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In the heart, the relative proportions of the 2 forms of the motor protein myosin heavy chain (MyHC) have been shown to be affected by a wide variety of pathological and physiological stimuli. Hearts that express the faster MyHC motor protein, alpha, produce more power than those expressing the slower MyHC motor protein, beta, leading to the hypothesis that MyHC isoforms play a major role in the determination of cardiac contractility. We showed previously that a significant amount of alphaMyHC mRNA is expressed in nonfailing human ventricular myocardium and that alphaMyHC mRNA expression is decreased 15-fold in end-stage failing left ventricles. In the present study, we determined the MyHC protein isoform content of human heart samples of known MyHC mRNA composition. We demonstrate that alphaMyHC protein was easily detectable in 12 nonfailing hearts. alphaMyHC protein represented 7.2+/-3.2% of total MyHC protein (compared with approximately 35% of the MyHC mRNA), suggesting that translational regulation may be operative; in contrast, there was effectively no detectable alphaMyHC protein in the left ventricles of 10 end-stage failing human hearts.

Background: Evaluation of gene expression in failing human heart has been limited by the availability of cardiac tissue., Methods and Results: We used the polymerase chain reaction (PCR) to assess gene expression in small quantities of failing and nonfailing human heart. PCR is a powerful new molecular biological tool that allows a small quantity of DNA to be amplified as much as 1 million-fold. Total RNA was extracted from 3-5 mg samples of human heart and reverse-transcribed to complementary DNA (cDNA). With selected oligonucleotide primers, we used PCR to amplify cDNAs encoding atrial natriuretic peptide, beta-myosin heavy chain, phospholamban, and cytoskeletal beta-actin. To quantify the relative levels of messenger RNA (mRNA) in human heart, a known amount of a control RNA was present in the reverse transcription and PCR reactions. The amount of mRNA in the sample could therefore be assessed in relation to the amount of control product. The control RNA was transcribed from a synthetic DNA template containing primers complementary to those used to amplify the cDNAs of interest. Atrial natriuretic factor mRNA could not be detected in nonfailing human heart but was abundant in ventricular myocardium from failing human heart. In contrast, steady-state levels of phospholamban mRNA decreased, whereas levels of beta-myosin heavy-chain mRNA were unchanged with heart failure., Conclusions: Alterations in gene expression in the failing human heart appear to be selective. In addition, the present study suggests that PCR provides a rapid and economical way to quantify the expression of multiple genes of interest in endomyocardial biopsy specimens and may therefore be used to advance our understanding of heart muscle disease.

Prenalterol (beta 1-agonist), denopamine (beta 1-agonist), and zinterol (beta 2-agonist) were partial agonists of adenylate cyclase (AC) stimulation in human ventricular myocardium obtained from nonfailing chambers whose beta 1/beta 2 receptor subtype ratio was approximately 80/20. At a concentration less than its low affinity (beta 2) Kl, betaxolol, a highly selective beta 1-antagonist, inhibited isoproterenol (non-selective agonist), denopamine, and prenalterol stimulation of AC, indicating that isoproterenol, denopamine, and prenalterol are all capable of stimulating AC through beta 1-receptor activation. At a concentration less than its low affinity (beta 1) Kl, ICI 118,551, a highly selective beta 2-agonist, inhibited both isoproterenol and zinterol stimulation of AC, indicating that isoproterenol and zinterol stimulate AC through beta 2-receptors. Zinterol stimulation of AC was mediated entirely by beta 2-receptors, inasmuch as 10(-7) M betaxolol had no effect on the zinterol dose-response curve and ICI 118,551 produced a degree of blockade (KB = 5.2 +/- 1.6 X 10(-9) M), consistent with the beta 2-receptor Kl of the latter (2.0 +/- .4 X 10(-9) M, p, not significant). In nonfailing myocardium, analysis of beta 1 versus beta 2 stimulation by the nonselective agonist isoproterenol revealed that the numerically small (19% of the total) beta 2 fraction accounted for the majority of the total adenylate cyclase stimulation. In failing ventricular chambers with a beta 1/beta 2 receptor subtype ratio reduced from 82/19 (nonfailing) to 64/36 (p less than 0.001) and a beta 1-receptor density reduced by 61% (p less than 0.001), maximal denopamine stimulation was reduced by 49% (p less than 0.001). Moreover, in preparations from failing heart, the component of denopamine stimulation that was inhibited by 10(-7) M betaxolol (beta 1 component) was reduced by 77% (p less than 0.05). Finally, in preparations derived from failing ventricular myocardium, beta 2-receptor density was not significantly decreased, but zinterol stimulation of AC was reduced by 32% (p less than 0.05). We conclude that heart failure results in subsensitivity to both selective beta 1 and beta 2 stimulation of adenylate cyclase, with beta 1 subsensitivity due to selective beta 1 receptor down-regulation and beta 2 subsensitivity due to partial uncoupling of beta 2 receptors from subsequent events in the beta 2-adrenergic pathway.