SummaryFour monoclonal antibodies (N1, N3, N4 and N5) with specificity for the N blood group have been produced from fusions in which human erythrocytes were used as the immunizing stimulus. N1 and N3 differed from N4 and N5 with regard to their serological specificities and pH optima. Antibody N3 has been assessed for its suitability as a blood grouping reagent by testing 10,767 blood donors in parallel with Vicia graminea anti‐N lectin or rabbit anti‐N. There was complete correlation between the results obtained with the monoclonal and conventional reagents providing strong evidence that N3 is a reliable and potent blood grouping reagent.
Summary The study included 138 A. sobria and 182 A. hydrophila isolated in Perth from samples of diarrhoeal or non‐diarrhoeal faeces or from domestic water. Strains were grouped i n relation to agglutination of human, horse, rat and guinea pig erythrocytes and the effect of sugars on haemagglutination. Agglutination of red cells of all four species (primary group 1) was most commonly associated with A. sobria , particularly those strains isolated from faeces of patients with diarrhoea. Most A. hydrophila associated with diarrhoea also belonged to group 1 but A. hydrophila from non‐diarrhoeal stools or from water most commonly agglutinated human and guinea pig cells but not horse erythrocytes (primary groups 2 and 3). Fucose‐resistant haemagglutination (FRHA) of strains in primary group 1 occurred with about 68% of the strains of A. sobria associated with diarrhoea. Mannose‐resistant haemagglutination (MRHA) was limited to 29% of strains of A. sobria associated with diarrhoea. The predominance of primary group 1 among strains of Aeromonas spp. associated with diarrhoea and the proportion of these strains showing FRHA suggest that haemagglutination of cells from human, horse, rat and guinea pig, particularly if fucose‐resistant, should be considered in a search for characteristics, which possibly contribute to virulence of Aeromonas spp.
Summary Specimens of normal and asthmatic lungs were studied at the electron microscopic level and the frequency and ultrastructural features of mast cells and their granules within the alveolar wall were assessed with morphometric techniques. The numerical density of mast cells per square millimetre of alveolar wall was 299 (SD = 258) in normal and 366 (SD = 260) in asthmatic lung. The mean area of the mast cell nucleated profile was 25·7 μm 2 (SD = 6·3) in normal lung and 29·8 (SD = 6·2) in asthmatic. The average number of secretory granules per single mast cell nucleated profile was 55 (SD = 13) in normal lung and 60 (SD = 12) in asthmatic lung. The diameter of the individual secretory granule was 338·9 nm (SD = 42·6) in normal and 345·6 (SD = 47·7) in asthmatic lung. The volume density of secretory granules in normal and asthmatic lung was 6·31 μm 3 and 5·81 μm 3 respectively. The mean diameter of the individual subunit (“scroll”) inside the secretory granule was 88·8 nm for both normal and asthmatic lung. In normal lung 64·2% of granules were of‘scroll’ and ‘combined’ type, and 35·8% of granules were “particulate” or ‘empty’. In specimens from asthmatic patients 40·3% of granules had ‘scroll’ or ‘combined’ structures and 59·7% were ‘particulate’ or ‘empty’. Our data suggest that there is no difference between the number of mast cells in normal and asthmatic lung. However, in pulmonary mast cells from asthmatic lung, degranulation is more common than in normal lung.
Depressed IgG responses to ovalbumin administered with the inflammatory adjuvant, Al(OH)3, have previously been demonstrated in mice immunized 5 to 13 days after infection with Nematospiroides dubius, when the parasite is encysted in the intestinal mucosa or becoming established in the lumen. This report extends these findings to show that primary IgE and delayed-type hypersensitivity (DTH) responses, the generation of antigen-primed cells detectable in syngeneic irradiated recipients or in vitro and the recall of immunological memory are correspondingly impaired. Co-transfer experiments using irradiated recipients demonstrated weak suppressor activity in spleen cells from infected mice which was not attributable to T cells. The results suggest that N. dubius administered before immunization can impair the development of many classes of antigen-primed lymphocytes. The implications of this finding with respect to parasite survival are discussed.
