a 5,9-year mean follow-up period) were retrospectively analyzed with a special focus on the possible peritoneal catheter complications after laparoscopical shunt insertion. In our department all the peritoneal catheters in ventriculoperitoneal shunting are inserted laparoscopically, when there is no contraindication for this technique. Results We had 0% peritoneal catheter misplacement rate with help of the laparoscopic technique. In two cases (0.49%) injury of the small bowel could be repaired immediately with no further action required. In two cases umbilical hernias have been accidently discovered and the repair of the hernias took place in the same surgical session. As this technique helps us to control the shunt position intraperitoneally by direct laparoscopic vision, the patients spared an extra radiation exposure, to control the position of the peritoneal catheter. A diagnostic laparoscopy is also possible if needed. The time of the operation is shortened in comparison with the needed time, which is mentioned in literature, for the open laparotomy and of course the needed anesthesia and its possible risks and complications decreased. Conclusion No revision surgeries were required because of any misplacement of the peritoneal catheter, no additional technique related risks compared to the open surgical technique, no abdominal x-rays were needed, the operation time is shortened and the dose of anesthesia needed is decreased and of course its possible side effects' rate is also decreased. Another great benefit of this technique is the possibility of accidental diagnosis of intra-abdominal pathologies. Also, if a surgical treatment of this accidentally discovered intra-abdominal pathologies is needed, it can take place in the same surgical session. No prolonged surgery time as the laparoscopic technique is much easier, controllable and fast.

RBOHF from Arabidopsis thaliana represents a multifunctional NADPH oxidase regulating biotic and abiotic stress tolerance, developmental processes and guard cell aperture. The molecular components and mechanisms determining RBOHF activity remain to be elucidated. Here we combined protein interaction studies, biochemical and genetic approaches, and pathway reconstitution analyses to identify and characterize proteins that confer RBOHF regulation and elucidated mechanisms that adjust RBOHF activity. While the Ca

The anatomy of the cavernous sinus is described controversially in a number of publications. In the present cadaveric study, the architecture of the dorsolateral wall of the cavernous sinus is studied microsurgically and histologically. Twenty cadaveric skulls have been dissected through a classical surgical frontotemporal approach. The temporal skull base was flattened and anatomical landmarks like the meningo-orbital band, superior orbital fissure, foramina rotundum, ovale, and spinosum were identified. Lateral of the trigeminal foramina, the dura was cut and the periosteal dural layer was separated from the meningeal layer, identifying an interdural zone. The length and the extent of this zone were evaluated. The dural architecture of the interdural incision zone was examined histologically. In all specimens, two dural layers lateral of the trigeminal foramina could be separated. The identified interdural incision zone extended in a length of 3.8–6.4 cm in the antero-posterior direction. The zone could be followed medially to the superior orbital fissure for 5.3 mm and lateral of the foramen spinosum for 6.4 mm. The separation of the dural layers allowed the approach to the superior border of the cavernous sinus through this interdural incision zone. The histological analysis of the interdural incision zone showed clearly the existence of two dural layers. The architecture of the temporal-fossa-dura allows the microsurgical separation of two meningeal dural layers through a length of 5–6 cm next to the trigeminal foramina. Opening this interdural incision zone allowed exploring the superior border of the cavernous sinus.

Objective There is a number of different methods to localize a glioma intraoperatively. Neuronavigation, intraoperative MRI, 5-aminolevulinic acid, as well as intraoperative sonography. Every method has its advantages and disadvantages. Low grade gliomas do not show a specific signal with 5-aminolevulinic acid and are difficult to distinguish macroscopically from normal tissue. In the present study we stress out the importance of intraoperative diagnostic ultrasound for localization of low grade gliomas. Methods We retrospectively evaluated the charts and MRIs of 34 patients with low grade gliomas operated in our department from 2011 until December 2014. The efficacy of ultrasound as an intraoperative navigational tool was assessed. In 15 patients ultrasound was used and in 19 not. Only histologically proven low grades gliomas (astrocytomas grade II) were evaluated. Results In none of the patients where ultrasound (combined with neuronavigation) was used (N = 15) to find the tumors, the target was missed, whereas the exclusive use of neuronavigation missed the target in 5 of 19 cases of small subcortical low grade gliomas. Conclusions Intraoperative ultrasound is an excellent tool in localizing low grade gliomas intraoperatively. It is an inexpensive, real time neuronavigational tool, which overcomes brain shift. Even when identifying the tumors with ultrasound is very reliable, the extend of resection and the decision to remove any residual tumor with the help of ultrasound is at the moment unreliable.

