Serum immunoconglutinins and complement in systemicLupus erythematosus

Summary IK activity titrated by the sedimentation method in sera from patients affected with SLE was found to be negatively correlated with C4 and C3 complement factors levels. The significance of this data is discussed.Index of the abbreviations used: SLE, systemicLupus eritematosus; IK, immunoconglutinins; CH50, Total haemolytic activity; C4b, Complement factor connect with the complex AgAbCl; C3b, Complement factor connect with the complex AgAbClC4C2; C3PA, C3 proactivator in the complement activation by the alternate pathway implicated; KAF, Conglutinogen activating factor inactive the C3b in C3c and C3d. Antrypol, factor that makes C3b KAF-unreactive.     

Haemolytic complement consumption by Parietaria pollen extracts in relation to peptide-bound flavonoids

Inhaled allergens from house dust, mites or animal danders activate human complement in vitro by engaging the C1-component through a non-antibody-dependent mechanism. These earlier findings are extended by showing that the allergenic components in extracts of Parietaria pollen are almost equally potent complement activators as those from house dust or mites. Spectroscopic evidence indicates that haemolytic complement consumption by the Parietaria allergens and their enzymic fragments is most likely related to post-translational side-chains comprising flavonoid derivatives. These adsorbed and/or peptide-bound tannin-like structures may also explain the exceptional stability of the high- and low-molecular mass allergenic components in Parietaria pollen extracts. Received 12 November 1996; received after revision 12 December 1996; accepted 17 December 1996     

Relation entre l'activité de la glande thyroÏde et la consommation d'oxygène chez les Téléostéens,Cichlidés

Summary The study of the relationship between the activity of the thyroid of Cichlid fishes and oxygen consumption has given the following results: The stimulation by thyroxine causes a significant increase of oxygen consumption (+35%) after a long treatment. This increase is not immediate; a latent period is observed till the gland is in repose. The antithyroid rapidly decreases the oxygen consumption (–24%). But this chemical blocking is not stable.     

Die Wirkung der Askorbinsäure auf die Katalaseinduktion in Mikroorganismenkulturen

Summary Catalase activity ofStaphylococcus aureus increases the effect of ascorbic acid. Namely the ascorbic acid decreases the dissolved oxygen content of the medium, and low oxygen tension is the natural inducer of catalase synthesis.     

In vivo effects of Sumithion on tissue respiration and enzyme activity in the fish,Etroplus maculatus

Summary LD₅₀ exposure of a teleost fish.Etroplus maculatus, to Sumithion depressed the rate of oxygen consumption, concomitantly with an inhibition in succinate dehydrogenase activity in the tissues, in the order gill>brain>liver>muscle. The effect of the same pesticide on the activity of acetylcholinesterase was quite interesting, with a maximal inhibitory effect on the brain followed by liver, muscle and gill, suggesting a tissue specificity, and a differential sensitivity of the enzyme towards the pesticide.I wish to express may gratitude to Dr K. Sasira Babu and Dr J. Hemalatha for their valuable discussions and encouragement.     

Complement factor H in host defense and immune evasion

Complement is the major humoral component of the innate immune system. It recognizes pathogen- and damage-associated molecular patterns, and initiates the immune response in coordination with innate and adaptive immunity. When activated, the complement system unleashes powerful cytotoxic and inflammatory mechanisms, and thus its tight control is crucial to prevent damage to host tissues and allow restoration of immune homeostasis. Factor H is the major soluble inhibitor of complement, where its binding to self markers (i.e., particular glycan structures) prevents complement activation and amplification on host surfaces. Not surprisingly, mutations and polymorphisms that affect recognition of self by factor H are associated with diseases of complement dysregulation, such as age-related macular degeneration and atypical haemolytic uremic syndrome. In addition, pathogens (i.e., non-self) and cancer cells (i.e., altered-self) can hijack factor H to evade the immune response. Here we review recent (and not so recent) literature on the structure and function of factor H, including the emerging roles of this protein in the pathophysiology of infectious diseases and cancer.     

