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Aminated beta-1,3-D-polyglucose activates salmon pronephros macrophages in vitro.

Salmon pronephros macrophages were stimulated in vitro using a novel immunomodulator, aminated beta-1,3-D-polyglucose. Activation of these cells was judged by the measurement of superoxide anion formation, pinocytotic activity, acid phosphatase activity and morphological criteria. Stimulated macrophages showed increased superoxide anion formation compared with unstimulated control cells as measured by reduction of Nitroblue tetrazoleum, and an increased level of acid phosphatase activity. Furthermore, stimulated macrophages showed increased pinocytotic activity and a significant increase in cell diameter and spreading.

Acid Phosphatase↗

cDNA cloning and its pronephros-specific expression of the Wilms' tumor suppressor gene, WT1, from Xenopus laevis.

The Wilms' tumor suppressor gene, WT1, plays a crucial role during early urogenital development in mammals. To elucidate the function of WT1 in other vertebrates, we isolated the Xenopus WT1 homolog (XeWT1) from a testis cDNA library. Comparison of the XeWT1 protein with other WT1 proteins revealed that the zinc finger domain of XeWT1 is identical to that of the human WT1 except for the first two amino acid residues. An alternative splice II site, located between the third and the fourth zinc finger is also conserved. In the transcriptional regulatory region, however, three domains, a glycine stretch, a proline stretch and one alternative splice site which are present in the mammalian WT1, are not conserved in amphibians. The XeWT1 gene is expressed in testis and kidney, and whole mount in situ hybridization revealed that the onset of the WT1 gene expression coincides with the early stages of pronephros development in Xenopus. These findings implicate the involvement of WT1 protein during urogenital development in amphibians as well as in mammals.

Amino Acid Sequence↗

Phagocytic response of macrophages from the pronephros of American plaice (Hipoglossoides platessoides) exposed to contaminated sediments from Baie des Anglais, Quebec.

Sediments of Baie des Anglais on the St. Lawrence estuary have a history of environmental contamination, but little information exists regarding their toxicity. The purpose of the present study was to determine the effects of contaminated Baie des Anglais sediments on American plaice (Hippoglossoides platessoides) immune function. Three sites in Baie des Anglais were selected which vary in proximity to local industries and in their sediment contaminant load. Sites 1 and 2 (within the bay) are the closest to shore and most heavily contaminated while sediments at Site 3, which is outside the bay, are the least contaminated. In the first experiment, American plaice were placed in cages at each site for three weeks and immune function was assessed by measuring the phagocytic activity of pronephric macrophages. At the time of sampling, plaice displayed pronephros cell immune response disturbances indicating that Site 1 and 2 were most toxic and Site 3 the least toxic. The results obtained for phagocytosis revealed that contaminants present in the sediments are bioavailable to fish, which came in contact with them and significantly affected their immune system. In the second experiment, sediments from the most toxic site, Site 1, were collected for a laboratory controlled experiment in which plaice were exposed for up to 3 months to these contaminated marine sediments, while the control group was exposed to relatively uncontaminated beach sand. At the end of the exposure period, plaice were transferred from contaminated sediment to beach sand and sampled one month later in order to determine if immune function had returned to control levels. The total number of macrophages decreased following three months of exposure, while the active macrophages had already decreased after the first month of exposure. Following the rehabilitation period a significant trend toward normal response was noted. Sediments from Baie des Anglais contain primarily less highly chlorinated PCBs and lower concentrations of the intermediate and highly chlorinated PCBs. The total concentration of PCBs (sum of 20 congeners) in the contaminated sediments was 1500 ng/g while in the beach sand, the levels were 13.6 ng/g dry weight. Only the low chlorinated PCB congeners were efficiently transferred from the sediments to the plaice liver. Together, these results suggest that the effect of chemical exposure on the phagocytosis of plaice macrophages may be reversible if the fish are returned to a non-contaminated habitat.

Adaptation, Physiological↗

Xenopus crescent encoding a Frizzled-like domain is expressed in the Spemann organizer and pronephros.

The Spemann organizer can be subdivided into head- and trunk-inducing tissues along the anteroposterior axis (Mangold, 1933. Naturwiisenschaften 43, 761-766; Spemann, 1931. Wilhelm Roux Arch. Entwicklungsmech. Org. 123, 389-517). Recent studies have suggested that head formation is brought about by repression of both Wnt and BMP signalling (Glinka et al., 1998. Nature 391, 357-362; Glinka et al., 1997. Nature 389, 517-519). Several Wnt inhibitors secreted from the head organizer region have been identified in Xenopus, such as Cerberus (Bouwmeester et al., 1996. Nature 382, 595-601), Frzb-1 (Leyns et al., 1997. Cell 88, 747-756; Lin et al., 1997. Proc. Natl. Acad. Sci. USA 94, 11196-11200), and Dkk-1 (Glinka et al., 1998. Nature 391, 357-362), supporting this two-inhibitor model. To isolate genes expressed in the head organizer, we screened a prechordal plate cDNA library by sequencing and expression pattern, and isolated the Xenopus ortholog of chick crescent encoding a Frizzled-like domain that is related to Wnt-binding regions of the Frizzled-family proteins. Expression of Xenopus crescent was first detected in the Spemann organizer region at the early gastrula stage and later in prechordal plate cells lining the boundary of mesoderm and ectoderm layers and in the anterior endoderm. At tailbud stages, the expression in the endomesoderm region was diminished, while expression in the pronephros became detectable. In animal cap assays, crescent gene was synergistically upregulated by coexpression of Xlim1, Ldb1, and Siamois, but not by Activin treatment.

