Marinobufagenin regulates permeability and gene expression of brain endothelial cells.
Marinobufagenin (MBG) is a cardiotonic steroid that increases in the circulation in preeclampsia. Preeclampsia and eclampsia are associated with cerebral edema. Therefore, we examined the effects of MBG on human brain microvascular endothelial cells (HBMEC) in vitro. MBG enhanced the permeability of HBMEC monolayers at 1-, 10-, and 100-nM doses, but had no effect at 0.1 nM. Agilent Human Gene Expression microarrays were utilized in these studies. MBG treatment (10 nM for 12 h) downregulated concentrations of the soluble VEGFR transcript sFLT by 59% but did not alter those of FLTv3 mRNA (determined by quantitative PCR). When treated and control HBMEC transcriptomes were interrogated on microarrays, 1,069 genes appeared to be regulated by MBG. Quantitative RT-PCR confirmed that MBG treatment upregulated ENKUR mRNA concentrations by 57%. Its protein product interacts with calmodulin and calcium channel proteins. MBG treatment downregulated several genes whose protein products are involved in cell adhesion (ITGA2B, FERMT1, CLDN16, and TMEM207) and cell signaling (GRIN2C, SLC8A1, and ESR1). The level of downregulation ranged from 22 to 66%. Altogether, MBG actively enhanced the permeability of HBMEC monolayers while downregulating genes involved in adhesion. MBG treatment had variable effects on ENKUR, GRIN2C, and SLC8A1 genes, all associated with calcium transport. These studies provide the basis for future investigations of MBG actions in normal physiology and disease.
Department of Animal Science, College of Veterinary Medicine and Biosciences, Texas A&M University, College Station, Texas;
Nancy H
NH
Ing
Department of Poultry Science, College of Agriculture and Life Sciences, Texas A&M University, College Station, Texas; and.
Luc
L
Berghman
Department of Poultry Science, College of Agriculture and Life Sciences, Texas A&M University, College Station, Texas; and.
Daad
D
Abi-Ghanem
the Department of Veterinary Pathobiology, College of Veterinary Medicine and Biosciences, Texas A&M University, College Station, Texas.
Kamran
K
Abbas
the Department of Veterinary Pathobiology, College of Veterinary Medicine and Biosciences, Texas A&M University, College Station, Texas.
Aditi
A
Kaushik
Department of Animal Science, College of Veterinary Medicine and Biosciences, Texas A&M University, College Station, Texas;
Penny K
PK
Riggs
the Department of Veterinary Pathobiology, College of Veterinary Medicine and Biosciences, Texas A&M University, College Station, Texas jbpuschett@aol.com.
Jules B
JB
Puschett
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism; D001921:Brain / Q000098:blood supply*; D002018:Bufanolides / Q000494:pharmacology*; D002148:Calmodulin-Binding Proteins / Q000235:genetics / Q000378:metabolism; D002316:Cardiotonic Agents / Q000494:pharmacology*; D002463:Cell Membrane Permeability / Q000187:drug effects* / Q000502:physiology; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D066298:In Vitro Techniques; D018092:Kainic Acid Receptors / Q000235:genetics / Q000378:metabolism; D040262:Receptors, Vascular Endothelial Growth Factor / Q000235:genetics / Q000378:metabolism; D019831:Sodium-Calcium Exchanger / Q000235:genetics / Q000378:metabolism; D046888:Tissue Array Analysis; D000097806:GluK2 Kainate Receptor
D048868:Adaptor Proteins, Signal Transducing; D002018:Bufanolides; D002148:Calmodulin-Binding Proteins; D002316:Cardiotonic Agents; D018092:Kainic Acid Receptors; D040262:Receptors, Vascular Endothelial Growth Factor; D019831:Sodium-Calcium Exchanger; D000097806:GluK2 Kainate Receptor; C493290:ENKUR protein, human; C110374:sodium-calcium exchanger 1; C093896:marinobufagenin
bufodienolides; human brain endothelial cells; marinobufagenin; microarray; quantitative PCR