RETRACTED ARTICLE: Functional analysis of newly identified RYR1 variants in patients susceptible to malignant hypertherm
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ORIGINAL ARTICLE
Functional analysis of newly identified RYR1 variants in patients susceptible to malignant hyperthermia Yuko Noda1 · Toshimichi Yasuda1 · Rieko Kanzaki1 · Hirotsugu Miyoshi2 · Keiko Mukaida3 · Satoshi Kamiya1 · Atsushi Morio1 · Hiroshi Hamada1 · Masashi Kawamoto4 · Yasuo M. Tsutsumi1 Received: 2 February 2020 / Accepted: 23 May 2020 © Japanese Society of Anesthesiologists 2020
Abstract Purpose This study aimed to evaluate whether the three ryanodine receptor type 1 (RYR1) variants (p.Ser2345Thr, p.Ser2345Arg, and p.Lys3367Arg) which we identified in Japanese malignant hyperthermia (MH) patients with a clinical grading scale rank of 6 were causative for MH. Methods We prepared human embryonic kidney (HEK)-293 cells transfected with wild-type RYR1 or one of the RYR1 variants, along with myotubes cultured from muscle pieces. Calcium kinetics were examined by calculating the 340/380-nm ratio under various caffeine and 4-chloro-m-cresol (4CmC) concentrations with the ratiometric dye Fura-2 AM. Half-maximal effective concentration ( EC50) values were calculated from dose–response curves. Statistical analysis was based on one-way analysis of variance with a Dunnett’s multiple comparison test, using a P value G 51 0
Process II: Muscle breakdown Process III: Respiratory acidosis Process IV: Temperature increase Process V: Cardiac involvement Other indicators
15
CK > 10,000 IU
15
PETCO2 > 55 mmHg
p.Ser2345Arg c.7035C > A 78 15 Generalized muscular rigidity 15 CK > 10,000 IU without succunylcholin 15 PETCO2 > 55 mmHg
15
Rapid increase in temperature Sinus tachycardia
15
Rapid reversal of acidosis with iv dantrolene
3 5
3
K+ > 6 mEq/L
15
PETCO2 > 55 mmHg
15
3
Rapid increase in temperature Sinus tachycardia
3
Rapid increase in temperature Sinus tachycardia
15
Atrial pH G) from Patient 3. Three prediction tools, namely Mutation Taster, PolyPhen-2, and SIFT, were employed prior to functional analyses to determine whether each variant had the potential to cause MH, and the results are shown in Table 2. p.Ser2345Thr and p.Ser2345Arg were both predicted to be “disease causing” by Mutation Taster and “probably damaging” by Polyphen-2, but p.Lys3367Arg was only predicted to be “disease causing” by Mutation Taster.
CICR rate testing
Table 2 Results of functional effects of each variant using prediction tools Mutation taster
PolyPhen-2
SIFT
p.Ser2345Thr
Disease causing
p.Ser2345Arg
Disease causing
p.Lys3367Arg
Disease causing
Possibly damag- Not scored ing Possibly damag- Not scored ing Benign Not scored
p.Ser2345Thr and p.Ser2345Arg were both predicted to be “disease causing” by Mutation Taster and “possibly damaging” by Polyphen-2, but p.Lys3367Arg was only predicted to be “disease causing” by Mutation Taster
Journal of Anesthesia
Fig. 1 Western blot image confirmed the presence of RYR1 protein in HEK293 cells transfected with wild-type or mutant RYR1
Responses of transfected HEK‑293 cells to caffeine Immunoblots of HEK293 cells revealed the expression of RyR1 in both
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