Abstract
To investigate the relevance of ·NO and oxyradicals in the blood-brain barrier (BBB), differentiated and well-proliferating brain capillary endothelial cells (BCEC) are required. Therefore, rat BCEC (rBCEC) were transfected with immortalizing genes. The resulting lines exhibited endothelial characteristics (factor VIII, angiotensin-converting enzyme, high prostacyclin/thromboxane release rates) and BBB markers (γ-glutamyl transpeptidase, alkaline phosphatase). The control line rBCEC2 (mock transfected) revealed fibroblastoid morphology, less factor VIII, reduced γ-glutamyl transpeptidase, weak radical defence, low prostanoid metabolism, and limited proliferation. Lines transfected with immortalizing genes (especially rBCEC4, polyoma virus large T antigen) conserved primary properties: epitheloid morphology, subcultivation with high proliferation rate under pure culture conditions, and powerful defence against reactive oxygen species (Mn-, Cu/Zn-superoxide dismutase, catalase, glutathione peroxidase, glutathione) effectively controlling radical metabolism. Only 100 μM H2O2 overcame this defence and stimulated the formation of eicosanoids similarly as in primary cells. Some BBB markers were expressed to a lower degree; however, cocultivation with astrocytes intensified these markers (e.g., alkaline phosphatase) and paraendothelial tightness, indicating induction of BBB properties. Inducible NO synthase was induced by a cytokine plus lipopolysaccharide mixture in all lines and primary cells, resulting in ·NO release. Comparing the cell lines obtained, rBCEC4 are stable immortalized and reveal the best conservation of properties from primary cells, including enzymes producing or decomposing reactive species. These cells can be subcultivated in large amounts and, hence, they are suitable to study the role of radical metabolism in the BBB and in the cerebral microvasculature.
| Original language | English |
|---|---|
| Pages (from-to) | 114-127 |
| Number of pages | 14 |
| Journal | Microvascular Research |
| Volume | 62 |
| Issue number | 2 |
| DOIs | |
| Publication status | Published - 2001 |
| Externally published | Yes |
Fingerprint
Keywords
- Blood-brain barrier
- Brain capillary endothelial cells
- Immortalization
- Nitric oxide
- Reactive oxygen species
ASJC Scopus subject areas
- Biochemistry
- Cardiology and Cardiovascular Medicine
Cite this
·NO and oxyradical metabolism in new cell lines of rat brain capillary endothelial cells forming the blood-brain barrier. / Blasig, Ingolf E.; Giese, Helga; Schroeter, Matthias L.; Sporbert, Anje; Utepbergenov, Darkhan I.; Buchwalow, Igor B.; Neubert, Katrin; Schönfelder, Gilbert; Freyer, Dorette; Schimke, Ingolf; Siems, Wolf Eberhard; Paul, Martin; Haseloff, Reiner F.; Blasig, Rosel.
In: Microvascular Research, Vol. 62, No. 2, 2001, p. 114-127.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - ·NO and oxyradical metabolism in new cell lines of rat brain capillary endothelial cells forming the blood-brain barrier
AU - Blasig, Ingolf E.
AU - Giese, Helga
AU - Schroeter, Matthias L.
AU - Sporbert, Anje
AU - Utepbergenov, Darkhan I.
AU - Buchwalow, Igor B.
AU - Neubert, Katrin
AU - Schönfelder, Gilbert
AU - Freyer, Dorette
AU - Schimke, Ingolf
AU - Siems, Wolf Eberhard
AU - Paul, Martin
AU - Haseloff, Reiner F.
