Functional aspects of creatine kinase isoenzymes in endothelial cells

To characterize the isoenzyme distribution of creatine kinase (CK) in endot helial cells (ECs) and its functional role during substrate depletion, ECs from aorta (AECs) and microvasculature (MVECs) of pig and rat were studied. In addition, high-energy phosphates were continuously monitored by P-31 NM R spectroscopy in pig AECs attached to microcarrier beads. CK activity per milligram of protein in rat AECs and MVECs (0.08 +/- 0.01 and 0.15 +/- 0.08 U/mg, respectively) was <3% of that of cardiomyocytes (6.46 +/- 1.02 U/mg) . Rat and pig AECs and MVECs displayed cytosolic BB-CK, but no MM-CK. Gel e lectrophoresis of mitochondrial fractions of rat and pig ECs indicated the presence of mitochondrial Mi-CK, mostly in dimeric form. The presence of Mi a-CK was demonstrated by indirect immunofluorescence staining using Mia-CK antibodies. When perifused with creatine-supplemented medium, phosphocreati ne (PCr) continuously increased with time (1.2 +/- 0.6 nmol.h(-1).mg protei n(-1)), indicating creatine uptake and CK activity. Glucose withdrawal from the medium induced a rapid decrease in PCr, which was fully reversible on glucose addition, demonstrating temporal buffering of an energy deficit. Be cause both cytosolic and mitochondrial CK isoforms are present in ECs, the CK system may also contribute to energy transduction ("shuttle hypothesis") .