Journal:

Article Title: Anti-Tumoral Activity of a Short Decapeptide Fragment of the Alzheimer's A? Peptide

doi: 10.1007/s10989-010-9198-8

Figure Lengend Snippet: Effect of 4-hydroxy tamoxifen on angiogenesis in vitro. a Representative pictures depicting the formation of capillary networks in response to a dose range of 4-hydroxy tamoxifen following the plating of human brain microvascular endothelial cells onto a layer of Matrigel. b The graph represents the quantification of capillary network length by image analysis and the amount of LDH activity detected in the culture medium surrounding the capillary like structures following 24 h of treatment with a dose range of 4-hydroxy tamoxifen. ANOVA revealed a significant main effect of 4-hydroxy tamoxifen on LDH released (P < 0.03) but no significant main effect on capillary formation (P = 0.743) and post hoc analysis showed significant differences for LDH released between control conditions and 10 µM of 4-hydroxy tamoxifen (P < 0.009)

Article Snippet: Capillary Morphogenesis Assay Human Brain Microvascular Endothelial Cells (HBMEC) (Sciencell, CA) were cultured in Endothelial Cell Growth Medium (Cell Applications, CA) containing 5% fetal bovine serum, 1% penicillin/streptomycin and 1% Endothelial Cell Growth Supplement (Sigma–Aldrich, MO).

Techniques: In Vitro, Activity Assay, Control

Journal: Frontiers in Microbiology

Article Title: The Mechanism of the Zika Virus Crossing the Placental Barrier and the Blood-Brain Barrier

doi: 10.3389/fmicb.2020.00214

Figure Lengend Snippet: The infectivity and virulence of ZIKV in JEG-3 and hCMEC/D3 cells. (A) The immunofluorescence images of ZIKV E protein revealed the population of ZIKV-infected cells in JEG-3 (MOI of 0.25 to 1) and hCMEC/D3 cells (MOI of 1 to 10) at 24 h post-infection. The ZIKV E protein antibody was recognized by an Alex488-secondary antibody (green). Nuclear DNA was stained with DAPI (blue). Scale bar, 20 μm. (B) The percentage of infected cells was quantified from (A) . (C) JEG-3 cells were infected ZIKV at MOI of 0.5 to 1, and hCMEC/D3 cells were infected ZIKV at MOI of 5 to 10 during 24 h. The virulence of ZIKV in JEG-3 and hCMEC/D3 was measured by a cell viability assay. The error bars represent standard deviations from three independent experiments. Statistical differences were obtained through t -tests. ** p < 0.01; *** p < 0.001.

Article Snippet: The immortalized human brain capillary endothelial cell line hCMEC/D3 (SCC066, Merck) was cultured in the EndoGRO TM -MV Complete Media Kit (Merck).

Techniques: Infection, Immunofluorescence, Staining, Viability Assay

Journal: Frontiers in Microbiology

Article Title: The Mechanism of the Zika Virus Crossing the Placental Barrier and the Blood-Brain Barrier

doi: 10.3389/fmicb.2020.00214

Figure Lengend Snippet: ZIKV crossed in vitro human physiological barriers. (A) A schematic diagram depicts a virus crossing in vitro barrier cells. (B) The immunofluorescence images of the ZIKV E protein revealed ZIKV-infected Vero cells cultured in a basal chamber through a JEG-3 barrier at a MOI of 0.5 or hCMEC/D3 barrier at MOI of 10 at 24 h post-infection. The ZIKV E protein antibody was recognized by an Alex488-secondary antibody (green). Nuclear DNA was stained with DAPI (blue). Scale bar, 50 μm. (C) FITC-dextran (40 kDa) was used to validate the permeability of a JEG-3 barrier at 24 h post-infection. The FITC-dextran of the basal chamber was collected and analyzed by a plate reader. The fold change of the FITC signal compared with the mock group was drawn in (C) . 12.5 μM of EDTA was used as a positive control. The permeability of an hCMEC/D3 barrier at 24 h ZIKV post-infection was measured in (D) . The error bars represent standard deviations from three independent experiments. * p < 0.05; ** p < 0.01; **** p < 0.0001.

Article Snippet: The immortalized human brain capillary endothelial cell line hCMEC/D3 (SCC066, Merck) was cultured in the EndoGRO TM -MV Complete Media Kit (Merck).

Techniques: In Vitro, Immunofluorescence, Infection, Cell Culture, Staining, Permeability, Positive Control

Journal: Frontiers in Microbiology

Article Title: The Mechanism of the Zika Virus Crossing the Placental Barrier and the Blood-Brain Barrier

doi: 10.3389/fmicb.2020.00214

Figure Lengend Snippet: ZIKV infection decreased expression of the tight junction protein ZO-1 and Occludin in JEG-3 but not hCMEC/D3. (A) Western blotting depicted that the amount of ZO-1 and Occludin decreased in JEG-3 cells at 24 h post-infection. The amount of ZO-1 and Occludin was normalized by β-actin and GAPDH, respectively. Comparison with the mock group was quantified in the right panel. Statistical differences were obtained through t -tests. ** p < 0.05. (B) Western blotting depicted that the expression of ZO-1 and Occludin were not affected in hCMEC/D3 cells with ZIKV infection. The amount of ZO-1 and Occludin was normalized by GAPDH, and quantified in the right panel. (C) The distribution of ZO-1 and Occludin in JEG-3 cells with/without ZIKV infection were imaged by confocal microscopy. The white arrows indicate the disruption of ZO-1 and Occludin. Scale bar, 20 μm. The tight junction integrity of ZO-1 and occludin were measured by the Skeletonize plug-in of ImageJ software. The length of segments in each group ( n = 50) were measured and quantified as right histograms. ** p < 0.01, *** p < 0.001.

