miR-210-3p enriched extracellular vesicles from hypoxic neuroblastoma cells stimulate migration and invasion of target cells.

BACKGROUND: Tumor hypoxia stimulates release of extracellular vesicles (EVs) that facilitate short- and long-range intercellular communication and metastatization. Albeit hypoxia and EVs release are known features of Neuroblastoma (NB), a metastasis-prone childhood malignancy of the sympathetic nervous system, whether hypoxic EVs can facilitate NB dissemination is unclear. METHODS: Here we isolated and characterized EVs from normoxic and hypoxic NB cell culture supernatants and performed microRNA (miRNA) cargo analysis to identify key mediators of EVs biological effects. We then validated if EVs promote pro-metastatic features both in vitro and in an in vivo zebrafish model. RESULTS: EVs from NB cells cultured at different oxygen tensions did not differ for type and abundance of surface markers nor for biophysical properties. However, EVs derived from hypoxic NB cells (hEVs) were more potent than their normoxic counterpart in inducing NB cells migration and colony formation. miR-210-3p was the most abundant miRNA in the cargo of hEVs; mechanistically, overexpression of miR-210-3p in normoxic EVs conferred them pro-metastatic features, whereas miR-210-3p silencing suppressed the metastatic ability of hypoxic EVs both in vitro and in vivo. CONCLUSION: Our data identify a role for hypoxic EVs and their miR-210-3p cargo enrichment in the cellular and microenvironmental changes favoring NB dissemination.

A Porous Gelatin Methacrylate-Based Material for 3D Cell-Laden Constructs.

3D constructs are fundamental in tissue engineering and cancer modeling, generating a demand for tailored materials creating a suitable cell culture microenvironment and amenable to be bioprinted. Gelatin methacrylate (GelMA) is a well-known functionalized natural polymer with good printability and binding motifs allowing cell adhesion; however, its tight micropores induce encapsulated cells to retain a non-physiological spherical shape. To overcome this problem, blended GelMa is here blended with Pluronic F-127 (PLU) to modify the hydrogel internal porosity by inducing the formation of larger mesoscale pores. The change in porosity also leads to increased swelling and a slight decrease in Young's modulus. All blends form stable hydrogels both when cast in annular molds and bioprinted in complex structures. Embedded cells maintain high viability, and while Neuroblastoma cancer cells typically aggregate inside the mesoscale pores, Mesenchymal Stem Cells stretch in all three dimensions, forming cell-cell and cell-ECM interactions. The results of this work prove that the combination of tailored porous materials with bioprinting techniques enables to control both the micro and macro architecture of cell-laden constructs, a fundamental aspect for the development of clinically relevant in vitro constructs.

Extending Thrombolysis in Acute Ischemic Stroke to Primary Care: Early Experiences with a Network-Based Teleneurology Approach.

Background and Purpose-Systemic thrombolysis represents the main proven therapy for acute ischemic stroke, but safe treatment is reported only in well-established stroke units. To extend the use of tissue plasminogen activator (tPA) treatment in primary care hospitals on isolated areas through telemedic was the purpose of specific initiatives in southern Umbria, Italy. Methods-The stroke center of Foligno established a telestroke network to provide consultations for three local hospitals in southern Umbria. The telemedic system consists of a digital network that includes a two-way video conference system and imaging sharing. The main network hospital established specialized stroke wards/teams in which qualified teams treat acute stroke patients. Physicians in these hospitals are able to contact the stroke centers 24 h per day. Quality data are available to support the safe implementation of the stroke procedures. Those available from governmental authorities and local datasets are volume of hospitalization, in-hospital mortality, 30-days mortality, and discharge setting. Objective of the study was to assess the annual hospitalization volume in both the hub and spoke hospitals for ischemic stroke and appraise the performance of the network after the introduction of the telestroke system. Results-A total of 225 systemic thrombolyses were performed in time period indicated above all hospitals. In the main spoke hospital, 41 procedures were performed after teleconsultations were made available. The thrombolysis rate in the hub hospital ranged between 10% in 2016 and 20% in 2019, while in the spoke hospital was below 5% in 2016 and raised to 15% in 2019. The statistically significant difference, in the number of procedures, between hub and spoke in the beginning of the observation time disappeared after introduction of the telestroke network. No increase of the mortality was found. Conclusions-The present data suggest that systemic thrombolysis indicated via stroke experts in the setting of teleconsultation shows similar complication rates to those reported in the National Institute of Neurological Disorders and Stroke trial. Therefore, tPA treatment is also safe in this context and can be extended to primary hospitals.

