Human SERCA2a levels correlate inversely with age in senescent human myocardium
Brian S. Cain, MDa,
Daniel R. Meldrum, MDa,
Kyung S. Joo, phDa,
Ju-Feng Wang, phDa,
Xianzhong Meng, MDa,
Joseph C. Cleveland, Jr., MDa,
Anirban Banerjee, PhDa and
Alden H. Harken, MDa
a Department of Surgery, University of Colorado Health Sciences Center, Denver, Colorado, USA
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Figure 1 Experimental protocols. All experiments were preceded by a 90-min period that allowed for stabilization of developed force. Simulated ischemia refers to incubation of trabeculae in substrate free, hypoxic Tyrode’s solution while pacing at 3 Hz. Normoxic perfusion and reperfusion refer to incubation of trabeculae in oxygenated Tyrode’s with substrate and pacing at 1 Hz. Ca2+ and wash indicate Ca2+ concentration changes to from 2.0 to 3.0 mmol/L and back to 2.0 mmol/L, respectively.
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Figure 2 A) Maximum rate of systolic contraction after simulated ischemia reperfusion injury (as measured by the ratio of +dF/dt before and after I/R) was better preserved in the younger versus older patients. B) Maximum rate of diastolic relaxation or compliance after simulated ischemia reperfusion injury (as measured by the ratio of –dF/dt before and after I/R) was better preserved in the younger versus the older patients. Dotted lines depict the 95% confidence limits.
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Figure 3 A) Inotropic response to 3.0 mmol/L Ca2+ concentration. Percent baseline developed force (%BDF) versus age group (senior, >60 years; junior,  60 years) is depicted. Increase in developed force after exposure to exogenous Ca2+ did not differ between senior and junior myocardium (p > 0.05). B) Maximum rate of systolic contraction after Ca2+ bolus (as measured by the ratio of +dF/dt before and after Ca2+ bolus) was increased in the younger patients; however, older patients were not able to increase their rates of systolic contraction. C) Maximum rate of diastolic relaxation or compliance after Ca2+ bolus (as measured by the ratio of –dF/dt before and after Ca2+ bolus) was increased in the younger patients; however, older patients were not able to increase their rates of diastolic relaxation. Dotted lines depict the 95% confidence limits.
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Figure 4 Immunoblot against SERC2a (110 kD), calsequestrin (63 kD), pentameric phospholamban (25 kD), and monomeric phospholamban (25 kD). Samples were each immunoprecipitated with anti-SERC2a, calsequestrin, or phospholamban selective antibodies, separated by SDS-PAGE on a 4–20% acrylamide gel, and then visualized by autoradiography. With increasing age, protein concentration of SERC2a and pentameric phospholamban decrease with age, while calsequestrin and monomeric phospholamban remain relatively constant.
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Figure 5 Protein concentration of SERCA2a (normalized to youngest patient) assessed by Western blot. SERCA2a protein content decreased with increasing age by linear regression (p < 0.003). Dotted lines depict the 95% confidence limits.
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Figure 6 Protein concentration of: A) SERCA2a, B) calsequestrin, C) pentameric phospholamban, and D) monomeric phospholamban, between junior (60 years) and senior (>60 years). SERCA2a and pentameric phospholamban were both decreased in the senior myocardium (*p > 0.05 vs. junior age group). Calsequestrin and monomeric phospholamban did not differ between these two groups (p > 0.05).
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Figure 7 A) SERCA2a normalized to calsequestrin protein levels revealed that the decrease in SERCA2a protein did not reflect a general loss of myocardial Ca2+ handling proteins with age (*p < 0.05 vs. junior age group), and B) SERCA2a normalized to monomeric phospholamban protein levels as an assessment of SERCA2a activity revealed that SERCA2a activity was decreased with age (*p < 0.05 vs. junior age group).
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