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PubMed2004

Microarray analysis of Lyn-deficient B cells reveals germinal center-associated nuclear protein and other genes associated with the lymphoid germinal center.

Lyn is the only member of the Src family expressed in DT40 B cells, which provide a unique model to study the singular contribution of this protein tyrosine kinase (PTK) family to cell signaling. In these cells, gene ablation of Lyn leads to defective B cell receptor signaling. Complementary DNA array analysis of Lyn-deficient DT40 cells shows that the absence of Lyn leads to down-regulation of numerous genes encoding proteins involved in B cell receptor signaling, proliferation, control of transcription, immunity/inflammation response, and cytoskeletal organization. Most of these expression changes have not been previously associated with Lyn PTK signaling. They include alterations in mRNA levels of germinal center-associated nuclear protein (germinal center-associated DNA primase) (GANP), CD74, CD22, NF-kappaB, elongation factor 1alpha, CD79b, octamer binding factor 1, Ig H chain, stathmin, and gamma-actin. Changes in GANP expression were also confirmed in Lyn-deficient mice, suggesting that Lyn PTK has a unique function not compensated for by other Src kinases. Because Lyn-deficient mice have impaired development of germinal centers in spleen, the decreased expression of GANP in the Lyn-deficient DT40 cell line and Lyn-deficient mice suggests that Lyn controls the formation and proliferation of germinal centers via GANP. GANP promoter activity was higher in wild-type vs Lyn-deficient cells. Mutation of the PU.1 binding site reduced activity in wild-type cells and had no effect in Lyn-deficient cells. The presence of Lyn enhanced PU.1 expression in a Northern blot. Thus, the following new signaling pathway has been described: Lyn-->PU.1-->GANP.

Animals↗

PubMed1980

Germinal centres and the B cell system. IV. Functional characteristics of rabbit appendix germinal centre (-derived) cells.

Functional capacities of rabbit appendix germinal centre cells were tested in appendectomized, 450 rads X-irradiated rabbits, reconstituted with autologous appendix germinal centre cells, which are obtained by mechanical separation--"stripping"--of the appendix. For comparison, functional capacities of appendix germinal centre derived cells were tested in rabbits, 450 rads X-irradiated with the appendix shielded. In both types of experimental systems, slow restoration of primary IgM responsiveness to Salmonella Java paratyphi-B (PAR) was observed; in contrast, the capacity for memory cell production (of both mu and gamma-type) to PAR was restored very early in both systems. These results suggest germinal centres to be antigen dependent microenvironments for selective as well as non-selective amplification of immature B cells, which after subsequent maturation become part of the pool of either memory or virgin mature B cells respectively.

Animals↗

PubMed1992

Cellular composition of germinal centers in lymph nodes after HIV-1 infection: evidence for an inadequate support of germinal center B lymphocytes by follicular dendritic cells.

Germinal centers in lymph nodes with follicular hyperplasia from 15 patients with HIV-1 infection were analyzed by qualitative and quantitative electron microscopical methods and compared with control follicular hyperplasia (FH). Using a pattern recognition method, two main clusters were recognized within the germinal centers of HIV and FH lymph nodes on the basis of the relative frequencies of small centroblast and centrocytes. All FH lymph nodes and 6 HIV-1 lymph nodes (HIV-Clu-1) were placed in cluster 1; 9 HIV-1 lymph nodes (HIV-Clu-2) formed cluster 2. Germinal centers in the HIV-Clu-2 lymph nodes were characterized by a cell composition of predominantly lymphoid blasts and decreased numbers of centrocytes, but without altered numbers of mitotic figures. The frequency distribution of ultrastructurally distinct FDC subtypes differed between these clusters. In HIV-Clu-2 the frequencies of FDC types with an undifferentiated and regressive morphology occurred at a higher frequency, whereas FDC types with a highly differentiated morphology had a lower frequency. We conclude that 9 out of 15 lymph nodes with HIV-1 associated follicular hyperplasia show changes in FDC morphology indicative of a less differentiated functional stage of FDC. The changes in FDC morphology are closely associated with changes in the germinal center B-cell population resulting in an inverted blast to the centrocyte ratio.

Dendritic Cells↗

PubMed1979

Photocontrol of the germination of onoclea spores: v. Analysis of germination processes by means of temperature.