Motor syndromes.

Motor disturbances alone or associated with other focal deficits are the most common symptoms suggesting a neurovascular event. An appropriate clinical assessment of these signs and symptoms may help physicians to better diagnose and to both better treat and predict outcome. In this paper the main clinical features of motor deficit are described together with other motor-related events such as ataxia and movement disturbances.

Lacunar versus non-lacunar syndromes.

Small-vessel disease is the accepted most frequent cause of lacunar stroke. The main clinical features seen with lacunar infarcts are motor and/or sensitive deficit, ataxic sign, without cortical involvement. A lacunar syndrome is generally the result of a small deep infarct within the territory of a single perforating artery with the maximum diameter on imaging of 15 mm. Recent studies have demonstrated alternative causes of lacunar stroke other than small-vessel disease (e.g. cardio embolism, atherosclerosis or other causes), especially in large lacunae, with a potential relevance on functional outcome. These findings suggest that lacunar stroke is not always a benign disease. Moreover, clinical features may be significant in terms of disability in lacunae in close proximity to crucial anatomical site. The following chapter reports the classical lacunar syndrome and discusses the debated etiology of lacunar stroke.

Acute myocardial infarction and heart failure in acute stroke patients: frequency and influence on clinical outcome.

In patients admitted for acute stroke, the clinical burden of acute myocardial infarction (AMI) and acute heart failure (AHF) is unclear. In these patients we evaluated: (1) the prevalence of AMI and AHF; (2) the effect of AMI and/or AHF on mortality at 3 months; (3) the risk factors for AMI and/or AHF. Consecutive patients admitted for acute stroke were prospectively assessed for the occurrence of AMI and AHF during the hospital stay. Mortality at 3 months was also assessed. Multiple logistic regression analysis was used to identify independent predictors for cardiovascular events and death. Eight hundred fourteen patients were included in the study, 685 of which had an ischemic stroke (84.2%). Fifty-three patients (6.5%) had an AMI and/or an AHF (13 AMI alone, 28 AHF alone and 12 AMI and AHF); all these events occurred in patients with ischemic stroke. At 3 months, 151 patients had died (18.8%). Among patients who had an AMI and/or AHF, 60.4% died as compared with 119 of the 750 patients who did not have these events (15.9%) (p < 0.0001). At logistic regression analysis, AMI and/or AHF were associated with increased mortality at 3 months (p = 0.001). History of angina (p = 0.003), AMI in the 3 months before admission (p < 0.0001), hyperglycemia (p = 0.047), and high NIHSS on admission (p < 0.0001) were associated with in hospital AMI and/or AHF. In acute stroke patients, AMI and AHF are common and associated with increased mortality at 3 months. Whether a timely and more careful management of these complications may improve clinical outcome should be further explored.

Gender differences in patients with acute ischemic stroke.

Stroke has a greater effect on women than men owing to the fact that women have more stroke events and are less likely to recover. Age-specific stroke rates are higher in men; however, because of women's longer life expectancy and the much higher incidence of stroke at older ages, women have more stroke events than men overall. The aims of this prospective study in consecutive patients were to assess whether there are gender differences in stroke risk factors, treatment or outcome. Consecutive patients with ischemic stroke were included in this prospective study at four study centers. Disability was assessed using a modified Rankin Scale score (>or=3 indicating disabling stroke) in both genders at 90 days. Outcomes and risk factors in both genders were compared using the chi(2) test. Multiple logistic regression analysis was used to identify any independent predictors of outcome. A total of 1136 patients were included in this study; of these, 494 (46%) were female. Women were statistically older compared with men: 76.02 (+/- 12.93) and 72.68 (+/- 13.27) median years of age, respectively. At admission, females had higher NIH Stroke Scale scores compared with males (9.4 [+/- 6.94] vs 7.6 [+/- 6.28] for men; p = 0.0018). Furthermore, females tended to have more cardioembolic strokes (153 [30%] vs 147 [23%] for men; p = 0.004). Males had lacunar and atherosclerotic strokes more often (146 [29%] vs 249 [39%] for men; p = 0.002, and 68 [13%] vs 123 [19%] for men; p = 0.01, respectively). The mean modified Rankin Scale score at 3 months was also significantly different between genders, at 2.5 (+/- 2.05) for women and 2.1 (+/- 2.02) for men (p = 0.003). However, at multivariate analysis, female gender was not an indicator for negative outcome. It was concluded that female gender was not an independent factor for negative outcome. In addition, both genders demonstrated different stroke pathophysiologies. These findings should be taken into account when diagnostic workup and treatment are being planned.

