Gel stretch method: a new method to measure constitutive properties of cardiac muscle cells

Abstract

Diastolic dysfunction is an important cause of congestive heart failure; however, the basic mechanisms causing diastolic congestive heart failure are not fully understood, especially the role of the cardiac muscle cell, or cardiocyte, in this process. Before the role of the cardiocyte in this pathophysiology can be defined, methods for measuring cardiocyte constitutive properties must be developed and validated. Thus this study was designed to evaluate a new method to characterize cardiocyte constitutive properties, the gel stretch method. Cardiocytes were isolated enzymatically from normal feline hearts and embedded in a 2% agarose gel containing HEPES-Krebs buffer and laminin. This gel was cast in a shape that allowed it to be placed in a stretching device. The ends of the gel were held between a movable roller and fixed plates that acted as mandibles. Distance between the right and left mandibles was increased using a stepper motor system. The force applied to the gel was measured by a force transducer. The resultant cardiocyte strain was determined by imaging the cells with a microscope, capturing the images with a CCD camera, and measuring cardiocyte and sarcomere length changes. Cardiocyte stress was characterized with a finite-element method. These measurements of cardiocyte stress and strain were used to determine cardiocyte stiffness. Two variables affecting cardiocyte stiffness were measured, the passive elastic spring and viscous damping. The passive spring was assessed by increasing the force on the gel at 1 g/min, modeling the resultant stress vs. strain relationship as an exponential [ς = A/ k( e kε− 1)]. In normal cardiocytes, A = 23.0 kN/m2 and k = 16. Viscous damping was assessed by examining the loop area between the stress vs. strain relationship during 1 g/min increases and decreases in force. Normal cardiocytes had a finite loop area = 1.39 kN/m2, indicating the presence of viscous damping. Thus the gel stretch method provided accurate measurements of cardiocyte constitutive properties. These measurements have allowed the first quantitative assessment of passive elastic spring properties and viscous damping in normal mammalian cardiocytes.

Aged bovine chondrocytes display a diminished capacity to produce a collagen-rich, mechanically functional cartilage extracellular matrix

“…For calf articular cartilage, fibril modulus Gbtained by unconfined compression [ 101 was found to be very close to dynamic peak modulus obtained during uniaxial tension testing [6]. In the same manner, the fibril modulus obtained for our samples, ranging from 0.12 MPa initially to 0.55 MPa after 3 weeks of culture, is similar to the reported tensile peak modulus of 0.16 MPa for 2% agarose gel submitted to a 15% stretch [41]. Thus even though the homogeneous nature of the fibril-reinforced biphasic model does not exactly represent the composite structure of our gels where chondrocyte islets are embedded in an agarose gel, its parameters are expected to reflect average bulk properties.…”

Role of microtubules versus myosin heavy chain isoforms in contractile dysfunction of hypertrophied murine cardiocytes

“…The methods used to measure cell shortening were described in MATE-RIALS AND METHODS. We showed previously (38) that for such gel-embedded cardiocytes there is Ͻ1% difference between gel strain and cell strain under these loading conditions. The embedded cardiocytes were studied at 32°C during electrically stimulated contractions (0.25 Hz, 5-ms pulse width, and voltage 10% above threshold).…”

“…The methods that we have used before (38,41) to characterize the constitutive properties of isolated cardiocytes were modified for the present purpose of examining the effects of variable afterloading on cardiocyte mechanics. The basis for and the details of this technique are given in the APPENDIX.…”

“…The gel was then placed in a custom-designed, computer-controlled stretching device and uniaxially stretched with forces (stresses) sufficient to increase gel length (strain) at a rate of 5 mm/s to a maximum strain of 10-15% of initial gel length without producing plastic deformation of the gel. We have described the basis for these techniques in detail elsewhere (38,41).…”

Dynamic Model for Characterizing Contractile Behaviors and Mechanical Properties of a Cardiomyocyte

“…The contractility of heart cells, as a label-free biomarker for heart diseases such as heart failure and cardiac arrhythmia, has been regarded as another important factor, leading to investigations of cardiac muscle mechanics directly in single, isolated heart cells (22,23). Single heart cells also work as the basic actuation elements of cardiomyocyte-based bio-syncretic robots, and the contractility and mechanics of single cardiomyocytes are fundamental to understanding the dynamics of bio-syncretic robots.…”