The physiological nature of photoinduced germination of Onoclea sensibilis L. spores was investigated by temporarily applying a range of temperatures, particularly 40 C, before and after short light treatment. Controls were germinated at 25 C.The preinduction phase, during which photosensitivity is maximally developed in the dark, is sensitive to temperature. Treatment at 40 C for 8 or more hours reduces the developed photosensitivity to a minimal level, but the inhibition by 40 C treatment is reversed slowly after subsequent incubation at 25 C in the dark. The postinduction phase, in which dark processes lead to stain uptake and eventually to visible protrusion, is also sensitive to temperature. Inhibition by 40 C occurs shortly after photoinduction, but disappears 6 or more hours after irradiation. Postinduction spores whose germination is inhibited by 40 C treatment recover the ability to germinate after subsequent incubation at 25 C plus a second light treatment. The inhibition and recovery take place faster in postinduction spores than in preinduction spores. In addition, escape from 40 C inhibition is found in the postinduction phase but not in the preinduction phase. Temperatures lower than 25 C exert slow inhibition of both pre- and postinduction processes, and 30 to 35 C act to stimulate germination.In comparison with our earlier work with anaerobiosis and cycloheximide, the postinduction step inhibited by 40 C can be located shortly after the step inhibited by anaerobiosis but before the cycloheximide sensitive step.

Journal Article↗

PubMed1988

Germination-specific cortex-lytic enzyme is activated during triggering of Bacillus megaterium KM spore germination.

Antibodies were raised against purified germination-specific cortex-lytic enzyme (GSLE) from spores of Bacillus megaterium KM which neutralized the ability of GSLE to germinate permeabilized spores. Western blotting of dormant spore and vegetative cell fractions separated by SDS-PAGE demonstrated that GSLE is spore-specific and that greater than 90% of the GSLE is associated with the dormant spore cortex peptidoglycan as a phosphorylated 63kD pro-form, which could only be visualized after lysozyme digestion of the peptidoglycan. During germination, the 63kD pro-form of GSLE is processed to release the active enzyme, which had an apparent molecular weight of 30kD. Inhibitor studies demonstrated that GSLE activation occurs as part of the commitment reaction and thus represents the first-identified enzymatic event to occur during germination triggering. Proteins that cross-react with anti-GSLE sera are present in spore fractions of other species.

Bacillus megaterium↗

PubMed1986

Germination of Bacillus cereus spores induced by purine ribosides and their analogs: effects of modification of base and sugar moieties of purine nucleosides on germination-inducing activity.

Purine riboside and some of its analogs were tested for their ability to induce germination of Bacillus cereus T spores. Hypoxanthine and adenine showed no germination-inducing activity either in the present or absence of D-ribose or its phosphorylated derivatives. Purine riboside and 18 analogs with modified purine base were all able to induce germination of the spores to various extents. In contrast to this, the requirement for the sugar moiety in the purine riboside appeared to be more stringent. Only those nucleosides that contained either D-ribose or deoxy-D-ribose, and certain species of azole derivatives such as 5-aminoimidazole-4-carboxamide covalently linked to the C(1') of the sugar actively induced germination.

Bacillus cereus↗

PubMed2004

Storage behavior of Chionanthus retusus seed and asynchronous development of the radicle and shoot apex during germination in relation to germination inhibitors, including abscisic acid and four phenolic glucosides.

Studies on seed storage of Chionanthus retusus Lindl. & Paxt. revealed an orthodox behavior, one which showed both desiccation and freezing tolerance. An epicotyl after-ripening dormancy was expressed in C. retusus seeds by slow growth of the shoot apex relative to more rapid growth of the radicle when seeds were germinated at 30/20 degrees C. Although these seeds exhibit radicle protrusion, they must be after-ripened for another 8-10 weeks at 30/20 degrees C in order to obtain normal shoot growth. Removal of the endosperm, however, quickly stimulated cotyledon and shoot emergence without the additional after-ripening. Water-soluble glucoside phenolics, GL-3, Nuzhenide, ligustroside and oleoside dimethyl ester are present at relatively high levels in endosperm of freshly harvested seeds. These glucoside phenolics are excreted from the endosperm during subsequent after-ripening. Embryo and endosperm tissue from seed germinating at 30/20 degrees C (germination being defined by protrusion of the radicle) had a 10 times lower abscisic acid (ABA) content than similar tissues from freshly harvested mature seed. However, no shoot growth occurred even with the 10-fold reduction in ABA and a concomitant increase in endogenous gibberellins A1, A4 and A20. Thus, epicotyl dormancy during the first 8 weeks of after-ripening at 30/20 degrees C may be controlled by factors other than high ABA, i.e., the slow development of the shoot apex following radicle protrusion may be controlled more by high levels of glucoside phenolics than by diminished ABA and elevated GA levels.

