A Circulating Myocardial Depressant Substance is Associated with Cardiac Dysfunction and Peripheral Hypoperfusion: In Vitro Model of Myocardial CeU Contractile Performance

In: Septic Shock

31 Oct 2014

In Vitro Model of Myocardial CeU Contractile Performance
A modification® of a previously described in vitro myocardial cell contractility system- was used to assay for MDS activity. Briefly, spontaneously beating newborn rat heart cell cultures were established as follows. Hearts from two-day-old rats were pooled and minced into small blocks of tissue. The cells were disaggregated with 0.05 percent trypsin in modified Hanks solution without Ca+ + or Mg+ + .

The myocardial cells were suspended in culture media consisting of the following: potassium-free balanced salt solution (Dulbeccos BSS-K) 65 percent; Medium 199 (Grand Island Biological Company) 25 percent, supplemented with 100 U/ml of penicillin, 100 m/ml of streptomycin, and 10 mM glutamine; and 10 percent newborn calf serum, heat-inactivated at 56°C for 30 min. The cells were then plated at a cell suspension of 5 x 10* cells/ml (2 ml per Petri dish). Cells were incubated at 37°C in a 100 percent humidified atmosphere of 95 percent Os and 5 percent COa. Growth medium was changed initially at 72 to 96 h and every 48 h thereafter. Latex microspheres (4.1 in diameter; Polysciences, Inc) were added with the first change of growth medium. Spontaneously beating cells were used in the assay five to nine days following inoculation of cells into the Petri dish. add comment
The extent of myocardial cell shortening during contraction was assayed by a modification of a previously described technique, employing latex microspheres and a video tracking system. The latex microspheres move in concert with the contracting cell and are tracked with a closed-loop video tracking system. The amplitude or extent of myocardial cell shortening is a function of the distance traveled by the latex microsphere during each contraction of the myocardial cell.


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