Amino Acid Sequence↗

Effects of creosote exposure on rainbow trout pronephros phagocyte activity and the percentage of lymphoid B cells.

An outdoor microcosm study was conducted in order to evaluate the kinetics of immunomodulation in rainbow trout (Oncorhynchus mykiss) exposed to liquid creosote. Fish were sampled on d 7, 14, 21, and 28 from microcosms dosed initially with 0, 3, and 10 microl/L of creosote. Pronephros leukocytes were monitored for phagocytic activity, oxidative burst, and surface immunoglobulin-positive (Slg+) B-cell counts. Oxidative burst was inhibited by creosote exposure; however, by sampling d 28, the burst response returned to near control levels. Phagocytic activity, on the other hand, was stimulated, peaking on sampling d 7, then returning to near control levels by d 28. Although control Slg+ B-cell counts were quite variable across sampling days, Slg+ B-cell counts were also elevated in creosote-exposed fish after seven days of exposure. Slg+ B-cell numbers decreased significantly to near control levels during the remainder of the study. The overall results from this study confirm that creosote has the potential to alter certain immune parameters, and emphasize the importance of monitoring changes in the immune system during exposure. Polycyclic aromatic hydrocarbons (PAHs), a major constituent of liquid creosote, are the suspected immune-altering agents.

Animals↗

CHARACTERIZATION OF MACROPHAGES AND NEUTROPHILIC GRANULOCYTES FROM THE PRONEPHROS OF CARP (CYPRINUS CARPIO)

To analyse the functional activity of different leucocyte types, carp pronephros cells were separated on Percoll density gradients and by use of fluorescence-activated cell sorting. Cell populations were characterised by light and electron microscopy and by flow cytometry. Fractions enriched in macrophages and neutrophilic granulocytes were subsequently analysed for phagocytic activity in vitro by quantification of the uptake of Escherichia coli bacteria or yeast cells, and for respiratory burst response by measurement of the production of the reactive oxygen intermediates O2· and H2O2. Both cell types showed very active in vitro phagocytosis and production of both O2· and H2O2. When activated with phorbol myristate acetate or bacteria, it was evident that the neutrophilic granulocytes were significantly more active than the macrophages. Analysis of single-cell respiratory burst activity in fish phagocytes was investigated after preloading of cells with dihydrorhodamine123. Cells were subsequently separated and analysed for fluorescence using flow cytometry. Both the macrophage-enriched fraction and the granulocyte-enriched fraction appeared to consist of active and inactive subpopulations. In comparison with the inactive populations, active populations had characteristic high forward/sideward scatter profiles.

Journal Article↗

Development of the chick embryo mesoblast: pronephros, lateral plate, and early vasculature.

The early development of the mesoblast in the intermediate and lateral regions of the chick embryo was examined with the scanning and transmission electron microscope. It was found that primary mesenchyme here becomes condenses into epithelial structures that emerge in a metameric pattern. Viewed in developmental sequence, the intermediate mesoblast condenses into a narrowing cord of axially oriented cells which divert medially at regular intervals into the intersegmental interfaces of somitomeres and somites. These cells give rise to the vascular channels of the posterior cardinal vein as well as to tubular elements of the pronephros. Intermediate mesenchyme cells become epithelial, forming zonular junctional complexes apically and depositing patchy basal lamina over their basal surfaces. The lateral plate mesenchyme organizes similarly into somatic and splanchnic epithelial sheets that utilize the body coelom as their lumenal surface. Cells of the lateral plate extend filopodia basally that interweave with adjacent cells, fibrillar extracellular matrix, as well as with interstitial bodies. The pattern in the lateral plate is subtly ribbed as bands of mesoblast undulate along the axis. The central region of each band is raised while ther are grooves created along lines of band abutment, corresponding to intersegmental clefts in the paraxial region and reflecting an underlying metameric pattern. These grooves are usually demarked medially by the protrusion of short segments of adjacent intermediate mesoblast. Most of the remaining primary mesenchyme develops into a non-metameric vascular epithelium, which forms a prominent anastamosing plexus between splanchnic mesoderm and endoderm. It is proposed that the emergence of primary mesenchyme into patterened epithelial anlage facilitates the distribution of neural crest cells introduced subsequently.

Animals↗