Glycoprotein differences among cells of the 14-day embryonic chick neural retina

In order to test the hypothesis that the progressive layering and differentiation of cell types during the development of the neural retina are associated with cell surface alterations we have separated distinct cell populations from the 14-day embryonic chick retina. Cells of these populations have been shown to differ in associative behavior and intramembrane particle content. We now report that these cells differ in cell surface glycoproteins. Proteins were labeled with two different extrinsic labels and one metabolic label. We used enzymatic transfer to galactose from UDP-gal to cellular acceptors, and borotritide reduction after galactose oxidation as extrinsic labels. Glucosamine incorporation was used as the metabolic label. In all these cases, we were able to identify bands on electrophoretic gels which were unique to individual populations.     

Heterogeneity of neural progenitor cells revealed by enhancers in the nestin gene

Using transgenic embryos, we have identified two distinct CNS progenitor cell-specific enhancers, each requiring the cooperation of at least two independent regulatory sites, within the second intron of the rat nestin gene. One enhancer is active throughout the developing CNS, while the other is specifically active in the ventral midbrain. These experiments demonstrate that neural progenitor cells in the midbrain constitute a unique subpopulation based upon their ability to activate the midbrain regulatory element. Our finding of differential enhancer activity from a gene encoding a structural protein reveals a previously unrecognized diversity in neural progenitor cell populations.     

Unique retina cell phenotypes revealed by immunological analysis of recoverin expression in rat retina cells

OBJECTIVE: This study was undertaken to determine the impact of previous cardiac surgery on the presentation, management, and outcome of late dissection of the ascending aorta. PATIENTS AND METHODS: From 1976 to 1998, type A dissection developed in 56 patients with a history of previous cardiac surgery. Interval from first operation to type A dissection was 49 +/- 47 months (0.3-180 months). Previous operations were coronary artery bypass grafting (n = 40), aortic valve replacement (n = 8), and other (n = 8). RESULTS: Type A dissection was acute in 34 patients and chronic in 22. In acute dissection, aortic insufficiency occurred in 50%, malperfusion in 12%, and rupture in 18%; 2 patients (6%) were in hemodynamically unstable condition because of rupture. Of patients with previous coronary bypass grafting, 98% had preoperative coronary angiography. Type A dissection was treated by supracoronary tube graft (84%), Bentall procedure (14%), or local repair (2%). Strategies for managing previous coronary bypass grafting included reimplantation of proximal anastomoses with a button of native aorta (29 patients), interposition graft to pre-existing saphenous vein grafts (9 patients), and new saphenous vein grafts (20 patients). Eight hospital deaths occurred (14%). CONCLUSIONS: We conclude that (1) patients having type A dissection late after cardiac surgery infrequently have cardiac tamponade and hemodynamic collapse; (2) patients with previous coronary bypass grafting require coronary angiography, because operative management must account for pre-existing coronary artery disease; and (3) operative mortality is low, and this may be attributable to preoperative hemodynamic stability, delineation of coronary anatomy in those with previous coronary bypass grafting, and operative treatment of coronary artery disease.     

A simple determination of steroidal alkaloid glycosides by thin-layer chromatography immunostaining using monoclonal antibody against solamargine

A method of determination for solasodine glycosides by using thin-layer chromatography (TLC) immunostaining was investigated. Solasodine glycosides separated by silica gel TLC were transferred to a polyvinylidene difluoride membrane. The membrane was treated with sodium periodate solution followed by bovine serum albumin (BSA), resulting in a solasodine glycoside-BSA conjugate. Conjugation was confirmed by matrix-assisted laser desorption/ionization mass spectrometry. Individual spots were stained by monoclonal antibody against solamargine. Immunostaining of solasodine glycosides was more sensitive compared to other stainings. The newly established immunostaining method can be extended to analysis of the distribution of solasodine glycoside in the plant body.     

Taxonomic relationships among spotted fever group rickettsiae as revealed by antigenic analysis with monoclonal antibodies

The spotted fever group (SFG) is made up of more than 20 different rickettsial species and strains. Study of the taxonomic relationships among the group has been attempted by phenotypic, genotypic, and phylogenetic analyses. In this study, we determined taxonomic relationships among the SFG rickettsiae by comparative analysis of immunogenic epitopes reactive against a panel of monoclonal antibodies. A total of 98 monoclonal antibodies, which were directed against epitopes on the major immunodominant proteins or on the lipopolysaccharide-like antigens of strains of Rickettsia africae, Rickettsia conorii, Rickettsia massiliae, Rickettsia akari, Rickettsia sibirica, and Rickettsia slovaca, were used in the study. The distribution and expression of the epitopes among 29 SFG rickettsiae and Rickettsia bellii were assessed by determination of reaction titers in a microimmunofluorescence assay. The results were scored as numerical taxonomic data, and cluster analysis was used to construct a dendrogram. The architecture of this dendrogram was consistent with previous taxonomic studies, and the implications of this and other findings are discussed.     

