Cytochemical demonstration of sites of hydrogen peroxide generation and increased vascular permeability in isolated pig hearts after ischaemia and reperfusion

Isolated pig hearts, subsequently perfused with pig or human blood, were prepared for the cytochemical demonstration of sites of hydrogen peroxide generation and increased vascular permeability. Oxidant stress was associated with ultrastructural changes commonly seen following myocardial reperfusion. In addition, the precipitation of cerium perhydroxide following perfusion with physiological saline containing cerium chloride suggested the vascular endothelium and leukocytes as sources of oxidants. This was associated with rapid penetration of horseradish peroxidase through the intercellular clefts of the vascular endothelium into the interstitial space, suggesting increased vascular leakiness at these sites. The rapid penetration of horseradish peroxidase was observed at all monitored periods of reperfusion with pig or human blood. This indicates that the increased permeability occurred during the ischaemic period and continued during reperfusion. Morphological damage was greatest in pig hearts reperfused with whole human blood and this was attenuated if the blood was preabsorbed to remove antibodies prior to reperfusion. We conclude that oxidant stress was initiated during ischaemia and continued during reperfusion in this model.     

Effect of postischaemic hypothermia on the mitochondrial damage induced by ischaemia and reperfusion in the gerbil

In order to test the effect of hypothermia on mitochondrial function damage following cerebral ischaemia/reperfusion, Mongolian gerbils were submitted to 30 min bilateral carotid occlusion and 2 h of reperfusion at 37 degreesC or 30 degreesC. After normothermic (37 degreesC) ischaemia/reperfusion, significant decreases in mitochondrial state 3 (+ADP) oxygen consumption (-42.2%), complex II-III activity in synaptosomes (-31.7%) and complex IV were measured, in both free mitochondria and synaptosomes (-30.3% and -27. 8% respectively). However, following hypothermic (30 degreesC) reperfusion, both respiration rates and all enzyme activities remained at levels not significantly different from those in the sham operated controls.     

Goblet cell carcinoid of the appendix. An ultrastructural and histochemical study

In a patient with a goblet cell carcinoid tumor of the appendix, light and electron microscopical studies demonstrated mucinous material and enterochromaffin granules within the same cell. Transitions between cells containing primarily mucin and cells containing numerous argentaffin granules were observed. Fluorescence studies demonstrated that the tumor cells contain biogenic amines.     

Contractile dysfunction caused by normothermic ischaemia and KCL arrest in the isolated pig heart: a 31P NMR study

The aims of this study were to assess (1) whether contractile dysfunction caused by ischaemia under hyperkalaemic conditions ("cardioplegic ischaemia") is associated with impaired energy production or abnormalities in regulation of contractility and (2) whether hyperkalaemia itself contributes to contractile dysfunction. We used 31P and 23Na NMR spectroscopy in conjunction with measurements of mechanical function and oxygen consumption in Langendorff perfused pig hearts to evaluate the mechanism of contractile failure caused by (1) total global cardioplegic (17 mM [K+]) ischaemia (36 degrees C, 50 min KCl arrest, 45 min ischaemia, 20 min reflow with high KCl) and (2) KCl arrest alone (115 min) without flow cessation. KCl arrest plus ischaemia and subsequent reperfusion (Group I) resulted in decreases in ATP (mean +/- S.D.; 61 +/- 13% of initial, n = 5; P < 0.01) and pressure-rate product (PRP) (31 +/- 9%, n = 17; P = 0.0001) while phosphocreatine (PCr), Pi, total creatine (Cr) and intracellular Na+ levels were unaffected. KCl arrest itself (Group II, n = 6) did not affect PCr, ATP or total Cr levels but decreased the PRP to 59 +/- 12% (P < 0.001). Oxygen consumption rates (Vo2) were reduced in both groups to similar levels (67 +/- 18, P < 0.01 and 77 +/- 13%, P < 0.02, respectively). The efficiency of energy conversion to mechanical work (PRP/delta VO2) decreased to 51 +/- 15 (P < 0.001) and 67 +/- 13% (P < 0.012) of initial levels, respectively. To assess metabolic and contractile reserves of post-ischaemic (n = 7) and KCl-treated (n = 3) hearts, the effects of isoproterenol (Iso) and increased Ca2+ were compared with those in normal beating hearts (Group III, n = 3). In all groups treatment with Iso (0.1 micron) greatly increased PRP (to 526 +/- 116, 203 +/- 16 and 198 +/- 8% of the level prior to stimulation (baseline), P < 0.01, respectively) and Vo2 (162 +/- 9, 153 +/-16 and 128 +/-10% of baseline, P < 0.05, Respectively). Increasing [Ca2+] from 1 to 1.66 mM produced less stimulation than Iso: PRP increased to 195 +/- 23, 156 +/- 13 and 163 +/- 22% (P < 0.05) and Vo2 increased to 138 +/- 22 (P < 0.05), 115 +/- 4 and 120 +/- 10% of baseline in Groups I, II and III, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)     

