Control of neonatal human dermal fibroblast migration on poly(lactic‐co‐glycolic acid)‐coated surfaces by electrotaxis

Many types of cells respond to applied direct current electric fields (dcEFs) by directional cell migration, a phenomenon called galvanotaxis or electrotaxis. In this study, electrotaxis was used to control cell migration. We designed a new electrotaxis incubator and chamber system to facilitate long‐term (> 12 h) observation and to allow for alterations to the direction of the current. Poly(lactic‐co‐glycolic acid) (PLGA) was coated onto surfaces to mimic a commonly used tissue‐engineering scaffolding environment. Neonatal human dermal fibroblasts (nHDFs) were grown on PLGA‐coated surfaces and exposed to EFs at increasing currents in the range 0–1 V/cm. These cells migrated toward the cathode during 3 h of dcEF stimulation; however, the migration speed decreased with increasing electric fields. Cells exposed to dcEFs in the range 1–2 V/cm showed no changes to migration speed or x forward migration indices (xFMIs) and the cells continued to move toward the cathode. nHDFs showed directional migration towards the cathode in direct current (dc) EFs (1 V/cm) and they moved in the opposite direction when the polarity of the dcEF was reversed. Reorganization of the actin cytoskeleton and polarization of the Golgi apparatus were evaluated by immunostaining, which showed that the actin cytoskeleton elongated towards the cathode and the Golgi apparatus polarized in the direction of the dcEF. This study revealed that cell migration could potentially be controlled on PLGA scaffolds through electrotaxis. Copyright © 2015 John Wiley & Sons, Ltd.     

Pharmacologic Rate versus Rhythm‐Control Strategies in Atrial Fibrillation: An Updated Comprehensive Review and Meta‐Analysis

Background : In patients with atrial fibrillation (AF), ventricular rate control with medications has been found to be noninferior in preventing clinical events, compared to a strategy converting patients to sinus rhythm and maintaining it with medications. Guidelines have accepted rate control as an acceptable therapeutic option. Most of the prior studies excluded patients without significant left ventricular dysfunction, or permanent AF. Methods : The authors searched the PubMed, CENTRAL, and EMBASE databases for randomized controlled trials from 1966 to 2011. Trials included were direct head‐to‐head comparisons of rate‐ and rhythm‐control strategy using pharmacological means. The primary outcome assessed was risk of all‐cause mortality. We also assessed other pooled clinical endpoints using a random effects model (Mantel‐Haenszel) between rate and rhythm‐control strategies. Results : Ten studies (total N = 7,867) met inclusion/exclusion criteria. In‐hospital mortality was not different between groups (P = 0.31). The rates of stroke, systemic embolism, worsening heart failure, myocardial infarction, and bleeding were also similar. However, rates of rehospitalization were much lower with a rate‐control strategy (P = 0.007). An exploratory analysis in patients younger than 65 years revealed a rhythm‐control strategy was superior to rate control in the prevention of all‐cause mortality (P = 0.0007). Conclusions : This systematic review suggests no difference in clinical outcomes with a rate or rhythm‐control strategy with AF. However, rehospitalization rates appear to be lower with pharmacological rate control for all ages, while finding support for rhythm control in younger patients. (PACE 2013; 36:122–133)     

