Comprehensive analysis of the in vitro and ex ovo hemocompatibility of surface engineered iron oxide nanoparticles for biomedical applications.

A set of biomedically relevant iron oxide nanoparticles with systematically modified polymer surfaces was investigated regarding their interaction with the first contact partners after systemic administration such as blood cells, blood proteins, and the endothelial blood vessels, to establish structure-activity relationships. All nanoparticles were intensively characterized regarding their physicochemical parameters. Cyto- and hemocompatibility tests showed that (1) the properties of the core material itself were not relevant in short-term incubation studies, and (2) toxicities increased with higher polymer mass, neutral = anionic < cationic surface charge and charge density, as well as agglomeration. Based on this, it was possible to classify the nanoparticles in three groups, to establish structure-activity relationships and to predict nanosafety. While the results between cyto- and hemotoxicity tests correlated well for the polymers, data were not fully transferable for the nanoparticles, especially in case of cationic low molar mass polymer coatings. To evaluate the prediction efficacy of the static in vitro models, the results were compared to those obtained in an ex ovo shell-less hen's egg test after microinjection under dynamic flow conditions. While the polymers demonstrated hemotoxicity profiles comparable to the in vitro tests, the size-dependent risks of nanoparticles could be more efficiently simulated in the more complex ex ovo environment, making the shell-less egg model an efficient alternative to animal studies according to the 3R concept.

Rapid formation of plasma protein corona critically affects nanoparticle pathophysiology.

In biological fluids, proteins bind to the surface of nanoparticles to form a coating known as the protein corona, which can critically affect the interaction of the nanoparticles with living systems. As physiological systems are highly dynamic, it is important to obtain a time-resolved knowledge of protein-corona formation, development and biological relevancy. Here we show that label-free snapshot proteomics can be used to obtain quantitative time-resolved profiles of human plasma coronas formed on silica and polystyrene nanoparticles of various size and surface functionalization. Complex time- and nanoparticle-specific coronas, which comprise almost 300 different proteins, were found to form rapidly (<0.5 minutes) and, over time, to change significantly in terms of the amount of bound protein, but not in composition. Rapid corona formation is found to affect haemolysis, thrombocyte activation, nanoparticle uptake and endothelial cell death at an early exposure time.

Recent developments and perspectives on gene therapy using synthetic vectors.

Nonviral vector technology is attracting increasing importance in the biomedical community owing to unique advantages and prospects for the treatment of severe diseases by gene therapy. In this review, synthetic vectors that allow the controlled design of efficient and biocompatible carriers are highlighted. The current benefits, potentials, problems and unmet needs of synthetic gene delivery systems, as well as the strategies to overcome the obstacles are also discussed. Common design principles and structure-activity trends have been established that are important for stable and targeted transport to regions of interest in the body, efficient uptake into cells as well as controlled release of drugs inside the cells, for example, in specialized compartments. The status quo of the use of these systems in preclinical and clinical trials is also considered.

Immunodepression after aneurysmal subarachnoid hemorrhage.

