Cellular and humoral immune responses to Plasmodium falciparum gametocyte antigens in malaria-immune individuals. Limited response to the 48/45-kilodalton surface antigen does not appear to be due to MHC restriction

We have examined immune responses to a cultured Plasmodium falciparum gametocyte lysate and to an affinity-purified preparation of the 48/45-kDa gamete surface Ag in a group of 30 malaria immune individuals and in 24 Europeans with no previous exposure to malaria. Cellular responses were assessed in vitro by lymphoproliferation and production of IFN-gamma; antigamete antibodies were detected by immunofluorescence, Western blotting, and competitive ELISA. Cells from all the malaria immune donors responded to the gametocyte lysate in both assays while cells from nonimmune donors gave only weak proliferative responses. Antigamete antibodies were detected in the serum of all the immune donors but not in serum from nonimmunes. Nonimmune donors were completely unresponsive to the purified 48/45-kDa surface Ag while cells from 40% of immune donors responded by either proliferation or IFN-gamma production. Only 3 of 30 immune donors had detectable antibodies to the 48/45-kDa Ag. Class II HLA type was determined for 27 of the immune donors but no relationship between HLA-DR or -DQ and responsiveness to the 48/45-kDa Ag was discerned. The possible reasons for limited recognition of this gamete surface Ag are discussed.     

Induction of morphological transformation by coal-dust extract in BALB/3T3 A31-1-13 cell line

The transforming activity of coal dust extracts was studied using BALB/3T3 clone A31-1-13 cells. Coal-dust extracts, both nitrosated and nonnitrosated, induced cell transformation in a dose-response manner. However, the transformation frequency was higher with the nitrosated than with the nonnitrosated extract. All transformed cell lines derived from coal-dust extract-induced foci showed biological characteristics of neoplastic transformation such as loss of contact inhibition and anchorage-independent growth. These results appear to support a hypothesis of coal mine dust causation of gastric cancer in coal miners.     

Positive and Negative Regulation of the Differentiation of Ventral Mesoderm for Erythrocytes in Xenopus laevis

To elucidate the mechanism of determination and regulation of hemopoiesis in the early Xenopus embryo, explants of dorsal and ventral mesoderm from various stage embryos were cultured alone or combined with various tissues derived from the same stage embryo. Western blot analysis of larvae-specific globin expression using monoclonal antibody L5.41 revealed that extensive erythropoiesis occurred in the explants of ventral mesoderm from st. 22 tailbud embryo, but not in those of dorsal mesoderm. Experiments using combined explants at this stage demonstrated that the in vitro differentiation of erythrocytes in the ventral mesoderm could be completely inhibited by the dorsal tissue, including neural tube, notochord, and somite mesoderm, but not by other mesoderms, gut endoderm, or forebrain. Subsequent explant studies showed that the notochord alone is sufficient for this inhibition. Furthermore, the ventral mesoderm explant from the st. 10⁺ early gastrula embryo was not able to differentiate into erythroid cells. However, small amounts of globin were expressed if ventral mesoderm of this stage was combined with animal pole cells which were mainly differentiated to epidermis. This stimulation was enhanced when both tissues were excised together without separation, while none of the other parts of st. 10⁺ embryo had this stimulatory effect. These observations found in the combined explants suggest that in vivo interactions between the ventral mesoderm and adjacent tissues are important for normal development of erythroid precursor cells.     

Localisation of Formyl-Peptide Receptor 2 in the Rat Central Nervous System and Its Role in Axonal and Dendritic Outgrowth