Acute hyperglycemia and early hemorrhagic transformation in ischemic stroke.

BACKGROUND: Hyperglycemia has been claimed to be associated with hemorrhagic transformation (HT) in patients with acute ischemic stroke treated with thrombolysis. The aim of this study was to assess whether the admission blood glucose level is related to HT in a prospective study in consecutive patients with acute ischemic stroke. METHODS: Consecutive patients admitted for ischemic stroke to 4 Italian hospitals were included in this prospective cohort study. RESULTS: Among 1,125 consecutive patients included in the analysis, 98 (8.7%) had HT: 62 (5.5%) had hemorrhagic infarction (HI) and 36 (3.2%) parenchymal hematoma (PH). A blood glucose level >110 mg/dl was found in 42.4% of the patients, a level between 110 and 149 mg/dl in 25.2%, and a level >150 mg/dl in 17.2%. At 3 months, 7 patients were lost at follow-up, 326 patients (29.2%) were disabled (modified Rankin score > or = 3) and 129 died (11.5%). PH was associated with an increased risk of death or disability (OR 15.29, 95% CI 2.35-99.35). However, this was not the case for HT overall and HI. At logistic regression analysis, PH was predicted by high levels of admission blood glucose (OR 1.01, 95% CI 1.00-1.01 for 1 added mg/dl). The rate of PH was 2.1% in patients with <110 mg/dl, 3.6% in patients with a level between 110 and 149 mg/dl and 6.4% in patients with a level >150 mg/dl. The curve estimation regression model showed a significant linear increase in the risk of PH related to an increase in blood glucose levels (R(2) = 0.007, p = 0.007). CONCLUSIONS: Hyperglycemia during acute ischemic stroke predisposes to PH, which in turn determines a non-favorable outcome at 3 months. This relationship seems to be linear.

Effect of carotid stenosis on the prognostic value of admission blood pressure in patients with acute ischemic stroke.

BACKGROUND: Data on the correlation between blood pressure (BP) on admission and clinical outcome in patients with acute stroke are conflicting. The aims of the present study in consecutive patients with acute ischemic stroke were to evaluate: (a) the relationship between systolic or diastolic BP on admission and mortality at 3 months; (b) the role of carotid artery disease ipsilateral to the index stroke on this relationship. METHODS: Consecutive patients admitted to four Italian hospitals with objectively diagnosed ischemic stroke were included in this prospective study. RESULTS: A total of 1467 patients (mean age 72.6+/-13.2 years; males 53.3%) with acute ischemic stroke were evaluated. At 3 months, 13 patients were lost at follow-up and 133 had died (9.2%). In patients with systolic BP <140mmHg, mortality was 11% (40/362), in patients with systolic BP between 140 and 179mmHg 8.3% (68/823) and in patients with systolic BP >or=180mmHg 9.2% (25/269). Patients with systolic BP <140mmHg were more likely to die within 90 days (OR 3.4; 95% CI 1.4-8.5, p=0.008, after adjusting for other risk factors) when compared to those with systolic BP between 140 and 179mmHg (reference group with OR=1.0). Systolic BP >or=180mmHg was not associated with increased mortality (OR 0.4; 95% CI 0.1-1.3, p=0.13). Two-hundred and thirty patients out of 1278 with anterior circulation stroke had a stenosis (>or=50% on ultrasonography) or an occlusion of the internal carotid ipsilateral to the index stroke. At 3 months, 29 patients had died (12.7%). In patients with systolic BP <140mmHg, mortality was 20.4% (11/54), in those with systolic BP between 140 and 179mmHg 13.0% (16/123) and in those with systolic BP >or=180mmHg 3.9% (2/51). Patients with systolic BP >or=180mmHg were less likely to die within 90 days (OR 0.1; 95% CI 0.0-0.5, p=0.022 after adjusting for other risk factors) compared to those with systolic BP between 140 and 179mmHg. Systolic BP <140mmHg was not associated with increased mortality (OR 6.3; 95% CI 0.8-48.0, p=0.08). CONCLUSIONS: In the overall population, low BP was an independent risk factor for mortality. In patients with stenosis >/=50% or occlusion of the carotid ipsilateral to the index stroke, high BP was associated with a lower mortality at 3 months.

Non-destructive and non-invasive analyses shed light on the realization technique of ancient polychrome prints.

