Cranial neural crest cells contribute to connective tissue in cranial muscles in the anuran amphibian, Bombina orientalis.

The contribution of cranial neural crest cells to the development and patterning of cranial muscles in amphibians was investigated in the phylogenetically basal and morphologically generalized frog, Bombina orientalis. Experimental methods included fluorescent marking of premigratory cranial neural crest and extirpation of individual migratory streams. Neural crest cells contributed to the connective tissue component, but not the myofibers, of many larval muscles within the first two branchial arches (mandibular and hyoid), and complex changes in muscle patterning followed neural crest extirpation. Connective tissue components of individual muscles of either arch originate from the particular crest migratory stream that is associated with that arch, and this relationship is maintained regardless of the segmental identity-or embryonic derivation-of associated skeletal components. These developmental relations define a pattern of segmentation in the head of larval anurans that is similar to that previously described in the domestic chicken, the only vertebrate that has been thoroughly investigated in this respect. The fundamental role of the neural crest in patterning skeleton and musculature may represent a primitive feature of cranial development in vertebrates. Moreover, the corresponding developmental processes and cell fates appear to be conserved even when major evolutionary innovations-such as the novel cartilages and muscles of anuran larvae-result in major differences in cranial form.

Cranial neural crest cell migration in the Australian lungfish, Neoceratodus forsteri.

A crucial role for the cranial neural crest in head development has been established for both actinopterygian fishes and tetrapods. It has been claimed, however, that the neural crest is unimportant for head development in the Australian lungfish (Neoceratodus forsteri), a member of the group (Dipnoi) which is commonly considered to be the living sister group of the tetrapods. In the present study, we used scanning electron microscopy to study cranial neural crest development in the Australian lungfish. Our results, contrary to those of Kemp, show that cranial neural crest cells do emerge and migrate in the Australian lungfish in the same way as in other vertebrates, forming mandibular, hyoid, and branchial streams. The major difference is in the timing of the onset of cranial neural crest migration. It is delayed in the Australian lungfish in comparison with their living sister group the Lissamphibia. Furthermore, the delay in timing between the emergence of the hyoid and branchial crest streams is very long, indicating a steeper anterior-posterior gradient than in amphibians. We are now extending our work on lungfish head development to include experimental studies (ablation of selected streams of neural crest cells) and fate mapping (using fluoresent tracer dyes such as Dil) to document the normal fate as well as the role in head patterning of the cranial neural crest in the Australian lungfish.

PDGFB regulates the development of the labyrinthine layer of the mouse fetal placenta.

PDGFB is a growth factor which is vital for the completion of normal prenatal development. In this study, we report the phenotypic analysis of placentas from mouse conceptuses that lack a functional PDGFB or PDGFRbeta gene. Placentas of both types of mutant exhibit changes in the labyrinthine layer, including dilated embryonic blood vessels and reduced numbers of both pericytes and trophoblasts. These changes are seen from embryonic day (E) 13.5, which coincides with the upregulation of PDGFB mRNA levels in normal placentas. By E17, modifications in shape, size, and number of the fetal blood vessels in the mutant placentas cause an abnormal ratio of the surface areas between the fetal and the maternal blood vessels in the labyrinthine layer. Our data suggest that PDGFB acts locally to contribute to the development of the labyrinthine layer of the fetal placenta and the formation of a proper nutrient-waste exchange system during fetal development. We point out that the roles of PDGFB/Rbeta signaling in the placenta may be analogous to those in the developing kidney, by controlling pericytes in the labyrinthine layer and mesangial cells in the kidney.

Neurotrophin-7: a novel member of the neurotrophin family from the zebrafish.

A novel member of the neurotrophin family, zebrafish neurotrophin-7 (zNT-7), was isolated from the zebrafish Danio rerio. The amino acid sequence of zNT-7 is more closely related to that of fish nerve growth factor (NGF) and neurotrophin-6 (NT-6) than to that of any other neurotrophin. zNT-7 is, however, equally related to fish NGF and NT-6 (65% and 63% amino acid sequence identity, respectively) indicating that it represents a distinct neurotrophin sequence. zNT-7 contains a 15 amino acid residue insertion in a beta-turn region in the middle of the mature protein. Recombinant zNT-7 was able to bind to the human p75 neurotrophin receptor and to induce tyrosine phosphorylation of the rat TrkA receptor tyrosine kinase, albeit less efficiently than rat NGF. zNT-7 did not interact with rat TrkB or TrkC, indicating a similar receptor specificity as NGF. We propose that a diversification of the NGF subfamily in the neurotrophin evolutionary tree occurred during the evolution of teleost fishes which resulted in the appearance of several additional members, such as zNT-7 and NT-6, structurally and functionally related to NGF.