Agonist-induced internalization and desensitization of the human nociceptin receptor expressed in CHO cells

In this study, we examined agonist-induced internalization, recycling and signalling (measure of cAMP levels) of the cloned human nociceptin receptor (hNOP) expressed in CHO-K1 cells. Internalization was proven by a receptor-binding assay on viable cells. The agonist nociceptin/orphanin FQ (NC) promoted rapid internalization of the hNOP receptor (approximately 78% of cell surface receptors were lost after 2 min exposure to 1 microM NC) in a clathrin- and ATP-dependent manner. Internalization was more rapid and marked in CHO-K1 cells than, as we previously reported, in SK-N-BE cells. This difference may be related to higher levels of beta-arrestin isoforms detected in CHO-K1 than in SK-N-BE cells. hNOP receptor internalization was partially reversible and recycling occurred in the presence of the agonist; receptor recycling was dependent on okadaic acid-sensitive phosphatases and was blocked by monensin. Confocal microscopy analysis confirmed the internalization and the recycling back to the plasma membrane of an epitope-tagged hNOP receptor expressed in CHO-K1 cells. These receptors underwent rapid desensitization upon agonist challenge: NC efficacy in inhibiting forskolin-stimulated cAMP production was significantly reduced 10 min after exposure and correlated with the rate of receptor internalization. Moreover, we observed that blockade of hNOP receptor recycling by monensin would cause a more prolonged and relevant desensitization of this receptor. Thus, the dynamic cycle between hNOP receptor activation, internalization and recycling determines the activity of this receptor on the cell surface.     

17 Beta-estradiol-sensitivity of cultured myometrial cells

The estrogen sensitivity of cells cultured from the rat myometrium was studied by growing the cells in the absence or presence of 1 nM 17 beta-estradiol. Following a time lag of approximately 10 days, exposure to estrogen resulted in increased incorporation of radiothymidine by the cells. Estrogen treatment also decreased isoproterenol-dependent and GTP-dependent adenylate cyclase activity, but had no effect on basal activity. These cultured cells have been shown previously to have some properties of uterine smooth muscle. The effects estrogen has in vitro, therefore, may reflect important properties in vivo that account for the mechanism by which the sex steroid decreases the sensitivity of the myometrium to isoproterenol.     

Glioma cell activation by Alzheimer's peptide Abeta1-42, alpha1-antichymotrypsin,and their mixture

We compared the effects ofAlzheimer's peptide (Abeta1-42), a,-antichymotrypsin (ACT) and an ACT/Abeta1-42 mixture on human glioma DK-MG cells. The solution of Abeta (5 microM) formed by 2-h incubation at room temperature induced tumour necrosis factor-alpha (TNF-alpha) and interleukin (IL)-6 levels by 55 and 45%, respectively, and increased gelatinase B activity by 67%, while exposure of cells to the ACT/Abeta1-42 mixture (1:10 molar ratio ACT: Abeta1-42) under the same experimental conditions showed no effect on IL-6 levels or gelatinase B activity, but strongly induced TNF-alpha (by 190%), compared to the controls. Stimulation of the cells with Abeta1-42 alone, but not with ACT, increased by about 20% low-density lipoprotein (LDL) uptake and mRNA levels for LDL receptor and HMG-CoA reductase, while the ACT/Abeta1-42 mixture significantly increased LDL uptake (by 50%), up-regulated mRNA levels for LDL receptor and HMG-CoA reductase by 48 and 63%, respectively, and increased lipid accumulation by about 20-fold. These data suggest a possible new role for Abeta in Alzheimer's disease through its interaction with the inflammatory reactant, ACT.     

Lactate dehydrogenase and glutamate dehydrogenase activities in the circumventricular organs of rat brain following neonatal monosodium glutamate

Glutamate (glu) an excitatory neurotransmitter amino acid, is present in high concentrations in the mammalian central nervous system and is the most abundant amino acid in our daily diet. In the present study the activities of lactate dehydrogenase (LDH) and glutamate dehydrogenase (GDH) were evaluated in the circumventricular organs (CVO) of the brain in 25-day-old rats following MSG administration at a dose of 4 mg/g b.wt during the first ten days of life. The results show the LDH activity increased to 265% of that in the control (p<0.001), whereas GDH activity was significantly decreased (p<0.05), The great elevation in LDH, a cytoplasmic marker enzyme, is apparently due to cytoskeletal changes brought about as a consequence of glu toxicity, whereas lowered GDH activity indicates altered glu homostasis in the blood-brain-barrier deficient areas following neonatal exposure to glu.     

Formation of lipoperoxide in the retina of rabbit exposed to high concentration of oxygen.

When rabbit was exposed to high concentrations of oxygen, lipoperoxide in the retina was increased at 12 h of the exposure, after which period amplitude of electro-retinogram decreased. The degeneration was observed in the visual cell layer of the retina of the exposed animal.The exposure increased lipoperoxide in isolated retina. These data show the intervention of lipoperoxide in retinal degeneration by exposure to high concentration of oxygen.     

Formation of lipoperoxide in the retina of rabbit exposed to high concentration of oxygen

Summary When rabbit was exposed to high concentration of oxygen, lipoperoxide in the retina was increased at 12 h of the exposure, after which period amplitude of electro-retinogram decreased. The degeneration was observed in the visual cell layer of the retina of the exposed animal. The exposure increased lipoperoxide in isolated retina. These data show the intervention of lipoperoxide in retinal degeneration induced by exposure to high concentration of oxygen.     

