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. Author manuscript; available in PMC: 2023 Sep 2.
Published in final edited form as: Circ Res. 2022 Aug 11;131(6):510–527. doi: 10.1161/CIRCRESAHA.122.321351

Figure 6. Cardiac metabolism and ischemic tolerance in Arrdc4-knockout hearts.

Figure 6.

A-C. Protein expression of GLUT1 was analyzed by Western blot analysis in the membrane fraction and total tissue homogenates from the mouse heart. Pan-cadherin and GAPDH served as loading control for membrane and cytosolic proteins (t-test). D. The Arrdc4-knockout (KO) or wild-type (WT) heart was subjected to global hypoxia (with buffer equilibrated with 95% N2, 5% CO2) or normoxia (95% O2, 5% CO2) in the Langendorff perfusion system. The perfusate was switched from Krebs-Henseleit (KH) buffer to a solution containing [3H] 2-deoxy-D-glucose. The heart was perfused with the radiolabeled glucose for 20 min and washed with the standard KH buffer for 5 min. The radioactivity was measured in whole heart homogenates by liquid scintillation counting (Mann-Whitney U test). E. Myocardial glycogen storage was quantified in heart homogenates at baseline and normalized by heart weight (Mann-Whitney U test). F. Exogenous oxidation rates of [3H]-palmitate were determined by measuring the release of 3H2O into the perfusion buffer and normalized by myocardial uptake of [3H]-palmitate. P=N.S. between the genotypes (Mann-Whitney U test). G. Cellular ATP content per heart weight was measured in isolated hearts following 60 min of normoxic or hypoxic perfusion (t-test). H and M. Representative tracing of left ventricular (LV) pressure in the WT or Arrdc4-KO heart under hypoxic perfusion or following global ischemia-reperfusion. I-L and N-Q. Arrdc4-KO hearts maintained better cardiac mechanical function during hypoxic perfusion (1 hr) or after ischemia (15 min) and reperfusion, as shown by the higher LV dP/dt maximum (I or N) and minimum (J or O) and lower LV end-diastolic pressure (K or P) than those in WT hearts. Heart rate slowed down during hypoxic perfusion (L) or after reperfusion (Q) in both genotypes (the P-value indicates the comparison between Arrdc4-KO hypoxia or ischemia vs. WT hypoxia or ischemia by two-way ANOVA with repeated measures followed by Bonferroni test).