Five polychrome prints representing famous painters, such as Albrecht Dürer, were analyzed using a non-destructive and non-invasive methodology as required by the artwork typology. The diagnostic strategy includes X-ray fluorescence (XRF), reflectance micro-infrared (microFTIR) and micro-Raman (microRaman) spectroscopy. These prints were realized with a la poupée method that involves application of the polychrome inks on a single copper plate, before the printing process. A broad range of compounds (i.e., cinnabar, red lead, white lead, umber earth, hydrated calcium sulfate, calcium carbonate, amorphous carbon, and Prussian blue) was employed as chalcographic inks, using linseed oil as a binding medium. Gamboge was identified in the delicate finishing brush touches realized in watercolor.

Early hemorrhagic transformation of brain infarction: rate, predictive factors, and influence on clinical outcome: results of a prospective multicenter study.

BACKGROUND AND PURPOSE: Early hemorrhagic transformation (HT) is a complication of ischemic stroke but its effect on patient outcome is unclear. The aims of this study were to assess: (1) the rate of early HT in patients admitted for ischemic stroke, (2) the correlation between early HT and functional outcome at 3 months, and (3) the risk factors for early HT. METHODS: Consecutive patients with ischemic stroke were included in this prospective study in 4 study centers. Early HT was assessed by CT examination performed at day 5+/-2 after stroke onset. Study outcomes were 3-month mortality or disability. Disability was assessed using a modified Rankin score (> or = 3 indicating disabling stroke) by neurologists unaware of the occurrence of HT in the individual cases. Outcomes in patients with and without early HT were compared by chi(2) test. Multiple logistic regression analysis was used to identify predictors for HT. RESULTS: Among 1125 consecutive patients (median age 76.00 years), 98 (8.7%) had HT, 62 (5.5%) had hemorrhagic infarction, and 36 (3.2%) parenchymal hematoma. At 3 months, 455 patients (40.7%) were disabled or died. Death or disability was seen in 33 patients with parenchymal hematoma (91.7%), in 35 patients with hemorrhagic infarction (57.4%) as compared with 387 of the 1021 patients without HT (37.9%). At logistic regression analysis, parenchymal hematoma, but not hemorrhagic infarction, was independently associated with an increased risk for death or disability (OR 15.29; 95% CI 2.35 to 99.35). At logistic regression analysis, parenchymal hematoma was predicted by large lesions (OR 12.20, 95% CI 5.58 to 26.67), stroke attributable to cardioembolism (OR 5.25; 95% CI 2.27 to 12.14) or to other causes (OR 6.77; 95% CI 1.75 to 26.18), high levels of blood glucose (OR 1.01; 95% CI 1.00 to 1.01), and thrombolytic treatment (OR 3.54, 95% CI 1.04 to 11.95). CONCLUSIONS: Early HT occurs in about 9% of patients. Parenchymal hematoma, seen in about 3% of patients, is associated with an adverse outcome. Parenchymal hematoma was predicted by large lesions attributable to cardioembolism or other causes, high blood glucose, and treatment with thrombolysis.

Clinical benefit of early anticoagulation in cardioembolic stroke.

BACKGROUND: Nonvalvular atrial fibrillation is the most common source of cardiac embolism with a high reported risk of stroke and a high stroke-related mortality. A common clinical dilemma in patients with acute stroke is whether the detection of one of the major cardiac sources of emboli requires an early anticoagulation to reduce early stroke recurrence and mortality. METHODS: In this review, we report on the results of clinical trials that have investigated the efficacy of early treatment for acute cardioembolic stroke. RESULTS: Large clinical trials demonstrate that there is no evidence supporting the administration of heparin in patients with acute ischemic stroke within 48 h from stroke onset. CONCLUSIONS: The results of recent studies showing an advantage of the very early administration of heparin (<3 h from stroke onset) should encourage clinicians to perform further trials on the efficacy of an early administration of heparin in acute cardioembolic stroke.

Need for extensive diagnostic work-up for patients with lacunar stroke.