The reported incidence rate of bile duct injury (BDI) during laparoscopic cholecystectomy (LC) is 0.3%. However, routine use of intraoperative cholangiography (IOC) is a controversial, due to the additional cost and radiation exposure. The aim of this study was to assess the application of fluorescence cholangiography (FC) in comparison to IOC and to LC without any intraoperative imaging.This prospective study included 230 patients undergoing LC in our institution. The subjects were divided into two groups. In the first group, with 170 patients, both FC and IOC were performed following a standardised protocol. In second group, with 60 patients, FC was compared to LC without any intraoperative imaging. The data were then analysed with respect to procedure time and identification of predefined anatomical structures.The mean age and body mass index in the first group were 54.4 ± 15.7 years and 27.9 ± 5.7 kg/m², respectively. The mean operative time was 67.6 ± 23.3 min. FC was performed more rapidly than IOC (1.5 ± 0.9 vs.7.3 ± 5.0 min) and visualised the cystic duct (DC) in 67.5% of patients and the common bile duct (DHC) in 66.2% of patients before dissection of Calot's triangle. During dissection, DC and DHC were detected in 95.9% and 71.2% of patients, respectively. BMI 25 kg/m² and male gender significantly reduced the identification rate of DC before dissection of Calot's triangle. Bile leakage from the liver bed after cholecystectomy was found in 3 cases (1.8%) by FC. In 2 patients (1.2%), IOC visualised the DC joining directly to the right hepatic duct. In 1 of these 2 cases (0.6%), the anatomical variation was identified first by FC. Intraductal filling defects were detected in 9 patients (5.3%) using IOC, compared to 1 patient (0.6%) using FC. In the second group, the visualisation rates of DC and DHC were 80.0 and 53.3%, respectively, with FC and 60.0 and 43.3%, respectively, during LC without any imaging. Surgeons confirmed an increase in safety in 70.0% of patients using FC.FC is a simple procedure for non-invasive real-time visualisation of bile duct anatomy during LC. Earlier identification of biliary anomalies and bile leakage increases the operative safety and enables immediate care. In obese patients, FC has limited validity.Die Inzidenzrate einer Gallengangverletzung (BDI) im Rahmen einer laparoskopischen Cholezystektomie (LC) beträgt 0,3%. Eine routinemäßige Anwendung der intraoperativen Röntgencholangiografie (IOC) wird jedoch aufgrund der zusätzlichen Kosten und der Strahlenbelastung kontrovers diskutiert. Ziel dieser Studie war die klinische Evaluation der Fluoreszenzcholangiografie (FC).Die vorliegende prospektive Studie umfasste 230 Patienten, die sich einer LC unterzogen haben. 170 Patienten erhielten sowohl eine FC als auch eine IOC entsprechend einem standardisierten Protokoll. In weiteren 60 Fällen wurden die Ergebnisse der FC mit einer LC ohne eine simultane Cholangiografie verglichen. Anschließend erfolgte die Analyse der Daten hinsichtlich des zeitlichen Aufwands und der Identifikation von biliären Strukturen.Das Durchschnittsalter und der Durchschnitts-BMI betrugen 54,4 ± 15,7 Jahre und 27,9 ± 5,7 kg/m² bei einer mittleren Operationszeit von 67,6 ± 23,3 min. Der zeitliche Aufwand der FC war dabei deutlich geringer als derjenige der IOC (1,5 ± 0,9 vs.7,3 ± 5,0 min). Die Detektionsraten für Ductus cysticus (DC) sowie Ductus hepaticus communis (DHC) im Rahmen einer FC betrugen vor Dissektion am Calot-Dreieck 67,5 bzw. 66,2% und währenddessen 95,9 bzw. 71,2%. Ein BMI 25 kg/m² und männliches Geschlecht reduzierten signifikant die Darstellbarkeit der Strukturen. Der Nachweis einer galligen Sekretion aus dem Gallenblasenbett gelang in 3 Fällen (1,8%) durch eine FC. In 2 Fällen (1,2%) wurde der Abgang des DC aus dem Ductus hepaticus dexter durch IOC dargestellt und in einem Fall (0,6%) bereits durch FC beschrieben. Intraduktale Aussparungen ließen sich bei 9 Patienten (5,3%) mithilfe einer IOC und bei einem Patienten (0,6%) durch eine FC nachweisen. Bei Verzicht auf eine zusätzliche intraoperative Diagnostik betrugen die Detektionsraten von DC und DHC60,0 bzw. 43,3%, mithilfe einer FC80,0 bzw. 53,3%. Dementsprechend bestätigten die Operateure in 70,0% der Fälle eine Erhöhung der operativen Sicherheit durch eine FC.Die FC ist eine einfache, nicht invasive Methode zur Echtzeitdarstellung der Gallengangsanatomie während einer LC. Eine schnelle Identifizierung von anatomischen Normvarianten und galliger Sekretion aus dem Gallenblasenbett erhöht die operative Sicherheit und ermöglicht die simultane Versorgung. Bei Vorliegen einer intraabdominalen Adipositas besitzt die Untersuchung eine limitierte Aussagekraft.

Rosette-forming glioneuronal tumors (RGNTs) are rare glioneuronal tumors of the fourth ventricle region that preferentially affect young adults. Despite their histologic similarity with pilocytic astrocytomas (PAs), RGNTs do not harbor KIAA1549-BRAF fusions or BRAF mutations, which represent the most common genetic alteration in PAs. Recently, mutations affecting the hotspot codons Asn546 and Lys656 of fibroblast growth factor receptor 1 (FGFR1) have been described in PAs. They are considered to be the most frequent mechanism of mitogen-activated protein kinase activation, alternative to KIAA1549-BRAF fusion and BRAF mutations. To uncover possible molecular similarities between RGNTs and PAs, we performed a mutational study of FGFR1 in 8 RGNTs. An FGFR1 N546K mutation and an FGFR1 K656E mutation were found in the tumors of 2 patients. Notably, the patient with an FGFR1 K656E mutated RGNT had undergone a resection of a diencephalic pilocytic astrocytoma with pilomyxoid features 5 years before the discovery of the fourth ventricle tumor; the mutational analysis uncovered the presence of the same FGFR1 K656E mutation in the diencephalic tumor. These results indicate that, in addition to histologic similarities, at least a subgroup of RGNTs may show close molecular relationships with PAs. Whether FGFR1 mutated RGNTs represent a specific subset of this rare tumor entity remains to be determined.

There are a number of different surgical approaches in middle cerebral artery (MCA) aneurysm surgery. Evolution from the classical pterional approach towards smaller modified approaches took place over the years. In the present report, we describe a new modified approach in the treatment of MCA aneurysms, which is almost exclusively subfrontal. A modified approach was used on three patients with MCA bifurcation aneurysms. Craniotomy was subfrontal and suprapterional with minimal dissection of the temporal muscle and no drilling of the pterion. In all three cases, after establishing proximal control and dissecting the M1 carefully, retraction of the frontal lobe elevated the sylvian fissure and allowed opening of the fissure. The aneurysm could be identified easily and clips were applied. There was no infection and complete aneurysm clipping was achieved in all 3 patients. The described minimal craniotomy to the MCA through a subfrontal-suprapterional approach allows dissection of peripheral MCA bifurcation aneurysms without any problems.

A 57-year-old man with a history of chemotherapy because of cutaneous lymphoma presented with an orbital apex syndrome. The cranial computed tomography scan revealed a tumour in the orbital apex, extending intradurally. With a suspected diagnosis of a neoplastic lesion, the patient underwent orbital surgery with optic nerve decompression. Histology revealed an aspergilloma. No other foci were seen and treatment with antifungals was started. In immunocompromised patients with intracranial tumours, infection is always a major consideration in the differential diagnosis, even if the reason for immunosuppression (in this case chemotherapy) dates back several months. Misdiagnosing an orbital apex lesion as a cancer and treating patients primarily with corticosteroids can be life threatening. Removal or biopsy of such lesions is essential in further treatment since antifungals have to be administered as fast as possible.

Posttrauma apoptosis resistance of neutrophils (PMN) is related to overshooting immune responses, systemic inflammatory response syndrome (SIRS) and multiple organ failure (MOF). Recently, we have shown that the apoptosis resistance in circulating PMN from severely injured patients which is known to be mediated by high serum levels of pro-inflammatory cytokines can be overcome by the activation of Fas death receptor. Here, we aimed to study whether stimulation of surface Fas leads to the inactivation of hyperactivated PMN from critically ill patients with SIRS. PMN from 23 multiple trauma patients (mean injury severity score (ISS) 34±1.9) were isolated at day 1 after admission to the trauma center. PMN from 17 volunteer blood donors served as controls. Neutrophil activity has been determined after ex vivo short (1 h) and long-term (4 h) stimulation of freshly isolated PMN with immobilized agonistic anti-Fas antibodies. We found neutrophil chemotactic migration in response to IL-8, phagocytosis and oxidative burst to be significantly inhibited in control cells already after short-term (1 h) Fas stimulation. In contrast, inactivation of trauma PMN by agonistic anti-Fas antibodies was found to be efficient only after long-term (4 h) incubation of cells with agonistic antibodies. Thus, in trauma PMN down-regulation of neutrophil activity seems to be delayed when compared to cells isolated from healthy controls, suggesting impaired susceptibility for Fas stimulation in these cells. Interestingly, whereas Fas-mediated inhibition of phagocytosis and oxidative burst could be prevented by the broad range caspase inhibitor t-butoxycarbonyl-aspartyl(O-methyl)-fluoromethyl ketone (BocD-fmk), the chemotactic activity in response to IL-8 was unaffected. In conclusion, we demonstrate that stimulation of neutrophil Fas does not only initiate apoptosis but also induces inhibition of neutrophil functions, partially by non-apoptotic signaling.