Diversity of human anti-D monoclonal antibodies revealed by reactions with chimpanzee red blood cells

Fifty-three human anti-D monoclonal antibodies (mAbs) revealed a striking diversity of reactions in tests with panels of chimpanzee red blood cells (RBCs) of various R-C-E-F blood group phenotypes (counterparts of the human Rh-Hr groups). The reactivities of these antibodies, which depended on the agglutination technique used, could be classified into four main types. These patterns of reactivity of anti-D mAbs with chimpanzee RBCs showed only limited correlation with types of reactions observed with human D variant RBCs. Primate red cells may, therefore, constitute an independent test system for subclassification of human monoclonal antibodies. Comparison of reactivities of human anti-D mAbs with chimpanzee and human D variant RBCs confirms the homology between the chimpanzee Rc, and the human D antigens. The chimpanzee Rc shares with human D the epitopes epD5, epD6/7 and epD8, but lacks epitopes epD1, epD2, epD3 and epD4 of the Rh mosaic, thus resembling the human D variants IVb and Vc.     

Proliferative potentials of glioma cells and vascular components determined with monoclonal antibody MIB-1

We present four cases of esophageal rupture (three iatrogenic, one Boerhaave syndrome) to demonstrate the difficulty in diagnosis and therapy. The current literature is discussed and conclusions are drawn as regards the modus operandi. All our patients were operated on. The site of esophageal rupture was always closed with a primary suture and substantial irrigation and drainage were performed. In two cases the suture line was in addition covered with fibrin glue or by an omentum flap, respectively. All patients survived and recovered was unremarkable. Our own results and a subsequent analysis of literature allow the following conclusions. At an early stage of esophageal rupture surgical intervention is indicated. The method of choice is primary closure of the rupture site by suture, possibly combined with a muscle or omentum flap. In cases of delayed diagnosis with advanced mediastinitis, suture of the rupture site should also be striven for. Additional coverage is advisable in these cases. Resection procedures with or without reconstruction should be done only in exceptional cases before of the high surgical risk.     

Characterization of Reed-Sternberg cells with granulocyte specific monoclonal antibody

Monoclonal antibody Leu M1 represents a highly specific marker to locate granulocyte antigen in Reed-Sternberg cells in Hodgkin's disease. Except the L&H variants of reed-sternberg cells in lymphocyte predominance variety in which antigens are probably sialylated. All the cases of non-Hodgkin's lymphoma were negative with this marker, because of absence of antigen. Therefore this specific marker characterizes granulocyte origin of reed-sternberg cells.     

Structural interlock between ligand-binding site and stalk-like region of beta1 integrin revealed by a monoclonal antibody recognizing conformation-dependent epitope

Integrin activation and sebsequent ligand binding to it are regulated by intracellular mechanisms called inside-out signaling, which are not fully understood and are accompanied by dynamic structural changes of the integrin molecule itself. A monoclonal antibody recognizing a conformation-dependent epitope on human beta1 integrin was produced and characterized in detail. This antibody, AG89, reacted with human integrin beta1 chain regardless of the alpha subunit. AG89 can recognize resting state beta1 integrin on the cells, but the reactivity is increased approximately 2-fold upon integrin activation by activating anti-beta1 antibodies and approximately 3-fold by Mn2+. Furthermore, occupation of the ligand-binding pocket by a soluble ligand (RGD peptide for alpha(v)beta1 and CS-1 peptide for alpha4beta1) resulted in maximum binding of AG89, indicating that the epitope for AG89 is exposed during the conformational changes of beta1 integrin upon activation/ligation. Epitope mapping by using interspecies chimeric beta1 revealed that the epitope for AG89 lies within residues 426-587, which corresponds to the cysteine-rich repeat structure located in the middle of the beta1 chain. The fact that binding of AG89 itself could activate the resting beta1 integrin indicates that exposure of the AG89 epitope in the membrane-proximal stalk-like domain and "opening" of the ligand-binding pocket at the outermost domain are physically linked. We propose that the integrin "signaling" is mediated by this direct physical transduction of conformational information along the integrin molecule.     

