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Updated: Jun 12, 2026
Assessment of Sarcoplasmic Reticulum Calcium Reserve and Intracellular Diastolic Calcium Removal in Isolated Ventricular Cardiomyocytes
Published on: September 18, 2017
NADPH oxidase-derived superoxide impairs calcium transients and contraction in aged murine ventricular myocytes
Uwe Rueckschloss1, Marten Villmow, Udo Klöckner
1Julius Bernstein Institute of Physiology, Martin Luther University Halle, Germany. uwe.rueckschloss@medizin.uni-halle.de
Abstract:
Since aging increases oxidative stress, we analyzed the contribution of reactive oxygen species (ROS) to the contractile dysfunction of aged ventricular myocytes and investigated whether short-term interference with ROS formation could normalize contractile performance. Isolated ventricular myocytes from young (2-4 months) and aged (24-26 months) male mice (C57BL/6) were used. We analyzed sarcomere shortening and calcium transients (Indo-1 fluorescence) of voltage clamped ventricular myocytes and myofilament ATPase activity (malachite green assay). Expression of calcium handling proteins (Western blots) and NADPH oxidase subunits (real-time PCR) was quantified, as well as NADPH oxidase activity (lucigenin chemiluminescence). We found that aged myocytes showed decelerated shortening/relengthening without changes in fractional shortening. Calcium transient decay was similarly decelerated, but the amplitude of calcium transients was increased with aging. Calcium sensitivity of myofilaments of aged myocytes was reduced. These age-dependent changes occurred without altered calcium handling protein expression but were reversed by the superoxide scavenger tiron. Aged myocytes showed increased NADPH oxidase expression and activity. Pharmacological inhibition of NADPH oxidase (diphenylene iodonium; apocynin) normalized age-dependent deceleration of shortening/relengthening. In summary, we show that increased superoxide formation by upregulated NADPH oxidase contributes significantly to age-dependent alterations in calcium handling and contractility of murine ventricular myocytes.
Insights
Aging impairs heart cell contraction due to increased reactive oxygen species (ROS). Reducing ROS formation with antioxidants or NADPH oxidase inhibitors can restore normal heart cell function in older mice.
Area of Science:
- Cardiology
- Cellular Physiology
- Aging Research
Background:
- Aging is associated with increased oxidative stress and impaired cardiac function.
- Reactive oxygen species (ROS) are implicated in age-related cellular dysfunction.
- Ventricular myocyte contractility declines with age, impacting overall cardiac performance.
Purpose of the Study:
- To investigate the role of ROS in age-dependent contractile dysfunction of ventricular myocytes.
- To determine if modulating ROS formation can normalize contractile performance in aged myocytes.
- To identify the specific mechanisms, including NADPH oxidase, involved in age-related oxidative stress in cardiac cells.
Main Methods:
- Isolated ventricular myocytes from young and aged mice were studied.
- Sarcomere shortening, calcium transients, and myofilament ATPase activity were measured.
- Expression and activity of NADPH oxidase and calcium handling proteins were quantified.
- The effects of superoxide scavengers (tiron) and NADPH oxidase inhibitors (diphenylene iodonium, apocynin) were assessed.
Main Results:
- Aged myocytes exhibited slower shortening and relengthening, with increased calcium transient decay and amplitude.
- Myofilament calcium sensitivity was reduced in aged myocytes.
- Increased NADPH oxidase expression and activity were observed in aged myocytes.
- Superoxide scavenger tiron and NADPH oxidase inhibition normalized age-dependent contractile deficits.
Conclusions:
- Increased superoxide formation by upregulated NADPH oxidase significantly contributes to age-dependent alterations in cardiac myocyte calcium handling and contractility.
- Targeting ROS production presents a potential therapeutic strategy for age-related cardiac dysfunction.
- These findings highlight the critical role of oxidative stress in the aging heart.
