The effect of neuraminidase on cell surface and beat rate of aggregated myocardial cells



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Texas Tech University


Morphological as well as physiological changes occur in aggregated embryonic chick myocardial cells after treatment with the sialic acid removing enzyme, neuraminidase (NMDase). Aggregates of seven day embryonic chick ventricular cells were prepared according to a procedure developed by DeHaan, including a plating sequence to remove most of the contaminating cell types. While muscle cells made up the majority (90%) of the core of the aggregate, there was an incomplete layer of fibroblasts (10%) covering the periphery. These beating spherical clumps of cells were then treated with two highly purified preparations of Clostridium perfringens neuraminidase. Roth preparations of enzyme were contaminated with variable amounts of a protein which electrophoretically comigrated with a known standard of C^. perfringens phospholipase C. Therefore, phospholipase C was used as a control in both the beat rate and cell surface studies. The morphological effect of NMDase on the available myocardial cells present at the aggregate's periphery was visualized using polycationic ferritin controlled by pH and acid methylation procedures. NMDase treatment (1.2 Units/ml for 2 hrs. at 37° C) on both prefixed and postfixed aggregates showed a reduction in stain at the cell surface. Although lower concentrations or shorter treatment times with NMDase showed little morphological change from controls, physiological differences in aggregate beat rate could be detected with only 0.03 U/ml after ten minutes. NMDase concentrations, 0.03, 0.5, and 1.2 U/ml, showed net decreases in beat rates over a sixty minute incubation. Within this period, the first thirty minutes displayed definitive decreases in aggregate beat rate while the rest of the hour was typified by arrhythmic contractions. This study demonstrates that NMDase reduces the number of anionic sites at the cell surface of embryonic myocardial cells and also reduces the aggregate beat rate.