Elevated DNase activity and caspase expression in association with apoptosis in failing ischemic sheep left ventricles

Abstract

We examined the DNase activity as well as the expression of the caspases in left ventricles of failing sheep hearts and compared it with samples from normal controls. The sheep model of chronic ischemic heart failure was developed by intracoronary microembolisation. Moderate heart failure was defined as a decrease of left ventricular (LV) ejection fraction (EF) to 35% or less and was stable for four weeks after the last embolisation. The failing hearts were harvested from these animals six months later. Bovine pancreas DNase I was used as a standard for the DNase zymograms. In normal sheep left ventricles, a low level of the DNase I‐like activity was detected (1.16 ± 0.17 pg/100 μg protein, n = 20), whereas it was elevated (3.64 ± 0.49 pg/100 μg, n = 32, p < 0.001) in the ischemic failing LV samples. The identity of the DNase in sheep is yet to be fully characterised. However, we also detected the expression of caspase‐2 and caspase‐3 by Western blotting. There was a 2.2‐fold increased expression of caspase‐2 p < 0.001= and a 2.6‐fold increased expression of caspase‐3 (p < 0.001) in failing left ventricles compared to normal samples.

P2X1 receptors are closely associated with connexin 43 in human ventricular myocardium

“…Western blotting was performed as described previously [27]. Proteins were extracted from homogenized tissues in a buffer containing 5 mM NaH 2 PO 4 (pH 7.4), 0.1 mM EDTA, 0.4 mM dithiothreitol (DTT), 5 Al/ml phenylmethylsulfonyl fluoride and 5 Al/ml protease inhibitor cocktail including chymostatin and leupeptin.…”

Heart failure and apoptosis: Electrophoretic methods support data from micro- and macro-arrays. A critical review of genomics and proteomics

“…It is clear that DNase I is upregulated in the failing heart sample. These data have been confirmed using paced canine left ventricle [3] as well as Heat shock protein DOWN DOWN another animal model of human heart failure, produced by microembolization in sheep left ventricle [5]. DNase I activity can also be monitored spectrophotometrically [6].…”

Remodeling of the chronic severely failing ischemic sheep heart after coronary microembolization: functional, energetic, structural, and cellular responses

“…Expression of FasL and caspases-8, -3, and -2. The protein expressions of FasL and caspases were determined by Western blotting as previously described (15,16). Briefly, protein extracts from the sheep LV were separated by SDS-PAGE.…”