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Characterization of ryanodine receptor-mediated calcium release in human B cells: relevance to diagnostic testing for malignant hyperthermia.

Abstract：

BACKGROUND:Mutations in the ryanodine type 1 receptor (RyR1) are causative for malignant hyperthermia. Studies in human B lymphocytes have shown that measurement of RyR1-mediated intracellular Ca(2+) (Ca(2+)(i)) release can differentiate between normal and malignant hyperthermia-susceptible individuals. The authors have further developed the B-cell assay by pharmacologically characterizing RyR1-mediated Ca release in two normal human B-cell lines and demonstrating increased sensitivity of lymphocytes to the RyR1 agonist 4-chloro-m-cresol (4-CmC) in the porcine model of MH. METHODS:Ca(2+)(i) was measured fluorometrically using fura-2 in populations of cells in suspension or with fluo-4 in single cells using confocal microscopy. The Dakiki and PP normal human B cell lines were used, as well as lymphocytes obtained from normal and malignant hyperthermia-susceptible pigs. 4-CmC was used to elicit RyR1-mediated Ca release; all experiments were performed in the absence of external Ca(2+). RESULTS:EC(50) values for 4-CmC were 0.98 and 1.04 mm for Dakiki and PP cells, respectively, demonstrating reproducibility. The 4-CmC-induced increase in Ca(2+)(i) was eliminated by thapsigargin and was unaffected by xestospongin C. The Ca(2+)(i) increase was separable from mitochondrial stores and was inhibited by azumolene. Caffeine did not induce Ca(2+)(i) release, but ryanodine depleted intracellular stores by 50%. Lymphocytes from pigs carrying the Arg614Cys mutation in RyR1 showed increased sensitivity to 4-CmC (EC(50) = 0.47 vs. 0.81 mm for cells derived from normal animals). CONCLUSIONS:RyR1-mediated Ca(2+) signals can be pharmacologically distinguished from other intracellular sources in human B cells, and alterations of RyR1 function can be successfully detected using Ca(2+) release from intracellular stores as an end point.

journal_name

Anesthesiology

journal_title

Anesthesiology

authors

McKinney LC,Butler T,Mullen SP,Klein MG

doi

10.1097/00000542-200606000-00014

subject

Has Abstract

pub_date

2006-06-01 00:00:00

pages

1191-201

issue

eissn

0003-3022

issn

1528-1175

pii

00000542-200606000-00014

journal_volume

104

pub_type

杂志文章

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Characterization of ryanodine receptor-mediated calcium release in human B cells: relevance to diagnostic testing for malignant hyperthermia.