Complement Factor C5a Inhibits Apoptosis of Neutrophils-A Mechanism in Polytrauma?

Life-threatening polytrauma results in early activation of the complement and apoptotic system, as well as leukocytes, ultimately leading to the clearance of damaged cells. However, little is known about interactions between the complement and apoptotic systems in PMN (polymorphonuclear neutrophils) after multiple injuries. PMN from polytrauma patients and healthy volunteers were obtained and assessed for apoptotic events along the post-traumatic time course. In vitro studies simulated complement activation by the exposure of PMN to C3a or C5a and addressed both the intrinsic and extrinsic apoptotic pathway. Specific blockade of the C5a-receptor 1 (C5aR1) on PMN was evaluated for efficacy to reverse complement-driven alterations. PMN from polytrauma patients exhibited significantly reduced apoptotic rates up to 10 days post trauma compared to healthy controls. Polytrauma-induced resistance was associated with significantly reduced Fas-ligand (FasL) and Fas-receptor (FasR) on PMN and in contrast, significantly enhanced FasL and FasR in serum. Simulation of systemic complement activation revealed for C5a, but not for C3a, a dose-dependent abrogation of PMN apoptosis in both intrinsic and extrinsic pathways. Furthermore, specific blockade of the C5aR1 reversed C5a-induced PMN resistance to apoptosis. The data suggest an important regulatory and putative mechanistic and therapeutic role of the C5a/C5aR1 interaction on PMN apoptosis after polytrauma.

Complement C5a Induces Pro-inflammatory Microvesicle Shedding in Severely Injured Patients.

Initially underestimated as platelet dust, extracellular vesicles are continuously gaining interest in the field of inflammation. Various studies addressing inflammatory diseases have shown that microvesicles (MVs) originating from different cell types are systemic transport vehicles carrying distinct cargoes to modulate immune responses. In this study, we focused on the clinical setting of multiple trauma, which is characterized by activation and dysfunction of both, the fluid-phase and the cellular component of innate immunity. Given the sensitivity of neutrophils for the complement anaphylatoxin C5a, we hypothesized that increased C5a production induces alterations in MV shedding of neutrophils resulting in neutrophil dysfunction that fuels posttraumatic inflammation. In a mono-centered prospective clinical study with polytraumatized patients, we found significantly increased granulocyte-derived MVs containing the C5a receptor (C5aR1, CD88) on their surface. This finding was accompanied by a concomitant loss of C5aR1 on granulocytes indicative of an impaired cellular chemotactic and pro-inflammatory neutrophil functions. Furthermore, in vitro exposure of human neutrophils (from healthy volunteers) to C5a significantly increased MV shedding and C5aR1 loss on neutrophils, which could be blocked using the C5aR1 antagonist PMX53. Mechanistic analyses revealed that the interaction between C5aR1 signaling and the small GTPase Arf6 acts as a molecular switch for MV shedding. When neutrophil derived, C5a-induced MV were exposed to a complex ex vivo whole blood model significant pro-inflammatory properties (NADPH activity, ROS and MPO generation) of the MVs became evident. C5a-induced MVs activated resting neutrophils and significantly induced IL-6 secretion. These data suggest a novel role of the C5a-C5aR1 axis: C5a-induced MV shedding from neutrophils results in decreased C5aR1 surface expression on the one hand, on the other hand it leads to profound inflammatory signals which likely are both key drivers of the neutrophil dysfunction which is regularly observed in patients suffering from multiple traumatic injuries.

Functional immune monitoring in severely injured patients-A pilot study.

