High-frequency ultrasound detection of the temporal evolution of protein cross linking in myocardial tissue

The progressive increase in stiffening of the myocardium associated with th e aging process and abetted by comorbid conditions such as diabetes may be linked to an excessive number of collagen cross links within the myocardial extra-cellular matrix. To determine whether ultrasound can delineate chang es in the physical properties of heart tissue undergoing cross linking, we employed a model in which increased cross linking was induced by treating r at myocardial tissue with specific chemical fixatives. Rat hearts (n=5 each group) were arrested at end-diastole, insonified (30 to 50 MHz) fresh with in a few minutes of excision in a phosphate buffered solution, placed in a fixative (10% formalin or 2.5% glutaraldehyde) and insonified at 30-minute intervals thereafter for 24 hours. Ultrasonic attenuation increased in tiss ues cross linked with formalin (maximal change: 27.2 +/- 3.4 dB/cm) and glu taraldehyde (maximal change: 40.2 +/- 5.6 dB/cm) over a 24-hour period. The frequency dependence of the attenuation coefficient increased as a functio n of the extent of collagen cross links in formalin (maximal change: 0.8 +/ - 0.3 dB/cm-MHz) and glutaraldehyde (maximal change: 0.9 +/- 0.6 dB/cm-MHz) . This study represents the first time that the precise time course of myoc ardial protein cross linking in situ has been characterized by using real t ime monitoring, and the physiologic effect has been delineated on microscop ic material properties.