Covalently cross-linked dimers and oligomers composed of 2-4 subunits of monoclonal human IgG1 were prepared by incubation of purified monomeric IgG1 with glutaraldehyde followed by gelfiltration chromatography. Monomers, dimers, and oligomers then were labeled with 125I and used to compare the binding properties of IgG Fc receptors on human peripheral blood monocytes and polymorphonuclear leukocytes (PMN). Binding of IgG1 to monocytes at 37°C and of IgG1 polymers to PMN at 4°C could be readily measured and were found to be reversible and saturable. Scatchard plots of binding were linear in each instance. Monocytes bound a mean of 20,200±6,800 molecules/cell of IgG1 monomer at saturation and comparable amounts of dimer or oligomer. The mean association constant (Ka) for binding of IgG1 monomer to monocytes was 8.6 × 108M−1 and the Ka for binding of dimer and oligomer were three-to fivefold greater.
Roger J. Kurlander, Janet Batker
It has been shown that active immunization of rats with the capsular polysaccharide of Bacteroides fragilis protects these animals against abscess development following intraperitoneal challenge with this species. Passive transfer of hyperimmune globulin from immunized animals to nonimmune recipients provided protection against B. fragilis bacteremia in challenged animals, but did not confer protection against abscess development. On the other hand, adoptive transfer of spleen cells from immunized animals to nonimmunized recipients resulted in protection against abscesses following challenge with B. fragilis. These data suggested that a T cell-dependent immune response was involved in protection against abscess development after immunization with B. fragilis capsular antigen.
Andrew B. Onderdonk, Richard B. Markham, Dori F. Zaleznik, Ronald L. Cisneros, Dennis L. Kasper
Antileukocyte antibodies in sera from patients with systemic lupus erythematosus (SLE) were characterized by determining cross-reacting specificies with the antigens defined by OKT3, OKT4, OKT8, OKM1 and anti-Ia hybridoma antibodies (Abs). T cells were prepared by sheep erythrocyte (E) rosetting after removal of adherent cells. T cells, or non-T cells, were preincubated with SLE sera at 4°C and then with monoclonal Abs. Binding by specific monoclonal Abs was assessed by two methods: rosetting with ox erythrocytes conjugated with goat anti-mouse IgG and also in the fluorescence-activated cell sorter using fluorescein isothiocyanate-conjugated goat anti-mouse IgG. Using the rosetting method, we found that sera from SLE can block the binding of monoclonal mouse hybridoma anti-Ia Abs to T cells; the blocking of other monoclonal Abs was not consistent. Using fluorescence-activated cell sorter analysis, preincubation with SLE sera lowered the intensity of staining and total percentage of either T or non-T cells stained by monoclonal anti-Ia Abs. Blocking of anti-Ia Abs binding by SLE sera was not histocompatibility leukocyte antigen (HLA)-DR restricted and was not due to Fc receptor binding. These results indicated that antibodies in SLE sera react with structures contiguous to or identical with Ia determinants. Anti-Ia activities in SLE sera correlate with SLE disease activity. In addition, there was a significant negative correlation between anti-Ia blocking activity in the sera and the percentage of Ia-positive T cells in the blood of SLE patients. Antibodies in SLE sera with anti-Ia blocking activity may play an important role in immune dysregulation in SLE patients.
Kunio Okudaira, Robert P. Searles, Jan L. Ceuppens, James S. Goodwin, Ralph C. Williams Jr.
A human neutrophil protease, initially termed neutral peptide-generating protease, has been shown to cleave angiotensin II directly from angiotensinogen and has been identified as leukocyte cathepsin G. When purified neutrophils were disrupted by nitrogen cavitation and fractionated by differential centrifugation, 44 and 24% of the angiotensin II-generating activity was in the lysosomal and undisrupted cell fractions, respectively. Cytochalasin B-treated human neutrophils stimulated with N-formyl-L-methionyl-L-leucyl-L-phenylalanine released beta-glucuronidase, lysozyme, and angiotensin II-generating protease in a dose-dependent fashion, consistent with localization of this protease to the neutrophil granule. Individually purified angiotensin II-generating protease and cathepsin G had similar proteolytic and esterolytic activity for angiotensinogen and N-benzoyl-L-tyrosine ethyl ester on a weight basis, exhibited identical mobilities by SDS-gradient polyacrylamide gel electrophoresis and pH 4.3 disc-gel electrophoresis, and gave precipitin lines of antigenic identity on Ouchterlony analysis with goat antibody to the angiotensin II-generating protease. Thus, the angiotensin II-generating protease of human neutrophils has been identified as cathepsin G on the basis of subcellular localization, substrate specificity, physicochemical characteristics, and antigenic identity.