After severe trauma, the resulting excessive inflammatory response is countered by compensatory anti-inflammatory mechanisms. The systemic inflammatory response to trauma enhanced by inappropriately timed surgical second hits may be detrimental for the patient. On the other hand, overwhelming anti-inflammatory mechanisms may put patients at increased risk from secondary local and systemic infections. The ensuing sepsis and organ dysfunction due to immune dysregulation remain the leading causes of death after injury. To date, there are no clinically applicable techniques to monitor the pro-/anti-inflammatory immune status of the patients and the remaining ability to react to microbial stimuli. Therefore, in the present study, we used a highly standardized and easy-to-use system to draw peripheral whole blood from polytraumatized patients (ISS ≥ 32, n = 7) and to challenge it with bacterial lipopolysaccharide. Secreted cytokines were compared with those in samples from healthy volunteers. We observed a significant decrease in the release of monocyte-derived mediators. Surprisingly, we detected stable or even increased concentrations of cytokines related to T cell maturation and function. For clinical practicability, we reduced the incubation time before supernatants were collected. Even after an abbreviated stimulation period, a stable release of almost all analysed parameters in patient blood could be detected. In conclusion, the data are indicative of a clinically well-applicable approach to monitor the immune status in severely injured patients in a short time. This may be used to optimize the timing of necessary surgical interventions to avoid a boost of proinflammation and reduce risk of secondary infections.

Deep hypothermic circulatory arrest or tepid regional cerebral perfusion: impact on haemodynamics and myocardial integrity in a randomized experimental trial.

OBJECTIVES: Organ protective management during aortic arch surgery comprises deep hypothermic (18°C) circulatory arrest (DHCA), or moderate hypothermia (28°C/ 'tepid') with regional cerebral perfusion (TRCP). The aim of this experimental study was to evaluate the effect of distinct organ protective management on hemodynamic performance and myocardial integrity. METHODS: Ten male piglets were randomized to group DHCA (n = 5) or TRCP (n = 5) group and operated on cardiopulmonary bypass (CPB) with 60 min of aortic cross-clamping. Blood gas analysis was performed throughout the experiment. Haemodynamic assessment was performed using a thermodilution technique before and after CPB. Myocardial biopsies were taken 2 h after CPB and evaluated using terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick-end labelling assay and western blot analysis. RESULTS: At reperfusion, levels of central venous saturation were significantly higher (P = 0.016) and levels of lactate significantly lower (P = 0.029) in the DHCA group. After CPB, thermodilution measurements revealed higher stroke volume and lower peripheral resistance in the TRCP group (P = 0.012 and 0.037). At the end of the experiment, no significant differences regarding laboratory and haemodynamic parameters were evident. All specimens showed enrichment of terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick-end labelling-positive cells exclusively at the left ventricular subendocardium with no difference between groups and equal concentrations of cyclo-oxygenase-2. CONCLUSIONS: TRCP is associated with decreased peripheral resistance and higher stroke volume immediately after CPB. However, this beneficial effect is contrasted by signs of lower body hypoperfusion, which is expressed by lower central venous saturations and higher lactate levels. Distinct strategies of organ protection did not seem to affect apoptotic/necrotic and inflammatory changes in the left ventricular myocardium.

Repair of Cerebrospinal Fluid Leakage Using a Transfrontal, Radial Adipofascial Flap: An Individual Approach Supported by Three-Dimensional Printing for Surgical Planning.

BACKGROUND: Leakage of cerebrospinal fluid (CSF) because of large prolactinomas represents a complex issue. Because of limited anatomic space, multiple leakage sites, and scarce locally available tissue for repair, surgical possibilities are limited. We report an initial case of using a radial fasciocutaneous flap applied subfrontally to cover a large skull base defect, supported by preoperative three-dimensional (3D) printing for surgical planning. CASE DESCRIPTION: A 29-year-old woman developed severe nuchal pain that was caused by destruction of large parts of her skull base by a prolactinoma. After occipitocervical fusion, medical treatment showed good tumor response but led to CSF leakage after 12 months. An endoscopic approach and ventriculoperitoneal shunt implantation failed to stop the leakage. A 3D model of the skull improved the understanding of the expanded osseous destruction and multiple CSF leakage sites and supported surgical planning. For an extensive coverage of the former clivus and sella region, an intracranially applied radial flap was planned. Dopamine-agonist medication was increased before the operation. Intraoperatively, the flap was brought into position subchiasmatically and wound around the pituitary stalk. CSF leakage was not observed on follow-up (10 weeks). Long-term follow-up will determine the effectiveness of this method.