Article Snippet: The immortalized human brain capillary endothelial cell line hCMEC/D3 (SCC066, Merck) was cultured in the EndoGRO TM -MV Complete Media Kit (Merck).

Techniques: Infection, Expressing, Western Blot, Confocal Microscopy, Software

Journal: Frontiers in Microbiology

Article Title: The Mechanism of the Zika Virus Crossing the Placental Barrier and the Blood-Brain Barrier

doi: 10.3389/fmicb.2020.00214

Figure Lengend Snippet: ZIKV infection decreased the accumulation of tight junction proteins through post-transcription regulation. (A) The mRNA levels of ZO-1, Occludin and ZIKV-E in JEG-3 cells upon ZIKV infection were quantified by RT-qPCR. Data were normalized by GAPDH. The fold changes of mRNA were compared with the mock group. The mRNA levels of ZO-1, Occludin and ZIKV-E in hCMEC/D3 upon ZIKV infection are shown in (B) . The error bars represent standard deviations from three independent experiments. (C) Western blotting depicts the accumulation of ZO-1 and Occludin in JEG-3 cells with various inhibitors treatments upon ZIKV infection. The amount of ZO-1 and Occludin was normalized by GAPDH. The fold changes of protein expression compared with the mock group in each term were quantified in the right panel. The error bars represent standard deviations from three independent experiments. * p < 0.05.

Article Snippet: The immortalized human brain capillary endothelial cell line hCMEC/D3 (SCC066, Merck) was cultured in the EndoGRO TM -MV Complete Media Kit (Merck).

Techniques: Infection, Quantitative RT-PCR, Western Blot, Expressing

Journal: Frontiers in Microbiology

Article Title: The Mechanism of the Zika Virus Crossing the Placental Barrier and the Blood-Brain Barrier

doi: 10.3389/fmicb.2020.00214

Figure Lengend Snippet: Detection of ZIKV transcytosis. (A) Fluorescence intensity profiles of elution from a Sephadex G-25 size-exclusion column is shown. An Atto647N-ZIKV particle was purified within the fraction layer 6 to 10 from gel filtration (solid squares). No fluorescence signal was detected in Atto647N-free ZIKV (solid triangles) within the same fraction layers. The 40 nm fluorescence microspheres were used as a size marker (solid circles). (B) Plaque assays were performed to determine the infectivity of the Atto647N-ZIKV. Dye-free ZIKV was used as a control. Both virus samples were purified from the same fractions of the Sephadex G-25 column. (C) Atto647N-ZIKV was used to validate virus crossing efficiency in JEG-3 and hCMEC/D3 barriers at 37 or 4 °C. (D) FITC-dextran was used to validate the permeability of JEG-3 and hCMEC/D3 barriers in (C) condition. The error bars represent standard deviations from three independent experiments. ** p < 0.01, *** p < 0.001.

Article Snippet: The immortalized human brain capillary endothelial cell line hCMEC/D3 (SCC066, Merck) was cultured in the EndoGRO TM -MV Complete Media Kit (Merck).

Techniques: Fluorescence, Purification, Filtration, Marker, Infection, Permeability

Journal: Frontiers in Microbiology

Article Title: The Mechanism of the Zika Virus Crossing the Placental Barrier and the Blood-Brain Barrier

doi: 10.3389/fmicb.2020.00214

Figure Lengend Snippet: Visualization of ZIKV transcytosis. The ZX images of confocal microscopy depicted the distribution of atto647-ZIKV from the apical side to the basolateral side within JEG-3 cells (A) and hCMEC/D3 cells (B) at 0, 30, and 60 min post-infection. The white arrows indicated att647-ZIKV particles (red) (A,B) . Nucleus was stained by DAPI staining (blue in A ), and cell membrane was stained by WGA488 (green) (A,B) .

Article Snippet: The immortalized human brain capillary endothelial cell line hCMEC/D3 (SCC066, Merck) was cultured in the EndoGRO TM -MV Complete Media Kit (Merck).

Techniques: Confocal Microscopy, Infection, Staining

Journal: Frontiers in Microbiology

Article Title: The Mechanism of the Zika Virus Crossing the Placental Barrier and the Blood-Brain Barrier

doi: 10.3389/fmicb.2020.00214

Figure Lengend Snippet: ZIKV transcytosis across JEG-3 and hCMEC/D3 barriers can be reduced by endocytosis and microtubule inhibitors. (A) The cell viabilities of JEG-3 and hCMEC/D3 with/without various inhibitors treatment were analyzed by a SRB assay. (B) The permeability of JEG-3 and hCMEC/D3 barriers with/without various inhibitors treatment were validated by detecting FITC-dextran across cell barriers. (C) Atto647-ZIKV was used to validate virus crossing efficiency in JEG-3 and hCMEC/D3 barriers with/without inhibitors treatment. (D) A plaque assay was used to measure the virus titers across JEG-3 and hCMEC/D3 barriers with/without various inhibitor treatments. All results are depicted as fold changes compared to the non-treatment control. The error bars represent standard deviations from three independent experiments. * p < 0.05; ** p < 0.01; **** p < 0.0001.

Article Snippet: The immortalized human brain capillary endothelial cell line hCMEC/D3 (SCC066, Merck) was cultured in the EndoGRO TM -MV Complete Media Kit (Merck).

Techniques: Sulforhodamine B Assay, Permeability, Plaque Assay