Responses of the pancreatic digestive enzyme secretion to amino acids, glucose and cholecystokinin in chicks

1. Pancreatic secretion of digestive enzymes, amylase, trypsinogen and chymotrypsinogen, was studied in response to the wing vein injection of digestive end products, various single amino acids and glucose, with or without cholecystokinin (CCK) in chicks. 2. Among amino acids administered, only phenylalanine significantly (P less than 0.05) increased trypsinogen and chymotrypsinogen secretions, while other amino acids did not. 3. Simultaneous injection of single amino acids with CCK increased digestive enzyme secretion to various extents depending on the kind of amino acids whereas the injection of glucose with CCK did not affect when compared with that of CCK alone. 4. By varying doses, a synergetic action of CCK plus amino acid on the secretion of pancreatic digestive enzymes was observed at 0.5 mM for valine and 5 mM for arginine.     

Amelioration of experimental autoimmune encephalomyelitis with anti-OX40 ligand monoclonal antibody: a critical role for OX40 ligand in migration, but not development, of pathogenic T cells

OX40 (CD134) and its ligand (OX40L) have been implicated in T cell activation and migration. In this study, we examined the contribution of these molecules to the pathogenesis of experimental autoimmune encephalomyelitis (EAE) by administering a neutralizing mAb against murine OX40L (RM134L) to proteolipid protein (139-151) peptide-induced EAE in SJL mice. Administration of RM134L effectively ameliorated the disease in both actively induced and adoptively transferred EAE models. Histological examination showed that the RM134L treatment greatly reduced mononuclear cell infiltration into the spinal cord. The RM134L treatment did not inhibit the development of pathogenic T cells, given that proliferative response and IFN-gamma production by draining lymph node cells were not reduced or rather enhanced upon restimulation with proteolipid protein (139-151) in vitro, and these cells effectively transferred EAE to naive SJL mice. Flow cytometric analyses showed that the RM134L treatment inhibited the accumulation of OX40-expressing CD4(+) T cells and the migration of adoptively transferred CD4(+) T cells in the spinal cord. Immunohistochemical staining showed that OX40L was most prominently expressed on endothelial cells in the inflamed spinal cord. These results suggest that the OX40/OX40L interaction plays a critical role for the migration of pathogenic T cells into the CNS in the pathogenesis of EAE.     

Genetic relationship and distribution of the Japanese wild boar (Sus scrofa leucomystax) and Ryukyu wild boar (Sus scrofa riukiuanus) analysed by mitochondrial DNA

Mitochondrial genetic variations were used to investigate the relationships between two Japanese wild boars, Japanese wild boar (Sus scrofa leucomystax) and Ryukyu wild boar (S.s. riukiuanus). Nucleotide sequences of the control (27 haplotypes) and cytochrome b (cyt-b) regions (19 haplotypes) were determined from 59 Japanese wild boars, 13 Ryukyu wild boars and 22 other boars and pigs. From phylogenetic analyses, the mtDNA of Ryukyu wild boar has a distinct lineage from that of Japanese wild boar, which was classified into the Asian pig lineage. This result suggests that the Ryukyu wild boar has a separate origin from the Japanese wild boar.     

cDNA cloning of the two subunits of phospholipase A2 inhibitor PLIgamma from blood plasma of the Chinese mamushi, Agkistrodon blomhoffii siniticus

Three phospholipase A2 (PLA2) inhibitors (PLI) have been purified from the blood plasma of the Chinese mamushi, Agkistrodon blomhoffii siniticus; 1 of these, PLIgamma, contains 2 homologous subunits, PLIgamma-A and PLIgamma-B. The cDNAs encoding these 2 subunits of PLIgamma were isolated from a liver cDNA library by using fragments from polymerase chain reaction amplifications as probes and sequenced. The respective nucleotide sequences encoded 19-residue signal sequences, followed by 181-residue proteins. The calculated molecular masses were 20123 and 20150 Da for the PLIgamma-A and PLIgamma-B subunits, respectively; and PLIgamma-A included a N-linked carbohydrate site at Asn-157. The sequences of these subunits contained 2 internal repeats of disulfide-bonding pattern characteristic to those of urokinase-type plasminogen activator receptor and members of the Ly-6 superfamily. A phylogenetic analysis comparing the amino acid sequences of PLIgamma-A and PLIgamma-B with those for other snakes revealed that the gene duplication leading to these 2 subunits occurred before the divergence of Viperidae and Elapidae.     