Stereotactic biopsy in elderly patients: risk assessment and impact on treatment decision.

To evaluate risk profile, diagnostic yield and impact on treatment decision of stereotactic biopsy (SB) in elderly patients with unclear cerebral lesions. In this single center retrospective analysis we identified all patients aged ≥70 years receiving SB between January 2005 and December 2015. Demographic data, Karnofsky Performance Status (KPS), histology, comorbidity (by CHA2DS2-VASc Score) and use of anticoagulation were retrieved. We scrutinized diagnostic yield, procedural complications (mortality, transient and permanent morbidity), hospitalization time and therapeutic consequence. For correlation analysis Chi-Square, Mann-Whitney rank sum test and binary regression were used. Two hundred and thirty patients were included. In 229 patients SB was technically successful. Median age was 74 (70-87) years, 56.1% of patients were male and median preoperative KPS was 80% (30-100). Median CHA2DS2-VASc Score was 4 (1-9), with 29.6% receiving anticoagulation. Median hospital stay was 8 (2-29) days. Pathological diagnosis was conclusive in 97% revealing neoplastic lesions in 91.7% (high-grade glioma 62.6%, lymphoma 18.3%, metastasis 4.8%, low-grade glioma 3.0% and other tumors 3.0%) and non-neoplastic lesions in 5.3% of cases. Procedure-related mortality was 0.4%, transient and permanent morbidity occurred in 19 patients (8.3%) and eight patients (3.5%). Complication rate was not associated with any of the above-mentioned parameters. Adjuvant therapy was initiated in 171 (74.3%) patients. Decision against disease-specific therapy was only influenced by preoperative KPS (p < 0.001). SB in elderly patients is characterized by a favorable risk profile and high diagnostic yield, allowing tissue based therapeutic consequences even in patients with high comorbidity and anticoagulant medication.

Impact of treatment on survival of patients with secondary glioblastoma.

Data concerning treatment of secondary glioblastoma evolving from previously treated WHO II or III grade tumors are very scarce. The aim of this study was to evaluate the impact of surgical resection and adjuvant treatment on survival in patients with secondary glioblastoma. Thirty-nine patients with secondary glioblastoma evolving from previously treated lower grade gliomas between 2004 and 2015 were included. We evaluated the extent of resection, pathological parameters, adjuvant treatment, as well as survival after malignant transformation. The primary tumor grade was WHO II in 16 (41.0%) and WHO III in 23 (59.0%) patients. Median age was 43 years (range 23-67). Median KPS was 80 (range 60-100) before surgery, and 70 (range 50-100) after surgery. Gross total resection (GTR) of contrast-enhancing disease was achieved in 19 (48.7%) patients. Adjuvant treatment was radio-chemotherapy in 23 (59.0%), radiotherapy in three (7.7%), chemotherapy in five (12.8%) and none in eight (20.5%) patients. Median survival was 11 months (range 1-35) in the entire group. Time since initial diagnosis and previous treatment did not correlate with survival after glioblastoma. Failed GTR, poor KPS after surgery, and no adjuvant treatment were prognostic factors for shorter survival in univariate analysis (p < 0.0001, p = 0.028 and p = 0.003). In selected patients, complete resection and adjuvant treatment may prolong survival in spite of multiple previous therapies.

Myocardial Protection During Aortic Arch Repair in a Piglet Model: Beating Heart Technique Compared With Crystalloid Cardioplegia.