Direct inhibition of phospholipid scrambling activity in erythrocytes by potassium ions

The exposure of phosphatidylserine (PS) at the cell surface plays a critical role in blood coagulation and serves as a macrophage recognition moiety for the engulfment of apoptotic cells. Previous observations have shown that a high extracellular [K⁺] and selective K⁺ channel blockers inhibit PS exposure in platelets and erythrocytes. Here we show that the rate of PS exposure in erythrocytes decreases by ~50% when the intracellular [K⁺] increases from 0 to physiological concentrations. Using resealed erythrocyte membranes, we further show that lipid scrambling is inducible by raising the intracellular [Ca²⁺] and that K⁺ ions have a direct inhibitory effect on this process. Lipid scrambling in resealed ghosts occurs in the absence of cell shrinkage and microvesicle formation, processes that are generally attributed to Ca²⁺-induced lipid scrambling in intact erythrocytes. Thus, opening of Ca²⁺-sensitive K⁺ channels causes loss of intracellular K⁺ that results in reduced intrinsic inhibitory effect of these ions on scramblase activity. Received 11 September 2008; received after revision 17 October 2008; accepted 27 October 2008     

Inhibition of IgM antibody-mediated aggregation of Trypanosoma gambiense in the presence of complement.

This paper deals with the immune reaction between Trypanosoma gambiense and monoclonal IgM mouse antibody at equivalence with or without rabbit complement. Antibody-mediated trypanosome clumps formed in the absence of complement, and were readily dissociated by complement to become free. In the presence of complement, on the other hand, T. gambiense were not aggregated by the antibody. Free parasites adhered readily to cultured peritoneal macrophages. Complement-mediated dissociation of the clumped trypanosomes in the equivalence area released a large number of previously bound surface antigens. These antigens were capable of binding again to fresh IgM antibody. Experimental results further indicated that the complement system caused a functional alteration, changing the multivalent nature of the IgM antibody in the immune complex into a univalent one. This phenomenon is of great advantage to the infected host in clearing pathogens in vivo, as it allows more antibodies to attach to trypanosomes and subsequently initiate complement activity.     

Inhibition of IgM antibody-mediated aggregation ofTrypanosoma gambiense in the presence of complement

This paper deals with the immune reaction betweenTrypanosoma gambiense and monoclonal IgM mouse antibody at equivalence with or without rabbit complement. Antibody-mediated trypanosome clumps formed in the absence of complement, and were readily dissociated by complement to become free. In the presence of complement, on the other hand,T. gambiense were not aggregated by the antibody. Free parasites adhered readily to cultured peritoneal macrophages. Complement-mediated dissociation of the clumped trypanosomes in the equivalence area released a large number of previously bound surface antigens. These antigens were capable of binding again to fresh IgM antibody. Experimental results further indicated that the complement system caused a functional alteration, changing the multivalent nature of the IgM antibody in the immune complex into a univalent one. This phenomenon is of great advantage to the infected host in clearing pathogens in vivo, as it allows more antibodies to attach to trypanosomes and subsequently initiate complement activity.     

Activation of complement by trypanosomes

Summary Factors exhibiting anti-complementary activity released from trypanosomes after incubation at 20°C were described. The active material was shown to consume the first component of bovine complement. While the anticomplementary factor(s) from T. lewisi could activate bovine, human and guinea pig complement, the factor(s) from T. congolense was observed to activate bovine complement, but not guinea pig and only slightly human complement. The roles of complement activating factor(s) of trypanosomes in the pathology of the disease are discussed.This project is supported by National Research Council of Canada grant A 0068 and a grant from the International Development Research Centre.     

Development of in vitro toxicity tests with cultures of freshly isolated rat hepatocytes