M G Tonnesen, M S Klempner, K F Austen, B U Wintroub
30% of patients with essential hypertension have a decreased adrenal response to angiotensin II (A II) on a low but not a high sodium intake. They also have a compensatory increase in the activity of the renin-angiotensin system best documented in a sodium-restricted state.
Gordon H. Williams, Lynne M. Braley, Alphonsa Menachery
The Na+-K+ pump in the erythrocytes of a mordibly obese patient shows a unique constellation of functional abnormalities. The number of pump units, measured by [3H]ouabain binding to intact cells, as well as the enzymatic activity of the (Na+-K+)-dependent ATPase in erythrocyte membranes were found to be markedly increased compared with control cells (18-fold and 14-fold, respectively). There was a concomitant fivefold increase in the rate of pump-mediated uptake of 86Rubidium (a K analogue); this was balanced by an increased rate of 86Rb efflux. In striking contrast to normal cells, however, a major portion of this efflux (80%) was inhibited by ouabain, and thus appeared to be mediated by the Na+-K+ pump.
Mario Deluise, Jeffrey S. Flier
The net exchange of glucose and lactate across the leg and the splanchnic bed and the arterialdeep venous (A-DV) differences for these substrates in the forearm were determined in healthy subjects during 3-3.5 h of leg exercise (bicycle ergometer) at 58% maximum O2 uptake and during a 40-min post-exercise recovery period.
Gunvor Ahlborg, Philip Felig
Increased sympathetic nervous system activity has been demonstrated in established one-kidney one-clip hypertension in the rat. We have found that renal denervation in this model results in an attenuation of hypertension, unassociated with alterations in sodium or water balance or renin activity. To determine whether the depressor effect of renal denervation is associated with changes in peripheral sympathetic nervous system activity, sham operation (n = 12), renal denervation (n = 13), or unclipping (n = 13) was carried out 2 wk after the onset of one-kidney one-clip hypertension. Normotensive unine-phrectomized age- and sex-matched rats were used as controls (n = 14). Renal denervation resulted in a significant decrease in systolic blood pressure (201±7 to 151±6 mm Hg), while unclipping lowered systolic blood pressure to normotensive levels (130±6 mm Hg). 8 d after operation plasma norepinephrine and mean arterial pressure before and after ganglionic blockade with 30 mg/kg hexamethonium bromide were measured in conscious, unrestrained, resting animals, as indices of peripheral sympathetic nervous system activity. Plasma norepinephrine was significantly higher in hypertensive sham-operated rats (422±42 pg/ml) compared with normotensive controls (282±25 pg/ml) (P < 0.01). Both renal denervation and unclipping restored plasma norepinephrine to normal levels (273±22 and 294±24 pg/ml, respectively). Ganglionic blockade in hypertensive sham-operated animals resulted in a significantly greater decrease in mean arterial pressure than occurred in renal denervated, unclipped, or control rats. The data suggest that the depressor effect of renal denervation or unclipping in the one-kidney one-clip hypertensive rat is associated with a decrease in peripheral sympathetic nervous system activity.
Richard E. Katholi, Sherry R. Winternitz, Suzanne Oparil
Human alveolar macrophages (AM) have recently been reported to ingest and kill a strain of Staphylococcus (502A) in the absence of opsonins. To further investigate the mechanism of non-opsonic recognition, we studied phagocytosis of 23 clinical and laboratory strains of S. aureus and Staphylococcus epidermidis by AM, and by blood polymorphonuclear leukocytes (PMN) and monocytes (MN). In the absence of opsonins, AM phagocytized 18 protein A-positive but not 5 protein A-negative strains of staphylococci, and the efficiency of phagocytosis directly correlated with the amount of protein A present in the bacterial cell wall (r = 0.86, P less than 0.001). Furthermore, AM rosetted around protein A-coated Sepharose beads, but not around beads without protein A. In contrast, PMN did not phagocytize nonopsonized staphylococci, and did not rosette around either type of Sepharose. MN phagocytized protein A-positive staphylococci, but much less efficiently than AM, and showed some rosetting around protein A-coated Sepharose. The nature of the AM receptor for protein A-positive staphylococci was studied. The surface of AM was positively stained with fluorescein-conjugated antibody to human IgG, but not with IgA- or IgM-specific conjugates. No such surface-immunoglobulins were detected on PMN, and MN were only weakly positive for surface IgG. Pretreatment of AM with F(ab')2 fragments specific for human IgG (anti-Fc) inhibited subsequent phagocytosis of protein A-positive staphylococci. There was no evidence that the AM surface IgG was aggregated or immunecomplexed. From these studies we conclude that human AM possess cytophilic IgG antibodies, which can function as receptors for phagocytosis of protein A-positive staphylococci.
