Magnesium-calcium exchange in cardiac troponin C bound to cardiac troponin I

Understanding the process of Ca(2+)/Mg(2+)exchange during muscle excitation and relaxation is fundamental to elucidating the mechanism of Ca(2+)-regulated muscle contraction. During the resting phase, the C-domain of cardiac troponin C may be occupied by either Ca(2+)or Mg(2+). Here, complexes of recombinant cardiac troponin C(81-161) and the N terminus of cardiac troponin I, representing residues 33-80, were generated in the presence of saturating Mg(2+). Heteronuclear multi-dimensional nuclear magnetic resonance experiments were used to obtain backbone assignments of the Mg(2+)-loaded complex. In the presence of cardiac troponin I, the affinity of site IV for Mg(2+)is increased. Comparison of Mg(2+)and Ca(2+)-loaded complexes reveals that chemical shift differences are primarily localized to metal-binding sites III and IV, defining positions within these sites that have distinct Ca(2+)/Mg(2+)conformations. The observed transition from the Mg(2+)-loaded to Ca(2+)-loaded form demonstrates that sites III and IV fill simultaneously with Ca(2+)displacing Mg(2+). However, even in the absence of excess Ca(2+), Mg(2+)does not readily displace Ca(2+)in the isolated binary complex. Thus, the Mg(2+)-loaded conformer may only represent a small fraction of the total cardiac troponin C found in the sarcomere.     

Exercise-induced release of troponin

It is well established that regular physical activity reduces cardiovascular disease risk; however, numerous studies have demonstrated postexercise elevations in cardiac troponin (cTn), indicative of cardiac injury in apparently healthy individuals. The prevalence of these findings in different exercise settings and population groups, as well as potential underlying mechanisms and clinical significance of exercise-induced cTn release are not yet quite determined. The present review will discuss the cTn response to exercise in light of developing cTn assays and the correlation between postexercise cTn release and cardiac function. Additionally, recent data regarding the potential link between strenuous endurance exercise and its relationship with unfavorable cardiac effects in athletes, as well as the management of patients presenting at emergency care after sport events will be briefly reviewed.     

High-sensitive cardiac troponin T

Cardiac troponin is the preferred biomarker for the diagnosis of acute myocardial infarction (AMI). The recent development of a high-sensitive cardiac troponin T (hs-cTnT) assay permits detection of very low levels of cTnT. Using the hs-cTnT assay improves the overall diagnostic accuracy in patients with suspected AMI, while a negative result also has a high negative predictive value. The gain in sensitivity may be particularly important in patients with a short duration from symptom onset to admission. Measurement of cardiac troponin T with the hs-cTnT assay may provide strong prognostic information in patients with acute coronary syndromes, stable coronary artery disease, heart failure and even in the general population; however, increased sensitivity comes at a cost of decreased specificity. Serial testing, as well as clinical context and co-existing diseases, are likely to become increasingly important for the interpretation of hs-cTnT assay results.     

心脏肌钙蛋白I在急性心肌梗死中的临床意义

目的：探讨心脏肌钙蛋白I（cTnI)在急性心肌梗死（AMI）中的临床意义。方法：60例AMI,其中Q波型AMI42例,非Q波型AMI18例,分别于入院即刻、24小时、72小时、7天抽血,测定cTnI、CK－MB,随访半年。结果：（1）cTnI阳性率在AMI即刻、24小时、72小时、7天的Q波型AMI和非Q波型AMI依次分别为23．8％,100％,90．48％,26．19％;11．11％,77．78％,55．56％,16．61％。Q波型AMI的cTnI水平明显依次高于CK－MB的11．9％、90．48％、11．97％、0％;（2）第7天持续阳性的14例中,4例死于心脏骤停,5例发生心衰,5例发生心绞痛。结论：（1）AMI后24小时cTnI阳性率高于CK－MB;（2）cTnI阳性持续预后不佳。     

The cardiac-specific N-terminal region of troponin I positions the regulatory domain of troponin C

