Evaluation of methods for estimating infarct size by myosin LC2: comparison with cardiac enzymes

The relationship between histologically determined infarct size and release or peak levels of circulating cardiac enzymes and myosin light chain 2 (LC2) was studied. Myocardial infarction was produced by ligating the left anterior descending coronary artery in 18 conscious closed-chest dogs. Creatine phosphokinase (CPK), cytosolic and mitochondrial isozymes of aspartate transaminase (sAST and mAST) in the plasma, and LC2 in the serum were measured serially until 10 days after infarction, when infarct size was determined histologically [range 4.0-38.8% of the left ventricular weight (%LV)]. Infarct size correlated most closely with LC2 release (r = 0.82, P less than 0.001) and less closely with peak sAST (r = 0.59, P less than 0.01), peak mAST (r = 0.49, P less than 0.05), peak CPK (r = 0.22), and CPK release (r = 0.14). The correlation between infarct size and CPK release was improved by limiting the analysis to the dogs with infarct size of less than 20% LV (n = 11, r = 0.53, P less than 0.1). Because, among cardiac enzymes and LC2, CPK activity decayed most rapidly in the lymph fluid when incubated in vitro, degeneration of CPK in the lymph stream may contribute to the nonlinear relationship between infarct size and CPK release.

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In vitro chamber specification during embryonic stem cell cardiogenesis. Expression of the ventricular myosin light chain-2 gene is independent of heart tube formation.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(19)74594-3 ◽

1993 ◽

Vol 268 (33) ◽

pp. 25244-25252

Author(s):

W C Miller-Hance ◽

M LaCorbiere ◽

S J Fuller ◽

S M Evans ◽

G Lyons ◽

...

Keyword(s):

Stem Cell ◽

Light Chain ◽

Myosin Light Chain ◽

Embryonic Stem ◽

Tube Formation ◽

Heart Tube ◽

Myosin Light Chain 2 ◽

Ventricular Myosin ◽

Heart Tube Formation

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Serotonin increases L-type Ca2+ current and SR Ca2+ content through 5-HT4 receptors in failing rat ventricular cardiomyocytes

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.01375.2006 ◽

2007 ◽

Vol 293 (4) ◽

pp. H2367-H2376 ◽

Cited By ~ 16

Author(s):

Jon Arne Kro Birkeland ◽

Fredrik Swift ◽

Nils Tovsrud ◽

Ulla Enger ◽

Per Kristian Lunde ◽

...

Keyword(s):

Myosin Light Chain ◽

Troponin I ◽

Contractile Function ◽

Left Ventricular ◽

Inotropic Response ◽

Receptor Stimulation ◽

Ventricular Cardiomyocytes ◽

Myosin Light Chain 2 ◽

Intracellular Ca ◽

Male Wistar Rats

Rats with congestive heart failure (CHF) develop ventricular inotropic responsiveness to serotonin (5-HT), mediated through 5-HT2A and 5-HT4 receptors. Human ventricle is similarly responsive to 5-HT through 5-HT4 receptors. We studied isolated ventricular cardiomyocytes to clarify the effects of 5-HT on intracellular Ca2+ handling. Left-ventricular cardiomyocytes were isolated from male Wistar rats 6 wk after induction of postinfarction CHF. Contractile function and Ca2+ transients were measured in field-stimulated cardiomyocytes, and L-type Ca2+ current ( ICa,L) and sarcoplasmic reticulum (SR) Ca2+ content were measured in voltage-clamped cells. Protein phosphorylation was measured by Western blotting or phosphoprotein gel staining. 5-HT4- and 5-HT2A-receptor stimulation induced a positive inotropic response of 33 and 18% (both P < 0.05) and also increased the Ca2+ transient (44 and 6%, respectively; both P < 0.05). ICa,L and SR Ca2+ content increased only after 5-HT4-receptor stimulation (57 and 65%; both P < 0.05). Phospholamban serine16 (PLB-Ser16) and troponin I phosphorylation increased by 26 and 13% after 5-HT4-receptor stimulation ( P < 0.05). 5-HT2A-receptor stimulation increased the action potential duration and did not significantly change the phosphorylation of PLB-Ser16 or troponin I, but it increased myosin light chain 2 (MLC2) phosphorylation. In conclusion, the positive inotropic response to 5-HT4 stimulation results from increased ICa,L and increased phosphorylation of PLB-Ser16, which increases the SR Ca2+ content. 5-HT4 stimulation is thus, like β-adrenoceptor stimulation, possibly energetically unfavorable in CHF. 5-HT2A-receptor stimulation, previously studied in acute CHF, induces a positive inotropic response also in chronic CHF, probably mediated by MLC2 phosphorylation.

