The role of the CD134-CD134 ligand costimulatory pathway in alloimmune responses in vivo

The CD134-CD134 ligand (CD134L) costimulatory pathway has been shown to be critical for both T and B cell activation; however, its role in regulating the alloimmune response remains unexplored. Furthermore, its interactions with other costimulatory pathways and immunosuppressive agents are unclear. We investigated the effect of CD134-CD134L pathway blockade on allograft rejection in fully MHC-mismatched rat cardiac and skin transplantation models. CD134L blockade alone did not prolong graft survival compared with that of untreated recipients, and in combination with donor-specific transfusion, cyclosporine, or rapamycin, was less effective than B7 blockade in prolonging allograft survival. However, in combination with B7 blockade, long-term allograft survival was achieved in all recipients (>200 days). Moreover, this was synergistic in reducing the frequency of IFN-gamma-producing alloreactive lymphocytes and inhibiting the generation of activated/effector lymphocytes. Most impressively, this combination prevented rejection in a presensitized model using adoptive transfer of primed lymphocytes into athymic heart transplant recipients. In comparison to untreated recipients (mean survival time (MST): 5.3 +/- 0.5 days), anti-CD134L mAb alone modestly prolonged allograft survival (MST: 14 +/- 2.8 days) as did CTLA4Ig (MST: 21.5 +/- 1.7 days), but all grafts were rejected within 24 days. Importantly, combined blockade further and significantly prolonged allograft survival (MST: 75.3 +/- 12.7 days) and prevented the expansion and/or persistence of primed/effector alloreactive T cells. Our data suggest that CD134-CD134L is a critical pathway in alloimmune responses, especially recall/primed responses, and is synergistic with CD28-B7 in mediating T cell effector responses during allograft rejection. Understanding the mechanisms of collaboration between these different pathways is important for the development of novel strategies to promote long-term allograft survival.     

Gangliosides from human melanoma tumors impair dendritic cell differentiation from monocytes and induce their apoptosis

Gangliosides are ubiquitous membrane-associated glycosphingolipids, which are involved in cell growth and differentiation. Most tumor cells synthesize and shed large amounts of gangliosides into their microenvironment, and many studies have unraveled their immunosuppressive properties. In the present study we analyzed the effects of GM3 and GD3 gangliosides, purified from human melanoma tumors, on the differentiation of monocyte-derived dendritic cells (MoDC). At concentrations close to those detected in the sera from melanoma patients, both gangliosides dose-dependently inhibit the phenotypic and functional differentiation of MoDC, as assessed by a strong down-regulation of CD1a, CD54, CD80, and CD40 Ags and impaired allostimulatory function on day 6 of culture. Furthermore, GM3 and GD3 gangliosides decreased the viable cell yield and induced significant DC apoptosis. Finally, addition of GD3 to differentiating DC impaired their subsequent maturation induced by CD154. The resulting DC produced low amounts of IL-12 and large amounts of IL-10, a cytokine pattern that might hamper an efficient antitumor immune response. In conclusion, the results demonstrate that gangliosides impair the phenotypic and functional differentiation of MoDC and induce their apoptosis, which may be an additional mechanism of human melanoma escape.     

Phylogeny of the lichen genus Placopsis and its allies based on Bayesian analyses of nuclear and mitochondrial sequences

The phylogenetic relationships of the lichen genus Placopsis and related genera in the Agyriales were analyzed using molecular data. We obtained a total of 66 new sequences from the nuclear ITS, LSU and the mitochondrial SSU rDNA. Phylogenetic analyses were conducted in a Bayesian and a maximum-parsimony framework. Our analyses show that Placopsis is paraphyletic with members of Orceolina nesting within the genus. A morphological character supporting the Placopsis-Orceolina clade is the non-amyloid ascus. The section Aspiciliopsis as defined by sunken fruiting bodies is not supported, but the type species of Aspiciliopsis is more closely related to Orceolina. This clade shares apothecia with reduced amphithecia as apomorphic character. We suggest resurrecting the generic name Aspiciliopsis. Trapelia is the sister genus to Placopsis and Aspiciliopsis/Orceolina.     

Expression and secretion of ficolin beta by porcine neutrophils

Ficolins are collagenous lectins that bind N-acetylglucosamine (GlcNAc) as well as some bacterial surfaces, and may have opsonic and complement-activating functions. Ficolin alpha in porcine plasma binds Actinobacillus pleuropneumoniae serotype 5 (APP) in a GlcNAc-dependent manner. In the present study, we discovered that porcine neutrophils, but not platelets or mononuclear cells, contained a different ficolin that migrated as a 39-kDa band on SDS-PAGE and resembled a minor component of plasma ficolins that binds APP. However, neutrophil ficolins (pI range 6.4-7.4) were readily distinguished from plasma ficolin alpha (pI 5.2-5.8) by 2D PAGE. Neutrophil ficolin was consistent with ficolin beta by pI and peptide mass fingerprinting with matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry. Porcine neutrophils expressed ficolin beta, but not ficolin alpha, as determined by RT-PCR. Ficolin beta was present in the membrane and cytoplasmic fractions of nonactivated neutrophils, but the majority of ficolin beta was secreted upon activation with PMA. Ficolin alpha readily bound to intact APP, but ficolin beta did not under the same conditions. These studies demonstrate that neutrophils express ficolin beta and secrete it when activated; however, ficolin beta may have different binding functions than ficolin alpha.     