Characterization of human neutrophils adherent to organic polymers.

The adherence of human neutrophils to surfaces from organic polymers (Pellethane, two types of polyvinyl chloride (PVC), polypropylene and polyethylene) is associated to a different extent with O2- formation. No comparable differences were observed for the liberation of the granular enzyme elastase. Only cells attached to Pellethane responded strongly after a second stimulation by opsonized zymosan, N-formyl-methionyl-leucyl-phenylalanine (FMLP) and aggregated IgG. The expression of related antigens on adherent cells was measured by a cell-ELISA (enzyme-linked immunosorbent assay). Cells adherent to five various untreated organic polymers show no differences of their Fc gamma- and C3bi-receptors. A considerable increase of all investigated cellular antigens was observed for neutrophils adherent to films precoated with autologous plasma. The adherence rates and the viability for cells adherent to different materials were similar and may not account for the observed differences in O2- production during the primary adherence reaction and a further secondary stimulation.

[An immunomonitoring program for supervising transplant patients]. / Ein Immun-Monitoringprogramm zur Uberwachung transplantierter Patienten.

Lymphocytes and monocytes with differential and activation antigens were determined by means of the flow cytometry by using monoclonal antibodies in the peripheral blood of patients after organ transplantation. Examinations of the course showed that in this way the acute activity is seized by the cellular immunosystem. Acute reject crises, systemic virus infections, as well as septic states lead to characteristic changes and these verify the diagnostic assertion of this immunomonitoring programme.

[Analysis of T-cell subpopulations and expression patterns of lymphocytic activation markers in patients in the early phase following kidney transplantation using laser flow cytometry]. / Analyse von T-Zellsubpopulationen und Expressionsmustern lymphozytärer Aktivierungsmarker bei Patienten in der Frühphase nach Nierentransplantation mittels Laser-Durchflusszytometrie.

In 10 patients after kidney transplantation the T cell subpopulations (CD4, CD8) and the number of activated lymphocytes (IL-2R and transferrin receptor+, 4F2+, HLA-DR+, CD3+) in the peripheral blood were determined. In 9 out of 9 cases with rejection a clear immune activation could observed. In patients with CMV-associated rejection crisis the CD4/CD8 ratio was significantly higher compared with patients with rejection crisis but without CMV infection. In all patients with CMV infection (n = 5) an increase of activated lymphocytes was observed. In a clinical-immunological monitoring the dynamics of the CD4/CD8 ratio, the expression of activation marker (especially CD3+, HLA-DR+ and 4F2+) and the absolute number of lymphocyte subpopulations should included.

Effect of cycloxygenase and lipoxygenase inhibitors on the chemiluminescence response of human neutrophils.

The chemiluminescence response of human neutrophils activated by zymosan and WGA is different in the presence of the amplifiers lucigenin and luminol. Inhibitors of arachidonic acid metabolism and scavengers of reactive oxygen species demonstrate that the generation of cellular chemiluminescence is connected with the activation of the lipoxygenase and cycloxygenase pathway. The effect on the chemiluminescence induced by WGA was always stronger than that of chemiluminescence activated by zymosan. Indomethacin, ETYA and two experimental lipoxygenase-inhibitors cause in some concentrations an increase of chemiluminescence response. The estimation of the influence of new substances on the cellular chemiluminescence stimulation and its comparison with standard drugs should be useful to enable a first in vitro characterization.

The influence of interferon-gamma and various phagocytic stimuli on the expression of MHC-class II antigens on human monocytes--relation to the generation of reactive oxygen intermediates.

Human monocytes show a dose-dependent decrease of the MHC-class II antigen expression (HLA-DR and HLA-DQ) after addition of zymosan or Staphylococcus aureus Cowan I particles. Interferon-gamma did not prevent this process. The expression of MHC-class I antigens was not affected. The dose-response and kinetic curves showed individual differences. An association between the capacity to form reactive oxygen intermediates and the downregulation of MHC-class II antigen expression was observed. In addition, after digestion of the phagocytosed particles interferon-gamma could restore the MHC-class II antigen expression on the cultured monocytes. The possible biological significance of these interactions between interferon-gamma and phagocytosis for the function of monocytes/macrophages in the local inflammatory process is discussed.

The influence of phagocytic stimuli on the expression of HLA-DR antigens; role of reactive oxygen intermediates.