The contents of endogenous noradrenaline (NA) in rabbit gingiva and dental pulp were approximately 15-fold greater than either adrenaline (A) or dopamine (DA). Whereas catecholamine contents of maxillary incisor pulps were similar to those in mandibular incisor pulps, variations existed between gingiva excised from different regions. Both reserpine pretreatment and chronic sympathetic denervation greatly decreased the NA contents. These findings support the view that NA is the sympathetic neurotransmitter in dental pulp and gingiva.
SummaryThe genetic relatedness of ten Murray Valley encephalitis virus (MVE) isolates from Australia has been examined by comparing Haelll and Taql restriction digest profiles of cDNA to virion RNA. The isolates were from the Murray Valley region of south‐eastern Australia and from the Ord River region of Western Australia and spanned a period of 23 years (1951‐1974). The isolates generated closely similar restriction digest profiles. The extent of similarity suggested that the level of nucleotide sequence divergence between any pair of Australian MVE isolates is probably around 1%. The genetic homogeneity of the MVE isolates contrasts with results obtained for Ross River virus, an alphavirus, using an identical methodological approach; we propose that this difference results from the important role of birds in the life cycle of MVE.Four MVE isolates from three fatal human cases showed small genetic differences on from the other. These isolates did not have a common restriction digest profile which distinguished them from strains obtained from other sources (e.g., from mosquitoes or a heron) The data do not support the view that clinical cases of MVE infection in humans are to a particular strain of virus although this has not been rigorously excluded.The two available MVE isolates from Papua New Guinea (PNG) were from the Sepik and Port Moresby regions. They generated Haelll and Taql restriction digest profiles which were different both from each other and from those of the Australian type. Genetic divergence between the two PNG isolates was estimated to be ≈ 6%; divergence between either of PNG isolates and the Australian type was >6%. Our data suggest that the evolution of MVE i n Australia and PNG has proceeded independently and that circulating Australian MVE strains are not systematically re‐seeded from regions of endemicity in PNG.Studies on the relatedness of MVE and two close antigenic relatives, Japanese encephalitis virus (JE) and Alfuy virus (ALF), showed that the genetic relatedness between any isolate and JE or ALF is less than that between the most divergent of the MVE isolates, including those from Papua New Guinea.Some epidemiologieal implications of the results are discussed.
Human neutrophils incubated with sodium arachidonate generated hydroxyl radicals. The radical formed an adduct with the spin trap 5', 5-dimethyl-l-pyrroline-N-oxide (DMPO) and was subsequently detected by electron spin resonance (ESR) spectroscopy. The ESR signal was inhibited by mannitol and superoxide dismutase but not by catalase. Removal of glucose from the reaction mixture or the presence of glucose metabolic inhibitors including 2-deoxy-D-glucose and 3-O-methyl-D-glucose did not affect the production of hydroxyl radical by the neutrophils. The ESR signal was, however, inhibited by the lipoxygenase inhibitors nordihydroguaiaretic acid and N-ethylmaleimide. The involvement of lipoxygenase in the production of hydroxyl radical was demonstrated by the trapping of the radical with DMPO in a reaction mixture of soybean lipoxygenase and arachidonic acid (AA). These findings support our previous postulation that the metabolism of AA via the lipoxygenase pathway is a source of hydroxyl radical in stimulated neutrophils.
MK351A, a tyrosyl analogue of enalaprilic acid (MK422) is a potent inhibitor of angiotensin-converting enzyme (ACE). MK351A was radioiodinated with 125I and used to develop a radio inhibitor binding assay for human serum ACE. 125I MK351A associated rapidly and reversibly with human serum ACE (T 1/2 = 1/2 h). Bound 125I MK351A was displaced by an excess of cold MK351A, to give non-specific binding of less than 1%. Scatchard analysis of binding was linear (r = -0.99, n = 6, p less than 0.001), indicating a single class of binding site. ACE was estimated in serum from normal patients and patients with sarcoid by the radio inhibitor binding assay and by enzyme kinetic assay using Hip-His-Leu as substrate. The two methods for ACE estimation correlated closely (r = 0.87, n = 82, p less than 0.001). The radio inhibitor binding assay for human serum ACE is a simple, sensitive and specific assay which utilizes novel assay methodology.