BACKGROUND AND PURPOSE: immunodepression after stroke is associated with complications like high infection rate, but its role in aneurysmal subarachnoid hemorrhage (aSAH) is unclear. This pilot study aimed to assess the presence of immunodepression and its association with infections after aSAH. METHODS: sixteen aSAH patients were enrolled in a prospective study on immune function in a single institution. Detailed immune monitoring (peripheral blood leukocyte subsets, monocyte human leukocyte antigen-DR expression, ex vivo lipopolysaccharide-induced monocytic, Concanavalin A-induced lymphocytic cytokine secretion) was performed until day 10 after aSAH. Occurrence of infection was assessed within 14 days after aSAH. RESULTS: sixteen consecutive aSAH patients (53.1 ± 10.2 years; mean ± SD) met the inclusion criteria, classified as asymptomatic (World Federation of Neurological Surgeons; median, 1; quartile, 1-1; n=7) and symptomatic (median, 4; quartile, 3-5; n=9), all presenting with acute neurological deficits, and 5 of these had additional delayed cerebral ischemia. T-lymphopenia, impaired ex vivo lymphocytic/monocytic cytokine secretion, and decreased monocyte human leukocyte antigen-DR expression occurred over all World Federation of Neurological Surgeons grades but persisted beyond day 3 only in symptomatic patients. Pneumonia (67%; P=0.011) was more frequent in symptomatic patients. Already at day 1, patients with pneumonia showed significantly lower T-cell counts and mitogen-induced interferon-γ production compared to patients without infections. CONCLUSIONS: a pronounced SAH-induced immunodepression was observed early after aSAH but persisted only in symptomatic patients. Immunodepression was associated with a high incidence of pneumonia. Early diagnosis of immunodepression might allow targeted treatment to prevent infectious complications after aSAH.

Relevance of cerebral interleukin-6 after aneurysmal subarachnoid hemorrhage.

BACKGROUND: This study examines the inflammatory response via interleukin-6 (IL-6) in aneurysmal subarachnoid hemorrhage (aSAH) patients and its association with their clinical course (occurrence of acute focal neurological deficits, AFND; and delayed cerebral ischemia, DCI). METHODS: A total of 38 consecutive aSAH patients were studied prospectively within 14 days after admission and classified as asymptomatic (n = 9; WFNS grade 1 (1-2), median and quartiles) and symptomatic (n = 29; WFNS grade 4 (2-5)); the latter presenting with AFND (n = 13), DCI (n = 10) or both (n = 6). Levels of pro-inflammatory cytokine IL-6 were determined in cerebral extracellular fluid (ECF, using cerebral microdialysis), cerebrospinal fluid (CSF) and plasma for 10 days after aSAH. Additionally, C-reactive protein (CRP) levels were measured in plasma. RESULTS: High IL-6 levels in CSF, ECF and plasma were found in all patients, reflecting a pronounced local inflammatory response after aSAH, followed only in symptomatic patients by a delayed systemic inflammation (CRP P < 0.025, days 7-9 after aSAH). In all compartments, IL-6 levels appeared to be higher in symptomatic patients, accompanied also by a higher ECF lactate-pyruvate ratio (P = 0.04). Cerebral, but not plasma IL-6, levels were indicative of the development of DCI in symptomatic patients (ECF P = 0.003; CSF P = 0.001). CONCLUSIONS: A pronounced initial cerebral inflammatory state was observed in patients of all WFNS grades, suggesting that IL-6 elevations are not necessarily detrimental. Cerebral, but not plasma IL-6, levels were predictive of the development of delayed ischemic deficits in symptomatic patients, suggesting that CSF or ECF are the best sampling media for future studies.

High ICP as trigger of proinflammatory IL-6 cytokine activation in aneurysmal subarachnoid hemorrhage.

OBJECTIVE: There is a rising debate about the role of inflammation in the pathogenesis of complications after aneurysmal subarachnoid hemorrhage (SAH) such as intracranial hypertension (intracranial pressure, ICP >20 mmHg). This study aimed to analyse the origin of interleukin-6 (IL-6) in respect to ICP and cerebral metabolism in SAH patients. METHODS: Prospectively, IL-6 was measured in three compartments, the extracellular fluid (ECF) monitored by cerebral microdialysis (MD), cerebrospinal fluid (CSF) and plasma for 10 days after SAH (days 0-4, three times daily; days 5-10, two times daily). Patients were classified having intracranial hypertension (n=7) or normal ICP (n=17) during 10 days after bleeding. Glasgow outcome scale (GOS) was assessed after 3 and 6 months. RESULTS: Patient groups were comparable for age, WFNS and Fisher grade. Intracranial hypertension was associated with an inflammatory response, indicating activation of the inflammatory cascade in the brain (ECF) and systemic circulation with high IL-6 and C-reactive protein (CRP) plasma levels after SAH, the latter associated with unfavourable outcome. The data suggest the ECF but not the CSF as main origin of IL-6 in the systemic circulation in the presence of intracranial hypertension in SAH. DISCUSSION: Intracranial hypertension is associated with a strong activation of the inflammatory cascade in the brain and systemic circulation, and might be underestimated as proinflammmatory trigger in the pathogenesis of complications after SAH. Future therapies targeting anti-inflammatory response in plasma may help to reduce the inflammatory cascade responsible for development of intracranial hypertension.