H A Verbrugh, J R Hoidal, B Y Nguyen, J Verhoef, P G Quie, P K Peterson
To study antibody (Ab) biosynthesis in rheumatoid arthritis (RA), the immunoglobulin (Ig)M anti-Fc, anti-Fab′, and antistreptokinase-streptodornase (SKSD) produced by peripheral blood lymphocytes (PBL) were measured at intervals from 1 to 19 d in culture. PBL from 17 seropositive patients with active RA and 30 age-matched controls were evaluated. Within the first 24 h, PBL from six of eight patients released >30 ng IgM anti-Fc, even in the absence of pokeweed mitogen (PWM). This early release of Ab was blocked by cycloheximide. With or without PWM, PBL from normal donors did not release IgM anti-Fc until after 3-5 d in vitro. By day 9, unstimulated PBL from seven patients made > 100 ng IgM anti-Fc. Un-stimulated PBL from normals never made >95 ng of this Ab.
Holly H. Birdsall, Roger D. Rossen
The mechanism of protection of type-specific antipneumococcal antibody and complement in bacteremia was investigated with purified rabbit antibody and a guinea pig model of pneumococcal bacteremia. IgG and IgM were isolated from the sera of rabbits immunized with type 7 pneumococci (Pn), and their binding to Pn was quantitated. The number of antibody-binding sites on the pnuemococcal capsule was also determined. Pn were incubated with various amounts of the immunoglobulin preparations before intravenous injection into nonimmune guinea pigs. Whereas 120 molecules of IgM per Pn were sufficient to enhance bloodstream clearance of Pn, 1,400 molecules of IgG per bacterium were required to produce this effect. As the amount of either IgG or IgM added to the Pn was increased, the rate of bloodstream clearance accelerated. In striking contrast, greater than 1,000 molecules of IgM had no effect on the rate of clearance in C4-deficient guinea pigs, which cannot activate complement via the classic pathway. Similarly, 5,000 molecules of IgG had only minimal effect in C4-deficient guinea pigs, and 24,000 molecules of IgG had no effect in guinea pigs depleted of complement by cobra venom factor. Thus, the in vivo opsonic effects of both IgG and IgM anticapsular antibody are mediated via their ability to activate complement. IgG anti-pneumococcal cell wall antibody, raised by intravenous injection of rabbits with unencapsulated Pn, had no effect on the rate of bloodstream clearance of Pn or on the polymorphonuclear leukocyte killing of type 7 Pn in an in vitro bacterial assay. Because the opsonic effects of anticapsular antibody required complement activation, the ability of anticell wall IgG to activate complement was compared with the two classes of anticapsular antibody. As judged by depletion of C3 and C4 from guinea pig serum, as well as by the fixation of radiolabeled C3 to Pn, IgM anticapsular antibody was the best complement activator. However, anticell wall IgG was somewhat more active than anticapsular IgG in each of these tests of complement activation and fixation. When equivalent amounts of C3 were fixed to Pn by each of the three antibodies, Pn sensitized with IgG and IgM anticapsular antibodies caused immune adherence, whereas Pn sensitized with anticell wall IgG did not. This may explain the failure of anticell wall antibody of mediate complement-dependent phagocytosis of Pn in vivo or in vitro. Although anticell wall IgG is capable of activating complement and fixing C3 to Pn, it is not opsonic; the most likely reason is that the nonopsonic antibody mediates C3 deposition in sites on the Pn that cannot interact efficiently with phagocytic cell C3 receptors.
E J Brown, S W Hosea, C H Hammer, C G Burch, M M Frank
Patterns of protein synthesis by genital skin fibroblasts from three unrelated normal individuals and three unrelated patients with complete testicular feminization were compared to two-dimensional gel electrophoresis. cell lines were maintained in monolayer culture and pulse labeled with [35S]methionine. Cells were lysed in 9 M urea, and aliquots of 20 microliters subjected to isoelectric focussing and polyacrylamide gel electrophoresis followed by autoradiography. Gels of control fibroblasts showed two proteins (mol wt approximately 45,000, approximately 85,000; pKi approximately 5.0) markedly more prominent than on gels from affected fibroblasts. This pattern was unaltered by prior exposure to dihydrotestosterone, suggesting differences in constitutive proteins of the fibroblast cells. Parallel studies demonstrated a marked reduction in the ability of fibroblasts from patients with complete testicular feminization to bind androgens in vitro compared with those of normal individuals. The relationship between these proteins, androgen receptors, and androgen insensitivity requires further investigation.
