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OBJECTIVE: Isolated rat hearts were used to determine whether structural changes in the coronary microcirculation caused by diabetes mellitus occur soon after the onset of hyperglycemia and whether early changes might be reversed by return to normoglycemic conditions. METHODS: Three experimental groups of hearts were used. The perfusate was Krebs-Henseleit medium with the addition of (a) 25 mmol/L glucose for 60 minutes, (b) as (a) followed by 5 mmol/L glucose for 30 minutes, and (c) hearts from diabetic rats perfused for 15 minutes. All the hearts were fixed and processed for electron microscopy. Measurements were made of various capillary dimensions and basal lamina thickness. RESULTS: The high glucose concentration caused an overall reduction in capillary dimensions which was partly reversed by reperfusion with 5 mmol/L glucose. Diabetic hearts showed enlarged whole capillary and luminal area and thinning of the endothelial cells without any change in cross-sectional area. Endothelial gaps, consistent with increased permeability, were observed in the high glucose group but not in diabetic hearts. The basal lamina was thickened in all three experimental groups compared with controls. CONCLUSIONS: The results show that morphological changes in the microcirculation occur much earlier than was previously thought and, with the possible exception of the basal lamina thickening may be reversible.

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The aim of this study was to investigate the effect of a moderate degree of hypoxia on coronary vascular permeability to various lipophobic solutes. Using the multiple indicator dilution method the permeability-surface area (PS) products were determined for 125I-albumin, 125I-insulin and 57Co-cyanocobalamin in perfused rat hearts (flow approximately 10 ml.min-1.g-1) either with well-oxygenated (pO2 approximately 96 kPa) or hypoxic (pO2 approximately 45 kPa) solutions. The PS products for albumin, insulin and cyanocobalamin during the well-oxygenated equilibration period were 0.20 +/- 0.03, 0.29 +/- 0.06 and 2.0 +/- 0.3 ml.min-1.g-1 (mean +/- SE), respectively, relative to 131I-gamma-globulin. The PS products for these solutes 15 min after the induction of hypoxia were 1.3 +/- 0.3, 0.8 +/- 0.1 (p < 0.05) and 2.1 +/- 0.2 (p < 0.05), respectively. In hearts perfused with well-oxygenated solution for 75 min, the PS products for these solutes remained stable throughout the period of the study. Electron-microscopic examination of hypoxic tissues showed the presence of endothelial gaps of approximately 1 micron which were underlined by an intact basal lamina. We conclude that a moderate degree of hypoxia produces a large increase in permeability of albumin and insulin but has no effect on the PS products for cyanocobalamin and that the endothelial gaps are the likely mechanism of the observed increase in permeability.

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1. The multiple indicator dilution method was used to study the transvascular movement of gamma-globulin, bovine serum albumin, insulin and cyanocobalamin in the isolated rat heart. 2. Perfusion of the heart with well-oxygenated solution for 75 min (constant flow) did not produce a significant change either in the total area under the dilution curve or the 'leakage index' (an arbitrary measure of transvascular flux) for all the tracers. 3. Perfusion of the heart with hypoxic solution produced a significant increase in leakage of gamma-globulin of 38.6 +/- 18, 48.5 +/- 17.6, 60.5 +/- 24 and 58 +/- 20% after 15, 30, 45 and 60 min, respectively, compared with the well-oxygenated equilibration period. Permeability- surface area products (PS) for the smaller diffusible solutes, therefore, could not be estimated. 4. The flux of albumin, insulin and cyanocobalamin in response to hypoxia was similar to that of gamma-globulin. 5. Ultrastructural examination of well-oxygenated hearts revealed that Monastral Blue-labelled albumin remained within the lumen and that endothelial integrity remained intact. 6. Conversely biopsies from hypoxic hearts showed that the labelled albumin had passed to the interstitium through gaps (approximately 3 microns) in venular endothelium. 7. The results showed that, in intact hearts, hypoxia produced gaps in the endothelium of venules and that these gaps could be the possible route for transvascular leakage of macromolecules.

