Effect of Diet and Cold Exposure on Norepinephrine Turnover in Brown Adipose Tissue of the Rat

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Abstract

Brown adipose tissue (BAT) is an important site of adaptive changes in thermogenesis in the rat. The sympathetic nervous system, which richly supplies BAT, is thought to play an important role in the regulation of BAT thermogenesis because catecholamines stimulate and beta adrenergic blocking agents inhibit oxygen consumption in this tissue. The present studies were carried out to assess directly sympathetic activity in BAT in response to cold exposure and to changes in dietary intake, both of which alter heat production in the rat. Sympathetic activity was determined from the rate of norepinephrine (NE) turnover in interscapular brown adipose tissue (IBAT) after preliminary experiments validated the use of NE turnover techniques in IBAT. Acute exposure to 4°C increased NE turnover in IBAT 4- to 12-fold compared with ambient temperature controls, depending upon the interval over which the turnover measurement was made, while in the heart NE turnover doubled in response to the same cold stimulus. In animals exposed to cold continuously for 10 d before study, NE turnover measurements in IBAT and in the heart were elevated comparably to those obtained during acute exposure. Alterations in feeding were also associated with changes in NE turnover in IBAT. Fasting for 2 d decreased NE turnover in IBAT (-35% from 29.2±4.2 ng NE/h to 18.9±5.9) and in heart (-52%). In animals fed a “cafeteria” diet, a model of voluntary overfeeding in the rat, NE turnover was increased in both IBAT (+108% from 24.8±4.5 ng NE/h to 51.7±6.8) and heart (+66%). Because ganglionic blockade exerted a greater effect on NE turnover in IBAT in cafeteria-fed rats than in controls, the increase in NE turnover in IBAT with this overfeeding regimen reflects enhanced central sympathetic outflow. Thus NE turnover techniques can be satisfactorily applied to the assessment of sympathetic nervous system activity in IBAT.

Authors

James B. Young, Elizabeth Saville, Nancy J. Rothwell, Michael J. Stock, Lewis Landsberg

Effects of dietary cholesterol and fatty acids on plasma lipoproteins.

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The effects of dietary cholesterol and fatty acids on low density and high density lipoproteins (LDL and HDL) were studied in 20 young men. After 2-3 wk of evaluations on ad lib. diets, basal diets, which consisted of 15% protein, 45% carbohydrates, 40% fat, and 300 mg/day of cholesterol, were given for 4-5 wk (Basal). The ratio of dietary polyunsaturated to saturated fatty acids (P/S) for different groups of subjects were 0.25, 0.4, 0.8, or 2.5. 750 and 1,500 mg/d of cholesterol were added to the basal diets as 3 and 6 eggs, respectively. Total cholesterol and LDL cholesterol were lower in all subjects on the basal diets than on the ad lib. diets. Addition of 750 mg cholesterol to the diet with P/S = 0.25-0.4 raised LDL cholesterol by 16 +/- 14 mg/dl to 115% of basal diet values (n = 11, P less than 0.01); 1,500 mg increased LDL cholesterol by 25 +/- 19 mg/dl to 125% (n = 9, P less than 0.01). On the diet with P/S = 0.8, 750 mg produced insignificant increases in LDL cholesterol, but 1,500 mg produced increases of 17 +/- 22 mg/dl to 115% of basal (n = 6, P less than 0.02). On the P/S = 2.5 diet, neither 750 nor 1,500 mg produced significant changes. Thus, both the cholesterol contents and P/S ratios of diets were important in determining LDL levels. The lipid and apoprotein compositions, flotation rates, molecular weights, and binding by cellular receptors of LDL were virtually unchanged by the addition of cholesterol to the diets high in saturated fat. These diets, therefore, caused an increase in the number of LDL particles of virtually unchanged physical and biological properties. On the diet with low P/S ratio, HDL2 rose, whereas this effect was absent on diets with high P/S ratios. The response of LDL to dietary manipulations is consonant with epidemiologic data relating diets high in cholesterol and saturated fat to atherogenesis. The response of HDL2, however, is opposite to that of its putative role as a negative risk factor. Further work is needed to clarify this interesting paradox.

