Alba Vieites-Prado

In vitro and in vivo ocular safety and eye surface permanence determination by direct and Magnetic Resonance Imaging of ion-sensitive hydrogels based on gellan gum and kappa-carrageenan

Gellan gum, kappa-carrageenan and alginates are natural polysaccharides able to interact with different cations that can be used to elaborate ion-activated in situ gelling systems for different uses. The interaction between fluid solutions of these polysaccharides and cations presents into the tear made these biopolymers very interesting to elaborate ophthalmic drug delivery systems. The main purpose of this study is to evaluate the ability of mixtures of these polymers to obtain ion-activated ophthalmic in situ gelling systems with optimal properties for ocular use. To achieve this purpose different proportion of the biopolymers were analyzed using a mixture experimental design evaluating their transparency, mechanical properties and bioadhesion in the absence and presence of simulated tear fluid. Tear induces a rapid sol-to-gel phase transition in the mixtures forming a consistent hydrogel. The solution composed by 80% of gellan gum and 20% kappa-carrageenan showed the best mechanical and mucoadhesive properties. This mixture was evaluated for rheological behavior, microstructure, cytotoxicity, acute corneal irritancy, ex-vivo and in vivo ocular toxicity and in vivo corneal contact time using Magnetic Resonance Images (MRI) techniques. Result indicates that the system is safe at ophthalmic level and produces an extensive ocular permanence higher than 6h.

Intraarterial route increases the risk of cerebral lesions after mesenchymal cell administration in animal model of ischemia

Mesenchymal stem cells (MSCs) are a promising clinical therapy for ischemic stroke. However, critical parameters, such as the most effective administration route, remain unclear. Intravenous (i.v.) and intraarterial (i.a.) delivery routes have yielded varied outcomes across studies, potentially due to the unknown MSCs distribution. We investigated whether MSCs reached the brain following i.a. or i.v. administration after transient cerebral ischemia in rats, and evaluated the therapeutic effects of both routes. MSCs were labeled with dextran-coated superparamagnetic nanoparticles for magnetic resonance imaging (MRI) cell tracking, transmission electron microscopy and immunohistological analysis. MSCs were found in the brain following i.a. but not i.v. administration. However, the i.a. route increased the risk of cerebral lesions and did not improve functional recovery. The i.v. delivery is safe but MCS do not reach the brain tissue, implying that treatment benefits observed for this route are not attributable to brain MCS engrafting after stroke.

Blood glutamate grabbing does not reduce the hematoma in an intracerebral hemorrhage model but it is a safe excitotoxic treatment modality

Recent studies have shown that blood glutamate grabbing is an effective strategy to reduce the excitotoxic effect of extracellular glutamate released during ischemic brain injury. The purpose of the study was to investigate the effect of two of the most efficient blood glutamate grabbers (oxaloacetate and recombinant glutamate oxaloacetate transaminase 1: rGOT1) in a rat model of intracerebral hemorrhage (ICH). Intracerebral hemorrhage was produced by injecting collagenase into the basal ganglia. Three treatment groups were developed: a control group treated with saline, a group treated with oxaloacetate, and a final group treated with human rGOT1. Treatments were given 1 hour after hemorrhage. Hematoma volume (analyzed by magnetic resonance imaging (MRI)), neurologic deficit, and blood glutamate and GOT levels were quantified over a period of 14 days after surgery. The results observed showed that the treatments used induced a significant reduction of blood glutamate levels; however, they did not reduce the hematoma, nor did they improve the neurologic deficit. In the present experimental study, we have shown that this novel therapeutic strategy is not effective in case of ICH pathology. More importantly, these findings suggest that blood glutamate grabbers are a safe treatment modality that can be given in cases of suspected ischemic stroke without previous neuroimaging.

Hyperthermia in human ischemic and hemorrhagic stroke: similar outcome, different mechanisms.