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The Wnt Inhibitor sFRP2 Enhances Mesenchymal Stem Cell Engraftment, Granulation Tissue Formation and Myocardial Repair

Blood ◽

10.1182/blood.v112.11.1365.1365 ◽

2008 ◽

Vol 112 (11) ◽

pp. 1365-1365

Author(s):

Maria Paula Alfaro ◽

Matthew Pagni ◽

Alicia Vincent ◽

Michael F. Hill ◽

Ethan Lee ◽

...

Keyword(s):

Infarct Size ◽

Granulation Tissue ◽

Myocardial Infarct ◽

Left Ventricular ◽

Human Mscs ◽

Myocardial Repair ◽

Acute Myocardial Infarct ◽

Intramyocardial Injection

Abstract Cell-based therapies using bone marrow-derived mesenchymal stem cells (MSCs) for organ regeneration are being pursued for cardiac disease, orthopedic injuries and biomaterial fabrication. The molecular pathways that regulate MSC-mediated regeneration or enhance their therapeutic efficacy are, however, poorly understood. In an attempt to elucidate a way to strengthen the regenerative potential of MSCs, preliminary studies in our lab were performed comparing MSCs isolated from wildtype and regenerative mouse strains. The MRL/MpJ mouse has been described as a “super healer” mouse that is able to repair soft tissue with minimal scaring. MSCs were isolated from the MRL/MpJ mouse (MRL-MSCs) and from C57/Bl6 mice (WT-MSCs) and their differing qualities assessed. Compared to WT-MSCs, MRL-MSCs demonstrated increased proliferation in vitro. We utilized a Poly-vinyl alcohol (PVA) sponge model of repair stimulation to assess their capacity to generate wound repair tissue. We observed that the MRL-MSCs demonstrated increased in vivo engraftment, experimental granulation tissue reconstitution, and tissue vascularity. The MRL-MSCs also reduced infarct size and improved cardiac function as compared to WT-MSCs in a murine acute myocardial infarct model. Genomic and functional analyses indicated a downregulation of the canonical Wnt pathway in MRL-MSCs characterized specifically by upregulation of secreted frizzled related proteins (sFRPs). In vitro proliferation studies confirmed that recombinant sFRP2 mediated enhanced proliferation of both mouse and human MSCs. Based on these observations, we hypothesized that sFRP2 served an important role in MSC-mediated repair and regeneration. We generated WT-MSCs overexpressing sFRP2 (sFRP2-MSCs) by retroviral transduction to test this hypothesis. sFRP2-MSCs maintained their ability for multilineage differentiation in vitro and proliferated faster than the vector only control MSCs (GFP-MSCs). When implanted in vivo in the PVA sponge model, the sFRP2-MSCs recapitulated the MRL phenotype by mediating greater, more vascularized granulation tissue. Moreover, periinfarct intramyocardial injection of sFRP2-MSCs resulted in reduced infarct size, favorable remodeling and better preserved left ventricular function following acute myocardial infarct in mice. These findings implicate sFRP2 as a key molecule for the biogenesis of a superior regenerative phenotype of MSCs.

