This study was designed to determine the effect of
cyclocreatine on the release of neutrophil
chemotactic factors (NCF) from isolated rabbit hearts. We tested the hypothesis that if
ischemia is important for the formation of NCF from the myocardium, then blocking (or delaying) ischemic changes with
cyclocreatine should inhibit the release of NCF. Two models were used, including (1) perfusion of rabbit hearts (Langendorff apparatus) with oxygenated (95%
oxygen)
Krebs-Henseleit buffer (K-H
buffer) containing 5%
cyclocreatine for 120 minutes, and (2) incubating hearts with
phosphate-buffered saline (PBS) containing 5%
cyclocreatine for 120 minutes. For both models, rabbits were injected intravenously with 10 ml of 5%
cyclocreatine solution 30 minutes before the animals were killed and the hearts removed. Control rabbits were injected with 5%
creatine solution or saline for 30 minutes before perfusing hearts with K-H
buffer or incubating with PBS. Chemotactic activity was assayed in the perfusates and supernatants using modified Boyden chambers and rabbit peritoneal neutrophils as
indicator cells. The
chemoattractant f-Met-Leu-Phe (f-MLP) was the positive control for a 100% response rate. Isolated hearts perfused with
cyclocreatine showed significantly lower chemotactic activity (ie, 1.24 +/- 1% f-MLP; P less than 0.0001) compared to hearts perfused with K-H
buffer (129 +/- 18%) or
creatine (227 +/- 42%) (mean +/- standard error). Similar results were obtained using incubated hearts. Next the effect of
cyclocreatine on neutrophils in the Boyden chamber was determined and it was found that it did not alter neutrophil migration, which excludes a direct inhibitory effect on the cells. Furthermore supernatant from
cyclocreatine-treated hearts did not inhibit neutrophil chemotaxis to C5a, indicating absence of a chemotaxis inhibitor in this preparation. Results of these studies suggest that the observed low activity recovered in perfusate and supernatant of
cyclocreatine-treated hearts is a result of reduction in the synthesis and/or release of the factors from myocardial tissues. Similar to previously established data,
cyclocreatine treatment significantly preserved myocardial
nucleotide levels (ie,
adenosine triphosphate and
creatine phosphate), which supports our hypothesis that the formation of NCF is
ischemia dependent and that maintaining elevated levels of myocardial energy
nucleotides reduced
chemotactic factor release.