Macromolecular Leakage Research Articles

Histamine-induced protein leakage in hypertensive rats: inhibition by verapamil.

Hypertension has been associated with an enhanced transport of macromolecules from the vasculature to the interstitium. The first objective of this study was to determine if, under control conditions, there is an enhanced leakage of macromolecules from the cremaster vasculature of the hypertensive rat. The second objective was to determine if the response to a mediator of macromolecular leakage (histamine) was altered in the renovascular hypertensive rat. A third objective was to determine if a calcium entry blocker, verapamil, could inhibit histamine-induced leakage and, if so, was the sensitivity to verapamil different in the renovascular hypertensive rat. Rats were anesthetized with pentobarbital, and the cremaster preparation was used for in vivo television microscopy studies. Fluorescein isothiocyanate was tagged to rat serum albumin (FITC-RSA), and the leakage of this albumin from the vasculature to the interstitium was quantitated by the use of fluorescent microscopy techniques. There was no difference during control conditions in macromolecular leakage between the normotensive and hypertensive rats. However, histamine induced a greater leakage in the renovascular hypertensive rat than in the normotensive controls. In addition, verapamil, in the presence of normal calcium levels, inhibited the histamine-induced leakage in the hypertensive rats but not in the normotensive controls. These data suggest that enhanced macromolecular leakage during hypertension may be due to an increased sensitivity to mediators of protein leakage. These agents may produce protein leakage by enhancing entry of extracellular calcium into endothelial cells.

Differential macromolecular leakage from the vasculature of tumors

Tumor-induced neovascularization is essential for invasion, metastases, and exponential growth of solid tumors. The authors studied the differences in macromolecular leakage from the neovasculature of a fast-growing, early-metastasizing tumor, the Walker 256 carcinosarcoma, and a slow-growing, nonmetastasizing tumor, a rat chondrosarcoma. A 1-mm3 piece of the Walker 256 carcinoma or the chondrosarcoma was implanted in the cremaster muscle of rats. Five days after surgery the cremaster muscle with the implanted tumor was placed in a special bath containing Krebs solution such that the circulation and nerves from the animal to the cremaster were intact. Fluorescein isothiocyanate-labeled rat serum albumin (FITC-RSA) was injected (intra-arterially) into each rat to permit visualization of the vasculature by fluorescent microscopy. A closed-circuit television system was used to quantitate macromolecular leakage as a change in interstitial fluorescent intensity. Data are given as a relative fluorescent intensity (mean +/- standard error of the mean) in an area of the cremaster with tumor-induced neovascularization. These studies demonstrated that the vasculature induced by rapidly growing Walker 256 carcinosarcoma leak albumin freely when compared with the vasculature induced by the slow-growing chondrosarcoma. Furthermore, there was a significant increase in fluorescent intensity (albumin leakage) in the Walker tumor from 1 minute (24 +/- 3.0) to 30 minutes (49 +/- 5.6). In the normal cremaster area there was a significantly lower fluorescent intensity in the interstitium and a very slight increase with time (4 +/- 1.5 at 1 minute vs. 7 +/- 1.4 at 30 minutes). One interpretation of these data is that the mechanisms responsible for protein leakage from the vasculature of the Walker tumor may be involved in the fast growth and metastases of this tumor as compared with slower-growing tumors such as the chondrosarcoma.

Prevention of transcoronary macromolecular leakage after ischemia-reperfusion by the calcium entry blocker nisoldipine. Direct observations in isolated rat hearts.

Coronary microvascular damage appears to play a role in reperfusion injury after myocardial ischemia. This study was designed to afford direct viewing of the effects of myocardial ischemia-reperfusion on the coronary microcirculation and to determine whether pretreatment with the calcium blocker nisoldipine would attenuate any microvascular damage during reperfusion. Four groups of isolated rat hearts were perfused with a solution that contained red cells and fluorescent albumin, but was essentially free of platelets and leukocytes. Group I served as a nonischemic control. Group II hearts were subjected to 30 minutes of no-flow ischemia followed by reperfusion. Group III hearts were pretreated with nisoldipine (1 microgram/min) for 5 minutes before ischemia, and group IV hearts were treated with nitroglycerin (93 micrograms/min) before and after ischemia to mimic the vasodilation caused by nisoldipine. Perfused coronary capillarity and transcoronary extravasation of plasma albumin were measured by direct visualization techniques before and after ischemia. For group I, there was no significant change in coronary resistance, perfused capillarity, or transcoronary extravasation with time. For both groups II and IV, ischemia-reperfusion caused no increase in coronary resistance, but a significant decrease in perfused capillarity and a marked increase in transcoronary extravasation of fluorescent albumin (P less than 0.05). The nisoldipine group (group III) demonstrated a similar decrease in perfused capillarity but no increase in protein extravasation during reperfusion. These results indicate that, in the heart, platelets and/or leukocytes are not absolutely necessary to induce either the no-reflow phenomenon or the permeability damage observed during reperfusion after ischemia. The protective effect of treatment with nisoldipine appeared to be independent of vasodilation. We speculate that this calcium blocker reduced endothelial uptake of calcium during reperfusion, preventing endothelial deformation and formation of interendothelial gaps.