SummaryFive groups of male rhesus monkeys, Gp. I–sham‐vasectomized stock diet‐fed (SVS), Gp. II–vasectomized stock diet‐fed (VS), Gp. III–sham‐vasectomized atherogenic diet‐fed (SVA), Gp. IV–vasectomized atherogenic diet‐fed (VA) and Gp. V–atherogenic diet‐fed vasectomized monkeys (AV) were observed for a period of 1½ years. Vasectomy per se had no effect on the level of serum or tissue lipids either alone or in combination with atherogenic diet feeding. Following vasectomy the involvement of aorta by fatty lesions was significantly increased and this occurred in animals of all four experimental groups when compared with the control animals. The maximum increase was noted in group IV. Again, there was a significant increase in the frequency of aortic plaques in groups II, III and IV while it was increased in coronary arteries of all the experimental animals (Gps. II, III, IV and V). Group IV monkeys showed higher frequency of plaques as compared to group V animals. Although the plaque height was increased in all the vessels of the experimental animals, significant differences were observed only in the coronary arteries of group II monkeys and in intra‐cranial cerebral vessels of group III animals. This study shows that vasectomy per se tends to increase atherosclerosis in rhesus monkeys.
Pulmonary macrophages from normal subjects and asthmatic patients were examined for the presence of sugar residues on their surface. The technique of bronchoalveolar lavage was employed to obtain cell samples. Ultrastructural and cytophotometric methods were used for studying the patterns of lectin binding by these two groups of macrophages. Three lectins, Concanavalin A (Con A), Wheat germ agglutinin (WGA) and Ricinus communis agglutinin (RCA), were used in this investigation. Pulmonary macrophages from both normal and asthmatic persons revealed a high level of Con A, WGA and RCA binding. The distribution of the electron dense reaction product on the macrophage surfaces was relatively uniform. Quantitative cytophotometric studies showed that the level of binding of Con A by macrophages from both groups was approximately the same. Similar results were obtained with WGA--the difference between macrophages from normal and asthmatic persons was not statistically significant. In the case of RCA, macrophages from asthmatic patients showed a higher level of lectin binding than macrophages from normal persons. The conclusion is made that macrophages from asthmatic persons have more D-galactose residues on their surface.
Depletion in the hepatic concentration of reduced glutathione (GSH) may potentiate liver injury resulting from toxic metabolites. Patients who have had severe haemorrhage frequently develop hepatic dysfunction and we have shown that the hepatic GSH level is decreased in these patients. The present study has examined the time-course effect of a 30% haemorrhage on the hepatic GSH level by sequential biopsies of the liver. Acute and chronic experiments were performed and, in both, the hepatic concentration of GSH fell during the first 6 h after haemorrhage; this fall was followed by a significant rebound elevation at 24 h. In the chronic haemorrhage experiment the hepatic GSH level was normal at 1 week after haemorrhage. Thus, the susceptibility of the liver to toxic metabolite injury after haemorrhage persists for less than 24 h in this experimental model and haemorrhage appears to have no long term effect on the hepatic GSH concentration.
Five conscious rabbits each received a single injection of almitrine bismesylate via a marginal ear vein. Measurements were made of ventilation and breathing pattern using a barometric method, and blood gases, pH and lactate concentration ([Lac]) were measured from arterial blood samples throughout the hour following the injection. Almitrine caused an immediate increase in ventilation through an increase in tidal volume (VT), frequency (f) showing a small decline. After 15 min f increased to above control level and VT declined. Slower, deeper breathing was reinstated by 40 min post-injection. Large changes in blood gases occurred in the first 15 min, with PO2 raised by 30 Torr and PCO2 lowered by 20 Torr. These recovered to be within 10 and 7.5 Torr of control at the end of 1 h. [Lac] increased steeply at first, then declined towards control levels. The acid-base situation was mixed, comprising a respiratory alkalosis, resulting from CO2 washout, and a metabolic acidosis from high [Lac]. These results are discussed in the context of assessing the primary and secondary effects of almitrine and recognition of the possibility that the drug may act at more than one site to alter respiration.