Downregulation of calpastatin in rat heart after brief ischemia and reperfusion

The activities of calpain and its endogenous inhibitor, calpastatin, were measured in the soluble fraction of perfused rat heart after ischemia for 5-20 min and reperfusion for up to 30 min. The method for m-calpain measurement was modified: washing of the DEAE-cellulose column with 0.18 M NaCl instead of 0.15 M NaCl increased the m-calpain activity 12.5-fold. Ischemia for 20 min followed by reperfusion for 30 min did not affect the m-calpain activity but decreased the calpastatin activity. m-Calpain was enriched in the nucleus-myofibril fraction but was not further translocated on ischemia-reperfusion. Mu-calpain was below the limit of detection on immunoblotting or casein zymography, but its mRNA was substantially expressed, as detected on Northern blotting. Casein zymography also revealed a novel Ca2+-dependent protease without the typical characteristics of mu- or m-calpain. The immunoblotting of myocardial fractions showed that calpastatin was proteolyzed on ischemia-reperfusion. The calpastatin proteolysis was suppressed by a calpain inhibitor, Ac-Leu-Leu-norleucinal. Calpastatin may sequester calpain from its substrates in the normal myocardium, but may be proteolyzed by calpain in the presence of an unidentified activator in the early phase of calpain activation during ischemia-reperfusion, resulting in the proteolysis of calpastatin and then other calpain substrates.     

Receptors for Helix pomatia agglutinin on normal and neoplastic lymphocytes -- a histochemical study using paraffin embedded tissue sections

In order to investigate the distribution and significance of receptors for Helix pomatia agglutinin (HPA), paraffin sections from several reactive and 70 neoplastic lymphoid tissues were utilized for a histochemical study after neuraminidase treatment. Lymphocytes in the germinal center and thymic cortex were mostly negative for the receptors, while most of the lymphocytes in the thymus-dependent area, some of those in the primary follicle or in the marginal zone and most of the plasma cells in the medulla possessed the receptors on their surfaces. Nodular lymphomas, thymomas, and diffuse lymphoblastic lymphomas were mostly negative in contrast to the positive reaction in diffuse poorly differentiated lymphomas of B-cell type and plasmocytomas. Diffuse histiocytic lymphomas and diffuse mixed lymphocytic and histiocytic lymphomas occasionally possessed the receptors, irrespective of their immunological markers. Hodgkin's cells were negative on their surface, but were occasionally positive in their cytoplasms ans seen in case of histiocytes. From these results, HPA receptors could be neither T-cell nor B-cell differentiation marker. The possibility that HPA-receptor-negative lymphocytes may hav a quick turn-over rate or a short life span is also discussed.     