Authors

G Schonfeld, W Patsch, L L Rudel, C Nelson, M Epstein, R E Olson

Clonal Expression of the Tn Antigen in Erythroid and Granulocyte Colonies and Its Application to Determination of the Clonality of the Human Megakaryocyte Colony Assay

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Abstract

To evaluate whether exposure of Tn determinants at the surface of human erythrocytes, platelets, and granulocytes could arise from a somatic mutation in a hemopoietic stem cell, burst-forming unit erythroid (BFU-E) colonies, colony-forming unit granulocyte-macrophage (CFU-GM), and colony-forming unit-eosinophil (CFU-Eo) were grown from a blood group O patient with a typical Tn syndrome displaying two distinct populations (Tn+ and Tn-) of platelets, granulocytes, and erythrocytes. A large number of colonies was observed. Individual colonies were studied with a fluorescent conjugate of Helix pomatia agglutinin (HPA). A sizeable fraction of each of the erythroid and granulocytic colonies appeared to consist exclusively of either HPA-positive or HPA-negative cells, thereby demonstrating the clonal origin of those exhibiting the Tn marker. Similar results were obtained from a second patient. These findings establish that the HPA labeling of Tn cells is an accurate marker permitting assessment of the clonality of the human megakaryocyte (MK) colony assay. For the study of MK cultures a double-staining procedure using the HPA lectin and a monoclonal antiplatelet antibody (J-15) was applied in situ to identify all MK constituting a colony. Our results, obtained in studies of 133 MK colonies, provide definitive evidence that the human MK colony assay is clonal because all MK colonies were exclusively composed of Tn+ and Tn- MK. Furthermore, the distribution of MK within a single colony was shown to be seminormal with a mean at 6 MK, isolated MK typically being absent in culture.

Authors

William Vainchenker, Ugo Testa, Jeanne Françoise Deschamps, Annie Henri, Monique Titeux, Janine Breton-Gorius, Henri Rochant, Douglas Lee, Jean-Pierre Cartron

Differing uptake of emulsion triglyceride by the fed and fasted rat liver.

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Abstract

The recycling perfusion of a fasted rat liver with an apoprotein E-enriched synthetic triglyceride emulsion revealed a significantly greater hepatic uptake of both the apoprotein and the triglyceride than did the liver of a chow-fed animal. This greater hepatic triglyceride uptake by the perfused fasted liver in comparison to the fed was also noted for emulsions containing no added apoprotein or supplemented with both the E and CIII-1 proteins. However, no difference in the uptake of the triglyceride emulsion was seen for the fed and fasted livers when evaluated by a nonrecycling single pass perfusion. The isolated hepatocyte plasma membranes from fasted rats failed to demonstrate enhanced binding of apoprotein or lipid when compared to those from fed animals. If the residual E loaded triglyceride emulsion was recovered from the recycling perfusates of fed and fasted livers and evaluated in a non-recycling single-pass system, the emulsion from the fasted perfusion was cleared as facilely as previously, whereas that from the fed was less actively cleared. The emulsions retrieved from the perfusion of the fed liver contained significantly more protein than did the fasted; in particular apo C. This apparent alteration of emulsion apoproteins by the fed liver possibly results in a less active hepatic retrieval and may be important in downregulating the entry of lipoprotein triglyceride in the postabsorptive liver.

Authors

S H Quarfordt, J Hanks, F Shelburne, B Schirmer

Role for Endogenous and Acquired Peroxidase in the Toxoplasmacidal Activity of Murine and Human Mononuclear Phagocytes

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Oxygen products generated by the respiratory burst of mononuclear phagocytes are microbicidal to intracellular pathogens including Toxoplasma gondii. The toxicity of one of these products, H2O2, is markedly amplified by the granule peroxidase of circulating phagocytes in the presence of a halide. Eosinophil peroxidase (EPO) binds firmly to the surface of T. gondii and such organisms remain viable as determined by vital staining, uptake of 2-deoxyglucose, and survival and replication in human fibroblasts. They are, however, rapidly killed by the addition of H2O2 and iodide under conditions in which control organisms are unaffected. We have used EPO bound to T. gondii to explore the role of peroxidase in the toxoplasmacidal activity of mononuclear phagocytes. Resident mouse peritoneal macrophages lack a granule peroxidase and have a weak respiratory burst; toxoplasma survive and replicate within these cells. However, these cells acquire significant toxoplasmacidal activity, as assessed microscopically and by the inhibition of uracil uptake, when organisms are coated with EPO before ingestion, an effect which is decreased by the hemeprotein inhibitors, aminotriazole and azide. EPO on the surface of Toxoplasma does not increase their ingestion by macrophages or the associated respiratory burst. Monocytes from patients with hereditary myeloperoxidase deficiency have a significant toxoplasmacidal defect that is abolished when EPO-coated organisms are used. In contrast, the toxoplasmacidal defect of monocytes from chronic granulomatous disease patients is unaffected by surface-bound EPO. In these studies, replication of surviving intracellular organisms varied inversely with the magnitude of the respiratory burst: replication was greatest in fibroblasts, slightly less in resident macrophages, and least in monocytes; it was significantly greater in chronic granulomotous disease than in normal or myeloperoxidase-deficient monocytes. These studies support a role for oxygen products and endogenous peroxidase in the optimal killing of T. gondii by monocytes and demonstrate that peroxidase-negative phagocytes can utilize peroxidase on the surface of ingested organisms to augment microbicidal activity.

