Abstract 17932: Diabetic Cardiomyopathy in Mice is Characterized by Defects in Myocyte Turnover Measured by Microdosing of 14C-Labeled Thymidine
Insulin-dependent diabetes mellitus was induced in mice by streptozotocin administration (175 mg/kg bw). Blood glucose was measured daily and when it reached 450 mg/dl, ~5-7 days later, 300 pCi 14C-thymidine was given intramuscularly to label forming cardiomyocytes. Control mice were similarly treated. This protocol results in a pulse labeling of replicating cells, since thymidine disappears from the circulation in less than one hour; the half-life of this nucleotide is ~15 minutes. Control and diabetic mice were sacrificed at 1, 3, 5, 10, 20, 30, 60, and 90 days after the injection of 14C-thymidine and the heart was collected. Left ventricular myocytes were isolated by enzymatic digestion, purified by differential centrifugation, and the DNA was extracted for the measurement of the kinetics of 14C concentration by Accelerator Mass Spectrometry. This is the only methodology that allows the assessment of trace-amount of 14C in the DNA. In non-diabetic control hearts, 14C concentration in the myocyte DNA increased progressively from 1 to 10 days, and remained essentially constant up to 90 days. These findings indicate that a significant number of new myocytes was formed in 10 days by activation and differentiation of progenitor cells. The regenerated myocytes had a lifespan of at least 90 days as demonstrated by the lack of changes in 14C from 10 to 90 days. This process excludes the contribution of mature myocytes reentering the cell cycle and dividing, since this mechanism of myocyte renewal would result in a drop in 14C concentration and could never account for the 2.5-fold increase in 14C in the myocyte DNA from 1 to 10 days. Importantly, myocyte formation was almost completely abrogated by diabetes, a condition characterized by a modest increase in 14C concentration at one day with no further changes from 3 to 10 days, and a significant decrease at 30, 60 and 90 days. The loss in 14C labeling reflected an equivalent loss in cardiomyocytes, a major determinant of the diabetic myopathy. Based on BrdU incorporation in cycling myocytes over a predefined time period, the increase in 14C from 1 to 10 days in control animals corresponded to generation of 180,000 new cells. In conclusion, myocyte turnover is relatively high in mouse heart but is markedly affected by type 1 diabetes.
- © 2013 by American Heart Association, Inc.