Mechanical cues and substrate interaction affect the manner in which cells adhere, spread, migrate and form tissues. With increased interest in tissue-on-a-chip and co-culture systems utilizing porous membranes, it is important to understand the role of disrupted surfaces on cellular behavior. Using a transparent glass membrane with defined pore geometries, we investigated endothelial fibronectin fibrillogenesis and formation of focal adhesions as well as development of intercellular junctions. Cells formed fewer focal adhesions and had shorter fibronectin fibrils on porous membranes compared to non-porous controls, which was similar to cell behavior on continuous soft substrates with Young’s moduli seven orders of magnitude lower than glass. Additionally, porous membranes promoted enhanced cell-cell interactions as evidenced by earlier formation of tight junctions. These findings suggest that porous membranes with discontinuous surfaces promote reduced cell-matrix interactions similarly to soft substrates and may enhance tissue and barrier formation.

Typical in vitro barrier and co-culture models rely upon thick semi-permeable polymeric membranes that physically separate two compartments. Polymeric track-etched membranes, while permeable to small molecules, are far from physiological with respect to physical interactions with co-cultured cells and are not compatible with high-resolution imaging due to light scattering and autofluorescence. Here we report on an optically transparent ultrathin membrane with porosity exceeding 20%. We optimize deposition and annealing conditions to create a tensile and robust porous silicon dioxide membrane that is comparable in thickness to the vascular basement membrane (100–300 nm). We demonstrate that human umbilical vein endothelial cells (HUVECs) spread and proliferate on these membranes similarly to control substrates. Additionally, HUVECs are able to transfer cytoplasmic cargo to adipose-derived stem cells when they are co-cultured on opposite sides of the membrane, demonstrating its thickness supports physiologically relevant cellular interactions. Lastly, we confirm that these porous glass membranes are compatible with lift-off processes yielding membrane sheets with an active area of many square centimeters. We believe that these membranes will enable new in vitro barrier and co-culture models while offering dramatically improved visualization compared to conventional alternatives.

Porous substrates have gained increased usage in cell studies and tissue mimetic applications because they can partition distinct cell types while still allowing important biochemical crosstalk. In the presented work, we investigated how porous substrates with micron and submicron features influence early cell migration and the associated ECM establishment, which can critically affect the rate of cell coverage on the substrate and the ensuing tissue organization. We showed through time-lapse microscopy that cell speed and migratory distance on membranes with 0.5 μm pores were nearly two-fold of those observed on nonporous membranes, while values on membranes with 3.0 μm pores fell in between. Although the cell directionality ratio and the persistence time was unaffected by the presence of pores, the cells did exhibit directionality preferences based on the hexagonal pore patterning. Fibronectin fibrillogenesis exhibited a distinct inverse relationship to cell speed, as the fibrils formed on the nonporous control were significantly longer than those on both types of porous substrates. We further confirmed on a per cell basis that there is a negative correlation between fibronectin fibril length and cell speed. The observed trade-off between early cell coverage and ECM establishment thus warrants consideration in the selection or the engineering of the ideal porous substrate for tissue mimetic applications and may help guide future cell studies.

IntroductionFor patients undergoing mechanical thrombectomy, numerous (>3) thrombectomy passes may be harmful. However, non-recanalization leads to poor outcomes. For patients requiring multiple thrombectomy passes to achieve reperfusion, it remains unclear if the risk/benefit ratio favors recanalization.ObjectiveTo test the hypothesis that the benefits afforded by successful reperfusion outweigh the risk conveyed by the numerous passes required.MethodsWe retrospectively reviewed prospectively collected data for patients presenting to a comprehensive stroke center with anterior circulation large vessel occlusion (ACLVO) and undergoing thrombectomy requiring more than one pass over 24 months. We stratified patients into three groups: group 1 (successful reperfusion in 2–3 passes), group 2 (successful reperfusion in ≥4 passes), and group 3 (unsuccessful reperfusion).Results250 patients with ACLVO constituted the study cohort. Despite similar demographics, group 2 patients had better clinical outcomes than those in group 3 at 24 hours (National Institutes of Health Stroke Scale (NIHSS) score 13.5 vs 19.1, p<0.001) and at 90 days (modified Rankin Scale score 0–2 rates of 31.1% vs 0.0%, p=0.006) On multivariate logistic regression analysis, age (p=0.034), Alberta Stroke Program Early CT Score (p<0.01), NIHSS score (p=0.02), and parenchymal hematoma type 2 (p=0.015) were significant predictors of functional independence among those who achieved successful reperfusion, but the number of passes required did not predict outcome for these patients (p=0.74).ConclusionPatients who achieve successful reperfusion after many passes have better clinical outcomes than those who do not, despite the number of passes and procedural time required. The number of passes required to achieve successful reperfusion beyond the first pass is not a predictor of functional independence.

No abstract