α-Hederin, a natural triterpene saponin and its derivative kalopanaxsaponin I (ksI) exhibit cytotoxicity against various cancer cell lines and IN VIVO tumors. We studied the genetic variants contributing to the activity of these two anticancer compounds. Cell lines derived from 30 trios of European descent (Centre d'Etude du Polymorphisme Human, CEPH; CEU) and 30 trios of African descent (Yoruban, YRI) were used. Cytotoxicity was determined as inhibition of cell growth at increasing concentrations of α-hederin or ksI for 24 h. In comparison to the European, the Yoruban populations revealed a higher sensitivity to α-hederin and to ksI that can be attributed to several unique SNPs. These SNPs are located near 111 and 130 genes in the European and the Yoruban populations, respectively, raising the possibility that some of these genes contribute to the differential sensitivity to these compounds.

To develop highly concentrated therapeutic antibodies enabling convenient subcutaneous application, well stabilizing pharmaceutical formulations with low viscosities are considered to be key. The purpose of this study is to select specific amino acid combinations that reduce and balance aggregation, fragmentation and chemical degradation, and also lower viscosity of highly concentrated liquid antibodies. As a model, the therapeutically well-established antibody trastuzumab (25->200 mg mL-1 ) in liquid formulation is used. Pre-testing of formulations based on a stabilizing and protecting solutions (SPS®) platform is conducted in a thermal unfolding model using differential scanning fluorimetry (DSF) and accelerated aging at 37 and 45 °C. Pre-selected amino acid combinations are further iteratively adjusted to obtain stable highly concentrated antibody formulations with low viscosity. Size exclusion chromatography (SE-HPLC) reveals significantly lower aggregation and fragmentation at specific amino acid:sugar and protein:excipient ratios. Dynamic viscosities

The systemic inflammatory response syndrome and subsequent organ failure are mainly driven by activated neutrophils with prolonged life span, which is believed to be due to apoptosis resistance. However, detailed underlying mechanisms leading to neutrophil apoptosis resistance are largely unknown, and possible therapeutic options to overcome this resistance do not exist. Here we report that activated neutrophils from severely injured patients exhibit cell death resistance due to impaired activation of the intrinsic apoptosis pathway, as evidenced by limited staurosporine-induced mitochondrial membrane depolarization and decreased caspase-9 activity. Moreover, we found that these neutrophils express high levels of antiapoptotic Mcl-1 and low levels of proapoptotic Bax protein. Mcl-1 up-regulation was dependent on elevated concentrations of GM-CSF in patient serum. Accordingly, increased Mcl-1 protein stability and GM-CSF serum concentrations were shown to correlate with staurosporine-induced apoptosis resistance. However, cross-linking of neutrophil Fas by immobilized agonistic anti-Fas IgM resulted in caspase-dependent mitochondrial membrane depolarization and apoptosis induction. In conclusion, the observed impairment of the intrinsic pathway and the resulting apoptosis resistance may be overcome by immobilized agonistic anti-Fas IgM. Targeting of neutrophil Fas by immobilized agonistic effector molecules may represent a new therapeutic tool to limit neutrophil hyperactivation and its sequelae in patients with severe immune disorders.

Neutrophil extracellular traps (NETs) have recently been discovered as a central part of antimicrobial innate immunity. In the meanwhile, evidence accumulated that NETs are also generated upon non-infectious stimuli in various clinical settings. In acute or chronic inflammatory disorders aberrantly enhanced NET formation and/or decreased NET degradation seems to correlate with disease outcome. This review summarizes current knowledge about the relation of NETs in a broad spectrum of clinical settings. Specifically, we focus on the importance of NETs as a predictive marker in severely ill patients and further, we speculate about the potential pathophysiology of NETs.

Study Design. A prospective analysis. Objective. The idea of this study was to evaluate a new miniature robotic system providing passive guidance for pedicle screw placement at the lumbar spine. Special focus was laid on the postoperative accuracy of screw placement. and Background Data. Recent technical developments lead to a minimization of pedicle screw fixation techniques. However, the use of navigational tech!: niques is still under controversy. Methods. Patients selected for a minimal invasive posterior lumbar interbody fusion received a spiral computer tomographic scan before surgery. The miniature hexapod robot was mounted to the spinous process and the system moves to the exact entry point according to the trajectory of the surgeon's preoperative plan. After minimal invasive screw placement all patients received routinely a postoperative spiral computer tomographic scan. Screws placed exactly within the pedicle were evaluated as group A, screws deviating

The release of "neutrophil extracellular traps" (NETs) has been identified as a novel immune response in innate immunity. Neutrophil extracellular traps are composed of neutrophil-derived circulating free DNA (cf-DNA), histones, and neutrophil cytoplasm-derived proteins such as proteases. Here, we studied the putative predictive value of plasma cf-DNA/NETs for the development of sepsis and mortality after multiple trauma. In a prospective pilot study with 45 multiple trauma (Injury Severity Score>16) patients, cf-DNA was directly quantified in plasma. Blood samples were sequentially obtained daily from admission to our Trauma Center until day 10. Because of limited intensive care unit (ICU) stay of less than 3 days, 8 patients have been excluded, resulting in 37 patients that were evaluated. Time kinetics of cf-DNA/NETs was compared with C-reactive protein (CRP), interleukin (IL) 6, leukocyte counts, and myeloperoxidase. The severity of the injury was calculated on the basis of the Injury Severity Score, as well as Multiple Organ Dysfunction Score, Sequential Organ Failure Assessment, and Simplified Acute Physiology Score II on ICU. Initially high cf-DNA/NETs values (>800 ng/mL) with recurrent increased values between days 5 to 9 were associated with subsequent sepsis, multiple organ failure, and death. In conjunction with cf-DNA/NETs, IL-6 was significantly elevated after admission. However, the development of a second hit was not indicated by IL-6. In contrast to cf-DNA/NETs, no difference in CRP kinetics was observed between patients with and without development of sepsis. Circulating free DNA/NETs kinetics rather followed kinetics of Multiple Organ Dysfunction Score, Sepsis-related Organ Failure Assessment, leukocyte counts, and partially of myeloperoxidase. Circulating free DNA/NETs seems to be a valuable additional marker for the calculation of injury severity and/or prediction of inflammatory second hit on ICU. However, a large clinical trial with severely injured patients should confirm the prognostic value of neutrophil-derived cf-DNA/NETs.