Effects of ischaemia and reperfusion on vasoactive neuropeptide levels in the canine infrarenal aortic revascularization model

Infrarenal aortic cross-clamping is associated with remote vascular events, including myocardial infarction and renal insufficiency. The purpose of this study was to determine whether hindlimb ischaemia and reperfusion associated with infrarenal aortic cross-clamping results in the production of vasoactive regulatory neuropeptides. A canine model of infrarenal aortic cross-clamping was used for the study. Serial blood samples were drawn, prior to, at the time of, and serially following placement of the clamp and subsequent release of the clamp and reperfusion. Ischaemia resulted in increased mean(s.e.m.) plasma levels of neuropeptide Y (NPY) (initial 10.0(1.8) pmol/l versus ischaemia 24.7(2.3) pmol/l, P < 0.001) and vasoactive intestinal polypeptide (VIP) (initial 2.53(0.5) pmol/l versus ischaemia, 7.3(1.3) pmol/l, P < 0.05). Reperfusion produced three-fold elevation of VIP (initial 2.5(0.5) pmol/l versus reperfusion 9.6(1.5) pmol/l, P < 0.001), two-fold elevation in the plasma levels of endothelin-1 (initial 1.3(0.1) pmol/l versus reperfusion maximum 2.5(0.3) pmol/l, P < 0.01) and NPY (initial 10.0(0.8) pmol/l versus reperfusion maximum 23.9(2.3) pmol/l, P < 0.001). Ischaemia and reperfusion did not alter calcitonin gene-related peptide (CGRP) (a potent vasodilator) levels. Endothelin-1 (ET-1) plasma levels were also increased following haemorrhagic shock (initial 1.3(0.1) pmol/l versus exsanguination 3.4(0.4) pmol/l, P < 0.001), but not during ischaemia (initial 1.3(0.1) pmol/l versus ischaemia maximum 1.7(0.2) pmol/l, P = 0.7). It was concluded that vasoactive regulatory peptides are released following ischaemia, reperfusion and shock in the canine infrarenal aortic revascularization model and, therefore could contribute to remote vascular events observed with infrarenal aortic cross-clamping.     

A new approach to the understanding of the mechanism of ischemia/reperfusion damage in the heart and the effects of anti-ischemic drugs

The classical understanding of the mechanism of anti-anginal or anti-ischemic drugs is an increase in blood supply to the heart and/or a decrease in oxygen consumption of the heart, maintaining energy balance in the heart between supply and demand and hence maintaining the tissue levels of high-energy phosphates. This scheme is reasonable. During reperfusion following ischemia, however, there is more serious damage to the heart, although the tissue levels of high-energy phosphates increase. This is probably because toxic substances are generated in the heart during ischemia/reperfusion. We propose that both lysophosphatidylcholine and palmitoyl-L-carnitine that accumulate in the myocardium during ischemia/reperfusion are candidates for the toxic substances that accelerate ischemia/reperfusion damage to the heart. Therefore, drugs that have anti-lysophosphatidylcholine and/or anti-palmitoyl-L-carnitine effects are promising for the treatment of ischemic heart diseases. We found that K-7259, a novel derivative of dilazep having a minimal effect on the normal heart, is a drug that attenuates the deleterious effects of both lysophosphatidylcholine and palmitoyl-L-carnitine on the heart, and therefore attenuates the ischemia/reperfusion damage.     

Histological and ultrastructural observations on the development of the lung of the fetal pig

Lungs of fetal pigs having gestational ages ranging from 80 to 115 days were examined histologically and by electron microscopy. At 80 days bronchial epithelium was ciliated but bronchiolar cells were not and bronchial mucosal glands were absent. Peripheral regions consisted predominantly of mesenchymal tissue with glandular alveoli. 92 days marked the transition from the immature to the more mature lung type. Bronchial glands appeared and began to grow from the epithelium into the lamina propria, bronchiolar epithelial cells acquired cilia, and alveoli were becoming irregular in shape and had thinner interalveolar septa. Close contact between capillaries and alveolar epithelium established the blood-air barrier at many points. Differentiation of alveolar epithelium into types I and II pneumonocytes occurred at this stage and lamellated osmiophilic inclusion bodies were present in type II cells for the first time. The number of lamellated bodies increased progressively to term at 115 days.     