The cardiac isoform of troponin I (cTnI) has a unique 31-residue N-terminal region that binds cardiac troponin C (cTnC) to increase the calcium sensitivity of the sarcomere. The interaction can be abolished by cTnI phosphorylation at Ser22 and Ser23, an important mechanism for regulating cardiac contractility. cTnC contains two EF–hand domains (the N and C domain of cTnC, cNTnC and cCTnC) connected by a flexible linker. Calcium binding to either domain favors an “open” conformation, exposing a large hydrophobic surface that is stabilized by target binding, cTnI[148–158] for cNTnC and cTnI[39–60] for cCTnC. We used multinuclear multidimensional solution NMR spectroscopy to study cTnI[1–73] in complex with cTnC. cTnI[39–60] binds to the hydrophobic face of cCTnC, stabilizing an alpha helix in cTnI[41–67] and a type VIII turn in cTnI[38–41]. In contrast, cTnI[1–37] remains disordered, although cTnI[19–37] is electrostatically tethered to the negatively charged surface of cNTnC (opposite its hydrophobic surface). The interaction does not directly affect the calcium binding affinity of cNTnC. However, it does fix the positioning of cNTnC relative to the rest of the troponin complex, similar to what was previously observed in an X-ray structure [Takeda S, et al. (2003) Nature 424(6944):35–41]. Domain positioning impacts the effective concentration of cTnI[148–158] presented to cNTnC, and this is how cTnI[19–37] indirectly modulates the calcium affinity of cNTnC within the context of the cardiac thin filament. Phosphorylation of cTnI at Ser22/23 disrupts domain positioning, explaining how it impacts many other cardiac regulatory mechanisms, like the Frank–Starling law of the heart.The balance between contraction and relaxation must be carefully regulated in the heart. Impaired relaxation can lead to diastolic heart failure, whereas systolic failure is characterized by insufficient contractility. Despite having different etiologies, both forms of heart failure are similar in terms of prevalence, symptoms, and mortality (1). Of all of the signaling pathways that regulate contractile function, the best studied is sympathetic β₁-adrenergic stimulation (2), which leads to cardiomyocyte cAMP production and activation of protein kinase A (PKA). Downstream phosphorylation of L-type calcium channels and phospholamban increases calcium fluxes, whereas phosphorylation of sarcomeric proteins, cardiac troponin I (cTnI), cardiac myosin binding protein-C, and titin (3) regulates the calcium-induced mechanical response.In human cTnI, Ser22 and Ser23 are the residues most consistently phosphorylated (4, 5). (There are some numbering inconsistencies in the literature, and we will refer to Ser22/23 instead of Ser23/24 to account for physiologic removal of the N-terminal methionine residue.) Originally identified as PKA targets, Ser22/23 are now known to be phosphorylated by other kinases, including PKG, PKCβ, PKCδ, and PKD1 (6), showing it to be an important locus at which multiple signaling pathways converge. Phosphorylation at cTnI Ser22/23 decreases the calcium sensitivity of the cardiac sarcomere (7). High levels of phosphorylation are seen in healthy individuals, but decreased phosphorylation levels occur in a number of pathologic states, including heart failure with reduced ejection fraction, heart failure with preserved ejection fraction, dilated cardiomyopathy, and hypertrophic cardiomyopathy (5, 8). Although dephosphorylation is likely a compensatory mechanism in many cases, it may be a disease-driving dysregulation in others.Other regulatory mechanisms are strongly influenced by the phosphorylation state of Ser22/23. The Frank–Starling law of the heart, also known as length-dependent activation or stretch activation, is more pronounced when Ser22/23 are phosphorylated (9, 10). In contrast, Ser5 (11) or Ser41/43 (12, 13) phosphorylation has more of an impact when Ser22/23 are unphosphorylated. Finally, some mutations that cause familial dilated cardiomyopathy have been shown to mitigate the effect of Ser22/23 phosphorylation (14). Despite the physiologic importance of Ser22/23 phosphorylation in regulating cardiac calcium sensitivity, the extent of its modulatory capacity has remained elusive.Ser22/23 lie within the cardiac-specific N-terminal region, cTnI[1–31], not present in the skeletal muscle isoforms. cTnI[1–209] forms long stretches of helical structure along a winding course that binds to troponin C, troponin T, and actin–tropomyosin. The X-ray structure of the cardiac troponin complex (15) did not include cTnI[1–34], so the structure of this region has not been determined, although there have been some preliminary investigations (16, 17). It is known that cTnI[1–31] interacts with cTnC in its unphosphorylated state (18), but phosphorylation abolishes this interaction, having an effect similar to truncation or removal of cTnI[1–31] (19). Our present study provides a detailed analysis of the structure and dynamics of cTnI[1–73] in complex with cTnC using solution NMR spectroscopy, highlighting its unique mechanism of action and physiologic implications.     