Hyperthermia is a predictor of poor outcome in ischemic (IS) and intracerebral hemorrhagic (ICH) stroke. Our aim was to study the plausible mechanisms involved in the poor outcome associated to hyperthermia in stroke. We conducted a case-control study including patients with IS (n = 100) and ICH (n = 100) within the first 12 hours from symptom onset. Specifically, IS and ICH patients were consecutively included into 2 subgroups, according to the highest body temperature within the first 24 hours: Tmax <37.5°C and Tmax ≥37.5°C, up to reach 50 patients per subgroup of temperature for both IS and ICH patients. Body temperature was determined at admission and every 4 hours during the first 48 hours. Main outcome variable was poor functional outcome (modified Rankin scale score >2) at 3 months. Serum levels of glutamate and active MMP-9 were measured at admission. Our results showed that Tmax ≥37.5°C within the first 24 hours was independently associated with poor outcome in both IS (OR, 12.43; 95% CI, 3.73–41.48; p<0.0001) and ICH (OR, 4.29; 95% CI, 1.32–13.91; p = 0.015) after adjusting for variables with a proven biological relevance for outcome. However, when molecular markers levels were included in the logistic regression model, we observed that glutamate (OR, 1.01; 95% CI, 1.00–1.02; p = 0.001) and infarct volume (OR, 1.06; 95% CI, 1.01–1.10; p = 0.015) were the only variables independently associated to poor outcome in IS, and active MMP-9 (OR, 1.04; 95% CI, 1.00–1.08; p = 0.002) and National Institute of Health Stroke Scale (NIHSS) at admission (OR, 1.29; 95% CI, 1.13–1.49; p<0.0001) in ICH. In conclusion, these results suggest that although the outcome associated to hyperthermia is similar in human IS and ICH, the underlying mechanisms may be different.

Positron Emission Tomography for the Development and Characterization of Corneal Permanence of Ophthalmic Pharmaceutical Formulations.

Purpose This work is aimed at describing the utility of positron emission tomography/computed tomography (PET/CT) as a noninvasive tool for pharmacokinetic studies of biopermanence of topical ocular formulations. Methods The corneal biopermanence of a topical ophthalmic formulation containing gellan gum and kappa carragenan (0.82% wt/vol) labeled with 18Fluorine (18F) radiotracers (18F-FDG and 18F-NaF) was evaluated by using a dedicated small-animal PET/CT, and compared with the biopermanence of an aqueous solution labeled with the same compounds. Regions of interest (ROIs) were manually drawn on the reconstructed PET images for quantifying the radioactivity concentration in the eye. The biopermanence of the formulations was determined by measuring the radioactivity concentration at different times after topical application. Additionally, cellular and ex vivo safety assays were performed to assess the safety of the performed procedures. Results Differences were observed in the ocular pharmacokinetics of the two formulations. After 1.5 hours of contact, 90% of the hydrogel remained in the ocular surface, while only 69% of the control solution remained. Furthermore, it was observed that flickering had a very important role in the approach of the trial. The application of 18F-FDG in the eye was neither irritating nor cytotoxic for human corneal epithelial cells. Conclusions The use of small-animal PET and 18F radiotracers in ocular pharmacokinetics of ophthalmic formulations is feasible and could be a safe method for future ocular pharmacokinetic studies in humans.

Role of adipocytokines in the pathophysiology of migraine. A cross-sectional study.

We are writing with regard to the letter sent by Hyun Jin Min and Kyung Soo Kim (1) in connection with our recently published work in Cephalalgia, ‘‘Role of adipocytokines in the pathophysiology of migraine. A cross-sectional study’’ (2). First of all, we want to thank the authors for the attention and time devoted to the analysis of our work and all their valuable comments. We acknowledge there were two minor transcription mistakes in our paper regarding mean age and prevalence of females in Table 1 and the acronym used for high-sensitivity C-reactive protein in Table 2. In our study, there was no statistically significant difference in levels of leptin and adiponectin between patients suffering migraine with aura and migraine without aura. We did not expect to see a difference between these two groups, considering that we assume the same pathophysiology under both types of migraine and that this difference has not been previously evaluated or reported (3,4). Regarding the effect of BMI and hemodilution on adipocytokine levels, our results are already adjusted by BMI, age and sex by logistic regression analysis. Therefore, the possible effect of hemodilution on adypocitokine levels should be already controlled. Moreover, if hemodilution had an important effect on levels of leptin or adiponectin, levels of both molecules would be inversely correlated with BMI, and not positively correlated in the case of leptin and inversely correlated in the case of adiponectin. Finally, all the studies mentioned by the authors (5–7) refer to tumor markers and obesity; in our study we try to correlate adipocytokine levels with BMI, without comparing values in obese and non-obese subjects. Last, levels of adiponectin in our sample are different in patients and controls, and also different when we analyze the subgroup of chronic migraineurs (CM) and the subgroup of episodic migraineurs (EM). Mean levels of adiponectin are lower in EM patients than in healthy controls, but this difference does not have statistical signification; therefore, we should assume these values as equal. Besides, values in EM in our sample are more scattered than in healthy controls if we look at standard deviation values. This variability suggests that there may be great clinical and physiopathological variations in the group of patients that we categorize as EM compared with the CM group and healthy controls. Previous studies also report very diverse levels of adipocytokine in migraineurs and controls: Bernecker et al. did not find statistically significant differences in ADP levels between migraineurs and controls(8,9); Peterlin et al. (10) analyzed results in EM and CM separately, but did not compare episodic migraineurs and controls; a later study by Duarte et al. (11) compared EM and CM with no significant differences, and more recently Dearborn et al. (9) also found higher levels in migraineurs, including CM and EM, but again did not compare levels in EM and controls. There is also a great variability in the levels of ADP reported in the mentioned studies, probably due to the different ADP multimers and its relation to inflammation, which make the study of this molecule and its role in migraine much more complex (12).