Arachidonic acid and docosahexaenoic acid (DHA) released by the action of phospholipases A₂ (PLA₂) on membrane phospholipids may be metabolized by lipoxygenases to the anti-inflammatory mediators lipoxin A4 (LXA4) and resolvin D1 (RvD1), and these can bind to a common receptor, formyl-peptide receptor 2 (FPR2). The contribution of this receptor to axonal or dendritic outgrowth is unknown. The present study was carried out to elucidate the distribution of FPR2 in the rat CNS and its role in outgrowth of neuronal processes. FPR2 mRNA expression was greatest in the brainstem, followed by the spinal cord, thalamus/hypothalamus, cerebral neocortex, hippocampus, cerebellum and striatum. The brainstem and spinal cord also contained high levels of FPR2 protein. The cerebral neocortex was moderately immunolabelled for FPR2, with staining mostly present as puncta in the neuropil. Dentate granule neurons and their axons (mossy fibres) in the hippocampus were very densely labelled. The cerebellar cortex was lightly stained, but the deep cerebellar nuclei, inferior olivary nucleus, vestibular nuclei, spinal trigeminal nucleus and dorsal horn of the spinal cord were densely labelled. Electron microscopy of the prefrontal cortex showed FPR2 immunolabel mostly in immature axon terminals or ‘pre-terminals’, that did not form synapses with dendrites. Treatment of primary hippocampal neurons with the FPR2 inhibitors, PBP10 or WRW4, resulted in reduced lengths of axons and dendrites. The CNS distribution of FPR2 suggests important functions in learning and memory, balance and nociception. This might be due to an effect of FPR2 in mediating arachidonic acid/LXA4 or DHA/RvD1-induced axonal or dendritic outgrowth.     

Metabolic signatures of four major histological types of lung cancer cells

Introduction

Histologically lung cancer is classified into four major types: adenocarcinoma (Ad), squamous cell carcinoma (SqCC), large cell carcinoma (LCC), and small cell lung cancer (SCLC). Presently, our understanding of cellular metabolism among them is still not clear.

Objectives

The goal of this study was to assess the cellular metabolic profiles across these four types of lung cancer using an untargeted metabolomics approach.

Methods

Six lung cancer cell lines, viz., Ad (A549 and HCC827), SqCC (NCl-H226 and NCl-H520), LCC (NCl-H460), and SCLC (NCl-H526), were analyzed using liquid chromatography quadrupole time-of-flight mass spectrometry, with normal human small airway epithelial cells (SAEC) as the control group. The principal component analysis (PCA) was performed to identify the metabolic signatures that had characteristic alterations in each histological type. Further, a metabolite set enrichment analysis was performed for pathway analysis.

Results

Compared to the SAEC, 31, 27, 34, 34, 32, and 39 differential metabolites mainly in relation to nucleotides, amino acid, and fatty acid metabolism were identified in A549, HCC827, NCl-H226, NCl-H520, NCl-H460, and NCl-H526 cells, respectively. The metabolic signatures allowed the six cancerous cell lines to be clearly separated in a PCA score plot.

Conclusion

The metabolic signatures are unique to each histological type, and appeared to be related to their cell-of-origin and mutation status. The changes are useful for assessing the metabolic characteristics of lung cancer, and offer potential for the establishment of novel diagnostic tools for different origin and oncogenic mutation of lung cancer.

     

Transcriptome and toxin family analysis of the paralysis tick, Ixodes holocyclus

The Australian paralysis tick (Ixodes holocyclus) secretes neuropathic toxins into saliva that induce host paralysis. Salivary glands and viscera were dissected from fully engorged female I. holocyclus ticks collected from dogs and cats with paralysis symptoms. cDNA from both tissue samples were sequenced using Illumina HiSeq 100?bp pair end read technologies. Unique and non-redundant holocyclotoxin sequences were designated as HT2–HT19, as none were identical to the previously described HT1. Specific binding to rat synaptosomes was determined for synthetic HTs, and their neurotoxic capacity was determined by neonatal mouse assay. They induced a powerful paralysis in neonatal mice, particularly HT4 which produced rapid and strong respiratory distress in all animals tested. This is the first known genomic database developed for the Australian paralysis tick. The database contributed to the identification and subsequent characterization of the holocyclotoxin family that will inform the development of novel anti-paralysis control methods.     

