Abstract
The effect of 3-[1-(p-chlorobenzyl)-5-(isopropyl)-3-tert-butylthioindol-2-yl]-2, 2-dimethylpropanoic acid (MK-886), a leukotriene biosynthesis inhibitor, on Ca2+ mobilization in Madin- Darby canine kidney cells has been examined by fluorimetry using fura-2 as a Ca2+ indicator. MK-886 at 0.5 to 25 μM concentration dependently increased [Ca2+]i. The [Ca2+]i increase comprised an immediate initial rise and a slowly decaying phase. Ca2+ removal inhibited the Ca2+ signals by reducing both the initial rise and the decay phase, suggesting that MK-886 activated Ca2+ influx and Ca2+ release. In Ca2+-free medium, 10 μM MK-886 still increased [Ca2+]i after pretreatment with carbonylcyanide m-chlorophenylhydrazone (CCCP; 2 μM), a mitochondrial uncoupler, and thapsigargin (1 μM), an endoplasmic reticulum Ca2+ pump inhibitor. Conversely, pretreatment with MK-886 abolished CCCP- and thapsigargin-induced Ca2+release. This suggests that 10 μM MK-886 released Ca2+from the endoplasmic reticulum, mitochondria, and other stores. The addition of 3 mM Ca2+ increased [Ca2+]i after pretreatment with 10 μM MK-886 for 700 s in Ca2+-free medium, indicating that MK-886 induced capacitative Ca2+ entry. This capacitative Ca2+ entry was partly inhibited by SKF96365 (50 μM), by econazole (25 μM), and by inhibiting phospholipase A2with aristolochic acid (40 μM) but not by inhibiting phospholipase D with 0.1 mM propranolol. MK-886 (10 μM)-induced Ca2+release was not altered by inhibiting phospholipase C with U73122 (2 μM) but was inhibited by 50% by suppressing phospholipase D and phospholipase A2 with propranolol (0.1 mM) and aristolochic acid (40 μM), respectively.
Footnotes
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Send reprint requests to: Chung-Ren Jan, Ph.D., Department of Medical Education and Research, Veterans General Hospital-Kaohsiung, 386 Ta Chung 1st Rd, Kaohsiung, Taiwan 813. E-mail:crjan{at}isca.vghks.gov.tw
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↵1 This work was supported by grants from the National Science Council (NSC88-2314-B-075B-003), Veterans General Hospital-Kaohsiung (VGHKS89-13), and VTY Joint Research Program, Tsou's Foundation (VTY88-P3-24) to C.-R.J.
- Abbreviations:
- MK-886
- 3-[1-(p-chlorobenzyl)-5-(isopropyl)-3-tert-butylthioindol-2-yl]-2, 2-dimethylpropanoic acid
- DMEM
- Dulbecco's modified Eagle's medium
- ER
- endoplasmic reticulum
- fura-2/AM
- 1-[2-(5-carboxyoxazol-2-yl)-6-aminobenzofuran-5-oxy]-2-(2′-amino-5′-methylphenoxy)-ethane-N,N,N,N-tetraacetic acid pentaacetoxymethyl ester
- IP3
- inositol 1,4,5-trisphosphate
- MDCK
- Madin-Darby canine kidney
- U73122
- 1-(6-((17β-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione
- U73343
- 1-(6-((17β-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-2,5-pyrrolidine-dione
- CCCP
- carbonylcyanide m-chlorophenylhydrazone
- SKF96365
- 1-[β-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]-1H-imidazole hydrochloride
- Received August 2, 1999.
- Accepted March 14, 2000.
- The American Society for Pharmacology and Experimental Therapeutics
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