TGF-Beta Modulates the Integrity of the Blood Brain Barrier In Vitro, and Is Associated with Metabolic Alterations in Pericytes
Abstract
:1. Introduction
2. Material and Methods
2.1. Cell Culture
2.2. Isolation of Primary Porcine Brain Microvascular Endothelial Cells
2.3. The In Vitro BBB Model
2.4. Immunocytochemistry and Immunofluorescence
2.5. Freeze Fracture Electron Microscopy
2.6. Metabolic Analysis
2.7. 1H-NMR Spectroscopy-Based Metabolomics of HBVP Intracellular Metabolites
2.8. RNA Sequencing
2.9. Combined Metabolomics and RNA-Seq Pathway Analysis
2.10. Statistics
3. Results
3.1. Co-Cultures of Primary Endothelial Cells and Pericytes Develop a Tight BBB
3.2. TGF-β Treatment of HBVP Negativels Modelates the Integrity of the BBB
3.3. TGF-β Treatment Leads to a Distinct Metabolite Profile, Pathway Alterations, and Gene Expression Patterns in HBVP
3.4. Joint-Pathway Analysis Illustrates Further Pathways Dysregulated upon TGF-β Treatment
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
ADAMTS6 | ADAM metallopeptidase with thrombospondin type 1 motif, 6 |
AJ | adherent junction |
ANOVA | analysis of variance |
BBB | blood–brain barrier |
BCAA | branched amino acid |
BMVEC | brain microvascular endothelial cells |
BSA | bovine serum albumin |
CM | conditioned medium |
CNS | central nerve system |
DMEM | Dulbecco’s Modified Eagle’s Medium |
ECGS | endothelial cell growth supplements |
EGF | epidermal growth factor |
EM | epithelial cell growth medium |
EMT | epithelial-to-mesenchymal transition |
ESM1 | endothelial cell-specific molecule 1/endocan |
FC | fold change |
FCS | fetal calf serum |
FGF | fibroblast growth factor |
FDR | false discovery rate |
FOXS1 | forkhead box S1 |
GBM | glioblastoma |
GJ | gap junction |
GPR183 | G protein-coupled receptor 183 |
HBVP | human brain microvascular pericytes |
HRP | horseradish peroxidase |
IGF-1 | insulin-like growth factor 1 |
KEGG | Kyoto Encyclopedia of Genes and Genomes |
mTORC1 | mammalian target of rapamycin complex 1 |
NOX4 | NADPH oxidase 4 |
NVU | neurovascular unit |
o/n | overnight |
OxPhos | oxidative phosphorylation |
PBMVEC | porcine brain microvascular endothelial cells |
PCA | principal component analysis |
PGS | pericyte growth supplement |
PLL | poly-L-lysine |
PM | pericyte medium |
PMET | plasma membrane electron transport |
PPARGC1AC | peroxisome proliferator-activated receptor gamma, coactivator 1 alpha |
PRR5 | proline rich 5 |
P/S | penicillin/streptomycin |
RT | room temperature |
SLC46A3 | solute carrier family 46, member 3 |
SD | standard deviation |
SEM | standard error of the mean |
SPOCK1 | sparc/osteonectin, cwcv and kazal-like domains proteoglycan 1 |
SV-GA | SV40 large T-antigene immortalized astrocytic cells |
TEER | transendothelial electric resistance |
TGF-β | transforming growth factor beta |
TJ | tight junction |
VEGF | vascular endothelial growth factor |
ZO | zonula occludens |
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Schumacher, L.; Slimani, R.; Zizmare, L.; Ehlers, J.; Kleine Borgmann, F.; Fitzgerald, J.C.; Fallier-Becker, P.; Beckmann, A.; Grißmer, A.; Meier, C.; et al. TGF-Beta Modulates the Integrity of the Blood Brain Barrier In Vitro, and Is Associated with Metabolic Alterations in Pericytes. Biomedicines 2023, 11, 214. https://doi.org/10.3390/biomedicines11010214
Schumacher L, Slimani R, Zizmare L, Ehlers J, Kleine Borgmann F, Fitzgerald JC, Fallier-Becker P, Beckmann A, Grißmer A, Meier C, et al. TGF-Beta Modulates the Integrity of the Blood Brain Barrier In Vitro, and Is Associated with Metabolic Alterations in Pericytes. Biomedicines. 2023; 11(1):214. https://doi.org/10.3390/biomedicines11010214
Chicago/Turabian StyleSchumacher, Leonie, Rédouane Slimani, Laimdota Zizmare, Jakob Ehlers, Felix Kleine Borgmann, Julia C. Fitzgerald, Petra Fallier-Becker, Anja Beckmann, Alexander Grißmer, Carola Meier, and et al. 2023. "TGF-Beta Modulates the Integrity of the Blood Brain Barrier In Vitro, and Is Associated with Metabolic Alterations in Pericytes" Biomedicines 11, no. 1: 214. https://doi.org/10.3390/biomedicines11010214