Abscisic Acid↗

PubMed1980

Quantitative determination of germines. The rate of transformation of germine-3,16-diacetate in buffered solutions of pH 7.4.

Germine (GER), germine-3-monoacetate (GMA) and germine-3,16-diacetate (GDA) when treated with 9:1 mixture of concentrated sulfuric acid and glacial acetic acid yield a deep purple color with characteristic ultraviolet absorption spectra. Combination of the color reaction with TLC makes possible the quantitative determination of these compounds in buffered solutions of pH 7.4 with a > 90% yield and satisfactory accuracy. It was observed that GDA incubated in such solutions is slowly transformed into GMA. The transformation of GDA to GMA was significantly faster with higher than with lower concentrations of GDA. There was no detectable breakdown of GMA under the same experimental conditions.

Animals↗

PubMed1989

Hungarian surveillance of germinal mutations. Lack of detectable increase in indicator conditions caused by germinal mutations following the Chernobyl accident.

The Hungarian surveillance of germinal mutations is based on three indicator conditions seen in offspring, i.e., 15 sentinel anomalies, Down syndrome and component anomaly pairs of unidentified multiple congenital anomalies. It is an "opportunistic program," because the necessary data are available from the Hungarian Congenital Malformation Registry. This system is described and the criteria of a good registry are summarized. The analysis of indicator conditions caused by germinal mutations did not reveal any measurable mutagenic effects in Hungary following the accident at the Chernobyl nuclear power plant. The pros and cons of germinal mutation surveillance are discussed.

Abnormalities, Multiple↗

PubMed1974

Effects of Germination-promoting Substances Given in Conjunction with Red Light on the Phytochrome-mediated Germination of Dormant Lettuce Seeds (Lactuca sativa L.).

Ethylene or thiourea can substitute for gibberellic acid but not for red light in breaking the secondary dormancy induced by extended dark storage of fully hydrated lettuce seeds (Lactuca sativa cv. Grand Rapids). After 10 days of storage, ethylene, thiourea, or gibberellic acid applied either separately or in any combination in conjunction with red light induced near maximal germination. When applied separately without red light, none of the substances promoted germination of seeds stored 10 days. Combinations of any two or all three of the substances in the absence of red light induced some germination but no combination was as effective as any single substance given with red light.

Journal Article↗

PubMed1975

Photocontrol of the Germination of Onoclea Spores: II. Analysis of Germination Processes by Means of Anaerobiosis.

The oxygen requirements during the three phases of photoinduced germination of Onoclea sensibilis L. spores were analyzed by temporarily applying nitrogen atmosphere. The dark preinduction phase, during which the spores imbibe water and establish sensitivity to irradiation, involves an oxidative process which can be reversibly inhibited and stimulated by nitrogen and air, respectively. The induction phase of germination is characterized by a pure photochemical reaction, independent of temperature and oxygen. The postinduction phase, when the photoproduct triggers dark processes eventually leading to the protrusion of the rhizoidal or protonematal cells, involves an oxidative process which occurs within the first 10 hours of this phase. This oxidative process differs in kinetic characteristics from that in the preinduction phase. The oxidative process is inhibited by nitrogen treatment, but following nitrogen inhibition the ability of the spores to germinate can be reinstated by a long period of air intervening between the nitrogen treatment and a second irradiation. This suggests that enzymes or reactants which are needed in the postinduction process decay under anaerobic conditions and are resynthesized when the spores are transferred to air. Spores take up acetocarmine stain towards the latter part of the postinduction phase. Stain uptake is apparently succeeded very closely by cell division, and some time later by protrusion of the germling cells.

Journal Article↗

PubMed1975

Lipid metabolism during bacterial growth, sporulation, and germination: kinetics of fatty acid and macromolecular synthesis during spore germination and outgrowth of Bacillus thuringiensis.

