Histochemical and Fluorescent Techniques for Detection of Early Myocardial Ischemia Following Experimental Coronary Artery Occlusion: A Comparative and Quantitative Study

Abstract

Several workers have used histochemical, enzymatic, and fluorescent meth ods to diagnose early myocardial ischemia, but the problem of unequivocal de tection of early ischemia still remains an enigma to pathologists. In the present study, the left coronary artery was ligated in an animal model, rat, in order to produce myocardial ischemia at different time intervals, from five minutes to six hours. Fluorescent techniques and tetrazolium staining of myocardial suc cinic dehydrogenases have been used to detect onset of ischemia with the pur pose of identifying a sensitive technique for use in routine pathologic specimens.

Nitroso-blue tetrazolium and triphenyl tetrazolium chloride staining of myocardium showed loss of dehydrogenases within five to twenty minutes of ligation of the coronary artery. This loss was consistent and progressively in creased at longer time intervals, the mean ischemic area mapped being 25.74 mm² and 66.87 mm² at five to twenty minutes and six hours respectively. Such comparison of ischemic area of myocardium at different time intervals has not been reported earlier.

Autofluorescence in formalin-fixed, hematoxylin and eosin-stained sections showed positive fluorescence only after fifty to seventy-five minutes of ischemia and was patchy in distribution in the left ventricular wall even up to six hours of ligation.

Examination of myocardium under fluorescent light after acridine orange staining proved to be more sensitive than autofluorescence for detecting ische mia. At five to twenty minutes, the mean ischemic area was 18.67 mm² and by six hours it increased to 27.48 mm² .

Friedman two-way analysis of variance, applied for comparative evaluation of techniques, showed that tetrazolium salts were more sensitive and the mean ischemic area was significantly greater than the zone outlined by acridine or ange (p < .05 at five to twenty minutes; p < .01 at fifty to seventy-five minutes; p < .001 at one and one half to two hours and p < .025 at two and one fourth to three hours).

Autofluorescence was significantly less than acridine orange fluorescence (p < .05 at five to twenty minutes and < .01 at six hours). Such statistical evalu ation has been done for the first time in the present study and has highlighted the value of acridine orange and tetrazolium salts in detection of early ischemia.