The effect of mannitol on reperfusion after severe induced subtotal ischemia of the leg. A hemodynamic, biochemical and metabolic study in a pig hind limb model

Revascularization of acutely ischemic muscle tissue is followed by edema, decreased oxygen utilization, increased vascular resistance and massive efflux of intracellular compounds indicating loss of cellular integrity, with resultant irreversible damage. In this pig hindlimb study, 8 pigs were submitted to standardized subtotal ischemia of one leg and mannitol was administered i.v. prior to and during 2.5 hours of reperfusion. Compared to 9 controls, the mannitol treated pigs had increased blood flow to the legs, increased oxygen consumption and decreased release of intracellular compounds (CK). This indicates that mannitol attenuates the ischemia/reperfusion syndrome. Muscle energy metabolic parameters showed a similar response to ischemia for both mannitol-treated pigs and controls and no differences in recovery were detected during 2.5 hours of reperfusion between the groups.     

Effect of ascorbic acid on infarct size in experimental focal cerebral ischaemia and reperfusion in a primate model

The iliac artery-ureteral fistula is a rare complication which becomes symptomatic with life-endangering bleeding. A typical feature is its intermittent occlusion, which makes diagnosis very difficult. Without knowledge of the correct diagnosis the rate of morbidity, through functional loss of the kidneys, and of mortality can be extremely high. In the following a case is described in which, after radiotherapy and chemotherapy following sigmoid resection due to carcinoma, urinary congestion of the remaining functional kidney occurred during the course of treatment. Following the placing of an endoureteral stent an iliac-ureteral fistula with massive bleeding developed. It was demonstrated by means of angiography. For the first time, therapy with endoluminal stent grafts was successfully applied.     

Scintigraphic assessment of silent myocardial ischaemia after early infarction using myocardial SPET imaging with 201Tl and 123I-MIBG

To test the hypothesis that myocardial sympathetic denervation reflects silent myocardial ischaemia early after infarction, 12 patients with myocardial infarction but without post-infarction angina pectoris underwent single photon emission tomography (SPET) at rest with 201Tl and 123I-metaiodobenzylguanidine (MIBG) shortly after and 3 months after infarction. Short-axis SPET images at the basal, mid-ventricular and apical portions of the left ventricle were selected, and each short-axis image was divided into eight segments. Tracer uptake in each of the 24 segments was scored using a 4-point scale. The total score in each segment was calculated as the defect score for each image, and the difference between the total defect score for the 201Tl and 123I-MIBG images was calculated as the delta defect score. All 12 patients underwent exercise stress 201Tl scintigraphy 1 month after infarction, and they were divided into two groups: those patients with (Group A, n = 7) and those patients without (Group B, n = 5) transient perfusion defects in the peri-infarcted region without chest pain. For the 123I-MIBG defect score, a marked reduction at 3 months was observed in Group A (24 +/- 12 vs 13 +/- 6; P < 0.01), whereas the defect score remained unchanged in Group B (25 +/- 7 vs 23 +/- 8; N.S.). The delta defect score was significantly reduced in Group A (10 +/- 5 vs 6 +/- 4; P < 0.05), whereas it remained unchanged in Group B. The 123I-MIBG defect score early after infarction was higher than the exercise-induced 201Tl defect score (24 +/- 12 vs 20 +/- 9; P < 0.01), whereas at 3 months post-infarction it was lower than the exercise-induced 201Tl defect score (13 +/- 6 vs 20 +/- 9; P < 0.05). Moreover, effort chest pain during daily activities was noted in 5 of the 7 (71%) patients in Group A within 3 months post-infarction. The results of this study suggest that viable but denervated myocardium (mismatched 123I-MIBG defects) is present in peri-infarcted regions, and that myocardial sensory nervous disturbance, which may co-exist with sympathetic nervous denervation, may induce silent myocardial ischaemia in patients with myocardial infarction.