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Theoretical analysis on the relationship between left ventricular energetic efficiency and acute infarct size

IET Systems Biology ◽

10.1049/iet-syb.2011.0080 ◽

2013 ◽

Vol 7 (3) ◽

pp. 74-78 ◽

Cited By ~ 1

Author(s):

Takao Shimayoshi ◽

Yuki Hasegawa ◽

Mitsuharu Mishima ◽

Tetsuya Matsuda

Keyword(s):

Theoretical Analysis ◽

Infarct Size ◽

Left Ventricular ◽

Energetic Efficiency ◽

Acute Infarct ◽

The Relationship

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Role of protein kinase C in the phosphorylation of cardiac myosin light chain 2

Biochemical Journal ◽

10.1042/bj2940401 ◽

1993 ◽

Vol 294 (2) ◽

pp. 401-406 ◽

Cited By ~ 65

Author(s):

R C Venema ◽

R L Raynor ◽

T A Noland ◽

J F Kuo

Keyword(s):

Protein Kinase ◽

Light Chain ◽

Myosin Light Chain ◽

Heart Cells ◽

Adult Rat ◽

Kinase C ◽

Myosin Light Chain 2

The role of protein kinase C (PKC) in the phosphorylation of myosin light chain 2 (MLC2) in adult rat heart cells has been investigated. PKC-mediated phosphorylation of MLC2 in adult rat cardiac myofibrils in vitro occurs with a stoichiometry (0.7 mol of phosphate/mol of protein) similar to that mediated by myosin light chain kinase (MLCK). Two-dimensional tryptic phosphopeptide mapping of MLC2 following phosphorylation by PKC or MLCK in vitro yields the same major phosphopeptides for each protein kinase. These sites are also 32P-labelled in situ when isolated cardiomyocytes are incubated with [32P]P(i). 32P labelling of MLC2 in cardiomyocytes is increased by 5-fold in 10 min upon incubation with the phosphatase inhibitor calyculin A, demonstrating the existence of a rapidly turning over component of MLC2 phosphorylation in these cells. 32P label is completely removed from MLC2 when myocytes are exposed to 2,3-butanedione monoxime, an inhibitor of cardiac contraction known to desensitize the myofilaments to activation by Ca2+. 32P labelling of MLC2 is also decreased by 50-100% following exposure to the PKC-selective inhibitors calphostin C and chelerythrine, suggesting that PKC, and not MLCK, is primarily responsible for incorporation of rapidly turning over phosphate into MLC2 in situ. Taken together, these data implicate PKC in the phosphorylation of MLC2 in heart cells and support the hypothesis that phosphorylation of cardiac MLC2 has a role in determining myofibrillar Ca2+ sensitivity.

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Myocardial functional correlates of cardiac myosin light chain 2 phosphorylation

Journal of Applied Physiology ◽

10.1152/jappl.1990.68.6.2426 ◽

1990 ◽

Vol 68 (6) ◽

pp. 2426-2433 ◽

Cited By ~ 11

Author(s):

D. P. Fitzsimons ◽

P. W. Bodell ◽

K. M. Baldwin

Keyword(s):

Heart Rate ◽

Light Chain ◽

Myosin Light Chain ◽

Diastolic Pressure ◽

Left Ventricular ◽

Exercise Heart Rate ◽

Cardiac Myosin ◽

Myosin Light Chain 2 ◽

Pressure Development

In vitro and in situ studies have proposed a potentiation of submaximal force production after myosin light chain 2 (P-light chain) phosphorylation in mammalian striated muscle. The purpose of this study was to ascertain the relationship between the augmentation in left ventricular pressure development and cardiac myosin P-light chain phosphorylation at different times during and after submaximal treadmill exercise involving adult female Sprague-Dawley rats. In vivo hemodynamic measurements were monitored with an indwelling high-fidelity solid-state pressure transducer. Exercise heart rate, peak left ventricular (LV) pressure, and rate of LV pressure development/relaxation (LV +/- dP/dt) were significantly elevated compared with a normal sedentary group (P less than 0.001). Peak LV pressure remained significantly elevated throughout 20 min of postexercise recovery (P less than 0.01), and heart rate, LV end-diastolic pressure, and LV +/- dP/dt returned rapidly to preexercise values. Corresponding to these in vivo hemodynamic changes, increased levels of P-light chain phosphorylation were observed during both exercise (16%, P less than 0.01) and subsequent recovery periods (14%, P less than 0.02) compared with the NC group. A quasi-temporal relationship was observed between postexercise peak LV pressure potentiation and P-light chain phosphorylation. These results demonstrate that cardiac myosin P-light chain phosphorylation is associated, in part, with the augmentation of peak LV pressure observed during both exercise and recovery.

