Experimental pulmonary candidiasis

The initial interaction of Candida albicans with pulmonary tissue of B6D2/F₁ mice was investigated. The LD₅₀ for mice challenged intravenously (IV) was approximately 3 × 10⁵ yeasts, whereas the LD₅₀ by the intratracheal (IT) route was in excess of 10⁸ yeasts. Mice challenged IV died of progressive yeast growth in the kidneys. In contrast, mice challenged IT rapidly eliminated the entire inoculum by the first day after challenge. Resident pulmonary alveolar macrophages (PAM) killed upwards of 70% of C. albicans in an in vitro killing assay. At effector: target ratios favoring the effector cell population resident PAM were able to restrict the formation of yeast germ tubes to only 30% of the yeasts, whereas at equivalent ratios virtually all of the intracellular yeasts produced germ tubes. Evaluation of the ability of PAM, harvested from genetically different strains of inbred mice, to kill C. albicans in vitro showed that killing ability was a property of resident PAM from mice with the black 6 background. It was discovered that during the initial stages of infection in vivo, the expression of the F4/80 surface molecule was down regulated, and the expression of the Mac 1 surface molecule upregulated. There were no quantitative changes in expression of either Mac 2, Mac 3, Ly 5 or the 5C6 surface epitopes. Taken together, the data show that pulmonary tissue is quantitatively very resistant to C. albicans infection, because of the ability of resident PAM to rapidly phagocytize and kill yeasts. Killing of C. albicans by resident PAM may be a property of a subset of this mononuclear phagocyte population and was accompanied by alterations in the expression of surface molecules.Presented as part of the Everett S. Beneke Symposium in Mycology, May 27, 1988.     

The cytotoxicity of chrysotile asbestos fibers to pulmonary alveolar macrophages I. Effects of inhibitors of ADP-ribosyl transferase

Pulmonary alveoler macrophages exposedto very short chrysotile asbestos fibers present a typical cytotoxic response: extracellular releases of lactate dehydrogenase and -galactosidase, and a decrease in cellular ATP content. The objective of this study was to determine if nicotinamide and 3-aminobenzamide, two inhibitors of the ADP-ribosyl transferase, could modify the in vitro toxicity of chrysotilee fibers. After 30 min of pre-exposure with each of the two inihibitors, pulmonary alveolar macrophage monolayers were concominantly exposed for 18 hours to 50g of fibers. It was observed that, in a dose-effect relationship (5 to 30 mM), nicotinamide was very effective in reducing the extracellular liberation of the marker enzymes. At 30 mM, the enzyme releases in the medium had returned to control values; the restoration of cell viability was confirmed by ATP levels. Up to 5 mM 3-aminobenzamide did not provide any protection against chrysotile cytotoxicity. Nicotinic acid, a structural analogue of nicotinamide, but not an inhibitor of the ADP-ribosyl transferase, also showed no protective effect. Nicotinamide and 3-aminobenzamide increased the intracellular NAD⁺ pools, respectively by 350% and 250%. However, with or without additives, the chrysotile fibers caused a constant and significant decrease in NAD⁺ levels (40–55 pmoles). These results suggest that the inhibition of the nuclear ADP-ribosyl transferase is not the major mechanism by which nicotinamide protects pulmonary alveolar macrophages against the chrysotile asbestos fibers.Abbreviations 3-AB 3-aminobenzamide - ADPRT ADP-ribosyl transferase - -GAL -galactosidase - DTT dithiothreitol - FBS fetal bovine serum - FMN flavin mononucleotide - HEPES N-2-hydroxyethyl piperazine-N-2-ethanesulfonic acid - LDH lactate dehydrogenase - NAD⁺ nicotinamide adenine dinucleotide (oxidized form) - NADH nicotimide adenine dinucleotide (reduced forms) - NADPH nicotimide adenine dinucleotide phosphate (reduced form) - NAM nicotinamide - NIC nicotinic acid - ORS oxygen radical species - PAM pulmonary alveolar macrophages - S.E. standard error of the mean - TAPS tris (hydroxymethyl) methylamino-propane sulfonic acid - TRIS tris (hydroxymethyl) aminomethane - VSF very short chrysotile fibers     

Prevention by N-acetylcysteine of benzo[a]pyrene clastogenicity and DNA adducts in rats

The daily i.t. administration of benzo[a]pyrene (BP) to Sprague-Dawley rats, for 3 consecutive days, did not cause any toxicity or clastogenicity in bone marrow cells, as evaluated by monitoring the ratio of polychromatic to normochromatic erythrocytes and the frequency of micronucleated polychromatic erythrocytes. However, BP produced a considerable enhancement of binucleated and micronucleated pulmonary alveolar macrophages, as well as a significant increase in polymorphonucleates recovered by bronchoalveolar lavage. These effects were prevented by administering the thiol N-acetylcysteine (NAC) by gavage 5 h before each BP instillation. In addition, the i.t. treatment with BP resulted in the formation of BP diolepoxide (BPDE)-DNA adducts in lungs and liver, as assessed by synchronous fluorescence spectrophotometry, with fluorescence peaks of similar magnitude in the 2 tissues. Pretreatment with NAC by gavage completely prevented BPDE adducts to liver DNA and significantly decreased those to lung DNA.     

