A simplified method for identification of human cardiac myosin heavy-chain isoforms

2003 ◽  
Vol 37 (1) ◽  
pp. 27 ◽  
Author(s):  
Shengfu Piao ◽  
Fushun Yu ◽  
Michael J. Mihm ◽  
Peter J. Reiser ◽  
Patrick M. McCarthy ◽  
...  
Keyword(s):  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Myosin Heavy Chain Isoforms ◽  
Cardiac Myosin ◽  
Simplified Method

scholarly journals Electrophoretic separation and quantitation of cardiac myosin heavy chain isoforms in eight mammalian species

1998 ◽  
Vol 274 (3) ◽  
pp. H1048-H1053 ◽  
Author(s):  
Peter J. Reiser ◽  
William O. Kline
Keyword(s):  
Skeletal Muscle ◽  
Sample Preparation ◽  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Mammalian Species ◽  
Myosin Heavy Chain Isoforms ◽  
Glycerol Concentration ◽  
Cardiac Myosin ◽  
Homogenization Step

A protocol for sample preparation and gel electrophoresis is described that reliably results in the separation of the α- and β-isoforms of cardiac myosin heavy chain (MHC-α and MHC-β) in eight mammalian species. The protocol is based on a simple, nongradient denaturing gel. The magnitude of separation of MHC-α and MHC-β achieved with this protocol is sufficient for quantitative determination of the relative amounts of these two isoforms in mouse, rat, guinea pig, rabbit, canine, pig, baboon, and human myocardial samples. The sensitivity of the protocol is sufficient for the detection of MHC isoforms in samples at least as small as 1 μg. The glycerol concentration in the separating gel is an important factor for successfully separating MHC-α and MHC-β in myocardial samples from different species. The effect of sample load on MHC-α and MHC-β band resolution is illustrated. The results also indicate that inclusion of a homogenization step during sample preparation increases the amount of MHC detected on the gel for cardiac samples to a much greater extent than for skeletal muscle samples. Although the protocol described in this study is excellent for analyzing cardiac samples, it should be noted that the same protocol is not optimal for separating MHC isoforms expressed in skeletal muscle, as is illustrated.

scholarly journals Regulatory light chain phosphorylation augments length-dependent contraction in PTU-treated rats

2018 ◽  
Vol 151 (1) ◽  
pp. 66-76 ◽  
Author(s):  
Jason J. Breithaupt ◽  
Hannah C. Pulcastro ◽  
Peter O. Awinda ◽  
David C. DeWitt ◽  
Bertrand C.W. Tanner
Keyword(s):  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Light Chain ◽  
Myosin Heavy Chain Isoforms ◽  
Regulatory Light Chain ◽  
Cardiac Myosin ◽  
Myosin Heavy Chain Isoform ◽  
Cross Bridge ◽  
Detachment Rate ◽  
Length Dependent Activation

Force production by actin–myosin cross-bridges in cardiac muscle is regulated by thin-filament proteins and sarcomere length (SL) throughout the heartbeat. Prior work has shown that myosin regulatory light chain (RLC), which binds to the neck of myosin heavy chain, increases cardiac contractility when phosphorylated. We recently showed that cross-bridge kinetics slow with increasing SLs, and that RLC phosphorylation amplifies this effect, using skinned rat myocardial strips predominantly composed of the faster α-cardiac myosin heavy chain isoform. In the present study, to assess how RLC phosphorylation influences length-dependent myosin function as myosin motor speed varies, we used a propylthiouracil (PTU) diet to induce >95% expression of the slower β-myosin heavy chain isoform in rat cardiac ventricles. We measured the effect of RLC phosphorylation on Ca2+-activated isometric contraction and myosin cross-bridge kinetics (via stochastic length perturbation analysis) in skinned rat papillary muscle strips at 1.9- and 2.2-µm SL. Maximum tension and Ca2+ sensitivity increased with SL, and RLC phosphorylation augmented this response at 2.2-µm SL. Subtle increases in viscoelastic myocardial stiffness occurred with RLC phosphorylation at 2.2-µm SL, but not at 1.9-µm SL, thereby suggesting that RLC phosphorylation increases β-myosin heavy chain binding or stiffness at longer SLs. The cross-bridge detachment rate slowed as SL increased, providing a potential mechanism for prolonged cross-bridge attachment to augment length-dependent activation of contraction at longer SLs. Length-dependent slowing of β-myosin heavy chain detachment rate was not affected by RLC phosphorylation. Together with our previous studies, these data suggest that both α- and β-myosin heavy chain isoforms show a length-dependent activation response and prolonged myosin attachment as SL increases in rat myocardial strips, and that RLC phosphorylation augments length-dependent activation at longer SLs. In comparing cardiac isoforms, however, we found that β-myosin heavy chain consistently showed greater length-dependent sensitivity than α-myosin heavy chain. Our work suggests that RLC phosphorylation is a vital contributor to the regulation of myocardial contractility in both cardiac myosin heavy chain isoforms.

scholarly journals Identification of alpha- and beta-cardiac myosin heavy chain isoforms as major autoantigens in dilated cardiomyopathy.