Authors

Richard M. Locksley, Christopher B. Wilson, Seymour J. Klebanoff

In vitro studies of human pluripotential hematopoietic progenitors in polycythemia vera. Direct evidence of stem cell involvement.

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Previous in vitro studies on committed hematopoietic progenitors have suggested that polycythemia vera (PV) is a clonal disorder arising in a pluripotential hematopoietic stem cell. In this study, recently developed technics for clonal assay of a human multipotential progenitor cell (CFU-GEMM) were used to assess the functional characteristics of CFU-GEMM in 19 PV patients. These studies showed: (a) increased numbers of detectable CFU-GEMM in blood and bone marrow samples of PV patients as compared with normals (P less than 0.002 and P less than 0.02, respectively); (b) erythropoietic differentiation of PV CFU-GEMM without exogenous erythropoietin (Ep) in culture (in marked contrast to CFU-GEMM of both normals and subjects with secondary erythrocytosis which require exogenous Ep for terminal hemoglobinization of their erythroid component), a property shown by experiments with an anti-Ep antiserum to be related to increased sensitivity of PV CFU-GEMM to Ep; (c) increased megakaryocyte formation by PV CFU-GEMM as compared with normals (P less than 0.025); and (d) a linear relationship, extrapolating to the origin, between CFU-GEMM detected and cells cultured. These studies demonstrate that at least two clinical features of PV, increased erythropoiesis and megakaryocytopoiesis, are reflected in corresponding functional characteristics of PV CFU-GEMM, and provide direct evidence of distinctive pluripotential stem cell activity in this disorder.

Authors

R C Ash, R A Detrick, E D Zanjani

Insulin stimulation of adipose tissue lipoprotein lipase. Use of the euglycemic clamp technique.

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The role of insulin in the regulation of adipose tissue lipoprotein lipase activity in humans was investigated in 11 normal subjects and compared with the effects of 0.9% saline infusions in five control subjects. After a basal adipose tissue biopsy for lipoprotein lipase activity, insulin was rapidly infused to achieve and maintain serum levels of approximately 70 microunits/ml while plasma glucose was kept at basal concentrations. Free fatty acids in serum fell to 27 +/- 3% of basal by 20 min (t = 5.19, P less than 0.001) and triglycerides decreased to 77 +/- 3% of basal by 80 min (t = 3.76, P less than 0.01). Adipose tissue lipoprotein lipase activity failed to increase significantly above that measured in controls by the first 3 h of the study. By 6 h of the infusion a stimulatory effect of insulin on adipose tissue lipoprotein lipase was found (t = 3.94, P less than 0.01). There was no relationship between the amount of glucose infused and the insulin effect on the enzyme. The increase in adipose tissue lipoprotein lipase activity at 6 h, however, was inversely related to the basal lipase activity (r = -0.690, P less than 0.02). Thus, insulin appears to stimulate adipose tissue lipoprotein lipase activity in humans. This effect of insulin is delayed when compared with antilipolysis and the fall in plasma triglyceride. The inverse relationship between insulin-stimulated adipose tissue lipoprotein lipase activity and basal enzyme activity suggests that adipose tissue itself is the main regulator of the lipase response to insulin.

Authors

C N Sadur, R H Eckel

Intravascular activation of complement and acute lung injury. Dependency on neutrophils and toxic oxygen metabolites.