Automatic segmentation of anatomical structures in medical images is a valuable tool for efficient computer-aided radiotherapy and surgery planning and an enabling technology for dynamic adaptive radiotherapy. This paper presents the design, algorithms and validation of new software for the automatic segmentation of CT images used for radiotherapy treatment planning. A coarse to fine approach is followed that consists of presegmentation, anatomic orientation and structure segmentation. No user input or a priori information about the image content is required. In presegmentation, the body outline, the bones and lung equivalent tissue are detected. Anatomic orientation recognizes the patient's position, orientation and gender and creates an elastic mapping of the slice positions to a reference scale. Structure segmentation is divided into localization, outlining and refinement, performed by procedures with implicit anatomic knowledge using standard image processing operations. The presented version of algorithms automatically segments the body outline and bones in any gender and patient position, the prostate, bladder and femoral heads for male pelvis in supine position, and the spinal canal, lungs, heart and trachea in supine position. The software was developed and tested on a collection of over 600 clinical radiotherapy planning CT stacks. In a qualitative validation on this test collection, anatomic orientation correctly detected gender, patient position and body region in 98% of the cases, a correct mapping was produced for 89% of thorax and 94% of pelvis cases. The average processing time for the entire segmentation of a CT stack was less than 1 min on a standard personal computer. Two independent retrospective studies were carried out for clinical validation. Study I was performed on 66 cases (30 pelvis, 36 thorax) with dosimetrists, study II on 52 cases (39 pelvis, 13 thorax) with radio-oncologists as experts. The experts rated the automatically produced structures on the scale 1-excellent (no corrections necessary, maximum time saving), 2-good (corrections necessary for up to 1/3 of slices), 3-acceptable (major corrections necessary, but still time saving), 4-not acceptable (manual redrawing more efficient, no time saving). A rating

Up to now, only little is known about hydrocephalus (HC) in vein of Galen malformation (VGM). We want to present the different etiologies and our long-term experience (1992–2015) in the management of HC. Out of 44 treated children with VGM, we retrospectively reviewed all cases with HC. We analyzed the etiologies, our treatment results and complications. Twenty-one children (48 %) presented either with HC or developed it over time. In 21 % of those cases, high venous pressure was presumably the sole cause. Until 2009, seven of them received ventriculoperitoneal (VP) shunting; six of those resulted in severe postoperative complications. The remaining children have been treated successfully by endovascular embolization. Five out of the 44 children (11 %) developed HC after intraventricular hemorrhage. In four cases, those children were treated with positive results by using transient external ventricular drainages. In one case a VP shunt with highest valve pressure was inserted. Another four children (9 %) presented with aqueductal stenosis-related HC caused by either dilated venous outflow or space-occupying coil masses after embolization. The latter case was successfully treated by ventriculocisternostomy, whereas endovascular treatment decreased the venous outflow in size and thus resolved the HC in the other cases. In the remaining cases (7 %), atrophy due to melting brain syndrome led to HC ex vacuo. HC in VGM is a common phenomenon with several etiologies requiring different treatments. In most cases, embolization of the VGM as sole treatment is completely sufficient in order to decrease high venous pressure. However, certain other causes of HC should be treated in an interdisciplinary setting by specialized neurosurgeons.

The aim of this study was to adapt the ultrasonographical techniques developed for brain perfusion imaging to an intraoperative setting for topographic diagnosis of cerebral tumours. During surgery, the patients underwent contrast-enhanced ultrasonography (phase inversion harmonic imaging, bolus kinetic, fitted model function). Endocavity curved array (6.5EC10, 6.5 MHz) was used intraoperatively. The ultrasound contrast agent SonoVue (Bracco) was administered IV as a bolus injection. Off-line, time-intensity curves as well as perfusion maps were calculated and parameters such as peak intensity were locally extracted to characterise perfusion. Seven patients with brain tumours of different histologic types were subjected to contrast-enhanced ultrasonography during surgery. Tissue differentiation with contrast agent was superior to conventional B-mode ultrasound imaging. Intraoperative contrast-enhanced ultrasonography enabled visualisation of cerebral tumours in high spatial resolution.

Despite the fact that neutrophils are essential for the protection from invading pathogens, hyperactive neutrophils may elicit detrimental cerebral damage after severe trauma. The neutrophil interactions with the neurovascular unit entail endothelial dysfunction involving endothelial leakage, formation of edema, coagulation abnormalities, disturbed hemodynamics, tissue infiltration etc. These elements of the "whole body inflammation," designated systemic inflammatory response syndrome (SIRS) in conjunction with intracerebral proinflammatory activities, are important triggers of post-traumatic cerebral damage and mortality according to the "second hit" concept. From the immunologic point of view, the brain is an immune privileged site, known to resist autodestructive inflammatory activity much more efficiently than other organs because of the highly efficient diverse functions of the blood-brain barrier (BBB). However, both the underlying strategy of the BBB to maintain cerebral protecting functions against the post-traumatic neutrophil-mediated "second hit" and how activated neutrophils may overcome the BBB are currently unknown. Therefore, this review summarizes the current understanding of the "second hit," the BBB physiology, and its role in the maintenance of cerebral immune privilege, and discusses recent findings that may explain the pathophysiologic neutrophil-BBB interactions occurring after severe trauma, thus offering novel therapeutic options to protect from post-traumatic brain damage.

Background: In patients after anterior cervical discectomy (ACD) with fusion newly developed retrospondylophytes or incomplete decompression of the nerve root can cause recurrent radicular pain. Anterior cervical uncoforaminotomy (uncoforaminotomy) is an operative method which removes the causative degenerative pathology at the level of the neural foramen leaving untouched the inserted graft at this level. Method: Between February 2004 and April 2005, 7 patients underwent uncoforaminotomy after ACD with fusion for the treatment of recurrent cervical radiculopathy in our neurosurgical department. Prior to treatment patients received a computed tomography (CT) and a neurological examination. Anterior uncoforaminotomy was performed thereafter (for technical details see publication by Jho, 1996). A postoperative CT scan was done before discharge. Follow-up examination was performed eight weeks after surgery. Findings: Five patients underwent the operation at C5/6, one patient was operated at C6/7 and one patient had the operation at two levels (C5/6 and C6/7). At discharge six patients had excellent or good results. Conclusion: Uncoforaminotomy is a good method for the treatment of newly acquired spondylotic spurs in the foramen or incomplete osseous decompression after ACD with fusion and recurrent radicular pain.