Microvascular effects of anaphylatoxins C3a and C5a.

The direct microvascular effects of human C3a and C5a were investigated by using hamster cheek pouches prepared for intravital microscopy. Topical application of 10 nM C3a resulted in pronounced microcirculatory alterations, characterized by vasoconstriction, platelet aggregation, and an increase in macromolecular leakage at postcapillary venules, as assessed by extravasation of intravascular fluorescein-labeled dextran (m.w. 150,000). Exposure of the preparation to 500 nM COOH-terminal octapeptide analogs of C3a resulted in a microvascular response almost identical to that of C3a, supporting the view that the active site of this anaphylatoxin resided within the COOH-terminal portion. Changes similar to those caused by C3a were also induced by 20 or 100 nM C5a and, in addition, the higher concentration of C5a caused accumulation of polymorphonuclear leukocytes (PMNL) in small venules and somewhat prolonged vascular leakage from venules exhibiting PMNL accumulation. Histamine was found to partially mediate the vascular leakage induced by C3a and the initial (first 5 min) permeability response to the high concentration of C5a, whereas the subsequent leakage induced by the latter anaphylatoxin was unaffected by mepyramine pretreatment. In neutropenic and mepyramine-treated animals exposed to the high concentration of C5a, a partial reduction of both the early and the subsequent vascular leakage was seen, indicating that accumulated PMNL play a role in the prolonged phase of leakage. The pronounced microvascular alterations induced by low concentrations of C3a and C5a strengthen the view that these anaphylatoxins act as mediators of inflammatory reactions in which the complement system is activated.

Both protein and blood cells reduce coronary microvascular permeability to macromolecules.

A number of studies report that isolated Krebs-perfused hearts deteriorate with time, develop edema, and demonstrate a progressive increase in coronary vascular resistance. However, hearts perfused with filtered blood are more stable with regard to cardiac function and coronary resistance. The changes observed in the Krebs hearts may be due to a "permeability"-type edema. The purpose of this study was to systematically determine whether adding protein and blood cells to a Krebs perfusate affected coronary microvascular permeability to macromolecules. The rat heart preparation employed allowed direct visualization and quantification of transcoronary macromolecular leakage. We observed severe transcoronary leakage of fluorescent albumin (FITC-BSA) when FITC-BSA was later added after 20 min of perfusion with Krebs. Leakage was decreased by including 2 g/100 ml albumin (BSA) in the initial perfusate but was not further reduced by increasing the BSA concentration to 5 g/100 ml. However, adding washed blood cells to the initial perfusate did further reduce FITC-BSA leakage. The index of FITC-BSA exchange, the O/I ratio, was 0.70 +/- 0.02 (+/- SE) for Krebs perfusate, 0.55 +/- 0.03 for Krebs-BSA, and 0.45 +/- 0.02 for Krebs-BSA-blood cells, indicating significant effects for both protein and blood cells (P less than 0.05). The results suggest that both protein and blood cells are necessary to maintain the semipermeable characteristics of the coronary exchange vessels.

Quantitation of vasodilator-induced macromolecular leakage by in vivo fluorescent microscopy

In vivo television microscopy of the rat cremaster muscle was used to investigate the phenomenon of drug-induced protein leakage from the vasculature. Concentration-response curves of the vasodilators, isoproterenol, histamine, and nitroprusside, were used to determine if vasodilation of this skeletal muscle vasculature would lead to or be associated with protein leakage. This objective required development of the methodology to quantitate protein leakage that occurs in any one vascular area of the microcirculation. Fluorescein isothiocyanate-tagged rat serum albumin was injected into the rats (ia) and an argon laser was used for excitation of the fluorochrome. The fluoroscent image was recorded on videotape for later analysis. Vascular diameters were measured off the television monitor. Protein leakage was quantitated by determining the change in interstitial fluorescent intensity that occurred during the experiment. These studies demonstrated that protein leakage started within 1 min of topical application of histamine and peak response occurred in 5–7 min. Nitroprusside and histamine produced protein leakage but isoproterenol just produced vasodilation. Furthermore, significant differences in pD2 values (−log ED 50), which were used as a measure of drug sensitivity, indicated that protein leakage and vasodilation were probably not related phenomena for histamine and nitroprusside. Thus, this method of quantitation of protein leakage in the microcirculation of the rat cremaster muscle, gave reproducible data that can be used to determine the time course of protein leakage and to compare concentration-dependent phenomena.