G P Risbridger, B A Khalid, G L Warne, J W Funder
Catecholamines are postulated to regulate growth hormone (GH) secretion by their influence on the release of two hypothalamic substances, somatostatin, which inhibits GH release, and GH-releasing factor, as yet unidentified. Extensive pharmacologic studies in man and animals indicate a stimulatory effect of central norepinephrine and dopamine on GH, but the function of epiphephrine (EPI) is uncertain. Furthermore, many of the agents used to study the role of catecholamines in GH regulation are not selective in that they affect adrenergic as well as nor-adrenergic and/or dopaminergic neurotransmission. In the present investigation, central nervous system (CNS) EPI biosynthesis was selectively interrupted with the specific norepinephrine N-methyltransferase inhibitors, SK & F 64139 (Smith, Kline & French Laboratories) and LY 78335, (Eli Lilly & Co. Research Laboratories) and the effects of central EPI depletion on episodic GH secretion in the chronically cannulated rat model were determined. Inhibition of CNS EPI synthesis with SK & F 64139 caused complete suppression of episodic GH secretion and concomitantly reduced the EPI level in the hypothalamus without affecting dopamine or norepinephrine. Administration of LY 78335 produced similar effects on pulsatile GH. Morphine-induced, but not clonidine-induced, GH release also was blocked by SK & F 64139. These results indicate that (a) the central EPI system has a major stimulatory function in episodic GH release, (b) morphine-induced GH release is mediated by the central EPI system, and (c) clonidine stimulates GH release by activation of postsynaptic α-adrenergic receptors. Drugs that affect CNS adrenergic systems have a potential role in the diagnosis and treatment of disorders of GH secretion.
L. Cass Terry, W. R. Crowley, M. D. Johnson
Fibronectin is a major adhesive and opsonic glycoprotein found in plasma and tissues. Because this molecule appears to mediate a number of interactions between cells and extracellular matrix, and because the interstitial lung disease are characterized by marked derangements of the pulmonary extracellular matrix, we evaluated fibronectin in the lower respiratory tract in patients with these disorders. Fibronectin could be detected in the bronchoalveolar lavage fluid of normals (11/11), as well as those with noninterstitial lung diseases (18/18), idiopathic pulmonary fibrosis (21/21), sarcoidosis (20/20), and other interstitial lung diseases (22/22). Compared with normal and those with noninterstitial lung disease, the levels in bronchoalveolar lavage of patients with interstitial disease were significantly higher (P less than 0.01), all comparisons). This was true only for bronchoalveolar lavage fibronectin; plasma levels were similar in all study groups (P greater than 0.2, all comparisons). The lavage fluid fibronectin was intact antigenically and retained collagen binding capability, although in some cases of interstitial disease, the presence of lower molecular weight fragments suggested some degradation. Thus, fibronectin is a normal constituent of the epithelial fluid of the lower respiratory tract and is present in increased amounts in a significant number of individuals with interstitial lung disease.
S I Rennard, R G Crystal
The mechanism by which immune complexes deposit in vascular tissue is uncertain. Several human viruses, including herpes simplex virus, have recently been demonstrated to replicate in human endothelial cells. Such viruses may injure vascular tissue and could play a role in the pathogenesis of immune complex deposition. Therefore, we studied the expression of receptors for immune complexes containing IgG and C3 on endothelial cells after infection with herpes simplex virus type I.
Douglas B. Cines, Alan P. Lyss, Mahin Bina
Porphyria cutanea tarda and erythropoietic porphyria are disorders of heme synthesis that originate in the liver and bone marrow, respectively. Each is characterized by increased accumulation of uroporphyrin, I, by cutaneous photosensitivity, and in some patients by indurated plaques and scarring that resemble scleroderma. These scleroderma-like lesions occur in light-exposed and light-protected body areas. In these studies we evaluated the role of uroporphyrin I and of light in evoking the scleroderma-like cutaneous changes. Normal human skin fibroblasts were exposed to uroporphyrin I and to 400 nm radiation and the effect of these agents on collagen accumulation by the cells was determined. Radioactive tracer studies showed that uroporphyrin I caused a specific increase in the accumulation of newly synthesized collagen by fibroblast monolayer cultures, as verified by [3H]hydroxyproline and collagenase digestion assays. Collagen accumulation was stimulated 1.5- to 2.7-fold by uroporphyrin I, whereas noncollagenous protein accumulation was unchanged. The increased collagen accumulation was time and uroporphyrin I-concentration-dependent, and occurred both in the presence or absence of ultraviolet light exposure. Further studies demonstrated that the increased accumulation was not the result of decreased rates of collagen degradation nor was it due to changes in cell population growth parameters (generation times and saturation densities). No changes in morphology of the treated cells occurred. These studies indicate that porphyrins possess previously undemonstrated biological effects that are independent of their photosensitizing properties. This novel dark effect of uroporphyrin I may account for the sclerodermatous lesions seen in the skin of patients with porphyria cutanea tarda and erythropoietic porphyria.
