Functional Properties of RYR1 Mutations Identified in Swedish Patients with Malignant Hyperthermia and Central Core Disease
(2010) In Anesthesia and Analgesia 111(1). p.185-190- Abstract
- BACKGROUND: A diagnosis of malignant hyperthermia susceptibility by in vitro contraction testing can often only be performed at specialized laboratories far away from where patients live. Therefore, we have designed a protocol for genetic screening of the RYR1-cDNA and for functional testing of newly identified ryanodine receptor 1 (RYR1) gene variants in B lymphocytes isolated from peripheral blood samples drawn at local primary care centers. METHODS: B lymphocytes were isolated for the extraction of RYR1-mRNA and genomic DNA and for establishment of lymphoblastoid B cell lines in 5 patients carrying yet unclassified mutations in the RYR1. The B lymphoblastoid cell lines were used to study resting cytoplasmic calcium concentration, the... (More)
- BACKGROUND: A diagnosis of malignant hyperthermia susceptibility by in vitro contraction testing can often only be performed at specialized laboratories far away from where patients live. Therefore, we have designed a protocol for genetic screening of the RYR1-cDNA and for functional testing of newly identified ryanodine receptor 1 (RYR1) gene variants in B lymphocytes isolated from peripheral blood samples drawn at local primary care centers. METHODS: B lymphocytes were isolated for the extraction of RYR1-mRNA and genomic DNA and for establishment of lymphoblastoid B cell lines in 5 patients carrying yet unclassified mutations in the RYR1. The B lymphoblastoid cell lines were used to study resting cytoplasmic calcium concentration, the peak calcium transient induced by the sarco(endo)plasmic reticulum Ca-ATPase inhibitor thapsigargin, and the dose-dependent calcium release induced by the ryanodine receptor agonist 4-chloro-m-cresol. RESULTS: It was possible to extract mRNA for cDNA synthesis and to create B lymphocyte clones from all samples. All B lymphoblastoid cell lines carrying RYR1 candidate mutations showed significantly increased resting cytoplasmic calcium levels as well as a shift to lower concentrations of 4-chloro-m-cresol inducing calcium release compared with controls. CONCLUSIONS: Peripheral blood samples are stable regarding RNA and DNA extraction and establishment of lymphoblastoid B cell lines after transportation at ambient temperature over large distances by ordinary mail. Functional tests on B cells harboring the newly identified amino acid substitutions indicate that they alter intracellular Ca2+ homeostasis and are most likely causative of malignant hyperthermia. (Anesth Analg 2010;111:185-90) (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/1629389
- author
- Vukcevic, Mirko ; Broman, Marcus ; Islander, Gunilla LU ; Bodelsson, Mikael LU ; Ranklev Twetman, Eva LU ; Mueller, Clemens R. and Treves, Susan
- organization
- publishing date
- 2010
- type
- Contribution to journal
- publication status
- published
- subject
- in
- Anesthesia and Analgesia
- volume
- 111
- issue
- 1
- pages
- 185 - 190
- publisher
- Lippincott Williams & Wilkins
- external identifiers
-
- wos:000279281500032
- scopus:77954684290
- pmid:20142353
- ISSN
- 1526-7598
- DOI
- 10.1213/ANE.0b013e3181cbd815
- language
- English
- LU publication?
- yes
- id
- 37d8bf76-2edc-4762-aeea-4aff7a013ae1 (old id 1629389)
- alternative location
- http://www.anesthesia-analgesia.org/content/111/1/185.abstract
- date added to LUP
- 2016-04-01 14:42:58
- date last changed
- 2022-03-06 20:46:37
@article{37d8bf76-2edc-4762-aeea-4aff7a013ae1, abstract = {{BACKGROUND: A diagnosis of malignant hyperthermia susceptibility by in vitro contraction testing can often only be performed at specialized laboratories far away from where patients live. Therefore, we have designed a protocol for genetic screening of the RYR1-cDNA and for functional testing of newly identified ryanodine receptor 1 (RYR1) gene variants in B lymphocytes isolated from peripheral blood samples drawn at local primary care centers. METHODS: B lymphocytes were isolated for the extraction of RYR1-mRNA and genomic DNA and for establishment of lymphoblastoid B cell lines in 5 patients carrying yet unclassified mutations in the RYR1. The B lymphoblastoid cell lines were used to study resting cytoplasmic calcium concentration, the peak calcium transient induced by the sarco(endo)plasmic reticulum Ca-ATPase inhibitor thapsigargin, and the dose-dependent calcium release induced by the ryanodine receptor agonist 4-chloro-m-cresol. RESULTS: It was possible to extract mRNA for cDNA synthesis and to create B lymphocyte clones from all samples. All B lymphoblastoid cell lines carrying RYR1 candidate mutations showed significantly increased resting cytoplasmic calcium levels as well as a shift to lower concentrations of 4-chloro-m-cresol inducing calcium release compared with controls. CONCLUSIONS: Peripheral blood samples are stable regarding RNA and DNA extraction and establishment of lymphoblastoid B cell lines after transportation at ambient temperature over large distances by ordinary mail. Functional tests on B cells harboring the newly identified amino acid substitutions indicate that they alter intracellular Ca2+ homeostasis and are most likely causative of malignant hyperthermia. (Anesth Analg 2010;111:185-90)}}, author = {{Vukcevic, Mirko and Broman, Marcus and Islander, Gunilla and Bodelsson, Mikael and Ranklev Twetman, Eva and Mueller, Clemens R. and Treves, Susan}}, issn = {{1526-7598}}, language = {{eng}}, number = {{1}}, pages = {{185--190}}, publisher = {{Lippincott Williams & Wilkins}}, series = {{Anesthesia and Analgesia}}, title = {{Functional Properties of RYR1 Mutations Identified in Swedish Patients with Malignant Hyperthermia and Central Core Disease}}, url = {{http://dx.doi.org/10.1213/ANE.0b013e3181cbd815}}, doi = {{10.1213/ANE.0b013e3181cbd815}}, volume = {{111}}, year = {{2010}}, }