A quantitative analysis of cells in the ganglion cell layer of the chick retina

This study investigated the organization of cells in the ganglion cell layer (GCL) using Nissl staining, retrograde cell degeneration with axotomy of the optic nerve, and retrograde cell labeling by injections of horseradish peroxidase (HRP) into the optic nerve of chicks (posthatching day 1 and 8, P-1 and P-8). The total number of cells in the GCL was 6.1 x 10(6) (P-1) and 4.9 x 10(6) (P-8), and the cell density was 14,300 cells/mm2 (P-1) and 10,400 cells/ mm2 (P-8) on average. Two high-density areas, the central area (CA) and the dorsal area (DA), were observed in the central and dorsal retinas in both P-1 (22,000 cells/mm2 in CA, 19,000 cells/mm2 in DA) and P-8 chicks (19,000 cells/mm2 in CA, 12,800 cells/mm2 in DA). The cell densities in the temporal periphery (TP) and the nasal (NP) peripheral retinas were 7,800 cells/mm2 and 12,500 cells/mm2, respectively, in P-1 and 5,000 cells/ mm2 and 8,000 cells/mm2, respectively, in P-8 chicks. The cell density in the temporal periphery was 35% (P-8) lower than in the nasal periphery in both P-1 and P-8 chicks. Thirty percent (1.9 x 10(6) cells in P-1) of the total cells in the GCL were resistant to axotomy of the optic nerve. The distribution of the axotomy-resistant cells showed two high-density areas in the central and dorsal retinas, corresponding to the CA (5,800 cells/mm2) and the DA (3,200 cells/mm2). These cells also exhibited a center-peripheral increase (2,200 cells/mm2 in the TP) in P-1 chicks, but the high-density area was not found in the dorsal retina of P-8 chicks. From these data and the HRP study, the number of presumptive ganglion cells in P-8 chicks was estimated to be 4 x 10(6) (8,600 cells/mm2 on average), and the density in each area was 13,500 (CA), 10,200 (DA), and 4,300 (TP) cells/mm2. The peripheral/ center ratios of the density of ganglion cells were significantly different along the nasotemporal and dorsoventral axes. The density of ganglion cells decreased more rapidly toward the temporal periphery (TP/CA ratio: 0.47 in P-1 and 0.32 in P-8) than toward the nasal periphery (NP/CA ratio: 0.67 in P-1 and 0.52 in P-8). In contrast, there was no significant difference in the peripheral/center ratios between the dorsal retina (DP/CA ratio: 0.6 in P-1 and 0.56 in P-8) and ventral retina (VP/CA ratio: 0.58 in P-1 and 0.51 in P-8). A small peak in the density of the presumptive ganglion cells was detected in the dorsal retina of both P-1 chicks (10,800 cells/mm2) and P-8 chicks (10,200 cells/mm2). The HRP-labeled cells were small in the CA (M +/- SD: 35.7 +/- 9.1 microm2) and DA (40.0 +/- 11.3 microm2), and their sizes increased toward the periphery (63.4 +/- 29.7 microm2 in the TP) accompanied by a decrease in the cell density. However, the axotomy-resistant cells did not significantly increase in size toward the peripheral retina (12.2 +/- 2.2 microm2 in the CA, 15.2 +/- 3.2 microm2 in the DA, 15.1 +/- 3.8 microm2 in the TP). The characteristic distribution of ganglion cells could be related to visual behavior based upon the specialization of avian visual fields.     

Circulating immunoglobulin G1 antibody in patients with ulcerative colitis against the colonic epithelial protein detected by a novel monoclonal antibody