A Case‐Control Study on Cortical Thickness in Episodic Cluster Headache

Objective.— This study aims at investigating cortical thickness in cluster headache patients as compared with a healthy control group. Background.— The pathobiology of cluster headache is not yet fully understood, although a dysfunction of the hypothalamus has been suggested to be causal. Previous studies in migraine and trigeminal neuropathic pain have demonstrated changes in cortical thickness using cortex segmentation techniques, but no data have been published on cluster headache. Methods.— We investigated 12 men with episodic cluster headache during a phase without acute headache as well as age and sex‐matched healthy controls using high resolution T1‐weighted magnetic resonance imaging acquired at 3T and performed a categorical whole‐brain surface‐based comparison of cortical thickness between groups. Furthermore, a correlation analysis of disease duration and cortical thickness was conducted. Results.— In comparison with control subjects, we found a reduction of cortical thickness in the angular gyrus and the precentral gyrus in cluster headache patients contralaterally to the headache side. These reductions did not correlate with disease duration. The cortical thickness of an area within the primary sensory cortex correlated with disease duration. Conclusions.— This study demonstrates alterations in cortical thickness in cluster headache patients suggesting a potential role of cortical structures in cluster headache pathogenesis. However, it cannot be determined from this study whether the changes are cause or consequence of the disorder. The correlation of cortical thickness with disease duration in the somatosensory cortex may suggest disease‐related plasticity in the somatosensory system.     

Injectable freeze‐dried chitosan‐platelet‐rich‐plasma implants improve marrow‐stimulated cartilage repair in a chronic‐defect rabbit model

Bone‐marrow stimulation (BMS) improves knee‐joint function but elicits incomplete repair. Liquid chitosan (CS)–glycerol phosphate/blood clots have been shown to improve BMS‐based cartilage repair. Platelet‐rich‐plasma (PRP)—a rich source of growth factors and cytokines—improves recruitment and chondrogenic potential of subchondral mesenchymal stem cells. We hypothesised that repair response in a rabbit chronic‐defect model will improve when freeze‐dried CS/PRP is used to augment BMS. Bilateral trochlear defects created in New Zealand white rabbits were allowed to progress to a chronic stage over 4 weeks. Chronic defects were debrided and treated by BMS in second surgery, then augmented with PRP (BMS + PRP) or freeze‐dried CS/PRP implants (BMS + CS/PRP). The quality of 8‐week repair tissue was assessed by macroscopic, histological, and micro computed tomography (Micro‐CT) analysis. ICRS macroscopic scores indicated fibrocartilaginous or fibrous repair in control defects that were improved in the BMS + CS/PRP group. An overall improvement in repair in BMS + CS/PRP group was further confirmed by higher O'Driscoll scores, %Saf‐O and %Coll‐II values. Micro‐CT analysis of subchondral bone indicated ongoing remodelling with repair still underway. Quality and quantity of cartilage repair was improved when freeze‐dried CS/PRP implants were used to augment BMS in a chronic defect model.     

Red blood cell aging markers during storage in citrate‐phosphate‐dextrose–saline‐adenine‐glucose‐mannitol

BACKGROUND: It has been suggested that red blood cell (RBC) senescence is accelerated under blood bank conditions, although neither protein profile of RBC aging nor the impact of additive solutions on it have been studied in detail. STUDY DESIGN AND METHODS: RBCs and vesicles derived from RBCs in both citrate‐phosphate‐dextrose (CPD)–saline‐adenine‐glucose‐mannitol (SAGM) and citrate‐phosphate‐dextrose‐adenine (CPDA) were evaluated for the expression of cell senescence markers (vesiculation, protein aggregation, degradation, activation, oxidation, and topology) through immunoblotting technique and immunofluorescence or immunoelectron microscopy study. RESULTS: A group of cellular stress proteins exhibited storage time– and storage medium–related changes in their membrane association and exocytosis. The extent, the rate, and the expression of protein oxidation, Fas oligomerization, caspase activation, and protein modifications in Band 3, hemoglobin, and immunoglobulin G were less conspicuous and/or exhibited significant time retardation under storage in CPD‐SAGM, compared to the CPDA storage. There was evidence for the localization of activated caspases near to the membrane of both cells and vesicles. CONCLUSIONS: We provide circumstantial evidence for a lower protein oxidative damage in CPD‐SAGM–stored RBCs compared to the CPDA‐stored cells. The different expression patterns of the senescence markers in the RBCs seem to be accordingly related to the oxidative stress management of the cells. We suggest that the storage of RBCs in CPD‐SAGM might be more alike the in vivo RBC aging process, compared to storage in CPDA, since it is characterized by a slower stimulation of the recognition signaling pathways that are already known to trigger the erythrophagocytosis of senescent RBCs.     

