A new approach to the biochemical pathology of the vascular system, using time governed laminar elution and bioluminescence analyses.
Endothelial cells which cover the inner wall of blood vessels were extracted for bioluminescence analyses of nucleotides and enzymes. The contaminating blood was removed by heparinization and rinsing with ammonium chloride. The content of the endothelial cells of the rat aorta was reached by time governed laminar elution, using a saponin solution to disrupt the cell membranes. Uniformity of extraction was achieved with a Hamilton programmable pump. All the analyses of the eluted fractions showed a characteristic and reproducible peak. The activities of glucose-6-phosphate dehydrogenase and adenylate kinase were significantly higher in the diabetic animals whereas the amount of nucleotides did not differ between diabetic and control rats. The laminar elution technique combined with bioluminescence assay represents a new approach to studies of biochemical alterations in the endothelial cells. The method is also useful for extraction and analyses of other surface layers.
D000227:Adenine Nucleotides / Q000032:analysis; D000263:Adenylate Kinase / Q000032:analysis; D000818:Animals; D001011:Aorta; D003921:Diabetes Mellitus, Experimental / Q000201:enzymology / Q000378:metabolism*; D004727:Endothelium / Q000201:enzymology / Q000378:metabolism*; D005954:Glucosephosphate Dehydrogenase / Q000032:analysis; D006454:Hemoglobins / Q000032:analysis; D008163:Luminescent Measurements; D008291:Malate Dehydrogenase / Q000032:analysis; D008297:Male; D009711:Nucleotides / Q000032:analysis; D051381:Rats
D000227:Adenine Nucleotides; D006454:Hemoglobins; D009711:Nucleotides; D008291:Malate Dehydrogenase; D005954:Glucosephosphate Dehydrogenase; D000263:Adenylate Kinase