The biconcave disk shape and deformability of mammalian RBCs rely on the membrane skeleton, a viscoelastic network of short, membrane-associated actin filaments (F-actin) cross-linked by long, ...flexible spectrin tetramers. Nonmuscle myosin II (NMII) motors exert force on diverse F-actin networks to control cell shapes, but a function for NMII contractility in the 2D spectrin–F-actin network of RBCs has not been tested. Here, we show that RBCs contain membrane skeleton-associated NMIIA puncta, identified as bipolar filaments by superresolution fluorescence microscopy. MgATP disrupts NMIIA association with the membrane skeleton, consistent with NMIIA motor domains binding to membrane skeleton F-actin and contributing to membrane mechanical properties. In addition, the phosphorylation of the RBC NMIIA heavy and light chains in vivo indicates active regulation of NMIIA motor activity and filament assembly, while reduced heavy chain phosphorylation of membrane skeleton-associated NMIIA indicates assembly of stable filaments at the membrane. Treatment of RBCs with blebbistatin, an inhibitor of NMII motor activity, decreases the number of NMIIA filaments associated with the membrane and enhances local, nanoscale membrane oscillations, suggesting decreased membrane tension. Blebbistatin-treated RBCs also exhibit elongated shapes, loss of membrane curvature, and enhanced deformability, indicating a role for NMIIA contractility in promoting membrane stiffness and maintaining RBC biconcave disk cell shape. As structures similar to the RBC membrane skeleton exist in many metazoan cell types, these data demonstrate a general function for NMII in controlling specialized membrane morphology and mechanical properties through contractile interactions with short F-actin in spectrin–F-actin networks.
A new in vitro cerebrovascular occlusion model of the intracranial circulation was developed recently for testing thrombectomy devices. Using this model, we compared recanalization success associated ...with different modern endovascular thrombectomy approaches.
Model experiments were performed in 4 thrombectomy test groups: 1) primary or direct Stentriever thrombectomy with a conventional guide catheter (control group), 2) primary Stentriever thrombectomy with a balloon-guide catheter, 3) combined Stentriever-continuous aspiration approach, and 4) direct aspiration alone. Successful recanalization was defined as a TICI score of 2b or 3.
Seventy-one thrombectomy experiments were conducted. Similar rates of TICI 2b-3 scores were achieved with balloon-guide and conventional guide catheters (P = .34). The combined Stentriever plus aspiration approach and the primary aspiration thrombectomy resulted in significantly higher rates of TICI 2b or 3 than the conventional guide-catheter approach in the control group (P = .008 and P = .0001, respectively). The primary Stentriever thrombectomy with the conventional guide catheter showed the highest rate of embolization to new territories (53%).
Data from our in vitro model experiments show that the Stentriever thrombectomy under continuous aspiration and primary aspiration thrombectomy approaches led to the highest degree of recanalization.
Acute, inflammatory conditions associated with dysregulated complement activation are characterized by significant increases in blood concentration of reactive oxygen species (ROS) and ATP. The ...mechanisms by which these molecules arise are not fully understood. In this study, using luminometric- and fluorescence-based methods, we show that ligation of glycophorin A (GPA) on human red blood cells (RBCs) results in a 2.1-fold, NADPH-oxidase-dependent increase in intracellular ROS that, in turn, trigger multiple downstream cascades leading to caspase-3 activation, ATP release, and increased band 3 phosphorylation. Functionally, using 2D microchannels to assess membrane deformability, GPS-ligated RBCs travel 33% slower than control RBCs, and lipid mobility was hindered by 10% using fluorescence recovery after photobleaching (FRAP). These outcomes were preventable by pretreating RBCs with cell-permeable ROS scavenger glutathione monoethyl ester (GSH-ME). Our results obtained in vitro using anti-GPA antibodies were validated using complement-altered RBCs isolated from control and septic patients. Our results suggest that during inflammatory conditions, circulating RBCs significantly contribute to capillary flow dysfunctions, and constitute an important but overlooked source of intravascular ROS and ATP, both critical mediators responsible for endothelial cell activation, microcirculation impairment, platelet activation, as well as long-term dysregulated adaptive and innate immune responses.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Brain CTP is used to estimate infarct and penumbra volumes to determine endovascular treatment eligibility for patients with acute ischemic stroke. We aimed to assess the accuracy of a Bayesian CTP ...algorithm in determining penumbra and final infarct volumes.
Data were retrospectively collected for 105 patients with acute ischemic stroke (55 patients with successful recanalization TICI 2b/2c/3 and large-vessel occlusions and 50 patients without interventions). Final infarct volumes were calculated using DWI and FLAIR 24 hours following CTP imaging. RAPID and the Vitrea Bayesian CTP algorithm (with 3 different settings) predicted infarct and penumbra volumes for comparison with final infarct volumes to assess software performance. Vitrea settings used different combinations of perfusion maps (MTT, TTP, CBV, CBF, delay time) for infarct and penumbra quantification. Patients with and without interventions were included for assessment of predicted infarct and penumbra volumes, respectively.
RAPID and Vitrea default setting had the most accurate final infarct volume prediction in patients with interventions (Spearman correlation coefficient, mean infarct difference default versus FLAIR: 0.77, 4.1 mL, default versus DWI: 0.72, 4.7 mL, RAPID versus FLAIR: 0.75, 7.5 mL, RAPID versus DWI: 0.75, 6.9 mL). Default Vitrea and RAPID were the most and least accurate in determining final infarct volume for patients without an intervention, respectively (default versus FLAIR: 0.76, -0.4 mL, default versus DWI: 0.71, -2.6 mL, RAPID versus FLAIR: 0.68, -49.3 mL, RAPID versus DWI: 0.65, -51.5 mL).
Compared with RAPID, the Vitrea default setting was noninferior for patients with interventions and superior in penumbra estimation for patients without interventions as indicated by mean infarct differences and correlations with final infarct volumes.
Short, uniform-length actin filaments function as structural nodes in the spectrin-actin membrane skeleton to optimize the biomechanical properties of red blood cells (RBCs). Despite the widespread ...assumption that RBC actin filaments are not dynamic (i.e., do not exchange subunits with G-actin in the cytosol), this assumption has never been rigorously tested. Here we show that a subpopulation of human RBC actin filaments is indeed dynamic, based on rhodamine-actin incorporation into filaments in resealed ghosts and fluorescence recovery after photobleaching (FRAP) analysis of actin filament mobility in intact RBCs (~25-30% of total filaments). Cytochalasin-D inhibition of barbed-end exchange reduces rhodamine-actin incorporation and partially attenuates FRAP recovery, indicating functional interaction between actin subunit turnover at the single-filament level and mobility at the membrane-skeleton level. Moreover, perturbation of RBC actin filament assembly/disassembly with latrunculin-A or jasplakinolide induces an approximately twofold increase or ~60% decrease, respectively, in soluble actin, resulting in altered membrane deformability, as determined by alterations in RBC transit time in a microfluidic channel assay, as well as by abnormalities in spontaneous membrane oscillations (flickering). These experiments identify a heretofore-unrecognized but functionally important subpopulation of RBC actin filaments, whose properties and architecture directly control the biomechanical properties of the RBC membrane.
A simple, yet powerful magnetic‐levitation‐based device is reported for real‐time, label‐free separation, as well as high‐resolution monitoring of cell populations based on their unique magnetic and ...density signatures. This method allows a wide variety of cellular processes to be studied, accompanied by transient or permanent changes in cells' fundamental characteristics as a biological material.
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