Journal: bioRxiv

Article Title: Glycan-coated nanoparticles mimicking the ischemic glycocalyx scavenge the complement system conferring protection after experimental ischemic stroke

doi: 10.64898/2026.03.30.715069

Figure Lengend Snippet: A ) The experimental plan for lectins binding to ihBMECs at the end of hypoxia or after re-oxygenation in absence/presence/. B ) Sensorgrams obtained with Quartz Crystal Microbalance showing the binding of ConA (upper panels) and WGA (lower) injected at four different concentration (0.7-2-6-18 µg/mL) over chip-adherent ihBMECs. The data show decreased binding at the end of the 16h of hypoxia and increased binding after the 4h of re-oxygenation either in presence or absence of MBL compared to normoxic ihBMECs. C ) Microphotographs of MBL (red) deposited on normoxic (left) or hypoxic (right) ihBMEC after re-oxygenation. Nuclei in blue (DAPI), scale bars 10 µm.

Article Snippet: Immortalized human brain microvascular endothelial cells (ihBMECs) (5000 cells/cm 2 ) (Innoprot) were seeded on black 96-well μ-plates (ibidi, Germany) with optically clear flat bottom, suitable for fluorescence microscopy.

Techniques: Binding Assay, Injection, Concentration Assay

Journal: bioRxiv

Article Title: Glycan-coated nanoparticles mimicking the ischemic glycocalyx scavenge the complement system conferring protection after experimental ischemic stroke

doi: 10.64898/2026.03.30.715069

Figure Lengend Snippet: A ) MBL detection on the soft and hard corona samples obtained after preincubation of GNPs with human serum. The MBL signal decreased in the soft corona concomitantly with the three washes (Soft C1-3) and no signal was captured in the fourth wash (Soft C.4). The presence of MBL in the Hard Corona (Hard C., i.e. the proteins remaining after the washings because of their high affinity for the GNPs,) was strong for Man-GNPs (black arrow), and much less for Glc-GNPs (white arrow). B ) The experimental plan for testing sugar-GNPs localization on ihBMEC. C ) 3D microphotographs of Man-GNPs (red, reflectance microscopy) and F-actin (phalloidin, green) in normoxic (CTRL) or hypoxic (HYP) ihBMECs undergone re-oxygenation in the presence of 40 µg/mL Man-GNPs in 30% human serum. Man-GNPs were internalized in the cytoplasm of ihBMECs. Nuclei in blue (DAPI), scale bar 10 µm. D ) Normoxic (CTRL) or hypoxic (HYPOXIA) ihBMECs undergone re-oxygenation in the presence of 5, 20 or 40 µg/mL Man-GNPs in 30% HS were analyzed by reflectance confocal microscopy for Man-GNPs and MBL co-localization. Microphotographs show that Man-GNPs (white, reflectance microscopy) and hMBL (red) did not co-localize (as seen in magnification of white frame, scale bar 1 µm). Phalloidin in green, nuclei in blue (DAPI), scale bar 10 µm.

Article Snippet: Immortalized human brain microvascular endothelial cells (ihBMECs) (5000 cells/cm 2 ) (Innoprot) were seeded on black 96-well μ-plates (ibidi, Germany) with optically clear flat bottom, suitable for fluorescence microscopy.

Techniques: Microscopy, Confocal Microscopy

Journal: bioRxiv

Article Title: Glycan-coated nanoparticles mimicking the ischemic glycocalyx scavenge the complement system conferring protection after experimental ischemic stroke

doi: 10.64898/2026.03.30.715069

Figure Lengend Snippet: A) Microphotographs of MBL (red) deposited on normoxic (CTRL) or hypoxic (HYPOXIA) ihBMECs undergone re-oxygenation in the presence of 5, 20 or 40 µg/mL Man-GNPs in 30% HS (w/Man-GNPs). Nuclei in blue (DAPI), scale bar 200 µm. B ) MBL deposition, measured as fluorescence intensity, was greater on hypoxic than normoxic cells exposed to 30% HS. This increase was significantly reduced when ihBMECs were exposed to 5 and 20 µg/mL of Man-GNPs. Data as mean with individual values ± SD (n= 4). Two-way ANOVA followed by Tukey’s multiple comparisons, **p<0.001, *p<0.05. C ) Overexpression of ICAM-1 in hypoxic ihBMECs was significantly reduced when the cells were exposed to 20 µg/mL of Man-GNPs, to a similar extent than exposure to MBL depleted HS 30% (Δ MBL). D ) Overexpression of MMP-2 in hypoxic ihBMECs was partially counteracted by 20 µg/mL of Man-GNPs. E ) Expression of IL-1α was not significantly changed in presence of Man-GNPs with or without hypoxia. Data from 3 independent experiments, presented as mean with individual values ± SD (n= 4-12). Two-way ANOVA followed by Tukey’s multiple comparisons, ****p<0.0001, ***p<0.001, **p<0.01, *p<0.05.

