Copeland O'neal, Messer Andrew, Jabbour Andrew, Poggesi Corrado, Prasad Sanjay, Marston Steven
Imperial College London, London, United Kingdom.
Royal Brompton Hospital, and Imperial College London, London, United Kingdom.
Front Physiol. 2020 Mar 19;11:241. doi: 10.3389/fphys.2020.00241. eCollection 2020.
In previous studies of septal heart muscle from HCM patients with hypertrophic obstructive cardiomyopathy (HOCM, LVOT gradient 50-120 mmHg) we found that the level of phosphorylation of troponin I (TnI) and myosin binding protein C (MyBP-C) was extremely low yet samples from hearts with HCM or DCM mutations that did not have pressure overload were similar to donor heart controls. We therefore investigated heart muscle samples taken from patients undergoing valve replacement for aortic stenosis, since they have pressure overload that is unrelated to inherited cardiomyopathy. Thirteen muscle samples from septum and from free wall were analyzed (LVOT gradients 30-100 mmHg) The levels of TnI and MyBP-C phosphorylation were determined in muscle myofibrils by separating phosphospecies using phosphate affinity SDS-PAGE and detecting with TnI and MyBP-C specific antibodies. TnI was predominantly monophosphorylated and total phosphorylation was 0.85 ± 0.03 molsPi/mol TnI. This phosphorylation level was significantly different ( < 0.0001) from both donor heart TnI (1.6 ± 0.06 molsPi/mol TnI) and HOCM heart TnI (0.19 ± 0.04 molsPi/mol TnI). MyBP-C is phosphorylated at up to four sites. In donor heart the 4P and 3P species predominate but in the pressure overload samples the 4P species was much reduced and 3P and 1P species predominated. Total phosphorylation was 2.0 ± 0.2 molsPi/mol MyBP-C ( = 8) compared with 3.4 ± 0.07 ( = 21) in donor heart and 1.1 ± 0.1 ( = 10) in HOCM heart. We conclude that pressure overload may be associated with substantial dephosphorylation of troponin I and MyBP-C.
在先前对肥厚性梗阻性心肌病（HOCM，左心室流出道梯度为50 - 120 mmHg）的肥厚型心肌病（HCM）患者的间隔心肌研究中，我们发现肌钙蛋白I（TnI）和肌球蛋白结合蛋白C（MyBP - C）的磷酸化水平极低，但来自没有压力超负荷的HCM或DCM突变心脏的样本与供体心脏对照相似。因此，我们研究了因主动脉瓣狭窄接受瓣膜置换术患者的心肌样本，因为他们存在与遗传性心肌病无关的压力超负荷。分析了13个来自间隔和游离壁的心肌样本（左心室流出道梯度为30 - 100 mmHg）。通过使用磷酸盐亲和SDS - PAGE分离磷酸化形式并用TnI和MyBP - C特异性抗体进行检测，测定肌原纤维中TnI和MyBP - C的磷酸化水平。TnI主要为单磷酸化，总磷酸化水平为0.85±0.03 molPi/mol TnI。该磷酸化水平与供体心脏TnI（1.6±0.06 molPi/mol TnI）和HOCM心脏TnI（0.19±0.04 molPi/mol TnI）均有显著差异（<0.0001）。MyBP - C最多在四个位点磷酸化。在供体心脏中，4P和3P形式占主导，但在压力超负荷样本中，4P形式大幅减少，3P和1P形式占主导。总磷酸化水平为2.0±0.2 molPi/mol MyBP - C（n = 8），相比之下，供体心脏为3.4±0.07（n = 21），HOCM心脏为1.1±0.1（n = 10）。我们得出结论，压力超负荷可能与肌钙蛋白I和MyBP - C的大量去磷酸化有关。