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Phosphorylation of rodent cardiac myosin light chain 2: effects of exercise

Journal of Applied Physiology ◽

10.1152/jappl.1989.67.6.2447 ◽

1989 ◽

Vol 67 (6) ◽

pp. 2447-2453 ◽

Cited By ~ 23

Author(s):

D. P. Fitzsimons ◽

P. W. Bodell ◽

K. M. Baldwin

Keyword(s):

Phase Ii ◽

Light Chain ◽

Myosin Light Chain ◽

Left Ventricular Pressure ◽

Left Ventricular ◽

Ventricular Pressure ◽

Pharmacological Intervention ◽

Cardiac Myosin ◽

Myosin Light Chain 2 ◽

Pressure Development

The purpose of this study was to ascertain whether the degree of cardiac myosin light chain 2 (P-light chain) phosphorylation occurs as a function of changes in cardiovascular functional state as induced by 1) treadmill exercise (20-27 m/min, 0% grade for 20, 30, 45 min) (phase I) and 2) pharmacological intervention (phase II) in adult female Sprague-Dawley rats. It was hypothesized that cardiac myosin phosphorylation is regulated in accordance with time-dependent sustained elevations in myocardial work demands requiring alterations in either heart rate or left ventricular pressure development. Exercise heart rates (HR) and double products (HR x DP) were equivalent among the three exercise groups and were significantly elevated in comparison with the normal-rest (NR) group (P less than 0.05). In phase II, isoproterenol elicited higher HR, although the atenolol group exhibited a marked reduction in HR, mean arterial pressure, and double product relative to NR (P less than 0.05). Percent myosin P-light chain phosphorylation exhibited both a HR- and a work load-dependent modulation in P-light chain levels (-9% to +23% change) in the two phases of the study (P less than 0.05). These data are consistent with the view that the above responses are associated with modulations in intracellular calcium concentrations commensurate with the alterations in HR and left ventricular pressure. Also, elevations in P-light chain phosphorylation could serve to augment left ventricular pressure development under these functional states.

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p21-activated kinase improves cardiac contractility during ischemia-reperfusion concomitant with changes in troponin-T and myosin light chain 2 phosphorylation

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00612.2011 ◽

2012 ◽

Vol 302 (1) ◽

pp. H224-H230 ◽

Cited By ~ 17

Author(s):

Michelle M. Monasky ◽

Domenico M. Taglieri ◽

Bindiya G. Patel ◽

Jonathan Chernoff ◽

Beata M. Wolska ◽

...

Keyword(s):