Inpatient hyperglycemia following aneurysmal subarachnoid hemorrhage: relation to cerebral metabolism and outcome.

INTRODUCTION: Despite its clear association with impaired prognosis, it remains controversial whether hyperglycemia after aneurysmal subarachnoid hemorrhage (SAH) actively contributes to neuronal damage. This study aimed to identify a threshold for blood glucose predicting unfavorable outcome, and to evaluate differences in cerebral metabolism in normo and hyperglycemic SAH patients. METHODS: Prospectively, blood glucose and cerebral metabolism, measured by cerebral microdialysis, were evaluated in 178 patients (WFNS grade I-V; age 51.6 +/- 12.4 years) during days 1-7 after SAH. Patients were classified into groups with mean blood glucose levels 6.1 mmol/l (110 mg/dl) and 7.8 mmol/l (140 mg/dl). Glasgow Outcome Score was assessed after 12 months. RESULTS: Higher inpatient blood glucose was associated with impaired prognosis, with a threshold of 7.5 mmol/l (135 mg/dl) distinguishing best between favorable and unfavorable outcome. Inpatient glucose levels >6.1 mmol/l (110 mg/dl) were associated with higher cerebral lactate and lactate/pyruvate ratio (P < 0.05). Cerebral glucose was elevated only at blood levels >7.8 mmol/l (140 mg/dl). Inpatient glucose levels above 7.8 mmol/l (140 mg/dl) were independent predictors of unfavorable outcome and mortality. CONCLUSION: Blood glucose levels >7.8 mmol/l (140 mg/dl), but not levels >6.1 mmol/l (110 mg/dl), independently predicted unfavorable outcome. While blood glucose levels >6.1 mmol/l (110 mg/dl) were already associated with slight metabolic derangements, cerebral glucose increased only at blood levels >7.8 mmol/l (140 mg/dl). Considering the risks associated with tight glycemic control, a moderate regimen accepting blood glucose levels up to 7.8 mmol/l (140 mg/dl) might be more reasonable after SAH.

Cerebral microdialysis for detection of bacterial meningitis in aneurysmal subarachnoid hemorrhage patients: a cohort study.

INTRODUCTION: Bacterial meningitis (BM) is a severe complication in patients with aneurysmal subarachnoid haemorrhage (SAH). Clinical signs of meningitis are often masked by SAH-related symptoms, and routine cerebrospinal fluid (CSF) analysis fails to indicate BM. Microdialysis (MD) is a technique for monitoring cerebral metabolism in patients with SAH. A cohort study was performed to investigate the value of MD for the diagnosis of BM. METHODS: Retrospectively, 167 patients with SAH in an ongoing investigation on cerebral metabolism monitored by MD were analysed for the presence of BM and related MD changes. Diagnosis of BM was based on microbiological CSF culture or clinical symptoms responding to antibiotic treatment, combined with an increased CSF cell count and/or fever. Levels of MD parameters before and after diagnosis of BM were analysed and compared with the spontaneous course in controls. RESULTS: BM developed in 20 patients, of which 12 underwent MD monitoring at the time of diagnosis. A control group was formed using 147 patients with SAH not developing meningitis. On the day BM was diagnosed, cerebral glucose was lower compared with the value three days before (p = 0.012), and the extent of decrease was significantly higher than in controls (p = 0.044). A decrease in cerebral glucose by 1 mmol/L combined with the presence of fever >or= 38 degrees C indicated BM with a sensitivity of 69% and a specificity of 80%. CSF chemistry failed to indicate BM, but the cell count increased during the days before diagnosis (p < 0.05). CONCLUSIONS: A decrease in MD glucose combined with the presence of fever detected BM with acceptable sensitivity and specificity, while CSF chemistry failed to indicate BM. In patients with SAH where CSF cell count is not available or helpful, MD might serve as an adjunct criterion for early diagnosis of BM.