George Varigos, John R. Schiltz, David R. Bickers
The cellular infiltrate in the deeper layers of the rheumatoid synovium produces a substantial amount of immunoglobulin (Ig)G. Culture supernatants of synovial tissues from 31 patients with rheumatoid arthritis (RA) undergoing joint replacement or synovectomy have been analyzed for the subclass of IgG present. IgG3 was measured by separation with Staphylococcal Protein A chromatography, precipitation with specific anti-IgG3 antibody, and differential separation of IgG3 heavy chains using polyacrylamide gel electrophoresis. IgG from RA synovial cultures contained an average of 41% IgG3 (range, 8-97%) compared with 12% IgG3 (range, 6-17%) in the serum IgG of the same patients. A group of non-RA control lymphoid tissues (four lymph nodes and five tonsils) produced 23% of total IgG as the IgG3 subclass (range, 16-35%). An average of only 9% of the synovial IgG showed aggregation compatible with IgG-rheumatoid factor (IgG-RF). Purified IgG from some of the RA synovial culture supernatants also showed significant restriction when separated by isoelectric focusing. This restriction and the enrichment for the IgG3 subclass in the IgG from RA synovial cultures suggest that either an antigen in the inflamed joint is selectively stimulating an antibody in this subclass, or that significantly differences in the catabolic rate of this subclass are found in cultures of synovial tissue when compared with that occurring in intact patients.
W L Hoffman, M S Goldberg, J D Smiley
Several theories have been advanced to explain the elevation in urinary PCO2 during bicarbonate loading and include: (a) H+ secretion, (b) countercurrent system for CO2, (c) the "ampholyte" properties of bicarbonate, and (d) mixing of urine of disparate bicarbonate and butter concentrations. In this study microelectrodes were used to measure in situ and equilibrium pH (pHis and pHeq) and PCO2 in control and bicarbonate loaded rats before and after infusion of carbonic anhydrase. The disequilibrium pH method (pHdq = pHis - pHeq) was used to demonstrate H+ secretion. Control rats excreting an acid urine (pH = 6.04 +/- 0.06) failed to display a significant disequilibrium pH at the base (BCD), or tip (TCD) of the papillary collecting duct. Urine pH (7.54 +/- 0.12), and urine to blood (U-B) PCO2 increased significantly during NaHCO3 loading while PCO2 at the BCD and TCD also increased (95 +/- 4 and 122 +/- 4). Furthermore, an acid disequilibrium pH was present at both the BCD and TCD (-0.42 +/- 0.04 and -0.36 +/- 0.03) and was obliterated by carbonic anhydrase. Comparison of the PCO2 in the BCD or TCD with the adjacent vasa recta revealed similar values (r = 0.97). It is concluded that H+ secretion by the collecting duct into bicarbonate containing fluid with delayed dehydration of H2CO3, is the most likely determinant of the U-B PCO2 in alkaline urine. Similar values for PCO2 in the collecting duct and the adjacent vasa recta suggests trapping of CO2 in the medullary countercurrent system. The rise in PCO2 occurs both along the collecting duct and after exit from the papilla.