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Abstract

Intravascular activation of the complement system with cobra venom factor results in acute lung injury, which has been quantitated by increases in lung vascular permeability. Cobra venom factor preparations devoid of phospholipase A2 activity retain full lung-damaging capacity. The lung injury is associated with the preceding appearance of chemotactic activity in the serum coincident with the development of a profound neutropenia. The chemotactic activity is immunochemically related to human C5a. Morphologic studies have revealed discontinuities in the endothelial cell lining of lung alveolar capillaries, damage and/or destruction of endothelial cells in these areas, plugging of pulmonary capillaries with neutrophils that are in direct contact with vascular basement membrane, the presence of fibrin in alveolar spaces and in areas adjacent to damaged endothelial cells, and intraalveolar hemorrhage. Lung injury is dramatically attenuated in animals that have been previously neutrophil depleted. Teh intravenous injection of superoxide dismutase or catalase also provides significant protection from the pulmonary damage. Very little protection from the pulmonary damage. Very little protection is afforded by pretreatment of rats with antihistamine. These studies suggest that intravascular activation of the complement system leads to neutrophil aggregation and activation, intrapulmonary capillary sequestration of neutrophils, and vascular injury, which may be related to production of toxic oxygen metabolites by complement-activated neutrophils.

Authors

G O Till, K J Johnson, R Kunkel, P A Ward

Insulin Binding and Degradation by Luminal and Basolateral Tubular Membranes from Rabbit Kidney

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Insulin influences certain metabolic and transport renal functions and is avidly degraded by the kidney, but the relative contribution of the luminal and basolateral tubular membranes to these events remains controversial. We studied 125I-insulin degradation [TCA and immunoprecipitation (IP) methods] and the specific binding of the hormone by purified luminal (L) and basolateral (BL) tubular membranes. These were prepared from rabbit kidney cortical homogenates by differential and gradient centrifugation and ionic precipitation steps in sequence, which resulted in enrichment vs. homogenate of marker enzymes' activities (sodium-potassium-activated adenosine triphosphatase for BL and maltase for L) of 8- and 12-fold, respectively. Both fractions degraded insulin avidly and bound the hormone specifically without saturation even at pharmacologic concentrations (10 μM). At physiologic insulin concentrations (0.157 nM) BL membranes degraded substantial amounts of insulin (44.2±2.6 and 40.7±2.2 pg/mg protein per min by the TCA and IP methods, respectively), even though at lesser rates (P < 0.001) than the luminal fraction (67.2±2.3 and 75±6.2 pg/mg protein per min, respectively); the rate of insulin catabolism by BL membranes was significantly higher (P < 0.001) than that which could be attributed to their contamination by luminal components [12.2±1.9 pg/mg per min (TCA method), or 13.7±1.9 pg/mg per min (IP method)]. Competition experiments suggested that insulin-degrading activity in both fractions includes both specific and nonspecific components. In contrast to degradation, insulin binding by both membranes was highly specific for native insulin and was severalfold higher in BL than L membranes [17.5±1.3 vs. 4.5±0.4 fmol/mg protein (P < 0.001) at physiologic insulin concentrations]. Despite the marked difference in the binding capacity for insulin by the two membranes, the patterns of labeled insulin displacement by increasing amounts of unlabeled hormone were superimposable (50% displacement required ∼3 nM), suggesting that their receptors' affinity for insulin was similar. These observations provide direct evidence that interaction of insulin with the kidney involves binding and degradation of the hormone at the peritubular cell membrane.

Authors

Zvi Talor, Dimitrios S. Emmanouel, Adrian I. Katz

An antiidiotypic antibody that recognizes the beta-adrenergic receptor.

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Antialprenolol rabbit antibodies were fractionated on an acebutolol affinity resin, followed by L-propranolol elution so as to separate a class of binding sites that mimic the beta-adrenergic receptor. Allotype-identicaL rabbits were immunized with this fraction. After 6 mo, antisera exhibited antiidiotypic activity inhibiting [3H]alprenolol binding to the original antibody and to rabbit antiacebutolol antibodies, which had a spectrum of ligand-binding properties identical to the original idiotype. Those antisera demonstrating the original idiotype. Those antisera demonstrating the most potent antiidiotypic activity also blocked [3H]alprenolol binding to the beta-adrenergic receptor of turkey membrane, canine pulmonary membrane, and rat reticulocyte. An idiotype affinity-purified fraction showed similar activity, inhibiting beta-receptor binding with a calculated dissociation constant (KD) of 53 nM. Isoproterenol-mediated adenylate cyclase activity was also inhibited in a competitive manner. The universality of recognition of these antiidiotypic antisera indicate that the three-dimensional structure of a receptor's binding site can be modeled by a subset of an elicited antibody population.

