Cellular membrane fluidity measurement by fluorescence polarization in indomethacin-induced gastric cellular injury in vitro.
BACKGROUND
Gastric complications of indomethacin involve generation of reactive oxygen species, which induce gastric mucosal injury via lipid peroxidation of cell membranes. Peroxidation by reactive oxygen species alters the amounts of unsaturated fatty acids in the cell membrane and thus affects membrane fluidity. Indomethacin-induced lipid peroxidation can thus be detected by measuring cellular membrane fluidity by the fluorescence polarization (FP) method. The aim of this study was to elucidate the usefulness of the FP method for detecting indomethacin-induced gastric cellular injury in RGM-1 cells.


METHODS
Indomethacin-treated RGM-1 cells were investigated by conventional cytotoxicity assay, fluorometry of diphenyl-1-pyrenylphosphine (DPPP) to detect lipid peroxidation, and FP. The effects of both a radical scavenger and an initiator on membrane fluidity change (MFC) in RGM-1 cells were examined. The sensitivity of FP in detecting cellular injury was compared with those of DPPP fluorometry and conventional cytotoxicity measurements.


RESULTS
Indomethacin caused an increase in MFC as determined by FP before cytotoxicity was detected by conventional methods. The increase in MFC was associated with increased membrane phospholipid peroxidation (MPP) but not with a prostaglandin deficiency, and the increases in both MFC and MPP were prevented by vitamin E.


CONCLUSIONS
The FP method is potentially useful for detecting cellular injury in vitro.
Kaneko
Tsuyoshi
T
Division of Gastroenterology, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Ten-nohdai, Tsukuba 305-8575, Japan.
Matsui
Hirofumi
H
Shimokawa
Osamu
O
Nakahara
Akira
A
Hyodo
Ichinosuke
I
D000818:Animals; D002460:Cell Line; D004847:Epithelial Cells / Q000737:chemistry / Q000187:drug effects; D005454:Fluorescence Polarization / Q000379:methods*; D005753:Gastric Mucosa / Q000187:drug effects*; D007213:Indomethacin / Q000633:toxicity*; D008560:Membrane Fluidity / Q000187:drug effects*; D008563:Membrane Lipids / Q000378:metabolism; D009943:Organophosphorus Compounds / Q000032:analysis; D010743:Phospholipids / Q000378:metabolism; D011721:Pyrenes / Q000032:analysis; D051381:Rats; D012680:Sensitivity and Specificity; D014810:Vitamin E / Q000494:pharmacology
D008563:Membrane Lipids; D009943:Organophosphorus Compounds; D010743:Phospholipids; D011721:Pyrenes; C066322:diphenyl-1-pyrenylphosphine; D014810:Vitamin E; D007213:Indomethacin