Cerebral Pericytes and Endothelial Cells Communicate through Inflammasome-Dependent Signals
Abstract
:1. Introduction
2. Results
2.1. Characterization of Inflammasome Activation in CECs
2.2. Activation of CECs by Pericytes Exposed to Inflammasome Activator Stimuli
2.3. Activation of Brain Pericytes by CECs Exposed to Inflammasome Activator Stimuli
3. Discussion
4. Materials and Methods
4.1. Isolation and Culture of Brain Endothelial Cells and Pericytes
4.2. Inflammatory Activation of Brain Endothelial Cells and Pericytes
4.3. Coculture Systems for the Study of Endothelial–Pericyte Communication
4.4. Measurement of Transendothelial Electrical Resistance (TEER)
4.5. Sample Preparation and Western Blot
4.6. Immunofluorescence and Fluorescence Microscopy
4.7. Real-Time PCR
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Primary Antibody | Cat. No. | Application |
---|---|---|
anti-IL-1β g. polyclonal | AF-401-NA (R&D Systems/Bio-Techne) | WB: 1:500 in 1% BSA in TBS-T |
anti-claudin-5 m. monoclonal | 35-2500 (Thermo Fisher Scientific) | WB: 1:300 in TBS-T |
IF: 1:50 in 1% BSA in PBS | ||
anti-occludin g. polyclonal | sc-8145 (Santa Cruz Biotechnology) | WB: 1:500 in TBS-T |
anti-β-actin m. monoclonal | AC-15 (Merck-Sigma) | WB: 1:1000 in 1% BSA in TBS-T |
anti-ZO-1 r. polyclonal | 61-7300 (Thermo Fisher Scientific) | IF: 1:50 in 1% BSA in PBS |
anti-NLRP3 g. polyclonal | GTX88190 (GeneTex, Hsinchu City, Taiwan) | IF: 1:200 in 1% BSA in PBS |
Secondary Antibody | Cat. No. | Application |
HRP-conjugated g. anti-mouse IgG (H + L) | 610094 (BD Biosciences, San Jose, CA, USA) | WB: 1:3000 in TBS-T |
HRP-conjugated g. anti-rabbit IgG (H + L) | 7074 (Cell Signaling Technology) | WB: 1:4000 in TBS-T |
Alexa Fluor® 594 AffiniPure g. anti-mouse IgG (H + L) | 115-585-003 (Jackson Immuno Research, Ely, UK) | IF: 1:600 in 1% BSA in PBS |
Alexa Fluor® 647 AffiniPure g. anti-rabbit IgG (H + L) | 111-605-003 (Jackson Immuno Research) | IF: 1:600 in 1% BSA in PBS |
Alexa Fluor® 488 Cross-Adsorbed d. anti-goat IgG (H + L) | A-11055 (Thermo Fisher Scientific) | IF: 1:600 in 1% BSA in PBS |
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Kozma, M.; Mészáros, Á.; Nyúl-Tóth, Á.; Molnár, K.; Costea, L.; Hernádi, Z.; Fazakas, C.; Farkas, A.E.; Wilhelm, I.; Krizbai, I.A. Cerebral Pericytes and Endothelial Cells Communicate through Inflammasome-Dependent Signals. Int. J. Mol. Sci. 2021, 22, 6122. https://doi.org/10.3390/ijms22116122
Kozma M, Mészáros Á, Nyúl-Tóth Á, Molnár K, Costea L, Hernádi Z, Fazakas C, Farkas AE, Wilhelm I, Krizbai IA. Cerebral Pericytes and Endothelial Cells Communicate through Inflammasome-Dependent Signals. International Journal of Molecular Sciences. 2021; 22(11):6122. https://doi.org/10.3390/ijms22116122
Chicago/Turabian StyleKozma, Mihály, Ádám Mészáros, Ádám Nyúl-Tóth, Kinga Molnár, Laura Costea, Zsófia Hernádi, Csilla Fazakas, Attila E. Farkas, Imola Wilhelm, and István A. Krizbai. 2021. "Cerebral Pericytes and Endothelial Cells Communicate through Inflammasome-Dependent Signals" International Journal of Molecular Sciences 22, no. 11: 6122. https://doi.org/10.3390/ijms22116122