Authors

C J Homcy, S G Rockson, E Haber

Mononuclear cells from patients with the hyperimmunoglobulin E-recurrent infection syndrome produce an inhibitor of leukocyte chemotaxis.

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The chemotactic responsiveness of the neutrophils of 10 patients with the hyperimmunoglobulin E-recurrent infection syndrome (HIE) were compared with neutrophils from normal volunteers over a 10-mo period. HIE neutrophils as a group displayed significantly less chemotactic motility than control neutrophils. The data from individual patients were variable, being normal or abnormal on different days. Mononuclear cells from HIE patients, when cultured for 24 h in the absence of serum or a mitogen, produced a factor that inhibited normal neutrophil and monocyte chemotaxis. Mononuclear cells from normal volunteers with and without atopy or from patients with parasites or bacterial infections did not produce such an inhibitory factor. The production of this chemotactic inhibitory factor in vitro was variable over time, but it correlated with the presence of an in vitro neutrophil chemotactic defect. The chemotactic inhibitory factor was partially purified and was found to contain protein, to be stable at 56 degrees C, and to have a molecular weight of approximately 61,000. Irreversible inhibitors of serine esterases do not inactivate the factor. The factor is produced by esterase-negative mononuclear cells and is not toxic to neutrophils. This chemotactic inhibitory factor may be the basis of the variable chemotactic defect in HIE neutrophils.

Authors

H Donabedian, J I Gallin

Regulation of hemoglobin AIc formation in human erythrocytes in vitro. Effects of physiologic factors other than glucose.

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Abstract

The formation of hemoglobin AIc was studied in intact human erythrocytes in vitro. Satisfactory methods were developed for maintaining erythrocytes under physiologic conditions for greater than 8 d with less than 10% hemolysis. Hemoglobin AIc levels were determined chromatographically on erythrocyte hemolysates after removal of reversible components by incubation for 6 h at 37 degree C. Hemoglobin AIc concentration was found to increase linearly with time during 8 d of incubation. The rate of formation of hemoglobin AIc increased linearly as glucose concentration was increased from 40 to 1,000 mg/dl. Deoxyhemoglobin was glycosylated twice as rapidly as oxyhemoglobin. The rate of hemoglobin AIc formation was further increased by elevated 2,3-diphosphoglycerate levels, an effect that was most marked with deoxyhemoglobin. We conclude that the nonenzymatic glycosylation of hemoglobin is influenced by factors other than glucose, including oxygen tension and 2,3-diphosphoglycerate levels.

Authors

R J Smith, R J Koenig, A Binnerts, J S Soeldner, T T Aoki

Preservation of mesangium and immunohistochemically defined antigens in glomerular basement membrane isolated by detergent extraction.

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To define the characteristics of isolated glomerular basement membrane (GBM), immunohistochemical and morphometric analyses have been carried out on rat and human tissues. Site-specific arrays of antigens were identified in detergent-isolated GBM in a distribution similar to that observed in intact kidney. In the human, fibronectin, procollagen IV, and collagen V were observed along the internal aspect of GBM continuous with antigenic sites in the mesangium. Another array of antigens was identified in the GBM but not within the mesangium--Goodpasture's antigen, bovine lens capsule type IV collagen, and amyloid P component. In addition, sites reactive with rabbit antiserum to laminin were present on both sides of the lamina densa as well as within the mesangial region. Actomyosin, a presumed mesangial cell antigen persisted in the mesangium of isolated GBM. Mesangial matrix was identified in detergent-isolated GBM in an amount equivalent to that present in intact glomeruli. Sonicated GBM contained the same antigens but it was not possible to quantitate the amount of mesangial material by immunofluorescence or morphometric analysis. The thickness of the lamina densa was greater in sonicated and detergent-treated rat GBM preparations than in native rat kidney. These studies demonstrated that isolated GBM is heterogeneous with respect to its antigenic constituents and in addition contains mesangial matrix, which is morphologically and immunohistochemically distinct from peripheral GBM.