In vitro culture of the strobilar stage of Echinococcus granulosus from protoscoleces of human, camel, cattle, sheep and goat origin from Kenya and buffalo origin from India

Protoscoleces from human, camel, cattle, sheep, goat (all from Kenya) and buffalo (from India) hydatid cysts were cultured under identical conditions in vitro using the diphasic culture system of Smyth (1979b). Organisms from all sources grew and segmented in culture. Genital anlagen developed in all cultured worms but further genital differentiation occurred only in cultures of cattle (testes) and camel (testes and genital pore) material. The possible significance of these results is discussed in relation to the general epidemiology of hydatid disease and the potential infectivity of the different strains to man.     

IL-33-ST2 pathway regulates AECII transdifferentiation by targeting alveolar macrophage in a bronchopulmonary dysplasia mouse model

Evidence points to the indispensable function of alveolar macrophages (AMs) in normal lung development and tissue homeostasis. However, the importance of AMs in bronchopulmonary dysplasia (BPD) has not been elucidated. Here, we identified a significant role of abnormal AM proliferation and polarization in alveolar dysplasia during BPD, which is closely related to the activation of the IL-33-ST2 pathway. Compared with the control BPD group, AMs depletion partially abolished the epithelialmesenchymal transition process of AECII and alleviated pulmonary differentiation arrest. In addition, IL-33 or ST2 knockdown has protective effects against lung injury after hyperoxia, which is associated with reduced AM polarization and proliferation. The protective effect disappeared following reconstitution of AMs in injured IL-33 knockdown mice, and the differentiation of lung epithelium was blocked again. In conclusion, the IL-33-ST2 pathway regulates AECII transdifferentiation by targeting AMs proliferation and polarization in BPD, which shows a novel strategy for manipulating the IL-33–ST2-AMs axis for the diagnosis and intervention of BPD.     

Detection of simian immunodeficiency virus DNA in macrophages from infected rhesus macaques.

We have examined the frequency of infection of monocyte-derived and alveolar macrophages isolated from rhesus macaques inoculated with simian immunodeficiency virus (SIVmac) utilizing a semiquantitative PCR methodology. Animals were inoculated with either pathogenic (SIVmac239) or nonpathogenic (SIVmac1A11) molecularly cloned viruses of SIVmac, or with uncloned pathogenic SIVmacBIOL. The frequency of SIV DNA in macrophages was highest early after infection and at terminal stages of disease, whereas during the asymptomatic period, SIV DNA was present at very low levels in macrophages.     

Phosphoprotein intermediate in the ca2+-dependent atpase reaction of macrophage plasma membrane

ATPase activity and phosphorylation by [γ-³²P] ATP of isolated plasma membrane of alveolar macorphages are stimulated in a parallel fashion by physiologic concentrations of Ca²⁺, with half-maximal activating effect of this ion at (3–7) × 10^?7 M. For various membrane preparations, a direct proportionality exists between Ca²⁺-dependent ATPase activity and amount of ³²P incorporated. Labeling of membrane attains the steady-state level by 10 sec at 0°C, and is rapidly reversed by adenosine diphosphate (ADP). K⁺ decreases the amount of membrane-bound ³²P, mainly by enhancing the rate of dephosphorylation of the ³²P-intermediate. Hydroxylamine causes a release of about 90% of ³²P bound to the membrane, thus indicating that the ³²P-intermediate contains an acyl-phosphate bond. When the labeled plasma membrane is solubilized and electrophoresed on acrylamide gels in the presence of sodium dodecyl sulphate, the radioactivity appears to be largely associated with a single protein fraction of 132,500 ± 2,000 apparent molecular weight. These features of the macrophage Ca²⁺-ATPase suggest that the enzyme activity might be part of a surface-localized Ca²⁺-extrusion system, participating in the regulation of Ca²⁺-dependent activities of the macrophage.     

NLRP3 regulates alveolar bone loss in ligature‐induced periodontitis by promoting osteoclastic differentiation

ObjectivesNLRP3 inflammasome is a critical part of the innate immune system and plays an important role in a variety of inflammatory diseases. However, the effects of NLRP3 inflammasome on periodontitis have not been fully studied.Materials and methodsWe used ligature‐induced periodontitis models of NLRP3 knockout mice (NLRP3^KO) and their wildtype (WT) littermates to compare their alveolar bone phenotypes. We further used Lysm‐Cre/Rosa^nTnG mouse to trace the changes of Lysm‐Cre⁺ osteoclast precursors in ligature‐induced periodontitis with or without MCC950 treatment. At last, we explored MCC950 as a potential drug for the treatment of periodontitis in vivo and in vitro.ResultsHere, we showed that the number of osteoclast precursors, osteoclast differentiation and alveolar bone loss were reduced in NLRP3^KO mice compared with WT littermates, by using ligature‐induced periodontitis model. Next, MCC950, a specific inhibitor of the NLRP3 inflammasome, was used to inhibit osteoclast precursors differentiation into osteoclast. Further, we used Lysm‐Cre/Rosa^nTnG mice to demonstrate that MCC950 decreases the number of Lysm‐Cre⁺ osteoclast precursors in ligature‐induced periodontitis. At last, treatment with MCC950 significantly suppressed alveolar bone loss with reduced IL‐1β activation and osteoclast differentiation in ligature‐induced periodontitis.ConclusionOur findings reveal that NLRP3 regulates alveolar bone loss in ligature‐induced periodontitis by promoting osteoclastic differentiation.