OBJECTIVE: Small-vessel disease is the most frequent cause of lacunar stroke. The aims of this study were to evaluate: 1) alternative causes of lacunar stroke other than small-vessel disease; 2) functional outcome of lacunar strokes due to small vessel disease compared to lacunar strokes due to alternative causes. METHODS: Consecutive patients with first-ever ischemic lacunar stroke were prospectively followed-up for at least 3 months. At discharge patients were divided into 2 groups: lacunar stroke due to small vessel disease (L-SVD) and lacunar stroke due to possible other etiologies (L-non SVD) (e. g. cardioembolism, atherosclerosis or other causes). Main outcome measures were the combined end point of death or disability (mRS >or= 3) and recurrent stroke at the end of follow-up. RESULTS: 535 patients with acute stroke were seen during the study period. Out of these, 196 patients (126 males) with a mean age of 71.6 years (SD = 10) had lacunar stroke. L-SVD was seen in 142 patients (72.4 %) and L-nonSVD in 54 patients (27.6 %). After 16.4 months, 12 patients had died (6.2%; annual mortality rate 4.4 %), 63 were disabled (32.5 %) and 27 had stroke recurrence (13.9%; annual recurrence rate 9.9 %). Forty-nine patients with L-SVD (34.7 %) and 26 with L-nonSVD (49%) had died or become disabled. Sixteen patients with L-SVD (11.3 %) and 11 with L-nonSVD (20.4 %) had stroke recurrence. On multivariate analysis, L-nonSVD (OR = 2.87, 95% CI 1.08-7.65; p = 0.034) and age (OR = 1.07, 95 % CI 1.02-1.12; p = 0.01) were associated to more severe outcome. L-nonSVD was independently associated with recurrence (OR = 5.03, 95% CI 1.54-16.44; p = 0.006). CONCLUSIONS: Twenty-seven percent of patients with lacunar stroke have causes other than small vessel disease. These patients have a severe outcome in terms of recurrence,mortality or disability. These findings support the need for a comprehensive diagnostic work-up for patients with lacunar stroke.

Efficacy and safety of anticoagulant treatment in acute cardioembolic stroke: a meta-analysis of randomized controlled trials.

BACKGROUND AND PURPOSE: The role of anticoagulant treatment for acute cardioembolic stroke is uncertain. We performed an updated meta-analysis of all randomized trials to obtain the best estimates of the efficacy and safety of anticoagulants for the initial treatment of acute cardioembolic stroke. METHODS: Using electronic and manual searches of the literature, we identified randomized trials comparing anticoagulants (unfractionated heparin or low-molecular-weight heparin or heparinoids), started within 48 hours, with other treatments (aspirin or placebo) in patients with acute ischemic cardioembolic stroke. Two reviewers independently selected studies and extracted data on study design, quality, and clinical outcomes, including death or disability, all strokes, recurrent ischemic stroke, and cerebral symptomatic bleeding. Odds ratios for individual outcomes were calculated for each trial and data from all the trials were pooled using the Mantel-Haenszel method. RESULTS: Seven trials, involving 4624 patients with acute cardioembolic stroke, met the criteria for inclusion. Compared with other treatments, anticoagulants were associated with a nonsignificant reduction in recurrent ischemic stroke within 7 to 14 days (3.0% versus 4.9%, odds ratio 0.68, 95% CI: 0.44 to 1.06, P=0.09, number needed to treat=53), a significant increase in symptomatic intracranial bleeding (2.5% versus 0.7%, odds ratio 2.89; 95% CI: 1.19 to 7.01, P=0.02, number needed to harm=55), and a similar rate of death or disability at final follow up (73.5% versus 73.8%, odds ratio 1.01; 95% CI: 0.82 to 1.24, P=0.9). CONCLUSIONS: Our findings indicate that in patients with acute cardioembolic stroke, early anticoagulation is associated with a nonsignificant reduction in recurrence of ischemic stroke, no substantial reduction in death and disability, and an increased intracranial bleeding.

First-ever stroke and outcome in patients admitted to Perugia Stroke Unit: predictors for death, dependency, and recurrence of stroke within the first three months.

Predictors of poor outcome after first-ever stroke within 3 months in consecutive patients admitted to our Stroke Unit were defined. Factors included age, sex, risk factors, occurrence of transient ischemic attacks, extension of cerebral infarction, presumed cause of stroke, clinical findings, and demographic characteristics. Multiple regression models were used to analyze predictors of mortality, dependency and stroke recurrence. A total of 435 patients with first-ever stroke were included. Of these, 358 patients had ischemic stroke and 77 hemorrhagic stroke. Three-month mortality rate was 20.5%. After the same period, 24.6% of survivors were dependent (mRS > or = 3) and 5.0% of patients had recurrent stroke. Age, the presence of atrial fibrillation, impaired consciousness on admission, and stroke severity were related to mortality. The presence of stroke due to an undetermined cause or small vessel disease was associated with lower mortality. Partial anterior circulation syndrome or lacunar syndrome were both related to better outcome. The best predictors for dependency after 3 months were age and stroke severity. The only variable identified as the best predictor for recurrence was the presence of diabetes mellitus.