Overcoming Heparin-Associated RT-qPCR Inhibition and Normalization Issues for microRNA Quantification in Patients with Acute Myocardial Infarction
Methodology / validation published in Thromb Haemost (2018)
Abstract
BACKGROUND: Cardiac-enriched micro ribonucleic acids (miRNAs) are released into the circulation following ST-elevation myocardial infarction (STEMI). Lack of standardized approaches for reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) data normalization and presence of RT-qPCR inhibitors (e.g. heparin) in patient blood samples have prevented reproducible miRNA quantification in this cohort and subsequent translation of these biomarkers to clinical practice. MATERIALS AND METHODS: Using a RT-qPCR miRNA screening platform, we identified and validated an endogenous circulating miRNA as a normalization control. In addition, we assessed the effects of in vivo and in vitro anticoagulant drugs administration (heparin and bivalirudin) on three RT-qPCR normalization strategies (global miRNA mean, exogenous spike-in control [cel-miR-39] and endogenous miRNA control). Finally, we evaluated the effect of heparin and its in vitro inhibition with heparinase on the quantification of cardiac-enriched miRNAs in STEMI patients. RESULTS: miR-425-5p was validated as an endogenous miRNA control. Heparin administration in vitro and in vivo inhibited all RT-qPCR normalization strategies. In contrast, bivalirudin had no effects on cel-miR-39 or miR-425-5p quantification. In vitro RNA sample treatment with 0.3 U of heparinase overcame heparin-induced over-estimation of cardiac-enriched miRNA levels and improved their correlation with high-sensitivity troponin T. CONCLUSION: miRNA quantification in STEMI patients receiving heparin is jeopardized by its effect on all RT-qPCR normalization approaches. Use of samples from bivalirudin-treated patients or in vitro treatment of heparin-contaminated samples with heparinase are suitable alternatives for miRNA quantification in this cohort. Finally, we reinforce the evidence that cardiac-enriched miRNAs early after myocardial reperfusion reflect the severity of cardiac injury.
Abstract sourced from PubMed (NCBI) for the cited record. See the original publication for the authoritative version.
Resumen
RT-qPCR miRNA-quantification methodology study showing bivalirudin (leech-derived DTI) does not inhibit RT-qPCR reactions unlike heparin. Bivalirudin-treated samples enable accurate cardiac-miRNA biomarker quantification in STEMI patients.
Por qué esto importa para la hirudoterapia
Este estudio de validación examinó cómo los fármacos anticoagulantes —específicamente la heparina y la bivalirudina— afectan la cuantificación de microARN basada en RT-qPCR en muestras de sangre de pacientes con infarto agudo de miocardio con elevación del segmento ST (STEMI), identificando miR-425-5p como control endógeno de normalización y demostrando que el tratamiento con heparinasa o el uso de muestras tratadas con bivalirudina pueden superar la inhibición de la RT-qPCR inducida por heparina. El resumen discute la heparina y la bivalirudina como los anticoagulantes estudiados, pero no menciona la hirudina, las sanguijuelas ni ninguna sustancia derivada de sanguijuela. En consecuencia, no existe una conexión defendible con la hirudoterapia o el secretoma de la sanguijuela basándose únicamente en el resumen, y cualquier relevancia con el ámbito de la ASH sería, en el mejor de los casos, indirecta. El estudio es una validación metodológica centrada en mejorar la medición de biomarcadores de miARN en pacientes cardiológicos.
Citación
Overcoming Heparin-Associated RT-qPCR Inhibition and Normalization Issues for microRNA Quantification in Patients with Acute Myocardial Infarction.
Coelho-Lima J et al. · Thrombosis and haemostasis, 2018
Contexto clínico relacionado
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Añadido a la biblioteca ASH: May 27, 2026 · Última actualización del sitio: 18 de junio de 2026