Article Snippet: Immortalized human brain microvascular endothelial cells (ihBMECs) (5000 cells/cm 2 ) (Innoprot) were seeded on black 96-well μ-plates (ibidi, Germany) with optically clear flat bottom, suitable for fluorescence microscopy.

Techniques: Fluorescence, Over Expression, Expressing

Journal: bioRxiv

Article Title: Glycan-coated nanoparticles mimicking the ischemic glycocalyx scavenge the complement system conferring protection after experimental ischemic stroke

doi: 10.64898/2026.03.30.715069

Figure Lengend Snippet: A) The experimental plan to generate ihBMECs’ normoxic or hypoxic conditioned medium (NORM CM, HYP CM, respectively) and co-cultures of hIPSC-derived neurons, astrocytes and microglia. B ) Microphotographs of GFAP (astrocytes, green), MAP-2 (neurons, red) exposed for 24h to NORM CM or HYP CM +/− Man-GNPs. White arrows point to damaged neurons, i.e. circular cells without dendrites. Nuclei in blue (DAPI), scale bar 10 µm. C ) The quantification of stained volumes (in µm 3 ) showed a decrease of MAP-2 volumes in co-cultures exposed to HYP CM, which was counteracted by Man-GNPs. Data as mean ± SD. Each value is a random field of view (FOV) selected automatically from the overview image. Two-way ANOVA for repeated measures followed by Sidak’s multiple comparisons, ****p<0.0001 (n= 16 FOVs from two experimental replicates, empty rectangles indicate the mean of each replicate). D ) Microphotographs of GFAP (green) and nuclei (DAPI, blue) with a yellow line along which we calculated the FWHM reported in the graph. Width of the first ramification emerging from astrocytic soma was calculated at gray level’s half maximum (HM) and was larger in HYP CM compared to NORM CM or HYP CM + Man-GNPs. Data as mean gray levels of 8 cells per group ± SEM. Two-way ANOVA followed by Tukey’s multiple comparisons, ***p<0.001. Scale bars 10 µm. E ) Microphotographs of GFAP (green), β3-tubulin (neurons, red) and Iba1 (microglia, purple) exposed for 24h to NORM CM or HYP CM +/− Man-GNPs. Dashed squares indicate the magnified views of microglia on the right panels. Nuclei in blue (DAPI), scale bar 100 µm in full images, 20 µm in magnifications. White traces in the magnifications correspond to the Iba1 skeletonized signal. F ) The quantification of microglia morphological parameters showed increased number of branches and junctions after HYP CM exposure, which was counteracted by Man-GNPs. Data as violin plot. Each dot is individual microglia. Kruskal-Wallis test, **p<0.01, ***p<0.001 (n= 25-40 cells from 3 FOVs placed in one well). G ) Histograms of frequency distributions of the morphological parameters in E, shown with automatically chosen bin size.

Article Snippet: Immortalized human brain microvascular endothelial cells (ihBMECs) (5000 cells/cm 2 ) (Innoprot) were seeded on black 96-well μ-plates (ibidi, Germany) with optically clear flat bottom, suitable for fluorescence microscopy.

Techniques: Derivative Assay, Staining

Journal: International Journal of Molecular Sciences

Article Title: Modulation of the Blood–Brain Barrier by Sigma-1R Activation

doi: 10.3390/ijms25105147

Figure Lengend Snippet: PRE-084 increases mitochondrial Ca 2+ in RBMVEC. PRE-084 (5, 10, 20 µM) produced a dose-dependent increase in mitochondrial Ca 2+ (Mito Ca 2+ ) in RBMVEC loaded with the mitochondrial Ca 2+ indicator, Rhod-2 AM. Pretreatment with Sigma-1R antagonists BD 1047 (25 µM) or NE 100 (5 µM) markedly decreased the effect of PRE-084 (10 µM); n = 40–55 cells/treatment group. * p < 0.05; #, not statistically significant.

Article Snippet: Rat brain microvascular endothelial cells (RBMVEC) from Cell Applications, Inc. (San Diego, CA, USA) were cultured in rat brain endothelial basal medium enriched with endothelial growth supplement, according to the manufacturer’s instructions at 37 °C and 5% CO 2 , as previously reported [ , ].

Techniques: Produced

Journal: International Journal of Molecular Sciences

Article Title: Modulation of the Blood–Brain Barrier by Sigma-1R Activation

doi: 10.3390/ijms25105147

Figure Lengend Snippet: PRE-084 increases mitochondrial superoxide and cytosolic reactive oxygen species in RBMVEC. ( A ), PRE-084 (5 µM, 10 µM, and 20 µM) produced a concentration-dependent increase in MitoSOX fluorescence, indicating an increase in mitochondrial superoxide (Mito ROS). Pretreatment with Sigma-1R antagonists, BD 1047 (25 µM) or NE 100 (5 µM) prevented the increase in Mito ROS by PRE-084 (10 µM). ( B ), PRE-084 (5 µM, 10 µM, and 20 µM) produced a concentration-dependent increase in CM-H 2 -DCFDA fluorescence intensity, indicating an increase in cytosolic ROS (Cyto ROS) production. The response to PRE-084 (10 µM) was abolished by pretreatment with BD 1047 (25 µM) or NE 100 (5 µM); n = 43–55 cells/treatment group. * p < 0.05; #, not statistically significant.