Object Cage subsidence occurs after anterior cervical discectomy and fusion (ACDF). The aim of this prospective study was to evaluate subsidence and total segmental height after implantation of a newly designed Wing titanium cage. Furthermore, alignment of the entire cervical spine was analyzed 2 years after surgery. Methods Fifty-four patients (26 women and 28 men) whose mean age was 48.3 years underwent ACDF. Follow-up examinations were performed at discharge and 6, 12, and 24 months postoperatively by an independent investigator. The clinical course was evaluated using the visual analog pain scale and the Prolo scales. Measurements of subsidence and total segmental height were conducted, and the alignment of the entire cervical spine was classified using two methods. In 54 patients 64 levels were fused. The patients noted a significant reduction of pain, and scores on both Prolo scales were significantly improved. At the 2-year follow-up examination, subsidence was present in 30 of the 67 fused segments. There was a statistically significant correlation between subsidence and the presence of posterior spondylosis at the initial surgery. Furthermore, there was a significant correlation between reduction of total segmental height and the presence of subsidence; however, subsidence did not prevent the development of a solid bone arthrodesis (fusion rate 98%) or have an adverse effect on the alignment of the cervical spine. Conclusions Titanium Wing cage–augmented ACDF was associated with comparatively good long-term results. Subsidence was present but did not cause clinical complications. Furthermore, radiological studies demonstrated that the physiological alignment of the cervical spine was preserved and a solid bone arthrodesis was present at 2 years after surgery.

PURPOSE. In addition to neuroinvasive disease, West Nile virus (WNV) infection is frequently associated with self-limiting chorioretinitis and vitritis. However, the mechanisms of ophthalmic WNV infection are rarely investigated, in part because of the lack of reliable in vitro models. The authors therefore established the first model of ocular WNV infection and investigated interaction of WNV with IFN signal-transduction mechanisms. METHODS. Human retinal pigment epithelial (RPE) cells were infected with WNV strain NY385-99 at a multiplicity of infection of 5. Virus replication was evaluated by virus titers at different times after infection. The susceptibility of RPE cells to WNV infection was confirmed by transmission electron microscopy. IFN- expression was assessed by quantitative real-time PCR and by measurements of antiviral activity in cell culture supernatants. IFN signaling was evaluated by phosphorylation of transducer and activator of transcription 1 and 2 (STAT1/2) proteins, with immunoblot analysis. RESULTS. RPE cells appeared to be highly sensitive to WNV infection. Maximum viral titers were found 24 hours after infection, followed by a continuous decline during the course of infection. WNV infection of RPE cells was followed by increased IFN- expression associated with IFN signaling and subsequent inhibition of WNV replication. CONCLUSIONS. In this study, the first cell culture model of ophthalmic WNV infection was developed and characterized in RPE cells, and the molecular mechanisms of WNV infection were studied. The data suggest that WNV induces a general antiviral state in RPE cells. This general antiviral state correlates with WNV-induced IFN signaling in retinal cells. (Invest Ophthalmol Vis Sci. 2006;47:645‐651) DOI:10.1167/iovs.05-1022

In the European countries there is a lack of young doctors. This shortage limits the possibilities for recruiting skilled surgeons for the microsurgical disciplines. Complicating the situation is the fact that most students do not decide on their area of specialization until the late clinical semesters. The authors present a new program of microsurgical training that dispenses completely with animal training. In addition, a scoring system is presented that enables instructors to compare the microsurgical skills of different students for the purpose of further statistical analysis. This scoring system could be used for the evaluation of potential microsurgical candidates as well as for other purposes, e.g. scientific projects. A total of 36 students with an average age of 24.2 were trained, with good-to-excellent results. Microsurgical training of young students can be recommended. A point worth exploring in the future is whether there are different learning curves for different age groups.

Systemic administration of immune modulating antibodies may play an important role in reducing neutrophil hyperactivity, for example, in patients undergoing cardiac surgery with extracorporeal circulation or in trauma patients. However, this strategy has extremely high costs and is often associated with severe adverse effects. We developed the Leukocyte-Inhibition-Module (LIM), an extracorporeal circulation (ECC) device housing a polyurethane matrix with covalently bound Fas (CD95; APO-1) stimulating antibodies to rapidly prevent neutrophil hyperactivation. A feasibility study with 14 patients undergoing cardiac surgery with the use of immunogenic ECC without (n = 5) and with (n = 9) LIM (venous line) was performed. Our data show that the usually observed ECC associated perioperative increase in neutrophils (control) was prevented by LIM (p = 0.023). Moreover, the increase of the proinflammatory markers tumor necrosis factor (TNF)-alpha and polymorphonuclear elastase was limited by LIM (p = 0.038 and p = 0.002). In both groups, no significant changes in liver enzymes or in clotting were detected after surgery, and up to 12 months follow up, no unusual complications were reported. This study shows for the first time to our knowledge the feasibility, efficacy, and safety of a new cost effective, immune management strategy in patients with aberrant immune activation by exposing the blood stream to immobilized agonistic anti-Fas antibodies.

During the development of new navigation systems, the ergonomic aspects of the accompanying software are seldom taken into account. The VN system is a navigation system working with real and previously stored neuroendoscopic images. The latter enable the module to execute virtual back movement, e.g. in case of bleeding. Several other modules are offered. The aim of the study was the ergonomic analysis of different modules and their learning curves in cadaveric heads. The endoscope was navigated by the neurosurgeon while another person operated the computer. A total of 128 experiments were performed with two software versions. When the landmark tracking module was used, a real learning curve could be observed. By contrast, testing the measurement module did not produce a learning curve. A significant reduction of the time required by the three modules investigated could be observed with software version 2. The module for virtual back movement works best with a minimum time of 20 s for image storage. During machine-human interactions ergonomic software use is important, especially if operative procedures are performed.

Severe acute respiratory syndrome (SARS) is caused by the SARS coronavirus (SCV). The disease appeared in the Guandong province of southern China in 2002. The epidemic affected > 8422 patients and caused 908 deaths in 29 countries on 5 continents. Several treatment modalities were tried with limited success to treat SARS and a variety of experimental drugs are under development. Type I interferons (IFNs-alpha/beta) were suggested as potential candidates to treat SARS. Several animal and human coronaviruses, including SCV, were shown to be sensitive to IFNs both in vitro and in vivo. A pilot clinical report showed effectiveness of IFN-alpha for the treatment of SARS patients. This review summarises antiviral activities of IFNs with special regard to SARS, and reviews the published clinical and experimental data describing the use of IFNs for SARS.

Recently, the term oncomodulation has been proposed to express the ability of human cytomegalovirus (HCMV) to modify tumor cell biology, a phenomenon that is independent from transformation. Because past studies have failed to show that HCMV can transform normal human cells, HCMV has not been regarded as an oncogenic tumor virus. However, recent investigations have revealed a high frequency of HCMV in tumor cells of malignancies such as colon cancer, malignant glioma, prostatic intraepithelial neoplasia, and carcinoma. Data from experiments with HCMV-infected tumor cell lines have highlighted the oncomodulatory potential of HCMV and provided important insights into the patho-mechanisms associated with aberrant signaling pathways and transcription factor and/or tumor suppressor function of the host cell.

