[Expression and role of inositol 1,4,5-trisphosphate receptor and ryanodine receptor in a human lens epithelial cell line].
OBJECTIVE
To determinate the existence of the inositol 1,4,5-triphosphate receptor (IP(3R)) and the ryanodine receptor (RyR), identify their isoforms and assess their functions during calcium ion concentration modulation in a human epithelial cell line (LEC-B3).


METHODS
LEC-B3 cell line was defrosted and subcultured, the 3rd passage cells were chosen as object when they became confluent. Then the total cellular RNA was extracted from the cells, the expression of mRNA of IP(3R) and RyR isoforms was analyzed by polymerase chain reaction. Ca(2+) ion changes in the same passage cells were observed and analyzed with laser scanning confocal microscope when acetylcholine (Ach), atropine, caffeine, ryanodine, and procaine were added to the extracellular Ca(2+), Mg(2+) free solutions with ethyleneglycol bis ether tetracetic acid or Ca(2+), Mg(2+) involved solutions.


RESULTS
The IP(3R) isoform I, III and RyR isoform I, III mRNA were expressed, the expression of isoform IP(3R) III > IP(3R)I (P < 0.05); RyR III > RyRI (P < 0.01). 10 micro mol/L Ach, 10 mmol/L caffeine and 50 micro mol/L ryanodine all could induce the intracellular free Ca(2+) concentration increase, but the incubation periods, ranges and lasting time of the increases are different: when Ach acted, the duration of calcium increase in calcium involved solutions was a little longer than in calcium free solutions; the amplitude of calcium oscillations induced by Ach was much higher than caffeine and ryanodine, while the onset of the later is slower than the former. Besides, 1 micro mol/L atropine could block the Ach effects, 50 micro mol/L procaine could inhibit the ryanodine effects.


CONCLUSIONS
(1) The IP(3R) and RyR gene transcription coexist in the human lens epithelial cell line: LEC-B3, and their isoforms have specificity. (2) Different agonists can induce different calcium changes. (3)The increase of Ca(2+) concentration induced by caffeine or ryanodine suggested the involvement of both the cyclic adenosine diphosphate ribose and the inositol 1,4,5-triphosphate systems in the regulation of intracellular calcium in LEC-B3, and IP(3R) is more sensitive to free Ca(2+) modulation than RyR.
Qu
Bo
B
Department of Ophthalmology, First Affiliated Hospital of China Medical University, Shenyang 110001, China. qubo24@hotmail.com
Zhang
Jin-song
JS
D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000096:biosynthesis* / Q000235:genetics; D002460:Cell Line; D004847:Epithelial Cells / Q000378:metabolism*; D006801:Humans; D007231:Infant, Newborn; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007908:Lens, Crystalline / Q000166:cytology*; D012333:RNA, Messenger / Q000096:biosynthesis; D018160:Receptors, Cytoplasmic and Nuclear / Q000096:biosynthesis* / Q000235:genetics; D019837:Ryanodine Receptor Calcium Release Channel / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism
D015220:Calcium Channels; C506926:ITPR1 protein, human; D053496:Inositol 1,4,5-Trisphosphate Receptors; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D019837:Ryanodine Receptor Calcium Release Channel; D002118:Calcium