Isolation and Quantification of Nanovesicles in Isoproterenol-Induced Myocardial Infarcted Rats

Abstract

Aviwe Ntsethe, Irene Mackraj, Saravanakumar Murugesan and Arunagiri Pandiyan

Introduction and Aim: Myocardial infarction (MI) is one of the leading causes of death worldwide. The pathogenesis and aetiology of MI is still unclear and there is currently no cure for MI. Cardiac troponin is the only well-known cardiac-specific marker for the diagnosis of MI but due to the delayed release of troponin in the circulation, a novel cardiac biomarker is needed in the early stages of development of MI to reduce MI mortality. Recently, it has been reported that exosome concentration is highly regulated by stress and cardiovascular diseases (CVDs). We assessed the hypothesis that exosome secretion by the injured cardiomyocytes is increased during MI and thereby serve as biomarker for MI. The aim of this study was to quantify exosomes in an isoproterenol (ISO)-induced MI rats.
Method: Twelve rats were divided into two groups (group A and B). Group-A (n=6) was the normal control rats and group-B (n=6) was the ISO-treated group. Group-B animals were injected with isoproterenol (85mg/kg/bw) for two consecutive days to induce MI. Blood pressure (BP), heart rate (HR) and body weight were monitored for 7 days in all animals prior the ISO injection and throughout the experiment. After second ISO-injection, all animals were sacrificed and blood, heart tissues were obtained. Histopathological analysis was performed in heart tissue samples and levels of cardiac markers (creatine kinase-MB, lactate dehydrogenase and troponin T) were measured from the serum. Exosomes were isolated from the plasma by differential ultracentrifugation. Exosomes were quantified and characterized using nanoparticle tracking analysis (NTA), transmission electron microscope (TEM) study and ELISA for the quantification of circulating exosomal protein (CD63).
Result and Discussion: ISO injection caused the development of MI and was confirmed by the increase in BP, Cardiac markers (cardiac troponin T, lactate dehydrogenase and creatine kinase). NTA together with TEM analysis revealed particle sizes of the exosome and showed an elevated number of the exosome in ISO-treated animals. Levels of exosomal protein (CD63) were also increased in ISO-treated animals.
Conclusion: The concentration of exosomes was increased in MI rats indicating that circulating exosomes may be used as a novel diagnostic marker for MI.

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