Human peripheral blood mononuclear cells show after one day of culture with opsonized zymosan a decreased expression of HLA-DR (but not HLA-A, B, C) antigens, which can be prevented by the scavenger of reactive oxygen intermediates beta-carotene, or superoxide dismutase or indomethacin. Mononuclear cells of a patient with a heterozygous form of chronic granulomatous disease show no alterations in HLA-DR antigen expression after culture with opsonized zymosan. Possible roles of reactive oxygen intermediates or prostaglandins in the modulation of HLA-DR antigens are discussed.

[Determination of superoxide dismutase in immune cells by lucigenin-mediated chemiluminescence]. / Bestimmung der Superoxid-Dismutase-Aktivität in Immunzellen mittels Lucigenin-vermittelter Chemolumineszenz.

DMSO induces the generation of superoxide radicals in solutions of Na2CO3-Na2EDTA. The formation of superoxide can be detected by lucigenin-dependent chemiluminescence. The inhibition of the chemiluminescence reaction by superoxide dismutase can be used for the sensitive measurement of enzymatic activity of this enzyme. Human mononuclear cells compared with neutrophils demonstrate a three times elevated activity of superoxide dismutase.

[Chemiluminescence, formation of thermostable E-rosettes and DNA synthesis of human mononuclear cells induced by various lectins]. / Chemolumineszenz, Bildung thermostabiler E-Rosetten und DNS-Synthese humaner mononukleärer Zellen induziert durch verschiedene Lektine.

Various plant lectins differentiate in their ability to induce early (chemiluminescence and thermostable rosette formation) and later (DNA synthesis) activation events of human mononuclear cells. Both phytohaemagglutinin and concanavalin A induced a cell activation detectable by the three investigated parameters. Wheat germ agglutinin was characterized as an efficient stimulator in both assays of early events whereas the induction of DNA synthesis measured by [3H]-thymidine incorporation was low. In contrast, pokeweek mitogen did evoke only a weak chemiluminescence signal, but induced a small number of thermostable rosettes and a high [3H]-thymidine incorporation. The results obtained from experiments with mononuclear cells of twelve healthy donors did not show a firm correlation between the investigated three parameters of cell activation.

Investigation of the chemiluminescence response of human neutrophils and mononuclear cells.

The activation of chemiluminescence (CL) of human phagocytes was investigated by the use of the amplifiers, luminol and lucigenin, in parallel studies. The optimal doses for opsonized zymosan, WGA and HAIG were determined, and the integral CL of polymorphonuclear and mononuclear cells from healthy probands and patients with enzyme defects related to the cellular O2 metabolism were compared. Both cell types showed striking differences with respect to the kinetics and integral responses and in dependence on the amplifier after stimulation with the various activators. These results suggest that there is no intimate relationship between O2-release and production of the luminol reactive oxygen species.

Histamine and immune reactions. 3. Inhibition of early events of human lymphocyte activation by histamine.

Histamine inhibits the chemiluminescence response of human lymphocytes to Concanavalin A by interfering with early triggering events. The inhibition seems to be mediated via H2 receptors, acts immediately, and is reversible by washing. A longer incubation in a histamine-containing medium induces a refractory state of the cells. The inhibitory effect described here is compared with the inhibitory action of histamine in the lymphocyte transformation test.

Circulating immune complexes after cadaver kidney transplantation.

351 sera from 27 human recipients of renal allografts and 21 healthy blood donors were assayed for circulating immune complexes by the Clq solid-phase radioimmune assay. Increased Clq-binding activity (Clq-BA) was detected in pretransplant sera from 5 patients with chronic pyelonephritis (PN) and 3 patients with chronic glomerulonephritis (GN). A significant decrease of Clq-BA immediately after transplantation could not be found. 6 weeks after transplantation only 2 patients of the PN group showed increased Clq-BA. Serial studies in 17 patients with rejection crises did not show any correlation between the level of serum Clq-BA and the occurrence of rejections. Furthermore, no correlation could be found between the occurrence of complement-dependent lymphocytotoxic antibodies measured by the 51Cr release technique and the level of serum Clq-BA. In contrast, our results show that the probability of graftectomy or graft failure is significantly higher, at least in the early phase after transplantation, when the serum Clq-BA is lowered for several weeks.

[Radioimmunological detection of soluble immune complexes in serum (author's transl)]. / Radioimmunologischer Nachweis von löslichen Immunkomplexen im serum.

Immune complexes could be measured in sera with the use of plastfixed Clq and 125J-labeled anti-Ig-antibody. The sensitivity of the system is 0.1 microgram aggregated human IgG, the working range is between 0.1 and 10 micrograms per 0.5 ml. In 58% of the sera from patients with systemic lupus erythematosus was found an increase in the immune complex content.