Ito H, Miller S C, Akimoto H, Torti S V, Taylor A, Billingham M E, Torti F M
Department of Medicine, Stanford University Medical Center, CA 94305.
J Mol Cell Cardiol. 1991 Oct;23(10):1117-25. doi: 10.1016/0022-2828(91)90201-v.
Doxurubicin is an effective and widely used chemotherapeutic agent. However, use of this drug is often limited by its cardiotoxic side effects. We have observed that an early event accompanying doxorubicin cardiomyopathy is a selective decrease in levels of muscle gene transcripts in cardiac tissue (Ito et al., Proc. Natl. Acad. Sci. USA 87: 4275-4279). Since this decrease precedes ultrastructural evidence of cardiac damage, measurements of muscle transcripts might assist in the clinical evaluation of doxorubicin cardiotoxicity. We have therefore assessed the utility of the polymerase chain reaction in the measurement of mRNA in control and doxorubicin-treated animals. These measurements were performed on small tissue samples that simulate endomyocardial biopsies. We measured cardiac alpha-actin transcripts as a fraction of ferritin heavy chain transcripts using the method described by Chelly et al. (Nature 333: 858-860, 1988). 0.5 micrograms of total RNA, an amount equivalent to that obtainable from a typical endomyocardial biopsy, was efficiently co-amplified with cardiac alpha-actin and ferritin heavy chain specific primers. The cardiac alpha-actin/ferritin heavy chain ratio calculated from the PCR results correlated well (R = 0.981) with results obtained using Northern blot analysis of 10 micrograms RNA. The correlation was maintained over a wide range of cardiac alpha-actin transcript abundance. These results show that mRNA from cardiac tissue can be estimated by the polymerase chain reaction, even from a small, endomyocardial biopsy-sized sample.
阿霉素是一种有效且广泛应用的化疗药物。然而，该药物的使用常常受到其心脏毒性副作用的限制。我们观察到，伴随阿霉素性心肌病的一个早期事件是心脏组织中肌肉基因转录物水平的选择性降低（伊藤等人，《美国国家科学院院刊》87: 4275 - 4279）。由于这种降低先于心脏损伤的超微结构证据出现，肌肉转录物的测量可能有助于阿霉素心脏毒性的临床评估。因此，我们评估了聚合酶链反应在测量对照动物和阿霉素处理动物的mRNA中的效用。这些测量是在模拟心内膜活检的小组织样本上进行的。我们使用谢利等人（《自然》333: 858 - 860, 1988）描述的方法，将心脏α - 肌动蛋白转录物作为铁蛋白重链转录物的一部分进行测量。0.5微克总RNA（相当于从典型心内膜活检中可获得的量）与心脏α - 肌动蛋白和铁蛋白重链特异性引物有效地共同扩增。由PCR结果计算出的心脏α - 肌动蛋白/铁蛋白重链比值与使用10微克RNA进行Northern印迹分析得到的结果相关性良好（R = 0.981）。在广泛的心脏α - 肌动蛋白转录物丰度范围内，这种相关性都得以保持。这些结果表明，即使是从小的、心内膜活检大小的样本中，也可以通过聚合酶链反应来估计心脏组织的mRNA。