Is continuous insulin treatment safe in aneurysmal subarachnoid hemorrhage?

OBJECTIVES: To investigate the long-term effect of continuous insulin infusion for glucose control on cerebral metabolism in aneurysmal subarachnoid hemorrhage (SAH) patients. METHODS: Prospective, nonrandomized study of 31 SAH patients in the ICU (52 +/- 10 years, WFNS Grade 2.9 +/- 1.6). A microdialysis catheter was inserted into the vascular territory of the aneurysm. Metabolic changes during 4 days after onset of insulin infusion were analyzed. Blood glucose levels >140 mg/dL after clinical stabilization were treated with intravenous insulin. RESULTS: 24 patients were treated with intravenous insulin. Though no insulin-induced hypoglycemia occurred, cerebral glucose decreased on days 1-4 after insulin onset without reaching critical levels. Glycerol, a marker of membrane degradation, showed a reversible increase on day 1 while the lactate/pyruvate ratio remained stable and glutamate even decreased indicating absence of severe cerebral crisis following insulin infusion and excluding ischemia as a cause for cerebral glucose depletion. CONCLUSIONS: Concerning cerebral metabolism, long-term continuous insulin infusion appears to be safe as long as cerebral glucose levels do not fall below the physiological range. In view of the high incidence of hyperglycemia and need for insulin treatment, future studies on the effect of insulin on cerebral metabolism in SAH patients are desirable.

Hyperglycemia and cerebral glucose in aneurysmal subarachnoid hemorrhage.

OBJECTIVE: To determine whether hyperglycemia exerts deleterious effects via cerebral energy metabolism and to illuminate the effects of cerebral high/low glucose in patients with aneurysmal subarachnoid hemorrhage. DESIGN AND SETTING: Prospective, nonrandomized single-center study over a 2-year period in an intensive care unit at a primary-level university hospital. PATIENTS: 28 subarachnoid hemorrhage patients (age 53 +/- 10 years, WFNS grade 2.8 +/- 1.5) classified as asymptomatic (n = 5) or symptomatic with acute focal or delayed ischemic neurological deficits (n = 23). MEASUREMENTS AND RESULTS: Hyperglycemia (> 7.8 mmol/l; >140 mg/dl) was more frequent in symptomatic patients and was reflected in higher glycerol concentrations than in asymptomatic patients. In all patients a microdialysis catheter was inserted into the tissue at risk; dialysates were collected hourly for 10 days. Cerebral low-glucose episodes (0.6 mmol/l) and high-glucose episodes (>2.6 mmol/l) occurred independently of blood glucose levels. During high-glucose episodes cerebral microdialysate levels were normal, while cerebral low glucose, occurring more frequently in symptomatic patients, was associated with severe cellular distress (increase in lactate/pyruvate ratio, glutamate, glycerol) and with unfavorable outcome if combined with hyperglycemia. CONCLUSIONS: Although hyperglycemia was more frequent in symptomatic patients and associated with high glycerol levels, hyperglycemia was not related to cerebral high glucose. It appears that the association of adverse outcome with acute-phase hyperglycemia is not mediated by cerebral glucose metabolism. Cerebral low glucose was associated with severe metabolic distress and may present a target for therapy to improve clinical outcome.

Insulin-related decrease in cerebral glucose despite normoglycemia in aneurysmal subarachnoid hemorrhage.