Measurement equivalence of touch‐screen computerized and paper‐based diabetes‐specific quality‐of‐life questionnaires

Current advances in technology have enabled the development of a computer‐based questionnaire that provides advantages over the paper‐based mode of administration, such as automatic data entry, storage and calculations. However, before implementing a computer‐based questionnaire, its equivalence with the original paper‐based questionnaire must first be demonstrated. The purpose of this study was to evaluate the measurement equivalence of the computerized Diabetes‐Specific Quality‐of‐Life questionnaire (cD‐QOL) with its original paper‐based counterpart. A two‐period crossover design was used in this study. The measurement equivalence was evaluated using quadratic weighted kappa coefficients, intraclass correlations and Cronbach's alpha comparisons. The cD‐QOL was equivalent to its original paper‐based counterpart. Participants preferred the cD‐QOL over the paper‐based questionnaire and reported that it was easy to use.     

MRI tumor characterization using Gd‐GlyMe‐DOTA‐perfluorooctyl‐mannose‐conjugate (Gadofluorine M™), a protein‐avid contrast agent

The rationale and objectives were to define the MRI tumor‐characterizing potential of a new protein‐avid contrast agent, Gd‐GlyMe‐DOTA‐perfluorooctyl‐mannose‐conjugate (Gadofluorine M?; Schering AG, Berlin, Germany) in a chemically induced tumor model of varying malignancy. Because of the tendency for this agent to form large micelles in water and to bind strongly to hydrophobic sites on proteins, it was hypothesized that patterns of dynamic tumor enhancement could be used to differentiate benign from malignant lesions, to grade the severity of malignancies and to define areas of tumor necrosis. Gadofluorine M, 0.05 mmol Gd kg^?1, was administered intravenously to 28 anesthetized rats that had developed over 10 months mammary tumors of varying degrees of malignancy as a consequence of intraperitoneal administration of N‐ethyl‐N‐nitrosourea (ENU), 45–250 mg kg^?1. These tumors ranged histologically from benign fibroadenomas to highly undifferentiated adenocarcinomas. Dynamic enhancement data were analyzed kinetically using a two‐compartment tumor model to generate estimates of fractional plasma volume (fPV), apparent fractional extracellular volume (fEV*) and an endothelial transfer coefficient (K^PS) for this contrast agent. Tumors were examined microscopically for tumor type, degree of malignancy (Scarff–Bloom–Richardson score) and location of necrosis. Eighteen tumor‐bearing rats were successfully imaged. MRI data showed an immediate strong and gradually increasing tumor enhancement. K^PS and fEV*, but not fPV obtained from tumors correlated significantly (p < 0.05) with the SBR tumor grade, r = 0.65 and 0.56, respectively. Estimates for K^PS and fEV* but not fPV were significantly lower in a group consisting of benign and low‐grade malignant tumors compared with the group of less‐differentiated high‐grade tumors (1.61 ± 0.64 vs 3.37 ± 1.49, p < 0.01; 0.45 ± 0.17 vs 0.78 ± 0.24, p < 0.01; and 0.076 ± 0.048 vs 0.121 ± 0.088, p = 0.24, respectively). It is concluded that the protein‐avid MRI contrast agent Gadofluorine M enhances tumors of varying malignancy depending on the tumor grade, higher contrast agent accumulation for more malignant lesions. The results show potential utility for differentiating benign and low‐grade malignant lesions from high‐grade cancers. Copyright © 2006 John Wiley & Sons, Ltd.     