Lactate dehydrogenase, E.C.1.1.1.27 (LDH) from the simian malarial parasite, Plasmodium knowlesi, and from normal rhesus monkey erythrocytes has been purified using Blue Sepharose affinity chromatography, and the properties of the purified enzyme from these two sources have been compared. The enzyme from the host and parasite were different in their kinetic properties, viz., substrate and pH optima, thermostability and isoenzymic behaviour. Partially purified LDH of the erythrocytes resolved into three isomeric bands on polyacrylamide gel electrophoresis, whereas the parasite LDH moved as a single enzyme band of different mobility from the host LDH. The molecular mass of the parasite enzyme was estimated as 117,500 daltons.
SummaryWe investigated the distribution of the nuclear encoded mitochondrial enzymes, carbamylphosphate synthetase (CPS; EC 6.3.4.16) and ornithine transcarbamylase (OTC; EC 2.1.3.3) in liver by immunocytochemistry on ultrathin sections using the protein A‐gold technique. Both enzymes were found to be present as aggregates in the cytoplasm of hepatocytes, in association with ER membranes adjacent to mitochondria. Clusters of the enzymes were also found inside the mitochondria. The aggregation of these enzymes was found only with antibodies to CPS and OTC and not with antibodies against albumin or with IgG from unimmunized serum, nor were aggregates found in cells other than hepatocytes. The results are suggestive of localized uptake of clusters of enzyme or co‐translational uptake of enzyme at discrete localizations and that endoplasmic reticulum (ER) associations may be necessary for uptake of the precursor forms of CPS and OTC. The possible involvement is discussed of micropinosomes which are seen associated with inner membrane, intermembrane space and outer membrane in mitochrondria obtained from a perinuclear pellet where ER and mitochondria are frequently found in close association.
The functional characteristics of peritoneal cells (PC's) acquired in response to infection with Nematospiroides dubius were studied to identify changes which could be associated with the high level, long-term survival of this parasite or explain its transient immunosuppressive properties. The number of cells in the peritoneal cavity increased steadily during the infection, and the population displayed a transient hyperresponsiveness to inflammatory stimulation. N. dubius did not affect the ability of antigen-pulsed PC's to induce humoral responses or delayed-type hypersensitivity (DTH) in primed recipients, but lymphoproliferative responses induced in vitro were depressed by PC's from infected donors. In addition, unpulsed PC's from N. dubius-infected mice depressed proliferative responses to parasite antigens by autologous mesenteric lymph node cells. These effects are attributable to increased lymphocytostatic activity in the PC's, which peaked one week after infection and correlated with the expansion of a population of large plastic-adherent vacuolated cells with elevated acid phosphatase activity (activated macrophages). Comparable inhibitory cells did not develop in the same period of time during a short-lived infection with Nippostrongylus brasiliensis.
Prior treatment (priming) with a weakly immunogenic dose of Type III pneumococcal polysaccharide (SIII) results in the development of an antigen-specific state of unresponsiveness termed low-dose paralysis which is believed to be mediated by suppressor T cells. The present findings show that the passive administration of functionally distinct non-complement-fixing (NCF) IgM anti-SIII antibodies either in monoclonal form or from protein-A absorbed immune serum could significantly suppress the direct plaque-forming cell (PFC) response to an immunogenic dose of SIII administered concurrently. The degree of suppression was comparable with that induced by low-dose paralysis. Low-dose paralysis was consistently induced in athymic (nude) mice 4 days, but not 3 days, after priming with a low dose of SIII, and was associated with the delayed appearance of NCF-IgM anti-SIII in the serum of athymic mice. In contrast, low-dose paralysis was readily induced in normal BALB/c mice 3 days after priming when NCF-IgM anti-SIII antibodies were present. Comparable inhibiton of the direct anti-SIII PFC response was observed when Concanavalin A (Con A) or NCF-IgM anti-SIII serum was administered with SIII antigen. That Con A and NCF-IgM anti-SIII together did not produce additive suppression was attributed to the adsorption of NCF-IgM anti-SIII antibodies to Con A. Complement-dependent single radial haemolysis mediated by CF hybrid IgM/A or CF-IgM anti-SIII serum was blocked by monoclonal NCF-IgM or IgA anti-SIII antibodies and indicated that each of the antibodies was specific for the same SIII-determinant. Evidence is presented to show that low-dose paralysis in the CF-IgM response to SIII is not mediated by suppressor T cells but can be attributed to highly avid NCF-IgM anti-SIII antibodies, formed preferentially to low doses of SIII, being able to reduce the immunogenicity of SIII administered subsequently. We propose that low-dose paralysis to SIII is the result of an immunobiological function of highly avid NCF-IgM anti-SIII antibodies which not only confer resistance against capsulated pneumococci but preferentially bind soluble SIII-antigen to reduce its immunogenicity and thereby protect specific CF-IgM positive B cells from being rendered tolerant by direct contact with higher doses of SIII antigen.