Prognostic use of cardiac troponin T and troponin I in patients with heart failure

BACKGROUND: Troponin T (cTnT) and troponin I (cTnI) are present in the sera of some heart failure (HF) patients and have potential importance as prognostic markers. OBJECTIVE: To prospectively evaluate the prognostic value of cTnT and cTnI in well-characterized HF patients and clarify their relationship to other clinical markers of HF severity. METHODS: cTnT and cTnI were measured in 78 HF patients (45 inpatients, 33 outpatients) who were followed up prospectively for 12 months. RESULTS: Plasma cTnT (> or =0.02 ng/mL) and cTnI (> or =0.3 ng/mL) were detected in 51% and 46% of patients, respectively. These patients were more likely to be inpatients (70% versus 45% for cTnT, 75% versus 43% for cTnI, P<0.05 for both), have a higher plasma creatinine (153 versus 119 micromol/L for cTnT; 157 versus 118 micromol/L for cTnI, P<0.05) and lower plasma sodium (134 versus 138 mmol/L for both, P<0.05). At 12 months, they were more likely to have died or undergone cardiac transplantation (41% versus 14%, P=0.01 for cTnT; 43% versus 15%, P=0.004 for cTnI). After adjustment for New York Heart Association class, plasma sodium and inpatient status, a significant association with events was still evident for both troponins. CONCLUSIONS: Both cTnT and cTnI are strongly associated with other clinical indicators of HF severity and remain independent predictors of prognosis after adjustment for these factors. These results indicate a potential role for cTnT and cTnI in the clinical management of HF patients.     

Baseline troponin level: key to understanding the importance of post-PCI troponin elevations.

AIMS: The adverse prognostic significance of biomarker elevations after percutaneous coronary intervention (PCI) is well established. However, often baseline troponin values are not included in the analysis or sensitive criteria are not employed. Accordingly, we assessed the timing and magnitude of post-PCI troponin T (cTnT) levels and their relationships to outcomes in patients with and without pre-PCI baseline cTnT elevations using a sensitive assay and sensitive cut-off values. METHODS AND RESULTS: cTnT was measured at baseline (pre-PCI), 8 and 16 h post-PCI in 2352 patients. A cTnT elevation was defined as > or =0.03 ng/mL. No baseline cTnT elevations were detected in 1619 patients undergoing mostly (97%) non-urgent procedures (cTnT = 0.01 +/- 0.002 ng/mL; mean +/- SD). 733 patients had baseline cTnT elevations. Only the baseline troponin value had prognostic importance. Patients with elevated cTnT baseline levels had a higher overall cumulative 12-month death/MI rate of 11.1% compared with those without elevated baseline levels of 4.7% (P < 0.05). Neither the timing nor the magnitude of the post-procedure cTnT elevations was predictive of long-term death/MI rates when baseline elevations were included in the analysis. Similar findings were observed for baseline creatine kinase-MB (CK-MB) levels. Late increases in cTnT levels (16 h post-PCI) presaged in-hospital events only. CONCLUSION: Long-term prognosis is most often related to the baseline pre-PCI troponin value and not the biomarker response to the PCI. These results support a re-evaluation of the use of biomarker data in relation to PCI.     

Comparison of new point-of-care troponin assay with high sensitivity troponin in diagnosing myocardial infarction

Objectives

The aim of this study is to compare a new improved point of care cardiac troponin assay (new POC-cTnI) with 1. its predecessor (old POC-cTnI) and 2. a high sensitivity assay (hs-cTnI) for the diagnosis of acute myocardial infarction (AMI) and for major adverse cardiac events (MACE) by 30 days.

