Lois C. Armiger

Histological assessment of orthotopic aortic valve leaflet allografts: its role in selecting graft pre-treatment

Summary Histopathological studies of human cardiac valve grafts recovered at autopsy or re‐operation, together with long‐term clinical follow‐up of valve graft recipients, have indicated that the success of grafts is largely dependent upon the extent to which they are replaced by host fibrous connective tissue. To find the valve preparation technique with least inhibitory effect on tissue ingrowth after grafting, various sterilizing and storage procedures were evaluated using a series of aortic valve leaflet allografts in dogs. To facilitate evaluation, a method for rapidly assaying relative degrees of colonization of grafts was first devised. Application of this method has unequivocally identified a newly‐formulated antibiotic solution as the pre‐treatment most compatible with host tissue ingrowth.

The effect of ischaemia on the function and fine structure of the microvasculature of myocardium.

Summary Seventeen mongrel dogs were subjected to ligation of the circumflex branch of the left coronary artery. At intervals from 10 to 300 minutes after ligation, 10 ml of a sodium fluorescein and colloidal thorium hydroxide solution were injected via an intra‐arterial catheter placed distal to the ligature. Up to 20 minutes after ligation this tracer solution evenly perfused the left ventricular wall including the posterior papillary muscle (PPM). During 25 to 50 minutes there was a progressive patchy loss of perfusion into the PPM although the sub‐epicardial region was evenly perfused. After 60 or more minutes of ischaemia it was not possible to inject this tracer solution into the subendocardial region. Electron microscopic examination indicated that this loss of vascular competence was possibly due to vasospasm of some vessels in the marginal zone between the perfused and non‐perfused areas and was unlikely to be due to intravascular thrombosis, endothelial cell swelling, or to compression of vessels due to cell or tissue oedema.

Effects of glutamic acid on cardiac function and energy metabolism of rat heart during ischaemia and reperfusion

The effects of exogenous glutamate (20 mM) on myocardial energy metabolism and cardiac function during low-flow ischaemia and subsequent reperfusion were studied in isolated working rat hearts. Hearts were made severely ischaemic for 60 min by reducing the perfusion rate to 0.17 ml/min, and then reperfused for 30 min. Low-flow ischaemia resulted in a 50% reduction of myocardial ATP, a 70% reduction of both creatine phosphate (CP) and GTP, and a 250% rise in AMP. After reperfusion, CP was restored to normal levels but ATP and GTP remained significantly low. All hearts failed completely to recover cardiac pump function. The addition of glutamate to the perfusate during low-flow ischaemia had no significant effect on myocardial high-energy phosphates (HEP) but slightly increased succinate production. Subsequent reperfusion without added glutamate resulted in the recovery of 62% of pre-ischaemic aortic flow rate, as well as restoration of myocardial ATP and GTP to 70% of their control values and of creatine phosphate to supranormal levels. Reperfusion with added glutamate did not raise HEP levels any further but did increase recovery of cardiac function to 92% or more of pre-ischaemic values. Thus, by mechanism(s) which are not yet clear but which may include an increase in HEP via anaerobic succinate production, elevated levels of exogenous glutamate exert a highly beneficial effect on the post-ischaemic recovery of cardiac function.

An occurrence of Labyrinthula in New Zealand Zostera

Summary This paper reports the occurrence of a persistent and systematic dying-off of Zostera in the vicinity of Auckland, and the detection of the slime mould Labyrinthula in partially discoloured leaf material from affected areas. The general similarity of the situation to the wasting disease epidemic which took place in the Northern Hemisphere some three decades ago is pointed out, and the ecological implications arc briefly discussed.

Viability studies of human valves prepared for use as allografts

The preimplantation viability status of pulmonary and aortic valves prepared for use as allografts by the methods in current use at Green Lane Hospital, Auckland was determined by autoradiography and culture. The valves were obtained from cadaver donors, disinfected in antibiotic solution and stored by cryopreservation. A group of 45 banked valves considered unsuitable for clinical use was assayed initially and very few were found to have viable fibroblasts in their leaflets. A series of 29 valves collected at postmortem examination then was assayed sequentially after each phase of the preparation procedure. Valves obtained within 24 hours of donor death usually retained considerable viability. However, in all but a minority of cases this declined markedly after antibiotic treatment and further still after cryopreservation, so that most valves were nonviable when thawed.

An experimental evaluation of staining techniques for the detection of early ischaemic injury to the myocardium.

Summary A series of experimental infarcts of the posterior papillary muscle of the canine heart was used to assess the value of 6 special stains in the histological detection of early myocardial infarction. The infarcts were of 5–720 min duration and were compared not only with normal control myocardium but also with normal myocardium autolyzed for similar periods of time. All tissue was stained with H & E, PAS, PAS‐diastase, PTAH, Massons trichrome, Connors modification of the acid fuchsin method, Puchtlers PAS‐navy blue, and Lies haematoxylin‐basic fuchsin‐picric acid. The zone of severely altered myofibres which separated normal from ischaemic tissue in infarcts aged 2 or more hours was demonstrated by all but PAS. Normal, border, and ischaemic zones of heart muscle were clearly differentiated only by PTAH and PAS‐navy blue. PAS distinguished normal from glycogen‐depleted ischaemic myocardium after only 40–60 min, but this change was also seen in autolyzed tissue.