BACKGROUND: Selective myocardial perfusion enables repair of congenital aortic arch obstruction without cardiac arrest. This study was inspired by the lack of prospective controlled studies of the beating heart (BH) technique compared with cold crystalloid cardioplegia (CC) regarding effects on myocardial performance, ischemic damage, and serum electrolyte levels. METHODS: In a prospective study, 20 male piglets weighing 11.1 ± 1.3 kg were operated on using cardiopulmonary bypass (CPB) and underwent 60 minutes of aortic cross-clamping. According to prospective randomization, myocardial protection included either a BH modification with selective myocardial perfusion using an individual roller pump or CC. Hemodynamic performance was evaluated by a conductance catheter technique before and after CPB and calculated in relation to baseline levels. Laboratory analysis included blood levels of troponin T and serum electrolytes. RESULTS: Eighteen piglets entered analysis. There were significantly higher slopes of end-systolic pressure-volume relations (168% ± 92% versus 89% ± 16%; p = 0.046) and preload-recruitable stroke work (PRSW) (139% ± 37% versus 103% ± 31%; p = 0.040) in BH piglets compared with those who underwent CC. Laboratory analysis during reperfusion revealed higher levels of troponin T (1.31 ± 0.28 ng/mL versus 0.49 ± 0.17 ng/mL; p < 0.01) and sodium (131 ± 4 mmol/L versus 120 ± 8 mmol/L; p = 0.003) and lower levels of potassium (4.8 ± 0.4 mmol/L versus 6.4 ± 1.0 mmol/L; p = 0.001) with BH compared with CC, whereas no significant differences were calculated at the end of the experiment. CONCLUSIONS: The BH technique is associated with improved contractility compared with standard CC. There is comparable ischemic damage in both groups, with an earlier rise in blood levels of troponin T after BH and more fluctuation of serum electrolytes with CC. Evidence of ischemic changes should dissuade one from using the BH technique imprudently.

Improved contractility with tepid modified full blood cardioplegia compared with cold crystalloid cardioplegia in a piglet model.

OBJECTIVES: Experience regarding warm blood cardioplegia according to Calafiore results from its broad use in adult patients. In this experimental study, tepid (28°C) modified full blood cardioplegia (MBC) was adopted for paediatric use and compared with cold crystalloid cardioplegia (CCC). METHODS: Twenty male piglets (mean weight: 11.1 ± 1.0 kg) were operated on cardiopulmonary bypass (CPB) in moderate hypothermia (28°C) and randomized to MBC (n = 8) or CCC (n = 12) for 60 min aortic cross-clamping. Blood levels of myocardial proteins [N-terminal pro-brain natriuretic peptide (NT-pro-BNP), myoglobin, creatine kinase type MB and troponin-I] were investigated at the beginning of the experiment and after CPB. Haemodynamic measurements included thermodilution and conductance-catheter technique inserted through the left ventricle-apex. Pressure-volume loop analysis was performed with dobutamine-stress test and inflow occlusion, enabling preload independent evaluation of myocardial performance. Changes of measured data post-CPB were calculated in relation to baseline-levels (%). RESULTS: Baseline and operative data in both groups were similar. During the experiment, cardiac markers showed no significant variations between groups. Pressure-volume loop analysis during stress test revealed a significantly higher preload independent contractility (slope of end-systolic pressure-volume relation: Ees) with MBC compared with CCC (MBC: 123 ± 35% [confidence interval (CI95): 93-153] vs CCC: 78 ± 34% [CI95: 54-102]; P = 0.042), whereas cardiac output was not significantly different between groups {MBC: 122 ± 16% [95% confidence interval (CI95): 109-135] vs CCC: 105 ± 17% [CI95: 93-116]; P = 0.069}. CONCLUSION: This randomized animal study proves feasibility and safety of MBC for paediatric use. Haemodynamic evaluation and cardiac markers did not show inferiority to standard CCC. Moreover, MBC seems to be associated with superior contractility post bypass, which encourages us to use MBC in paediatric patients in the near future.