Methods

This is a single centre observational study, set in Christchurch Hospital, New Zealand. Patients presenting to the emergency department with non-traumatic chest pain underwent blood sampling at 0 h and 2 h post presentation for analysis with the 3 cTnI assays for the outcome of AMI and for analysis using an accelerated diagnostic protocol (ADP-normal 2 h troponins, normal electrocardiograms and Thrombolysis In Myocardial Infarction (TIMI) score of 0 or ≤ 1) for 30 day MACE.

Results

Of 962 patients, 220 (22.9%) had AMI. Old POC-cTnI was least sensitive at 70.0% (65.4–73.9%) by 2 h (p < 0.001). New POC-cTnI, sensitivity 93.6% (89.9–96.2%) had similar sensitivity to hs-cTnI, sensitivity 95.0% (91.5–97.3%) (p = 0.508). There were 231 (24.0%) patients with 30 day MACE. When used as part of the ADP, all assays had 100% (98.0–100%) sensitivity using TIMI = 0. Sensitivities of new POC-cTnI ADP, 98.3% (95.4–99.4%), old POC-cTnI, 96.5% (93.2–98.4%) and hs-cTnI, 98.7% (96.0–99.7%) were similar (p = 0.063–0.375) using TIMI ≤ 1.

Conclusions

A new POC-cTnI has improved sensitivity for AMI and MACE compared with its predecessor and comparable sensitivity to a high sensitivity assay. Now that sensitivities of the POC assay are improved, the new assay may be a useful alternative to central laboratory assays when rapid turn-around times are not possible.     

Interaction of levosimendan with cardiac troponin C in the presence of cardiac troponin I peptides

The interaction between troponin C (TnC) and troponin I (TnI) is essential for the regulation of muscle contraction. There are several binding sites for TnI on TnC that are differentially occupied depending on the phase of the contraction/relaxation cycle. TnI and TnC interact in an antiparallel fashion with each other. The C-domain of cTnC and the N-domain region of cTnI(residues 33-70) always interact under physiological conditions, whereas the interaction between regulatory regions of TnC and TnI (residues 128-166) is calcium dependent. Previously, it has been shown that levosimendan, a calcium sensitizer used as a treatment for acute heart failure, can interact with both domains of isolated cTnC. To understand which interaction is relevant for the mechanism of calcium sensitization, we used a more complete troponin model obtained by complexing cTnI(32-79) and cTnI(128-180) with calcium-saturated cTnC(CS). The cTnI peptides bound to cTnC(CS) to form a 1:1:1 complex. The interaction of levosimendan with this complex was followed by 1H-(15)N heteronuclear correlation spectroscopy. It was clear that based on chemical shift changes, cTnI(32-79) blocked the levosimendan interaction sites on the C-domain, whereas cTnI(128-180) did not compete with levosimendan for the binding site on the N-domain. Hence, the effective binding site of levosimendan on cTnC resulting in the calcium-sensitizing effect is located in the regulatory domain (N-domain).     

血清肌钙蛋白T及I对急性心肌梗死早期诊断的临床价值

目的 :观察急性心肌梗死 (AMI)患者入院前后血清肌钙蛋白T(cTnT)、肌钙蛋白I(cTnI)的变化 ,探讨cTnT、cTnI对早期诊断AMI的价值。方法 :对 10 0例患者 (AMI 5 0例、不稳定型心绞痛 5 0例 )和 2 0例健康人进行了血清cTnT、cTnI、肌酸激酶 (CK)和肌酸激酶同工酶 (CK MB)检测。结果 :①血清cTnT、cTnI、CK和CK MB检测AMI的敏感度和特异度分别为 80 %和 90 %、85 %和 92 %、92 %和 6 1%、75 %和 75 %。②AMI患者血清cTnT、cTnI浓度平均升高 (17.5± 5 .7)和 (16 .2± 4 .8)倍 ,显著高于CK(7.6± 3.1)倍、CK MB (6 .5± 2 .9)倍 (P <0 .0 1) ;③AMI后 3h内cTnT、cTnI阳性检出率为 5 0 .0 %、4 0 .0 % ,明显高于CK MB(2 4 .0 % )和CK(2 0 .0 % ) ;AMI 5d后cTnT、cTnI阳性检出率为 70 .0 %、6 6 .0 % ,而CK MB和CK仅为 4 .0 %和 8.0 %。结论 :血清cTnT、cTnI能早期确切诊断AMI ,具有较宽的时间诊断窗口 ,是心肌梗死早期诊断较敏感和特异的血清标志物     