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A technique has been developed to monitor changes in synovial vascular tone and leakage of macromolecules, two of the cardinal features of inflammation. The technique permits not only quantitative sequential measurements of the leakage of 125I-albumin from the circulation into the synovium, but also semiquantitative continuous monitoring of the leakage changes. It allows simultaneous continuous monitoring of changes in synovial vascular tone as well. Vascular protein leakage was monitored semiquantitatively by measuring the radioactivity of 125I-albumin in synovial perfusate when passed through a flow cell consisting of a plastic tubing coil situated inside the well of a gamma detector. Quantitative measurements were provided by collecting and counting the synovial perfusate for 125I-albumin. Synovial vascular tone was monitored continuously by measurement of the change in joint radiation emitted by in vivo-labelled 99m-Tc erythrocytes. The in vivo labelling procedure yielded an 86% +/- 9% labelling efficiency. The degree of erythrocyte labelling was stable throughout the course of the experiment. Using this technique, the abilities of histamine to induce changes in synovial vascular tone and protein leakage were examined. Intraarticular infusion of 1 ng/mL histamine produced a 61% +/- 24% increase in synovial vascular blood volume, but did not significantly increase 125I-albumin leakage above that produced by Krebs solution alone. Higher concentrations produced a concentration-related increase in the leakage of 125I-albumin, which was not accompanied by appreciable increases in synovial vascular blood volume above the basal line. The simultaneous and continuous monitoring provided by this technique has revealed an apparent dissociation between vascular tone and vascular protein leakage in response to intraarticular histamine.

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Increased levels of amidase acting on a tissue-kallikrein selective substrate, Val.Leu.Arg.pNA, with an activity optimum at pH9, were detected in blood-free inflamed tissues from adjuvant arthritic rats (p less than 0.01). The component of this activity resistant to inhibition by soybean trypsin inhibitor (SBTI) also greatly increased (p less than 0.05). Both the SBTI-sensitive and SBTI-resistant components were inhibited by aprotinin (93% and 72% respectively). Kallikrein-like amidase also increased in inflamed synovia from seropositive rheumatoid, and osteoarthritic dogs when compared with healthy canine synovia. This increase was parallelled by an increase in kinin-forming enzyme which was also measured in rheumatoid and healthy animals and this activity was inhibited 72% by aprotinin. Total kallikrein-like amidase also increased 989% (p less than 0.05) in synovia from seropositive rheumatoid human patients, compared with healthy synovial tissue. Evidence is presented indicating that the origin of this enzymic activity may be plasma kallikrein.

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A single sub-plantar injection of 1% carrageenan in saline produced a bi-phasic paw volume increase, phase 1 plateauing at 0.5-1 h, phase 2 at 3-6 h. Subtraction of the non-specific contralateral paw response produced by saline injection revealed a monophasic carrageenan response. Mepyramine maleate at a dose maximally effective on histamine-induced paw oedema (1 mg X kg-1 i.p.) failed to suppress carrageenan-induced inflammation. Maximally effective doses of cimetidine HCl (1-100 mg X kg-1) suppressed the response by no more than 53%. In animals treated with cimetidine HCl (1 mg X kg-1), pretreatment with indomethacin (2.5 mg X kg-1) further suppressed the mean oedema response by 59% compared with cimetidine alone. The indomethacin dose suppressed mean volume increase by 41%. Mepyramine alone or combined with cimetidine showed no anti-inflammatory activity. Following pretreatment with indomethacin (2.5 mg X kg-1), mepyramine maleate further suppressed the carrageenan oedema by 41% compared with indomethacin alone. While supporting a role for prostanoid formation, or other indomethacin-sensitive mechanisms, the results show that the mechanism of carrageenan-induced rat paw oedema also involves histamine acting largely through H2-receptors, although a role for H1-receptors can not be excluded. The interaction between mepyramine and indomethacin is discussed.

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Adjuvant arthritis in rats was induced by a single subcutaneous injection of Freund adjuvant into a hind paw. The injected paws' mean volume increased continuously by 80 +/- 26% after 6 h (acute local response). The injected paw displayed a biphasic response peaking at day 1-2 decreasing to day 5, then increased to day 14 by 123% while the contralateral non-injected paw showed little increase until day 10 and increased by 42% at day 14. The lateral diameter of both knee-joints also increased biphasically, peaking at day 4, decreasing to day 6, then increasing to day 14 (early and late systemic phase). Mepyramine maleate, 1 mg.kg-1 (i.p.) daily, suppressed only the early systemic phase by 50%. Daily cimetidine hydrochloride, 1 mg.kg -1.day-1, almost completely abolished the inflammatory response in the knee joints up to day 12, while at day 14 the suppression was 71%. The increase in mean paw volume was also suppressed by orally administered cimetidine. Intraperitoneal doses of cimetidine (3.5 mumol.kg-1) totally suppressed paw oedema produced by subplantar injection of histamine (1.8 mM) but had no action on equiactive dose of the specific H1-agonist, 2-pyridylethylamine. Mepyramine at 3.5 mumol.kg-1 or greater produced no more than 50% suppression of the histamine response. A component of both adjuvant and histamine-induced responses thus appears to be histamine H2-receptor mediated.

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