title: 12(S)-HETE increases intracellular Ca(2+) in lymph-endothelial cells disrupting their barrier function in vitro; stabilization by clinical drugs impairing calcium supply.
abstract: Secretion of 12(S)-HETE by breast cancer emboli provokes "circular chemorepellent induced defects" (CCIDs) in the adjacent lymphatic vasculature facilitating their intravasation and lymph node metastasis which determines prognosis. Therefore, elucidating the mechanism of lymph endothelial cell (LEC) wall disintegration may provide cues for anti-metastatic intervention. The role of intracellular free Ca(2+) for CCID formation was investigated in LECs using MCF-7 or MDA-MB231 breast cancer cell spheroids in a three-dimensional cell co-culture model. 12(S)-HETE elevated the Ca(2+) level in LEC by activating PLC/IP3. Downstream, the Ca(2+)-calmodulin kinase MYLK contributed to the phosphorylation of Ser19-MLC2, LEC contraction and CCID formation. Approved clinical drugs, lidoflazine, ketotifen, epiandrosterone and cyclosporine, which reportedly disturb cellular calcium supply, inhibited 12(S)-HETE-induced Ca(2+) increase, Ser19-MLC2 phosphorylation and CCID formation. This treatment strategy may reduce spreading of breast cancer through lymphatics.
authors.affiliation: Department of Clinical Pharmacy and Diagnostics, University of Vienna, Althanstrasse 14, A-1090 Vienna, Austria.
authors.forename: Chi Huu
authors.initials: CH
authors.lastname: Nguyen
authors.affiliation: Department of Clinical Pharmacy and Diagnostics, University of Vienna, Althanstrasse 14, A-1090 Vienna, Austria.
authors.forename: Stefan
authors.initials: S
authors.lastname: Brenner
authors.affiliation: Clinical Institute of Pathology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.
authors.forename: Nicole
authors.initials: N
authors.lastname: Huttary
authors.affiliation: Department of Pharmacognosy, University of Vienna, Althanstrasse 14, A-1090 Vienna, Austria.
authors.forename: Yuanfang
authors.initials: Y
authors.lastname: Li
authors.affiliation: Department of Pharmacognosy, University of Vienna, Althanstrasse 14, A-1090 Vienna, Austria; Institute of Genetics and Animal Breeding of the Polish Academy of Sciences, Postępu 36A Street, Jastrzebiec 05-552, Poland.
authors.forename: Atanas Georgiev
authors.initials: AG
authors.lastname: Atanasov
authors.affiliation: Department of Pharmacognosy, University of Vienna, Althanstrasse 14, A-1090 Vienna, Austria.
authors.forename: Verena M
authors.initials: VM
authors.lastname: Dirsch
authors.affiliation: Clinical Institute of Pathology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.
authors.forename: Silvio
authors.initials: S
authors.lastname: Holzner
authors.affiliation: Clinical Institute of Pathology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.
authors.forename: Serena
authors.initials: S
authors.lastname: Stadler
authors.affiliation: Department of Clinical Pharmacy and Diagnostics, University of Vienna, Althanstrasse 14, A-1090 Vienna, Austria.
authors.forename: Juliane
authors.initials: J
authors.lastname: Riha
authors.affiliation: Clinical Institute of Pathology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.
authors.forename: Sigurd
authors.initials: S
authors.lastname: Krieger
authors.affiliation: Clinical Institute of Pathology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.
authors.forename: Danijela
authors.initials: D
authors.lastname: Milovanovic
authors.affiliation: Department of Clinical Pharmacy and Diagnostics, University of Vienna, Althanstrasse 14, A-1090 Vienna, Austria; Clinical Institute of Pathology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.
authors.forename: Adryan
authors.initials: A
authors.lastname: Fristiohardy
authors.affiliation: Clinical Institute of Pathology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.
authors.forename: Ingrid
authors.initials: I
authors.lastname: Simonitsch-Klupp
authors.affiliation: Institute of Medical Genetics, Medical University of Vienna, Waehringer Strasse 10, Vienna, A-1090 Austria.
authors.forename: Helmut
authors.initials: H
authors.lastname: Dolznig
authors.affiliation: Department of Medical and Chemical Laboratory Diagnostics, Medical University of Vienna, Waehringer Guertel 18-20, Vienna A-1090, Austria.
authors.forename: Philipp
authors.initials: P
authors.lastname: Saiko
authors.affiliation: Department of Medical and Chemical Laboratory Diagnostics, Medical University of Vienna, Waehringer Guertel 18-20, Vienna A-1090, Austria.
authors.forename: Thomas
authors.initials: T
authors.lastname: Szekeres
authors.affiliation: Department of Clinical Pharmacy and Diagnostics, University of Vienna, Althanstrasse 14, A-1090 Vienna, Austria; Clinical Institute of Pathology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.
authors.forename: Benedikt
authors.initials: B
authors.lastname: Giessrigl
authors.affiliation: Department of Clinical Pharmacy and Diagnostics, University of Vienna, Althanstrasse 14, A-1090 Vienna, Austria.
authors.forename: Walter
authors.initials: W
authors.lastname: Jäger
authors.affiliation: Clinical Institute of Pathology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria. Electronic address: georg.krupitza@meduniwien.ac.at.
authors.forename: Georg
authors.initials: G
authors.lastname: Krupitza
mesh_terms: D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid / Q000494:pharmacology*; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology*; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D065096:Calcium Chelating Agents / Q000494:pharmacology; D020013:Calcium Signaling / Q000187:drug effects*; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D024722:Cardiac Myosins / Q000378:metabolism; D002465:Cell Movement; D018920:Coculture Techniques; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D005260:Female; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D008207:Lymphatic Metastasis; D042601:Lymphatic Vessels / Q000187:drug effects* / Q000378:metabolism; D061986:MCF-7 Cells; D018994:Myosin Light Chains / Q000378:metabolism; D009219:Myosin-Light-Chain Kinase / Q000235:genetics / Q000378:metabolism; D010539:Permeability; D010766:Phosphorylation; D034622:RNA Interference; D012694:Serine; D018874:Spheroids, Cellular; D013997:Time Factors; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism
chemical_list: D002121:Calcium Channel Blockers; D065096:Calcium Chelating Agents; D002135:Calcium-Binding Proteins; D018994:Myosin Light Chains; C107184:myosin light chain 2; D012694:Serine; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D015544:Inositol 1,4,5-Trisphosphate; C499378:MYLK protein, human; D009219:Myosin-Light-Chain Kinase; D010738:Type C Phospholipases; D024722:Cardiac Myosins; D002118:Calcium
keywords: 12(S)-HETE; Endothelial integrity disruption; Intracellular Ca(2+); MLC2; MYLK; PLC/IP(3)