Immunopathology of Psoriasis: A Comparison with Other Parakeratotic Lesions

Characteristic in vivo binding of IgG and C3 to the stratum corneum of psoriatic lesions has been implicated as a significant event in the pathogenesis of the disease. Immunofluorescent findings of psoriatic lesions are compared with other parakeratotic and nonparakeratotic lesions. Parakeratotic lesions studied include verrucae vulgares in patients with psoriasis, verrucae vulgares in patients without psoriasis, actinic keratoses and lichen simplex chronicus. Psoriatic lesions and other parakeratotic lesions demonstrate similar immunopathological phenomena within the involved stratum corneum: (1) A high percentage of the lesions stain positively with one or more of the immunoglobulins (IgG, IgM and IgA). (2) No predominance of any immunoglobulin class is found. (3) The predominant sites of immunoglobulin deposition are within the regions of parakeratosis. (4) C3 deposition is found in a high percentage of the lesions and is present at the sites of immunoglobulin binding. By contrast only a small percentage of nonparakeratotic lesions display stratum corneum fluorescence. These data are consistent with macromolecular leakage through the lesional microvasculature and high affinity binding of immunoglobulins by parakeratotic stratum corneum. Similar immunofluorescent findings in psoriatic lesions and nonpsoriatic parakeratotic lesions suggest that immunoglobulin binding in psoriatic lesional stratum corneum is not a significant event in the pathogenesis of psoriatic lesions.

Cytochemistry of brain amyloid in adult dementia.

A cytochemical study of 14 cases of adult dementia revealed the presence of gamma globulin in the amyloid of three cases of congophilic angiopathy by means of immunofluorescence microscopy. In these cases, the additional identification of human albumin is regarded to indicate a non-specific macromolecular leak in the blood-brain barrier. Both reactions are inhibited by prior absorption with the appropriate serum protein. Twelve of the 14 cases had congophilic amyloid deposits which were not affected by permanganate pre-treatment, so that immunoglobulin content remains a possibility, despite the negative immune reaction. Alcianophilia was studied at a varying pH and electrolyte concentration, but these findings do not appear to have nosologic significance. The three positive cases are characterized by a rapid terminal decline. The heterogeneity of amyloid and the significance of immunoamyloid in the pathogenesis of adult dementia is discussed.

Simultaneous measurements of macromolecular leakage and arteriolar blood flow as altered by PGE 1 and β2-receptor stimulant in the hamster cheek pouch

Alterations in arteriolar blood flow and macromolecular leakage after prostaglandin (PGE 1) administration in the presence and absence of a β-receptor stimulant (terbutaline) were investigated in the hamster cheek pouch preparation by direct observation. After iv injection of FITC-dextran ( M w = 150,000 ; 25 mg 100 g −1 body wt), macromolecular leakage was quantified as the number of leakage sites per square centimeter appearing in the cheek pouch upon topical application of PGE 1 (0.1, 1.0, 10.0, and 100.0 ng ml −1) in the presence and in the absence of terbutaline (1.0 μg ml −1) in the superfusion solution. Concomitantly, arteriolar blood flow was calculated from measurements of red blood cell velocity and vessel diameter. PGE 1 produced a dose-dependent increase in macromolecular leakage during normal superfusion while this effect was reduced significantly when terbutaline was present. PGE 1 produced an increase in arteriolar blood flow which was not affected by terbutaline. Terbutaline, alone, reduced macromolecular leakage and increased arteriolar blood flow. Thus, it is concluded that terbutaline counteracts PGE 1-induced macromolecular leakage. This effect is not mediated through a reduction of the PGE 1-induced increase in arteriolar blood flow.

Intravital and electron microscopic study of bradykinin-induced vascular permeability changes using FITC-dextran as a tracer.

Fluorescein-labelled dextran (FITC-dextran) of molecular weight 145,000 was used to study vascular permeability to macromolecules by intravital and electron microscopy. Anaesthetised hamsters prepared for intravital observation of the cheek pouch microvasculature were given an intravenous injection of FITC-dextran. Leakage of macromolecules was induced by topical application of bradykinin to the cheek pouch microvasculature and observed in fluorescent light. Leakages occurred only from postcapillary venules of a diameter well below 50 micrometer. The cheek pouch preparation was rapidly fixed by immersion and samples of tissue with intravitally identified leakages of FITC-dextran were studied by electron microscopy. FITC-dextran appeared as black precipitates in the vascular lumen and also outside the lumen in bradykinin-treated animals. In most animal; gaps were found between endothelial cells and these gaps contained dextran precipitates. The results support much other evidence that bradykinin induces macromolecular leakage by opening gaps between endothelial cells in postcapillary venules.