Contraction-band necrosis: patterns of distribution in the myocardium and their diagnostic usefulness in sudden cardiac death

&NA; Experimentally‐produced acute regional myocardial infarcts of 2 or more hours duration show a characteristic pattern of myocardial cell alteration known as “contraction‐band necrosis”. To investigate the potential usefulness of this feature in assessing the myocardial status in sudden cardiac death, detailed histological examinations of the coronary arteries and midventricular myocardium were carried out on hearts from 70 unselected cases of sudden cardiac death. Contraction‐band necrosis was frequently encountered and occurred in 3 main patterns which could be correlated with the coronary artery pathology and the case history: (1) a regional distribution consistent with early subendocardial or transmural infarction, not yet characterized by coagulative necrosis, associated in most cases with a recent thrombotic event in the relevant supply artery (27.1%); (2) adjacent to pre‐existing infarction (recent or healed) or to subendocardial fibrosis resulting from severe coronary artery disease, consistent with recent extension of ischemic injury (18.6%); (3) a global, full‐transmural distribution, consistent with reperfusion injury from delayed resuscitation in the absence of significant coronary artery pathology (8.6%). Thus, examination of the myocardium for a specific pattern of contraction‐band necrosis may frequently facilitate the definitive diagnosis of sudden cardiac death.

Effects of ischaemia on vasculature

The effects of ischaemia on the structure and function of coronary vessels have been evaluated most thoroughly in the open-chest anaesthetised dog. In the beating heart the flow into the capillary bed is controlled by precapillary sphincters in the terminal arterioles. Short periods (up to 20 min) of ischaemia result during reperfusion in vasodilatation which is mediated through increased tissue adenosine levels. Longer periods of ischaemia (60 or more min) result in degenerative vascular changes and no-reflow during reperfusion. Endothelial cells swell, lose their pinocytotic vesicles and form spherical cytoplasmic protrusions into the capillary lumina. Prolonged ischaemia (over 3 hours) causes ruptures in microvascular walls. Such vessels become permeable to large tracers, e.g. carbon particles, and reperfusion results in haemorrhage. Capillaries in ischaemic myocardium are collapsed by compression resulting from intra- and extracellular oedema and contracture of myocytes. Erythrocytes plug collapsed microvessels. However, in vitro observations have shown that no-reflow may result from ischaemia even without erythrocyte plugging. Capillary compression is the main determinant of no-reflow after ischaemia. Degenerative ischaemic changes in vascular walls further contribute to vascular incompetence in vivo.

Comparative biochemistry and fine structure of atrial and ventricular myocardium during autolysis in vitro

SummaryIn a previous study we found that the development of fine structural alteration in atrial myocardium made ischaemicin vivo was slower than has been observed for ventricular myocardium. To explore possible reasons for this, parallel samples of atrial (A) and ventricular (V) myocardium undergoing autolysis (ischaemic necrosis)in vitro at 37°C were studied for up to 2 hours. At 15-minute intervals tissue was snap-frozen for measurement of pH, lactate, and adenine metabolites by HPLC. In half the experiments comparable specimens were taken for electron microscopic examination as well. Fine structural alteration developed less uniformly and more slowly in A than in V. The most striking metabolic differences between A and V were:(1)A had a consistently higher tissue pH and lower lactate level(2)The sum of the adenine + hypoxanthine metabolites was essentially constant bu significantly different for each (A=5.04±0.12 (s.e.m.), V=7.71±0.15 (s.e.m.) μmol/g wet tissue weight)(3)Initial ATP levels were lower (40% less) in A(4)The maximum accumulation of AMP was higher in A, despite its smaller pool of adenine metabolites(5)Both adenosine and inosine showed slower rates of change in A. These results suggest that during early, severe ischaemic injury A and V show differing activities of 5′-nucleotidase.

Changes in the Contractile state, fine structure and metabolism of cardiac muscle cells during the development of rigor mortis

SummaryTransmural slices from the left anterior papillary muscle of dog hearts were maintained for 120 min in a moist atmosphere at 37° C. At 15-min intervals tissue samples were taken for estimation of adenosine triphosphate (ATP) and glucose-6-phosphate (G6P) and for electron microscopic examination. At the same times the deformability under standard load of comparable regions of an adjacent slice of tissue was measured. ATP levels fell rapidly during the first 45 min to a relative plateau which was maintained from 45 to 75 min after excision of the heart. During a subsequent further decline in ATP, the mean deformability of myocardium fell from 30 to 12% indicating the development of rigor mortis. Conversely, G6P levels increased during the first decline in adenosine triphosphate but remained relatively steady thereafter. Whereas many of the myocardial cells fixed after 5 min contracted on contact with glutaraldehyde, all cells examined after 15 to 40 min were relaxed. A progressive increase in the proportion of contracted cells was observed during the rapid increase in myocardial rigidity. During this late contraction the cells showed morphological evidence of irreversible injury. These findings suggest that ischaemic myocytes contract just before actin and myosin become strongly linked to maintain the state of rigor mortis.