Circulation ◽  
1992 ◽  
Vol 85 (5) ◽  
pp. 1734-1742 ◽  
Author(s):  
A L Caforio ◽  
M Grazzini ◽  
J M Mann ◽  
P J Keeling ◽  
G F Bottazzo ◽  
...  
Keyword(s):  
Dilated Cardiomyopathy ◽  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Myosin Heavy Chain Isoforms ◽  
Cardiac Myosin

scholarly journals Electrophoretic Mobility of Cardiac Myosin Heavy Chain Isoforms Revisited: Application of MALDI TOF/TOF Analysis

2011 ◽  
Vol 2011 ◽  
pp. 1-9 ◽  
Author(s):  
Petra Arnostova ◽  
Petr L. Jedelsky ◽  
Tomáš Soukup ◽  
Jitka Zurmanova
Keyword(s):  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Myosin Heavy Chain Isoforms ◽  
Cardiac Myosin ◽  
Thyroid Status ◽  
Mobility Rate ◽  
Sds Page ◽  
Myhc Isoforms ◽  
Lower Mobility ◽  
Altered Thyroid

The expression of two cardiac myosin heavy chain (MyHC) isoforms in response to the thyroid status was studied in left ventricles (LVs) of Lewis rats. Major MyHC isoform in euthyroid and hyperthyroid LVs had a higher mobility on SDS-PAGE, whereas hypothyroid LVs predominantly contained a MyHC isoform with a lower mobility corresponding to that of the control soleus muscle. By comparing the MyHC profiles obtained under altered thyroid states together with the control soleus, we concluded that MyHCαwas represented by the lower band with higher mobility and MyHCβby the upper band. The identity of these two bands in SDS-PAGE gels was confirmed by western blot and mass spectrometry. Thus, in contrast to the literature data, we found that the MyHCαpossessed a higher mobility rate than the MyHCβisoform. Our data highlighted the importance of the careful identification of the MyHCαand MyHCβisoforms analyzed by the SDS-PAGE.

Human cardiac myosin heavy chain isoforms in fetal and failing adult atria and ventricles

2001 ◽  
Vol 280 (4) ◽  
pp. H1814-H1820 ◽  
Author(s):  
Peter J. Reiser ◽  
Michael A. Portman ◽  
Xue-Han Ning ◽  
Christine Schomisch Moravec
Keyword(s):  
Heart Failure ◽  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Ischemic Cardiomyopathy ◽  
Myosin Heavy Chain Isoforms ◽  
Left Ventricular ◽  
Human Myocardium ◽  
Cardiac Myosin ◽  
Fetal Stage

The goal of this study was to test the hypothesis that the relative amounts of the cardiac myosin heavy chain (MHC) isoforms MHC-α and MHC-β change during development and transition to heart failure in the human myocardium. The relative amounts of MHC-α and MHC-β in ventricular and atrial samples from fetal (gestational days 47–110) and nonfailing and failing adult hearts were determined. The majority of the fetal right and left ventricular samples contained small relative amounts of MHC-α (mean < 5% of total MHC). There was a small significant decrease in the level of MHC-α in the ventricles between 7 and 12 wk of gestation. Fetal atria expressed predominantly MHC-α (mean > 95%), with MHC-β being detected in most samples. The majority of adult nonfailing right and left ventricular samples had detectable levels of MHC-α ranging from 1 to 10%. Failing right and left ventricles expressed a significantly lower level of MHC-α. MHC-α comprised ∼90% of the total MHC in adult nonfailing left atria, whereas the relative amount of MHC-α in the left atria of individuals with dilated or ischemic cardiomyopathy was ∼50%. The differences in MHC isoform composition between fetal and nonfailing adult atria and between fetal and nonfailing adult ventricles were not statistically significant. We concluded that the MHC isoform compositions of fetal human atria are the same as those of nonfailing adult atria and that the ventricular MHC isoform composition is different between adult nonfailing and failing hearts. Furthermore, the marked alteration in atrial MHC isoform composition, associated with cardiomyopathy, does not represent a regression to a pattern that is uniquely characteristic of the fetal stage.

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