Cardiac surgery with cardiopulmonary bypass (CPB) is associated with neutrophil activation, inflammation, and consecutive edema. The impairment of endothelial junction molecules, and thus, hyperpermeability elicited by the interaction of activated neutrophils with endothelial cells may be important in this regard. Cocultures with human endothelial cells and neutrophils from 10 cardiac surgery patients with CPB were used to evaluate the role of neutrophils in modifications of the endothelial zonula adherens molecules VE-cadherin and beta-catenin. Laser scan microscopic analyses showed that neutrophils, which were isolated after the beginning of CPB, significantly impaired intracellular redistribution of endothelial beta-catenin with regard to membrane association (p.0002) and staining pattern (p.0001). VE-cadherin localization was not found to be significantly modified. Western blots with total cell extracts showed that amounts of beta-catenin did not vary significantly after co-culture with activated neutrophils. Activated neutrophils during cardiac surgery with CPB may induce endothelial dysfunction by impairing beta-catenin localization and thus contribute to endothelial hyperpermeability.

Brain abscesses are life-threatening and sometimes difficult to detect. A brain abscess after placement, manipulation of a bone anchored hearing aid, or a periauricular implant for fixation of an ear prosthesis has never been reported in the literature.A 42-year-old man suffered from a right-sided temporodorsal brain abscess after change of a bone anchored hearing aid abutment. The fixture itself had been inserted 8 years before without any complications in the peri- or postoperative period. A CT-guided puncture of the abscess could be performed via the screw-hole in the temporal bone after removal of the fixture, and the patient was treated with antibiotics.The outcome of the procedure was good without neurologic deficits for the patient.The insertion of periauricular screw implants bears the risk of meningeal lesions as well as a small risk of purulent intracranial and intracerebral complications perioperatively or in the context of later manipulations. Minimally invasive therapy of such brain abscesses can be performed by removal of the foreign body, CT-guided puncture, and antibiotic medication.

Cardiac surgery with cardiopulmonary bypass (CPB) results in transient cerebral swelling in most patients. Cognitive decline occurs in 24-57% of patients and 2-5% experience stroke. Serum levels of S-100B, a potential marker of increased blood-brain barrier permeability, increase during and early after surgery. The authors studied the effects of the novel Na /H exchange inhibitor cariporide (HOE642) on postoperative serum levels of S-100B and neuron-specific enolase (NSE) in 53 patients at high risk undergoing coronary artery bypass grafting. Patients were randomly assigned to one of four groups: I, placebo; II, 20 mg cariporide; III, 80 mg cariporide; IV, 120 mg cariporide). In addition, the leukocyte activation marker myeloperoxidase (MPO) and malondialdehyde (MDA), a marker for lipid peroxidation, were evaluated by enzyme-linked immunoassay. Postoperatively, five patients experienced transient ischemic attack or stroke. S-100B levels increased from 0.43 microg/l +/- 0.33 before operation to 2.27 microg/l +/- 0.69 1 hour after surgery in the placebo group. Preoperative S-100B levels in the HOE642 groups did not differ from the placebo group whereas, 1 hour after surgery, levels were significantly lower in groups II, III, and IV (1.63 microg/l +/- 0.2, 1.27 microg/l +/- 0.27, and 0.90 microg/l +/- 0.21, respectively). NSE, MPO, and MDA serum levels did not differ among groups. These findings may stimulate larger clinical studies to examine the effects of HOE642 on cerebral swelling and neurologic/cognitive outcome of cardiac surgery with CPB.

Dear Editor, We report the case of a 44-year-old female patient with lumbo-ischialgic pain projecting to dermatomes S1. Conservative treatment was unsuccessful and the pain progressed until the patient was unable to walk. After lumbar computed tomography (CT) evaluation, a radicular compression due to listhesis could be seen in the level L5/S1 (Meyerding Grade I, Fig. 1a). Radicular decompression and screw fixation from L5 to S1 followed. The postoperative CT shows good screw position and dural decompression (Fig. 1b). At this point it is important to mention that the patient had allergies against sodium thiosulphate (dental products), 1,3-diphenylguanidine (rubber) and nickel (II) sulphate. No titanium allergy. Immediately post-surgery, the patient still complained of lumbo-ischialgic pain with a 7/10 visual analogue scale (VAS) score. Because of pain, ambulation was impossible (despite good postoperative results on CT). Inspection of the wound showed oedema, redness and hyperthermia resembling the clinical signs of wound infection. After microbiological screening (Staphylococcus epidermidis, Streptococcus oralis, Streptococcus faecalis and Staphylococcus haemoliticus positive), we started antibiotic treatment even in absence of leucocytosis or increased C-reactive protein. The pain persisted and a lumbar magnetic resonance imaging (MRI) 2 weeks post-surgery failed to show signs of post-operative infection. Irrespective of absence of inflammatory proteins in the serum and/or leucocytosis and under antibiotic treatment, there was putrid wound excretion. Surgical wound debridment followed. Intraoperativelly there was no dislocation of the screws. There were no signs of infection under the skin (i.e. epidural or lumbar bone). After wound-debridment there was still no improvement of pain (VAS 6/10). Creactive protein was normal. Because of persistence of symptoms, another lumbar CT followed 7 weeks postsurgery. No signs of screw dislocation were seen, but at this time there was subcutaneous air identifiable (subcutaneous infection?). MRI and serum failed to show infection. In a second wound debridment, granulation tissue was removed epidurally. The whole time the patient received antibiotic treatment. After two wound revisions we performed an epicutaneus allergy test. A titan screw like the ones used for fixation in the patient’s case was adhered for 48 h on the patient’s upper arm. After 48 h the patient exhibited an allergic reaction of the type IV (allergic contact dermatitis) (Fig. 2). Steel and PEEK cage did not resemble allergic reactions on the patient’s skin. In a third revision we changed the titanium screws with steel screws and a PEEK cage was placed in the intervertebral space L5/S1. Histopathologically, fat and muscle tissue from the wound showed lymphocytic inflammation and chronic inflammatory infiltrates. Immediately after the replacement of the titanium screws the patient showed an improvement of her symptoms (VAS 1/10), and could be discharged with minimal postoperative pain. In the MELISA (Memory Lymphocyte Immunostimulation Assay) test and the lymphocyte transformation test, no Rashad El Habony and Yucel Binay contributed equally

INTRODUCTION: Owing to increasing numbers of decompressive craniectomies in patients with malignant middle cerebral artery infarction, cranioplastic surgery becomes more relevant. However, the current literature mainly consists of retrospective single-centre (evidence class III) studies. This leads to a wide variability of technical approaches and clinical outcomes. To improve our knowledge about the key elements of cranioplasty, which may help optimising clinical treatment and long-term outcome, a prospective multicentre registry across Germany, Austria and Switzerland will be established. METHODS: All patients undergoing cranioplastic surgery in participating centres will be invited to join the registry. Technical methods, materials, medical history, adverse events and clinical outcome measures, including modified Rankin scale and EQ-5D, will be assessed at several time points. Patients will be accessible to inclusion either at initial decompressive surgery or when cranioplasty is planned. Scheduled monitoring will be carried out at time of inclusion and subsequently at time of discharge, if any readmission is necessary, and at follow-up presentation. Cosmetic results and patient satisfaction will also be assessed. Collected data will be managed and statistically analysed by an independent biometric institute. The primary endpoint will be mortality, need for operative revision and neurological status at 3 months following cranioplasty. ETHICS AND DISSEMINATION: Ethics approval was obtained at all participating centres. The registry will provide reliable prospective evidence on surgical techniques, used materials, adverse events and functional outcome, to optimise patient treatment. We expect this study to give new insights in the treatment of skull defects and to provide a basis for future evidence-based therapy regarding cranioplastic surgery. TRIAL REGISTRATION NUMBER: This trial is indexed in the German Clinical Trials Register (DRKS-ID: DRKS00007931). The Universal Trial Number (UTN) is U1111-1168-7425. peerReviewed