Dispersal biology of Liatris scariosa var. novae-angliae (Asteraceae), a rare New England grassland perennial

Propagule dispersal biology is a crucial avenue of research for rare plant species, especially those adapted to disturbance, such as northern blazing star (Liatris scariosa var. novae-angliae), a rare, early-successional New England grassland perennial. We examined the dispersal ability of northern blazing star propagules collected from 14 populations covering the entire latitudinal range of the taxon. Multiple regression demonstrated that dispersal ability, as measured by drop time in still air and flight distance in a low-speed wind tunnel, decreased significantly with propagule size and achene length, and increased with achene width and (for flight distance) pappus length. We used this multiple regression model to test for differences in predicted dispersal capability among maternal families, populations, and inland, coastal, and island habitats. Dispersal capability differed significantly among families and populations but not regions, and allometric relationships between morphological measurements were consistent across populations. Overall, dispersal capability was negatively correlated with germination success in a common greenhouse environment. However, germination success for a given dispersal ability, as well as achene shape, differed among populations. These results suggest specific populations to be targeted for management efforts promoting dispersal and establishment.     

Heterogeneous selection at specific loci in natural environments in Arabidopsis thaliana

Genetic variation for quantitative traits is often greater than that expected to be maintained by mutation in the face of purifying natural selection. One possible explanation for this observed variation is the action of heterogeneous natural selection in the wild. Here we report that selection on quantitative trait loci (QTL) for fitness traits in the model plant species Arabidopsis thaliana differs among natural ecological settings and genetic backgrounds. At one QTL, the allele that enhanced the viability of fall-germinating seedlings in North Carolina reduced the fecundity of spring-germinating seedlings in Rhode Island. Several other QTL experienced strong directional selection, but only in one site and seasonal cohort. Thus, different loci were exposed to selection in different natural environments. Selection on allelic variation also depended upon the genetic background. The allelic fitness effects of two QTL reversed direction depending on the genotype at the other locus. Moreover, alternative alleles at each of these loci caused reversals in the allelic fitness effects of a QTL closely linked to TFL1, a candidate developmental gene displaying nucleotide sequence polymorphism consistent with balancing selection. Thus, both environmental heterogeneity and epistatic selection may maintain genetic variation for fitness in wild plant species.     

Cyclo19,31[D-Cys19]-uPA19-31 is a potent competitive antagonist of the interaction of urokinase-type plasminogen activator with its receptor (CD87)

Urokinase-type plasminogen activator (uPA) represents a central molecule in pericellular proteolysis and is implicated in a variety of physiological and pathophysiological processes such as tissue remodelling, wound healing, tumor invasion, and metastasis. uPA binds with high affinity to a specific cell surface receptor, uPAR (CD87), via a well defined sequence within the N-terminal region of uPA (uPA19-31). This interaction directs the proteolytic activity of uPA to the cell surface which represents an important step in tumor cell proliferation, invasion, and metastasis. Due to its fundamental role in these processes, the uPA/uPAR-system has emerged as a novel target for tumor therapy. Previously, we have identified a synthetic, cyclic, uPA-derived peptide, cyclo19,31uPA19-31, as a lead structure for the development of low molecular weight uPA-analogues, capable of blocking uPA/uPAR-interaction [Burgle et al., Biol. Chem. 378 (1997), 231-237]. We now searched for peptide variants of cyclo19,31uPA19-31 with elevated affinities for uPAR binding. Among other tasks, we performed a systematic D-amino acid scan of uPA19-31, in which each of the 13 L-amino acids was individually substituted by the corresponding D-amino acid. This led to the identification of cyclo19,31[D-Cys19]-uPA19-31 as a potent inhibitor of uPA/uPAR-interaction, displaying only a 20 to 40-fold lower binding capacity as compared to the naturally occurring uPAR-ligands uPA and its amino-terminal fragment. Cyclo19,31[D-Cys19]-uPA19-31 not only blocks binding of uPA to uPAR but is also capable of efficiently displacing uPAR-bound uPA from the cell surface and to inhibit uPA-mediated, tumor cell-associated plasminogen activation and fibrin degradation. Thus, cyclo19,31[D-Cys19]-uPA19-31 represents a promising therapeutic agent to significantly affect the tumor-associated uPA/uPAR-system.