Autoimmunity has been implicated in the pathogenesis of ulcerative colitis (UC). Several studies have shown amplified immunoglobulin G1 (IgG1) antibody response in UC; however the immunoreactive antigen(s) is unknown. To study this antigen(s), mucosal colonic extract was prepared by sonication, ultracentrifugation followed by ion exchange chromatography in fast protein liquid chromatography. The fraction (enriched colonic peptide), that was most reactive to a novel monoclonal antibody, 7E12H12 (IgM isotype), was isolated and used to examine the immunoreactivity against the patients' serum samples. Two hundred and thirteen coded samples from 111 patients with UC (symptomatic and untreated (63), symptomatic and treated (26), remission (22)); 47 with Crohn's disease (CD) (40 were symptomatic and untreated, and 30 had colonic disease); 29 with acute diarrhoea caused by specific pathogen(s); 10 with systemic lupus erythematosus, and 16 normal subjects were examined against the enriched colonic peptide by IgG subtype specific enzyme linked immunosorbent assays (ELISAs). Total IgG antibody reactivity was significantly (p < 0.01) higher only in symptomatic and untreated UC patients compared with each of the non-UC group, but the sensitivity was only 50%. IgG2 and IgG3 reactivities were not different among various groups. The IgG1 antibody reactivity against the enriched colonic peptide, however, differentiated UC patients from CD and each of the other non-UC groups. Seventy nine per cent of the patients with UC, treated or untreated, symptomatic or in remission, had significantly (p < 0.0001) higher IgG1 antibody against the enriched colonic peptide when compared with each of the other non-UC groups. Only 12% of CD serum samples and none of the other control serum samples reacted. Using purified serum IgG1 and 7E12H12-IgM, by 7E12H12 reactive peptide indeed reacts with UC-IgG1 antibody but not with control IgG1.     

Proliferative potential in pituitary adenomas: measurement by monoclonal antibody MIB-1

The proliferative potential of 45 pituitary adenomas was compared with their biological behaviour as determined by immunohistochemical studies, radiological findings, and clinical manifestations. The PCI (proliferating cell index) as measured using antibody MIB-1 in this study ranged from 0.05 to 4.80%, with an average PCI of 1.49 +/- 0.19% (mean +/- standard error of the mean). There was no significant correlation between proliferation and hormonal state, maximum size, intra-adenomatous haemorrhage, or invasiveness. However, a PCI > or = 1.5% appeared to correlate with the likelihood of tumour regrowth (regrowth rate: 50%); for PCIs < 1.5%, the rate was 16%. Regrowth adenomas had a higher mean MIB-1 PCI than non-regrowth adenomas [2.34 +/- 0.58% (SE) versus 1.14 +/- 0.16%, p < or = 0.05]. MIB-1 PCIs may provide information that is useful for planning follow-up studies and treatment after surgical resection.     

Characterization of human IgG1 monoclonal antibody against gangliosides expressed on tumor cells

In this study, our intention was to describe the decision making of nurses practicing in intensive care, and the differences of nurses' decision making in Canada, Finland, Northern Ireland, Switzerland, and the United States. The instrument used in the study was a 56-item Likert-type questionnaire that has been used in previous studies and has proved to be a reliable tool. The target group comprised a nonrandom sample of nurses (N = 314) from five countries. The samples are not representative; therefore, the results in these cases cannot be generalized. The results showed that the decision making of nurses practicing in intensive care was broadly based, and that there were some country differences in data collection, problem definition, and planning. In contrast, decision making related to the implementation and evaluation of nursing is quite similar in the different countries. Canada and the United States on the one hand, and Finland, Northern Ireland, and Switzerland on the other, showed more similarities with each other in data collection, problem definition, and nursing planning related to decision making. Neither experience nor nurse's knowledge structure was associated with different decision-making approaches.     

Antigenic variation of the Epstein-Barr virus nuclear antigen EBNA1 as revealed by monoclonal antibodies

To obtain specific immunological probes for studying molecular mechanisms involved in cell renewal, cell differentiation, and pattern formation in intact and regenerating planarians, we have produced a hybridoma library specific for the asexual race of the fresh-water planarian Dugesia (Girardia) tigrina. Among the 276 monoclonal antibodies showing tissue-, cell-, cell subtype-, subcellular- and position-specific staining, we have found monoclonal antibodies against all tissues and cell types with the exception of neoblasts, the undifferentiated totipotent stem-cells in planarians. We have also detected position-specific antigens that label anterior, central, and posterior regions. Patterns of expression uncovered an unexpected heterogeneity among previously thought single cell types, as well as interesting cross-reactivities that deserve further study. Characterization of some of these monoclonal antibodies suggests they may be extremely useful as molecular markers for studying cell renewal and cell differentiation in the intact and regenerating organism, tracing the origin, lineage, and differentiation of blastema cells, and characterizing the stages and mechanisms of early pattern formation. Moreover, two position-specific monoclonals, the first ones isolated in planarians, will be instrumental in describing in molecular terms how the new pattern unfolds during regeneration and in devising the pattern formation model that best fits classical data on regeneration in planarians.     