Freshly isolated and cultured hepatocytes were analyzed by two-parameter flow cytometry. The combined analysis of DNA and cellular protein content allowed the contribution of ploidy classes and of subpopulations within a ploidy class to be defined. Analysis of hepatocytes during exposure to dimethylsulfoxide (DMSO), phenobarbital (PB), low oxygen tension (4% O2) or fetal calf serum (FCS), provided insight into the dynamic response of individual ploidy classes as a function of culture time. By analogy with the age-dependent ploidy shifts in vivo, hepatocyte-cultures shift towards adult animals during exposure to DMSO and towards young animals when cultured at low pO2 (4% O2). FCS and phenobarbital disturb this constitutive ploidy balance. FCS increased the 2 N cell population, where stem cells probably respond to the proliferative stimuli provided by growth factors in the serum. Phenobarbital affects the liver-specific 4 N hepatocytes, which agrees with effects seen in liver after exposure in vivo. It is suggested that drug-induced pathological alterations in ploidy in hepatocyte cultures could serve as indicators of compounds, such as liver tumor promoters, which interfere with cell differentiation in liver. The heterotypic cell-cell interaction of freshly isolated hepatocytes with isolated, in vitro cultured, rat liver epithelial cells in co-cultures proved to be a valuable concept in toxicity testing: aldrin epoxidase, an enzyme system involved in xenobiotic metabolism, was stabilized for more than two weeks. After exposure to the three chemicals, 2-acetylaminofluoren, procarbazine and cyproterone-acetate, a preferential toxicity for each compound and cell population was established. Thus heterotypic cell cultures can considerably increase the amount of information available from in vitro studies. The final concept, combining monitoring of cellular DNA (ploidy) and protein content in hepatocyte cultures during and after exposure to a given test compound at tissue oxygen tension with the heterotypic cell-cell interaction, would create a more in vivo-like culture system. This would enhance the predictability of hepatocyte cultures and contribute to a more widespread use of the test system and as a result help to reduce the number of whole-animal tests.     

Development of in vitro toxicity tests with cultures of freshly isolated rat hepatocytes

Summary Freshly isolated and cultured hepatocytes were analyzed by two-parameter flow cytometry. The combined analysis of DNA and cellular protein content allowed the contribution of ploidy classes and of subpopulations within a ploidy class to be defined. Analysis of hepatocytes during exposure to dimethylsulfoxide (DMSO), phenobarbital (PB), low oxygen tension (5% O₂) or fetal calf serum (FCS), provided insight into the dynamic response of individual ploidy classes as a function of culture time. By analogy with the age-dependent ploidy shifts in vivo, hepatocyte-cultures shift towards adult animals during exposure to DMSO and towards young animals when cultured at low pO₂ (4% O₂). FCS and phenobarbital disturb this constitutive ploidy balance. FCS increased the 2 N cell population, where stem cells probably respond to the proliferative stimuli provided by growth factors in the serum. Phenobarbital affects the liver-specific 4 N hepatocytes, which agrees with effects seen in liver after exposure in vivo. It is suggested that drug-induced pathological alterations in ploidy in hepatocyte cultures could serve as indicators of compounds, such as liver tumor promoters, which interfere with cell differentiation in liver. The heterotypic cell-cell interaction of freshly isolated hepatocytes with isolated, in vitro cultured, rat liver epithelial cells in co-cultures proved to be a valuable concept in toxicity testing: aldrin epoxidase, an enzyme system involved in xenobiotic metabolism, was stabilized for more than two weeks. After exposure to the three chemicals, 2-acetylaminofluoren, procarbazine and cyproterone-acetate, a preferential toxicity for each compound and cell population was established. Thus heterotypic cell cultures can considerably increase the amount of information available from in vitro studies.The final concept, combining monitoring of cellular DNA (ploidy) and protein content in hepatocyte cultures during and after exposure to a given test compound at tissue oxygen tension with the heterotypic cell-cell interaction, would create a more in vivo-like culture system. This would enhance the predictability of hepatocyte cultures and contribute to a more widespread use of the test system and as a result help to reduce the number of whole-animal tests.     

Etude de l'activité anticomplémentaire du sérum décomplémenté de rats hypo-, normo- et hyperglycémiques

Summary The anticomplementary activity of decomplemented rat serum was determined using a technique of inhibition of guinea-pig complement in a 50% hemolysis system. The anticomplementary activity of the serum varies inversely to the glycemic state of the rats, being highest in the serum from hypo, and lowest in that of hyperglycemic animals.     

Superoxide dismutase activity in the Spanish population

Superoxide dismutase is an enzyme that catalyzes the dismutation of superoxide radicals to hydrogen peroxide and molecular oxygen. This superoxide radical is produced by all aerobic cells as a normal metabolic intermediate of molecular oxygen, and is dangerous for the cell because it induces the inactivation of various enzymes, lipid peroxidation and mutations. Superoxide dismutase can therefore be considered as a protective enzyme. The purpose of this work was to determine the level of superoxide dismutase activity in the Spanish population, and to study the factors that influence this activity. The superoxide dismutase activity of 2397 individuals was determined using the method described by Minami and Yoshikawa. The superoxide dismutase activity level in the adult Spanish population was found to be 4.16 +/- 0.89 Units/ml of blood. No significant variations with respect to sex were detected. But it was observed that the superoxide dismutase activity level was 9% higher in the young urban Spanish population.