cTnT、cTnI在急性病毒性心肌炎诊断中的价值

目的 探讨心肌损伤指标肌钙蛋白T(cTnT)、肌钙蛋白I(cTnI)对急性病毒性心肌炎的临床诊断价值。方法116例首次诊断为急性病毒性心肌炎患者,在其发病的一至二周内,同时定量检测血清cTnT、cTnI和心肌酶谱系列肌酸磷酸激酶(CK)及其同功酶(CK-MB)、血清谷草转氨酶(sGOT)、乳酸脱氢酶(LDH)并作比较。结果 116例急性病毒性心肌炎患者中,共有53例cTnT异常升高,阳性率为45.7%;共有75例cTnI异常升高,阳性率为64.7%;而CK、CK-MB、sGOT、LDH检测阳性率分别为10.3%、7.8%、6.9%、12.9%。上述结果分别与cTnT及cTnI结果作卡方检验均有显著差异(P<0.01)。结论 病毒性心肌炎患者在急性期心肌损伤指标检测中,cTnT、cTnI比传统的CK、CK-MB、sGOT、LDH有更好的敏感度,是及时反映心肌损伤的良好的观察指标。     

Stunned peri-infarct canine myocardium is characterized by degradation of troponin T, not troponin I

OBJECTIVE: Degradation of cardiac troponin I (cTnI) has been proposed to represent the underlying molecular mechanism responsible for post-ischemic contractile dysfunction of viable but 'stunned' myocardium. However, this concept is largely derived from models of brief, sublethal ischemia essentially devoid of necrosis, and there is speculation that defects in cTnI may be model-dependent. Accordingly, our primary aim was to evaluate the integrity of cardiac troponins-i.e., cTnI, as well as cTnT and cTnC-in viable but stunned peri-infarct tissue. In addition, we addressed the as-yet unexplored issue of whether the profound reduction of infarct size evoked by brief preconditioning ischemia (PC) was accompanied by a favorable attenuation in ischemia/reperfusion-induced degradation of cTnI, cTnT or cTnC in the remaining viable subepicardium. METHODS: Anesthetized open-chest dogs received 10 min of PC ischemia or a comparable control period, followed by 1 h of sustained coronary occlusion and 3 h of reperfusion. Subepicardial biopsies from the center of the soon-to-be ischemic territory were obtained at baseline and at 30 min and 3 h post-reflow, and myofilament protein integrity (intact cTnI, cTnT and cTnC, as well as degradation bands and covalent complexes) were assessed by Western immunoblotting. In addition, in all dogs, wall thickening was measured by echocardiography, collateral blood flow was assessed during sustained occlusion by injection of radiolabeled microspheres, and infarct size was delineated by tetrazolium staining. RESULTS: Although PC was, as expected, cardioprotective (infarct size of 2 +/- 1% of the risk region vs. 17 +/- 6% in controls; p < 0.05), both control and PC groups exhibited profound and comparable contractile dysfunction following reflow (mean wall thickening reduced to 20-22% of baseline values). There was, however, no significant degradation of cTnI in the viable but stunned, peri-infarct tissue. We did observe degradation of cTnT in the stunned subepicardium, an effect that was attenuated in dogs that received antecedent PC ischemia. However, there was no correlation between post-ischemic wall thickening and the immunoreactivity of the intact cTnT band, or wall thickening and the intensity of the cTnT degradation products. CONCLUSIONS: Our results suggest cTnI degradation is not a universal determinant of post-ischemic myocardial stunning. Moreover, the dissociation between cTnT degradation and wall thickening argue against a direct 'cause-and-effect' relationship between proteolysis of cTnT and acute, post-ischemic contractile dysfunction of stunned peri-infarct myocardium.