Heterogeneity among muscle precursor cells in adult skeletal muscles with differing regenerative capacities

Skeletal muscle has a remarkable capacity to regenerate after injury, although studies of muscle regeneration have heretofore been limited almost exclusively to limb musculature. Muscle precursor cells in skeletal muscle are responsible for the repair of damaged muscle. Heterogeneity exists in the growth and differentiation properties of muscle precursor cell (myoblast) populations throughout limb development but whether the muscle precursor cells differ among adult skeletal muscles is unknown. Such heterogeneity among myoblasts in the adult may give rise to skeletal muscles with different regenerative capacities. Here we compare the regenerative response of a masticatory muscle, the masseter, to that of limb muscles. After exogenous trauma (freeze or crush injuries), masseter muscle regenerated much less effectively than limb muscle. In limb muscle, normal architecture was restored 12 days after injury, whereas in masseter muscle, minimal regeneration occurred during the same time period. Indeed, at late time points, masseter muscles exhibited increased fibrous connective tissue in the region of damage, evidence of ineffective muscle regeneration. Similarly, in response to endogenous muscle injury due to a muscular dystrophy, widespread evidence of impaired regeneration was present in masseter muscle but not in limb muscle. To explore the cellular basis of these different regenerative capacities, we analyzed the myoblast populations of limb and masseter muscles both in vivo and in vitro. From in vivo analyses, the number of myoblasts in regenerating muscle was less in masseter compared with limb muscle. Assessment of population growth in vitro indicated that masseter myoblasts grow more slowly than limb myoblasts under identical conditions. We conclude that the impaired regeneration in masseter muscles is due to differences in the intrinsic myoblast populations compared to limb muscles.     

Developmentally regulated spontaneous activity in the embryonic chick retina

Even before birth and the onset of sensory experience, neural activity plays an important role in shaping the vertebrate nervous system. In the embryonic chick visual system, activity in the retina before vision has been implicated in the refinement of retinotopic maps, the elimination of transient projections, and the survival of a full complement of neurons. In this study, we report the detection of a physiological substrate for these phenomena: waves of spontaneous activity in the ganglion cell layer of the embryonic chick retina. The activity is robust and highly patterned, taking the form of large amplitude, rhythmic, and wide-ranging waves of excitation that propagate across the retina. Activity waves are most prominent and organized between embryonic days 13-18, coinciding with the developmental period during which retinal axons refine their connections in their targets. The spatial and temporal features of the patterns observed are consistent with the role of activity patterns in shaping eye-specific projections and retinotopic maps but inconsistent with the hypothesis that they specify lamina-specific projections in the tectum. Antagonists of glutamatergic and glycinergic transmission and of gap junctional communication suppress spontaneous activity, whereas antagonists to GABAergic transmission potentiate it. Based on these results, we propose that spontaneous activity in the ganglion cells is regulated by chemical inputs from both bipolar and amacrine cells and by gap junctional coupling involving ganglion cells.     

JSJ-1, an anti-spermidine monoclonal antibody with potential clinical applications

Thirtysix patients with persistent gestational trophoblastic disease were analysed in The Clinic of Cancer of Female Reproductive System. Using transvaginal USG, the volume of pathological foci within myometrium, diametre of theca lutein ovarian cysts and uterus length before, during and after the treatment were registred. In order to check the effectiveness of USG examination in the monitoring of GTD treatment particular USG factors were compared with hCG level. On 28 patients (77.7%) in USG examination pathological changes in uterus and ovaries were seen. The relation between hCG level and tumor volume, uterus length and theca lutein ovarian cysts was stated. That proves the usefulness of transvaginal USG examination in the monitoring of treatment of patients with persistent gestational trophoblastic disease.     

Characterisation of chicken monocytes, macrophages and interdigitating cells by the monoclonal antibody KUL01

The distribution, function and ontogeny of the mononuclear phagocyte system of the chicken were characterised using the monoclonal antibody (MAb) KUL01. KUL01 specifically recognises chicken monocytes, macrophages and interdigitating cells, as well as activated microglia cells. Its tissue distribution allowed to discriminate KUL01 from all earlier described MAb, reactive with mononuclear phagocytes. The specificity of KUL01 for mononuclear phagocytes was further confirmed in functional assays: KUL01-positive macrophages in spleen and liver actively took up colloidal carbon, while monocytes and spleen and gut macrophages contained non-specific esterase and acid phosphatase activities characteristic for antigen-processing. Further, it was demonstrated that KUL01-reactive peripheral blood monocytes express MHC-II, but not CD4. In all tissues investigated, the same morphological subtypes of macrophages were detected in chicken at similar localisations as in mammals, indicating a high degree of conservation between the mononuclear phagocyte system of the chicken and of mammals.