Intracellular localization of group II phospholipase A2 in rat vascular smooth muscle cells and its possible relationship to eicosanoid formation.
We investigated the localization of group II phospholipase A2 (PLA2-II) in rat vascular smooth muscle cells (VSMCs) by applying immunofluorescence and immunoelectron microscopy with its polyclonal antibody. In unstimulated cells, no immunolabelling was detected in the cells. On the other hand, in the cells stimulated with tumor necrosis factor (TNF) and/or forskolin (FK), intense fluorescence was detected in the cytoplasm. The immunoperoxidase reactions were detected in the cisternae of rough endoplasmic reticulum (rER), trans-cisternae of Golgi apparatus, and small vesicles beneath the plasma membrane. Western blot analysis showed VSMCs secrete PLA2-II after stimulation. Secreted PLA2-II was associated with the plasma membrane and extracellular matrix. Colchicine inhibited PLA2-II synthesis and its secretion to the extracellular space. These observations indicate that in VSMCs PLA2-II is synthesized at rER. transported to Golgi apparatus, discharged into extracellular space via the small vesicles, and microtubules may concern with its process. Furthermore, in VSMCs treated with TNF or TNF + FK, prostaglandin E2 formation was also increased. Actinomycin D and cycloheximide inhibited the potentiation of the prostaglandin E2 formation induced by TNF or TNF + FK, indicating that both RNA and protein synthesis are required for the potentiation. These results suggest an involvement of PLA2-II in the prostaglandin formation.
Kurihara
H
H
Shionogi Research Laboratories, Shionogi & Co., Ltd., Osaka, Japan.
Nakano
T
T
Takasu
N
N
Arita
H
H
D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D003078:Colchicine / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D015232:Dinoprostone / Q000096:biosynthesis*; D005455:Fluorescent Antibody Technique; D007124:Immunoenzyme Techniques; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000201:enzymology*; D010741:Phospholipases A / Q000037:antagonists & inhibitors / Q000378:metabolism*; D054467:Phospholipases A2; D051381:Rats; D011919:Rats, Inbred Strains; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D014409:Tumor Necrosis Factor-alpha; D003609:Dactinomycin; D005576:Colforsin; D003513:Cycloheximide; D010741:Phospholipases A; D054467:Phospholipases A2; D015232:Dinoprostone; D003078:Colchicine