Prior Freezing In Liquid Nitrogen Has Minimal Impact On The Contractile Properties Of Chemically Permeabilized Human Myocardium

Experiments measuring the contractile properties of human myocardium are important for translational research but complicated by the logistical difficulties of acquiring specimens. Accordingly, many groups perform assays using samples that are acquired from patients at one hospital and shipped to another institution for experiments. This necessitates freezing the samples and performing subsequent assays using chemically permeabilized preparations. This study tested how prior freezing impacts contractile properties. Samples of left ventricular tissue were obtained from patients who were having a Ventricular Assist Device implanted. All patients gave informed consent and the study was approved by the Institutional Review Board. Half of each sample was chemically permeabilized within ∼30 minutes of being removed from the patient and used immediately for contractile assays performed at 37C. The other half of the sample was snap-frozen in liquid nitrogen and maintained at −180C for at least 3 months. These specimens were subsequently thawed, chemically permeabilized, and tested in a second series of experiments. Linear mixed models tested whether freezing had a statistically significant impact on contractile properties (n=39 preparations from n=7 patients). Isometric force measured in pCa 4.5 solution was 20.3±1.7 kN m−2 in never-frozen samples and 17.5±1.8 kN m−2 in previously-frozen samples (p=0.07). Passive force measured in pCa 9.0 solution and Hill coefficients were not influenced by prior freezing (p=0.14 and 0.27 respectively). pCa50 in never frozen samples (6.11±0.04) was greater (p<0.001) than that measured after prior freezing (5.99±0.04) but the magnitude of the effect was only ∼0.1 pCa units. We conclude that prior freezing has minimal impact on the contractile properties that can be measured using chemically permeabilized human myocardium.