Topographic Diagnosis of Gastrointestinal Protein Loss by Means of Radioiodine-labelled Albumin and IgG

A method has been worked out to determine the site of plasma protein loss in proteinlosing gastroenteropathies. It uses the ratio between faecal excretion of 125I and 131I following simultaneous i.v. injection of 125I-IgG and 131I-albumin. 165 cases were studied (Menetrier's disease, 12 cases; intestinal lymphangiectasia, 19; adult coeliac disease, 6; other small-intestinal diseases, 5; chronic inflammatory bowel disease, 115 (83 of whom had Crohn's disease and 32 ulcerative colitis); and neoplastic diseases, 8 cases). Gastrointestinal macromolecular leakage was detected and semiquantitated by means of 51CrCl3 (33 cases) or 59Fe-iron dextran (90 cases). Faecal 59Fe clearance was significantly (p 200g/day) in the other groups. Faecal radioiodide ratio (125I/131I) was inapplicable in 2...

Bradykinin and prostaglandin E 1, E 2 and F 2α-induced macromolecular leakage in the hamster cheek pouch

The hamster cheek pouch prepared for intravital observations on macromolecular permeability with fluorescein labelled dextran was used in four series of 5 hamsters each, all pretreated with indomethacin. Bradykinin, PGE 1, PGE 2 and PGF 2α increased macromolecular leakage at postcapillary venules, and this leakage was reversible on removal of agent. A linear relation was found between the logarithmic value of dose of bradykinin and the mean number of leakage sites. No tachyphylaxis to bradykinin was seen. The effect of either PGE 1, PGE 2 or PGF 2α applied simultaneously with bradykinin was to significantly (p<0.05) potentiate the bradykinin response. Bradykinin and these prostaglandins appeared to have the same site of action for their effect of increasing permeability, e.g. the postcapillary venule.

Inhibition of bradykinin induced macromolecular leakage from post-capillary venules by a beta2-adrenoreceptor stimulant, terbutaline.

Acta Physiologica ScandinavicaVolume 101, Issue 4 p. 504-506 Inhibition of Bradykinin Induced Macromolecular Leakage from Post-Capillary Venules by a β2-Adrenoreceptor Stimulant, Terbutaline E. Svensjö, E. Svensjö Department of Experimental Medicine, Pharmacia AB, Uppsala and the Research Department, Draco AB, Lund, SwedenSearch for more papers by this authorC. G. A. Persson, C. G. A. Persson Department of Experimental Medicine, Pharmacia AB, Uppsala and the Research Department, Draco AB, Lund, SwedenSearch for more papers by this authorG. Rutili, G. Rutili Department of Experimental Medicine, Pharmacia AB, Uppsala and the Research Department, Draco AB, Lund, SwedenSearch for more papers by this author E. Svensjö, E. Svensjö Department of Experimental Medicine, Pharmacia AB, Uppsala and the Research Department, Draco AB, Lund, SwedenSearch for more papers by this authorC. G. A. Persson, C. G. A. Persson Department of Experimental Medicine, Pharmacia AB, Uppsala and the Research Department, Draco AB, Lund, SwedenSearch for more papers by this authorG. Rutili, G. Rutili Department of Experimental Medicine, Pharmacia AB, Uppsala and the Research Department, Draco AB, Lund, SwedenSearch for more papers by this author First published: December 1977 https://doi.org/10.1111/j.1748-1716.1977.tb06038.xCitations: 78AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Citing Literature Volume101, Issue4December 1977Pages 504-506 RelatedInformation

The effect of inhibition of prostaglandin synthesis on microvascular haemostasis and macromolecular leakage

A study has been made of haemostasis and permeability in rabbits in which prostaglandin synthesis inhibition was induced by indomethacin. Prostaglandin activity of the mesentery increased with time after preparation. Prostaglandin synthesis was completely inhibited and micromolecular leakage in post-capillary venules diminished after indomethacin treatment. PGE 1 prolonged haemostatic plug formation time whereas indomethacin decreased venular haemostatic plug formation time, abolishing the difference normally seen between arterioles and venules. It is suggested that this difference is due to prostaglandin induced inhibition of platelet aggregation on the venular side of the microcirculation.