INTRODUCTION: Hyperglycaemia following aneurysmal subarachnoid hemorrhage (SAH) is associated with complications and impaired neurological recovery. The aim of this study was to determine the effect of insulin treatment for glucose control on cerebral metabolism in SAH patients. METHODS: This prospective, nonrandomized study was conducted in 31 SAH patients in an intensive care unit (age 52 +/- 10 years, World Federation of Neurological Surgeons grade 2.9 +/- 1.6). A microdialysis catheter was inserted into the vascular territory of the aneurysm after clipping. Blood glucose levels above 140 mg/dl were treated with intravenous insulin and the microdialysates were analyzed hourly for the first 12 hours of infusion. RESULTS: No hypoglycaemia occurred. Twenty-four patients were treated with insulin for glucose control. Higher age and World Federation of Neurological Surgeons score were risk factors for need for insulin treatment (P < 0.05). Although blood glucose remained stable after initiation of insulin infusion, insulin induced a significant decrease in cerebral glucose at 3 hours after onset of the infusion until the end of the observation period (P < 0.05), reflecting high glucose utilization. The lactate:pyruvate ratio and glutamate did not increase, excluding ischaemia as possible cause of the decrease in glucose. Glycerol tended toward higher values at the end of the observation period (9 to 12 hours), reflecting either tissue damage after SAH or the beginning of cellular distress after insulin infusion. CONCLUSION: Higher SAH grade was among the risk factors for need for insulin. Intensive glycaemic control using insulin induced a decrease of cerebral glucose and a slight increase in glycerol, though blood glucose remained normal. Future studies might detect relevant metabolic derangements when insulin treatment starts at low cerebral glucose levels, and may allow us to design a strategy for avoidance of insulin-induced metabolic crisis in SAH patients.

Impact of hyperglycemia on neurological deficits and extracellular glucose levels in aneurysmal subarachnoid hemorrhage patients.

OBJECTIVE: Hyperglycemia after aneurysmal subarachnoid hemorrhage (SAH) is associated with serious complications. Blood glucose may indicate a target for therapy to prevent delayed ischemic neurological deficits (DIND) and improve outcome. The objective of this study was to investigate energy metabolism in the extracellular/cerebrospinal fluid and blood in relation to outcome. METHODS: Prospective non-randomized study was carried out in the intensive care unit (ICU) of university hospital (n = 170 aneurysmal SAH patients, age: 51.0 +/- 12.6 years old). Following approval by the ethics committee, a microdialysis catheter was inserted into the vascular territory of the aneurysm after clipping. Patients were studied for 165 +/- 84 hours and classified according to the presence of neurological symptoms as asymptomatic (n = 66) and symptomatic (n = 104): acute focal neurological deficits (AFND, n = 61) and delayed ischemic neurological deficits (DIND, n = 43). The microdialysates were analysed hourly for energy metabolites. Daily morning blood glucose and cerebrospinal fluid (CSF) levels (glucose and lactate) were determined. Six-month Glasgow outcome scale (GOS) was assessed. RESULTS: Hyperglycemia on admission and high blood glucose levels on the following days were significantly related to the presence of symptoms, most pronounced in patients with poor outcome (p<0.05). In symptomatic patients (high blood glucose), the lowest extracellular fluid (ECF) glucose concentrations were found, most pronounced in the AFND group (1.0 +/- 1.2 mmol/l). The anaerobic metabolites lactate, lactate/pyruvate ratio (LPR) and lactate/glucose ratio (LGR) were higher in symptomatic patients (p<0.001) indicating cerebral metabolic distress. CSF concentrations of glucose and lactate were of no specific value. CONCLUSION: This study confirms the relevance of hyperglycemia to neurological outcome in SAH patients. Cerebral glucose was significantly lower in AFND patients despite hyperglycemic blood levels. More detailed works are necessary to select risk patients for optimized targeted therapy to avoid insulin-induced cerebral metabolic crisis.