Objective and design: The beneficial effects of leukocyte filtration on the outcome of cardiac surgery with cardiopulmonary bypass (CPB) is probably due to the limitation of pathogenesis mediated by over-stimulated neutrophils. However, the influence of leukocyte filtration on the functional neutrophil activity has not been studied in detail. Therefore, by using different filtration timing strategies we determined neutrophil effector functions before and after the filter passage as well as blood surrogate markers for neutrophil activation.¶Methods: We randomly assigned 80 cardiac surgery patients to four groups (n = 20 each) without (I) and with three different filtration timing strategies (II–IV). As functional end points neutrophil phagocytic activity and oxidative burst upon ex vivo stimulation with E.coli were analyzed from blood of 31 patients whereas polymorphonuclear elastase (PMNE), myeloperoxidase, and malondialdehyde (MDA), a marker for lipid peroxidation was determined in plasma samples from 80 patients. Blood was harvested immediately before and behind the filter (Pall LG6) at different times during CPB.¶Results: We found that none of the filtration strategies either reduced the number of neutrophils capable of eliciting phagocytic activity and oxidative burst or the activity per cell. In contrast, PMNE and MPO levels in peripheral venous blood were found to be significantly increased in groups II–IV compared with group I throughout the entire filtration period in all patients. MDA was not enhanced in the filter groups.¶Conclusions: Our results show that the leukocyte depletion filter in the arterial line of the heart-lung machine failed to limit neutrophil stimulation but rather augmented PMNE plasma levels. We speculate that augmented PMNE and MPO levels mainly stem from neutrophils that are captured within the mesh of the leukocyte filter.

Leukocyte filtration has evolved as an important technique in cardiac surgery with cardiopulmonary bypass to prevent pathogenic effector functions mediated by activated leukocytes. The underlying mechanisms that result in an improvement of laboratory variables as well as clinical outcome are not resolved yet. Moreover, the optimum strategy for the use of current filtration technology has not been systematically evaluated. This paper, therefore, reviews how activated leukocytes may lead to tissue damage, summarizes the known effects of leukocyte filtration on clinical outcome and laboratory parameters, and deals with current experimental and clinical efforts to further limit the pathogenic effects of leukocytes in cardiac surgery.

The human CMV (HCMV) is a persistent virus that may cause severe inflammatory responses especially in immunocompromised hosts. In different cell types, HCMV infection leads to the activation of the pleiotropic transcription factor, NF-κB, which triggers virus replication but also propagates cell-mediated inflammatory mechanisms that largely depend on PG synthesis. We investigated the interactions of HCMV and the NF-κB-dependent PG synthesis pathway in cultures of retinal pigment epithelial (RPE) cells that are known to be infected in HCMV retinitis patients. Unlike in other cell types, HCMV increased neither NF-κB activity nor p65 and p105/50 mRNA levels in RPE cells. Both TNF-α and phorbol ester 12,0-tetradecanoylphorbol 13-acetate (TPA) enhanced NF-κB activity but only TPA increased HCMV replication. Cyclooxygenase-2 expression and PGE2 release was increased by TPA and TNF-α but not by HCMV infection. Stimulatory activity of TPA on HCMV replication was suppressed by protein kinase C inhibitors and inhibitors of p42/44 and p38 mitogen-activated protein kinases but not by NF-κB inhibitors. In conclusion, HCMV circumvents the NF-κB route in favor of the protein kinase C-dependent mitogen-activated protein kinase pathway in RPE cells. This virus/host cell interaction might be a mechanism that promotes HCMV persistence in immune-privileged organs such as the eye.

Improved myocardial protection and cardiopulmonary bypass (CPB) have limited, but not abolished, intraoperative myocardial damage due to surgical reperfusion injury after release of the aortic crossclamp. In this double-blind, randomized study, we evaluated whether short-term leukocyte filtration during reperfusion may further reduce myocardial damage. Thirty-eight patients with coronary artery disease were randomly assigned to CPB with (group I; n = 19) or without leukocyte filtration (group II; n = 19). There was no difference in bypass time or crossclamp time between the groups. No patient in group I required catecholamines, whereas three patients in group II were supported with adrenaline or dobutamine on the first and second postoperative day. In addition, troponin T plasma levels were lower in group I ( p < 0.05), whereas other markers for tissue injury (CK, CK-MB, LDH, S- GOT and S100B) did not differ. In conclusion, leukocyte filtration during reperfusion may further improve CPB by reducing myocardial damage.

Thrombospondin-1 (TSP-1) is a potent inhibitor of angiogenesis. It has been shown that promoter sequences of the TSP-1 gene can be transactivated by the wild-type tumor suppressor protein p53. As human cytomegalovirus (HCMV) infection inactivates wild-type p53 of various cell types, we investigated whether HCMV infection is associated with reduced TSP-1 production. We found, in conjunction with accumulated p53, that TSP-1 mRNA and protein expression was significantly reduced in HCMV-infected cultured human fibroblasts. To determine whether the observed TSP-1 suppression depends on p53 inactivation, the p53-defective astrocytoma cell line U373MG was infected with HCMV. In these cells TSP-1 expression was also significantly reduced by HCMV infection whereas expression of the p53 mutant variant remained unaltered. In both cell lines the decreased expression of TSP-1 mRNA occurred early after infection (4 hours), indicating that HCMV inhibits TSP-1 transcription during the immediate-early phase of infection before HCMV DNA replication. Inhibition of HCMV DNA synthesis by ganciclovir did not influence TSP-1 reduction whereas the antisense oligonucleotide ISIS 2922, complementary to HCMV immediate-early mRNA, completely prevented the HCMV-mediated TSP-1 suppression. These findings strongly suggest a novel role for HCMV in the modulation of angiogenesis due to p53-independent down-regulation of TSP-1 expression.

In this retrospective investigation 7 patients operated on for recurrent trigeminal neuralgia via a suboccipital enlarged burr hole trepanation were evaluated. The intraoperative findings indicate that neo-compressive effects due to foreign material used during prior surgery are more important than adhesions around the nerve. Furthermore, the results of recurrent operations are encouraging as 6 of the 7 patients were painfree without additional neurological deficits.