In vivo eye surface residence determination by high-resolution scintigraphy of a novel ion-sensitive hydrogel based on gellan gum and kappa-carrageenan

&NA; In last years, sensitive hydrogels have become a breakthrough in ophthalmic pharmaceutical technology aimed at developing new strategies to increase the residence time of active substances. In a previous paper, we qualitatively demonstrated the capacity of a new ion sensitive hydrogel to increase the residence time. Nevertheless, the clearance of the gel from the ocular surface was not quantifiable with the used methodology. The aim of the present work was to use a well‐established approach based on scintigraphy to quantitatively estimate the residence time of the previously proposed hydrogel. The rat corneal residence time of a topic ophthalmic formulation containing gellan gum and kappa carragenan (0.82% w/v) labeled with 99mTc‐DTPA radiotracer was evaluated and compared with the residence of an aqueous solution. Ophthalmic safety studies such as eye irritation or passage through the cornea were also carried out. After 1.5 h of contact, 77% of the hydrogel remained in the ocular surface, presenting kinetics of disappearance one‐phase decay and a half time of 262 min. We conclude that the novel ophthalmic hydrogel developed with kappa carrageenan and gellan gum remains for long periods of time on the corneal surface, presenting a drop that fits an exponential decay. Graphical abstract Figure. No caption available.

CGRP and PTX3 as Predictors of Efficacy of Onabotulinumtoxin Type A in Chronic Migraine: An Observational Study

The aim of this study is to find a relation between several biomarkers in peripheral blood and outcome after treatment with onabotulinumtoxin A (OnabotA).

CM352 Reduces Brain Damage and Improves Functional Recovery in a Rat Model of Intracerebral Hemorrhage

Background Intracerebral hemorrhage (ICH) is an acute neurological disorder with high mortality and no effective treatment. In addition to the initial bleeding event, rebleeding and hematoma expansion are associated with poor outcome in these patients. We studied the effectiveness of the new antifibrinolytic agent CM352, a short‐half‐life matrix metalloproteinase inhibitor, for achieving early hemostasis and improving functional recovery in a rat model of collagenase‐induced ICH. Methods and Results ICH was induced by striatal injection of collagenase, and 1 hour later, rats received an intravenous injection of saline (n=6) or CM352 (1 mg/kg, n=6). Hematoma (basal and after 3 and 24 hours) and lesion (14 days) volumes were quantified on T2‐weighted (T2) magnetic resonance images. Neurological and functional recovery was evaluated by using Bederson score and a cylinder test (basal, 24 hours, and 14 days). Early treatment (1 hour) with CM352 was efficient reducing hematoma expansion at 3 hours (P<0.01) and, more markedly, at 24 hours (P<0.01). Decreased bleeding after antifibrinolytic treatment was accompanied by reduced interleukin‐6 levels at 3 hours (P<0.05) and smaller lesion volume at 14 days (P<0.01). CM352 drastically reduced sensorimotor impairment (cylinder test) after ICH in rats at 24 hours (P<0.01) and 14 days (P<0.01). Similarly, it also attenuated neurological deficit (Bederson scale) at 24 hours (P<0.01) and 14 days (P<0.01). Interestingly, late (3 hours) CM352 administration also resulted in reduced lesion size and better functional outcome. Conclusions CM352, a new antifibrinolytic agent and matrix metalloproteinase inhibitor, effectively prevented hematoma growth and reduced lesion size in ICH in association with improved functional and neurological recovery.

Vectorized nanodelivery systems for ischemic stroke: a concept and a need

Neurological diseases of diverse aetiologies have significant effects on the quality of life of patients. The limited self-repairing capacity of the brain is considered to be the origin of the irreversible and progressive nature of many neurological diseases. Therefore, neuroprotection is an important goal shared by many clinical neurologists and neuroscientists. In this review, we discuss the main obstacles that have prevented the implementation of experimental neuroprotective strategies in humans and propose alternative avenues for the use of neuroprotection as a feasible therapeutic approach. Special attention is devoted to nanotechnology, which is a new approach for developing highly specific and localized biomedical solutions for the study of the multiple mechanisms involved in stroke. Nanotechnology is contributing to personalized neuroprotection by allowing us to identify mechanisms, determine optimal therapeutic windows, and protect patients from brain damage. In summary, multiple aspects of these new players in biomedicine should be considered in future in vivo and in vitro studies with the aim of improving their applicability to clinical studies.