T D DuBose Jr
Urine was observed to flow intermittently in the collecting ducts of the extrarenal papilla of antidiuretic rats. The purpose of this investigation was to test Reinking and Schmidt-Nielsen's hypothesis that intermittent flow plays an important role in the production of maximally concentrated urine. Samples of collecting duct fluid were obtained from the base and tip of the papilla by micropuncture through the intact ureter. Fluid osmolality rose sharply from base, 894±120 mosmol/kg H2O−1 (mean±SE), to tip, 1,667±114 (P<0.001), a distance of only 2 mm, and was due exclusively to reabsorption of water. After excision of the ureter, which abolished intermittent flow, osmolality fell modestly at the base to 723±82 mosmol/kg H2O−1 (P < 0.02), but strikingly at the tip to 1,012±103 (P < 0.001). The pelvic ureter was paralyzed by topical verapamil and dimethylsulfoxide, which abolished intermittent flow. Osmolality of urine at the tip was not changed (1,959±184 mosmol/kg H2O−1 before, vs. 1,957±126 after paralysis). The ureter was severed just beyond the papillary tip, a maneuver which preserved intermittent flow but abolished urinary reflux over the papilla. Urinary osmolality fell from 1,876±134 mosmol/kg H2O−1 to 1,284±115 (P < 0.005). These findings demonstrate that when the ureter is intact, over half of the increase in urinary osmolality above isotonicity occurs in the terminal one-fourth of the medullary collecting duct and is due exclusively to water reabsorption (no net solute addition). It is the continuity of the ureter, rather than intermittent flow due to ureteral peristalsis, which is essential for the formation of a maximally concentrated urine.
Richard E. Oliver, Denis R. Roy, Rex L. Jamison
Micropuncture and microcatheterization studies have been used extensively to investigate the pathophysiologic alterations in renal function induced by urinary tract obstruction. The present isolated tubule microperfusion studies were designed to examine the intrinsic alterations in segmental nephron function induced by 24 h of bilateral (BUO) and unilateral (UUO) urinary tract obstruction.
Michael J. Hanley, Karen Davidson
Whereas the cardiac effects of digitalis glycosides have been extensively studied, less is known of the extracardiac effects of the drug, in particular the effects on vascular capacity. We investigated the effects of parenteral ouabain on vascular capacity in the dog with particular emphasis on transhepatic resistance and its interaction with splanchnic and total intravascular capacity. We studied 49 dogs on total cardiopulmonary bypass in which the splanchnic and extrasplanchnic circulations could be separately perfused and drained, and the portal vein could be vented to systemic venous pressure. The results indicate: (a) ouabain produces a net central displacement of blood at 30 min after administration of 150 +/- 70 ml (SEM), (b) this displacement occurs despite a substantial increase in transhepatic resistance, although the early rise in transhepatic resistance may delay the net displacement of blood, and (c) the decrease of overall vascular capacity is due to an effect of ouabain on the capacitance vessels of both the splanchnic and extrasplanchnic circulations. The peripheral vascular capacity effects of ouabain may therefore contribute to overall cardiac performance.
M R Goldman, S W Wolk, D L Rutlen, W J Powell Jr
Autologous immune complex nephropathy (AICN), an experimental model for human membranous glomerulopathy, is characterized by marked heterogeneity in function from glomerulus to glomerulus. However, the fraction of the filtered load of fluid reabsorbed by the proximal tubule remains nearly constant from nephron to nephron, despite wide variation in single nephron glomerular filtration rate (SNGFR). To define the physiological mechanisms responsible for this marked variation in SNGFR values within a given kidney and for the remarkable preservation of glomerulotubular balance, the various determinants of fluid exchange across glomerular and peritubular capillary networks were evaluated in Munich-Wistar rats with AICN. For comparison, similar measurements were obtained in rats with the functionally more homogeneous lesion of heterologous immune complex nephropathy. In AICN rats studied ∼5 mo after injection of renal tubule epithelial antigen (Fx1A), a high degree of glomerulus-proximal tubule balance was found, despite marked variations in SNGFR values within a single kidney. These changes were associated with marked heterogeneity in immunoglobulin and complement deposition within and among glomeruli. Although mean capillary hydraulic pressure and Bowman's space hydraulic pressure ranged widely from glomerulus to glomerulus, the mean glomerular transcapillary hydraulic pressure difference was remarkably uniform among these functionally diverse glomeruli and could not, therefore, be implicated as the cause of the dispersion in SNGFR values. The two remaining determinants of SNGFR, namely, glomerular plasma flow rate (QA) and ultrafiltration coefficient (Kf), varied markedly from glomerulus to glomerulus, but always in direct proportion to SNGFR, and proved to be responsible for the marked variation in SNGFR.