The timing and kinetics of fatty acid synthesis are delineated for Bacillus thuringiensis spore germination and outgrowth by analyzing [U-14C]acetate and [2-3H]glycerol incorporation into chloroform-methanol-extractable and trichloroacetic acid-precipitable lipids. In addition to measurement of pulsed and continuous labeling of fatty acids, monitoring the incorporation of radioactive phenylalanine, thymidine, and uridine from the onset of germination through first cell division provides a profile of biochemical activities related to membrane differentiation and cellular development. Upon germination, ribonucleic acid synthesis is initiated, immediately followed by rapid and extensive fatty acid synthesis that in turn precedes protein, deoxyribonucleic acid and triglyceride synthesis. Significantly, formation of fatty acids from acetate exhibits further developmental periodicity in which a large transient increase in fatty acid synthetic activity coincides with the approach of cell division. Radiorespirometric analyses indicates only slight oxidative decarboxylation of acetate and corroborates the extreme involvement of acetate in specific fatty acid biosynthetic reactions throughout cellular modification. These findings graphically demonstrate an intimate association of fatty acid metabolism with commitment to spore outgrowth and subsequent cell division.

Acetates↗

PubMed2005

Expression of the human germinal center-associated lymphoma (HGAL) protein, a new marker of germinal center B-cell derivation.

We identified the human germinal center-associated lymphoma (HGAL) in gene-expression profiling studies of diffuse large B-cell lymphoma (DLBCL). The expression of HGAL correlated with survival in patients with DLBCL. The HGAL gene is the human homolog of M17, a mouse gene expressed specifically in normal germinal center (GC) B cells. We generated a monoclonal antibody against the HGAL protein and show that HGAL is expressed in the cytoplasm of GC lymphocytes and in lymphomas of GC derivation. Among 727 lymphomas tested by immunohistochemistry on tissue microarrays, HGAL staining was found in follicular lymphomas (103 of 107), Burkitt lymphomas (40 of 40), mediastinal large B lymphomas (7 of 8), and in DLBCLs (103 of 151). Most marginal zone lymphomas lacked HGAL staining. Lymphocyte-predominant Hodgkin lymphomas (12 of 17) and, surprisingly, classical Hodgkin lymphomas (78 of 107) were found to be positive. Hierarchical clustering of comparative immunohistologic results in DLBCLs demonstrates that the expression of HGAL is similar to 2 other GC-associated proteins, BCL6 and CD10, but different from 2 markers associated with a non-GC phenotype, MUM1/IRF4 and BCL2. The restricted expression and GC specificity of HGAL protein suggest that it may have an important role in the diagnosis of specific lymphomas, and, potentially in the identification of subtypes associated with different prognoses.

Animals↗

PubMed2006

Expression of the human germinal center-associated lymphoma (HGAL) protein identifies a subset of classic Hodgkin lymphoma of germinal center derivation and improved survival.

The human germinal-center-associated lymphoma (HGAL) gene and its cognate protein are expressed in a germinal center (GC)-specific manner. Its expression in classic Hodgkin lymphoma (cHL) prompted us to address whether HGAL expression could distinguish biologically distinct subgroups of cHL. Tissue microarrays from 145 patients treated with curative intent showed HGAL staining in 75% and was closely correlated with MUM1/IRF4 (92%) expression. BCL6 (26%), CD10 (0%), BCL2 (31%), Blimp1 (0.02%), and Epstein-Barr virus (EBV) (20%) showed no specific correlation; neither did phospho-STAT6, a key mediator of IL-4 and IL-13 signaling that induces HGAL and is implicated in cHL pathogenesis. In our study cohort, the 5-year overall survival (OS) correlated with young age (less than 45 years, P < .001), low stage (stage I and II, P = .04), and low International Prognostic Score (P = .002). In univariate analysis, HGAL expression was associated with improved OS (P = .01) and failure-free survival (FFS) (P = .05) but was not independent of other factors in multivariate analysis of OS or FFS. The expression of the GC-specific marker HGAL in a subset of cHL suggests that these cHLs retain characteristics of GC-derived lymphomas. The association with improved OS in univariate but not multivariate analysis suggests that HGAL expression is related to known clinical parameters of improved survival.

Adult↗

PubMed2001

B-cell development in progressively transformed germinal centers: similarities and differences compared with classical germinal centers and lymphocyte-predominant Hodgkin disease.