Cyclosporin A reduces the mitotic activity of allosensitized lymphocytes, but fails to limit emigration of these cells into the donor organ. However, the modulation of both lymphocyte proliferation and infiltration are desirable characteristics of immunosuppressive therapy. The calcium-channel blocker, verapamil, has recently been shown to effectively prevent the transmigration of CD4+ and CD8+ T cells through allogeneic endothelium. Mibefradil (Ro 40-5967) represents a new generation of calcium antagonists with high potency and long-term activity. To evaluate the immunosuppressive potential of this drug, the influence of mibefradil on lymphocyte adhesion to, horizontal locomotion along, and penetration through allogeneic endothelium (HUVEC) was performed. When lymphocytes were prestimulated for 24 hr with mibefradil, adhesion and penetration were dose-dependently reduced. The adhesion ID50 values were 3.4 microM (CD4+ T cells) versus 9.2 microM (CD8+ T cells) and 2.1 microM (CD4+ T cells) versus 3.9 microM (CD8+ T cells) with regard to penetration. Mibefradil also effectively blocked horizontal locomotion. Specific down-regulation of T-cell binding to the P-selection receptor (ID50: CD4+ T cells, 0.8 microM: CD8+ T cells, 1.2 microM) and to the intracellular adhesion molecule-1 (ICAM-1) receptor (ID50: CD4+ T cells, 1.9 microM; CD8+ T cells, 1.5 microM) by mibefradil seems to be responsible for the decreased adhesion and penetration rates. Reduction of intracellular F-actin in T lymphocytes could diminish cell locomotion. In conclusion, the potent suppressive properties of mibefradil support its use as a co-medication in cyclosporin A-based immunosuppressive therapy.

Primary cell cultures, such as isolated epithelial cells, neuronal cells, or hepatocytes are characterized by a very low mitotic activity. Monitoring of small changes in cell numbers requires staining with a DNA-specific dye with an extremely high sensitivity and a low inter- and intraassay variability. For this purpose, an ultrasensitive in vitro assay has been developed based on the fluorescent nucleic acid stain PicoGreen. PicoGreen has been shown to detect as little as 0.5 ng pure DNA or 10(2) cells (interassay SD < 10%, intraassay SD < 5%). This is far above the limit of sensitivity of conventional fluorochromes, such as Hoechst 33342 or propidium iodide. To obtain optimum efficacy of PicoGreen, cells were digested with papain for 20 h at 60 degrees C prior to staining. Under these conditions, the slope factor was calculated to be 0.105 relative fluorescence units (RFU)/cell, which is far superior to the slope factor of Hoechst 33342 (0.0137 RFU/cell) or propidium iodide (0.0077 RFU/cell). Analysis of the blank values revealed a very low autofluorescence of PicoGreen, which is only 1/50th of the autofluorescence of Hoechst 33342 and 1/5th of the autofluorescence of propidium iodide. Additional coating of the culture plates with extracellular matrix proteins to prevent cellular dedifferentiation did not influence the high sensitivity of PicoGreen. In conclusion, the PicoGreen-assay seems to be the method of choice when the growth capacity of primary cell cultures needs to be analyzed with high accuracy.

We studied the frequency of patients who had chronic subdural haematomas (CSDH) and Huntington's disease (HD) in a 1-year study period. In our department a total of 58 patients with CSDH were treated. Four patients (6.9% of them) had HD. Surgical evacuation of the haematoma was performed in all four cases with the use of a twist drill trepanation without a drainage system.

CMV disease is an important peristent factor for enhanced morbidity and mortality in immunocompromised patients. In high-risk patients the prevention of primary infection or reactivation of latent infection is endeavoured by means of various antiviral strategies. However, the prevention of CMV disease by immunomodulators appears to be a hopeful additional tool for the treatment of immunocompromised patients and ought to be further investigated.

This review summarizes current strategies for the treatment of human cytomegalovirus (CMV) infection/diseases in high-risk patients such as transplant recipients and AIDS patients. Since the major drugs ganciclovir (Cytovene), foscarnet (Foscavir) and cidofovir (Vistide) are frequently associated with severe side effects and the formation of viral resistance, it should be endeavored to develop better strategies in anti-CMV treatment. Moreover, blocking of the viral replication does not always resolve the manifestations which are often linked with CMV-associated immunopathomechanisms. Thus, the efficacy of the available drugs is also discussed in the light of their ability to modulate inflammatory components of the cell-mediated immune system.

Chromosome analysis of brain tumours can provide important pathobiological data; however, cytogenetic tools are so far not routinely applied for diagnosis. In the present study 25 paraffin embedded stereotactic biopsies from 19 glioma patients were studied using in situ hybridization of chromosome #10 and #15 using biotinylated pericentromeric probes. Numerical changes of chromosome #10 are frequent alterations in glioblastoma. Quantification of chromosome #15 served as a control in order to exclude artificial monosomies or nonspecific changes. The number of chromosomes in at least 200 cells were counted for each specimen. 18 of 25 biopsies could be evaluated quantitatively. The small volume of probes was not a limiting factor for analysis. Quantification of "nonspecific" chromosome #15 revealed single spots in 22-41% of all cells in the 18 biopsies. Chromosome #10 showed single spots in a range between 34 and 44% of counted nuclei in 13/18 biopsies. In 5 out of 18 biopsies 51-60% monosomies were found: in this subgroup were 4 high grade gliomas. These cases were interpreted as monosomy of chromosome #10. The results demonstrate feasibility and quantitative evaluability of cytogenetic analysis in stereotactic biopsy material using in situ hybridization.

The biliary epithelium is a major target for allograft-directed immune responses during rejection crises after liver transplantation. This paper deals with in vitro studies on the immunogenetic potential of cultured biliary epithelial cells (BECs) to elicit an allogeneic cellular immune response. Therefore, BECs were cocultured with syngeneic and allogeneic lymphocytes in order to study lymphocyte activation. The respective lymphocytes [3H]thymidine incorporation was a measure for the proliferative activity. While syngeneic peripheral blood lymphocytes (PBL) never exhibited BEC-induced proliferation allogeneic PBL were significantly (P0.05) activated in all experiments (n = 6). In experiments with purified subpopulations CD8+ cells but not CD4+ cells proved to be activated by BECs. In time kinetics (n = 5) the maximum of the BEC-induced proliferation was on day 9 while the endothelial cell-induced proliferation was found to be 2 days earlier on day 7 after onset of the experiments (P0.05). BEC-induced proliferation was accompanied by induced IL-2 secretion (300 pg/ml) by activated lymphocytes as determined by ELISA. Stimulation of BECs with 500 U/ml interferon-gamma, 1000 U/ml interferon-alpha or blocking expression of HLA molecules on the surface membrane of BECs by monoclonal antibodies did not alter BEC-induced allogeneic lymphocyte proliferation. Monoclonal antibodies against CD8+ but not CD4+ suppressed proliferative activity of PBL and CD8+ cells by 40 and 45%, respectively. Overall, these results provide evidence that BECs may induce CD8+ lymphocyte activation in vivo and therefore might play a crucial role in triggering immune responses related to liver transplant rejection episodes.