Light Chain ◽

Myosin Light Chain ◽

Diastolic Pressure ◽

Troponin T ◽

Ex Vivo ◽

Ischemia Reperfusion ◽

Cardiac Contractility ◽

Left Ventricular ◽

Myosin Light Chain 2 ◽

P21 Activated Kinase

p21-activated kinase 1 (Pak1) is a serine/threonine kinase that activates protein phosphatase 2a, resulting in the dephosphorylation of cardiac proteins and increased myofilament Ca2+ sensitivity. Emerging evidence indirectly indicates a role for Pak1 in ischemia-reperfusion (I/R), but direct evidence is lacking. We hypothesize that activation of the Pak1 signaling pathway is a cardioprotective mechanism that prevents or reverses the detrimental effects of ischemic injury by inducing posttranslational modifications in myofilament proteins that ultimately improve cardiac contractility following ischemic insult. In the present study, we subjected ex vivo hearts from wild-type (WT) and Pak1-knockout (KO) mice to 20 min of global cardiac ischemia followed by 30 min of reperfusion. In the absence of Pak1, there was an exacerbation of the increased end-diastolic pressure and reduced left ventricular developed pressure occurring after I/R injury. ProQ analysis revealed an increase in troponin-T phosphorylation at baseline in Pak1-KO hearts compared with WT. Significantly decreased myosin light chain 2 (MLC2) phosphorylation in Pak1-KO hearts compared with WT after I/R injury was confirmed by Western immunoblotting. These data indicate that Pak1-KO hearts have reduced recovery of myocardial performance after global I/R injury concomitant with changes in troponin-T and MLC2 phosphorylation. Finally, a protein-protein association between Pak1 and MLC2, and Pak1 and troponin-T, was determined by coimmunoprecipitation. Thus, results of our study provide a basis for targeting a novel pathway, including Pak1, in the therapies for patients with ischemic events.

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Abstract 2826: Microvascular Obstruction on Cardiac MRI is Associated with an Early Peak Troponin

Circulation ◽

10.1161/circ.118.suppl_18.s_782 ◽

2008 ◽

Vol 118 (suppl_18) ◽

Author(s):

James C Weaver ◽

David Rees ◽

Ananth M Prasan ◽

Jane A McCrohon

Keyword(s):

Infarct Size ◽

Functional Recovery ◽

Cardiac Mri ◽

Microvascular Obstruction ◽

Area Under The Curve ◽

Primary Angioplasty ◽

Left Ventricular ◽

End Diastolic Volume ◽

Early Peak ◽

The Relationship

Background : Microvascular obstruction (MVO) post STEMI, as defined on cardiac MRI (CMR), reflects slow flow in the microvasculature. It has been postulated that MVO may result in impaired washout of troponin (Tn) and therefore a later peak. Aim : To use CMR to explore the relationship between MVO and the pattern of TnI release after STEMI. Methods: After successful primary angioplasty serial TnI measurements were taken at 6, 14 and 24 hours. Gadolinium (Gd) enhanced CMR was performed at 4.7±2 days and again at 191±17 days. Infarction was defined as a hyperenhanced region 10 minutes after intravenous 0.1mmol/kg Gd and MVO defined as a hypoenhanced region in the centre of this infarcted segment. A 17 segment model was used for wall motion assessment and functional recovery. Results: 26 patients, age 57 ±11 yrs, were included in the analysis. On early scanning infarct size was 22.1±13% and MVO was 4.3± 4% of LV mass. The 81% of patients with MVO had an earlier peak TnI than those without MVO (p=0.005). Patients with severe MVO, defined as the highest quartile of MVO extent, all had the earliest peak of 6hrs. Those with MVO had higher TnI at 6hrs (p=0.002), 14hrs (p=0.027) and area under the curve (AUC) (p=0.014). TnI levels at all time points in the first 24 hours had a very strong correlation with MVO extent (r=0.80 – 0.88, p<0.001). This was stronger than between TnI and infarct size (r=0.68 – 0.78, p<0.001). Equally an early peak TnI (6hrs compared with 14hrs) was associated with a higher peak TnI level (p=0.013) and AUC (p=0.019). In the 17 patients with follow up scans infarct involution was 19.8±13% and the relationship between TnI level and infarct size did not change significantly. As MVO increased there was less functional recovery (r=−0.49, p=0.046) and increased left ventricular end diastolic volume (r=0.512, p=0.036) at 6 months. Of the 5 patients who developed dyskinetic segments 3 had severe MVO and 4 an early peak TnI. Conclusion: MVO presence and extent is associated with an early peak in TnI after reperfused STEMI and more extensive myonecrosis as evidenced by cumulative (AUC) TnI release. These findings favour more complete myonecrosis within the MVO region, rather than impaired washout, as the predominant pathological basis for the relationship between MVO and TnI kinetics.

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