Article Snippet: Rat brain microvascular endothelial cells (RBMVEC) from Cell Applications, Inc. (San Diego, CA, USA) were cultured in rat brain endothelial basal medium enriched with endothelial growth supplement, according to the manufacturer’s instructions at 37 °C and 5% CO 2 , as previously reported [ , ].

Techniques: Produced, Concentration Assay, Fluorescence

Journal: International Journal of Molecular Sciences

Article Title: Modulation of the Blood–Brain Barrier by Sigma-1R Activation

doi: 10.3390/ijms25105147

Figure Lengend Snippet: PRE-084 decreases the electrical resistance of the RBMVEC monolayer. ( A ), Examples of decreases in RBMVEC normalized resistance produced by PRE-084 (1–20 µM) determined with ECIS over a period of 150 min (left panel) and comparison of the decrease in amplitude of normalized resistance (right panel). ( B ), Examples of the decrease in normalized resistance produced by PRE-084 (10 µM) in the absence and presence of Sigma-1R antagonists, BD 1047 (25 µM) and NE 100 (5 µM) (left panel), and comparison of the amplitude of decrease in normalized resistance in each condition (right panel); n = 6 experiments/treatment group. * p < 0.05.

Article Snippet: Rat brain microvascular endothelial cells (RBMVEC) from Cell Applications, Inc. (San Diego, CA, USA) were cultured in rat brain endothelial basal medium enriched with endothelial growth supplement, according to the manufacturer’s instructions at 37 °C and 5% CO 2 , as previously reported [ , ].

Techniques: Produced, Comparison

Journal: International Journal of Molecular Sciences

Article Title: Modulation of the Blood–Brain Barrier by Sigma-1R Activation

doi: 10.3390/ijms25105147

Figure Lengend Snippet: PRE-084 produced a disruption in tight and adherens junction proteins and F-actin in RBMVEC. Distribution of tight junction accessory protein ZO-1, adherens junction protein VE-cadherin, and cytoskeleton component F-actin in the control (untreated) RBMVEC (top panels) and RBMVEC treated with PRE-084 (10 µM, 30 min, bottom panels). Nuclei are stained with DAPI. Treatment with PRE-084 produced a disruption in ZO-1 and VE-cadherin, reorganization of F-actin, and the formation of intercellular gaps (arrows). Microscope objective 63X oil.

Article Snippet: Rat brain microvascular endothelial cells (RBMVEC) from Cell Applications, Inc. (San Diego, CA, USA) were cultured in rat brain endothelial basal medium enriched with endothelial growth supplement, according to the manufacturer’s instructions at 37 °C and 5% CO 2 , as previously reported [ , ].

Techniques: Produced, Disruption, Control, Staining, Microscopy

Journal: Cancer research

Article Title: Targeting brain-adaptive cancer stem cells prohibits brain metastatic colonization of triple-negative breast cancer

doi: 10.1158/0008-5472.CAN-17-2994

Figure Lengend Snippet: A. Oncomine database analysis of PCDH7 expression in normal human tissue (Neurogenetics 2006 7:67–80). B. Western Blot analysis of PCDH7 in independent brain metastasis-derived tumorspheres (TS1 and TS2) and corresponding brain-seeking (Br) cell lines. C. Protein expression of PCDH7 in TNBC patient brain metastasis tumorspheres (BM-TS1 and 2) and various cell models. MCF7, SUM159, SKBR3: human breast cancer cell lines; BoM1833: MB231 bone-seeking cell line; LM4175: MB231 lung seeking cell line; NMA: primary normal mouse astrocytes; NHA: normal human astrocytes; HBVEC: human brain microvascular endothelial cells. D. Receiver Operating Characteristic (ROC) curve for PCDH7 expression in primary breast tumor samples of brain metastatic patients using the combined 368 microarray data (MSK-82 and EMC-286 cohort). E. Kaplan–Meier curves showing the brain metastasis-free survival of patients with positive or negative PCDH7 expression in the combined cohort of 368 breast cancer patients, P=1.21×10−5 determined by log rank test. F. Representative PCDH7 immunohistochemistry staining of matched patient tissue sections of brain metastasis, lung metastasis and primary breast tumors. Scale bar: 20µm.

Article Snippet: Normal human astrocytes and human brain microvascular endothelial cells were purchased from Lonza Group Ltd (Allendale, NJ).

Techniques: Expressing, Western Blot, Derivative Assay, Microarray, Immunohistochemistry, Staining