Protective effects of PF‐4708671 against N‐methyl‐d‐aspartic acid‐induced retinal damage in rats

We previously demonstrated that rapamycin, an inhibitor of the mammalian target of rapamycin (mTOR), protects against N‐methyl‐d ‐aspartic acid (NMDA)‐induced retinal damage in rats. Rapamycin inhibits mTOR activity, thereby preventing the phosphorylation of ribosomal protein S6, which is a downstream target of S6 kinase. Therefore, we aimed to determine whether PF‐4708671, an inhibitor of S6 kinase, protects against NMDA‐induced retinal injury. Intravitreal injection of NMDA (200 nmol/eye) caused cell loss in the ganglion cell layer and neuroinflammatory responses, such as an increase in the number of CD45‐positive leukocytes and Iba1‐positive microglia. Surprisingly, simultaneous injection of PF‐4708671 (50 nmol/eye) with NMDA significantly attenuated these responses without affecting phosphorylated S6 levels. These results suggest that PF‐4708671 and rapamycin likely protect against NMDA‐induced retinal damage via distinct pathways. The neuroprotective effect of PF‐4708671 is unlikely to be associated with inhibition of the S6 kinase, even though PF‐4708671 is reported to be a S6 kinase inhibitor.     

Memantine delayed N‐methyl‐D‐aspartate ‐induced convulsions in neonatal rats

Memantine (1‐amino‐3,5‐dimethyladamantane) is a moderate‐affinity uncompetitive antagonist of N‐methyl‐d‐aspartate (NMDA) receptors. In this study, we have explored the effect of memantine against N‐methyl‐d‐aspartate (NMDA)‐induced seizures in neonatal rats. Here, we evaluated various behavioral seizure abnormalities in neonatal rats (Sprague–Dawley; postnatal day 9) after an intraperitoneal administration of NMDA. Further, we explored whether an acute administration of memantine could protect these neonates against different phases of convulsions induced by NMDA. In a separate study, we have compared the effect of levetiracetam in the same animal model. Exogenous administration of NMDA (30 mg/kg., i.p.) in neonatal rats resulted in arrest of activity, emprosthotonos curvature (trunk is bent forward by the entire muscles), myoclonic jerks, and forelimb/hindlimb clonus. The clonus phase in neonates was followed by loss of righting reflex and continuous seizures (for more than 5 min) suggesting status epilepticus, tonic extension, and death. Pretreatment of memantine hydrochloride (10–30 mg/kg., i.p.) dose‐dependently delayed the onset of different phases of convulsions induced by NMDA. Memantine at the highest dose was found to be ataxic in rat neonates, while lower doses were free of any observed behavioral signs of toxicity. Levetiracetam (25 mg/kg., i.p.) when administered 30 min before the NMDA challenge blocked only the jerk phase and did not affect other phases of NMDA‐induced convulsions. These data indicated that memantine and other safer uncompetitive NMDA receptor antagonists may be protective in the management of neonatal seizures.     

A hydrolytically‐tunable photocrosslinked PLA‐PEG‐PLA/PCL‐PEG‐PCL dual‐component hydrogel that enhances matrix deposition of encapsulated chondrocytes

In this study, a series of photocrosslinked hydrogels were designed composed of both poly(lactide)‐poly(ethylene glycol)‐poly(lactide) (PEL) and poly(ε‐caprolactone)‐poly(ethylene glycol)‐poly(ε‐caprolactone) (PEC) macromers. The PEL/PEC hydrogels at ratios of 100:0, 75:25, 50;50, 25:75 and 0:100 were studied for their degradation characteristics and their ability to support chondrogenesis of encapsulated chondrocytes. Difference in hydrolytic susceptibility between copolymers led to different degradation patterns where higher PEC content correlated with slower degradation. Increased chondrogenic gene expression was observed in chondrocyte‐laden hydrogels within a 4‐week culture period. Biochemical and histological evaluations revealed significant accumulation of extracellular matrix proteins such as glycosaminoglycans and collagen in the 50/50 hydrogel owing to appropriate tuning of hydrogel degradation. These results demonstrate that the dual‐component photocrosslinked hydrogel system is suitable for use as scaffold to support chondrogenesis and, moreover, the tunability of these systems opens up possibilities for use in different cell culturing applications. Copyright © 2014 John Wiley & Sons, Ltd.