AU - Blasig, Rosel
PY - 2001
Y1 - 2001
N2 - To investigate the relevance of ·NO and oxyradicals in the blood-brain barrier (BBB), differentiated and well-proliferating brain capillary endothelial cells (BCEC) are required. Therefore, rat BCEC (rBCEC) were transfected with immortalizing genes. The resulting lines exhibited endothelial characteristics (factor VIII, angiotensin-converting enzyme, high prostacyclin/thromboxane release rates) and BBB markers (γ-glutamyl transpeptidase, alkaline phosphatase). The control line rBCEC2 (mock transfected) revealed fibroblastoid morphology, less factor VIII, reduced γ-glutamyl transpeptidase, weak radical defence, low prostanoid metabolism, and limited proliferation. Lines transfected with immortalizing genes (especially rBCEC4, polyoma virus large T antigen) conserved primary properties: epitheloid morphology, subcultivation with high proliferation rate under pure culture conditions, and powerful defence against reactive oxygen species (Mn-, Cu/Zn-superoxide dismutase, catalase, glutathione peroxidase, glutathione) effectively controlling radical metabolism. Only 100 μM H2O2 overcame this defence and stimulated the formation of eicosanoids similarly as in primary cells. Some BBB markers were expressed to a lower degree; however, cocultivation with astrocytes intensified these markers (e.g., alkaline phosphatase) and paraendothelial tightness, indicating induction of BBB properties. Inducible NO synthase was induced by a cytokine plus lipopolysaccharide mixture in all lines and primary cells, resulting in ·NO release. Comparing the cell lines obtained, rBCEC4 are stable immortalized and reveal the best conservation of properties from primary cells, including enzymes producing or decomposing reactive species. These cells can be subcultivated in large amounts and, hence, they are suitable to study the role of radical metabolism in the BBB and in the cerebral microvasculature.
AB - To investigate the relevance of ·NO and oxyradicals in the blood-brain barrier (BBB), differentiated and well-proliferating brain capillary endothelial cells (BCEC) are required. Therefore, rat BCEC (rBCEC) were transfected with immortalizing genes. The resulting lines exhibited endothelial characteristics (factor VIII, angiotensin-converting enzyme, high prostacyclin/thromboxane release rates) and BBB markers (γ-glutamyl transpeptidase, alkaline phosphatase). The control line rBCEC2 (mock transfected) revealed fibroblastoid morphology, less factor VIII, reduced γ-glutamyl transpeptidase, weak radical defence, low prostanoid metabolism, and limited proliferation. Lines transfected with immortalizing genes (especially rBCEC4, polyoma virus large T antigen) conserved primary properties: epitheloid morphology, subcultivation with high proliferation rate under pure culture conditions, and powerful defence against reactive oxygen species (Mn-, Cu/Zn-superoxide dismutase, catalase, glutathione peroxidase, glutathione) effectively controlling radical metabolism. Only 100 μM H2O2 overcame this defence and stimulated the formation of eicosanoids similarly as in primary cells. Some BBB markers were expressed to a lower degree; however, cocultivation with astrocytes intensified these markers (e.g., alkaline phosphatase) and paraendothelial tightness, indicating induction of BBB properties. Inducible NO synthase was induced by a cytokine plus lipopolysaccharide mixture in all lines and primary cells, resulting in ·NO release. Comparing the cell lines obtained, rBCEC4 are stable immortalized and reveal the best conservation of properties from primary cells, including enzymes producing or decomposing reactive species. These cells can be subcultivated in large amounts and, hence, they are suitable to study the role of radical metabolism in the BBB and in the cerebral microvasculature.
KW - Blood-brain barrier
KW - Brain capillary endothelial cells
KW - Immortalization
KW - Nitric oxide
KW - Reactive oxygen species
UR - http://www.scopus.com/inward/record.url?scp=0034741553&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0034741553&partnerID=8YFLogxK
U2 - 10.1006/mvre.2001.2318
DO - 10.1006/mvre.2001.2318
M3 - Article
C2 - 11516240
AN - SCOPUS:0034741553
VL - 62
SP - 114
EP - 127
JO - Microvascular Research
JF - Microvascular Research
SN - 0026-2862
IS - 2
ER -