I. Ichikawa, J. R. Hoyer, M. W. Seiler, B. M. Brenner
The purpose of the present study was to define myocardial and blood thallium-201 (Tl-201) kinetics after infusion of dipyridamole in normal canine myocardium and in myocardium distal to a coronary artery stenosis. Miniature radiation detector probes were implanted in the left ventricle in 39 open-chest dogs. A balloon constrictor was placed around the proximal left circumflex coronary artery. Electromagnetic flow probes were positioned proximally around both the left circumflex and left anterior descending coronary arteries. In five control dogs (group 1) the balloon occluder was not inflated; in 12 dogs (group 2) a mild stenosis was created such that resting flow was not reduced, yet the hyperemic response after 10 s of total occlusion was partially attenuated; in nine dogs (group 3) a moderate stenosis was created such that resting flow was not reduced, yet the hyperemic response was completely eliminated; and in 13 dogs (group 4) a severe stenosis was created such that resting flow was reduced. After intravenous dipyridamole (0.08 mg/kg . min-1 x 4 min), 1.5 mCi Tl-201 was injected intravenously and probe counts were collected continuously for 4 h. The mean 4-h fractional myocardial Tl-201 clearance for nonstenotic zones was 0.35, 0.27 for group 2 stenotic zones, 0.19 for group 3 stenotic zones, and 0.05 for group 4 stenotic zones (P less than 0.0001). After reaching peak activity, myocardial Tl-201 activity cleared biexponentially with a final decay constant lambda 2 = 0.0017 +/- 0.0001 min-1 (SE) for nonstenotic zones, 0.0011 +/- 0.0001 min-1 for group 2 stenotic zones, and 0.0006 +/- 0.0001 min-1 for group 3 stenotic zones (P less than 0.01). Group 4 stenotic zone Tl-201 clearances were negligible (decay constant essentially zero). Blood Tl-201 activity decayed triexponentially with a final blood lambda 3 = 0.0018 +/- 0.0001 min-1, which was almost identical to the final myocardial lambda 2 decay constant. Thus, the rate of myocardial Tl-201 clearance can distinguish between coronary stenoses of graded hemodynamic severity. These results may be applicable to quantitative techniques for determining myocardial Tl-201 clearance rates on serial clinical images after dipyridamole administration.
R D Okada, J A Leppo, C A Boucher, G M Pohost
The effect of biliary diversion on intestinal apolipoprotein (apoA)-I and high density lipoprotein formation was studied in mesenteric lymph fistula rats. Bile diversion was produced by an exteriorized catheter that allowed interruption and reconstitution of the enterohepatic circulation. Bile diversion reduced lymph cholesterol output from 0.47±0.05 μmol/h to 0.17±0.03 μmol/h (P < 0.025), and lymph triglyceride output from 3.6±0.3μmol/h to 0.6±0.05 μmol/h (P < 0.025) after 24 h. This was due to depletion of lymph chylomicrons and very low density lipoprotein (VLDL). Despite the reduced lipid outputs, lymph apoA-I output was maintained during biliary diversion (basal: 119±15 μg/h; diverted 140±20 μg/h, n = 12). During biliary diversion, high density lipoprotein (HDL) were maintained in mesenteric lymph as shown by lipoprotein and immunoelectrophoresis. Bile diversion altered the lipid composition of lymph HDL. Bile-diverted lymph HDL was depleted in total cholesterol and has a greater phospholipid/cholesterol ester ratio than basal lymph HDL. Lymph HDL contained discoidal particles when examined by negative stain electron microscopy. Bile diversion was associated with a reduction in the size of discoidal HDL particles (basal, nondiverted, 165±7Å (n = 112) compared with diverted 126±5Å (n = 98, P < 0.025). Experiments were then carried out to determine the source of the apoA-I and HDL found in lymph from bile-diverted animals. The transfer of HDL from plasma into lymph was determined by the intravenous infusion of 125I-apoA-I labeled HDL into lymph fistula rats. In both nonbile-diverted and diverted rats, the specific activity of apoA-I in the HDL fraction of lymph was 23% of the specific activity of apoA-I in plasma HDL, indicating that the major portion (75%) of mesenteric lymph apoA-I did not come from plasma filtration. In other experiments the intraduodenal infusion of [3H]leucine to bile fistula, lymph fistula rats resulted in relative fivefold increase in the specific activity in apoA-I in lymph HDL when compared with the specific activity of apoA-I in plasma HDL from the same animal. We conclude that intestinal apoA-I secretion is maintained during biliary diversion and that synthesis of this apoprotein occurs in the absence of chylomicron formation. We also conclude that discoidal HDL are present in mesenteric lymph despite reduced triglyceride absorption and secretion into lymph.
