Aldrich Kennedy, Velez-Irizarry Deborah, Fenger Clara, Schott Melissa, Valberg Stephanie J
Mary Anne McPhail Equine Performance Center, Large Animal Clinical Sciences, College of Veterinary Medicine, Michigan State University, East Lansing, MI, United States of America.
Equine Integrated Medicine, PLC, Lexington, KY, United States of America.
PLoS One. 2021 Feb 10;16(2):e0244556. doi: 10.1371/journal.pone.0244556. eCollection 2021.
Recurrent exertional rhabdomyolysis (RER) is a chronic muscle disorder of unknown etiology in racehorses. A potential role of intramuscular calcium (Ca2+) dysregulation in RER has led to the use of dantrolene to prevent episodes of rhabdomyolysis. We examined differentially expressed proteins (DEP) and gene transcripts (DEG) in gluteal muscle of Thoroughbred race-trained mares after exercise among three groups of 5 horses each; 1) horses susceptible to, but not currently experiencing rhabdomyolysis, 2) healthy horses with no history of RER (control), 3) RER-susceptible horses treated with dantrolene pre-exercise (RER-D). Tandem mass tag LC/MS/MS quantitative proteomics and RNA-seq analysis (FDR <0.05) was followed by gene ontology (GO) and semantic similarity of enrichment terms. Of the 375 proteins expressed, 125 were DEP in RER-susceptible versus control, with 52 ↑DEP mainly involving Ca2+ regulation (N = 11) (e.g. RYR1, calmodulin, calsequestrin, calpain), protein degradation (N = 6), antioxidants (N = 4), plasma membranes (N = 3), glyco(geno)lysis (N = 3) and 21 DEP being blood-borne. ↓DEP (N = 73) were largely mitochondrial (N = 45) impacting the electron transport system (28), enzymes (6), heat shock proteins (4), and contractile proteins (12) including Ca2+ binding proteins. There were 812 DEG in RER-susceptible versus control involving the electron transfer system, the mitochondrial transcription/translational response and notably the pro-apoptotic Ca2+-activated mitochondrial membrane transition pore (SLC25A27, BAX, ATP5 subunits). Upregulated mitochondrial DEG frequently had downregulation of their encoded DEP with semantic similarities highlighting signaling mechanisms regulating mitochondrial protein translation. RER-susceptible horses treated with dantrolene, which slows sarcoplasmic reticulum Ca2+ release, showed no DEG compared to control horses. We conclude that RER-susceptibility is associated with alterations in proteins, genes and pathways impacting myoplasmic Ca2+ regulation, the mitochondrion and protein degradation with opposing effects on mitochondrial transcriptional/translational responses and mitochondrial protein content. RER could potentially arise from excessive sarcoplasmic reticulum Ca2+ release and subsequent mitochondrial buffering of excessive myoplasmic Ca2+.
复发性运动性横纹肌溶解症（RER）是一种病因不明的赛马慢性肌肉疾病。肌内钙（Ca2+）调节异常在RER中的潜在作用促使人们使用丹曲林来预防横纹肌溶解症的发作。我们对三组各5匹纯种赛马训练母马运动后臀肌中的差异表达蛋白（DEP）和基因转录本（DEG）进行了检测；1）易患但目前未发生横纹肌溶解症的马匹，2）无RER病史的健康马匹（对照组），3）运动前用丹曲林治疗的RER易感马匹（RER-D）。采用串联质量标签液相色谱/串联质谱定量蛋白质组学和RNA测序分析（FDR<0.05），随后进行基因本体论（GO）和富集术语的语义相似性分析。在表达的375种蛋白质中，125种在RER易感组与对照组之间存在差异表达，其中52种上调的DEP主要涉及Ca2+调节（N=11）（如RYR1、钙调蛋白、肌浆网钙结合蛋白、钙蛋白酶）、蛋白质降解（N=6）、抗氧化剂（N=4）、质膜（N=3）、糖（原）酵解（N=3），21种DEP存在于血液中。下调的DEP（N=73）主要存在于线粒体中（N=45），影响电子传递系统（28）、酶（6）、热休克蛋白（4）和收缩蛋白（12），包括Ca2+结合蛋白。RER易感组与对照组之间有812个DEG，涉及电子传递系统、线粒体转录/翻译反应，尤其是促凋亡的Ca2+激活线粒体膜通透性转换孔（SLC25A27、BAX、ATP5亚基）。上调的线粒体DEG其编码的DEP经常下调，语义相似性突出了调节线粒体蛋白质翻译的信号机制。用丹曲林治疗的RER易感马匹，可减缓肌浆网Ca2+释放，与对照组马匹相比未显示出DEG。我们得出结论，RER易感性与影响肌浆Ca2+调节、线粒体和蛋白质降解的蛋白质、基因和途径的改变有关，对线粒体转录/翻译反应和线粒体蛋白质含量产生相反影响。RER可能源于肌浆网Ca2+过度释放以及随后线粒体对过多肌浆Ca2+的缓冲作用。