Authors

M T Houser, J I Scheinman, J Basgen, M W Steffes, A F Michael

Evidence for Two Tissue-specific Pathways for In Vivo Thyroxine 5′-Deiodination in the Rat

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Abstract

Propylthiouracil (PTU) is a well known inhibitor of thyroxine (T4) to triiodothyronine (T3) conversion as evidenced by its effect in several in vitro systems and by the decrease in serum T3 caused by this drug in either rats or man receiving T4 replacement. However, the failure of PTU to decrease the intrapituitary T3 concentration and to completely blunt the serum T3 concentration in T4-replaced athyreotic rats suggest that there may be a PTU-insensitive pathway of T4 to T3 conversion in some tissues. To address this question, we have studied the in vivo effect of PTU treatment on the generation of [125I]T3 from [125I]T4 in the serum and cerebral cortex (Cx), cerebellum (Cm), liver (L), and anterior pituitary (P) of euthyroid rats. Whereas PTU decreased the concentration of [125I]T3 in the serum, L homogenates, and L nuclei after [125I]T4, it did not affect the concentration of [125I]T3 in homogenates or nuclei of Cx, Cm, or P. Iopanoic acid pretreatment significantly reduced the [125I]T3 concentration in serum, homogenates, and cell nuclei of all these organs. Neither agent affected the metabolism or tissue distribution of simultaneously injected [131I]T3. The presence of PTU in these tissues was evaluated by in vitro assessment of iodothyronine 5′-deiodinating activity using both [125I]rT3 and [125I]T4 as substrates. In agreement with the in vivo findings, generation of [125I]T3 from T4 in vitro was not affected by PTU in Cx, Cm, P but it was inhibited by 76% in L. However, rT3 5′-deiodination, known to be sensitive to PTU in these tissues, was inhibited in all four indicating that the PTU given in vivo was present in significant amounts. These results demonstrate that in rat Cx, Cm, and P unlike liver, PTU does not inhibit T4 to T3 conversion in vivo despite the presence of the drug in the tissues in amounts that significantly inhibit reverse T3 5′-deiodination. These results show that in vivo 5′-deiodination of T4 proceeds via a PTU-insensitive pathway in the central nervous system and pituitary, while this pathway is not quantitatively important in the L. This mechanism accounts for the “locally generated” T3 in central nervous system and pituitary and could also provide the approximately one-third of extrathyroidally produced T3 not blocked by PTU administration in athyreotic T4-replaced rat.

Authors

J. Enrique Silva, Jack L. Leonard, Frank R. Crantz, P. Reed Larsen

Long-lasting, specific immunologic unresponsiveness associated with cryptococcal meningitis.

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A sensitive radioimmunoassay and an antibody class-specific enzyme-linked immunosorbent assay were used to determine whether patients cured of cryptococcosis responded normally to immunization with cryptococcal capsular polysaccharide (CPS) and type III pneumococcal polysaccharide. 10 normal volunteers and 8 patients who had been cured of cryptococcal meningitis and who had been cured of cryptococcal meningitis and who had no serious underlying diseases were immunized with both antigens. Geometric mean titers to CPS measured by radioimmunoassay were 1:1 in both groups before vaccination, but were 1:3 in patients and 1:119 in controls following immunization (P less than 0.01, Student's t test). Analysis of the class-specific response to immunization with CPS found little anti-CPS IgG or IgA. Geometric mean postvaccination IgM titers were 1:31 in patients and 1:238 in controls (P less than 0.01). Responses to immunization with type III pneumococcal polysaccharide were similar in patients and controls, with IgA, IgM, and IgG mean titers of 1:1129, 1:369, and 1:158 in patients and 1:1504, 1:1039, and 1:163 in controls (P greater than 0.2 for each antibody class). Cured cryptococcal meningitis is often associated with prolonged specific immunologic unresponsiveness.

Authors

D K Henderson, J E Bennett, M A Huber

Glucocorticoid and Mineralocorticoid Effects on Adrenocorticotropin and β-Endorphin in the Adrenalectomized Rat