Progressively transformed germinal centers (PTGCs) are histologic structures mainly composed of small resting B cells and intermingled proliferating centroblast-like cells. The B-cell differentiation processes within PTGCs and their relation to classical germinal centers (GC) and to lymphocyte-predominant Hodgkin disease (LPHD), with which PTGCs are often associated, are largely unknown. To address these issues, single small resting (Ki67-) and proliferating (Ki67+) centroblast-like cells were isolated from 7 PTGCs of 5 lymph nodes, and rearranged immunoglobulin genes were amplified and sequenced. Most small resting B cells were clonally unrelated, and most carried unmutated immunoglobulin gene rearrangements resembling mantle zone B cells. Small resting B cells with mutated immunoglobulin gene rearrangements may represent centrocytes, memory B cells, or both. Among the centroblast-like Ki67+ cells, expanded B-cell clones were observed in 6 of 7 PTGCs analyzed. Clonally related V region genes showed extensive intraclonal diversity, and the mutation pattern indicated stringent selection of the cells for the expression of functional antigen receptors. Thus, somatic hypermutation, clonal expansion, and selection occur also in the disorganized PTGC microenvironment, as in classical GCs. In lymph nodes affected by PTGCs, no clonal expansion across the borders of individual PTGCs was observed, distinguishing PTGCs from LPHD.

Adolescent↗

PubMed2004

Comparative study on the modulation of IgE and cytokine production by Phellinus linteus grown on germinated brown Rice, Phellinus Linteus and germinated brown rice in murine splenocytes.

We compared the immunomodulating activities in mice of extracts from Phellinus linteus grown on germinated brown rice (PB), Phellinus linteus (PL) alone, and germinated brown rice (BR) alone. The PL, BR and PB-treated mice were administered with the respective extract (2 mg/head/day) by oral gavage for 4 weeks. All extracts markedly decreased the IgE production and allergic responses in serum and splenocytes. PL and PB increased the proportion of CD4(+) but not CD8(+) T cells in splenocytes. Cytokine production was significantly augmented in all treated mice; the concentration of IFN-gamma was greater in the PL, BR and PB mice than in the control group. The concentration of IL-10 was lower in the BR group than in the other groups. These results may be related to the suppression of IgE production. We conclude that PB modulated the immune responses of IgE production and Th1/Th2 cytokine secretion in murine splenocytes.

Animals↗

PubMed1994

Characterization of a major nucleoplasmin-like germinal vesicle protein which is rapidly phosphorylated before germinal vesicle breakdown in Spisula solidissima.

Oocytes of the surf clam, Spisula solidissima, are arrested at the G2/M boundary of meiotic prophase I. At this stage, they possess a prominent germinal vesicle (GV), comprising about 25% of the total oocyte volume, in which pools of mRNAs and proteins that facilitate the rapid rounds of cell division which occur early in development are stored. We have isolated and characterized an abundant 49-kDa phosphoprotein, localized exclusively to the GV, which shares properties with nucleoplasmin. Like nucleoplasmin, this 49-kDa protein is a heat-stable, highly acidic phosphoprotein [containing over 32% (Glx + Asx) with an isoelectric point of about 4.0] and is soluble in 80% ammonium sulfate. In contrast to the pentameric nucleoplasmin, much of this protein is isolated as a disulfide-linked multimer which migrates at 120 kDa. In addition, a fraction of the 49-kDa protein is associated via disulfide bonds to a doublet of 42-kDa proteins. In vivo, this 49-kDa protein is phosphorylated prior to germinal vesicle breakdown (GVBD) and at 5 min after oocyte activation rapidly incorporates 32P to 60% of maximal level, suggesting that this protein plays a major role in the cascade of events which lead to oocyte activation and GVBD.

Animals↗

PubMed1996

Effect of cadmium on germination, amylases and rate of respiration of germinating pea seeds.

Growth of embryonic axis of germinating pea seeds (Pisum sativum cv. Bonneville) was significantly inhibited by as low as 0.25 mM cadmium and the elongation of the radicle was affected more severely than that of the plumule. Total amylolytic activity, as well as activities of alpha- and beta-amylases, diminished progressively with increasing concentrations of the metal in the media. The deleterious effect on alpha-amylase persisted throughout, whereas beta-amylase activity recovered with time. The rate of respiration of seeds, measured by oxygen uptake, was markedly impeded by cadmium and the phase of rapid and linear development of respiratory activity (after 3 days of imbibition) was almost completely, suppressed in the presence of higher concentrations (1 mM) of the metal. The observed impact of cadmium on starch mobilization and on respiratory activity are discussed in relation to its adverse effects on seed germination.

Journal Article↗