The influence of variations in protocol on the induction and early stages of maintenance of allograft tolerance in neonatal rats was studied. Allogeneic bone marrow cells, the anti-recipient activity of which was specifically reduced because of their origin from immunologically tolerant donors, possessed diminished tolerogenic capacity. This was commonly manifested by development of an unreactive state in only one of the two parameters of reactivity--allograft rejection and graft-versus-host reactivity--that were monitored. A similar deviation from the occurrence of unreactivity in both forms that characterizes tolerant rats was observed in animals which had been inoculated with syngeneic thymus cells at the time of induction of tolerance. Attempts to demonstrate a requirement for an extended period of exposure of young rats to semi-allogeneic bone marrow cells before specific unreactivity on the part of host cells became irreversible were unsuccessful.
Summary As part of a project to study the effect of copper deficiency (CD) on bone development in young dogs, the composition and metabolism of proteoglycans (PGs) and extractability of collagens in the epiphyseal growth plate cartilage (EGPC) and articular cartilages (AC) were investigated. Copper deficiency was induced by feeding ammonium tetra‐thiomolybdate (TTM) a copper antagonist. The collagen of cartilages from TTM‐treated animals was significantly more soluble in 0‐5 M saline than control tissues. While no distinction between TTM‐treated and control cartilages was evident in terms of PG content or extractability under associative (0‐5 M‐GuHCl) or dissociative (4‐0 M‐GuHCl) conditions, the sedimentation behaviour of the PG aggregates following CsCl density gradient ultracentrifugation suggested less polydispersity of PGs in preparations from the TTM‐treated animals. Moreover, analysis of the PG monomers from EGPC of TTM animals showed galactosamine/glucosamine ratios higher than control preparations, suggesting a reduced keratan sulphate content in these preparations. Organ culture of EGPC showed a significant reduction in the incorporation of 35 S into PGs and of 3 H‐thymidine into DNA in the tissues of TTM‐treated animals relative to controls. From these findings we deduce that the catabolism of PGs and the extent of collagen cross‐linking in EGPC of TTM‐treated animals may be reduced relative to age‐matched control tissues.
Female guinea pigs, inoculated intravaginally with a herpes simplex virus-type 2 (HSV-2) strain M1, developed typical symptoms of a primary and recurrent genital herpes infection. Sera from HSV-2 infected guinea pigs taken during the primary or recurrent stages of the genital infection contained complement (C)-fixing antibody which reacted with an apparent type specificity to an early 4 h HSV-2 infected cell extract (AG-4) when compared to a 4 h HSV-1 infected cell extract. This C-fixing anti-AG-4 activity was shown to be associated with the IgG2 subclass and directed primarily against HSV-2 infected cell polypeptide (ICP)6 and ICP8. Furthermore, C-fixing anti-AG-4 levels remained constant after the primary infection and during recurrences for over 6 months. Thus, while the anti-AG-4 response in guinea pigs was of the IgG type, it varied from that found previously in human genital HSV-2 sufferers which was of the IgM type. Also, while an IgG anti-AG-4 response in human genital HSV-2 sufferers is associated with a reduction in the number of recurrences, this reduction is not apparent in the guinea pig model.