Immune islet killing mechanisms associated with insulin-dependent diabetes: in vitro expression of cellular and antibody-mediated islet cell cytotoxicity in humans

In previous work we have shown that some bacteria can bind to human lymphocytes and can be used to identify lymphocyte subpopulations in conventionally stained blood smears. These bacteria are of different species or genera, which makes it difficult to study the binding mechanism. Also, the main marker for B cells, Brucella melitensis, is of very small size and highly pathogenic. Here we show that B cells as well as some of the T cell subpopulations can be identified by different mutants obtained from a strain of an Escherichia coli. Two procedures were used to generate mutants. First, E. coli-YS57 (pro-his-trp-) was mutagenized with N-methyl-N'-nitro-N-nitrosoguanidine and the binding to mouse spleen cells was used as a selective pressure. Second, phage-resistant mutants of E. coli-YS57 were obtained and tested for the ability to bind to lymphocytes. Out of 10 strains selected by the former procedure, 5 bound to a significant number of human lymphocytes. All four phage-resistant mutants bound to human lymphocytes. Out of the total of nine mutants that bound to lymphocytes, six bound consistently, i.e., they bound to similar percentages of peripheral blood lymphocytes from different normal donors. One phage-resistant mutant, E. coli USC-106, bound only to B cells. The subpopulations of lymphocytes identified by the mutants were essentially the same as those identified by different species or genera of bacteria. We concluded that E. coli mutants can be obtained that identify human lymphocyte subpopulations and that one of these mutants recognizes B cells; these mutants may be used to study the nature of the receptors for bacteria on lymphocytes, which appear to have a lectin-like nature.     

Mutagenicity studies of ambient airborne particles. II. Comparison of extraction methods

Organic materials were extracted with acetone from airborne particles by shaking, soxhletion and sonication for varying durations. 4-h, 1-h and 1/8-min extractions by shaking, soxhletion and sonication, respectively gave maximum his+ revertants with the Ames Salmonella/microsome assay. In a comparative study of extraction methods, sonication gave the highest and soxhletion the lowest mutagenic response. It appears that sonication with acetone is the best procedure for the extraction of mutagens from airborne particles as shown by Ames assay and Arar assay systems in Salmonella typhimurium.     

Fluorescence demonstration of a cathepsin H-like protease in cardiac, skeletal and vascular smooth muscles

Summary A cathepsin H-like enzyme was localized by histochemical techniques in cardiac muscle, extensor digitorum longus and soleus skeletal muscles, and vascular smooth muscle. Using the specific exopeptidase activity of this enzyme against the substrate arg-4-methoxy--naphthylamide, valid histochemical assay conditions were developed. More fluorescent granules were observed in cardiac muscle than in the soleus and about equal amounts in vascular smooth muscle and the extensor digitorum longus. The reaction rate was enhanced by chloride ions and inhibited by 1mm p-chloromercuribenzoate. The maximal activity was observed between pH 5.5 and 6.0. Chemical fixation with periodate-lysine-paraformaldehyde preserved a small amount of enzymatic activity.     

Spontaneous sexual differentiation of Mesocestoides corti tetrathyridia in vitro

Barrett N. J., Smyth J. D. and Ong S. J. 1982. Spontaneous sexual differentiation of Mesocestoides corti tetrathyridia in vitro. International Journal for Parasitology12: 315–322. Tetrathyridia of Mesocestoides corti, from the body cavity of mice, maintained in the laboratory by intraperitoneal infection, were used for in vitro culture. In an initial experiment, after 50 days asexual multiplication in vitro one tetrathyridium spontaneously segmented and developed into a sexually mature adult. Further experiments were carried out in an attempt to determine the conditions favouring segmentation and sexual differentiation. A combination of 5 or 10 ml liquid medium S1OE.H (basically composed of CMRL 1066 and foetal calf serum with supplements) changed every 3 days, in a Leighton tube (19 × 105 mm), rotated at 38°C and gassed with 10 or 20% CO₂, containing between 100 and 200 tetrathyridia, has proved to be most suitable so far. Numerous adult worms with normal male and female genitalia have been obtained in this system. However, segmentation is sporadic, rather than consistent and only a few shelled eggs with hooked oncospheres have so far been obtained, suggesting that impregnation and fertilization in vitro is not fully comparable with that in vivo.