H. Robert Bearnot, Robert M. Glickman, Lee Weinberg, Peter H. R. Green, Alan R. Tall
The amount and type of cholecystokinin (CCK) in duodenal extracts and plasma of celiac patients and normal subjects was studied by radioimmunoassay and gel filtration. In both groups there were similar patterns of molecular forms in extracts of duodenal biopsies, but concentrations in celiac disease were significantly depressed. In boiling water extracts of duodenal mucosa from both groups a factor with the properties of the COOH-terminal octapeptide of cholecystokinin predominated, but there were also significant amounts of a larger molecular weight form. In acid extracts of mucosa a factor with the properties of the 33 or 39 residue form was identified in amounts that were approximately 25% those of CCK8; there were also similar amounts of an acid-soluble form that had an apparent molecular weight higher than CCK39. Plasma immunoreactive cholecystokinin was studied after concentration by immunoaffinity adsorption and fractionation by gel filtration. In normal subjects fasting CCK-like immunoreactivity was less than 0.8 pmol/liter, and after a light breakfast increased to 2.0 +/- 0.7 (range 1.0 to 4.8) pmol/liter; CCK8-like activity accounted for all the increased immunoreactivity. In five of six celiac patients the concentrations of both fasting and postprandial CCK-like immunoreactivity in plasma were undetectable (less than 0.8 pmol/liter). We conclude that diminished production and release of CCK could account for the impaired pancreatic and gall bladder responses to intraluminal stimuli in celiac disease.
J Calam, A Ellis, G J Dockray
To assess for possible inhibition of cellular transmethylation during adenine arabinoside (ara-A) therapy, S-adenosylhomocysteine hydrolase activity was analyzed in 10 patients with chronic hepatitis B virus infection. In six patients receiving ara-A, enzyme activity was suppressed to 0-2% of control erythrocyte enzyme activity. This decrease in enzyme activity was evident within 4 h of starting the drug infusion and continued for 7 d after cessation of therapy. S-adenosylhomocysteine hydrolase activity of peripheral mononuclear cells was also measured in two patients receiving ara-A. Suppression to as low as 3.5% of pretreatment levels was found; however, marked fluctuations with partial return of enzyme activity during therapy was also observed in mononuclear cells. Inhibition of an enzyme involved in transmethylation reactions was observed in patients during ara-A therapy. This could contribute to the side effects and antiviral properties of ara-A.
S L Sacks, T C Merigan, J Kaminska, I H Fox
The objectives of this investigation were: (a) to characterize the time and dose dependence of the effects of prostacyclin (PGI2) on renin release in healthy men; (b) to define whether PGI2-induced renin release is secondary to hemodynamic changes; (c) to determine the plasma and urine concentrations of 6-keto-PGF1α (the stable breakdown product of PGI2) associated with renin release induced by exogenous or pharmacologically enhanced endogenous PGI2. Intravenous PGI2 or 6-keto-PGF1α infusions at nominal rates of 2.5, 5.0, 10.0, and 20.0 ng/kg per min were performed in each of six normal human subjects; in three of them, PGI2 infusion was repeated after β-adrenergic blockade and cyclooxygenase inhibition. PGI2, but not 6-keto-PGF1α, caused a time- and dose-dependent increase of plasma renin activity, which reached statistical significance at 5.0 ng/kg per min and was still significantly elevated 30 min after discontinuing the infusion. Although combined propranolol and indomethacin treatment significantly enhanced the hypotensive effects of infused PGI2, it did not modify the dose-related pattern of PGI2-induced renin release.
Carlo Patrono, Francesco Pugliese, Giovanni Ciabattoni, Paola Patrignani
Total renal ammonia production and ammonia precursor utilization were evaluated in patients under normal acid-base balance and in patients with 24-h NH4Cl acidosis by measuring (a) ammonia excreted with urine and that added to renal venous blood, and (b) amino acid exchange across the kidney. In 24-h acidosis not only urinary ammonia excretion is increased, but also total ammonia production is augmented (P less than 0.005) in comparison with controls. By evaluating the individual role of acid-base parameters, urine pH and urine flow in influencing renal ammonia production, it was shown that the degree of acidosis and urine flow are likely major factors stimulating ammoniagenesis. Both urine pH and urine flow are determinant in the preferential shift of ammonia into urine. In 1-d acidosis, renal extraction of glutamine was not increased and the total ammonia produced/glutamine N extracted ratio was higher than in controls (P less than 0.005) and was inversely correlated with the log of arterial bicarbonate concentration (P less than 0.001). In the same condition, renal glycine and ornithine uptake took place; the more severe the acidosis, the greater was the renal extraction of these amino acids (P less than 0.001). These data indicate that at the early stages of metabolic acidosis, in spite of a brisk increase in ammonia production, the mechanisms responsible for the increased glutamine use, which are operative in chronic acidosis, are not activated and other ammonia precursors, besides glutamine, are probably used for ammonia production.
A Tizianello, G Deferrari, G Garibotto, C Robaudo, N Acquarone, G M Ghiggeri