Circulation, Vol 83, 1866-1872, Copyright © 1991 by American Heart Association
AM Feldman, PE Ray, CM Silan, JA Mercer, W Minobe and MR Bristow
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.
ARTICLES
Selective gene expression in failing human heart. Quantification of steady-state levels of messenger RNA in endomyocardial biopsies using the polymerase chain reaction
Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Md. 21205.
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