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Metabolic Syndrome and SIRT1 Mutation Impair Ca 2+ Channels in Coronary Smooth Muscle Cells of Ossabaw Miniature Swine
Author(s) -
Thamba Aish,
Reed John,
Strobel John,
Byrd James,
Alloosh Mouhamad,
Coutts Alex,
Sturek Michael
Publication year - 2022
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2022.36.s1.r6137
Subject(s) - caffeine , medicine , endocrinology , endoplasmic reticulum , mutation , metabolic syndrome , voltage dependent calcium channel , calcium , chemistry , biology , obesity , genetics , gene
Background Normal metabolic health and sirtuin1 (SIRT1) by its deacetylase activity maintain voltage‐gated Ca 2+ channel (VGCC) function that promotes gene expression to maintain contractile coronary smooth muscle (CSM) phenotype. MetS impairs VGCC function and sarcoplasmic reticulum (SR) Ca 2+ release, contributing to dedifferentiation of CSM to proliferative and osteogenic phenotypes. Hypothesis MetS and impaired SIRT1 will impair VGCC and SR Ca 2+ release channels in CSM. Methods Using CRISPR/Cas9 methodology a point mutation (SIRT1 L100P ) was made in Ossabaw miniature swine to mimic the naturally occurring mutation in humans and decrease SIRT1 activity, thereby resulting in hyperacetylation (Ac) of Ca 2+ transporters and impaired function. Four groups of pigs were used to analyze genotype and diet interactions: wild type lean, SIRT1 lean, wild type MetS, SIRT1 MetS. Pigs were age 4 months at the start and fed normal chow (lean) or atherogenic diet (MetS) for 7 months. CSM cells were enzymatically dispersed and Ca 2+ measured with fura‐2. Depolarization with 80 mM K+ assessed Ca 2+ influx through VGCC and the peak Ca 2+ response to 5 mM caffeine to open SR Ca 2+ release channels assessed the caffeine‐sensitive SR Ca 2+ store. Results Two‐way ANOVA showed SIRT1 mutation (p=0.02) and MetS diet (p<0.0001) significantly decreased VGCC function independently, but not additively or synergistically. Interaction of SIRT1 genotype and MetS diet was significant (p<0.0001). No significant effect of genotype or diet was observed on SR calcium store release. Conclusion SIRT1 L100P mutation is likely to contribute to coronary atherosclerosis and SIRT1 activators may be effective therapies.