Cytoplasmic domain is not essential for the cell adhesion activities of gicerin, an Ig-superfamily molecule

Gicerin is a cell adhesion molecule in the immunoglobulin (Ig) superfamily and is expressed abundantly during development in the nervous system. It has homophilic cell adhesion activity and also has heterophilic binding activity with NOF (neurite outgrowth factor) and mediates neurite extension. There are two isoforms of gicerin, one with a short (s-gicerin) and the other with a longer cytoplasmic domain (l-gicerin). We have reported that s-gicerin possesses stronger activities than l-gicerin during cell aggregation, in NOF-binding, and in neurite extension. In this study, we established cell lines which expressed a mutant-gicerin whose cytoplasmic domain was deleted and we compared the above three biological activities of the mutant-gicerin with those of s- and l-gicerin. We found that the mutant-gicerin retained all these activities, but the activities were weaker than those of s-gicerin and almost the same as those of l-gicerin. We concluded that the cytoplasmic domain of gicerin is not essential for optimal adhesive activities of gicerin, but might be involved in the regulation of its activities.     

High Efficiency Gene Transfer by Multiple Transfection Protocol

A high efficiency transfection protocol employing a common polycationic lipid is described. Using LipofectAMINE, a widely used transfection reagent, we transfected 293T cells with a plasmid harboring the -galactosidase (-gal) gene. The transfection efficiency was determined by direct staining for X-gal. The conventional transfection protocol achieved an efficiency of <40% while our protocol, which employs the repetition of transfection a few times, achieved an efficiency of approximately 80%. Thus, a dramatic increase in transfection efficiency can be obtained by simply repeating transfection with the use of a common polycationic lipid. This method will be useful in many molecular biological experiments.     

Over-expression of the testis-specific gene TSGA10 in cancers and its immunogenicity

The TSGA10 gene was originally isolated in normal testis by differential mRNA display. TSGA10 is located on chromosome 2q11.2 and consists of 19 exons extending over 3 kb. TSGA10 mRNA expression was investigated in normal and malignant tissues using quantitative real-time RT-PCR. It was predominantly expressed in the testis in adult normal tissues. In malignant tissues, TSGA10 was over-expressed in 4 of 20 hepatocellular carcinomas (HCC), 1 of 20 colon cancers, 7 of 20 ovarian cancers, 3 of 20 prostate cancers, 1 of 21 malignant melanomas, and 8 of 21 bladder cancers. Serological analysis revealed that 3 out of 346 patients with various types of cancer possessed antibody against recombinant TSGA10 protein. They included 2 patients with hepatocellular carcinoma and a patient with malignant melanoma.     

Secretory leukoprotease inhibitor and pulmonary surfactant serve as principal defenses against influenza A virus infection in the airway and chemical agents up-regulating their levels may have therapeutic potential

Influenza A virus (IAV) is one of the most common infectious pathogens in humans. Entry of this virus into cells is primarily determined by host cellular trypsin-type processing proteases, which proteolytically activate viral membrane fusion glycoprotein precursors. Human IAV and murine parainfluenza virus type 1 Sendai virus are exclusively pneumotropic, and the infectious organ tropism of these viruses is determined by the susceptibility of the viral envelope glycoprotein to cleavage by proteases in the airway. Proteases in the upper respiratory tract are suppressed by secretory leukoprotease inhibitor, and those in the lower respiratory tract are suppressed by pulmonary surfactant, which by adsorption inhibits the interaction between the proteases and viral membrane proteins. Although the protease activities are predominant over the activities of inhibitory compounds under normal airway conditions, intranasal administration of inhibitors was able to significantly suppress multi-cycles of viral replication in the airway. In addition, we identified chemical agents that could act as defensive factors by up-regulating the levels of the natural inhibitors and immunoglobulin A (IgA) in airway fluids. One of these compounds, ambroxol, is a mucolytic and anti-oxidant agent that stimulates the release of secretory leukoprotease inhibitor and pulmonary surfactant in the early phase, and IgA in the late phase of infection at an optimal dose, i.e. a dose sufficient to inhibit virus proliferation and increase the survival rate of animals after treatment with a lethal dose of IAV. Another agent, clarithromycin, is a macrolide antibiotic that increases IgA levels through augmentation of interleukin-12 levels and mucosal immunization in the airway. In addition to the sialidase inhibitors, which prevent the release of IAV from infected cells, inhibitors of the processing proteases and chemical agents that augment mucosal immunity and/or levels of the relevant defensive compounds may also ultimately prove to be useful as new anti-influenza agents.     

Cutting edge: a potent adjuvant effect of ligand to receptor activator of NF-kappa B gene for inducing antigen-specific CD8+ T cell response by DNA and viral vector vaccination

The ligand to receptor activator of NF-kappaB (RANK-L)/RANK interaction has been implicated in CD40 ligand/CD40-independent T cell priming by dendritic cells. In this report, we show that the coadministration of the RANK-L gene with a Trypanosoma cruzi gene markedly enhances the induction of Trypanosoma Ag-specific CD8(+) T cells and improves the DNA vaccine efficacy. A similarly potent adjuvant effect of the RANK-L gene on the induction of Ag-specific CD8(+) T cells was also observed when recombinant influenza virus expressing murine malaria Ag was used as an immunogen. In contrast, the coadministration of the CD40L gene was not effective in these systems. Our results demonstrated, for the first time, the potent immunostimulatory effect of the RANK-L gene to improve the CD8(+) T cell-mediated immunity against infectious agents.