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Immunoreactive ACTH (ir-ACTH) and immunoreactive β-endorphin (ir-βEP) were determined in plasma, anterior pituitary, neuro-intermediate lobe, and hypothalamus of sham-adrenalectomized rats, and adrenalectomized rats given six daily injections of vehicle (oil), dexamethasone, 9α-fluorocortisol or deoxycorticosterone. 6 d after adrenalectomy, anterior pituitary ir-ACTH and ir-βEP were double, and plasma levels approximately fivefold those in controls. Adrenalectomy did not alter hypothalamic levels of either peptide, or ir-βEP in neuro-intermediate lobe, in which tissue ir-ACTH was below detection limit at routine dilutions. Dexamethasone (0.2-200 μg/d) concurrently suppressed plasma ir-ACTH and ir-βEP, with a near maximal effect at 20 μg, and a half-maximal effect between 2 and 6 μg; similar dose-response characteristics were found for thymolysis. Step-wise increases in anterior pituitary content of both peptides were found, with no change in hypothalamic levels of either peptide, or neuro-intermediate lobe ir-βEP. 9α-fluorocortisol (0.2-200 μg/d) produced plasma, anterior pituitary, and hypothalamic effects equivalent to dexamethasone, but with one-tenth the potency. Unlike dexamethasone, higher doses of 9α-fluorocortisol significantly elevated neuro-intermediate lobe ir-βEP. Deoxycorticosterone (2-2,000 μg/d) produced no significant changes in plasma, anterior pituitary or hypothalamic levels of either peptide; like 9α-fluorocortisol, doses of >60 μg/d significantly elevated neuro-intermediate lobe ir-βEP. Whereas ir-ACTH and ir-βEP synthesis in and release from the anterior pituitary are under complex negative feedback glucocorticoid control, there exists a mineralocorticoid-specific effect on neuro-intermediate lobe content of ir-βEP.

Authors

Alan T. Lim, B. A. K. Khalid, Judith Clements, John W. Funder

Purified human plasma kallikrein aggregates human blood neutrophils.

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Abstract

Exposure of human blood polymorphonuclear leukocytes (PMN) to purified active plasma kallikrein resulted in PMN aggregation when kallikrein was present at concentrations ranging from 0.4 to 0.6 U/ml (0.18-0.27 microM). Kallikrein-induced PMN aggregation was not mediated through C5-derived peptides, because identical responses were observed whether or not kallikrein had been preincubated with an antibody to C5. Moreover, kallikrein was specific for aggregating PMN, because no aggregation was observed with Factor XII active fragments (23 nM), Factor XIa (0.6 U/ml or 15nM), thrombin (1.6 microM), plasmin (2 microM), porcine pancreatic elastase (2 microM), bovine pancreatic chymotrypsin (2 microM), or bradykinin (1 microM). Bovine pancreatic trypsin (2 microM) aggregated PMN, but to a lesser extent than kallikrein (0.18 microM). Kallikrein was a potent aggregant agent for PMN because similar responses were observed with kallikrein (0.5 U/ml or 0.23 microM) and an optimal dose (0.2 microM) of N-formyl-methionyl-leucyl-phenylalanine. In addition, PMN incubation with kallikrein resulted in stimulation of their oxidative metabolism as assessed by an increased oxygen uptake. Neutropenia and leukostasis observed in diseases associated with activation of the contact phase system may be the result of PMN aggregation by plasma kallikrein.

Authors

M Schapira, E Despland, C F Scott, L A Boxer, R W Colman

3 alpha, 17 beta-androstanediol glucuronide in plasma. A marker of androgen action in idiopathic hirsutism.

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Abstract

Biologically active androgens and peripheral androgen metabolites in plasma were measured in 25 women with idiopathic hirsutism (IH). Plasma testosterone was not significantly elevated. Free testosterone however was increased although the elevation was not impressive (10.9 +/- 6.6 SD vs. 3.3 +/- 1.5 ng/dl) and one-fourth of the cases had normal unbound testosterone. Dihydrotestosterone (DHT) values were elevated (23.5 +/- 14 vs. 12.5 +/- 3.59) but again over half of the values were within the normal range. In our series of mild to moderate cases, 3 alpha-diol was not at all discriminatory. However, plasma 3 alpha-diol glucuronide was markedly increased (604 +/- 376 vs. 40 +/- 10 ng/dl), and elevated in all but one mild case. Previous studies document that DHT is the important androgen in skin and formation of DHT and 3 alpha-diol is markedly increased in vitro in IH. Since 3 alpha-diol glucuronide is derived largely from extrasplanchnic events, beta-glucuronidase is present in skin, and androgen stimulates formation of the enzyme in extrasplanchnic tissue, we conclude that 3 alpha-diol glucuronide is a marker of peripheral androgen action and markedly elevated in IH.

Authors

R Horton, D Hawks, R Lobo